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New findings What is the central question of this study? The relevance of microRNA-22 (miR-22) has been indicated in coronary heart disease (CHD). How does it exert a protective role in CHD? What is the main finding and its importance? EZH2 inhibited transcription of the miR-22 promoter, thus modulating cell proliferation in human umbilical vein endothelial cells and vascular smooth muscle cells to induce CHD. Abstract MicroRNA-22 (miR-22) was indicated to modulate cell proliferation in human umbilical vein endothelial cells (HUVECs) under exposure to environmental toxicants. In the present study, we investigated the involvement of miR-22 in the mediation of HUVEC and vascular smooth muscle cell (VSMC) function, hence in the development of coronary heart disease (CHD). miR-22 expression was reduced in serum of CHD patients. Restoration of miR-22 decreased the proliferation, migration and invasion of VSMCs and increased apoptotic cells and inflammatory factors. In contrast, upregulation of miR-22 led to opposite trends in HUVECs. Chromatin immunoprecipitation and dual-luciferase assays validated that enhancer of zeste 2 polycomb repressive complex 2 subunit (EZH2) inhibited transcription of miR-22 promoter. EZH2, overexpressed in serum from CHD patients, diminished VSMC apoptosis, but facilitated HUVEC apoptosis. Luciferase reporter assays confirmed that thioredoxin-interacting protein (TXNIP) was a new direct target of miR-22. Overexpression of TXNIP blocked the function of miR-22 in HUVECs and VSMCs. Taken together, these findings will shed light on the role and mechanism of EZH2 in viability, migration, invasion and apoptosis via the miR-22/TXNIP axis in VSMCs and HUVECs, which might provide new insights into the treatment of CHD.

Leonurine (LEO) is pseudoalkaloid that has been isolated from motherwort. It has been found to have various biological activities, including an antioxidant capacity. This study aimed to confirm whether LEO could be used in porcine in vitro culture (IVC) medium for its antioxidant effect and related molecular mechanisms. The results showed that embryos in IVC medium supplemented with 40 μM LEO had an increased blastocyst formation rate, total cell number, and proliferation capacity and a low apoptosis rate. LEO supplementation decreased reactive oxygen species levels and increased glutathione levels. Moreover, LEO-treated embryos exhibited improved intracellular mitochondrial membrane potential and reduced autophagy. In addition, pluripotency related gene was up-regulated while apoptosis and autophagy related genes were down-regulated with LEO supplementation. These results suggest that LEO has a beneficial effect on pre-implantation embryo development by reducing oxidative stress and enhancing mitochondrial function.

The current experiment was performed to investigate whether luteolin affects the proliferation and apoptosis of keloid fibroblasts by regulating the expression of FRAT1 gene. Keloid fibroblasts were treated with luteolin at different concentrations. MTT method, western blot, flow cytometry, and real-time quantitative PCR (qPCR) were used to detect cell proliferation, cyclin D1 (CyclinD1), p21, B-cell lymphoma / leukemia-2 (Bcl-2), Bcl-2 related X protein (Bax), FRAT1 protein expression, apoptosis and ARHI mRNA expression. Keloid fibroblasts were transfected with si-FRAT1, or pcDNA-FRAT1 and treated with luteolin to observe their roles in cell proliferation and apoptosis. Compared with the control group, luteolin significantly reduced the keloid fibroblast activity, CyclinD1, Bcl-2, and FRAT1 protein levels, and obviously improved the cell apoptosis rate, p21 and Bax protein expression (P

B lymphocyte-induced maturation protein-1 (Blimp1) is a key regulator that promotes the terminal differentiation of mature B lymphocytes into plasma cells, and is essential for the survival of Multiple myeloma (MM)cells. However, the expression of Blimp1 in MM and its effect on the signaling pathway remain unknown. Studies have found that during long-term endoplasmic reticulum (ER) stress, activated ATF4 may also stimulate the CCAAT-enhancer-binding protein homologous protein (CHOP) gene, triggering the unfolded protein response (UPR) terminal apoptotic pathway in plasma cells. Moreover Aspirin can induce MM cell apoptosis through mitochondria and death receptor pathway. Therefore, we aim to explore whether Aspirin could induce AFT4/CHOP apoptosis pathway in MM by inhibiting Blimp1 expression, thereby promoting MM cell apoptosis and exerting anti-tumor effects.

Neochlorogenic acid (NCA), a natural compound found in honeysuckle, possesses prominent anti‑inflammatory and antitumor effects. Pingyangmycin (PYM) induces DNA damage and has been used for the treatment of oral and maxillofacial tumors. Oral care serves an important role in promoting wound healing during chemotherapy in patients with oral squamous cell carcinoma (OSCC). Therefore, the present study aimed to analyze the effects of NCA and PYM on OSCC cells and to investigate the potential underlying mechanism. Reverse transcription‑quantitative PCR and western blotting were conducted to analyze the expression levels of DNA topoisomerase II α (TOP2A) in different OSCC cell lines. TOP2A‑overexpression cells were constructed via transfection of TOP2A‑overexpression plasmids. Following NCA or PYM treatment, cell proliferation was assessed using Cell Counting Kit‑8 and colony formation assays, whereas cell apoptosis and the cell cycle distribution were assessed via TUNEL staining and flow cytometry, respectively. In addition, the expression levels of apoptosis‑ and cell cycle‑related proteins were detected via western blotting. Moreover, co‑immunoprecipitation (Co‑IP) was conducted to determine whether TOP2A interacted with CDK1. The results of the present study indicated that NCA treatment significantly enhanced the suppressive effects of PYM on OSCC cell proliferation and apoptosis. The results also indicated that PYM arrested the cell cycle in the G0/1 by regulating cyclin dependent kinase 1 (CDK1)/cyclin B1, which was enhanced by the cotreatment of NCA and PYM. In addition, NCA and PYA treatment altered the expression levels of apoptosis‑related proteins. The Co‑IP assay indicated that TOP2A interacted with CDK1. Moreover, TOP2A overexpression significantly reversed the effects of NCA and PYM treatment on OSCC cell proliferation and apoptosis. In addition, NCA significantly decreased PYM‑induced toxicity in normal oral epithelial cells. In conclusion, the results of the present study suggested that NCA may promote the inhibitory effects of PYM in OSCC via TOP2A.

Aim: Immunological checkpoint therapy is considered a powerful method for cancer therapy and acts by re-activating autologous T cells to kill the cancer cell. Myocarditis cases have been reported in cancer patients after immunological therapy, for example, nivolumab treatment is a monoclonal antibody that blocks programmed cell death-1/programmed cell death ligand-1 ligand interaction. This project provided insight into the inflammatory response as a benchmark to investigate the potential cardiotoxic effect of T cell response to the programmed cell death-1 (PD-1)/programmed cell death ligand-1 (PD-L1) axis in regulating cardiomyocyte injury in vitro. Methods and Results: We investigated cardiomyopathy resulted from the PD-1/PD-L1 axis blockade using the anti-PD-1 antibody in Rockefeller University embryonic stem cells-derived cardiomyocytes (RUES2-CMs) and a melanoma tumor-bearing murine model. We found that nivolumab alone did not induce inflammatory-related proteins, including PD-L1 expression, and did not induce apoptosis, which was contrary to doxorubicin, a cardiotoxic chemotherapy drug. However, nivolumab was able to exacerbate the immune response by increasing cytokine and inflammatory gene expression in RUES2-CMs when co-cultured with CD4+ T lymphocytes and induced apoptosis. This effect was not observed when RUES2-CMs were co-cultured with CD8+ T lymphocytes. The in vivo model showed that the heart function of tumor-bearing mice was decreased after treatment with anti-PD-1 antibody and demonstrated a dilated left ventricle histological examination. The dilated left ventricle was associated with an infiltration of CD4+ and CD8+ T lymphocytes into the myocardium. PD-L1 and inflammatory-associated gene expression were significantly increased in anti-PD-1-treated tumor-bearing mice. Cleaved caspase-3 and mouse plasma cardiac troponin I expressions were increased significantly. Conclusion: PD-L1 expression on cardiomyocytes suppressed T-cell function. Blockade of PD-1 by nivolumab enhanced cardiomyocyte inflammation and apoptosis through the enhancement of T-cell response towards cardiomyocytes.

To explore the expression of Blimp1, ATF4 and CHOP in bone marrow mononuclear cells from patients with multiple myeloma as well as the effect of aspirin on their expression.Sixty untreated patients with multiple myeloma and 30 patients with relatively normal bone marrow were selected. Mononuclear cells from the bone marrow fluid were separated using Ficoll separation solution. CD138Compared with plasma cells in normal group, the expression of Blimp1 mRNA in CD138Inhibition of Blimp1 expression in multiple myeloma cells can promote the expression of ATF4 and CHOP. Aspirin can inhibit the proliferation activity of myeloma cells by down-regulating Blimp1 expression in myeloma cells and up-regulating ATF4 and CHOP expression, therefore plays an anti-tumor rote.多发性骨髓瘤细胞中Blimp1、ATF4和CHOP的表达及阿司匹林对其表达的影响.探究多发性骨髓瘤（MM）患者骨髓单个核细胞中Blimp1、ATF4因子和CHOP蛋白的表达及阿司匹林对其表达的影响及意义.采用Ficoll分离液分离60例初诊未经治疗的多发性骨髓瘤患者和30例骨髓相对正常者骨髓液中的单个核细胞,免疫磁珠法分选CD138与相对正常组浆细胞比较，MM患者CD138抑制多发性骨髓瘤细胞中Blimp1的表达，可促进ATF4和CHOP的表达。阿司匹林可通过下调骨髓瘤细胞中Blimp1表达和上调ATF4和CHOP的表达抑制骨髓瘤细胞的增殖活性，发挥抗肿瘤活性.

The objective of this study was to synthesize the 9-/13-position substituted berberine derivatives and evaluate their cytotoxic and photocytotoxic effects against three human cancer cell lines. Among all the synthesized compounds, 9-O-dodecyl- (5e), 13-dodecyl- (6e), and 13-O-dodecyl-berberine (7e) exhibited stronger growth inhibition against three human cancer cell lines, (HepG2, HT-29 and BFTC905), in comparison with structurally related berberine (1). These three compounds also showed the photocytotoxicity in human cancer cells in a concentration-dependent and light dose-dependent manner. Through flow cytometry analysis, we found out a lipophilic group at the 9-/13-position of berberine may have facilitated its penetration into test cells and hence enhanced its photocytotoxicity on the human liver cancer cell HepG2. Further, in cell cycle analysis, 5e, 6e, and 7e induced HepG2 cells to arrest at the S phase and caused apoptosis upon irradiation. In addition, photodynamic treatment of berberine derivatives 5e, 6e, and 7e again showed a significant photocytotoxic effects on HepG2 cells, induced remarkable cell apoptosis, greatly increased intracellular ROS level, and the loss of mitochondrial membrane potential. These results over and again confirmed that berberine derivatives 5e, 6e, and 7e greatly enhanced photocytotoxicity. Taken together, the test data led us to conclude that berberine derivatives with a dodecyl group at the 9-/13-position could be great candidates for the anti-liver cancer medicines developments.

The objective of this study was to synthesize 9-/13-position substituted berberine derivatives and evaluated their cytotoxic and photocytotoxic effects against three human cancer cell lines. Among all the synthesized compounds, 9-O-dodecyl- (5e), 13-dodecyl- (6e) and 13-O-dodecyl-berberine (7e) exhibited stronger growth inhibition against three human cancer cell lines, (HepG2, HT-29 and BFTC905), in compare with structurally related berberine (1). These three compounds also showed the photocytotoxicity in human cancer cells in a concentration-dependent and light dose-dependent manner. Through flow cytometry analysis, we found out a lipophilic group at 9-/13-position of berberine may have facilitated its penetration into test cell and hence enhanced its photocytotoxicity on human liver cancer cell HepG2. Further, in cell cycle analysis, 5e, 6e and 7e induced HepG2 cells to arrest at S phase and caused apoptosis upon irradiation. In addition, photodynamic treatment of berberine (1) and its derivatives 5e, 6e and 7e again showed a significant photocytotoxic effects on HepG2 cells, induced remarkable cell apoptosis, greatly increased intracellular ROS level and the loss of mitochondrial membrane potential. These results over and again confirmed that berberine derivatives 5e, 6e and 7e greatly enhanced photocytotoxicity. Taking together, the test data led us to conclude that berberine derivatives with a dodecyl group at 9-/13-position could be great candidates for the anti-liver cancer medicines developments.

Skeletal muscle, a highly metabolic tissue, is particularly vulnerable to increased levels of saturated free fatty acids (FFAs). The role of autophagy in saturated FFAs-induced cellular senescence and insulin resistance in skeletal muscle remains unclear. Therefore, the present study was aimed to explore autophagic flux in cellular senescence and insulin resistance induced by palmitate in muscle cells, and whether resveratrol limited these responses. Our results showed that palmitate induced cellular senescence in both myoblasts and myotubes. In addition, palmitate delayed differentiation in myoblasts and inhibited expression of insulin-stimulated p-AKTSer473 in myotubes. The accumulations of autophagosome assessed by tandem fluorescent-tagged LC3 demonstrated that autophagic flux was impaired in both palmitate-treated myoblasts and myotubes. Resveratrol protected muscle cells from palmitate-induced cellular senescence, apoptosis during differentiation, and insulin resistance via ameliorating autophagic flux. The direct influence of autophagic flux on development of cellular senescence and insulin resistance was confirmed by blockage of autophagic flux with chloroquine. In conclusion, impairment of autophagic flux is crucial for palmitate-induced cellular senescence and insulin resistance in muscle cells. Restoring autophagic flux by resveratrol could be a promising approach to prevent cellular senescence and ameliorate insulin resistance in muscle. Keywords: Autophagic flux, Resveratrol, Cellular senescence, Insulin resistance

Colorectal cancer (CRC) is a common malignant gastrointestinal tumor with high mortality worldwide. Drug resistance and cytotoxicity to normal cells are the main causes of chemotherapeutic treatment failure in CRC. Therefore, extracting the bioactive compounds from natural products with anti-carcinogenic activity and minimal side-effects is a promising strategy against CRC. The present study aims to evaluate the anti-carcinogenic properties of avenanthramides (AVNs) extracted from oats bran and clarify the underlying molecular mechanisms. We demonstrated that AVNs treatment suppressed mitochondrial bioenergetic generation, resulting in mitochondrial swelling and increased reactive oxygen species (ROS) production. Further study indicated that AVNs treatment significantly reduced DDX3 expression, an oncogenic RNA helicase highly expressed in human CRC tissues. DDX3 overexpression reversed the ROS-mediated CRC apoptosis induced by AVNs. Of note, we identified Avenanthramide A (AVN A) as the effective ingredient in AVNs extracts. AVN A blocked the ATPase activity of DDX3 and induced its degradation by directly binding to the Arg287 and Arg294 residues in DDX3. In conclusion, these innovative findings highlight that AVNs extracts, in particular its bioactive compound AVN A may crack the current hurdles in the way of CRC treatment.

Background Increasing evidence has shown that mesenchymal stem cells (MSCs) yield a favorable therapeutic benefit for thermal burn skin wounds. Human amniotic MSCs (hAMSCs) derived from amniotic membrane have multilineage differentiation, immunosuppressive, and anti-inflammatory potential which makes them suitable for treating skin wounds. However, the exact effects of hAMSCs on the healing of thermal burn skin wounds and their potential mechanisms are not explored. Methods hAMSCs were isolated from amniotic membrane and characterized by RT-PCR, flow cytometry, immunofluorescence, and tumorigenicity test. We assessed the effects of hAMSCs and hAMSC conditional medium (CM) on wound healing in a deep second-degree burn injury model of mice. We then investigated the biological effects of hAMSCs and hAMSC-CM on the apoptosis and proliferation of heat stress-injured human keratinocytes HaCAT and dermal fibroblasts (DFL) both in vivo and in vitro. Next, we explored the underlying mechanisms by assessing PI3K/AKT and GSK3β/β-catenin signaling pathways in heat injured HaCAT and DFL cells after hAMSCs and hAMSC-CM treatments using PI3K inhibitor LY294002 and β-catenin inhibitor ICG001. Antibody array assay was used to identify the cytokines secreted by hAMSCs that may activate PI3K/AKT signaling pathway. Results Our results showed that hAMSCs expressed various markers of embryonic stem cells and mesenchymal stem cells and have low immunogenicity and no tumorigenicity. hAMSC and hAMSC-CM transplantation significantly promoted thermal burn wound healing by accelerating re-epithelialization with increased expression of CK19 and PCNA in vivo. hAMSCs and hAMSC-CM markedly inhibited heat stress-induced apoptosis in HaCAT and DFL cells in vitro through activation of PI3K/AKT signaling and promoted their proliferation by activating GSK3β/β-catenin signaling. Furthermore, we demonstrated that hAMSC-mediated activation of GSK3β/β-catenin signaling was dependent on PI3K/AKT signaling pathway. Antibody array assay showed that a panel of cytokines including PAI-1, C-GSF, periostin, and TIMP-1 delivered from hAMSCs may contribute to the improvement of the wound healing through activating PI3K/AKT signaling pathway. Conclusion Our results demonstrated that hAMSCs and hAMSC-CM efficiently cure heat stress-induced skin injury by inhibiting apoptosis of skin cells and promoting their proliferation through activating PI3K/AKT signaling pathway, suggesting that hAMSCs and hAMSC-CM may provide an alternative therapeutic approach for the treatment of skin injury. Electronic supplementary material The online version of this article (10.1186/s13287-019-1366-y) contains supplementary material, which is available to authorized users.

Sepsis-induced myocardial injury is a well-known cause of mortality. The cholinergic anti-inflammatory pathway (CHAIP) is a physiological mechanism by which the central nervous system regulates immune response through the vagus nerve and acetylcholine; the α7-nicotinic acetylcholine receptor (α7nAChR) is the main component of CHAIP; GTS-21, a synthetic α7nAChR selective agonist, has repeatedly shown its powerful anti-inflammatory effect. However, little is known about its effect on LPS-induced myocardial injury. We investigated the protective effects of GTS-21 on lipopolysaccharide (LPS)-induced cardiomyopathy via the cholinergic anti-inflammatory pathway in a mouse sepsis model. We constructed the model of myocardial injury in sepsis mice by C57BL/6 using LPS and determined the time of LPS treatment by hematoxylin-eosin (HE) and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL). C57BL/6 mice were randomized into five groups: blank control group, model group, α-bungarotoxin + LPS group, GTS-21 + LPS group, and α-bungarotoxin + GTS-21 + LPS group. The pathological results of myocardial tissue were detected by the HE method; the apoptosis rate was detected by the TUNEL method; the relative expressions of NF-κB p65, Caspase-3, Caspase-8, Bcl-2, Bax, p53, and a7nAChR were detected by real-time quantitative PCR (RT-PCR); and the protein expressions of IL-6, IL-1 β, TNF-α, and pSTAT3 were detected by western blot. The results showed that LPS-induced myocardial pathological and apoptosis changes were significant compared with the blank group, which was reversed by GTS-21; however, pretreatment with α-bungarotoxin obviously blocked the protective effect of GTS-21. NF-κB p65, Caspase-3, Caspase-8, Bax, p53, IL-6, IL-1β, TNF-α, and pSTAT3 were significantly increased in the model group, while a7nAChR and Bcl-2 were significantly decreased; GTS-21 treatment reversed that result, while pretreatment with α-bungarotoxin strengthened the result in the model. And pretreatment with α-bungarotoxin blocked the protective effect of GTS-21. GTS-21 can alleviate the LPS-induced damage in the heart via a7nAChR, and pretreatment with α-bungarotoxin obviously blocked the protective effect of GTS-21 on sepsis in mice.

A novel tumor-selective catalytic nanosystem that enables efficient chemodynamic therapy (CDT) and activatable fluorescence imaging in H2O2-rich tumor microenvironments has been developed.

5-Fluorouracil (5-FU)-based chemotherapy is the first-line recommended regimen in colorectal cancer (CRC), but resistance limits its clinical application. Andrographolide sulfonate, a traditional Chinese medicine, is mainly used to treat infectious diseases. In the present study, we reported that andrographolide sulfonate could significantly inhibit the growth of transplanted CT26 colon cancer in mice and improve survival when combined with 5-FU. Furthermore, TUNEL assay and immunohistochemistry analysis of proliferating cell nuclear antigen, Ki-67 and p-STAT3 confirmed that co-treatment could inhibit tumor proliferation and promote apoptosis. In tumor tissues of groups that received 5-FU and andrographolide sulfonate, CD4+ and CD8+ T cell infiltration was increased, and the expression of IFN-γ and Granzyme B detected by immunohistochemistry and qPCR was upregulated, reflecting improved antitumor immunity. Finally, we verified that 5-FU significantly activated the NLR Family Pyrin Domain Containing 3 (NLRP3) inflammasome in myeloid-derived suppressor cells (MDSCs) and that andrographolide sulfonate reversed this process to sensitize cells to 5-FU. In summary, andrographolide sulfonate synergistically enhanced antitumor effects and improved antitumor immunity by inhibiting 5-FU-induced NLRP3 activation in MDSCs. These findings provide a novel strategy to address 5-FU resistance in the treatment of CRC.

Smoking is identified as a risk factor for periodontitis. Carbon monoxide (CO)-releasing molecule-3 (CORM-3) is a compound that has demonstrated anti-inflammatory effects in vitro and in vivo studies. The present study aimed to investigate the effects of CORM-3 on the expression of inflammatory and osteoclastogenic cytokines in human periodontal ligament cells (PDLCs) stimulated by nicotine and lipopolysaccharide (LPS). The cells were pretreated with CORM-3 and then cultured in medium in the presence of nicotine and LPS. The mRNA and protein expression levels of prostaglandin E2 (PGE2), cyclooxygenase-2 (COX‑2), osteoprotegerin (OPG), receptor activator of nuclear factor-κB ligand (RANKL) and heme oxygenase-1 (HO-1) were evaluated using reverse transcription-quantitative polymerase chain reaction and western blot analysis. The mRNA and protein expression levels of these cytokines were also evaluated in PDLCs transiently transfected with HO-1 small interfering RNA (siRNA) in response to nicotine and LPS stimulation. CORM-3 attenuated the LPS- and nicotine-induced production of PGE2, COX-2 and RANKL in human PDLCs by releasing CO, and upregulated the expression of OPG. However, these effects of CORM-3 were abrogated when HO-1 siRNA was transiently transfected into the cells. These results demonstrate that CORM-3 exerts anti-inflammatory and anti-osteoclastogenic effects on nicotine- and LPS-stimulated human PDLCs via the HO-1 pathway, which suggests its promising potential for use in the treatment of inflammatory periodontal disease.

A natural ursolic compound, 2α,3β-dihydroxy-urs-12-en-28-oic acid (corosolic acid, CRA) was isolated from the root of Actinidia valvata Dunn. (A. valvata Radix). Since a large number of triterpenoid compound has marked anticancer effects toward various types of cancer cell lines in vitro, this study was carried out to investigate the anticancer effect of CRA in human gastric cancer cell line BGC823 cells and the underlying apoptotic mechanism of CRA was examined in BGC823 cell lines. The results showed that CRA significantly suppressed the viability of BGC823 cells in a concentration- and time-dependent manner. CRA also significantly increased the sub G1 population by cell cycle analysis in a concentration dependent manner. Exposure to CRA decreased p65, bcl-2, Fas, smac mRNA and protein expression, and increased IκBα, bax, survivin mRNA and protein expression. Results of immunofluorescence staining and EMSA further indicated CRA induced apoptosis by inhibiting nuclear translocation of nuclear factor NF-κB subunit p65. Consistently overall, our findings suggest that CRA induces apoptosis via inhibition of NF-κB (p65) expression level and activation of IκBα in BGC cells as a potent anticancer candidate for gastric cancer treatment.

Summary Although the TIM gene family plays important roles in immune responses, little is known about TIM regulation in the development of systemic lupus erythematosus (SLE). This study aimed to investigate the association of two TIM-4 single nucleotide polymorphisms (SNPs) rs6874202 (−1419G>A) and rs62382402 (−1609G>A) with SLE susceptibility in a Chinese Han population. The results showed no significant differences between patients with SLE and control group for rs6874202 and rs62382402 (p = .72, .53 respectively). However, the anti-dsDNA levels in serum from SLE patients with GG genotype of TIM-4 gene at -1419 site were significantly higher than those with GA and AA genotype (p = .0335), and C3 levels of SLE patients with GG and GA genotype were much lower than those with AA genotypes (p = .0187). Moreover, the apoptotic cell levels of SLE patients with AA and GG genotypes were significantly higher than those with GA genotypes in patients with SLE (p = .0393). In addition, the C3 concentration of SLE patients with the GG genotype of TIM-4 gene at -1609 site was found to be significantly higher than those with the GA genotype (p = .0129). The results imply that GG genotype of the TIM-4 gene at -1419 site might be associated with the disease activity of SLE.

Histone deacetylase (HDAC) inhibitors have been widely shown to result in cancer cell death. The present study investigated the mechanisms underlying the antitumor effects of the phytochemical trichostatin A (TSA), a classic pan-HDAC inhibitor, in 5,637 urinary bladder cancer cells. It was found that TSA caused cell cycle arrest at the G2/M and G1 phase accompanied by reduced expression of cyclin D1 and upregulated induction of p21. In addition, TSA induced morphological changes, reduced cell viability and apoptotic cell death in 5,637 cells through caspase-3 activation followed by PARP cleavage. The loss of mitochondrial membrane potential (MMP) indicated that TSA induced apoptosis in 5,637 cells through the intrinsic mitochondrial pathway. TSA significantly suppressed Akt activity at 12 h after treatment, suggesting that the apoptosis in the early phase was mediated by Akt inhibition. In addition, the protein level of transcription factor Sp1 was decreased at 24 h after TSA treatment, which likely led to the downregulation of survivin gene expression, and then contributed to the antitumor activity of TSA. Taken together, the present study delineated that TSA-induced growth inhibition and apoptosis in 5,637 cells was associated with pAKT inhibition and MMP loss at the early phase, followed by downregulation of Sp1 and survivin at the late phase of treatment.

5-FU is the first line therapy for colorectal cancer, however, treatment effect is often hampered by the development of drug resistance or toxicity at high doses. Andrographolide is a natural diterpenoid from Andrographis paniculata which has anti-bacterial, anti-antiviral and anti-inflammation activities. In the current study, we test the hypothesis that Andrographolide reverses 5-FU resistance in colorectal cancer and examine the underlying mechanism. In vitro and vivo studies indicated that Andrographolide treatment significantly re-sensitizes HCT116/5-FUR cells (HCT116 cells which are 5-FU resistant) to cytotoxicity of 5-FU. Mechanism analysis showed that Andrographolide/5-FU co-treatment elevated apoptosis level of HCT116/5-FUR cells with highly increased level of BAX. By using biotin-Andrographolide pull down and cellular thermal shift assay, we found out that Andrographolide can directly target to BAX. Andrographolide-BAX interaction prevented BAX degradation, enhancing mitochondria-mediated apoptosis thus reversed 5-FU resistance while BAX silence diminished this effect. Further, by analyzing patient samples who received 5-FU involved chemotherapy, we found that expression level of BAX is correlated with PFS. Our results here provide a novel combination treatment strategy, especially for patients with 5-FU-resistant tumors expressing low level of BAX. Meanwhile, we also proposed that BAX expression may be a predicted and prognosis marker of 5-FU involved chemotherapy.

Background Increasing evidence supports the anticancer effects of morin in vitro and in vivo . However, the role of morin-7-sulphate sodium (NaMoS), a water-soluble flavonoid derivative synthesized from morin remains unclear. The present study investigated the tumor suppression by NaMoS in mouse melanoma cells. Materials and methods We synthesized the flavonoid derivative morin-7-sulphate sodium according to the method described for quercetin-sulphate derivative, and further isolated, purified and identified the compound. Cell proliferation in vitro was assessed using a CCK-8 assay. The wound healing assay was performed to evaluate cell motility, and flow cytometry was used to detect cellular apoptosis. Protein levels of vimentin, matrix metalloproteinase 9 (MMP9), phosphorylation of Akt1/2/3 (p-Akt1/2/3), extracellular signal-regulated kinase 1/2 (p-ERK1/2) and Caspase3 in B16F10 cells were detected by immunohistochemistry and Western blot. Results The results suggest that cell proliferation was markedly decreased in NaMoS-treated groups (1, 10, 25, 50, 100, 500, 1000 μM) in a dose-dependent manner compared with the Control group and the IC 50 was 221.67 μM at 48 h. NaMoS at 200 μM concentration significantly inhibited the invasion and promoted apoptosis of B16F10 cells. Moreover, protein level of Caspase3 increased significantly in B16F10 cells treated by NaMoS. Immunohistochemistry and Western blot further confirmed that NaMoS decreased the expression of vimentin, MMP9, p-Akt1/2/3 and p-ERK1/2 in B16F10 cells. Conclusions This study provides robust evidence that NaMoS, a water-soluble flavonoid, manifests anticancer properties and may act as a signal transduction inhibitor in melanoma cells.

Osteoarthritis (OA) is a common degenerative joint disorder, which involves articular cartilage degeneration as well as joint inflammatory reactions. The recent studies have identified microRNA (miRNA) as one of the epigenetic mechanisms for the regulation of gene expression. Here we aim to reveal the role of miRNA in the regulation of gene expression in articular chondrocytes and its significance in the OA pathogenesis. In the present study, miRNA profiling was performed using OA cartilage and normal healthy cartilage tissues. As compared to their levels in normal cells and tissues, miR-27a expression was found to be upregulated in OA cartilage and IL-1β-treated articular chondrocytes. TUNEL staining, as well as flow cytometry with Annexin V-FITC/PI double labeling indicated that miR-27a inhibition reduced the apoptosis of IL-1β-treated articular chondrocytes. Bioinformatics prediction and the dual-luciferase reporter assay indicated that miR-27a targeted the 3'-UTR of the PI3K gene to silence it. The PI3K mRNA level in OA cartilage and IL-1β-treated articular chondrocytes was also downregulated, comparing with normal cells and tissues. Transfection of chondrocytes transfected with the miR-27a inhibitor upregulated the PI3K expression. This study demonstrated miR-27a is a regulator of the PI3K-Akt-mTOR axis in human chondrocytes and could participate in OA pathogenesis.

Epstein-Barr virus (EBV), the first human tumor virus to have been discovered, sustains an asymptomatic lifelong infection in ∼95% of the world's population. Reportedly, EBV infection induces the expression of specific cellular microRNAs (miRNAs), such as miR-155, miR-146a, miR-21, which can contribute to the persistence of latently infected cells. In this study, we investigated whether C-X-C chemokine receptor type 4 (CXCR4) is a cellular target of human miR-146a. We also investigated the role of miR-146a and CXCR4 in EBV-associated cells. The results indicate that miR-146a is more abundantly expressed in EBV-positive than in EBV-negative cells. MiR-146a down-regulated CXCR4 expression in a dose- and time-dependent manner. Phenotypic experiments detected miR-146a mimics that could suppress cell proliferation and cell migration and promote cell apoptosis by targeting CXCR4. In addition, miR-146a mimics suppressed cell survival by decreasing the population of G0/G1 phase cells. Latent membrane protein (LMP)1, an importance oncoprotein, can stimulate miR-146a and inhibit the CXCR4 expression. Our findings indicate that LMP1-miR-146a-CXCR4 axis functions as a regulator in the EBV-associated cells.

EBV-associated gastric carcinoma (EBVaGC) is a specific subgroup of gastric carcinoma, and the multifunctional transcriptional factor NF-κB may contribute to its tumorigenesis. In this study, we comprehensively characterized NF-κB signaling in EBVaGC using qRT-PCR, western blot, immunofluorescence assays, ELISA, and immunohistochemistry staining. NF-κB-signaling inhibitors may inhibit the growth of EBVaGC cells and induce significant apoptosis. IκBα is a key regulatory molecule, and repression of IκBα can contribute to aberrant NF-κB activation. Overexpression of LMP1 and LMP2A in the EBV-negative GC cell line SGC7901 could inhibit the expression of IκBα and induce NF-κB activation. These findings indicate that the canonical NF-κB signal is constitutively activated and plays an important role in EBVaGC tumorigenesis.

According to a report of the International Agency for Research on Cancer, arsenic and inorganic arsenic compounds are classified into Group 1 carcinogens with regard to human health. Epidemiological studies indicate that arsenic is one of the main risk factors for the development of bladder cancer. In the present study, arsenic‑altered gene expression in mouse bladder tissues and in human urothelial cells was compared. In the mouse model, sodium arsenite‑induced mouse urothelial hyperplasia and intracellular inclusions were present. Following DNA array analysis, four genes with differential expression were selected for quantitative real‑time PCR assay. The genes were the following: Cystathionine β‑synthase (CBS), adenosine A1 receptor (ADORA1), metastasis‑associated lung adenocarcinoma transcript 1 (MALAT1) and Wnt inhibitory factor 1 (Wif1). The results indicated a significant increase in the levels of Cbs and Adora1. The analysis of the DNA CpG methylation levels of the mouse Cbs and Adora1 genes revealed no significant change. In contrast to these observations, the four genes were further analyzed in the human normal urothelial cell line SV‑HUC1. The data indicated that WIF1 gene expression was decreased by sodium arsenite, whereas this was not noted for CBS, MALAT1 and ADORA1. Sodium arsenite decreased mRNA and protein expression levels of the WIF1 gene. In addition, the methylation levels of the WIF1 gene were increased. Sodium arsenite inhibited cell proliferation and promoted cell migration as demonstrated in cell functional assays. The gene status was compared in 8 human urothelial cell lines, and WIF1 mRNA expression levels were determined to be higher, whereas DNA CpG methylation levels were lower in SV‑HUC1 cells compared with those noted in the other 7 bladder cancer cell lines. In summary, the data indicated that sodium arsenite decreased WIF1 gene expression and promoted cell migration. The increased methylation levels of WIF1 DNA CpG could be a potential biomarker for bladder cancer.

Scope Oligonol has been shown to moderate mitochondrial biogenesis, protein synthesis, and protein degradation in diabetic mice in a previous study. It is therefore hypothesized that oligonol alleviated sarcopenia by regulating pathways involved in protein turnover and mitochondrial quality. Methods and results The 32-week-old senescence-accelerated mouse prone 8 (SAMP8) mice are fed with chow diet containing 200 mg kg-1 oligonol for 8 weeks. Oligonol supplementation increased skeletal muscle mass, cross-sectional areas, and grip strength in SAMP8 mice. Oligonol increased phosphorylation of AKT/mTOR/p70sk6, inhibited nuclear localization of FoxO3a and NFκB, and decreased transcription of MuRF-1 and MAFbx in skeletal muscle of SAMP8 mice. Downregulation of mitochondrial biogenesis genes (PGC-1α and Tfam) and mitochondrial fusion genes (Mfn2 and Opa1), loss of PINK1, overexpression of Atg13, LC3-II, and p62, and abundant accumulation of autophagosomes and lysosomes in skeletal muscle of SAMP8 mice are limited by oligonol. Furthermore, oligonol reduced expression of released cytochrome c and cleaved caspase-9 in skeletal muscle of SAMP8 mice. Conclusion Regulating pathways involved in protein synthesis and degradation, mitochondrial biogenesis, mitochondrial fusion/fission, autophagy, and mitochondria-dependent apoptosis by oligonol contribute to positive protein turnover and mitochondrial quality, thus increasing muscle mass and strength in SAMP8 mice.

The present study was performed to investigate the role of p38 mitogen‑activated protein kinase (MAPK) in the resorption of herniated intervertebral discs in 30 rats. In the non‑contained and p38 MAPK inhibition (p38i) groups, two coccygeal intervertebral discs (IVDs) were removed and wounded prior to relocation into the subcutaneous space of the skin of the back. In the contained group, the cartilage endplates maintained their integrity. Furthermore, SB203580 was injected intraperitoneally into the p38i group, whereas saline was injected into the other two groups. In the non‑contained group, the weight of the relocated IVDs decreased to a greater extent over time when compared with the contained and p38i groups. Phosphorylated p38, tumor necrosis factor‑α, and interleukin‑1β were observed to exhibit higher expression levels in the non‑contained group compared with the contained and p38i groups, at weeks 1 and 4 post‑surgery. The expression level of caspase‑3 and the densities of apoptotic disc cells were significantly higher in the non‑contained group compared with the contained and p38i groups at 4 weeks post‑surgery. In conclusion, p38 MAPK induces apoptosis in IVDs, while also accelerating the resorption of the relocated IVDs. Thus, p38 MAPK may be important in spontaneous resorption of IVDs.

// Chunxiao Liu 1,* , Yangjing Ye 1,* , Qian Zhou 1 , Ruijie Zhang 1 , Hai Zhang 1 , Wen Liu 1 , Chong Xu 1 , Lei Liu 2 , Shile Huang 2,3 and Long Chen 1 1 Jiangsu Key Laboratory for Molecular and Medical Biotechnology, Jiangsu Key Laboratory for Microbes and Functional Genomics, College of Life Sciences, Nanjing Normal University, Nanjing, PR China 2 Department of Biochemistry and Molecular Biology, Louisiana State University Health Sciences Center, Shreveport, LA, USA 3 Feist-Weiller Cancer Center, Louisiana State University Health Sciences Center, Shreveport, LA, USA * These authors have contributed equally to this work Correspondence to: Long Chen, email: // Shile Huang, email: // Keywords : rotenone, apoptosis, calcium ion, CaMKII, H2O2, mTOR, Pathology Received : October 11, 2015 Accepted : January 24, 2016 Published : February 03, 2016 Abstract Rotenone, a neurotoxic pesticide, induces loss of dopaminergic neurons related to Parkinson’s disease. Previous studies have shown that rotenone induces neuronal apoptosis partly by triggering hydrogen peroxide (H 2 O 2 )-dependent suppression of mTOR pathway. However, the underlying mechanism is not fully understood. Here, we show that rotenone elevates intracellular free calcium ion ([Ca 2+ ] i ) level, and activates CaMKII, resulting in inhibition of mTOR signaling and induction of neuronal apoptosis. Chelating [Ca 2+ ] i with BAPTA/AM, preventing extracellular Ca 2+ influx using EGTA, inhibiting CaMKII with KN93, or silencing CaMKII significantly attenuated rotenone-induced H 2 O 2 production, mTOR inhibition, and cell death. Interestingly, using TTFA, antimycin A, catalase or Mito-TEMPO, we found that rotenone-induced mitochondrial H 2 O 2 also in turn elevated [Ca 2+ ] i level, thereby stimulating CaMKII, leading to inhibition of mTOR pathway and induction of neuronal apoptosis. Expression of wild type mTOR or constitutively active S6K1, or silencing 4E-BP1 strengthened the inhibitory effects of catalase, Mito-TEMPO, BAPTA/AM or EGTA on rotenone-induced [Ca 2+ ] i elevation, CaMKII phosphorylation and neuronal apoptosis. Together, the results indicate that the crosstalk between Ca 2+ signaling and mitochondrial H 2 O 2 is required for rotenone inhibition of mTOR-mediated S6K1 and 4E-BP1 pathways. Our findings suggest that how to control over-elevation of intracellular Ca 2+ and overproduction of mitochondrial H 2 O 2 may be a new approach to deal with the neurotoxicity of rotenone.

Inactivated Sendai virus (hemagglutinating virus of Japan; HVJ) envelope (HVJ-E) induces anticancer immunity and cancer cell-selective apoptosis through the recognition of viral RNA genome fragments by retinoic acid-inducible gene-I (RIG-I). Here, we discovered that the "copy-back" type of defective-interfering (DI) particles that exist in the Cantell strain of HVJ induced the human PC3 prostate cancer cell death more effectively than the Sendai/52 strain or Cantell strain, which contain fewer DI particles. DI particle genomic RNA (~550 bases) activated proapoptotic genes such as Noxa and/or TNF-related apoptosis-inducing ligand (TRAIL) in human prostate cancer cells to induce cancer cell-selective apoptosis. DI particle-derived RNA was synthesized by in vitro transcription (in vitro transcribed (IVT)-B2). IVT-B2 RNA, which has a double-stranded region in its secondary structure, promoted a stronger anticancer effect than IVT-HN RNA, which does not have a double-stranded region in its secondary structure. The intratumoral transfection of IVT-B2 significantly reduced the volume of a human prostate tumor and induced tumor cell apoptosis in the xenograft mouse model. Moreover, the involvement of natural killer (NK) cells in IVT-B2-RNA-induced anticancer effects was also suggested. These findings provide a novel nucleic acid medicine for the treatment of cancer.

Epidemiologic studies show that the levels of air pollutants and particulate matter are positively associated with the morbidity and mortality of cardiovascular diseases. Here we demonstrate that the intratracheal instillation of multi-walled carbon nanotubes (MWCNTs), a standard fine particle, exacerbate doxorubicin (DOX)-induced cardiotoxicity in mice through altering gut microbiota and pulmonary and colonic macrophage phenotype. MWCNTs (25 μg/kg per day, 5 days a week for 3 weeks) promoted cardiotoxicity and apoptosis in the DOX (2 mg/kg, twice a week for 5 weeks)-treated C57BL/6 mice. MWCNTs exaggerated DOX-induced gut microbiota dysbiosis characterized by the increased abundances of Helicobacteraceae and Coriobacteriaceae. In addition, MWCNTs promoted DOX-induced M1-like polarization of colonic macrophages with an increase in TNF-α, IL-1β and CC chemokine ligand 2 in peripheral blood. Importantly, treatment with the antibiotics attenuated MWCNTs plus DOX-induced apoptosis of cardiomyocytes and M1-like polarization of colonic macrophages. The fecal microbiota transplantation demonstrated that MWCNTs exaggerated DOX-induced cardiotoxicity with M1-like polarization of colonic macrophages. The conditioned medium from MWCNTs-treated pulmonary macrophages promoted DOX-induced gut microbiota dysbiosis and colonic macrophage polarization. Furthermore, the co-culture of macrophages and fecal bacteria promoted M1-like macrophage polarization and their production of TNF-α and IL-1β, and thereby exacerbated the effects of MWCNTs. Moreover, IL-1β and TNF-α blockade, either alone or in combination attenuated MWCNTs-exacerbated cardiotoxicity. In summary, MWCNTs exacerbate DOX-induced cardiotoxicity in mice through gut microbiota and pulmonary and colonic macrophage interaction. Our findings identify a novel mechanism of action of inhaled particle-driven cardiotoxicity.

Most tumor cells take up more glucose than normal cells. Splicing dysregulation is one of the molecular hallmarks of cancer. However, the role of splicing factor in glucose metabolism and tumor development remains poorly defined. Here, we show that upon glucose intake, the splicing factor SRSF5 is specifically induced through Tip60-mediated acetylation on K125, which antagonizes Smurf1-mediated ubiquitylation. SRSF5 promotes the alternative splicing of CCAR1 to produce CCAR1S proteins, which promote tumor growth by enhancing glucose consumption and acetyl-CoA production. Conversely, upon glucose starvation, SRSF5 is deacetylated by HDAC1, and ubiquitylated by Smurf1 on the same lysine, resulting in proteasomal degradation of SRSF5. The CCAR1L proteins accumulate to promote apoptosis. Importantly, SRSF5 is hyperacetylated and upregulated in human lung cancers, which correlates with increased CCAR1S expression and tumor progression. Thus, SRSF5 responds to high glucose to promote cancer development, and SRSF5–CCAR1 axis may be valuable targets for cancer therapeutics., Changes in glucose metabolism can lead to tumor development, but the involvement of splicing factors is unclear. Here, the authors screened for SR proteins and identified SRSF5 stability is enhanced in response to glucose elevation to promote alternative splicing of CCAR1 which facilitates tumor growth.

Background Chemoresistance to temozolomide (TMZ) is a major challenge in the treatment of glioblastoma (GBM). We previously found that miR-519a functions as a tumor suppressor in glioma by targeting the signal transducer and activator of transcription 3 (STAT3)-mediated autophagy oncogenic pathway. Here, we investigated the effects of miR-519a on TMZ chemosensitivity and autophagy in GBM cells. Furthermore, the underlying molecular mechanisms and signaling pathways were explored. Methods In the present study, two stable TMZ-resistant GBM cell lines were successfully generated by exposure of parental cells to a gradually increasing TMZ concentration. After transfecting U87-MG/TMZ and U87-MG cells with miR-519a mimic or inhibitor, a series of biochemical assays such as MTT, apoptosis, and colony formation were performed to determine the chemosensitive response to TMZ. The autophagy levels in GBM cells were detected by transmission electron microscopy, LC3B protein immunofluorescence, and Western blotting analysis. Stable knockdown and overexpression of miR-519a in GBM cells were established using lentivirus. A xenograft nude mouse model and in situ brain model were used to examine the in vivo effects of miR-519a. Tumor tissue samples were collected from 48 patients with GBM and were used to assess the relationship between miR-519a and STAT3 expression. Results TMZ treatment significantly upregulated miR-519a in U87-MG cells but not in U87-MG/TMZ cells. Moreover, the expression of miR-519a and baseline autophagy levels was lower in U87-MG/TMZ cells as compared to U87-MG cells. miR-519a dramatically enhanced TMZ-induced autophagy and apoptotic cell death in U87-MG/TMZ cells, while inhibition of miR-519a promoted TMZ resistance and reduced TMZ-induced autophagy in U87-MG cells. Furthermore, miR-519a induced autophagy through modification of STAT3 expression. The in vivo results showed that miR-519a can enhance apoptosis and sensitized GBM to TMZ treatment by promoting autophagy and targeting the STAT3/Bcl-2/Beclin-1 pathway. In human GBM tissues, we found an inverse correlation between miR-519a and STAT3 expression. Conclusions Our results suggested that miR-519a increased the sensitivity of GBM cells to TMZ therapy. The positive effects of miR-519a may be mediated through autophagy. In addition, miR-519a overexpression can induce autophagy by inhibiting STAT3/Bcl-2 pathway. Therefore, a combination of miR-519a and TMZ may represent an effective therapeutic strategy in GBM. Electronic supplementary material The online version of this article (10.1186/s13045-018-0618-0) contains supplementary material, which is available to authorized users.

Epstein-Barr virus (EBV) is a ubiquitous oncogenic herpesvirus associated with various human tumors. Latent membrane protein (LMP) 1 is considered a key oncoprotein in some EBV-associated tumors, while it is absent in EBV-associated gastric carcinoma (EBVaGC). In the present study, we did not detect the mRNA or the protein expression of LMP1 in 10 EBVaGC tissue samples, but we found that miRNA-BART16 was more abundantly expressed in EBVaGC primary tissues than in cell lines. It has been reported that EBV-encoded miRNA-BART16 can target the 3'-untranslated regions of LMP1 and negatively regulate its expression. When a BART16 inhibitor or mimics were transfected into EBVaGC cell lines, both mRNA and protein levels were changed compared with those in controls. We further investigated the role of miRNA-BART16 in EBVaGC. According to the CCK8 assay, the proliferation of GT38 and GT39 cells increased after treatment with a miRNA-BART16 inhibitor. At the same time, the BART16 inhibitor caused increased accumulation of G2/M phase cells by bromodeoxyuridine and propidium iodide staining analysis. In addition, no obvious difference was observed in apoptosis after treatment with an inhibitor or mimics using annexin V staining. Our findings highlight the viral role of EBV-encoded miRNA-BART16 in regulating oncogenic LMP1 expression and a negative association with the proliferation of EBVaGC.

Medullary thyroid cancer (MTC) is a relatively rare thyroid cancer responsible for a substantial fraction of thyroid cancer mortality. More effective therapeutic drugs with low toxicity for MTC are urgently needed. Orphan nuclear receptor 4A1 (NR4A1) plays a pivotal role in regulating the proliferation and apoptosis of a variety of tumor cells. Based on the NR4A1 protein structure, 2-imino-6-methoxy-2H-chromene-3-carbothioamide (IMCA) was identified from the Specs compounds database using the protein structure-guided virtual screening approach. Computationally-based molecular modeling studies suggested that IMCA has a high affinity for the ligand binding pocket of NR4A1. MTT [3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide] and apoptosis assays demonstrated that IMCA resulted in significant thyroid cancer cell death. Immunofluorescence assays showed that IMCA induced NR4A1 translocation from the nucleus to the cytoplasm in thyroid cancer cell lines, which may be involved in the cell apoptotic process. In this study, the quantitative polymerase chain reaction results showed that the IMCA-induced upregulation of sestrin1 and sestrin2 was dose-dependent in thyroid cancer cell lines. Western blot showed that IMCA increased phosphorylation of adenosine 5′-monophosphate-activated protein kinase (AMPK) and decreased phosphorylation of ribosomal protein S6 kinase (p70S6K), which is the key enzyme in the mammalian target of rapamycin (mTOR) pathway. The experimental results suggest that IMCA is a drug candidate for MTC therapy and may work by increasing the nuclear export of NR4A1 to the cytoplasm and the tumor protein 53 (p53)-sestrins-AMPK-mTOR signaling pathway.

Background/Aims: Micro RNAs (miRNAs) play a very important role in myocardial ischemia/ reperfusion injury (MIRI), including in inflammation, apoptosis, and angiogenesis. Previous studies have demonstrated up-regulation of miR-327 in renal ischemia/reperfusion injury and MIRI. Via TargetScan, we found RP105 is a possible target gene of miR-327; our previous studies have also confirmed that RP105 acted as a cardioprotective protein in MIRI by reducing inflammation. However, the regulatory effect of miR-327 on RP105 has not previously been proposed. In our study, we aimed to identify the regulatory effect of miR-327 on RP105 protein in MIRI rats. Methods: Sixty male Sprague–Dawley rats were randomly divided into five groups, which were pre-treated with saline (sham and ischemia/reperfusion group), adenovirus-expressing miR-327-RNAi (Ad-miR-327-i group), control (Ad-NC group), or pri-miR-327 (Ad-miR-327 group) treatments. Three days later, the rat MIRI model was established by ischemia for 30 min, followed by reperfusion for 3 h. Myocardium and plasma were harvested and assessed. Results: miR-327 was increased by nearly 3-fold both in myocardium and plasma, which down-regulated RP105 in a 3′-untranslated region-dependent manner, and down-regulation of miR-327 via adenovirus transfection indirectly suppressed the TLR4/ TLR2-MyD88-NF-κB signaling axis activation via up-regulation of RP105, which subsequently resulted in reduced myocardial infarct size, attenuated cardiomyocyte destruction, and alleviated inflammation. In contrast, up-regulation of miR-327 induced the opposite effect. Conclusion: Down-regulation of miR-327 exerts a cardioprotective effect against MIRI by reducing inflammation, which may constitute a promising molecular therapeutic target for treating MIRI.

The nuclear receptor EcRB1, which is activated by the insect steroid hormone 20-hydroxyecdysone (20E), is reportedly phosphorylated by a protein kinase after 20E induction. However, the protein kinase has not been identified, and the significance of EcRB1 phosphorylation is unclear. In this study, we identified a protein kinase C δ (PKCδ) isoform (the E isoform) that phosphorylates EcRB1 in the lepidopteran Helicoverpa armigera, a serious agricultural pest worldwide, to promote apoptotic gene expression and apoptosis during metamorphosis. Through activation of the EcRB1/USP1 transcription complex by 20E, PKCδ expression was up-regulated in several tissues during the metamorphic stage. Knockdown of PKCδ caused failure to transition from larvae to pupae, prevented tissues from undergoing programmed cell death (PCD), and down-regulated the expression of the transcription factor Brz-7 and the apoptosis executors caspase-3 and caspase-6. The threonine residue at position 1343 of PKCδ was phosphorylated and was critical for its proapoptotic function. Overexpression of the PKCδ catalytic domain was localized to the nuclei in HaEpi cells, which increased caspase-3 activity and apoptosis. PKCδ directly phosphorylated a threonine residue at position 468 in the amino acid sequence of EcRB1. The phosphorylation of EcRB1 was critical for its heterodimeric interaction with the USP1 protein and for binding to the ecdysone response element. The data suggested that 20E up-regulates PKCδ expression to regulate EcRB1 phosphorylation for EcRB1/USP1 transcription complex formation, apoptotic gene transcription, and apoptosis.

Eleven novel naftopidil-related compounds that contain amide and indole groups were designed and synthesized. The biological effects of these compounds on three α1-adrenoceptor subtypes and cancerous human prostate cell lines (PC-3, DU-145, and LNCaP) were determined. Compounds 2, 3, 5, 11, and 12 exhibited an α1-adrenoceptor antagonistic activity, whereas compounds 9, 10, and 12 displayed moderate antiproliferative activities. Compound 3 exhibited a significant α(1D/1A) blocking activity in isolated rat tissues (97.7- and 64.6-fold selective for α(1D) and α(1A) compared with α(1B)) but not a relevant cytotoxic activity. Compound 12 demonstrated a potent and selective α(1D/1A) antagonistic activity (47.9- and 19.1-fold for α(1D) and α(1A) compared with α1B) and a potent antiproliferative activity in PC-3 cells (IC50 = 15.70 μM). Further testing confirmed that compound 12 inhibited the growth of PC-3 cells by inducing apoptosis and G0/G1 cell cycle arrest, which was mediated by α1-adrenoceptor. Therefore, compound 12 is a potential multipotent agent that can act as an effective α1-adrenoceptor subtype antagonist for treating benign prostatic hyperplasia and a preventive medication against human prostate cancer.

Hydrogen sulfide (H(2)S) has recently been identified as an endogenous gaseous signaling molecule. In the vascular system, the formation of H(2)S is catalyzed by cystathionine γ‑lyase (CSE). Previous studies have demonstrated the protective effects of H(2)S on ischemic injury in various types of tissue. However,, little is known about the role of H(2)S in diabetes-associated vascular diseases. Thus, the aim of the present study was to examine the possible role of H(2)S in high glucose-induced vascular dysfunction, and to explore the underlying mechanisms. Human umbilical vein endothelial cells (HUVECs) were isolated from human umbilical veins. The levels of H(2)S following treatment with various levels of glucose were determined and the secretion of endothelin-1 (ET-1) was measured by ELISA. The mRNA and protein expression of CSE in the HUVECs was determined by real-time RT-PCR and western blot analysis, respectively. Treatment with high glucose (25 mmol/l) for 48 h significantly increased the secretion of ET-1 by HUVECs, with the concomitant suppression of H(2)S production and CSE protein expression. The increase in exogenous H(2)S levels through the administration of sodium hydrosulfide (NaHS) attenuated the high glucose-induced downregulation of CSE protein expression, and significantly inhibited the secretion of ET-1. These results suggest that the downregulation of CSE protein expression and the subsequent decrease in H(2)S production play a role in high glucose-induced vascular dysfunction possibly by increasing the secretion of ET-1 by endothelial cells.

Rotenone, a common pesticide and inhibitor of mitochondrial complex I, induces loss of dopaminergic neurons and consequential aspects of Parkinson’s disease (PD). However, the exact mechanism of rotenone neurotoxicity is not fully elucidated. Here, we show that rotenone induced reactive oxygen species (ROS), leading to apoptotic cell death in PC12 cells and primary neurons. Pretreatment with catalase (CAT), a hydrogen peroxide-scavenging enzyme, attenuated rotenone-induced ROS and neuronal apoptosis, implying hydrogen peroxide (H2O2) involved, which was further verified by imaging intracellular H2O2 using a peroxide-selective probe H2DCFDA. Using thenoyltrifluoroacetone (TTFA), antimycin A, or Mito-TEMPO, we further demonstrated rotenone-induced mitochondrial H2O2-dependent neuronal apoptosis. Rotenone dramatically inhibited mTOR-mediated phosphorylation of S6K1 and 4E-BP1, which was also attenuated by CAT in the neuronal cells. Of interest, ectopic expression of wild-type mTOR or constitutively active S6K1, or downregulation of 4E-BP1 partially prevented rotenone-induced H2O2 and cell apoptosis. Furthermore, we noticed that rotenone-induced H2O2 was linked to the activation of caspase-3 pathway. This was evidenced by the finding that pretreatment with CAT partially blocked rotenone-induced cleavages of caspase-3 and poly (ADP-ribose) polymerase. Of note, zVAD-fmk, a pan caspase inhibitor, only partially prevented rotenone-induced apoptosis in PC12 cells and primary neurons. Expression of mTOR-wt, S6K1-ca, or silencing 4E-BP1 potentiated zVAD-fmk protection against rotenone-induced apoptosis in the cells. The results indicate that rotenone induction of H2O2 inhibits mTOR-mediated S6K1 and 4E-BP1/eIF4E pathways, resulting in caspase-dependent and -independent apoptosis in neuronal cells. Our findings suggest that rotenone-induced neuronal loss in PD may be prevented by activating mTOR signaling and/or administering antioxidants.

Oxidative stress is an important molecular mechanism underlying lung fibrosis. The mitochondrion is a major organelle for oxidative stress in cells. Therefore, blocking the mitochondrial signalling pathway may be the best therapeutic manoeuver to ameliorate lung fibrosis. Astaxanthin (AST) is an excellent antioxidant, but no study has addressed the pathway of AST against pulmonary oxidative stress and free radicals by the mitochondrion-mediated signalling pathway. In this study, we investigated the antioxidative effects of AST against H2 O2 - or bleomycin (BLM)-induced mitochondrial dysfunction and reactive oxygen species (ROS) production in alveolar epithelial cells type II (AECs-II) in vivo and in vitro. Our data show that AST blocks H2 O2 - or BLM-induced ROS generation and dose-dependent apoptosis in AECs-II, as characterized by changes in cell and mitochondria morphology, translocation of apoptotic proteins, inhibition of cytochrome c (Cyt c) release, and the activation of caspase-9, caspase-3, Nrf-2 and other cytoprotective genes. These data suggest that AST inhibits apoptosis in AECs-II cells through the ROS-dependent mitochondrial signalling pathway and may be of potential therapeutic value in lung fibrosis treatment.

The cardioprotection of the immature heart during cardiac surgery remains controversial due to the differences between the adult heart and the newborn heart. Lysophosphatidic acid (LPA) is a small bioactive molecule with diverse functions including cell proliferation and survival via its receptor: LPA1–LPA6. We previously reported that the expressions of LPA1 and LPA3 in rat hearts were much higher in immature hearts and then declined rapidly with age. In this study, we aimed to investigate whether LPA signaling plays a potential protective role in immature hearts which had experienced ischemia/reperfusion (I/R) injury. The results showed that in Langendorff-perfused immature rat hearts (2 weeks), compared to I/R group, LPA pretreatment significantly enhanced the cardiac function, attenuated myocardial infarct size and CK-MB release, decreased myocardial apoptosis and increased the expression of pro-survival signaling molecules. All these effects could be abolished by Ki16425, an antagonist to LPA1 and LPA3. Similarly, LPA pretreatment protected H9C2 from hypoxia-reoxygenation (H/R) induced apoptosis and necrosis in vitro. The mechanisms underlying the anti-apoptosis effects were related to activation of the phosphatidylinositol 3-kinase (PI3K)/protein kinas B (AKT) signaling pathways as well as phosphorylation of the downstream effector of AKT, glycogen synthase kinase 3 beta (GSK3β), through LPA1 and/or LPA3. What's more, we found that LPA preconditioning increased glucose uptake of H9C2 subjected to H/R by the activation of AMP-Activated Protein Kinase (AMPK) but not the translocation of GLUT4. In conclusion, our study indicates that LPA is a potent survival factor for immature hearts against I/R injuries and has the potential therapeutic function as a cardioplegia additive for infantile cardiac surgery.

The p38 pathway, which is important in mitogen-activated protein kinase (MAPK) family protein signaling, leads to mitochondrial dysfunction and activation of caspase-3. B-cell lymphoma 2 (Bcl‑2) family members are involved in the regulation of activities associated with the survival and death of neurons through apoptosis and have important functions in most types of apoptosis. In the present study, the effects of the p38 MAPK inhibitor SB203580 on N-methyl-d-aspartate (NMDA)-induced cerebral cortical neuron apoptosis were observed to further analyze the possible mechanisms of NMDA-induced neuronal death. Cultured primary cortical neurons were randomly divided into five groups: A control group, NMDA group and three SB203580 interventional groups. The lactate dehydrogenase (LDH) and MTT assays were employed to investigate the effects of the drugs on apoptosis. The morphology of apoptotic cells was observed using acridine orange/ethidium bromide (AO/EB) fluorescence staining. The expression levels of phospho‑(p)‑p38MAPK, Bcl‑2 and Bcl-2-associated X (Bax) were assessed by immunohistochemical methods and western blot analysis to investigate the possible underlying protective mechanisms. The cell viability markedly decreased following incubation with NMDA. The protein levels of cell death repressor Bcl-2 and the levels of Bcl-2/Bax were downregulated. The protein levels of p‑p38MAPK and cell death promoter Bax increased significantly in cells with NMDA treatment. However, these changes were inhibited by SB203580 treatment, particularly in the high‑dose group. Neuronal death induced by NMDA in primary cortical neurons was caused in part by apoptosis, which was mediated through the activation of the p38 signaling pathway by NMDA. SB203580 has neuroprotective effects against NMDA‑induced apoptosis.

Hinokitiol is found in the heartwood of cupressaceous plants and possesses several biological activities. Hinokitiol may play an important role in anti-inflammation and antioxidant processes, making it potentially useful in therapies for inflammatory-mediated disease. Previously, the suppression of tumor growth by hinokitiol has been shown to occur through apoptosis. Programmed cell death can also occur through autophagy, but the mechanism of hinokitiol-induced autophagy in tumor cells is poorly defined. We used an autophagy inhibitor (3-methyladenine) to demonstrate that hinokitiol can induce cell death via an autophagic pathway. Further, we suggest that hinokitiol induces autophagy in a dose-dependent manner. Markers of autophagy were increased after tumor cells were treated with hinokitiol. In addition, immunoblotting revealed that the levels of phosphoprotein kinase B (P-AKT), phosphomammalian target of rapamycin (P-mTOR), and phospho-p70 ribosomal s6 kinase (P-p70S6K) in tumor cells were decreased after hinokitiol treatment. In conclusion, our results indicate that hinokitiol induces the autophagic signaling pathway via downregulation of the AKT/mTOR pathway. Therefore, our findings show that hinokitiol may control tumor growth by inducing autophagic signaling.

Radiation-induced lung injury (RILI) is a potentially fatal and dose-limiting complication of thoracic radiotherapy. This study was to investigate the protective effects of grape seed pro-anthocyanidins (GSPs), an efficient antioxidant and anti-carcinogenic agent, on RILI. In our study, it was demonstrated that acute and late RILI was ameliorated after GSPs treatment possibly through suppressing TGF-β1/Smad3/Snail signalling pathway and modulating the levels of cytokines (interferon-γ, IL-4 and IL-13) derived from Th1/Th2 cells. In addition, a sustained high level of PGE2 was also maintained by GSPs treatment to limited fibroblast functions. As shown by electron spin resonance spectrometry, GSPs could scavenge hydroxyl radical (•OH) in a dose-dependent manner, which might account for the mitigation of lipid peroxidation and consequent apoptosis of lung cells. In vitro, GSPs radiosensitized lung cancer cell A549 while mitigating radiation injury on normal alveolar epithelial cell RLE-6TN. In conclusion, the results showed that GSPs protects mice from RILI through scavenging free radicals and modulating RILI-associated cytokines, suggesting GSPs as a novel protective agent in RILI.

Recent studies showed that hydrogen can be used as an effective radioprotective agent through scavenging free radicals. This study was undertaken to evaluate the radioprotective effects of hydrogen on immune system in mice. H(2) was dissolved in physiological saline using an apparatus produced by our department. Spleen index and histological analysis were used to evaluate the splenic structural damage. Spleen superoxide dismutase, GSH, MDA were measured to appraise the antioxidant capacity and a DCF assay for the measurement of radical oxygen species. Cell apoptosis was evaluated by an Annexin V-FITC and propidium iodide staining method as well as the apoptotic proteins such as Bcl-2, Bax, caspase-3 and c-caspase-3. CD4+ and CD8+ T cells subtypes were detected by flow cytometry with FITC-labelled antimouse CD4 and PE antimouse CD8 staining. Real-time PCR was utilized to determine the CD4+ T cell subtypes and related cytokines. Our study demonstrated that pre-treatment with H(2) could increase the spleen index and attenuate the radiation damage on splenic structure. Radical oxygen species level was also reduced by H(2) treatment. H(2) also inhibited radiation-induced apoptosis in splenocytes and down-regulated pro-apoptotic proteins in living mice. Radiation-induced imbalance of T cells was attenuated by H(2). Finally, we found that H(2) could regulate the polarization of CD4+ T cells and the level of related cytokines. This study suggests H(2) as an effective radioprotective agent on immune system by scavenging reactive oxygen species.

Worldwide, cervix carcinoma is among the most dangerous cancer types, and novel therapies are under development. Cancer treatments are often hampered because of lack of specificity. The chicken anemia virus-derived apoptin induces apoptosis selectively in tumor cells and leaves normal cells unharmed. Here, we have carried out in-vitro and in-vivo studies on the cytotoxic effect of apoptin in a cervix carcinoma model. Apoptin was fused to the protein transduction domain 4 (PTD4), enabling delivery of the fusion protein across cellular membranes. PTD4-apoptin protein is located in the nuclei of human cervical carcinoma HeLa cells and in the cytoplasm of normal cells L02. By MTT and flow cytometry analysis, we have proven that PTD4-apoptin protein induced apoptosis in the cervical carcinoma cells. PTD4-apoptin enhanced the level of active executioner caspase-3. Neither caspase-3 activation nor apoptin-induced accumulation of the mitochondrial outer-membrane protein Mfn-2 was affected by ectopic Bcl-2 expression. In contrast, apoptin-mediated AKT activation was inhibited by Bcl-2. In vivo, cervix carcinoma xenografts were treated for 7 days with PTD4-apoptin protein. The PTD4-apoptin treatment induced a decrease in the cervix carcinoma, whereas the PTD4-GFP protein-treated controls expanded significantly. TUNEL analysis showed that PTD4-apoptin protein induced apoptosis in cervix carcinoma cells, in contrast to the control PTD-GFP-treated ones. Our results indicate that apoptin is a potential anticancer agent for treating cervix carcinoma.

Magnolol, a hydroxylated biphenol compound isolated from the bark of Magnolia officinalis, has been shown to exhibit anti-proliferative effect in various cancer cells, including skin cancer cells. Methoxylation of magnolol appears to improve its anti-inflammatory activity, yet the effect of this modification on the agent’s antitumor activity remains unknown. In this work, we report that 2-O-methylmagnolol (MM1) displays improved antitumor activity against skin cancer cells compared to magnolol both in vitro and in vivo. The increased antitumor activity of MM1 appears to correlate with its increased ability to induce apoptosis. DNA microarray and network pathway analyses suggest that MM1 affects certain key factors involved in regulating apoptosis and programmed cell death. Interestingly, the level of the long non-coding (lnc) RNA of growth arrest-specific 5 (GAS5) was increased in MM1-treated cells, and inhibition of lncRNA GAS5 inhibited MM1-induced apoptosis. Conversely, overexpression of lncRNA GAS5 inhibited cell proliferation and promoted cell apoptosis in skin cancer cells. The expression of lncRNA GAS5 in the skin cancer tissues was found to be lower than that in the adjacent normal tissues in a majority of patients. Taken together, our findings suggest that MM1 has improved antitumor activity in skin cancer cells, and that this is due, at least in part, to the upregulation of lncRNA GAS5 and the enhancement of apoptosis.

Thyroid cancer is the most common endocrine malignancy, the global incidence rate of which is rapidly rising. Surgery and radioiodine therapies are common and effective treatments only for nonmetastasized primary tumors. Therefore, effective treatment modalities are imperative for patients with radioiodine-resistant thyroid cancer. Honokiol, a biophenolic compound derived from Magnolia spp., has been shown have diverse biological and pharmacological activities, including anti-inflammatory, antioxidative, antiangiogenic, and anticancer properties. In the present study, three human thyroid cancer cell lines, namely anaplastic, follicular, and poorly differentiated thyroid cancer cells, were used to evaluate the chemotherapeutic activity of honokiol. Cell viability, cell cycle, apoptosis, and autophagy induction were determined through flow cytometry and western blot analysis. We found that honokiol treatment can suppress cell growth, induce cell cycle arrest, and enhance the induction of caspase-dependent apoptosis and autophagy in cancer cells. Moreover, honokiol treatment modulated signaling pathways including Akt/mTOR, ERK, JNK, and p38 in the studied cells. In addition, the antitumorigenic activity of honokiol was also confirmed in vitro and in vivo. Our data provide evidence that honokiol has a unique application in chemotherapy for human thyroid cancers.