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Background: 3-hydroxy-3-methylglutaryl-coenzymOHBe A(HMG-CoA) lyase (HMGCL) catalyzes the cleavage of HMG-CoA into acetyl-CoA and acetoacetic acid and serves as a rate-limiting enzyme in the metabolism of ketone bodies. While HMGCL is involved in various biological processes, its specific role in osteosarcoma remains unclear., Methods: Using data from a public database of osteosarcoma patients, we investigated the expression and prognostic value of HMGCL. The effects of HMGCL on the proliferation, migration, and invasion of osteosarcoma cells were assessed using CCK-8 assays, wound healing tests, and transwell invasion assays. We explored and validated the specific molecular mechanisms by which HMGCL influences osteosarcoma through transcriptome sequencing. Finally, we established a subcutaneous tumor formation model in nude mice to investigate the function of HMGCL in vivo., Results: The expression of HMGCL is downregulated in osteosarcoma and correlates with the prognosis of osteosarcoma patients. Overexpression of HMGCL can inhibit the proliferation, migration, and invasion of osteosarcoma cells, as well as tumor growth in vivo. Through our investigation of the underlying mechanism, we found that HMGCL may inhibit the activation of the PI3K/AKT/mTOR signaling pathway via its product, β-HB. This inhibition promotes the phosphorylation of ULK1, thereby facilitating autophagy in osteosarcoma cells and enhancing the malignancy of the disease., Conclusion: HMGCL inhibits the activation of the PI3K/AKT/mTOR signaling pathway mediated by β-HB, thereby reducing the proliferation, migration, and invasion of osteosarcoma cells while promoting autophagy. HMGCL may represent a new target for the treatment of osteosarcoma, offering new hope for patients with this disease., Competing Interests: Declarations. Ethics approval and consent to participate: The Animal Center of Renmin Hospital of Wuhan University approved all experiments conducted on animals. Our animal experiments strictly follow the ARRIVE guidelines. Consent for publication: All authors agree to publication. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Previous studies have shown that nickel sulfate (NiSO 4 ) increases autophagy in thyroid cells and tissues. As an important organ of the endocrine system, the pancreas not only contributes to the exocrine function of digestion but also has the endocrine function of regulating blood sugar. However, it remains unknown whether NiSO 4 increases pancreatic autophagy. Bradykinin (BK) is an important component of the kallikrein-kinin system (KKS) and has many biological functions, such as reducing autophagy. The purpose of the present study was to explore the effects of BK on NiSO 4 -induced changes in pancreatic endocrine function. The present results demonstrate that NiSO 4 increases fasting blood glucose (FBG) within a certain range and decreases insulin levels in mice. Moreover, NiSO 4 triggers incomplete autophagy in MIN6 cells by upregulating microtubule-associated protein 1 light chain 3-II (LC3II) and Beclin 1 but downregulating p62. Mechanistically, NiSO 4 leads to abnormal activation of autophagy by inhibiting the PI3K/AKT/mTOR signaling pathway. Moreover, BK decreases FBG and increases insulin secretion in mice exposed to NiSO 4 . Light microscopy and transmission electron microscopy (TEM) analyses revealed that BK pretreatment partially restores MIN6 cell viability and decreases the number of autophagic bodies. BK significantly upregulates the protein levels of LC3II and Beclin1 but downregulates p62 in NiSO 4 -induced MIN6 cells. In addition, BK increases the levels of phosphorylated phosphatidylinositol 3-kinase (P-PI3K), phosphorylated protein kinase B (P-AKT) and mammalian target of rapamycin (mTOR). Most of these effects of BK are reversed by treatment with the HOE140 B2R inhibitor. The present results suggested that BK ameliorates NiSO 4 -induced pancreatic β-cell dysfunction through B2R-mediated activation of the PI3K/AKT/mTOR signaling pathway and inhibition of autophagy., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2025 Elsevier Inc. All rights reserved.)

Acute myocardial infarction (AMI) is a leading cause of mortality among cardiovascular diseases, yet effective therapies for AMI are limited. Previous studies have suggested cardioprotective effects of columbianadin (CBN), but its specific role in AMI and the underlying mechanisms remain unclear. This study aims to investigate whether CBN influences AMI and to elucidate the underlying mechanisms. We conducted a network pharmacology analysis to investigate the relationship between CBN and AMI. The AMI model was established by ligating the left anterior descending (LAD) artery in C57BL/6J mice, which were subsequently administered CBN. Hypoxic H9c2 cells were utilized to evaluate the effects of CBN in vitro. Our study revealed that CBN treatment significantly reduced myocardial infarction in AMI mice. It enhanced mitochondrial function and suppressed autophagy flux in hypoxic H9c2 cells. Furthermore, CBN downregulated the expression of LC3, Beclin1, and Atg 5 genes and proteins. In response to CBN treatment, the phosphorylation levels of PI3K, Akt, and mTOR increased. Notably, RAPA attenuated the protective effect of CBN in enhancing the survival of hypoxic H9c2 cells and abolished its regulation of autophagy-related proteins via the PI3K/Akt/mTOR signaling pathway. In conclusion, CBN reduces myocardial damage by suppressing autophagy via the PI3K/Akt/mTOR signaling pathway in AMI mice and hypoxic H9c2 cells., (© 2024 John Wiley & Sons Ltd.)

This study aims to reveal the effects of the herb pair Astragali Radix-Salviae Miltiorrhizae Radix et Rhizoma(AR-SMRR) on phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin(PI3K/Akt/mTOR) pathway and autophagy in the lung tissue of the rat model of acute lung injury(ALI). Fifty adult male SD rats were randomized into sham, model, autophagy inhibition(intraperitoneal injection of chloroquine at 10 mg·kg~(-1)), autophagy induction(intraperitoneal injection of rapamycin at 15 mg·kg~(-1)), and AR-SMRR(5 g·kg~(-1), gavage) groups. The rats in the sham group received intratracheal instillation of normal saline, and those in other groups received intratracheal instillation of lipopolysaccharide(LPS, 5 mg·kg~(-1)) for the modeling of ALI. Seven days before the operation, the rats in the sham and model groups were administrated with normal saline, and those in other groups with corresponding drugs. Specimens were collected 24 h after modeling. The pathological changes of the lung tissue were observed under a light microscope. The lung wet/dry weight ratio and the lactate dehydrogenase(LDH) activity and total protein concentration in the bronchoalveolar lavage fluid(BALF) were measured. Western blot was employed to measure the protein levels of microtubule-associated protein 1-light chain 3(LC3), beclin-1, p62, PI3K, Akt, and mTOR. Compared with the sham group, the model group showed increased histopathological score of the lung tissue, lung wet/dry weight ratio, and LDH activity and protein concentration in BALF. Autophagy inhibition further increased these indicators compared with the model group, while autophagy induction and AR-SMRR lowered the levels. In addition, AR-SMRR up-regulated the protein levels of LC3-Ⅱ and beclin-1, down-regulated the expression of p62, and inhibited the expression of p-PI3K, p-Akt, and p-mTOR in the lung tissue of ALI rats. The findings suggest that AR-SMRR can alleviate the lung injury and edema in the rat model of ALI induced by LPS by enhancing autophagy via down-regulating PI3K/Akt/mTOR signaling pathway.

Polycystic ovary syndrome (PCOS) is a disease in which endocrine metabolic abnormalities coexist with reproductive system abnormalities, with the main clinical manifestations including abnormal menstruation, hirsutism, acne, infertility, and obesity, and it is also a high risk for the development of many pregnancy complications, gynecological malignancies and other diseases. Therefore, timely intervention to prevent the progression of PCOS is of great significance for improving the quality of life of most female patients. Insulin resistance (IR) is one of the most common endocrine disorders in PCOS patients, with approximately 75% of PCOS patients experiencing varying degrees of IR. It is now believed that it is mainly related to the PI3K signaling pathway. The role of autophagy and apoptosis of ovarian granulosa cells (GCs) in the pathogenesis of PCOS has also been gradually verified in recent years. Coincidentally, it also seems to be associated with the PI3K signaling pathway. Our aim is to review these relevant studies, to explore the association between the IR, cellular autophagy and apoptosis in PCOS patients and the PI3K pathway. We summarize some of the drug studies that have improved PCOS as well. We have also found that proteomics holds great promise in exploring the pathogenesis of PCOS, and we have published our views on this., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Tong, Wu, Zhang and Yu.)

Argpyrimidine (APMD), a methylglyoxal-arginine-derived product, is one of the main products of diabetes mellitus. We aimed to systematically investigate the role of APMD in regulating autophagy activity, with a specific focus on the finding of APDM binding molecule, matching amino acid residues, autophagy flux and proteins, cell cycle arrest, cell skeleton and migration, PI3K/AKT/mTOR pathways, inflammatory signals, alveolar bone destruction, and inhibition verification. In this study, binding to 59/94/121 amino acid residues of advanced glycosylation end product receptor (RAGE), APMD suppressed PI3K/AKT/mTOR pathway to attenuate cell survival of periodontal ligament cells (PDLCs). Simultaneously, autophagy proteins ATG5, Beclin1, and LC3-II/I expression ratio were upregulated while P62/SQSTM was downregulated. Cell cycle arrested at G0/G1 with enhancing Cyclin D1/CDK4 and decreasing Cyclin A/CDK2 expression. Inhibition of autophagy abrogated APMD-induced cell cycle arrest. Furthermore, the inflammation regulation network of matrix metalloproteinase (MMP)-2, MMP-9, MAPKs and NF-κB pathways were activated by APMD. Rat periodontal models confirmed that APMD induced alveolar bone resorption, increased inflammatory infiltrates, and degraded collagen fibers through RAGE and PI3K. APMD-induced autophagy, G0/G1 arrest, pro-inflammatory signals activating and periodontal destruction were reversed by RAGE knockdown while aggravated by PI3K inhibitor. This study provides the first evidence that APMD bind to RAGE to regulate autophagy and cell cycle of PDLCs through the PI3K/AKT/mTOR pathway, thereby promoting periodontal destruction., (© 2022 Wiley Periodicals LLC.)

Objective: To assess effect of licochalcone A (LicA) on amyloid-β (Aβ) peptide fragment 25-35-induced nerve injury and reveal the potential molecular mechanisms involved., Methods: Viability of SH-SY5Y cells was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide assay after treatment with Aβ25-35 and/or LicA, following which apoptosis was detected by flow cytometry and Hoechst staining. Then, reactive oxygen species, glutathione, and superoxide dismutase were measured with flow cytometry and spectrophotometry. The ultrastructure of mitochondria was examined by transmission electron microscopy, and the biomarker proteins of autophagy, apoptosis, and phosphoinositide 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling pathway were measured with Western blotting., Results: LicA improved cell viability and decreased lactate dehydrogenase leakage remarkably in Aβ25-35-induced injury in SH-SY5Y cells. After treatment with LicA, reactive oxygen species, glutathione, and superoxide dismutase levels in cells all were significantly decreased, which indicated that LicA has an antioxidative effect on Aβ25-35-induced oxidative injury. LicA could also significantly reduce Aβ25-35-induced autophagy in SH-SY5Y cells. In the cells injured by Aβ25-35, LicA prevented the transformation from light chain protein 3-I to light chain protein 3-II and reduced the levels of proteins GRP78, GRP94, CHOP, and Bax, but increased the levels of antiapoptotic protein and phosphorylation of PI3K, Akt, and mTOR. These effects of LicA were restored or suppressed by mTOR inhibitor rapamycin or PI3K inhibitor LY294002., Conclusions: LicA protects SH-SY5Y cells against Aβ25-35-induced injury, wherein suppressed autophagy and activated PI3K/Akt/mTOR signaling pathway are involved, and mTOR-dependent autophagy at least plays some role., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Soluble fms-like tyrosine kinase-1 (sFlt-1), a circulating antiangiogenic protein, is involved in the pathogenesis of atherosclerosis (AS), and the underlying mechanism is still unclear. Here, we attempted to investigate the mechanism of action of sFlt-1 in AS. Human umbilical vein endothelial cells (HUVECs) were treated with oxidized low density lipoprotein (ox-LDL) to induce cell injury. ox-LDL treatment increased LC3-II/LC3-I ratio, Beclin-1 expression and GFP-LC3 puncta in HUVECs, suggesting that ox-LDL may induce autophagic flux impairment in HUVECs. ox-LDL-treated HUVECs displayed a decrease of sFlt-1 levels. Moreover, ox-LDL treatment reduced cell proliferation and elevated apoptosis in HUVECs, which was abrogated by sFlt-1 overexpression. Up-regulation of sFlt-1 repressed the activity of PI3K/AKT/mTOR signaling pathway and enhanced autophagy in HUVECs following ox-LDL treatment. Additionally, sFlt-1 overexpression-mediated increase of autophagy in ox-LDL-treated HUVECs was abolished by 3-methyladenine (autophagy inhibitor). 3-methyladenine abrogated the impact of sFlt-1 overexpression on proliferation and apoptosis in ox-LDL-treated HUVECs. This work confirmed that overexpression of sFlt-1 activated autophagy by repressing PI3K/Akt/mTOR signaling pathway, and thus alleviated ox-LDL-induced injury of HUVECs. Therefore, this study suggests that sFlt-1 may be a potential target for AS treatment., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Ligustrazine, an alkaloid monomer extracted from Chuanxiong Rhizoma, has the function of protecting nerve cells. However, the effect and mechanism of ligustrazine on retinal ischemia/reperfusion (I/R) injury still need to be clarified. In our study, retinal ganglion cells (RGC-5) were used to establish a retinal I/R injury model by anaerobic cultivation. Cell viability, autophagy, and apoptosis were evaluated by cell counting kit 8 assay, transmission electron microscopy, and TUNEL staining after treatment with ligustrazine, PI3K inhibitor Ly294002, and/or mTOR inhibitor rapamycin, respectively. Besides, the levels of PI3K/Akt/mTOR pathway and autophagy-related proteins were determined by western blot. Moreover, one-way ANOVA was adopted for inter-group comparisons of measurement data. Our results demonstrated that low-concentration ligustrazine significantly enhanced cell viability and suppressed cell autophagy and apoptosis of RGC-5 cells after I/R injury, suggesting the protective effect of low-concentration ligustrazine on retinal I/R injury. Moreover, the alleviating effect of ligustrazine on RGC-5 with retinal I/R injury was mechanistically associated with the activation of the PI3K/Akt/mTOR pathway. In conclusion, low-concentration ligustrazine has a significant protective effect on RGC-5 cells with retinal I/R injury by activating the PI3K/Akt/mTOR pathway.

Bladder cancer is the most common cancer in the urinary system which has threatened lives. Increasing evidence has shown that lncRNAs were expressed in cancer cells as a biomarker and might regulate cancer cell development and progression. It was reported that ADAMTS9-AS1 plays a suppressive role in tumor cell growth and proliferation in prostate cancer. However, it remains unclear whether ADAMTS9-AS1 influences the function of bladder cancer cells. In this study, we detected that ADAMTS9-AS1 is highly expressed in bladder cancer. We observed that the up-regulation of ADAMTS9-AS1 promoted cell proliferation, migration and invasion, and reduced the apoptosis and autophagy of 5637 and T42 cell lines. Meanwhile, the down-regulation of ADAMTS9-AS1 increased the apoptosis and autophagy, and suppressed their proliferation, migration and invasion. Interestingly, the up-regulation of ADAMTS9-AS1 was accompanied by the activation of PI3K/AKT/mTOR signaling pathway, while the down-regulation of ADAMTS9-AS1 lead to an opposite effect. Together, these results demonstrated that ADAMTS9-AS1 promotes bladder cancer cell invasion and migration, and negatively regulates bladder cancer cell apoptosis and autophagy, which might be through PI3K/AKT/mTOR signaling pathway. Targeting ADAMTS9-AS1 might become a potential therapeutic approach in treating bladder cancer., (Copyright © 2021. Published by Elsevier Ltd.)

Ethnopharmacological Relevance: Guizhi Fuling Wan (GFW) is a traditional Chinese medicine used to remove blood stasis and dissipate phlegm for treating gynecological diseases that was invented by Zhang Zhongjing in the Eastern Han dynasty. In recent years, GFW has been widely used to treat patients with polycystic ovary syndrome (PCOS). Clinical and animal studies have shown that it is effective in the treatment of PCOS, but its mechanism is unknown. Generally, it works by regulating autophagy via the PI3K/AKT/mTOR signaling pathway., Aim of the Study: This study investigated the effects and mechanism of GFW in PCOS rats with insulin resistance (IR) in order to provide better understanding of its observed clinical effects and a theoretical basis for the study of traditional Chinese medicine., Materials and Methods: Eighty-four female Sprague-Dawley rats were randomly divided into seven groups (n = 12 per group): 1) control, 2) PCOS model, 3) low-dose GFW, 4) medium-dose GFW, 5) high-dose GFW, 6) metformin, and 7) medium-dose GFW plus LY294002. In all non-control groups, we induced PCOS through daily letrozole combined with intragastric high-fat emulsion for 21 days. After treatment, rats were sacrificed and serum follicle-stimulating hormone (FSH), testosterone (T), progesterone, luteinizing hormone (LH), 17β-estradiol, fasting insulin (FINS), and fasting plasma glucose levels were measured by enzyme-linked immunosorbent assay (ELISA). The LH/FSH ratios and HOMA-IR values were calculated. Ovarian morphology was observed by hematoxylin and eosin staining, and all follicles were counted under a microscope. MDC-positive vesicles were used as markers to detect autophagy, and the expression levels of p62, Beclin1, and LC3-II were examined by immunostaining. Western blotting was used to measure PI3K/AKT/mTOR pathway activation, granulosa cell apoptosis, and autophagy., Results: Compared with the PCOS model group, GFW-treated rats had less atretic and cystic follicles, and more mature follicles and corpus lutea. The GFW-treated rats had lower serum T, LH, and FINS levels than the PCOS model group, as well as lower LH/FSH ratios and HOMA-IR values. GFW treatment resulted in significantly reduced levels of cleaved-Caspase-3, cleaved-Caspase-9, BAX, Beclin1, Atg5, and LC3-II. Phosphorylation of PI3K, AKT, and mTOR was significantly higher in GFW-treated rats compared with the PCOS model group. The phosphorylation of PI3K, AKT, and mTOR was decreased with the use of a PI3K antagonist., Conclusions: Our results indicate that GFW inhibited granulosa cell autophagy and promoted follicular development to attenuate ovulation disorder in PCOS-IR rats. This was associated with activation of the PI3K/AKT/mTOR signaling pathway., (Copyright © 2021 Elsevier B.V. All rights reserved.)

High pathogenicity island (HPI), which is widely distributed in Escherichia coli (E. coli), can enhance the pathogenicity of E. coli. Thus the HPI positive E. coli could pose a threat to human and animal health. It remains to be elucidated how HPI affects the virulence of pathogenic E. coli. Autophagy is an important mechanism to maintain cellular homeostasis and an innate immunity responses of organisms against pathogens. The interaction between pathogenic E. coli possessing HPI (E. coli HPI) and host autophagy system has not been reported. In this study, it was demonstrated that pathogenic E. coli induced autophagy in 3D4/21 macrophages and HPI was associated with enhanced autophagy through transmission electron microscopy, immunofluorescence and real-time PCR. The PI3K/Akt/mTOR pathway is an important negative regulatory pathway for autophagy. Through detecting the expression of key genes of PI3K/Akt/mTOR pathway, it was speculated that HPI enhanced the inhibition of the signaling pathway stimulated by pathogenic E. coli. Furthermore, HPI inhibited the secretion of IFN-γ, while the presence of HPI did not significantly affect the secretion of IL-1β. This work is the first attempt to explore the interplay between HPI carried by pathogenic E. coli and host cell autophagy. The findings might enable better understanding of the contribution of HPI to pathogenicity., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

The duration of denture use, oral hygiene, smoking and male sex were identified as risk factors for oral mucosal lesions. As it is well known, all the oral mucosal lesions associated with risk factors have an important degree of malignity. Chronic mechanical irritation can be another cause of oral cancer and it is produced by the constant action of a deleterious agent from the oral cavity. Autophagy represents a complex evolutionary conserved catabolic process in which cells self-digest intracellular organelles in order to regulate their normal turnover and remove the damaged ones with compromised function to further maintain homeostasis. Autophagy is modulated by mTOR kinase and indirectly by PI3K/AKT survival pathway. Due to its dual capacity to either induce cell death or promote cell survival, important evidence pointed that autophagy has a two-faced role in response to chemotherapy in cancer. In conclusion, understanding how to overcome cytoprotective autophagy and how to take advantage of autophagic cell death is critical in order to enhance the cancer cells sensitivity to particular therapeutic agents.

Objective: To observe the effect of electroacupuncture at "Baihui"(GV20) and "Shenshu"(BL23) on the expression of autophagy-related proteins in the hippocampus of rats with Alzheimer's disease (AD)，so as to explore its underlying mechanisms on improvement of AD., Methods: Forty-eight male SD rats were randomly divided into blank control group, model group, electroacupuncture group and sham electroacupuncture group, with 12 rats in each group. The AD rat model was establish by intraperitoneal injection of D-galactose for 6 weeks. Rats in the electroacupuncture group received electroacupuncture (50 Hz, 1 mA)at GV20 and BL23 for 20 min each time after daily intraperitoneal injection. Rats in the sham electroacupuncture group received acupuncture at the local skin of GV20 and BL23 without electricity. After the intervention, Morris water maze and open field test were used to evaluate the learning and cognitive ability of rats in each group. The transmission electron microscope was used to observe the numerical density of synaptic in hippocampus, and the immunohistochemical staining was used to observe the paired helical filament protein-1 (PHF-1) in the hippocampus. Western blot was used to detected the expression of autophagy-related proteins phosphoinositide-3-kinase (PI3K), protein kinase B (AKT), phosphorylated AKT (p-AKT), mammalian target of rapamycin (mTOR) in the hippocampus., Results: Compared with the blank control group, the escape latency of the rats in the model group increased from day 2 to day 5 ( P <0.01), and the ratio of the time through the quadrant of the original platform reduced （ P <0.01）, in the open field test the distance of exercise, the number of uprights and the rate of exercise time in the central area decreased ( P <0.01), meanwhile the density of hippocampus synapses decreased ( P <0.01), the positive expression of PHF-1 and the relative expression of PI3K, AKT, p-AKT, and mTOR all increased ( P <0.01). Compared with the model group, the escape latency of rats in the electroacupuncture group was shortened from day 2 to day 5 ( P <0.01), and the ratio of the time through the quadrant of the original platform meanwhile, the distance of the open field test, the number of uprights, and the rate of central area exercise time up-regulated ( P <0.01), the numerical density of hippocampus synatic increased ( P <0.01), the positive expression of PHF-1 and the relative expression of PI3K, AKT, p-AKT, and mTOR all down-regulated ( P <0.01). Compared with the model group, the expression of PI3K in the sham electroacupuncture group decreased ( P <0.05)., Conclusion: Electroacupuncture can improve learning and memory and cognitive impairment in AD rats, which may be associated with its effects in regulation of hippocampal autophagy and removal of neurofibrillary tangles by suppressing PI3K/AKT/mTOR signaling pathway.

Osteoarthritis is a degenerative joint disease. Currently, no effective therapeutic exists for osteoarthritis in the clinic setting. Inflammatory response and autophagy are key players in the occurrence and prognosis of osteoarthritis. In recent years, the regulation of inflammation and autophagy signal pathway has been touted as a potential treatment course for osteoarthritis. Saikosaponin D has anti-inflammatory and induces autophagy effects via inhibiting the nuclear transcription factor-κB, mTOR signaling pathways. Here in the report, we analyze and summarize recent evidences pertaining to the relationship between Saikosaponin and osteoarthritis. Published studies were scoured for in research databases, such as PubMed and Scopus with the keywords Saikosaponin and osteoarthritis. Phosphatidylinositol 3-kinase (PI3k)/Akt/mTOR signaling pathway is an important autophagy modulator, and can regulate chondrocytic autophagy, inflammation, and apoptosis. Saikosaponin D alleviates inflammation and regulates autophagy by inhibiting the PI3k/Akt/mTOR signaling pathway. Saikosaponin D could be a potential therapeutic drug for osteoarthritis., (© 2020 John Wiley & Sons, Ltd.)

This article has been retracted: please see Elsevier Policy on Article Withdrawal (http://www.elsevier.com/locate/withdrawalpolicy). This article has been retracted at the request of the Editor-in-Chief. Concern was raised about the reliability of the Western blot results in Figures 5B+D, 6B+D, 7B+D, and 8C, which appear to have a similar phenotype as many other publications, as detailed here: https://pubpeer.com/publications/70795908CC4CEFC1753E19583700F4; and here: https://docs.google.com/spreadsheets/d/1r0MyIYpagBc58BRF9c3luWNlCX8VUvUuPyYYXzxWvgY/edit#gid=262337249. In addition, a portion of Figure 6C, ‘miR-381 mimic’ group appeared to contain image similarities with Figure 6C, ‘si-RELN’ group. The journal requested that the corresponding author comment on these concerns and provide the raw data. The authors did not respond to this request and therefore the Editor-in-Chief decided to retract the article., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Daphnetin (DAP), an active ingredient extracted from Daphne odora, has pharmacological effects such as anti-inflammatory, antioxidation and anti-tumor properties. The current study aims to investigate the relationship between the anti-rheumatoid effect of DAP and the inhibition of both the PI3K/AKT/mTOR and autophagy signaling pathways. DAP inhibited the proliferation of CIA-FLS in a dose-dependent manner and induce apoptosis, accelerated the G1/G0 phase and inhibited the S phase. DAP reduced the phosphorylation of AKT and mTOR and the expression of Atg5, Beclin-1 and LC3-II/LC3-I in CIA-FLS induced by TNF-α. DAP also reduced the inflammatory response in CIA-FLS induced by TNF-α by inhibiting the cytokine expression of TNF-α, IL-6, TGF-β, IL-17, and INF-γ and promoting IL-10 expression. Overall, DAP inhibited the proliferation of CIA-FLS by down-regulating the PI3K/AKT/mTOR signaling pathway and inhibited autophagy in order to induces apoptosis, which may be potential therapeutic approach in treatment of RA., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Background: Autophagy is a self-degrading process. Previously, we showed that insulin-like growth factor binding protein-related protein 1 (IGFBPrP1) is a novel transforming growth factor β1 (TGFβ1)-interacting factor in liver fibrosis; the role of TGFβ1-mediated autophagy in hepatic stellate cells (HSCs) activation has been investigated. However, whether autophagy is regulated by IGFBPrP1 remains unknown., Aims: We investigated the interactions among IGFBPrP1, autophagy, and activation of primary rat HSCs., Methods: Primary HSCs were separated from Sprague Dawley rats by two-step enzymatic digestion, and then, we overexpressed or inhibited IGFBPrP1 expression in HSCs under serum-starved condition. Autophagy inducer rapamycin or inhibitor 3-methyladenine (3MA) was used to assess the relationship between autophagy and HSCs activation., Results: We observed the expression of activation marker α-SMA and autophagy markers such as LC3B and Beclin1, which were significantly increased in HSCs treated with adenovirus vector harboring the IGFBPrP1 gene (AdIGFBPrP1) compared to cells cultured under serum-starved. In comparison, HSCs treated with shIGFBPrP1 showed opposite results. Furthermore, HSCs activation and autophagy increased when cells were treated with rapamycin, whereas opposite results were obtained when cells were treated with 3MA. AdIGFBPrP1 treatment downregulated the phosphorylation of Akt and mTOR., Conclusion: Autophagy was induced in IGFBPrP1-treated primary HSCs, and IGFBPrP1-induced autophagy promoted the activation of HSCs and extracellular matrix expression, the underlying mechanism of which may involve the phosphatidylinositide 3-kinase/Akt/mTOR signaling pathway.

Objective: To observe the eliminating effects of moxibustion at "Baihui" (GV 20), "Fengfu" (GV 16) and "Dazhui" (GV 14) on amyloid β-peptide (Aβ) in brain of the amyloid precursor protein/presenili1 (APP/PS1) double-transgenic mice with Alzheimer's disease (AD) by regulating the phosphoinositide 3-kinases/protein kinase B/mammalian target of rapamycin (PI3K/Akt/mTOR) signaling pathway., Methods: A total of 60 APP/PS1 double-transgenic mice with AD were randomly divided into a model group, a moxibustion group, a rapamycin group and a combination group (treated with moxibustion and inhibitor), 15 mice in each group, another 15 male C57BL/6J mice with same age and background were selected as the control group. In the moxibustion group, pressing moxibustion was applied at "Baihui" (GV 20) while the mild moxibustion was applied at "Fengfu" (GV 16) and "Dazhui" (GV 14). The treatment was manipulated for 20 min each time, once a day for 2 weeks. In the rapamycin group, rapamycin (2 mg/kg) was given by intraperitoneal injection once a day for 2 weeks. On the basis of the treatment in the moxibustion group, 3-methyladenine (1.5 mg/kg) was given by intraperitoneal injection once a day for 2 weeks. The mice in the control and the model group received normal diet and no intervention was given for 2 weeks. Immunohistochemica method was used to measure the levels of Aβ 1-42 in the cerebral cortex and hippocampal, transmission electron microscopy was used to observe the formation of autophagosome in hippocampus, and Western blot method was used to observe the levels of PI3K, Akt, p-Akt, mTOR and p-mTOR in hippocampus., Results: Compared with the control group, the levels of Aβ 1-42 in the cerebral cortex and hippocampal were increased in the model group ( P <0.01). Compared with the model group, the levels of Aβ 1-42 in the cerebral cortex and hippocampal were decreased in the moxibustion group, the rapamycin group and the combination group (all P <0.01), compared with the moxibustion group, the levels of Aβ 1-42 in the cerebral cortex and hippocampal were increased in the combination group ( P <0.01), while there was no significant difference between the moxibustion group and the rapamycin group in the levels of Aβ 1-42 （ P >0.05）. Compared with the rapamycin group, the levels of Aβ 1-42 in the cerebral cortex and hippocampal were increased in the combination group ( P <0.01). In the model group, the cytoplasmic utophagic vacuoles and organelles of neuron were reduced. In the moxibustion group, the utophagic vacuoles were increased, and the organelles showed deformation and atrophy. In the rapamycin group, the utophagic vacuoles were widely disturbed and few deformed organelles were found. In the combination group, few utophagic vacuoles were found and additional organelles showed deformation and atrophy. Compared with the control group, the levels of PI3K、Akt、p-Akt、mTOR and p-mTOR were increased in the model group (all P <0.01). Compared with the model group, the levels of PI3K、Akt、p-Akt、mTOR and p-mTOR were reduced in the moxibustion group, the rapamycin group and the combination group (all P <0.01). Compared with the moxibustion group, the levels of PI3K、Akt、and p-mTOR were increased in the rapamycin group and the levels of PI3K、Akt、p-Akt、mTOR and p-mTOR were increased in the combination group (all P <0.01). Compared with the rapamycin group, the levels of PI3K、Akt、p-Akt、mTOR and p-mTOR were increased in the combination group ( P <0.01)., Conclusion: Moxibustion at acupoints of governor vessel can enhance the autophagy process on Aβ 1-42 in brain of the APP/PS1 double-transgenic AD mice, which may be associated with its effects on inhibiting the abnormal activation of PI3K/Akt/mTOR signaling pathway.

Aflatoxin B 1 (AFB 1 ) is a hazard environmental pollutants and the most toxic one of all the aflatoxins. AFB 1 can cause a serious impairment to testicular development and spermatogenesis, yet the underlying mechanisms remain inconclusive. Oxidative stress acts as a master mechanism of AFB 1 toxicity, and can promote autophagy. Abnormal autophagy resulted in testicular damage and spermatogenesis disorders. The objective of this study was to explore the effect of AFB 1 on autophagy in mice testis and its potential mechanisms. In this study, male mice were intragastrically administered with 0, 0.375, 0.75 or 1.5 mg/kg body weight AFB 1 for 30 days. We found that AFB 1 induced testicular damage, reduced serum testosterone level and impaired sperm quality accompanied with the elevation of oxidative stress and germ cell apoptosis. Interestingly, we observed increasing numbers of autophagosomes in AFB 1 -exposed mice testis. Meanwhile, AFB 1 caused testis abnormal autophagy with the characterization of increased expressions of LC3, Beclin-1, Atg5 and p62. Furthermore, AFB 1 downregulated the expressions of PI3K, p-AKT and p-mTOR in mice testis. Taken together, our data indicated AFB 1 induced testicular damage and promoted autophagy, which were associated with oxidative stress-related PI3K/AKT/mTOR signaling pathway in mice testis., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Fibroblasts excessive proliferation was considered as a decisive reason for epidural fibrosis, which was known as a serious complication of lumbar laminectomy. As a traditional Chinese medicine, triptolide (TP) was used to be proved effective in preventing several fibrosis scar formation diseases. However, little is known about the effect of TP on preventing epidural fibrosis and its possible mechanism. Here, we performed in vitro and in vivo experiments to detect the possible mechanism of TP in preventing epidural fibrosis. In vitro, the effect of TP on impacting fibroblasts proliferation activities was detected by CCK-8, cell cycle assay and EdU incorporation assay. Also, the expressions of cell proliferation protein markers and the expressions of p-PI3K, p-AKT, p-mTOR were detect by Western blot. Besides, the effect of TP on inducing fibroblast apoptosis and autophagy was tested by Western blots, flow cytometry, TUNEL staining, Transmission electron microscope (TEM) analysis and LC3 immunofluorescent staining. The results suggested that TP could suppress the activation of PI3K/AKT/mTOR signaling pathway. Meanwhile, TP could inhibit fibroblast proliferation and induce fibroblast apoptosis as well as autophagy, which was known as two cellular self-destructions. Furthermore, we speculated the possible molecular pathway, through which that TP could inhibit fibroblast proliferation, induce fibroblast apoptosis and autophagy. We used PI3-kinase activator (740Y-P) to activate the PI3K/AKT/mTOR signaling. Activation of PI3K/AKT/mTOR signaling pathway increase the proliferation of fibroblasts and suppressed the autophagy and apoptosis induced by TP. In vivo, we built epidural fibrosis models in rats and locally applied TP of various concentrations. Hematoxylin-eosin (HE) and Masson's trichrome were used to detect the effect of TP on reducing epidural fibrosis. And the results showed that TP could significantly reduce the surgery-induced epidural fibrosis. In conclusion, the results above shown that TP could reduce epidural fibrosis formation, and the potential mechanism might through inhibiting fibroblast proliferation and stimulating apoptosis and autophagy via suppressing PI3K/AKT/mTOR signaling pathway. It might provide a novel thought for reducing surgery-induced epidural fibrosis., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Aconitine, a major aconitum alkaloid, is well known for its high toxicity that induces severe arrhythmias and neurological symptoms. One mechanism of aconitine-induced toxic responses is the induction of apoptosis. Apoptosis and autophagy are interconnected processes and the two pathways share critical components. In this study, we investigated the role of autophagy in aconitine-induced toxicity using mouse model. 120 mice were randomly divided into 4 experimental groups (normal saline), low dose group (0.14 μmol/L), medium dose group (0.28 μmol/L) and high dose group (0.56 μmol/ L). 30 mice in each group were administered with aconitine (lavage) for 30 days. The livers were collected for analysis of autophagy-related proteins by Western blotting. The expression of LC3II/LC3I ratio and Beclin 1 were found to increase and then decrease with the highest expression at 10 days and the p62 showed a time-dependent decreases. Autophagy is regulated by the mTOR pathway, we further analyzed the effects of aconitine on this pathway and found aconitine inhibited, phosphorylation of p-PI3K, p-Akt and p-mTOR. The p-p70s6k and p-4EBP1 which are downstream of mTOR were concomitantly decreased. These results suggest that aconitine induce autophagy in mouse liver. The PI3K/Akt/mTOR signaling pathway is involved in the regulation of aconitine-induced autophagy in the liver of mice., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Ischemic heart disease (IHD) is a significant cause of cardiovascular diseases. MicroRNAs (miRNAs) have been thought to be critical regulators in the heart diseases. The present study was aimed to investigate the effect of miR-129-5p on the autophagy and apoptosis by targeting ATG14 as well as how miR-129-5p worked through the PI3K/AKT/mTOR signaling pathway in H 2 O 2 -induced H9c2 cells. H9c2 cells were induced by H 2 O 2 , after which the expression of miR-129-5p was decreased. Reverse transcription-quantitative polymerase chain reaction (qRT-PCR) was performed to detect the expression level of miR-129-5p in H9c2 cells. In addition, the expression of miR-129-5p and ATG14 were overexpressed or down-regulated after transfection. The transfection efficiency was verified by qRT-PCR. Cell viability, cell apoptosis, and the expression of autophagy and apoptosis-related proteins were determined by CCK-8, flow cytometry and western blotting, respectively. Furthermore, GFP fusion protein analysis was used to detect the expression level of LC3II which was related to autophagy. As a result, cell viability was decreased and cell autophagy was increased in H 2 O 2 -induced H9c2 cells. MiR-129-5p overexpression inhibited cell injury caused by H 2 O 2 in H9c2 cells which was certified by the increased cell viability and decreased cell autophagy and apoptosis. In addition, ATG14 was demonstrated to be a target of miR-129-5p which inhibited cell injury by down-regulation of ATG14. Moreover, phosphorylation of PI3K/AKT/mTOR pathway was activated by miR-129-5p overexpression or ATG14 inhibition to alleviate the autophagy and apoptosis in H 2 O 2 -induced H9c2 cells. In conclusion, this study indicated that miR-129-5p inhibited autophagy and apoptosis in H 2 O 2 -induced H9c2 cells partly by down-regulation of ATG14 through the activation of PI3K/AKT/mTOR pathway., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Cervical cancer (CC) is a malignant solid tumor, which is one of the main causes of morbidity and mortality in women. Given that autophagy is an important factor promoting tumor progression, we aim to investigate the functional role of miR-338 in autophagy and proliferation of cervical cancer. In our study, expression of miR-338 was validated by quantitative RT-PCR in 30 paired cervical cancer tissues and normal tissues. We performed MTT, colony formation and cell cycle assay to explore the effect of miR-338 on cell proliferation. The level of autophagy was evaluated by observing the expression of LC3 formation under fluorescence microscope and detected the LC3 expression by western blot. We used luciferase reporter assays to identify the target gene about miR-338. We not only found that the level of miR-338 is decreased in cervical cancer tissues and cells, but also negatively correlated with the protein level of ATF2. In turn, restoring the expression of miR-338 inhibited proliferation in Hela and SiHa cells. Further mechanistic study identified that ATF2 as a direct target of miR-338. Forced lowexpression of miR-338 directly led to increased the level of autophagy in cervical cancer cells, which was similar to the mTOR signaling inhibitor rapamycin. The western blot analysis show that inhibited miR-338 expression could decrease the p-mTOR and p-p70S6 expression. Thus, we infer that miR-338 decreases autophagy level in cervical cancer cells by activating mTOR signaling pathway. In summary, our study demonstrate that miR-338 could inhibites proliferation and autophagy by targeting ATF2 via mTOR signaling pathway on cervical cancer cells. These results suggest a potential application of miR-338 in cervical cancer as a novel mechanism of tumor therapeutic., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

This article has been retracted: please see Elsevier Policy on Article Withdrawal (http://www.elsevier.com/locate/withdrawalpolicy). This article has been retracted at the request of the Editor-in-Chief. Concern was raised about the reliability of the Western blot results in Figure 5D and 5F, which appear to have a similar phenotype as contained within another publication, as detailed here: https://pubpeer.com/publications/CD4DF7B6DCA28182EC6809846F3653; and here: https://docs.google.com/spreadsheets/d/1r0MyIYpagBc58BRF9c3luWNlCX8VUvUuPyYYXzxWvgY/edit#gid=262337249. Concerns were also raised about the provenance of the flow cytometry data in Figure 7A. In addition, suspected duplications were detected in Western blots contained within Figure 5E and 5F. The journal requested the corresponding author comment on these concerns and provide raw data. The corresponding author communicated that the raw data was not available and some of the results were not repeatable and therefore not solid enough to support the conclusions. The authors requested retraction of this article. The Editor-in-Chief assessed this case and decided to retract the article., (Copyright © 2018. Published by Elsevier Masson SAS.)

Background/aims: Oxidized low-density lipoprotein (oxLDL) promotes unregulated platelet activation in patients with dyslipidemic disorders. Although oxLDL stimulates activating signaling, researchers have not clearly determined how these events drive accelerated thrombosis. Here, we describe the mechanism by which ROS regulate autophagy during ox-LDL-induced platelet activation by modulating the PI3K/AKT/mTOR signaling pathway., Methods: For in vitro experiments, ox-LDL, the ROS scavenger N-acetylcysteine (NAC), the mTOR inhibitor rapamycin and the autophagy inhibitor 3-MA were used alone or in combination with other compounds to treat platelets. Then, platelet aggregation was evaluated on an aggregometer and platelet adhesion was measured under shear stress. The levels of a platelet activation marker (CD62p) were measured by flow cytometry, reactive oxygen species (ROS) levels were then quantified by measuring DCFH-DA fluorescence intensity via flow cytometry. Nitric oxide (NO) and superoxide (O2·-) levels were determined by the nitric acid deoxidize enzyme method and lucigenin-enhanced chemiluminescence (CL), respectively. Transmission electron microscopy was used to observe the autophagosome formation, immunofluorescence staining was employed to detect LC3 expression and western blotting was used to measure the levels of PI3K/AKT/mTOR pathway- and autophagy-related proteins., Results: Ox-LDL-induced platelets showed a significant increase in platelet aggregation and adhesion, CD62p expression, ROS level and O2·- content, with an elevated LC3II/LC3I ratio and Beclin1 expression, but a dramatic reduction in the levels of p62 and pathway-related proteins (all P < 0.05). However, platelet activation and autophagy were aggravated by the Rapamycin treatment, and decreased following treatment with NAC, 3-MA, or NAC and 3-MA, together with increased activity of the PI3K/AKT/mTOR pathway. Additionally, decreased platelet activation and autophagy were observed in platelets treated with NAC and Rapamycin or Rapamycin and 3-MA compared with platelets treated with Rapamycin alone, suggesting that both NAC and 3-MA reversed the effects of Rapamycin., Conclusion: Inhibition of ROS production may reduce autophagy to suppress ox-LDL-induced platelet activation by activating PI3K/AKT/mTOR pathway., (© 2018 The Author(s). Published by S. Karger AG, Basel.)

Objective: This study aims to investigate the effects of PI3K/AKt/mTOR signaling pathway on the proliferation and apoptosis in acute liver failure (ALF) by chaperone mediated autophagy (CMA)., Methods: The hepatocytes extracted from both normal rats and rats with ALF were assigned to control, acute injury, P13K agonist, and P13K inhibitor groups. Quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting were used as part of this investigation to detect the expression of PI3K/AKt/mTOR signaling pathway related-proteins (PI3K, AKt, mTOR), apoptosis related-proteins (Fas, Bax, Bcl-2), chaperone-mediated autophagy (CMA) marker proteins (LAMP-2A, HSC 70), p-PI3K, p-AKt, p-4E-BPI, and p-S6K. An MTT assay was used for analysis of cell proliferation after transfection. Flow cytometry is performed to detect the cell apoptosis., Results: In comparison to the normal group, the model group showed enhanced positive rate of PI3K, AKt, mTOR, increased expression levels of PI3K, AKt, mTOR, Fas, Bax, p-PI3K, p-AKt, p-4E-BPI and p-S6K, reduced expression levels of Bcl-2, LAMP-2A and HSC 70. The results in vitro experiment: compared with the acute injury group, the PI3K agonist group showed elevated expression levels of PI3K, AKt, mTOR, Fas, Bax, p-PI3K, p-AKt, p-4E-BPI and p-S6K, decreased expression levels of Bcl-2, LAMP-2A and HSC 70, inhibited cell proliferation, more arrested cells in G1 stage, and promoted cell apoptosis. Opposing this, the P13K inhibitor group exhibited an opposite trend., Conclusion: In conclusion, inhibition of the PI3K/AKt/mTOR signaling pathway plays a protective role in ALF by promoting CMA expression, which could arrest cell proliferation and promote cell apoptosis., (Copyright © 2017 Elsevier Masson SAS. All rights reserved.)

Background/aims: Lung cancer (LC) continues to be one of the most prevalent cancers around the world. During this study we aimed to investigate the involvement of endoplasmic reticulum stress (ERS) in autophagy, apoptosis, and chemotherapy resistance of mutant p53 LC cells., Methods: Immunohistochemistry was employed to help determine the p53 mutation status of cancer cells from 92 primary LC patients, who were subsequently assigned to either the mutant p53 (n = 39) or wild-type p53 group (n = 53)., Results: Mutant p53 cells exhibited increased expression of the C/EBP homologous protein (CHOP), glucose-regulated protein 78 (GRP78), and inositol-requiring enzyme-1α (IRE1α). The Mutant p53 cells were also found to be sensitive to chemotherapy and displayed decreased expression of PI3K, Akt, and mTOR. The mutant p53 cell lines were treated with tunicamycin to induce ERS and rapamycin in order to inhibit mTOR. Both agents increased the expression of CHOP, GRP78, IRE1α, LC3-II/LC3-I, Atg5, Atg7, caspase-3, caspase-12, cleaved caspase-3, cleaved caspase-12, as well as decreases in cell proliferation as well as the expression levels of PI3K, Akt, and mTOR. Enhanced levels of cell apoptosis and reduced chemotherapy resistance were also detected., Conclusion: The findings of our study suggest that ERS promotes autophagy and apoptosis, while acting to reduce chemotherapy resistance in mutant p53 LC cells by downregulating the PI3K/Akt/mTOR signaling pathway., (© 2017 The Author(s). Published by S. Karger AG, Basel.)

BACKGROUND: Studies have shown that metformin has anti-inflammatory, anti-tumor, anti-aging and vasoprotective effects, and can inhibit the progression of osteoarthritis, but its specific mechanism of action remains unclear. OBJECTIVE: To investigate the mechanism of metformin on cartilage protection in a rat model of osteoarthritis. METHODS: Forty male Sprague-Dawley rats were randomly divided into four groups (n=10 per group): blank, control, sham-operated, and metformin groups. The blank group did not undergo any surgery. In the sham-operated group, the joint cavity was exposed. In the model group and the metformin group, the modified Hulth method was used to establish the osteoarthritis model. At 1 day after modeling, the rats in the metformin group were given 200 mg/kg/d metformin by gavage, and the model, blank, and sham-operated groups were given normal saline by gavage. Administration in each group was given for 4 weeks consecutively. Hematoxylin-eosin staining, toluidine blue staining, and safranin O-fast green staining were used to observe the morphological structure of rat knee joints. Immunohistochemical staining and western blot were used to detect the protein expression of SOX9, type II collagen, a disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS5), Beclin1, P62, phosphatidylinositol 3-kinase (PI3K), p-PI3K, protein kinase B (AKT), p-AKT, mammalian target of rapamycin (mTOR), and p-mTOR in rat cartilage tissue. RESULTS AND CONCLUSION: The results of hematoxylin-eosin, toluidine blue and safranin O-fast green staining showed smooth cartilage surface of the knee joints and normal histomorphology in the blank group and the sham-operated group, while in the model group, there was irregular cartilage surface of the knee joint and cartilage damage, with a decrease in the number of chondrocytes and the content of proteoglycans in the cartilage matrix. In the metformin group, there was a significant improvement in the damage to the structure of the cartilage in the knee joints of the rats, and the cartilage surface tended to be smooth, with an increase in the number of chondrocytes and the content of proteoglycans in the cartilage matrix. Immunohistochemistry staining and western blot results showed that compared with the control and sham-operated groups, the expression of SOX9, type II collagen, and Beclin1 proteins in the cartilage tissue of rats in the model group was significantly decreased (P < 0.05). Conversely, the expression of ADAMTS5, P62, as well as p-PI3K, p-AKT, and p-mTOR proteins was significantly increased (P < 0.05). Furthermore, compared with the model group, the expression of SOX9, type II collagen, and Beclin1 proteins in the cartilage tissue of rats in the metformin group was significantly increased (P < 0.05), while the expression of ADAMTS5, P62, as well as p-PI3K, p-AKT, and p-mTOR proteins was significantly decreased (P < 0.05). To conclude, Metformin can improve the autophagy activity of chondrocytes and reduce the degradation of cartilage matrix in osteoarthritis rats by inhibiting the activation of PI3K/AKT/mTOR signaling pathway, thus exerting a protective effect on articular cartilage. [ABSTRACT FROM AUTHOR]

AIM: To explore the possible mechanism of Yiqi-Huoxue formula （YQHXF） in treating cerebral ischemia-reperfusion injury. METHODS: Male SD rats were randomly divided into six groups, namely, the sham, model, nimodipine, and low-, middle- and high-dose YQHXF groups. The middle cerebral artery occlusion/reperfusion （MCAO/R） model was established in all groups except the sham group. After successful modeling, the YQHXF low-, medium-, and high-dose groups were given 3. 8, 7. 5, and 15 g∙kg-1∙d-1 of YQHXF, respectively, by gavage, while the nimodipine group was given 12 mg∙kg-1∙d-1 of nimodipine tablets by gavage. The sham and model groups were given 10 mL∙ kg-1∙d-1 of distilled water by gavage. After 14 days of drug intervention, the rats were euthanized and the neurological function was evaluated. The infarct volume was assessed using 2, 3, 5-triphenyltetrazolium chloride （TTC） staining and brain histopathological changes were determined by hematoxylin-eosin （HE） staining. Transmission electron microscopy was used to investigate changes in autophagosomes, with immunofluorescence used to assess expression of microtubule-associated protein 1 light chain 3 （LC3） protein in the cerebral cortex, Western blot was used to measure protein levels of p-PI3K, PI3K, p-Akt, Akt, p-mTOR, mTOR, LC3B, p62, beclin-1, and Atg5, and RT-qPCR was used to determined LC3 and P62 mRNA expression. RESULTS: Compared with the sham group, the neural function scores of rats in the model group rats were significantly increased, and TTC staining revealed large areas of white cerebral infarction. There was severe pathological damage to the cerebral tissue in the ischemic cortical area, and large numbers of autophagosomes were seen inside the cells. Immunofluorescence staining showed significant numbers of LC3B-positive cells （P<0. 01）. Protein expression of beclin-1, Atg5, and LC3-II/LC3-I was significantly upregulated （P<0. 01）, while that of p62 was markedly downregulated （P<0. 01）. The expression of p-PI3K/PI3K, p-Akt/Akt, and p-mTOR/mTOR proteins was also significantly reduced （P<0. 01）. In addition, the mRNA expression of LC3 was significantly upregulated （P<0. 01）, with downregulation of P62 mRNA levels （P < 0. 01）. Compared with the model group, both the YQHXF medium- and highdose groups showed upregulated LC3-II/LC3-I values after 12 h of reperfusion （P<0. 01）, followed by downregulation of the ratios （P<0. 05） after 3, 7, and 14 days of reperfusion. Furthermore, after 14 days of reperfusion, compared with the model group, the middle- and high-dose YQHXF groups and the nimodipine group showed reduced neurological function scores（ P<0. 01）, reduced cerebral infarction volumes（ P<0. 01）, improvements in the pathological damage to cortical tissue, and reduced autophagosome formation to varying degrees. At the same time, the number of LC3B-positive cells was reduced （P<0. 01）. Protein expression of beclin-1, Atg5, and LC3-II/LC3-I was significantly downregulated, while that of p62 was upregulated （P<0. 01）. The mRNA expression of LC3 and p62 was consistent with the protein levels （P< 0. 01）. In addition, the expression of p-PI3K/PI3K, p-Akt/Akt, and p-mTOR/mTOR proteins was upregulated （P< 0. 01）. CONCLUSION: YQHXF can dynamically regulate autophagy in ischemic brain tissue, with inhibition of excess autophagy by activation of the PI3K/Akt/mTOR signaling pathway, thus reducing the infarct volume, alleviating brain damage, and promoting the recovery of neurological function. [ABSTRACT FROM AUTHOR]

Monocrotaline (MCT), a major pyrrolizidine alkaloid, is well-known for its high liver toxicity. Dysregulation of autophagy induced apoptosis can lead to various liver diseases, including those induced by chemical compounds. Therefore, we aim to explore whether autophagy might serve as a potential strategy for addressing liver apoptosis caused by MCT. In primary rat hepatocytes (PRHs), MCT significantly increased the number of autophagosomes and the expression levels of LC3II, Becline-1, and Atg5, while it decreased the expression of p62 in a concentration-dependent manner at doses of 100, 200, 300, and 400 µM. Western blot assays revealed MCT inhibited the phosphorylation levels of the PI3K/AKT/mTOR pathway. To elucidate the role of autophagy in mediating MCTinduced apoptosis, we further pretreated PRHs with the autophagy agonist Rapamycin and the inhibitors Bafilomycin A1 and Chloroquine, respectively, and assessed the apoptosis of PRHs induced by MCT. The results displayed that Rapamycin increased the apoptosis rate and the expression of cleaved caspase-3, whereas Bafilomycin A1 and Chloroquine reduced the apoptosis and the expression of cleaved caspase-3 in PRHs. This study confirms that autophagy enhances PRHs apoptosis induced by MCT. In summary, this study demonstrates that MCT-induced autophagy via inhibition of the PI3K/AKT/mTOR pathway can lead to apoptosis in PRHs. [ABSTRACT FROM AUTHOR]

Objective: To investigate influences of curcumin on LPS-induced autophagy in corneal stromal cells by regulating phosphatidylinositol 3-kinase (PI3K)/serine-threonine kinase (AKT)/mammalian target of rapamycin (mTOR) signaling pathway. Methods: Mouse corneal stromal cells were grouped into control group, LPS group (1 µg/µl LPS), L-Cur group (15 µmol/L curcumin), M-Cur group (30 µmol/L curcumin), H-Cur group (50 µmol/L curcumin), and H-Cur+740Y-P group (50 µmol/L curcumin+ 10 µmol/L PI3K activator 740Y-P). MTT was applied to detect effect of curcumin on corneal stromal cytotoxicity and cell viability; ELISA was applied to detect levels of inflammatory factors in corneal stromal cells; immunofluorescence staining was applied to detect levels of Beclin1 and LC3 in mouse corneal stromal cells; Western blot was applied to detect expressions of autophagy-related proteins and pathway-related proteins in corneal stromal cells. Results: 0~90 µmol/L curcumin had no obvious toxicity to corneal stromal cells. Compared with control group, A570 nm, levels of Beclin1 and LC3 II/I in LPS group were obviously decreased (P<0.05), levels of IL-6, IL-8, p-PI3K/PI3K, p-AKT/AKT and p-mTOR/mTOR were obviously increased (P<0.05); compared with LPS group, A570 nm,, levels of Beclin1 and LC3II/I in L-Cur group, M-Cur group, H-Cur group were obviously increased in turn (P<0.05), levels of IL-6, IL-8, p-PI3K/PI3K, p-AKT/AKT and p-mTOR/mTOR were obviously decreased in turn (P<0.05); H-Cur+740Y-P eliminated beneficial effect of curcumin on corneal stromal cells. Conclusion: Curcumin may promote LPS-induced autophagy in corneal stromal cells by down-regulating PI3K/AKT/mTOR signaling pathway. [ABSTRACT FROM AUTHOR]

Zhen Wang,1,&ast; Hui Xu,1,2,&ast; Zheng Wang,1,&ast; Yu Wang,3,&ast; Jieyao Diao,1 Juntao Chen,1 Yuchen Xie,4 Lijuan Zhang,5 Miaoxiu Li,6 Yanqin Bian,7 Yunfeng Zhou1 1College of Acupuncture and Massage, Henan University of Chinese Medicine, Zhengzhou, People’s Republic of China; 2Tuina Department, The Third Affiliated Hospital of Henan University of Chinese Medicine, Zhengzhou, People’s Republic of China; 3College of Computer Science, Xidian University, Xian, People’s Republic of China; 4Tuina Department, Henan Provincial Hospital of Traditional Chinese Medicine, Zhengzhou, People’s Republic of China; 5Rehabilitation Department, Jiaozuo Coal Industry (Group) Co. Ltd. Central Hospital, Jiaozuo, People’s Republic of China; 6College of Acupuncture and Massage, Shanghai University of Chinese Medicine, Shanghai, People’s Republic of China; 7Orthopaedic Research Laboratory, University of California, Davis, CA, USA&ast;These authors contributed equally to this workCorrespondence: Hui Xu; Yunfeng Zhou, Email 15036065036@163.com; zyf5680198@126.comBackground: Knee osteoarthritis (KOA) is on the rise due to lifestyle changes, obesity, and aging, yet effective treatments are lacking. Traditional Chinese manual therapy (Tuina) is promising for KOA. However, its mechanism remains unclear.Objective: This study aims to determine the effects of Tuina on a rat KOA model, focusing on the role of chondrocyte apoptosis and autophagy mechanisms.Methods: KOA was induced in rats by intra-articular injection of L-cysteine-activated papain into the right knee. Thirty-six male Sprague Dawley (SD) rats were randomly divided into blank, model control, Tuina, and positive drug groups. Paw withdrawal threshold tests, knee joint swelling, and passive range of motion assessed knee behavior. Cartilage tissue cytology, cytokine contents, and the mRNA and protein expression of PI3K/AKT/mTOR signaling pathway components were analyzed using HE and TUNEL staining, ELISA, RT-qPCR, and Western blotting, respectively. In addition, we used machine learning methods to conduct a secondary analysis of the dataset from the in vivo experiments in rats to verify the findings.Results: Tuina significantly relieved pain and joint swelling, and improved range of motion. Staining showed reduced articular cartilage destruction and apoptosis. Tuina reduced the serum levels of IL-1β, IL-17, MMP-3, and MMP-13. Tuina downregulated Bax, ULK1, Beclin-1, LC3-II/I and upregulated PI3K, AKT, mTOR, and BCL-2 in cartilage tissue. The machine learning results indicated an 83.33% accuracy for the prediction model, remaining stable through both uni- and multivariate analyses. Tuina yielded the best comprehensive efficacy on KOA as well as better rat behavior and PI3K/AKT/mTOR signaling pathway improvement effect than positive drugs, while its cytokine-reducing ability was comparable to that of positive drugs.Conclusion: Tuina can alleviate cartilage tissue injury in KOA, relieve inflammation, and reduce chondrocyte apoptosis and autophagy, the underlying mechanisms of which may be associated with activation of the PI3K/AKT/mTOR signaling pathway.Keywords: knee osteoarthritis, apoptosis, autophagy, tuina, PI3K/AKT/mTOR signaling pathway, in vivo experiment, machine learning

Objective: To investigate the effects of isoliensinine (Iso) on the proliferation, apoptosis and autophagy of colon cancer SW480 cells through PI3K/Akt/mTOR signaling pathway. Methods: Colon cancer SW480 cells were treated with 10, 20 and 40 μmol/L Iso, and the effects of Iso on the cell proliferation capacity, apoptosis and expressions of autophagy related proteins LC3 I, LC3II and p62 were detected by CCK-8, flow cytometry and Western blot, respectively. Then, SW480 cells were treated respectively with 20 μmol/L Iso and 25 μmol/L PI3K activator 740 Y-P, and the cells were divided into the control group, the 740 Y-P group, the Iso group and the Iso+740 Y-P group. The effects of 740 Y-P on the apoptosis and the expressions of LC3 I, LC3 II, p62, PI3K, p-PI3K, mTOR and p-mTOR proteins in each group were detected by flow cytometry and WB. Results: After treatments with 10, 20 and 40 μmol/L Iso, the proliferation capacity of SW480 cells was decreased significantly (all P<0.05); the apoptosis rate was increased significantly (all P<0.05); the expressions of LC3 II/LC3 I were up-regulated significantly (all P<0.05), and the expression of p26 protein was down-regulated significantly (all P<0.05). After treatments with Iso and 740 Y-P, compared with the control group, the apoptosis rate and LC3 II/LC3 I expression of the 740 Y-P group were decreased significantly (both P<0.05), while the expressions of p26, p-PI3K/PI3K and p-mTOR/mTOR were increased significantly (all P<0.05).The apoptosis rate and LC3 II/LC3 expression in the Iso group were increased (both P<0.05) and the expressions of p26, p-PI3K/PI3K and p-mTOR/mTOR were decreased (all P<0.05). Compared with the 740 Y-P group, the apoptosis rate and LC3 II/LC3 I expression were increased in the Iso+740 Y-P group (P<0.05), while the expressions of p26, p-PI3K/PI3K and p-mTOR/mTOR were decreased (all P<0.05). Compared with the Iso group, the apoptosis rate and LC3 II/LC3 I expression were decreased (both P<0.05), and the expressions of p26, p-PI3K/PI3K, and p-mTOR/mTOR were increased significantly (all P<0.05) in the Iso+740 Y-P group. Conclusion: Iso inhibits the proliferation and induces the apoptosis and autophagy of SW480 cells by inhibiting PI3K/Akt/mTOR signaling pathway. [ABSTRACT FROM AUTHOR]

Autophagy is a vital cellular process that functions to degrade and recycle damaged organelles into basic metabolites. This allows a cell to adapt to a diverse range of challenging conditions. Autophagy assists in maintaining homeostasis, and it is tightly regulated by the cell. The disruption of autophagy has been associated with many diseases, such as neurodegenerative disorders and cancer. This review will center its discussion on providing an in-depth analysis of the current molecular understanding of autophagy and its relevance to brain tumors. We will delve into the current literature regarding the role of autophagy in glioma pathogenesis by exploring the major pathways of JAK2/STAT3 and PI3K/AKT/mTOR and summarizing the current therapeutic interventions and strategies for glioma treatment. These treatments will be evaluated on their potential for autophagy induction and the challenges associated with their utilization. By understanding the mechanism of autophagy, clinical applications for future therapeutics in treating gliomas can be better targeted.

Copyright of Journal of Hainan Medical University is the property of Journal of Hainan Medical College Editorial Office and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Parkinson's disease (PD), as a chronic and progressive neurodegenerative disease, is associated with autophagy. This study focused on the regulation of sinomenine (SN) on autophagy in PD and its related mechanism. The PD mouse model was constructed by MPTP inducement, and the mouse motor function after modeling and SN treatment was examined by rotarod, grip strength, and foot printing tests. Tyrosine hydroxylase (TH)/LC3B-positive neurons in the substantia nigra pars compacta of mouse brains were detected by immunofluorescence. The expressions of proteins related to autophagy (Beclin1, p62, LC3-I and LC3-II) and phosphorylated phosphoinositide 3-kinase (PI3K)/AKT/mechanistic target of rapamycin kinase (mTOR) signaling pathway were measured by western blot. Rescue experiments were performed to determine the effects of MHY1485 (mTOR activator) on SN-treated PD mice. SN potentiated the motor ability in PD mice, promoted the survival of dopaminergic neurons, increased the protein expression level of Beclin1, LC3-II/LC3-I ratio and LC3B-positive neurons, lowered the protein expression level of p62 and inactivated PI3K/AKT/mTOR pathway in the substantia nigra tissue of mouse brains. Moreover, MHY1485 reversed the above effects of SN on PD mice via reactivating PI3K/AKT/mTOR pathway. SN augments the autophagy of dopaminergic neurons via inhibiting the PI3K/AKT/mTOR pathway and exerts a neuroprotective effect on PD mice. [ABSTRACT FROM AUTHOR]

Background: Low back pain is a global health problem that originated mainly from intervertebral disc degeneration (IDD). Autophagy, negatively regulated by the phosphatidylinositol 3‐kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling pathway, prevents metabolic and degenerative diseases by removing and recycling damaged cellular components. Despite growing evidence that autophagy occurs in the intervertebral disc, the regulation of disc cellular autophagy is still poorly understood. Methods: Annulus fibrosus (rAF) cell cultures derived from healthy female rabbit discs were used to test the effect of autophagy inhibition or activation on disc cell fate and matrix homeostasis. Specifically, different chemical inhibitors including rapamycin, 3‐methyladenine, MK‐2206, and PP242 were used to modulate activities of different proteins in the PI3K/Akt/mTOR signaling pathway to assess IL‐1β‐induced cellular senescence, apoptosis, and matrix homeostasis in rAF cells grown under nutrient‐poor culture condition. Results: Rapamycin, an inhibitor of mTOR complex 1 (mTORC1), reduced the phosphorylation of mTOR and its effector p70/S6K in rAF cell cultures. Rapamycin also induced autophagic flux as measured by increased expression of key autophagy markers, including LC3 puncta number, LC3‐II expression, and cytoplasmic HMGB1 intensity and decreased p62/SQSTM1 expression. As expected, IL‐1β stimulation promoted rAF cellular senescence, apoptosis, and matrix homeostatic imbalance with enhanced aggrecanolysis and MMP‐3 and MMP‐13 expression. Rapamycin treatment effectively mitigated IL‐1β‐mediated inflammatory stress changes, but these alleviating effects of rapamycin were abrogated by chemical inhibition of Akt and mTOR complex 2 (mTORC2). Conclusions: These findings suggest that rapamycin blunts adverse effects of inflammation on disc cells by inhibiting mTORC1 to induce autophagy through the PI3K/Akt/mTOR pathway that is dependent on Akt and mTORC2 activities. Hence, our findings identify autophagy, rapamycin, and PI3K/Akt/mTOR signaling as potential therapeutic targets for IDD treatment. [ABSTRACT FROM AUTHOR]

Abstract Background Low back pain is a global health problem that originated mainly from intervertebral disc degeneration (IDD). Autophagy, negatively regulated by the phosphatidylinositol 3‐kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling pathway, prevents metabolic and degenerative diseases by removing and recycling damaged cellular components. Despite growing evidence that autophagy occurs in the intervertebral disc, the regulation of disc cellular autophagy is still poorly understood. Methods Annulus fibrosus (rAF) cell cultures derived from healthy female rabbit discs were used to test the effect of autophagy inhibition or activation on disc cell fate and matrix homeostasis. Specifically, different chemical inhibitors including rapamycin, 3‐methyladenine, MK‐2206, and PP242 were used to modulate activities of different proteins in the PI3K/Akt/mTOR signaling pathway to assess IL‐1β‐induced cellular senescence, apoptosis, and matrix homeostasis in rAF cells grown under nutrient‐poor culture condition. Results Rapamycin, an inhibitor of mTOR complex 1 (mTORC1), reduced the phosphorylation of mTOR and its effector p70/S6K in rAF cell cultures. Rapamycin also induced autophagic flux as measured by increased expression of key autophagy markers, including LC3 puncta number, LC3‐II expression, and cytoplasmic HMGB1 intensity and decreased p62/SQSTM1 expression. As expected, IL‐1β stimulation promoted rAF cellular senescence, apoptosis, and matrix homeostatic imbalance with enhanced aggrecanolysis and MMP‐3 and MMP‐13 expression. Rapamycin treatment effectively mitigated IL‐1β‐mediated inflammatory stress changes, but these alleviating effects of rapamycin were abrogated by chemical inhibition of Akt and mTOR complex 2 (mTORC2). Conclusions These findings suggest that rapamycin blunts adverse effects of inflammation on disc cells by inhibiting mTORC1 to induce autophagy through the PI3K/Akt/mTOR pathway that is dependent on Akt and mTORC2 activities. Hence, our findings identify autophagy, rapamycin, and PI3K/Akt/mTOR signaling as potential therapeutic targets for IDD treatment.

To explore the effect and mechanism of hydrogen sulfide (H2S) on autophagy and angiogenesis in skin wound of diabetic rats. METHODS: Among 36 healthy 8-week-old male Sprague-Dawley rats, 12 rats were selected as control group, and the remaining rats were intraperitoneally injected with streptozotocin (STZ) to induce diabetic model and were randomly divided into diabetes mellitus (DM) group and NaHS (H2S donor) intervention (DM+NaHS) group, with 12 rats in each group. A skin trauma model was established by excising the skin of the back of rats in each group. The rats in DM+NaHS group were intraperitoneally injected with NaHS (56 µmol/kg), and the rats in control and DM groups were daily received the same volume of normal saline for 21 consecutive days. The healing of skin wound was measured on days 0, 7, 14 and 21 after operation. On the 21st day after surgery, the content of H2S in skin tissues was detected by C-7Az fluorescent probe, and the morphological changes and angiogenesis of wound tissues were observed by HE staining. The expression of CD31 was detected by immunofluorescence staining, and endothelial autophagy was detected by double staining of CD31 and beclin-1. The protein levels of cystathionine γ-lyase (CSE), CD31, microtubule-associated protein 1 light chain 3 (LC3), beclin-1, P62, Bcl-2, Bax, phosphatidylinositol 3-kinase (PI3K), protein kinase B (PKB/Akt) and mammalian target of rapamycin (mTOR) in wound tissues were determined by Western blot. Caspase-3 and propidium iodide (PI) staining was used to detect cell apoptosis, and apoptosis of vascular endothelial cells was determined with CD31 and TUNEL double immunofluorescence staining. RESULTS: Compared with DM group, the wound healing rate, H2S content and CSE protein expression were significantly increased in DM+NaHS group (P<0. 01), but still lower than those in control group (P<0.01). HE staining showed that the wound surface in DM group was thin and wide, with few capillary, while that in DM+NaHS group was thicker with lots of capillary and wound width was reduced. Compared with DM group, CD31 expression was markedly increased (P<0. 01), the fluorescence intensity of caspase-3 and PI was significantly decreased (P<0. 01, and CD31+/beclin-1+ as well as CD31+/TUNEL+ cells were decreased (P<0. 01) in DM+NaHS group. Western blot analysis showed that compared with DM group, the levels of beclin-1, Bax and LC3-II/LC3-I were significantly decreased (P<0. 01), while the levels of P62 and Bcl-2, as well as ratios of p-PI3K/PI3K, p-Akt/ Akt and p-mTOR/mTOR were significantly increased (P<0. 01) in DM+NaHS group. CONCLUSION: H2S can promote skin wound healing, which may be related to activation of PI3K/Akt/mTOR signaling pathway, inhibition of endothelial autophagy and apoptosis, and promotion of angiogenesis in diabetic rats. [ABSTRACT FROM AUTHOR]

• Intranasal insulin alleviates neurological damage caused by traumatic brain injury. • Autophagy and endoplasmic reticulum stress-mediated apoptosis are crucial players. • The neuroprotective effects are counteracted by rapamycin, an mTOR inhibitor. Intranasal insulin reduces lesion size and enhances memory capacity in traumatic brain injury (TBI) models, but the molecular mechanisms behind this neuroprotective action not yet understood. Here we used Feeney's free-falling method to construct TBI mouse models and administrated intranasal insulin, rapamycin, insulin and rapamycin, or normal saline to assess their effects on neurological functions, cerebral edema, and the expression of Iba1 in microglia through immunofluorescence assay. We also measured concentrations of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) in the brain using enzyme immunosorbent assay, investigated apoptosis with TUNEL staining and Western blotting, and evaluated autophagy, endoplasmic reticulum (ER) stress, and PI3K/Akt/mTOR signaling pathway with Western blotting. The autophagosome was assessed through transmission electron microscopy. Our findings demonstrated that intranasal insulin promoted neurological recovery, decreased brain swelling, and reduced injury lesions on days 1, 3, and 7 post TBI. Moreover, intranasal insulin reduced microglia activation and the concentration of IL-1β or TNF-α on the same days. Through Western blotting and transmission electron microscopy, we observed that intranasal insulin suppressed autophagy while activating the PI3K/AKT/mTOR signaling pathway on days 1 and 3 post TBI. TUNEL assay and Western blotting also indicated that intranasal insulin inhibited ER stress-mediated apoptosis. Interestingly, the mTOR inhibitor rapamycin partially blocked the pro-autophagy and anti-apoptosis effects of intranasal insulin both on days 1 and 3 post TBI. Our results suggest that intranasal insulin can ameliorate TBI by regulating autophagy and ER stress-mediated apoptosis through the PI3K/AKT/mTOR signaling pathway, providing a promising therapeutic strategy for TBI. [ABSTRACT FROM AUTHOR]

More and more evidence shows that the pathological mechanism of vascular dementia (VD) is closely related to oxidative stress injury, cell apoptosis, autophagy, inflammatory response, excitatory amino acid toxicity, synaptic plasticity change, calcium overload, and other processes. Edaravone dexborneol (EDB) is a new type of neuroprotective agent that can improve the neurological damage caused by an ischemic stroke. Previous studies showed that EDB has effects on synergistic antioxidants and induces anti-apoptotic responses. However, it remains unclear whether EDB can affect apoptosis and autophagy by activating the PI3K/Akt/mTOR signaling pathway and its impact on the neuroglial cells. In this study, we established the VD model of rats by bilateral carotid artery occlusion to explore the neuroprotective effect of EDB and its mechanism. Morris Water Maze test was applied to assess the cognitive function of rats. H&E and TUNEL staining were applied to observe the cellular structure of the hippocampus. Immunofluorescence labeling was used to observe the proliferation of astrocytes and microglia. ELISA was applied to examine the levels of TNF-α, IL-1β and IL-6, and RT-PCR was applied to examine their mRNA expression levels. Western blotting was applied to examine apoptosis-related proteins (Bax, Bcl-2, Caspase-3), autophagy-related proteins (Beclin-1, P62, LC3B), PI3K/Akt/mTOR signaling pathway proteins and their phosphorylation levels. The results indicated that EDB ameliorates learning and memory in rats subjected to the VD model, alleviates neuroinflammatory response by reducing the proliferation of the neuroglial cell and inhibits apoptosis and autophagy, which may be mediated by the PI3K/Akt/mTOR signaling pathway. [ABSTRACT FROM AUTHOR]

Abstract Background Osteoarthritis (OA) is a chronic degenerative joint disease characterized by cartilage degeneration and intra-articular inflammation. Daurisoline (DAS) is an isoquinoline alkaloid isolated from Rhizoma Menispermi, whose antitumor and anti-inflammatory pharmacological effects have been demonstrated, but the effects of DAS on OA have rarely been researched. In this study, we aimed to explore the potential role of DAS in OA and its partial mechanism. Materials and methods The cytotoxicity of H2O2 and DAS toward chondrocytes was detected by the Cell Counting Kit-8 assay. Safranin O staining was used to detect chondrocyte phenotype changes. Cell apoptosis was measured by both flow cytometry and quantitative analysis of the protein levels of the apoptosis-related factors Bax, Bcl-2 and cleaved caspase-3 by western blot. Western blotting and immunofluorescence were used to assess the expression of the autophagy-related proteins LC3, Beclin-1 and p62. In addition, key signal pathway targets and matrix-degrading indicators were measured by western blot. Results Our results indicated that H2O2 induced human chondrocyte apoptosis and activated autophagy in a dose-dependent manner. DAS treatment dose-dependently reversed the expression of apoptosis-related proteins (Bax, Bcl-2 and cleaved caspase3) and the apoptosis rate induced by H2O2. Western blot and immunofluorescence analyses showed that DAS decreased the H2O2-induced upregulation of the autophagy marker Beclin-1 and the LC3 II/LC3 I ratio and upregulated the p62 protein level. Mechanistically, DAS inhibited autophagy through the activation of the classical PI3K/AKT/mTOR signaling pathway and protected chondrocytes from apoptosis. In addition, DAS alleviated the H2O2-induced degradation of type II collagen and the high expression of matrix metalloproteinase 3 (MMP3) and MMP13. Conclusion Our research demonstrated that DAS alleviated chondrocyte autophagy caused by H2O2 through activation of the PI3K/AKT/mTOR signaling pathway and protected chondrocytes from apoptosis and matrix degradation. In conclusion, these findings suggest that DAS may serve as a promising therapeutic strategy for OA.

Copyright of Chinese Journal of Cancer Biotherapy is the property of Editorial Office of Chinese Journal of Cancer Biotherapy and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Dexmedetomidine has been shown to protect against cerebral ischemia-reperfusion injury (CIRI). Nevertheless, the precise mechanism is obscure. In order to explore the effect of dexmedetomidine pre-conditioning on autophagy against CIRI in rats, middle cerebral artery occlusion (MCAO) was conducted to establish cerebral ischemia-reperfusion (I/R) model in male SD rats with 2 h ischemia and 24 h reperfusion. Dexmedetomidine was delivered to rats at 10, 50 and 100 µg/kg doses respectively, and LY294002, a PI3K/Akt/mTOR pathway inhibitor, was administered at 10 mg/kg intraperitoneally 30 min before MCAO. Neurological deficit score was assessed and cerebral infarct size was detected by TTC staining. Morris water maze (MWM) was performed to estimate spatial learning and memory ability. Furthermore, to detect activity of PI3K/Akt/mTOR pathway and autophagy, p-Akt, p-mTOR, Beclin-1 and LC3 were measured by western blot. Our findings revealed that 50 and 100 µg/kg of dexmedetomidine pretreatment could improve the neurological deficit score and reduce cerebral infarct size after CIRI, while these effects were markedly suppressed by LY294002. In MWM test, dexmedetomidine was confirmed to shorten escape latency and increase times across platform after CIRI. Nevertheless, LY294002 pretreatment eliminated the improvement of dexmedetomidine on spatial learning and memory ability. Furthermore, dexmedetomidine pretreatment reduced ratios of Beclin-1 and LC3II/LC3I and elevated p-Akt/Akt and p-mTOR/mTOR after CIRI. However, above effects of dexmedetomidine were partly reversed by LY294002. Overall, dexmedetomidine pretreatment exerted neuroprotection against CIRI in rats by attenuating autophagy via the PI3K/Akt/mTOR pathway. [ABSTRACT FROM AUTHOR]

AIM: To investigate the effects of Xiaozhong-Zhitong mixture （XZM） on the ulcer wounds in diabetic rats, as well as on phosphatidylinositol 3-kinase（ PI3K）/protein kinase B（ PKB/Akt）/mammalian target of rapamycin （mTOR） signaling pathway and endothelial cells in rats. METHODS: Sprague-Dawley （SD） rats were randomly divided into the following 5 groups with 10 rats per group: blank group, model group, XZM group, XZM+PI3K inhibitor （PI3Ki） group and XZM+mTOR inhibitor（ mTORi） group. The healing of foot ulcers in diabetic rats was evaluated. The pathological changes in the granulation tissues of rats were studied using HE staining. The levels of p-PI3K, p-Akt and p-mTOR in tissues were detected by immunohistochemistry. The changes in vascular endothelial cells were observed by transmission electron microscopy. The apoptotic cells from granulation tissues were analyzed by fluorescence TUNEL assay. RESULTS: The local areas of granulation tissues in model group exhibited different degrees of thickening, necrosis and structural loss in the epidermal layer, and internal collagen fiber hyperplasia and necrosis were seen in the dermis. The collagen fiber structure disappeared, accompanied by infiltration of a large number of inflammatory cells. The blood vessels of granulation tissue exhibited congestion and dilatation, and the endothelial cells exhibited disordered structure with severe swelling mitochondria （ridge fracture and dissolution） and more autophagy in the cytoplasm. Compared with model group, the ulcer wound area of rats in XZM group was significantly reduced, with numerous scabs formed on the surface of the granulation tissue. The epidermis showed varying degrees of thickening. The collagen fibers in the dermis were interlaced, without inflammatory cell infiltration, vascular congestion and dilatation in the dermis, or granulation tissue hemorrhage. A very small amount of autophagy was observed in the cells, and the levels of p-PI3K, p-Akt and p-mTOR proteins increased （P<0. 05）. Apoptosis rates in the granulation tissue of XZM, XZM+PI3Ki and XZM+mTORi groups were significantly decreased （P<0. 05）. The mitochondria of endothelial cells in XZM+PI3Ki and XZM+mTORi groups showed more swollen and had more autophagosomes compared with XZM group. The apoptosis rate of endothelial cells was higher than that in XZM group. The fibrous tissue in the dermis was uneven and exhibited a similar degree of hyperplasia. The stratified squamous epithelium was thin or missing. There was still a large amount of inflammatory cell infiltration in the dermal tissues of rats in XZM+mTORi group. In addition, the protein levels of p-Akt and p-mTOR in PI3K group and XZM+mTORi group were significantly higher than those in model group （P<0. 05）, and those in PI3K group and XZM+mTORi group were significantly lower than those in XZM group （P<0. 05）. CONCLUSION: Xiaozhong-Zhitong mixture could repair wounds in a rat model of diabetic ulcer, alleviate the inflammatory response, and promote the healing of diabetic ulcer wound, which may be related to the regulation of PI3K/Akt/mTOR signaling pathway, as well as the autophagy and apoptosis of vascular endothelial cells. [ABSTRACT FROM AUTHOR]

Dulaglutide is a new type of hypoglycemic agent that agonizes glucagon-like peptide-1 receptor (GLP-1RA). It can be concluded from previous studies that a GLP-1RA can reduce apoptosis and regulate autophagy in the nervous system, while related research on dulaglutide in vascular dementia (VD) has not been reported. In our study, the VD rat model was established by bilateral carotid artery occlusion, and the results of the Morris water maze test (MWM) and open-field test showed that the application of dulaglutide could effectively reduce the cognitive decline of VD rats without changing the behavior in the open-field test, which was used to assess an anxiety-like phenotype. We applied HE staining and immunofluorescence labeling to show that dulaglutide treatment significantly alleviated neuronal damage in the hippocampal region of VD rats, and reduced microglial and astrocyte proliferation. Western blot results showed that dulaglutide reduced VD-induced neuronal apoptosis (BCL2/BAX, c-caspase3) and autophagy (P62, LC3B, Beclin-1), and upregulated phosphorylation of PI3K/Akt/mTOR signaling pathway. KEGG pathway analysis of RNA-Sequence results showed that the differentially expressed genes in the dulaglutide treatment group were significantly enriched in the mTOR signaling pathway, and the repressor of mTOR, Deptor, was down-regulated. In conclusion, this study suggested that dulaglutide may alleviate learning and memory impairment and neuron damage in VD rats by attenuating apoptosis, regulating autophagy, and activating the PI3K/Akt/mTOR signaling pathway in neurons, which may make it a promising candidate for the simultaneous treatment of VD and diabetes. [ABSTRACT FROM AUTHOR]

Background: Osteoarthritis (OA) is a chronic degenerative joint disease characterized by cartilage degeneration and intra-articular inflammation. Daurisoline (DAS) is an isoquinoline alkaloid isolated from Rhizoma Menispermi, whose antitumor and anti-inflammatory pharmacological effects have been demonstrated, but the effects of DAS on OA have rarely been researched. In this study, we aimed to explore the potential role of DAS in OA and its partial mechanism. Materials and methods: The cytotoxicity of H2O2 and DAS toward chondrocytes was detected by the Cell Counting Kit-8 assay. Safranin O staining was used to detect chondrocyte phenotype changes. Cell apoptosis was measured by both flow cytometry and quantitative analysis of the protein levels of the apoptosis-related factors Bax, Bcl-2 and cleaved caspase-3 by western blot. Western blotting and immunofluorescence were used to assess the expression of the autophagy-related proteins LC3, Beclin-1 and p62. In addition, key signal pathway targets and matrix-degrading indicators were measured by western blot. Results: Our results indicated that H2O2 induced human chondrocyte apoptosis and activated autophagy in a dose-dependent manner. DAS treatment dose-dependently reversed the expression of apoptosis-related proteins (Bax, Bcl-2 and cleaved caspase3) and the apoptosis rate induced by H2O2. Western blot and immunofluorescence analyses showed that DAS decreased the H2O2-induced upregulation of the autophagy marker Beclin-1 and the LC3 II/LC3 I ratio and upregulated the p62 protein level. Mechanistically, DAS inhibited autophagy through the activation of the classical PI3K/AKT/mTOR signaling pathway and protected chondrocytes from apoptosis. In addition, DAS alleviated the H2O2-induced degradation of type II collagen and the high expression of matrix metalloproteinase 3 (MMP3) and MMP13. Conclusion: Our research demonstrated that DAS alleviated chondrocyte autophagy caused by H2O2 through activation of the PI3K/AKT/mTOR signaling pathway and protected chondrocytes from apoptosis and matrix degradation. In conclusion, these findings suggest that DAS may serve as a promising therapeutic strategy for OA. [ABSTRACT FROM AUTHOR]

Objective To investigate the effect and mechanism of astragalus polysaccharides (APS) on autophagy in colorectal cancer. Methods The effects of various concentrations of APS (0.25 g/L, 0.5 g/L, 0.75 g/L and 1 g/L) on the proliferation of human colorectal cancer HCT-116 cells were detected by CCK-8 test. The effect of APS on the migration of HCT-116 cells was detected by cell scratch assay. The effect of APS on autophagy of colorectal cancer cells was detected by dansylcadaverine staining technique. Effects of APS on autophagy and PI3K/Akt/mTOR signaling pathway related proteins LC3B, p62, p-PI3K, p-Akt, p-mTOR, PI3K, Akt and mTOR in HCT-116 cells and colorectal cancer tumor-bearing nude mice were detected by Western blot assay. The expression of autophagy-related protein p62 in colorectal cancer tumors was detected by immunohistochemistry. Results APS inhibited HCT-116 cell proliferation in a dose-dependent manner (P＜ 0.05). APS could inhibit the proliferation and migration of HCT-116 cells by inducing autophagy (P＜0.05). The autophagy inhibitor 3-MA (2 mmol/L) not only attenuated the autophagy induction effect of APS on HCT-116 cells, but also attenuated the inhibitory effect of APS on the proliferation and migration of HCT-116 cells (P＜0.05). APS could increase the expression ratio of autophagy-related protein LC3BⅡ/Ⅰ and reduce the expression level of p62 protein in HCT-116 cells and colorectal cancer mice (P＜0.05). 3-MA could attenuate the increasing effect of APS on the expression ratio of LC3BⅡ/Ⅰ protein and reduce the inhibitory effect of APS on the expression of p62 protein in HCT-116 cells (P＜0.05). APS could reduce the expression ratio of PI3K/Akt/mTOR signaling pathway related proteins p-PI3K/PI3K, p-Akt/Akt and p-mTOR/ mTOR in HCT-116 cells and colorectal cancer tumor (P＜0.05). The pathway agonist 740Y-P (10 µmol/L) could attenuate the inhibitory effect of APS on expression levels of PI3K/Akt/mTOR signaling pathway-related proteins in HCT-116 cells (P＜0.05). Conclusion APS can induce autophagy activation in colorectal cancer tumor tissue and cells, and inhibit cell proliferation and migration of HCT-116 cells by inducing autophagy. The mechanism of action may be related to the inhibition of PI3K/Akt/mTOR signaling pathway. [ABSTRACT FROM AUTHOR]