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DNA topoisomerases are essential nuclear enzymes in correcting topological DNA errors and maintaining DNA integrity. Topoisomerase inhibitors are a significant class of cancer chemotherapeutics with a definite curative effect. Natural products are a rich source of lead compounds for drug discovery, including anti-tumor drugs. In this study, we found that narciclasine (NCS), an amaryllidaceae alkaloid, is a novel inhibitor of topoisomerase I (topo I). Our data demonstrated that NCS inhibited topo I activity and reversed its unwinding effect on p-HOT DNA substrate. However, it had no obvious effect on topo II activity. The molecular mechanism of NCS inhibited topo I showed that NCS did not stabilize topo-DNA covalent complexes in cells, indicating that NCS is not a topo I poison. A blind docking result showed that NCS could bind to topo I, suggesting that NCS might be a topo I suppressor. Additionally, NCS exhibited a potent anti-proliferation effect in various cancer cells. NCS arrested the cell cycle at G2/M phase and induced cell apoptosis. Our study reveals the antitumor mechanisms of NCS and provides a good foundation for the development of anti-cancer drugs based on topo I inhibition. [ABSTRACT FROM AUTHOR]

Recent reports have suggested that abnormal miR-29c expression in hippocampus have been implicated in the pathophysiology of some neurodegenerative and neuropsychiatric diseases. However, the underlying effect of miR-29c in regulating hippocampal neuronal function is not clear. In this study, HT22 cells were infected with lentivirus containing miR-29c or miR-29c sponge. Cell counting kit-8 (CCK8) and lactate dehydrogenase (LDH) assay kit were applied to evaluate cell viability and toxicity before and after TNF-α administration. Reactive oxygen species (ROS) generation and mitochondrial membrane potential (MMP) were measured with fluorescent probes. Hoechst 33258 staining and TUNEL assay were used to evaluate cell apoptosis. The expression of key mRNA/proteins (TNFR1, Bcl-2, Bax, TRADD, FADD, caspase-3, -8 and -9) in the apoptosis pathway was detected by PCR or WB. In addition, the protein expression of microtubule-associated protein-2 (MAP-2), nerve growth-associated protein 43 (GAP-43) and synapsin-1 (SYN-1) was detected by WB. As a result, we found that miR-29c overexpression could improve cell viability, attenuate LDH release, reduce ROS production and inhibit MMP depolarization in TNF-α-treated HT22 cells. Furthermore, miR-29c overexpression was found to decrease apoptotic rate, along with decreased expression of Bax, cleaved caspase-3, cleaved caspase-9, and increased expression of Bcl-2 in TNF-α-treated HT22 cells. However, miR-29c sponge exhibited an opposite effects. In addition, in TNF-α-treated HT22 cells, miR-29c overexpression could decrease the expressions of TNFR1, TRADD, FADD and cleaved caspase-8. However, in HT22 cells transfected with miR-29c sponge, TNF-α-induced the expressions of TNFR1, TRADD, FADD and cleaved caspase-8 was significantly exacerbated. At last, TNF-α-induced the decreased expression of MAP-2, GAP-43 and SYN-1 was reversed by miR-29c but exacerbated by miR-29c sponge. Overall, our study demonstrated that miR-29c protects against TNF-α-induced HT22 cells injury through alleviating ROS production and reduce neuronal apoptosis. Therefore, miR-29c might be a potential therapeutic agent for TNF-α accumulation and toxicity-related brain diseases. [ABSTRACT FROM AUTHOR]

Purpose: Ferroptosis is a form of regulated cell death that has the potential to be targeted as a cancer therapeutic strategy. But cancer cells have a wide range of sensitivities to ferroptosis, which limits its therapeutic potential. Accumulation of lipid peroxides determines the occurrence of ferroptosis. However, the type of lipid involved in peroxidation and the mechanism of lipid peroxide accumulation are less studied. Methods: The effects of fatty acids (10 μM) with different carbon chain length and unsaturation on ferroptosis were evaluated by MTT and LDH release assay in cell lines derived from prostate cancer (PC3, 22RV1, DU145 and LNCaP), colorectal cancer (HT-29), cervical cancer (HeLa) and liver cancer (HepG2). Inhibitors of apoptosis, necroptosis, autophagy and ferroptosis were used to determine the type of cell death. Then the regulation of reactive oxygen species (ROS) and lipid peroxidation by docosahexaenoic acid (DHA) was measured by HPLC–MS and flow cytometry. The avtive form of DHA was determined by siRNA mediated gene silencing. The role of lipoxygenases was checked by inhibitors and gene silencing. Finally, the effect of DHA on ferroptosis-mediated tumor killing was verified in xenografts. Results: The sensitivity of ferroptosis was positively correlated with the unsaturation of exogenously added fatty acid. DHA (22:6 n-3) sensitized cancer cells to ferroptosis-inducing reagents (FINs) at the highest level in vitro and in vivo. In this process, DHA increased ROS accumulation, lipid peroxidation and protein oxidation independent of its membrane receptor, GPR120. Inhibition of long chain fatty acid-CoA ligases and lysophosphatidylcholine acyltransferases didn't affect the role of DHA. DHA-involved ferroptosis can be induced in both arachidonate lipoxygenase 5 (ALOX5) negative and positive cells. Down regulation of ALOX5 inhibited ferroptosis, while overexpression of ALOX5 promoted ferroptosis. Conclusion: DHA can effectively promote ferroptosis-mediated tumor killing by increasing intracellular lipid peroxidation. Both ALOX5 dependent and independent pathways are involved in DHA-FIN induced ferroptosis. And during this process, free DHA plays an important role. [ABSTRACT FROM AUTHOR]

Glioma is a highly malignant type of intracranial tumor with a poor prognosis resulting from traditional chemo-resistance with temozolomide (TMZ). Luteolin has been detected to exert limited anti-tumor effects on gliomas, while valproic acid (VPA) is a common chemotherapy sensitizer in the treatment of tumors. In this study, three glioma cell lines including U251, LN229 and SNB19 were selected for evaluation of combined anti-tumor effects of VPA and luteolin via Cell Counting Kit-8 (CCK-8) assay, colony formation assay, wound-healing assay, flow cytometry and western blot assay. The results disclosed that VPA sensitized glioma cells to luteolin by repressing cell viability, colony formation and migration. Mechanically, VPA boosted cellular apoptosis and cell-cycle arrest by increased level of cleaved caspase-3/caspase-3, cleaved PARP/PARP and Bax/Bcl-2. In addition, VPA also facilitated cellular autophagy via the decline of p62, p-Akt/Akt and the accumulation of LC3-II. These findings suggested that VPA enhanced the anticancer effects of luteolin by strengthening apoptosis and autophagy via Akt signaling, which could be adopted as a novel therapy for glioma. [ABSTRACT FROM AUTHOR]

Acute myocardial infarction (AMI) is the main cause of death in the whole world. This study aimed to investigate whether forkhead box O4 (FoxO4) could negatively modulate ubiquitin specific peptidase 10 (USP10) transcription to aggravate the apoptosis and oxidative stress of hypoxia/reoxygenation (H/R)-induced cardiomyocytes through Hippo/YAP pathway. mRNA expression as well as protein expressions of USP10 and FoxO4 in H9C2 cells after H/R induction or transfection were respectively detected by Reverse transcription-quantitative (RT-q) PCR analysis and Western blot. The viability and apoptosis of H9C2 cells after H/R induction or transfection were respectively detected by CCK-8 and TUNEL assays. The expressions of lactate dehydrogenase (LDH), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and malondialdehyde (MDA) in H9C2 cells after H/R induction or transfection were analyzed using appropriate kits and intracellular reactive oxygen species (ROS) levels were detected using a ROS Assay Kit. Dual luciferase reporter assay and Chromatin Immunoprecipitation (ChIP) have adopted to confirm the combination of USP10 and FoxO4. Western blot was also used to analyze the expression of apoptosis-related proteins and Hippo/YAP pathway-related proteins. As a result, USP10 expression was decreased in H/R-induced H9C2 cells in a time-dependent manner. USP10 overexpression increased the viability and suppressed the apoptosis and oxidative stress of H/R-induced H9C2 cells. In addition, FoxO4 modulated USP10 transcription. FoxO4 expression was increased in H9C2 cells induced by H/R. FoxO4 overexpression could reverse the protective effects of USP10 overexpression on H/R-induced H9C2 cells by regulating the Hippo/YAP signaling pathway. In conclusion, FoxO4 negatively modulated USP10 transcription to aggravate the apoptosis and oxidative stress of H/R-induced H9C2 cells via blocking Hippo/YAP pathway. [ABSTRACT FROM AUTHOR]

Type 1 diabetes is characterized by an increase in blood glucose levels resulting from damage to β cells in pancreatic islets and the consequent absolute insufficiency of insulin. Animal models of type 1 diabetes were usually established using drugs toxic to β cells, such as streptozotocin (STZ). To assess the application of zebrafish larvae in diabetes research, we explore the effects of STZ on pancreatic islets and glucose metabolism in zebrafish larvae. STZ was microinjected into the pericardial cavity of zebrafish larvae on alternate days for three times. At 2 days after the whole series of STZ injection (12 dpf), free-glucose level in larvae tissue shows a significant increase, and the fluorescence signal in immunohistochemistry, which indicates the insulin expression, was significantly weaker compared with the solution-injected control. Obvious apoptosis signals were also observed in the location of pancreatic islet, and insulin content decreased to be undetectable in STZ-injected larvae. Gene expression level of ins decreased to half of the solution injection control and that of casp3a was upregulated by 2.20-fold. Expression level of glut2 and gck decreased to 0.312-fold and 0.093-fold, respectively. pck1 was upregulated by 2.533-fold in STZ-injected larvae. By tracking detection, we found the free-glucose level in STZ-injected larvae gradually approached the level of the solution injection control and the insulin content recovered at 6 days post-STZ injection (16 dpf). Consistent with the change of the glucose level, the regeneration rate of the caudal fin in the STZ-injected group decreased initially, but recovered and accelerated gradually finally at 8 days post-amputation (20 dpf). These results indicate the generation of a transient hyperglycemia model due to β-cell apoptosis caused by STZ, which is abated by the vigorous regeneration ability of β cells in zebrafish larvae. [ABSTRACT FROM AUTHOR]

Objective: Malignant glioma is a lethal brain tumor with a low survival rate and poor prognosis. New strategies are urgently needed to augment the chemotherapeutic effects of temozolomide (TMZ), the standard drug in glioma treatment. Carnosic acid (CA) has been reported to have anticancer, antioxidant and anti-infectious properties. In this study, we aimed to investigate the anticancer effects and the underlying mechanisms of CA in combination with TMZ in glioma cancer cells.Methods: The glioma cancer cells were treated with TMZ, CA, or TMZ + CA. We evaluated cell survival by CCK-8 assay, cell anchorage-independent survival by colony formation assay, cell migration by wound-healing assay, cell cycle and cell apoptosis by flow cytometry, and protein expression by western blot.Results: CA enhanced the cytotoxic effect of TMZ in glioma cancer cells. CA enhanced TMZ-induced inhibition of colony formation and cell migration and enhanced TMZ-induced cell cycle arrest and cellular apoptosis. Immunofluorescence suggested that CA in combination with TMZ triggered autophagy. Furthermore, CA promoted TMZ-induced cell cycle arrest and cellular apoptosis by Cyclin B1 inhibition and activation of PARP and Caspase-3, while CA promoted TMZ-induced cellular autophagy by p-AKT inhibition, p62 downregulation and LC3-I to LC3-II transition.Conclusion: These data suggest that the combination therapy of CA and TMZ strengthens the anticancer effect of TMZ by enhancing apoptosis and autophagy. [ABSTRACT FROM AUTHOR]

Polycystic ovary syndrome (PCOS) is a common endocrine disorder affecting women of reproductive age. Insulin resistance/hyperinsulinemia is a prevalent finding in women with PCOS, which indicates that insulin resistance/hyperinsulinemia may be an important player in the pathogenesis of the PCOS. However, the underlying mechanism of insulin resistance/hyperinsulinemia on the pathogenesis of the PCOS remains elusive. In this study, we found an increased high-mobility group box 1 (HMGB1) in the serum from women with PCOS having insulin resistance/hyperinsulinemia. Furthermore, we discovered that high concentration of insulin, which mimics insulin resistance model, promoted apoptosis in primary cultured rat ovarian granulosa cells (GCs) via its effect on the increase in extracellular HMGB1. Our data presented the first evidence that increased HMGB1 induced by insulin resistance/hyperinsulinemia promoted apoptosis of ovarian GCs, which provided new molecular basis for the PCOS pathogenesis. [ABSTRACT FROM PUBLISHER]

Intestinal ischemia reperfusion (IR) causes injury of distant critical organs. Remote intestinal ischemic preconditioning (IP) may confer the cytoprotection in critical organs including lung. The authors hypothesized that intestinal IP would be a prophylactic factor in the prevention of distant lung injury induced by IR. Rats were randomly divided into IR, IP, and Sham (S) group. Compared with IR group in the serum and lung tissue, MPO, MDA, TNF-α, and IL-1 levels were significantly decreased in the IP group. Following the same pattern, NO level in the serum and lung tissue was significantly increased in the IP group. And intestinal IP markedly abolished lung injury scores in contrast to IR group. Moreover, intestinal IP significantly attenuated caspase-3 expression, leading to the low expression of Bax and the high expression of Bcl-2. The present study showed that intestinal IP ameliorates the capacity of anti-oxygen free radical, inhibits the release of pro-inflammatory cytokines and alleviates apoptosis in IR-induced lung injury in rats. Intestinal IP may provide a novel prophylactic strategy for treatment of IR-induced lung injury. [ABSTRACT FROM AUTHOR]

Data have increasingly shown that interlukin-24 (IL-24) has growth suppression activity and can induce apoptosis in a broad spectrum of tumor cells. However, the therapeutic effect of IL-24 on human neuroblastoma has rarely been explored. In this study, we used a human neuroblastoma cell line (SH-SY5Y) to reveal the effect of adenovirus-mediated IL-24 (Ad-IL24) gene therapy for neuroblastoma. We showed that Ad-IL24 effectively inhibited the proliferation of SH-SY5Y cells in vitro by conspicuously inducing apoptosis. To further explore the molecular mechanism by which Ad-IL24 induced apoptosis in SH-SY5Y tumor cells, we found that Ad-IL24 increased the expression of Bax and promoted the activation of caspase-3, while decreasing Bcl-2 levels. We also demonstrated that Ad-IL24 significantly inhibited tumor growth in vivo in a xenograft neuroblastoma tumor in athymic nude mice. In summary, Ad-IL24 overexpression exerted potent antitumor activity via inducing apoptosis in neuroblastoma cells. Therefore, IL-24 has the potential to serve as an agent for gene therapy in the treatment of neuroblastoma. [ABSTRACT FROM AUTHOR]

Isoliensinine, liensinine and neferine are major bisbenzylisoquinoline alkaloids in the seed embryo of lotus (Nelumbo nucifera), and exhibit potential anti-cancer activity. Here, we explored the effects of these alkaloids on triple-negative breast cancer cells and found that among the three alkaloids isoliensinine possesses the most potent cytotoxic effect, primarily by inducing apoptosis. Interestingly, isoliensinine showed a much lower cytotoxicity against MCF-10A, a normal human breast epithelial cell line. Further studies showed that isoliensinine could significantly increase the production of reactive oxygen species (ROS) in triple-negative breast cancer cells, but not in MCF-10A cells. The isoliensinine-induced apoptosis could be attenuated by radical oxygen scavenger N-acetyl cysteine, suggesting that the cytotoxic effect of isoliensinine on cancer cells is at least partially achieved by inducing oxidative stress. We found that both p38 MAPK and JNK signaling pathways were activated by isoliensinine treatment and contributed to the induction of apoptosis. Furthermore, inhibitors or specific siRNAs of p38 MAPK and JNK could attenuate apoptosis induced by isoliensinine. However, only the p38 inhibitor or p38-specific siRNA blocked the elevation of ROS in isoliensinine-treated cells. Our findings thus revealed a novel antitumor effect of isoliensinine on breast cancer cells and may have therapeutic implications. [ABSTRACT FROM AUTHOR]