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Purpose: Paclitaxel is an anticancer drug for the treatment of non-small cell lung cancer (NSCLC). However, drug-resistance remains a major problem. Honokiol is a natural component which has been found to exhibit anti-tumor activity. Paclitaxel and honokiol have been reported to be able to induce paraptosis. The aim of this study was to investigate whether honokiol can reverse paclitaxel resistance by inducing paraptosis in NSCLC cells., Methods: NSCLC cell lines H1650 (paclitaxel-sensitive), H1299 and H1650/PTX (intrinsic and acquired paclitaxel-resistant, respectively) were used to assess the cytotoxic effects of paclitaxel and honokiol. Light and transmission electron microscopy were performed to detect cytoplasmic vacuolation. In vitro cell viability and clonogenic survival assays, as well as in vivo xenograft assays were conducted to test synergistic killing effects of paclitaxel and honokiol on NSCLC cells. Western blotting, flow cytometry and immunofluorescence were performed to evaluate paraptosis-regulating mechanisms., Results: We found that combination treatment with paclitaxel and honokiol synergistically killed H1650, H1299 and H1650/PTX cells by inducing paraptosis, which is characterized by cytoplasmic vacuolation. Moreover, paclitaxel/honokiol treatment resulted in a significant growth delay in H1299 xenograft tumors that showed extensive cytoplasmic vacuolation. Mechanistically, proteasomal inhibition-mediated endoplasmic reticulum (ER) stress and unfolded protein responses leading to ER dilation, and the disruption of intracellular Ca 2+ homeostasis and mitochondrial Ca 2+ overload resulting in mitochondrial disfunction, were found to be involved in paclitaxel/honokiol-induced paraptosis. Cellular protein light chain 3 (LC3) may play an important role in paclitaxel/honokiol induced cytoplasmic vacuolation and NSCLC cell death., Conclusions: Combination of honokiol and paclitaxel may represent a novel strategy for the treatment of paclitaxel-resistant NSCLC.

Non-small cell lung cancer (NSCLC) with epidermal growth factor receptor (EGFR) wild-type is intrinsic resistance to EGFR-tyrosine kinase inhibitors (TKIs). In this study, we assessed whether the combination of bisdemethoxycurcumin (BDMC) and icotinib could surmount primary EGFR-TKI resistance in NSCLC cells and investigated its molecular mechanism. Results showed that the combination of BDMC and icotinib produced potently synergistic growth inhibitory effect on primary EGFR-TKI-resistant NSCLC cell lines H460 (EGFR wild-type and K-ras mutation) and H1781 (EGFR wild-type and Her2 mutation). Compared with BDMC or icotinib alone, the two drug combination induced more significant apoptosis and autophagy via suppressing EGFR activity and interaction of Sp1 and HDCA1/HDCA2, which was accompanied by accumulation of reactive oxygen species (ROS), induction of DNA damage, and inhibition of cell migration and invasion. ROS inhibitor (NAC) and autophagy inhibitors (CQ or 3-MA) partially reversed BDMC plus icotinib-induced growth inhibitory effect on the NSCLC cells. Meanwhile, co-treatment with NAC attenuated the two drug combination-induced autophagy, apoptosis, DNA damage and decrease of cell migration and invasion ability. Also, 3-MA or CQ can abate the combination treatment-induced apoptosis and DNA damage, suggesting that there is crosstalk between different signaling pathways in the effect produced by the combination treatment. Our data indicate that BMDC has the potential to improve the treatment of primary EGFR-TKI resistant NISCLC that cannot be controlled with single-target agent, such as icotinib., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

G kinase-anchoring protein 1 (GKAP1) is a G kinase-associated protein that is conserved in many eutherians and is mainly expressed in the testis, especially in spermatocytes and round spermatids. The function of GKAP1 in the testis is largely unknown. Here, we revealed that deletion of GKAP1 led to an increase in sperm production with swollen epididymis, and germ cell apoptosis was found to decrease in GKAP1 knock-out mice. Further investigations showed that a deficiency of GKAP1 could partly change the cellular location of cGK-Iα and increase the amount of active cAMP response element-binding protein (CREB) in the nucleus. Therefore, the expression of a particular inhibitor of apoptosis proteins (IAPs) was upregulated because of the activation of CREB, and this increase in IAPs was associated with a decrease in the level of activated caspase-3. These results suggest that a deficiency of GKAP1 in mouse testis could increase sperm production through a reduction of the spontaneous apoptosis of germ cells in the testis, possibly because of a change in the activity of the cGK-Iα pathway., (© 2019 Wiley Periodicals, Inc.)

Sirtuin 1 (Sirt1) exerts its cardioprotective effects in various cardiovascular diseases via multiple cellular activities. However, the therapeutic implications of Sirt1 in hypoxic cardiomyocytes and the underlying mechanisms remain elusive. The present study investigated whether Sirt1 regulates autophagy and apoptosis in hypoxic H9C2 cardiomyocytes and in an experimental hypoxic mouse model. Right ventricular outflow tract biopsies were obtained from patients with cyanotic or acyanotic congenital heart diseases. Adenovirus Ad‑Sirt1 was used to activate Sirt1 and Ad‑Sh‑Sirt1 was used to inhibit Sirt1 expression in H9C2 cells, in order to investigate the effect of Sirt1 on cellular autophagy and apoptosis. SRT1720, a pharmacological activator of Sirt1 and EX‑527, a Sirt1 antagonist, were administered to mice to explore the role of Sirt1 in hypoxic cardiomyocytes in vivo. The levels of autophagy and apoptosis‑related proteins were evaluated using western blotting. Apoptosis was investigated by TUNEL staining and Annexin V/7‑aminoactinomycin D flow cytometry analysis. Heart tissue samples from cyanotic patients exhibited increased autophagy and apoptosis, as well as elevated Sirt1 levels, compared with the noncyanotic control samples. The data from the western blot analysis revealed that Sirt1 promoted autophagic flux and reduced apoptosis in hypoxic H9C2 cells. In addition, Sirt1 activated AMP‑activated protein kinase (AMPK), and the AMPK inhibitor Compound C abolished the effect of Sirt1 on autophagy activation. Further exploration of the mechanism revealed that Sirt1 protects hypoxic cardiomyocytes from apoptosis, at least in part, through inositol requiring kinase enzyme 1α (IRE1α). Consistent with the in vitro results, treatment with the Sirt1 activator SRT1720 activated AMPK, inhibited IRE1α, enhanced autophagy, and decreased apoptosis in the heart tissues of normoxic mice compared with the hypoxia control group. Opposite changes were observed in hypoxic mice treated with the Sirt1 inhibitor EX‑527. These results suggested that Sirt1 promoted autophagy via AMPK activation and reduced hypoxia‑induced apoptosis via the IRE1α pathway, to protect cardiomyocytes from hypoxic stress.

Despite advances in management, bladder cancer remains a principal cause of cancer‑associated complications. Tumor necrosis factor receptor superfamily member 14 (TNFRSF14) is dysregulated in certain types of cancer; however, limited data are available on the expression and function of TNFRSF14 in bladder cancer. In the present study, the aim was to evaluate the expression and biological functions of TNFRSF14 in bladder cancer. Firstly, the expression levels of TNFRSF14 in bladder cancer tissue were examined using The Cancer Genome Atlas (TCGA) database. Secondly, reverse transcription‑quantitative polymerase chain reaction was utilized to investigate the expression levels of TNFRSF14 in the T24, SW780 and EJ‑M3 bladder cancer cell lines. Transfection and Cell Counting kit‑8 (CCK‑8) assay was used to evaluate whether TNFRSF14 overexpression or silencing would have an effect on cell proliferation of T24 and EJ‑M3 cells. In addition, TNFRSF14‑induced apoptotic cells were identified using Annexin V‑fluorescein isothiocyanate and propidium iodide staining. Western blot analysis was used to detect proteins associated with the phosphatidylinositol 3‑kinase pathway. According to the TCGA dataset, the expression levels TNFRSF14 were decreased in bladder cancer tissue compared with in normal control samples. Patients with bladder cancer exhibiting low expression levels of TNFRSF14 had a worse prognosis compared to those with high expression levels of TNFRSF14. Overexpression of TNFRSF14 in T24 cells led to increased apoptosis and inhibited cell proliferation in vitro. Western blotting demonstrated that TNFRSF14 overexpression increased the expression levels of caspase3‑p17 in T24 cells, but significantly decreased the expression levels of phosphorylated (p)‑protein kinase B (AKT) and P70 S6 kinase (P70). TNFRSF14 silencing in EJ‑M3 cells enhanced cell growth, inhibited cell apoptosis, increased the expression levels of p‑AKT and P70, and decreased the expression levels of caspase3‑p17. In conclusion, TNFRSF14 may serve a tumor suppressive role in bladder cancer by inducing apoptosis and suppressing proliferation, and act as a novel prognostic biomarker for bladder cancer.

Malignant pheochromocytoma (PHEO) is diagnosed only when metastasis has occurred, making it less likely for patients to obtain the benefits of traditional chemotherapy. Anti-oncogene TP53 mutation has been detected in PHEO and is possibly related to disease progression. However, whether the upregulation of wild-type TP53 has antitumoral effects on PHEO remains completely unknown. In the present study, we used RNA activation (RNAa) technique to upregulate the expression of wild-type TP53 by transfecting synthetic dsP53‑285 into PHEO cell line PC12. We found that the upregulation of wild-type p53 blocked the transition of PC12 cells from the G0/G1 to the S phase, with induction of apoptosis. Additionally, the above-mentioned findings were attested in vivo. Most importantly, dsP53-285-induced antitumoral effects were reversible following co-transfection with siRNA that targeted p53 mRNA. Collectively, our results revealed that the upregulation of p53 and possibly other anti-oncogenes may provide a potential effective therapeutic strategy for PHEO.

Cadmium (Cd), an environmental and industrial pollutant, affects the nervous system and consequential neurodegenerative disorders. Recently, we have shown that celastrol prevents Cd-induced neuronal cell death partially by suppressing Akt/mTOR pathway. However, the underlying mechanism remains to be elucidated. Here, we show that celastrol attenuated Cd-elevated intracellular-free calcium ([Ca 2+ ] i ) level and apoptosis in neuronal cells. Celastrol prevented Cd-induced neuronal apoptosis by inhibiting Akt-mediated mTOR pathway, as inhibition of Akt with Akt inhibitor X or ectopic expression of dominant negative Akt reinforced celastrol's prevention of Cd-induced phosphorylation of S6K1/4E-BP1 and cell apoptosis. Furthermore, chelating intracellular Ca 2+ with BAPTA/AM or preventing [Ca 2+ ] i elevation using EGTA potentiated celastrol's repression of Cd-induced [Ca 2+ ] i elevation and consequential activation of Akt/mTOR pathway and cell apoptosis. Moreover, celastrol blocked Cd-elicited phosphorylation of CaMKII, and pretreatment with BAPTA/AM or EGTA enhanced celastrol's suppression of Cd-increased phosphorylation of CaMKII in neuronal cells, implying that celastrol hinders [Ca 2+ ] i -mediated CaMKII phosphorylation. Inhibiting CaMKII with KN93 or silencing CaMKII attenuated Cd activation of Akt/mTOR pathway and cell apoptosis, and this was strengthened by celastrol. Taken together, these data demonstrate that celastrol attenuates Cd-induced neuronal apoptosis via inhibiting Ca 2+ -CaMKII-dependent Akt/mTOR pathway. Our findings underscore that celastrol may act as a neuroprotective agent for the prevention of Cd-induced neurodegenerative disorders. J. Cell. Physiol. 232: 2145-2157, 2017. © 2016 Wiley Periodicals, Inc., (© 2016 Wiley Periodicals, Inc.)

Background: Cancer cells show increased glycolysis and take advantage of this metabolic pathway to generate ATP. The TP53-induced glycolysis and apoptosis regulator (TIGAR) inhibits aerobic glycolysis and protects tumor cells from intracellular reactive oxygen species (ROS)-associated apoptosis. However, the function of TIGAR in glycolysis and survival of acute myeloid leukemia cells remains unclear., Methods: We analyzed TIGAR expression in cytogenetically normal (CN-) AML patients and the correlations with clinical and biological parameters. In vivo and in vitro, we tested whether glycolysis may induce TIGAR expression and evaluated the combination effect of glycolysis inhibitor and TIGAR knockdown on human leukemia cell proliferation., Results: High TIGAR expression was an independent predictor of poor survival and high incidence of relapse in adult patients with CN-AML. TIGAR also showed high expression in multiple human leukemia cell lines and knockdown of TIGAR activated glycolysis through PFKFB3 upregulation in human leukemia cells. Knockdown of TIGAR inhibited the proliferation of human leukemia cells and sensitized leukemia cells to glycolysis inhibitor both in vitro and in vivo. Furthermore, TIGAR knockdown in combination with glycolysis inhibitor 2-DG led leukemia cells to apoptosis. In addition, the p53 activator Nutlin-3α showed a significant combinational effect with TIGAR knockdown in leukemia cells. However, TIGAR expression and its anti-apoptotic effects were uncoupled from overexpression of exogenous p53 in leukemia cells., Conclusions: TIGAR might be a predictor of poor survival and high incidence of relapse in AML patients, and the combination of TIGAR inhibitors with anti-glycolytic agents may be novel therapies for the future clinical use in AML patients.

Objectives: Circulating chronic lymphocytic leukemia (CLL) cells appear not to be overly utilizing aerobic glycolysis. However, recurrent contact with CLL cells in a stromal microenvironment leads to increased aerobic glycolysis and the cells' overall glycolytic capacity, which promotes cell survival and proliferation. TP53-induced glycolysis and apoptosis regulator (TIGAR) has been directly implicated in cellular metabolism in the control of glycolysis. TIGAR inhibits glycolysis and protects cells from intracellular reactive oxygen species (ROS)-associated apoptosis., Methods: TIGAR mRNA expression was investigated by quantitative PCR in 102 newly diagnosed CLL patients. Furthermore, the relationship between the expression of TIGAR and its clinical characteristics and prognosis were investigated. Moreover, we also investigated the correlation between TIGAR expression and apoptosis in primary CLL cells., Results: Our data revealed that TIGAR overexpression was correlated with the protection from spontaneous apoptosis in CLL cells, and is strongly associated with advanced Binet stage, unmutated immunoglobulin heavy-chain variable region (IGHV) status, CD38 positivity, β2-microglobulin and p53 aberrations. Higher expression of TIGAR was associated with shorter treatment-free survival (median: three months vs. 51 months, P=0.0108), worse overall survival (median: 74 months vs. not reached, P=0.0242), and the diverse responses to fludarabine-based chemotherapy. TIGAR expression in patients resistant to chemotherapy was significantly higher than in patients sensitive to chemotherapy (mean: 0.3859±0.1710 vs. 0.0974±0.0291, P=0.0290)., Conclusion: Taken together, our findings revealed that high TIGAR expression is closely correlated with worse clinical outcome in CLL patients, and depicted how bioenergetic characteristics could be therapeutically exploited in CLL., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Cadmium (Cd) is a highly toxic metal that affects the central nervous system. Recently we have demonstrated that inhibition of mTOR by rapamycin rescues neuronal cells from Cd-poisoning. Here we show that rapamycin inhibited Cd-induced mitochondrial ROS-dependent neuronal apoptosis. Intriguingly, rapamycin remarkably blocked phosphorylation of JNK, Erk1/2 and p38 in neuronal cells induced by Cd, which was strengthened by co-treatment with Mito-TEMPO. Inhibition of JNK and Erk1/2 by SP600125 and U0126, respectively, potentiated rapamycin's prevention from Cd-induced apoptosis. Consistently, over-expression of dominant negative c-Jun or MKK1 also potently improved the inhibitory effect of rapamycin on Cd neurotoxicity. Furthermore, pretreatment with SP600125 or U0126, or expression of dominant negative c-Jun or MKK1 enhanced the inhibitory effects of rapamycin or Mito-TEMPO on Cd-induced ROS. Further investigation found that co-treatment with Mito-TEMPO/rapamycin more effectively rescued cells by preventing Cd inactivation of PP2A than treatment with rapamycin or Mito-TEMPO alone. Over-expression of wild-type PP2A reinforced rapamycin or Mito-TEMPO suppression of activated JNK and Erk1/2 pathways, as well as ROS production and apoptosis in neuronal cells in response to Cd. The findings indicate that rapamycin ameliorates Cd-evoked neuronal apoptosis by preventing mitochondrial ROS inactivation of PP2A, thereby suppressing activation of JNK and Erk1/2 pathways. Our results underline that rapamycin may have a potential in preventing Cd-induced oxidative stress and neurodegenerative diseases., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Background: Apoptosis of endothelial cells caused by reactive oxygen species plays an important role in ischemia/reperfusion injury after cerebral infarction. Buyang Huanwu Decoction (BYHWD) has been used to treat stroke and stroke-induced disability, however, the mechanism for this treatment remains unknown. In this study, we investigated whether BYHWD can protect human umbilical vein endothelial cells (HUVECs) from H2O2-induced apoptosis and explored the underlying mechanisms., Methods: To investigate the effect of BYHWD on the apoptosis of HUVECs, we established a H2O2-induced oxidative stress model and detected apoptosis by Hoechst 33342 and propidium iodide staining. JC-1 and DCFH-DA assays,western blotting and electron microscopy were used to examine the mechanism of BYHWD on apoptosis., Results: Pretreatment with BYHWD significantly inhibited H2O2-induced apoptosis and protein caspase-3 expression in a concentration-dependent manner. In addition, BYHWD reduced reactive oxygen species production and promoted endogenous antioxidant defenses. Furthermore, loss of mitochondrial membrane potential and structural disruption of mitochondria were both rescued by BYHWD., Conclusions: BYHWD protects HUVECs from H2O2-induced apoptosis by inhibiting oxidative stress damage and mitochondrial dysfunction. These findings indicate that BYHWD is a promising treatment for cerebral ischemia diseases.

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.

Bisphenol-A-glycidyl methacrylate (Bis-GMA) released from dental resin materials causes various toxic effects on gingival epithelium. Thus the underlying mechanisms of its cytotoxicity should be elucidated for safety use. One potential cause of cell damage is the generation of reactive oxygen species (ROS) beyond the capacity of a balanced redox regulation. In this study, we found that exposure of human oral keratinocytes (HOKs) to Bis-GMA caused apoptosis and G1/S cell cycle arrest in parallel with an increased ROS level. Moreover, Bis-GMA induced a depletion of mitochondrial membrane potential, an increase in the Bax/Bcl-2 ratio, an activation of caspase-3 and altered expressions of cell cycle-related proteins (p21, PCNA, cyclinD1). Furthermore, the co-treatment of the ROS scavenger N-acetyl cysteine (NAC) obviously attenuated Bis-GMA-induced toxicity. Here we also evaluated the effects of Bis-GMA on the ROS-related PI3k/Akt pathway. We found that Bis-GMA inhibited the phosphorylation of Akt, whereas the amount of phosphorylated Akt was reverted to the control level in the presence of NAC. Our findings suggested that the toxic effects of Bis-GMA were related to ROS production and the antioxidant NAC effectively reduced Bis-GMA-mediated cytotoxicity., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Cadmium (Cd), an environmental contaminant, causes neurodegenerative disorders. Recently we have shown that rapamycin prevents Cd-induced neuronal cell death by inhibiting mTOR signaling pathway. Here we found that rapamycin exerted its prevention against Cd-induced neuronal cell death also partially via blocking Erk1/2 pathway. Inhibiting Erk1/2 with PD98059 or silencing Erk1/2 potentiated rapamycin's inhibition of Cd-induced phosphorylation of Erk1/2 and apoptosis in neuronal cells. Both PP2A and PTEN/Akt were involved in the regulation of Erk1/2 activation and cell death triggered by Cd. Inhibition of PP2A with okadaic acid or ectopic expression of dominant negative PP2A attenuated rapamycin's inhibition of Cd-induced phospho-Erk1/2 and apoptosis, whereas over-expression of wild-type PP2A enhanced rapamycin's effects; Over-expression of wild-type PTEN or dominant negative Akt, or inhibition of Akt with Akt inhibitor X strengthened rapamycin's inhibition of Cd-induced phospho-Erk1/2 and cell death. Furthermore, expression of a rapamycin-resistant and kinase-active mTOR (mTOR-T) blocked rapamycin's inhibitory effects on Cd-induced inhibition of PP2A, down-regulation of PTEN, and activation of Akt, leading to Erk1/2 activation and cell death, whereas silencing mTOR mimicked rapamycin's effects. The results uncover that rapamycin inhibits Cd activation of Erk1/2-mediated neuronal apoptosis through intervening mTOR-PP2A/PTEN signaling network.

Berberine, an isoquinoline alkaloid extracted from the dry root of Coptidis Rhizoma, has been found to exhibit marked anticancer effects on a panel of established cancer cells. Among the human osteosarcoma lines treated, MG‑63 cells were found to be the most sensitive. The present study investigated the potential genotoxic effect of berberine on MG‑63 human osteosarcoma cells. The effect of berberine on cell viability was determined using a 3-(4,5-dimethylthiazol-2-yl)-2,5‑diphenyltetrazolium bromide assay and cell apoptosis was analyzed by flow cytometry and a DNA ladder assay. γH2AX focus formation was used to detect DNA damage in MG-63 cells. Berberine induced a significant increase in apoptosis in MG-63 cells in a concentration- and time-dependent manner, as determined by DNA fragmentation analysis and flow cytometry. Furthermore, berberine induced significant concentration- and time-dependent increases in DNA damage compared with that in the negative control. In conclusion, these observations indicated that berberine induced apoptosis and DNA damage in MG‑63 cells.

Background: Hypoxia-inducible factor-1 alpha (HIF-1α) plays a critical role in tumor cell adaption to hypoxia by inducing the transcription of numerous genes. The role of HIF-1α in malignant retinoblastoma remains unclear. We analyzed the role of HIF-1α in WERI-Rb-1 retinoblastoma cells under hypoxic conditions., Methods: CoCl2 (125 mmol/L) was added to the culture media to mimic hypoxia. HIF-1α was silenced using siRNA. Gene and protein expression were measured by semi-quantitative RT-PCR and Western blotting. Cell cycle and apoptosis were analyzed by flow cytometry. Cell proliferation, adhesion and invasion were assayed using MTT, Transwell invasion, and cell adhesion assays respectively., Result: Hypoxia significantly upregulated HIF-1α protein expression and the HIF-1α target genes VEGF, GLUT1, and Survivin mRNA. HIF-1α mRNA expression was not affected by hypoxia. Transfection of the siRNA expression plasmid pRNAT-CMV3.2/Neo-HIF-1α silenced HIF-1α by approximately 80% in hypoxic WERI-Rb-1 cells. The knockdown of HIF-1α under hypoxic conditions downregulated VEGF, GLUT1, and Survivin mRNA. It also inhibited proliferation, promoted apoptosis, induced the G0/G1 phase cell cycle arrest, and reduced the adhesion and invasion of WERI-Rb-1 cells., Conclusion: HIF-1α plays a major role in the survival and aggressive phenotype of retinoblastoma cells under hypoxic conditions. Targeting HIF-1α may be a promising therapeutic strategy for human malignant retinoblastoma.

Lamellarin D (LamD) is a marine alkaloid with a pronounced cytotoxicity against a large panel of cancer cells, affecting cell growth and inducing apoptosis. However, the molecular mechanisms of action of this compound are poorly understood. In this study, the anticancer efficacy of LamD was investigated in human leukemia K562 cells. The results showed suppressed cell proliferation and induction of G0/G1-phase arrest,while expression of CDK1, and activity of smad3 and smad5 were reduced, but that of p27, p53 and STGC3 was increased. LamD induced cell apoptosis through activation of caspases-8/-3, inhibition of survivin and Bcl-2, suggesting that this compound may also act through a caspase-independent pathway. Moreover, LamD inhibited the secretion of TGF-β, IL-1β, IL-6, IL-8 and other inflammatory cytokines and the transcriptional activity of transcription factor NF-κB in human leukemia K562 cells. Taken together, our results suggest that LamD-mediated inhibition of leukemia cell proliferation may be related to the induction of apoptosis and the regulation of cell cycle, tumor- related gene expression and cytokine expression, which may provide a new way of thinking for the treatment leukemia.

This study aims to investigate whether the expression of heat shock protein 90 (HSP90) is associated with the malignant pheochromocytoma (PHEO) and the effects of 17-allylamino-17-demethoxygeldanamcyin (17-AAG) on the expression of vascular endothelial growth factor (VEGF) in PHEO cell line PC12. The expression of HSP90 was investigated in 38 paraffin-embedded samples of PHEO patients using immunohistochemistry. The time and concentration effects of 17-AAG were investigated in PHEO PC12 cells. Cell proliferation was measured by MTT assay and cell counting. Apoptosis was detected by flow cytometry. Positive staining for HSP90 was found in 14 of 17 malignant (82.35%) and in 5 of 21 (23.81%) benign PHEOs. There existed a significant statistical difference between the malignant group and the benign ones (P < 0.001). 17-AAG inhibited the proliferation of HCC cells in a time- and concentration-dependent manner. The apoptosis rates of PC12 cells after treatment with 0.1 μmol/L for 6, 12, 24, and 48 h were significantly higher than that in blank control group. 17-AAG significantly downregulated VEGF-165 protein level in PC12 cells. This study has confirmed that the specific HSP90 inhibitor 17-AAG can play a therapeutic role in malignant PHEO treatment, and HSP90 qualifies as a promising new target in malignant PHEO.

High-grade glioma is incurable and is associated with a short survival time and a poor prognosis. There are two forms of brain-derived neurotrophic factor (BDNF), proBDNF and mature BDNF, which exert opposite effects. Their diverse actions are mediated through two different transmembrane receptor signalling systems: p75NTR and TrkB. The important roles of the BDNF/TrkB signalling system in tumour cell proliferation and survival have been demonstrated. However, few studies have been able to distinguish mature BDNF from proBDNF due to the limitation of specific antibodies. Using specific proBDNF antibodies, we demonstrated that the proBDNF/p75NTR pathway appears to inhibit malignant glioma cell growth and migration. In the present study using specific mature BDNF antibodies, we found that mature BDNF inhibited C6 glioma cell apoptosis and increased cell growth and migration in vitro. Our data suggest that the counterbalance between mature BDNF and proBDNF may regulate tumour growth.

Gefitinib is the first targeted drug approved for non-small cell lung cancer (NSCLC) treatment. Clinical trails showed that patients with certain clinical and histologic characteristics (such as women, patients of East Asian descent, no history of smoking, and adenocarcinoma) had higher rates of response and overall survival. Despite excellent clinical response to gefitinib in certain NSCLC patients, nearly all patients who respond initially to gefitinib later develop drug resistance. Isothiocyanates have been shown to possess antitumor activity, inhibiting several types of cancer cells growth. However, there are limited studies on their effects on chemoresistance of cancer cells. In this report, we found that benzyl isothiocyanate (BITC) inhibited gefitinib-resistant human NSCLC cells growth by inducing apoptosis in a dose-dependent manner, and activated caspase-3. There were no effects of BITC on epidermal growth factor receptor and multidrug resistant proteins expression. BITC caused cell cycle arrest at G2/M phase, reactive oxygen species generation, and glutathione depletion. Akt activity and NFκB transcriptional activation were suppressed; mitogen-activated protein kinase and activator protein 1 (AP-1) were activated. Our results demonstrated that BITC overcame gefitinib resistance in lung cancer cells. The further understanding of the anti-resistance mechanism of BITC would contribute to establish it as a potent lead compound for the synthesis of novel anticancer drugs.

Objective: To establish rabbit eyeball rupture model by air gun in order to observe and analyze the early injury condition and reasons of retinal cell after eyeball rupture., Methods: Forty eight healthy rabbits were randomly divided into control group and 1, 3, 6, 12 and 24 h after injury groups. After anesthesia, the rabbit eyeball rupture model was established by air gun. Then the early pathological changes of rabbit retina were observed, and apoptotic index (AI), oncosis index (OI), the relationship between the expression amounts of apoptosis-related genes and AI were analyzed., Results: Obvious pathological lesion appeared in retina 6 h after injury. Irreversible damage occurred 12-24 h after injury. The results of AI and OI indicated that the OI peak appeared 6 h after injury and then gradually declined, while the AI increased with the prolongation of time, and the AI was higher than OI in 12 h after injury. Immunohistochemical results indicated that there was no obvious bcl-2 protein expression change. Compared with the control group and the 3, 6, 12 and 24 h after the injury groups, the expressions of p53 and Caspase-3 were significantly improved and peaked at 12 h (P<0.01). Positive correlation existed among p53, Caspase-3 expression amount and cell apoptosis amount., Conclusions: Apoptosis and oncosis of visual cells are the main reasons of retinal cell injury. p53 and Caspase-3 are the important factors in promoting the retinal cell apoptosis after eyeball rupture., (Copyright © 2013 Hainan Medical College. Published by Elsevier B.V. All rights reserved.)

Malignant glioma, also known as brain cancer, is the most common intracranial tumor, having an extremely high mortality and recurrence rate. The survival rate of the affected patients is very low and treatment is difficult. Hence, growth inhibition of glioma has become a hot topic in the study of brain cancer treatment. Among the various isothiocyanate compounds, it has been confirmed that benzyl isothiocyanate (BITC) can inhibit the growth of a variety of tumors, including leukemia, glioma and lung cancer, both inside and outside the body. This study explored inhibitory effects of BITC on human glioma U87MG cells, as well as potential mechanisms. It was found that BITC could inhibit proliferation, induce apoptosis and arrest cell cycling of U87MG cells. In addition, it inhibited the expression of SOD and GSH, and caused oxidative stress to tumor cells. Therefore, it is believed that BITC can inhibit the growth of U87MG cells outside the body. Its mechanism may be related to the fact that BITC can cause oxidative stress to tumor cells.

Objective: To investigate the expression and relationship of programmed cell death 5 (PDCD5) and cell apoptosis in the parotid gland after leading duct ligation in rat and elucidate the role of PDCD5 on the atophy of parotid gland., Methods: The Wistar rat model of leading duct ligation was established, and the samples of parotid gland were obtained from different time point (0, 1, 3, 5, 7, 14, 21, 30, 60, 90 and 120 d). The expression of PDCD5 protein was examined by immunohistochemistry. Cell apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL)., Results: The distribution of PDCD5 protein in normal parotid was in cytoplasm with uniformity. The expression of PDCD5 protein was significantly increased and reached the peak at 3 d (1.261 ± 0.048) following main duct ligation. PDCD5 was located both in cytoplasm and nuclear of parotid gland cells. The PDCD5 density in acinar cells was higher than that in duct cells at day 1 and 3 after duct ligation (P < 0.01). The apoptotic cells were obviously upregulated at 3 d after duct ligation. The apoptosis index observed in acinar cells [(21.750 ± 0.119)%] was more than that in duct cells [(5.720 ± 0.205)%]. The difference of apoptosis index between acinar cells and duct cells was statistically significant (P < 0.01). The increased PDCD5 levels were positively correlated with cell apoptosis induced by duct ligation., Conclusions: The expression of PDCD5 is associated with the atophy of the parotid gland after rat parotid duct ligation, indicating that PDCD5 might play an important role in apoptotic pathways after parotid duct ligation.

Objective: To explore the influence of extract of Ginkgo biloba (EGB) on the proliferation, apoptosis of ACC-2 cell and Survivin gene expression in adenoid cystic carcinoma (ACC) of lacrimal gland., Methods: ACC-2 cell in human with ACC of lacrimal gland was in vitro cultured. MTT method was used for cell proliferation detection. Annexin V/PI double-staining flow cytometer was used to detect cell apoptosis and cell cycle. Survivin gene expression was analyzed by RT-PCR and Western blotting., Results: EGB had inhibitory effect on the proliferation of ACC-2 cell with significant dose-effect relationship, and there was statistical difference when compared with the control group (P<0.01). The inhibitory concentration 50 % (IC(50)) is 88 mg/L. The flow cytometer test indicated that EGB can gradually increase ACC-2 cell in G(0)-G(1) stage and decrease it in G(2)-M and S stage. With the increase of dose, the apoptosis rate of ACC-2 cell was obviously increased (P<0.05 or P<0.01). EGB had certain inhibitory effect on Survivin gene expression of ACC-2 cell, and Survivin gene expression was decreased with the increasing of the EGB concentration (P<0.01)., Conclusions: EGB can effectively inhibit Survivin gene expression of ACC-2 cell in human with ACC of lacrimal gland, induce the apoptosis of ACC-2 cell and inhibit tumor cell proliferation., (Copyright © 2012 Hainan Medical College. Published by Elsevier B.V. All rights reserved.)

Dietary isothiocyanates have been shown to possess anti-tumour activity, inhibiting several types of cultured human cancer cell growth. However, there are limited studies on their effects on cancer cell metastasis. Our previous study showed that benzyl isothiocyanate (BITC) and phenethyl isothiocyanate (PEITC) suppressed human lung cancer cell metastasis potential. In the present study, we found BITC (7·5 and 10 μm) and PEITC (12·5 and 20 μm) induced highly metastatic human non-small cell lung cancer L9981 cell apoptosis in a dose-dependent manner. Caspase-3 was activated. They also caused cell cycle arrest at the G2/M phase, via modulation of cyclin B1 expression. The mitogen-activated protein kinase (MAPK) signalling pathway was involved. c-Jun N-terminal kinase, extracellular signal-regulated protein kinase 1/2 and p38 were activated in a dose-dependent manner; activator protein 1 (AP-1) transcriptional activation and cyclin D1 expression were repressed. Apoptosis and MAPK activation were abrogated by anti-oxidant N-acetyl cysteine (NAC), suggesting that cell death signalling was triggered by oxidative stress. Further microarray analysis evaluated the potential targeted genes related to apoptosis and the cell cycle. Our studies suggested that BITC and PEITC suppressed the metastasis potential of highly metastatic lung cancer cells by inducing apoptosis and cell cycle arrest, via targeting the MAPK/AP-1 pathway. This may provide a novel approach for metastasis therapy of lung cancer by dietary isothiocyanates and possibly other types of cancer.

Objective: To investigate the effects of ischemic postconditioning(IPo) on cellular apoptosis in acute renal ischemic-reperfusion injury (IRI) in rats., Methods: Model of acute renal ischemic-reperfusion injury (IRI) was constructed using Wistar rats. Forty healthy male rats were divided into 4 groups randomly,including group C,group IRI, group IPo and group Ado. Rats of group C received right nephronectomy and separation of left renal vessels. Before reperfusion, group IPo was perfused for 10 s, following ischemic reperfusion for another 10 s with 6 cycles. Group Ado was infused with adenosine intravenously 10 min before ischemia. Cellular apoptosis was determined with TUNEL (TdT-mediated dUTP Nick End Labelling), flowcytometry and histopathological staining., Results: Compared with group C, the apoptotic index increased predominatly in group IRI (P<0.05), whereas, apoptosis index decreased obviousely in group IPo, Ado (P<0.05). In addition, the apoptosis index between group IPo and Ado was not statistically different., Conclusion: Ischemic postconditioning could decrease cellular apoptosis induced by IRI in rat kidney.

Objective: To investigate the effects of the small interfering RNA plasmid-dextran magnetic nanoparticles (siRNA-DMN) combination with external magnetic fields on silencing survivin gene expression of bladder cancer cells and apoptosis when DMN used as gene carrier to transfer siRNA-survivin recombinant plasmid in vivo., Methods: The siRNA-survivin recombinant plasmid specific targeted survivin was synthesized in previous experiment. DMN were prepared by chemical coprecipitation method and used as gene carrier. The siRNA-DMN were constructed by static electricity of polylysine and transferred into human bladder cancer BIU-87 cells with the help of external magnetic fields. The growth inhibiting rate (IR) of BIU-87 cells was observed by 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl tetrazolium bromide's test and the apoptosis index (AI) was detected by transferase-mediated dUTP nick end labeling method. The relatively transcription levels of survivin mRNA and protein expression were respectively detected by semi-quantitive reverse transcription polymerase chain reaction and Western Blotting techniques., Results: The diameter, effective diameter and saturation magnetization of DMN-siRNA were about 10 - 12 nm, 94.8 nm and 0.19 emu/g, respectively. The IR (39.60%) and AI (28.72%), the relative expression of survivin mRNA and protein of siRNA-DMN combination with external magnetic fields on BIU-87 cells were significantly higher and lower than those in the control group and single siRNA-DMN group, respectively (P < 0.05)., Conclusions: The siRNA-survivin plasmid-DMN combination with external magnetic fields could effectively inhibit survivin expression and induce BIU-87 cells apoptosis which provided experimental basis for the magnetic targeting gene therapy of bladder tumor.

Objective: To explore the possible mechanism of apoptosis induced by photodynamic therapy (PDT) in human pancreatic cancer cells Capan-1 with 2-butylamino-2-demethoxy-hypocrellin B (BAHB) as photosensitizer., Methods: The localization of BAHB in Capan-1 cells was studied, apoptosis was determined by DNA gel electrophoresis after PDT. The mitochondria membrane potential (DYm) and cytochrome C release were observed by laser scan confocal microscopy and Western blotting., Results: The low concentration photosensitizer was mainly localized in mitochondria and also in lysosomes when the concentration is high. DNA ladder analysis showed characteristic of apoptosis. The mitochondria membrane potential (DYm) showed a loss of 30% around, after 6 hours by PDT under laser scan confocal microscopy, which is caused by a sudden increase in the permeability of mitochondria membrane accompanied with apoptosis. In Western blotting, cytochrome C release was observed from the mitochondria into the cytoplasm during BAHB-induced apoptosis., Conclusion: The research suggests that BAHB-induced apoptosis is related to photosensitization of mitochondria.

Human Kruppel-like factor 2 (KLF2) is a Cys(2)/His(2) zinc-finger-containing transcriptional factor, which is involved in multiple cellular pathways. Utilizing gene expression profiling to identify aberrantly expressed genes in ovarian cancer, we found that KLF2 was significantly and specifically downregulated in ovarian tumors. After reintroducing KLF2 into ovarian cancer cell lines, we observed decreased cell growth and increased sensitivity to DNA damage-induced apoptosis. Analysis of genes that could be potential targets of KLF2 revealed that KLF2 negatively regulated WEE1 expression. WEE1 encodes a tyrosine kinase that regulates the G2/M cell cycle transition. Expression of KLF2 markedly repressed the transcription of WEE1 by directly binding to an SP1/CPBP motif located between -252 bp and the start codon of the WEE1 promoter. Both activation and zinc-finger domains of KLF2 were required for this suppression of Wee1 expression. In addition, we demonstrated that Wee1 expression prevents cancer cells from undergoing apoptosis in response to DNA damage; however, this resistance was abolished by coexpression of KLF2, which inhibits WEE1 transcription. Thus, the level of WEE1 is regulated by KLF2 and enhanced KLF2 expression sensitizes cells to DNA damage-induced apoptosis.

Objective: To explore the inhibitory effect of Fuzheng Yiliu Granule (FYG) on growth and apoptosis of H22 tumor cell, and expressions of p53 and Caspase-3 gene., Methods: The tumor inhibitory rate of FYG on H22 tumor cell line was observed in vivo, cell apoptosis rate and cell cycle were determined by flow cytometry (FCM), and expressions of wild type p53 and Capase-3 mRNA were determined by RT-PCR., Results: FYG could inhibit the growth of H22 tumor cell at the dose of 12g/kg, 24g/kg, the maximal inhibitory rate up to 51.24% (P < 0.01). By FCM, it was shown that FYG could significantly enhance apoptosis rate of cell line H22, with the peak reached 15.84% (P < 0.01), and cause the tumor cell cycle being blocked at G0/G1 phase, with decrease of cells in S phase. RT-PCR illustrated that FYG could significantly up-regulate the level of p53 and Caspase-3 mRNA expression., Conclusion: FYG can significantly inhibit the growth of tumor cell in mice, its anti-tumor mechanisms may relate to the cell apoptosis induction, cell cycle regulation and wild type p53 and capase-3 gene expression enhancing.

Purpose: To investigate the effect of high dose methylprednisolone (MP) on retinal ganglion cells (RGC) apoptosis., Methods: One hundred and twenty six Wistar rats were divided into normal control, crush control and MP treatment groups. High dose MP and 0.9% sodium chloride were injected intravenously at 1 h after injury in the treatment group and crush control group, respectively. Apoptotic cells, Bcl-2 and Bax positive cells were located and calculated by terminal-deoxynucleotidyl transferase mediated dUTP-biotin nick end labeling (TUNEL), immunohistochemical method and retinal whole-mount combined with section technique at day 4, 7, 14 after crush, respectively., Results: The TUNEL-labeling cells, Bcl-2 and Bax positive cells were detected in the ganglion cell layer of retina. Few apoptotic cells, a few of Bcl-2 and Bax positive cells were observed in normal control groups. The numbers of positive cells increased gradually after crush. The number of apoptotic cells and Bax positive tells in treatment groups was less than that in crush group significantly (P < 0.05), while the number of Bcl-2 positive cells in treatment group was more than that in crush group at day 4, 7 and 14 after crush significantly (P < 0.05)., Conclusions: These results suggest that high dose MP can inhibit the apoptosis of RGC after optic nerve crush probably through up-regulation of Bcl-2 expression and down-regulation of Bax expression.

PPF1 encodes a putative calcium ion carrier that affects the flowering time of transgenic Arabidopsis by modulating Ca(2+) storage capacities in chloroplasts of a plant cell. In the current work, we found that differential expression of PPF1 might affect processes of programmed cell death (PCD) since DNA fragmentation was detected in senescencing apical buds of long day-grown G2 pea (Pisum sativum L.) plants, but was not in non-senescencing short day-grown counterparts at all growth stages. An animal inhibitor of caspase-activated DNase (ICAD) homologue was detected in short day-grown plant continuously throughout the whole experiment and only in early stages of long day-grown pre-floral G2 pea apical buds. DNA fragmentation was significantly inhibited in apical meristems of transgenic Arabidopsis that over-expressed the PPF1 gene when compared to that of either wild-type control or to PPF1 (-) plants. The expression of ICAD-like protein decreased to undetectable level at 45 dpg in apical tissues of PPF1 (-) Arabidopsis, which was much earlier than that found in PPF1 (+) or wild-type controls. In epidermal cells of PPF1 (-) plants, we recorded significantly earlier calcium transient prior to PCD. We suggest that the expression of PPF1, a chloroplast localized Ca(2+) ion channel may inhibit programmed cell death in apical meristems of flowering plants by keeping a low cytoplasmic calcium content that might inhibit DNA fragmentation in plant cells.

Ferroptosis is an iron-dependent programmed cell death associated with severe kidney diseases, linked to decreased glutathione peroxidase 4 (GPX4). However, the spatial distribution of renal GPX4-mediated ferroptosis and the molecular events causing GPX4 reduction during ischemia-reperfusion (I/R) remain largely unknown. Using spatial transcriptomics, we identify that GPX4 is situated at the interface of the inner cortex and outer medulla, a hyperactive ferroptosis site post-I/R injury. We further discover OTU deubiquitinase 5 (OTUD5) as a GPX4-binding protein that confers ferroptosis resistance by stabilizing GPX4. During I/R, ferroptosis is induced by mTORC1-mediated autophagy, causing OTUD5 degradation and subsequent GPX4 decay. Functionally, OTUD5 deletion intensifies renal tubular cell ferroptosis and exacerbates acute kidney injury, while AAV-mediated OTUD5 delivery mitigates ferroptosis and promotes renal function recovery from I/R injury. Overall, this study highlights a new autophagy-dependent ferroptosis module: hypoxia/ischemia-induced OTUD5 autophagy triggers GPX4 degradation, offering a potential therapeutic avenue for I/R-related kidney diseases. Understanding the role of GPX4 in cell ferroptosis at the interface of the inner cortex and medulla is crucial in the context of renal injury. Here, the authors demonstrate that the OTUD5 interaction with GPX4 is key in resisting ischemia/reperfusion-induced ferroptosis in renal cells, offering a new strategy for treating acute kidney injury. [ABSTRACT FROM AUTHOR]

Methamphetamine (METH) abuse is known to cause executive dysfunction. However, the molecular mechanism underlying METH induced executive dysfunction remains unclear. Go/NoGo experiment was performed in mice to evaluate METH-induced executive dysfunction. Immunoblot analysis of Nuclear factor-E2-related factor 2 (Nrf2), phosphorylated Nrf2 (p-Nrf2), heme-oxygenase-1 (HO-1), Glucose Regulated Protein 78(GRP78), C/EBP homologous protein (CHOP), Bcl-2, Bax and Caspase3 was performed to evaluate the levels of oxidative stress, endoplasmic reticulum (ER) stress and apoptosis in the dorsal striatum (Dstr). Malondialdehyde (MDA) levels and glutathione peroxidase (GSH-Px) activity was conducted to evaluate the level of oxidative stress. TUNEL staining was conducted to detect apoptotic neurons. The animal Go/NoGo testing confirmed that METH abuse impaired the inhibitory control ability of executive function. Meanwhile, METH down-regulated the expression of p-Nrf2, HO-1 and GSH-Px and activated ER stress and apoptosis in the Dstr. Microinjection of Tert-butylhydroxyquinone (TBHQ), an Nrf2 agonist, into the Dstr increased the expression of p-Nrf2, HO-1, and GSH-Px, ameliorated ER stress, apoptosis and executive dysfunction caused by METH. Our results indicated that the p-Nrf2/HO-1 pathway was potentially involved in mediating methamphetamine-induced executive dysfunction by inducing endoplasmic reticulum stress and apoptosis in the dorsal striatum. [ABSTRACT FROM AUTHOR]

Neurological dysfunction is a common complication of deep hypothermic circulatory arrest (DHCA). Endoplasmic reticulum (ER) stress plays a role in neuronal ischemia-reperfusion injury; however, it is unknown whether it contributes to DHCA-induced brain injury. Here, we aimed to investigate the role of ER stress in a rat DHCA model and cell hypothermic oxygen–glucose deprivation reoxygenation (OGD/R) model. ER stress and apoptosis-related protein expression were identified using Western blot analysis. Cell counting assay-8 and flow cytometry were used to determine cell viability and apoptosis, respectively. Brain injury was evaluated using modified neurological severity scores, whereas brain injury markers were detected through histological examinations and immunoassays. We observed significant ER stress molecule upregulation in the DHCA rat hippocampus and in hypothermic OGD/R PC-12 cells. In vivo and in vitro experiments showed that ER stress or activating transcription factor 6 (ATF6) inhibition alleviated rat DHCA-induced brain injury, increased cell viability, and decreased apoptosis accompanied by C/EBP homologous protein (CHOP). ER stress is involved in DHCA-induced brain injury, and the inhibition of the ATF6 branch of ER stress may ameliorate this injury by inhibiting CHOP-mediated apoptosis. This study establishes a scientific foundation for identifying new therapeutic targets for perioperative brain protection in clinical DHCA. [ABSTRACT FROM AUTHOR]

Objectives: Human papillomavirus 16 (HPV 16) E2 is a transcriptional regulator that plays a key role in regulating a variety of biological responses. Hematopoietic cell-specific protein 1-related protein X-1 (HAX-1) is a mitochondrial membrane protein, and the HAX-1 gene is involved in the occurrence, growth, invasion, and metastasis of various human malignant tumors. The purpose of this study was to investigate the relationships among HPV 16 E2, the role of HAX-1 gene, and the underlying intracellular apoptotic mechanism of human cervical squamous carcinoma cells (C33a and SiHa).Methods: In this study, HAX-1 expression was examined using real-time polymerase chain reaction, Western blot, and immunohistochemical staining analysis. Apoptosis of cells was assessed by flow cytometry. The mitochondrial function was assessed by the mitochondrial copy number, reactive oxygen species (ROS) generation, the mitochondrial membrane potential (ΔΨm), and mitochondrial morphology.Results: Our study demonstrated that the expression of the HAX-1 gene was significantly increased in human cervical carcinoma tissues relative to noncancerous cervix tissues. HPV 16 E2 inhibited HAX-1 protein expression. Overexpression of HAX-1 increased the mitochondrial copy number, decreased the production of ROS, and maintained the integrity of the mitochondrial membrane and morphology. So, enhanced expression of the HAX-1 gene could abrogate the HPV 16 E2-induced cell apoptosis.Conclusion: Therefore, these data support a mechanism that HAX-1 plays a crucial role in HPV 16 E2-induced human cervical squamous carcinoma cell apoptosis in a mitochondrial-dependent manner. [ABSTRACT FROM AUTHOR]