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The potential of adult adipose-derived stromal cells (ADSCs) to differentiate into astrocytes holds promise for future cell transplantation therapies. However, the growth of differentiated astrocytes is unstable, and their survival rate is low. Endoplasmic reticulum (ER) pathway mediated apoptosis is one of the causes of cell death, but whether there is ER stress response in the differentiation of ADSCs into astrocytes is still unclear. In this study, the expression of protein factors related to endoplasmic reticulum stress (ERS) and apoptosis, including GRP78, ATF6, PERK, CHOP, Caspase12, and Caspase3, was detected in cells. It was found that the expression of ERS pro-survival factors was highest in the ADSCs group and decreased with prolonged induction time. Conversely, the expression levels of pro-apoptotic factors increased with the extension of induction time. Thus, ERS occurs during the differentiation of ADSCs into astrocytes, and ERS can mediate apoptosis of ADSC-derived astrocytes.

Purpose: Diabetic cataract (DC) is a major cause of blindness worldwide. Prion protein (PRNP) was proved to be up-regulated and hypomethylated in DC samples. Here, we investigated whether PRNP was involved in DC progression in N6-methyladenosine (m6A)-dependent manner, and its potential mechanisms., Methods: Levels of genes and proteins were assayed using qRT-PCR and western blotting. Cell proliferation and apoptosis were determined using Cell Counting Kit-8 assay, 5-thynyl-2'-deoxyuridine (EdU) assay, and flow cytometry, respectively. Oxidative stress was analyzed by measuring the production of glutathione peroxidase (GSH-PX), superoxide dismutase (SOD), and malondialdehyde (MDA). The m6A modification was determined by RNA immunoprecipitation (Me-RIP) assay. The interaction between RBM15 (RNA binding motif protein 15) and PRNP was probed using RIP assay., Results: PRNP was highly expressed in DC patients and HG-induced HLECs. Functionally, PRNP deficiency reversed HG-induced apoptosis and oxidative stress in HLECs. Mechanistically, RBM15 induced PRNP m6A modification and directly bound to PRNP. Knockdown of RBM15 abolished HG-induced apoptotic and oxidative injury in HLECs, while these effects were rescued after PRNP overexpression., Conclusion: RBM15 silencing suppressed HG-induced lens epithelial cell injury by regulating PRNP in an m6A-mediated manner, hinting a novel therapeutic strategy for DC patients.

MicroRNAs (miRNAs) have been confirmed to play pivotal roles in hepatocellular carcinoma (HCC) carcinogenesis. However, the underlying function of microRNA-33b (miR-33b) in HCC remains unclear. Here, we found that miR-33b level was significantly reduced in both HCC tissues and tumor cell lines. Further, luciferase reporter assay and western blot analysis confirmed that Friend leukemia virus integration 1 (Fli-1) was a direct target of miR-33b. Overexpression of miR-33b dramatically suppressed HCC tumor cell proliferation and cell mobility, but facilitated tumor cell apoptosis in vitro. Besides, restoration of Fli-1 partially attenuated miR-33b-mediated inhibition of cell growth and metastasis via activating Notch1 signaling and its downstream effectors. Our findings demonstrate the important role of miR-33b/Fli-1 axis in HCC progression and provide novel therapeutic candidates for HCC clinical treatment., (© 2020 Wiley Periodicals, Inc.)

This investigation was intended to elucidate whether long noncoding RNA (lncRNA)-activated by transforming growth factor-β (ATB) interacting with miR-200c could mediate colorectal cancer (CRC) progression, offering potential strategies for diagnosing and treating CRC. Here totally 315 patients with CRC were recruited, and their CRC tissues and adjacent normal tissues were gathered. Concurrently, four colon cancer cell lines (ie, SW620, Lovo, HCT116, and SW480) and the human colon mucosal epithelial cell line (NCM460) were also purchased. Moreover, si-ATB, si-NC, miR-200c mimic, miR-200c inhibitor, and miR-NC were prepared for transfection into the CRC cells, and their effects on CRC cell lines were evaluated based on the conduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, colony formation assay, and flow cytometry assay. Eventually, the Luciferase reporter gene assay was carried out to judge if there existed a targeted relationship between ATB and miR-200c. The results of Cox regression analyses suggested that overexpressed lncRNA ATB, underexpressed miR-200c, poor tumor differentiation, lymph-vascular invasion, and perineural invasion were symbolic of shortened survival of the patients with CRC (all P < .05). Besides, transfection of pcDNA3.1-ATB and miR-200c inhibitor could boost the viability and proliferation of Lovo and SW620 cell lines (all P < .05). Meanwhile, the expressions of p53 and p21 were also reduced under treatments of pcDNA3.1-ATB and miR-200c inhibitor (P < .05). In addition, CDK2 seemed to reverse the contribution of miR-200c to intensifying viability and proliferation of Lovo and SW420 cell lines (P < .05). Furthermore, ATB might downregulate miR-200c expression by targeting it (P < .05), and CDK2 was subjected to dual regulation of both ATB and miR-200c (P < .05). In conclusion, the lncRNA ATB/miR-200c/CDK2 signaling was responsible for intensified proliferation and prohibited apoptosis of CRC cells, which might provide effective approaches for diagnosing and treating CRC., (© 2019 Wiley Periodicals, Inc.)

Polyphyllin has been reported to exhibit anticancer effects against various types of cancer via the proapoptotic signaling pathway. The aim of the present study was to investigate the role of the endoplasmic reticulum stress and death receptor signaling pathways in PPI-induced apoptosis of human hepatocellular carcinoma HepG2 cells. Analysis demonstrated that PPI could significantly inhibit the proliferation and induce apoptosis of HepG2 cells in a dose- and time-dependent manner. Investigation into the molecular mechanism of PPI indicated that PPI notably mediated ER stress activation via IRE-1 overexpression and activation of the caspase-12 to protect HepG2 cells against apoptosis. In addition, PPI markedly induced the expression of death receptors signaling pathways-associated factors, including tumor necrosis factor (TNF) receptor 1/TNF- α and FAS/FASL. Additionally, suppression of the death receptor signaling pathways with a caspase-8 inhibitor, Z-IETD-FMK, revealed an increase in the death rate and apoptotic rate of HepG2 cells. Collectively, the findings of the present study suggested that the ER stress and death receptor signaling pathways were associated with PPI-induced HepG2 cell apoptosis; however, endoplasmic reticulum stress may serve a protective role in this process. The combination of PPI and Z-IETD-FMK may activate necroptosis, which enhances apoptosis. Therefore, the results of the present study may improve understanding regarding the roles of signaling pathways in PPI regulated apoptosis and contribute to the development of novel therapies for the treatment of HCC.

Osteosarcoma is the most common primary malignant tumor of the bone found predominantly in children and teenagers and results in early metastasis and poor prognosis. The present study primarily focused on the impact of celastrol on apoptosis and autophagy of osteosarcoma HOS cells, as well as the related mechanisms. Following the appropriate treatment, the human osteosarcoma cell line HOS was assessed for viability, Ca2+ in cells, apoptosis and changes in cell morphology using Cell Counting Kit-8, flow cytometry, inverted phase contrast microscope, Hoechst staining and transmission electron microscopy. The expression levels of various proteins, including endoplasmic reticulum stress (ERS)-related proteins (Bip, PERK, p-PERK, IRE1α, calnexin, PDI and Erol‑Lα), apoptosis-related proteins (CHOP, cleaved caspase‑12), mitochondrial apoptosis-related proteins (Bax, Bcl-2 and cytochrome c), cleaved caspase-3, and autophagy-related proteins (LC3-Ⅰ, LC3-Ⅱ and P62) and β-actin, were assessed with western blotting. Celastrol significantly inhibited the viability of HOS cells in a dose-dependent manner and promoted the expression of ERS-related, apoptosis-related and mitochondrial apoptosis-related proteins. The ERS inhibitor tauroursodeoxycholate promoted celastrol-induced autophagy and apoptosis of HOS cells. Pretreatment with the PERK inhibitor GSK2656157 significantly promoted celastrol-induced death and attenuated HOS cell autophagy. Our results indicated that the ERS pathway and the mitochondrial pathway were involved in celastrol-induced apoptosis of HOS cells. The ERS/PERK pathway may protect HOS cells from apoptosis by celastrol and may play a complicated role in the process of autophagy.

MicroRNAs, such as miR-34, have been reported to influence age-related diseases. In this study, we explored the role of the miR-34a/Bcl-2 signaling pathway in age-related hearing loss (AHL). Using an AHL mouse model (C57BL/6), we found that the expression of miR-34a in the cochlea increased with age, whereas expression of Bcl-2 decreased. Increasing the amount of a miR-34a mimetic in a mouse auditory cell line (HEI-OC1) inhibited Bcl-2, leading to enhanced apoptosis; in contrast, miR-34a inhibition produced the opposite effect. Our results support a link between age-related cochlear hair cell apoptosis and miR-34a/Bcl-2 signaling. The latter may thus serve as a potential target for AHL therapy., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Osmolarity fluctuations are inevitable within the nucleus pulposus (NP). However, the effects of osmolarity on NP cell apoptosis within the organ-cultured disc remain unclear. The objective of this study was to investigate effects of different osmolarity levels (hypo-, iso-, and hyper-) and osmolarity modes (constant and cyclic) on NP cell apoptosis in a disc perfusion culture and to study the role of the ERK1/2 pathway in this regulatory process. Porcine discs were cultured for 7 days in different osmotic medium, including constant hypo-, iso-, and hyper-osmolarity (330, 430, and 550 mOsm/L, respectively) and cyclic-osmolarity (430 mOsm/L for 8 h, followed by 550 mOsm/L for 16 h). The role of the ERK1/2 pathway was investigated by using the pharmacological inhibitor U0126. NP cell apoptosis was analyzed by TUNEL staining, caspase-3 activity, gene expression of Bcl-2, Bax and caspase-3 and protein expression of cleaved caspase-3, and cleaved PARP. Our results showed that NP cell apoptosis was increased in hypo- and hyper-osmolarity cultures compared to iso- or cyclic-osmolarity culture, whereas the level of apoptosis in the iso-osmolarity culture was lower than that in the cyclic-osmolarity culture. When the ERK1/2 pathway was inhibited in the iso- and cyclic-osmolarity cultures, the level of NP cell apoptosis was significantly increased. In conclusion, the effects of osmolarity on NP cell apoptosis depend on the osmolarity level (hypo-, iso-, or hyper-) and osmolarity mode (constant or cyclic). Futhermore, inhibition of the ERK1/2 pathway promotes NP cell apoptosis in this process. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:86-92, 2017., (© 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.)

Hypothesis: The miR-34a/Bcl-2 signaling pathway may play a role in the mechanisms related to age-related hearing loss (AHL) in the auditory cortex., Background: The auditory cortex plays a key role in the recognition and processing of complex sound. It is difficult to explain why patients with AHL have poor speech recognition, so increasing numbers of studies have focused on its central change. Although micro (mi)RNAs in the central nervous system have recently been increasingly reported to be associated with age-related diseases, the molecular mechanisms of AHL in the auditory cortex are not fully understood., Methods: The auditory brainstem response was used to assess the hearing ability of C57BL/6 mice, and q-PCR, immunohistochemistry, and Western blotting were used to detect the expression levels of miR-34a and Bcl-2 in the mouse auditory cortex. TUNEL and DNA fragmentation were adopted to detect the apoptosis of neurons in the auditory cortex. To verify the relationship of miR-34a and Bcl-2, we transfected an miR-34a mimic or miR-34a inhibitor into primary auditory cortex neurons., Results: In this study, miR-34a/Bcl-2 signaling was examined in auditory cortex neurons during aging. miR-34a and apoptosis increased in the auditory cortex neurons of C57BL/6 mice with aging, whereas an age-related decrease in Bcl-2 was determined. In the primary neurons of the auditory cortex, miR-34a overexpression inhibited Bcl-2, leading to an increase in apoptosis. Moreover, miR-34a knockdown increased Bcl-2 expression and diminished apoptosis., Conclusion: Our results support a link between age-related apoptosis in auditory cortex neurons and miR-34a/Bcl-2 signaling, which may serve as a potential mechanism of the expression of AHL in the auditory cortex., (© 2017 S. Karger AG, Basel.)

Our objective is to study the relationship between the regulatory proteins Bcl-2/Bax and mitochondria-mediated apoptosis during the differentiation of adipose-derived stromal cells (ADSCs) into neurons. Immunocytochemistry and western blotting showed that the cells weakly expressed neuron-specific enolase (NSE) in the non-induced group and expressed NSE more strongly in the groups induced for 1 h, 3 h, 5 h and 8 h. NSE expression peaked at 5 h (P < 0.05), although there was no significant difference between 5 and 8 h (P > 0.05). Bcl-2 expression gradually decreased over time in the non-induced group (P < 0.05). However, Bax, caspase-9, Cyt-c and caspase-3 expression gradually increased and peaked at 8 h (P < 0.05). Transmission electron microscopy revealed karyopyknosis, chromatin edge setting, mitochondria swelling and cavitation in cells at 5 h, and the mitochondrial membrane potential decreased over time, as demonstrated by laser scanning confocal microscopy. After a 5 h induction, cells differentiated into typical neurons and expressed Bcl-2, which inhibited apoptosis. Bax showed a strong apoptosis-promoting capacity, leading to changes in the mitochondrial membrane potential and structure, and then triggered the caspase-independent apoptotic response through the mitochondrial pathway. At the same time, Cyt-c was directly or indirectly released from the mitochondria to the cytoplasm to trigger the caspase-dependent apoptotic response through the mitochondrial pathway. Therefore, Bcl-2/Bax play an important role in regulating caspase-dependent and caspase-independent apoptosis mediated by the mitochondrial pathway during the differentiation of ADSCs into neurons., Competing Interests: The authors have declared that no competing interests exist.

Pancreatic adenocarcinoma (PDA) is one of the most lethal human malignancies, and unresponsive to current chemotherapies. Here we investigate the therapeutic potential of phycocyanin as an anti-PDA agent in vivo and in vitro. Phycocyanin, a natural product purified from Spirulina, effectively inhibits the pancreatic cancer cell proliferation in vitro and xenograft tumor growth in vivo. Phycocyanin induces G2/M cell cycle arrest, apoptotic and autophagic cell death in PANC-1 cells. Inhibition of autophagy by targeting Beclin 1 using siRNA significantly suppresses cell growth inhibition and death induced by phycocyanin, whereas inhibition of both autophagy and apoptosis rescues phycocyanin-mediated cell death. Mechanistically, cell death induced by phycocyanin is the result of cross-talk among the MAPK, Akt/mTOR/p70S6K and NF-κB pathways. Phycocyanin is able to induce apoptosis of PANC-1 cell by activating p38 and JNK signaling pathways while inhibiting Erk pathway. On the other hand, phycocyanin promotes autophagic cell death by inhibiting PI3/Akt/mTOR signaling pathways. Furthermore, phycocyanin promotes the activation and nuclear translocation of NF-κB, which plays an important role in balancing phycocyanin-mediated apoptosis and autosis. In conclusion, our studies demonstrate that phycocyanin exerts anti-pancreatic cancer activity by inducing apoptotic and autophagic cell death, thereby identifying phycocyanin as a promising anti-pancreatic cancer agent.

Current treatment strategies for lung cancer cause undesirable side‑effects. Integrated medicine with a curative approach has become a common approach to the treatment strategy. Recent studies suggest that American alligator blood is effective in reducing colorectal cancer cell viability in vitro, but the mechanism remains unclear. In the present study, we aimed to study the anticancer activity of crocodile blood extracts on lung cancer cell line A549 and investigate the possible mechanisms involved. In vitro studies were utilized to investigate the effects on the cancer cells after incubation with the blood extracts. The active fraction that showed more efficacy in inhibiting cell growth was characterized in the supernatant (S2) from whole blood extracts. High performance liquid chromatography (HPLC) analysis revealed that S2 contained more polar moiety from whole blood. S2 induced DNA fragmentation. Cell cycle arrest in the G1/M phase was demonstrated and mitochondrial membrane permeability was disrupted. An increase in the generation of reactive oxygen species (ROS) and increased activities of caspase-3 and caspase-7 were detected. Furthermore, release of cytochrome c, upregulation of expression of Bax, p53, p21, Bid, cleaved forms of the caspase family and PARP along with downregulation of Bcl-2, PCNA, MDM2, caspase‑8, wild types of caspase family proteins and PARP were recorded after treatment with S2 fractions. Moreover, the PI3K/AKT survival pathway was downregulated by S2 fractions in the lung cancer cell line.

The present study was carried out to investigate the effect and mechanisms of aloe‑emodin (AE)-mediated photodynamic therapy (AE-PDT) on the human osteosarcoma cell line MG-63. After treatment with AE-PDT, the human osteosarcoma cell line MG-63 was tested for levels of viability, autophagy, reactive oxygen species (ROS) and apoptosis and changes in cell morphology with the Cell Counting Kit-8 (CCK‑8), monodansylcadaverine (MDC) and Hoechst staining and transmission electron microscopy. The expression of proteins including LC-3, cleaved caspase-3, Beclin-1, Bcl-2, p-JNK, t-JNK and β-actin was examined with western blotting. AE-PDT significantly inhibited the viability of the MG-63 cells in an AE-concentration- and PDT energy density-dependent manner. Autophagy and apoptosis of MG-63 cells was substantially promoted in the AE-PDT group compared to the control group, the AE alone group and the light emitting diode (LED) alone group. Inhibition of autophagy by 3-methyladenine (3-MA) (5 mM) and chloroquine (CQ) (15 µM) significantly promoted the apoptosis rate and improved the sensitivity of the MG-63 cells to AE-PDT. AE-PDT was found to induce the expression of ROS and p-JNK. ROS scavenger, N-acetyl-L-cysteine (NAC, 5 mM), was able to hinder the autophagy, apoptosis and phosphorylation of JNK, and JNK inhibitor (SP600125, 10 µM) significantly inhibited the autophagy and apoptosis, and attenuated the sensitivity of MG63 cells to AE-PDT. In conclusion, AE-PDT induced the autophagy and apoptosis of human osteosarcoma cell line MG-63 through the activation of the ROS-JNK signaling pathway. Autophagy may play a protective role during the early stage following treatment of AE-PDT.

Methylglyoxal (MG) is a reactive dicarbonyl compound, whose abnormal accumulation in diabetic patients exerts deleterious effects on cells and tissues. The β-cell is the main target cell of Type 2 diabetes, and its insulin secretion injury and cell apoptosis can be due to mitochondrial dysfunction. Previous studies have demonstrated MG induced β-cell apoptosis. However, little is known about the effect of MG on β-cell mitochondrial dysfunction. Phycocyanin (PC) has been demonstrated to possess various biological activities including the effects on diabetic models in vivo. The aim of this study was to determine the protective effect of PC against methylglyoxal (MG)-induced dysfunction in pancreatic β-cell INS-1 and also the mechanism. We demonstrated that MG induced mitochondrial dysfunction by the decline in ATP levels, and the increase of the level of intracellular reactive oxygen species (ROS). Furthermore, MG released cytochrome c and apoptosis-inducing factor (AIF) from the mitochondrion, induced changes in the expression of Bcl-2 family members, activated caspases and increased PARP cleavage. Interestingly, PC activated nuclear erythroid-related factor 2 (Nrf2), and Nrf2 activation as well as antioxidant enzymes HO-1 and glyoxalase 1 (Glo-1) were confirmed to be involved in the mechanisms underlying the protection of PC by RNA interference. Altogether, these results demonstrated that PC prevented mitochondrial-dependent apoptosis in MG-induced INS-1 cells and the effect was associated with Nrf2 activation.

The molecular mechanisms underlying age-related hearing loss are not fully understood, and currently, there is no treatment for this disorder. MicroRNAs have recently been reported to be increasingly associated with age-related diseases and are emerging as promising therapeutic targets. In this study, miR-34a/Sirtuin 1 (SIRT1)/p53 signaling was examined in cochlear hair cells during aging. MiR-34a, p53 acetylation, and apoptosis increased in the cochlea of C57BL/6 mice with aging, whereas an age-related decrease in SIRT1 was observed. In the inner ear HEI-OC1 cell line, miR-34a overexpression inhibited SIRT1, leading to an increase in p53 acetylation and apoptosis. Moreover, miR-34a knockdown increased SIRT1 expression and diminished p53 acetylation, and apoptosis. Additionally, resveratrol, an activator of SIRT1, significantly rescued miR-34a overexpression-induced HEI-OC1 cell death and significantly reduced hearing threshold shifts and hair cell loss in C57BL/6 mice after a 2-month administration. Our results support a link between age-related cochlear hair cell apoptosis and miR-34a/SIRT1/p53 signaling, which may serve as a potential target for age-related hearing loss treatment., (Copyright © 2015 Elsevier Inc. All rights reserved.)

To research the features of mitochondrial fusion and fission proteins and their relationship with apoptosis in the process of adipose-derived stromal cells (ADSCs) differentiation into astrocytes. Immunocytochemistry and Western-blotting were used to analyze the expression of glial fibrillary acidic protein (GFAP), mitochondria fusion-2 (Mfn2), mitochondrial fission-1 (Fis1) and cysteine aspartate specific protease-3 (Caspase-3). Flow cytometry for quantification of the number of apoptotic cells. Transmission electron microscopy (TEM) for observation of the ultrastructure. After induced for 48h, 7, 14 and 21 days showed expression of GFAP, reached the peak on the 7th day (P<0.05). The expression of Mfn2 was decreased with the induction time extending, reached the minimum on the 14th day (P<0.05). The expression of Fis1 and Caspase-3 was increased with the induction time extending, reached the peak on the 14th day (P<0.05, respectively). The rates of early apoptosis, late apoptosis or necrosis were increased gradually (P<0.05). Our findings suggest that in the process of ADSCs differentiation into astrocytes, mitochondrial fusion decreased while mitochondrial fission enhanced significantly; caspase-dependent apoptosis was one of the main reasons leading to cell death., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

The present study aims to investigate the pharmacological effect of the exopolysaccharides from Aphanothece halophytica GR02 (EPSAH) on the HeLa human cervical cancer cell line. HeLa cells were cultured in RPMI-1640-10% FBS medium containing with or without different concentrations of EPSAH. Cell viability was assessed by methylthiazol tetrazolium (MTT) assay. Cell apoptosis was elevated with Wright-Giemsa staining, AO/EB double staining, and DNA fragmentation assay. Apoptosis-associated molecules from cultured HeLa cells were quantified using Western blot analysis. Our results suggest that EPASH induces apoptosis in HeLa cells by targeting a master unfolded protein response (UPR) regulator Grp78. Grp78 further promotes the expression of CHOP and downregulates expression of survivin, which leads to activate mitochondria-mediated downstream molecules and p53-survivin pathway, resulting in caspase-3 activation and causing apoptosis. These findings provide important clues for further evaluating the potential potency of EPSAH for use in cancer therapy.

Background: The aim of this study is to investigate the effects of polyphenol extract from Phyllanthus emblica (PEEP) on cervical cancer cells and to explore the underlying mechanism., Methods: MTT assay was used to measure inhibition of proliferation of cervical cancer (HeLa) cells after treatment with PEEP at concentrations of 0, 50, 100, 150, and 200 mg/ml for 48 hours. HeLa cells were treated with PEEP (150 mg/ml) for 48 hours in the following analysis. Karyomorphism was assessed by immunofluorescence using DAPI staining, and cell apoptosis and cell cycle were assessed using flow cytometry. Three apoptotic marker proteins, namely, Fas, FasL, and cleaved caspase-8, were assessed by western blotting., Results: PEEP inhibited the growth of HeLa cells, and the optimum concentration of PEEP was 150 mg/ml. In addition, the karyomorphism of HeLa cells after treatment with PEEP was abnormal. Furthermore, PEEP induced arrest of the HeLa cell cycle at G2/M phase, and triggered apoptosis. PEEP also induced significant Fas and FasL activation, and cleavage of caspase-8., Conclusions: Our study indicates that PEEP is effective in inhibiting HeLa cell proliferation by inducing cell cycle arrest at G2/M phase and inducing apoptosis.

The role of cyclin B1 in the clinical therapeutic sensitivity of human esophageal squamous cell carcinoma (ESCC) remains to be defined. In this study, we found that elevated cyclin B1 expression attenuated the apoptosis induced by cisplatin or paclitaxel, while knockdown of cyclin B1 enhanced cisplatin or paclitaxel sensitivity in ESCC cells. Cyclin B1-mediated apoptosis may rely on the Bcl-2-dependent mitochondria-regulated intrinsic death pathway, and the antagonizing effect of cyclin B1 on chemotherapeutic agent-induced apoptosis was through PTEN/Akt pathway. Therefore, cyclin B1 might be a therapeutic target for the development of specific and efficient approaches in the treatment of ESCC.

Apoptosis of lens epithelial cell (LEC) plays an important role in cataract formation, and its prevention may be one of the therapeutic strategies in treating cataract. This study used human lens epithelial cell (hLEC) line SRA01/04 to investigate the protective effect and mechanism of phycocyanin on glactose-induced apoptosis in hLEC. hLECs were cultured in D/F(12)-10% FBS medium containing 125mM d-galactose with or without phycocyanin. Cell viability was assessed by methylthiazol tetrazolium (MTT) assay. Cell apoptosis was elevated with Wright-Giemsa staining, AO/EB double staining, and DNA fragmentation assay. Mitochondrial apoptosis-associated molecules and unfolded protein response-associated molecules from cultured SRA01/04 cells were quantified using protein blot analysis. The results demonstrated that phycocyanin suppressed SRA01/04 cells' morphologic changes and apoptosis induced by d-galactose, inhibited the expression and activation of caspase 3, alternated the Bax/Bcl-2 ratio, and down-regulated the level of p53, GRP78, and CHOP in d-galactose-treated SRA01/04 cells. These results suggest that phycocyanin might suppress d-galactose-induced hLEC apoptosis through two pathways: mitochondrial pathway, involving p53 and Bcl-2 family protein expression, and unfolded protein response pathway, involving GRP78 and CHOP expression., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Background: Celecoxib has a positive effect on human osteoarthritic cartilage, but the mechanisms remain unclear. The aim of this study was to test whether celecoxib could inhibit the apoptosis of chondrocyte and ameliorate type II collagen synthesis to relieve symptoms of OA (osteoarthritis)., Material/methods: 130 Wistar rats were randomly divided into 4 groups as celecoxib (CE), ibuprofen (IBP), indomethacin (IN) and normal saline group (NS). The osteoarthritis was induced by the excision of the left Achilles tendon. At the 3th, 6th, 9th month of treatment, the histological structure of articular cartilage was observed using HE staining. Type II collagen was examined using immunohistochemistry. Chondrocyte apoptosis was detected by TUNEL staining, and the change of ultra-microstructure of chondrocyte was examined through a transmission electron microscope., Results: CE reduced the OA-like histological changes and suppressed chondrocyte apoptosis. However, IN or IBP had deleterious effects on articular cartilage and enhanced the chondrocyte apoptosis. IBP promoted the expression of type II collagen, and IN inhibited its expression, but had no effect in the CE group., Conclusions: CE had favorable action on OA progression, and may be the ideal choice in the treatment of chronic destructive joint disease where anti-inflammatory drugs need to be used for a prolonged period.

Objectives: Human lactoferrin, an 80 kDa iron-binding glycoprotein, has antitumour effects. We have explored the potential therapeutic role of re-expressing human lactoferrin gene product in human breast cancer., Methods: A recombinant adenovirus expressing the human lactoferrin cDNA (ad-hLTF) was constructed and used to infect breast cancer cells., Key Findings: Seventy-two hours after infection, ad-hLTF had considerable cytotoxicity on MCF-7 cells. A time-course study showed that ad-hLTF infection of MCF-7 cells at 100 plaque-forming units per cell increased the number of cells in G(0) /G(1) phase and appeared markedly at Sub-G(1) apoptotic peak. The presence of apoptotic cells was confirmed using Annexin V-fluoresecein isothiocyanate apoptosis detection by flow cytometry. Ad-hLTF also resulted in a decrease of Bcl-2 protein and an increase in Bax protein., Conclusions: Ad-hLTF plays an important role in the induction of cell cycle arrest and apoptosis in MCF-7 cells. The results demonstrated that ad-hLTF could have potential benefits in the treatment of breast cancer., (© 2011 The Authors. JPP © 2011 Royal Pharmaceutical Society.)

A tetramethylpyrazine analogue, CXC195, was synthesized by the Boekelheide reaction, in which the second methyl group of tetramethylpyrazine (TMP) was replaced with (4,4'-fluorine) diphenyl-methyl-1-piperazidine, the active group of flunarizine. We have observed protective effects of CXC195 on vascular endothelial cell survival under oxidative stress in previous study. The aim of the present study was to investigate the effects of CXC195 against apoptosis induced by hydrogen peroxide in human umbilical vein endothelial cells (HUVECs). Accordingly, a biochemical approach to elucidate the apoptotic signal pathways was attempted. HUVECs were exposed to 150 muM H(2)O(2) for 12 h, resulting in an increase of apoptotic cells assessed by the nuclear staining assay and flow cytometry. Mitochondrial membrane potential was detected by retention of rhodamine123. The concentration of free intracellular calcium was determined by fura-2/AM fluorometry. Co-incubation with CXC195 reduced the percentage of apoptotic cells and inhibited the loss of mitochondrial membrane potential and intracellular calcium overload induced by H(2)O(2). Induction of p53, the activation of caspase-3 by H(2)O(2) which accompanying downregulation of bcl-2, was blocked by CXC195. In addition, CXC195 clearly improved phosphorylation levels of the antiapoptotic extracellular signal-regulated kinase-1/2 (ERK1/2) in cells undergoing oxidative damage. Moreover, CXC195 showed stronger effects on inhibition of apoptotic cells and loss of mitochondrial membrane potential and activation of phosphorylated ERK1/2 than TMP. These results suggest that CXC195 prevents reactive oxygen species-induced apoptosis through inhibition of the mitochondria-dependent caspase-3 pathway and ERK pathway to show a better beneficial effect in protecting endothelial cells than TMP.

Oxidative processes in the lenses are the most commonly found damaging factor for the development of cataracts. Hirudin, a most potent inhibitor of thrombin as an antithrombic drug, also have potential use in cataracts. In order to investigate the mechanisms of hirudin against galactose-induced cataract at the cellular level. We used recombinant hirudin variant III (rHV3) to study the protective effect of hirudin on galactose-mediated human lens epithelial cells injury. The human lens epithelial cells (hLECs) were cultured in D/F(12)-10% FBS medium containing 125 mM D-galactose with or without rHV3. Cell viability was assessed by methylthiazol tetrazolium (MTT) assay and propidium iodide (PI) staining in situ. Cell apoptosis was elevated with comet assay (single cell gel electrophoresis, SCGE), AO/EB double staining and Annexin-V/PI double staining assay. Reactive oxygen species (ROS) were quantified with 2',7'-dichlorofluorescein (DCF), and free glutathione (GSH) levels were measured with a commercial GSH quantification kit. Decreased viability and increased apoptosis of the hLECs were observed when incubated with 125 mM galactose. These hLECs also demonstrated the increased presence of ROS, whereas GSH was reduced. rHV3 blocked the induction of cell death, apoptosis and oxidative stress in hLECs. One mechanism may be through regulating intracellular ROS and GSH levels to inhibit apoptosis of the human lens epithelial cells.

Objective: To study whether As(2)O(3) has an apoptotic effect on human solid tumor cells, and the possible cellular and molecular mechanisms of this treatment using human esophageal squamous carcinoma cells (EC8712) as a model., Methods: DNA microarray, biochemical and cytological analyses were used., Results: The growth and survival of EC8712 cells were markedly inhibited by As(2)O(3) treatment at a concentration of 1, 2 and 4 micromol/L. EC8712 cells were obviously arrested at G2/M phase with As(2)O(3) treatment and apoptosis induced at micromolar As(2)O(3) concentrations, as shown by morphology, histogram related nuclear DNA contents, and DNA gel electrophoresis. As(2)O(3) activated caspase-3, which might be involved in the process of As(2)O(3), induced apoptosis in EC8712 cells., Conclusions: As(2)O(3) changes the expression of many genes at transcription level. The regulation of expression of many genes might be involved in the process of As(2)O(3) inducing apoptosis. These results suggest that As(2)O(3) can be clinically useful for solid tumor treatment.

Aim: To investigate regulative effects of thiol reagents, N-acetyl-l-cysteine (NAC) and natrii dimercaptosussinas (NDMS), catalase (CAT), and calcium chelator 2-[(2-bis-[carboxymethyl]-amino-5-methyl-phenoxy)-met]-6-methoxy-8-bis-[carboxy-methyl]-aminoquinoline (Quin 2) on apoptosis and telomerase activity induced by arsenic trioxide (As2O3) in three myelocytic leukemia cell lines., Methods: Flow cytometry was used to examine apoptosis and a PCR ELISA kit was used to detect telomerase activity., Results: As2O3 induced about 40 % - 60 % of apoptosis in NB4, K562, and HL-60 cells at the concentration of 0.6, 2.7, and 8.1 micromol/L respectively, as well as down-regulated telomerase activities in three cell lines. NAC 4 mmol/L, NDMS 200 micromol/L, CAT 80 kU/L, and Quin 2 20 micromol/L could down-regulate apoptosis variously induced by As2O3. NAC and CAT alone could decline telomerase activity in three cell lines and further decline telomerase activities that had been decreased by As2O3, whereas Quin 2 antagonized the decline in K562 and HL-60 cells., Conclusion: Thiol activity loss, free radical alteration, intracellular calcium changes, and decline of telomerase activity might be involved in As2O3-induced apoptosis. NAC, NDMS, CAT, and Quin 2 antagonized in some extent the effect of As2O3 on the three tested cell lines.

An immortal human hepatic cell line HL-7702 and human hepatoma cell line SMMC-7721 were treated with 3-30 microM SeO(2). SeO(2) at 30 microM markedly inhibited cell proliferation and viability, and prompted apoptosis of both normal hepatic and hepatoma cells after 48h treatment. SeO(2) could also down-regulate the Bcl-2 level, greatly in HL-7702 and slightly in SMMC-7721 cells, but up-regulate wild type P53 level a little in HL-7702 and significantly in SMMC-7721 cells. The Bcl-2/P53 value was closely correlated with the apoptotic rate as well as SeO(2) concentrations.