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To explore the effects and the possible mechanism of bone marrow mesenchymal stem cell (BMMSC) transplantation on apoptosis in rats cerebral cortex after cardiac arrest/cardiopulmonary resuscitation (CA/CPR).The BMMSC of 2 Sprague-Dawley (SD) rats aged 4-5weeks was extracted, and the 3rd passage was used in experimental study. Eighteen Sprague-Dawley (SD) rats were divided into sham group, model group (CA/CPR group) and intervention group (BMMSC group) according to random number table method, with 6 rats in each group. CPR was performed 6 minutes after asphyxia induced CA. In sham group, CA was not induced except performing general surgical procedure. At 1 hour after return of spontaneous circulation (ROSC), 0.5 mL phosphate buffered saline (PBS) was injected through tail vein in CA/CPR group. 2×10At 3 days after CPR, compared with sham group, the apoptosis of cerebral cortex cells was increased and brain damage was obvious, NDS score was decreased significantly (56.6±5.5 vs. 80.0±0.0, P0.05), and serum S100B was increased markedly (ng/L: 45.1±4.7 vs. 19.1±1.4, P0.05), apoptosis rate of cerebral cortex cells increased significantly [(52.9±11.8)% vs. (10.1±1.5)%, P0.05], the level of active caspase-8 expression in cerebral cortex was significantly higher (caspase-8/GAPDH: 0.689±0.047 vs. 0.330±0.108, P0.05), and there was no significant difference in active caspase-9 protein expression (caspase-9/GAPDH: 0.428±0.014 vs. 0.426±0.021, P0.05) in CA/CPR group. After BMMSC transplantation, GFP-labeled BMMSC were primarily detected in cerebral cortex, compared with CA/CPR group, the apoptosis of cerebral cortex cells and brain injury were significantly improved in BMMSC group, NDS score increased significantly (70.6±2.1 vs. 56.6±5.5, P0.05), serum S100B levels in BMMSC group were lower (ng/L: 32.0±3.2 vs. 45.1±4.7, P0.05), apoptosis rate of cerebral cortex cells decreased significantly [(31.1±3.4)% vs. (52.9±11.8)%, P0.05], and the active caspase-8 expression in cerebral cortex in BMMSC group was significantly decreased (caspase-8/GAPDH: 0.427±0.067 vs. 0.689±0.047, P0.05). The active caspase-9 expression in cerebral cortex in BMMSC group and CA/CPR group were not significantly different (caspase-9/GAPDH: 0.431±0.022 vs. 0.428±0.014, P0.05).BMMSC transplantation can alleviate rat brain damage after CA/CPR possibly by inhibiting the death receptor mediated apoptotic pathway to inhibit the apoptosis of brain cells.

To investigate the damage of rat alveolar type II epithelial cells(RLE-6 TN) caused by air fine particulate matter(PM_(2.5)) and its related mechanism.PM_(2.5) in the atmosphere of Weifang City in 2020 was collected and cell culture medium was used to prepare particulate suspension. RLE-6 TN cells were exposed to different concentrations(25, 50, 100, 200, 400 μg/mL) of particulate matter suspensions for 24 h. The morphological changes of RLE-6 TN cells were observed under inverted microscope, and the cell viability was determined by MTT method. The concentration of lactate dehydrogenase(LDH) in cell supernatant was determined by microplate method. DCFH-DA, Annexin V-FITC/PI, JC-1 probe and laser confocal fluorescence intensity were used to determine the levels of reactive oxygen species(ROS), apoptosis and mitochondrial membrane potential. Total superoxide dismutase(T-SOD), glutathione(GSH) and malondialdehyde(MDA) contents and activity levels in cells were determined by colorimetric method. Caspase-3 and Caspase-9 kit were used to detect the relative expression activity of apoptosis proteins.PM_(2.5) could lead to morphological changes of RLE-6 TN cells, enlarged cell space and decreased cell viability. Compared with the control group, there were statistically significant differences in each dose group(Plt;0.05). LDH concentration in the supernatant of ≥50 μg/mL infected group increased, and LDH concentration was ≥(377.82±29.84), which was significantly different from that of the control group(278.51±23.76)(Plt;0.05). The result of laser confocal detection of ROS showed that the intracellular green fluorescence increased gradually in the ≥50 μg/mL group, and the relative fluorescence intensity was ≥(2.77±0.18), which was statistically significant compared with the control group(Plt;0.05). The level of apoptosis was significantly increased compared with the control group(Plt;0.05). The level of mitochondrial membrane potential decreased gradually, and the level of mitochondrial membrane potential in the ≥25 μg/mL group was ≤(4.22±0.45), which was statistically different from that in the control group(6.16±0.49)(Plt;0.05). PM_(2.5) could reduce the levels of T-SOD and GSH, and the levels of T-SOD and GSH in ≥50 μg/mL exposed group were ≤(14.67±0.49) and ≤(433.29±39.24), respectively, significantly lower than those in control group((16.58±0.60) and(542.90±45.06))(Plt;0.05). MDA level increased with the increase of PM_(2.5) concentration. Compared with the control group(1.15±0.19), MDA level in ≥50 μg/mL exposed group was ≥(1.72±0.13), with statistical significance(Plt;0.05). The activity levels of Caspase-3 and Caspase-9 increased in ≥100 μg/mL group, and the activity levels were ≥(1.62±0.27) and ≥(1.23±0.06), respectively, compared with the control group, the differences were statistically significant(Plt;0.05).Exposure to a certain concentration of PM_(2.5) can induce oxidative stress of rat alveolar type II epithelial cells, reduce the membrane potential, and eventually lead to cell apoptosis.

To investigate the effect and mechanism of artesunate (ARTS) combined with cytarabine(Ara-C) and/or daunorubicin (DNR) on the proliferation and apoptosis of MV4-11 human mixed-lineage leukemia rearranged（MLL-r） acute myeloid leukemia (AML) cell line.CCK-8 assay was used to detect the proliferation effect of individual or in combination of ARTS, DNR, Ara-C on MV4-11 cells. The ICThe inhibition effect of ARTS, Ara-C and DNR on the proliferation of MV4-11 were all dose-dependently (The in vitro study showed that tri-combination of ARTS+Ara-C+DNR and bi-combination of ARTS+Ara-C could inhibit the proliferation and promote apoptosis of MV4-11 cell line. The inhibition effect of these two combinations were significantly superior to that of the traditional Ara-C+DNR treatment. The mechanism underlying this finding may be identified by the down regulation of Caspase-3, while no altered expression was observed of Caspase-9, DR4 and DR5.青蒿酯联合阿糖胞苷±柔红霉素对MLL基因重排白血病细胞株MV4-11凋亡的影响及其机制研究.探讨青蒿酯（ARTS）联合柔红霉素（DNR）±阿糖胞苷（Ara-C）对MLL基因重排（MLL-r）急性髓系白血病(AML)细胞株MV4-11细胞增殖、凋亡的影响及作用机制.采用CCK-8法检测ARTS、DNR、Ara-C单药及联用对MV4-11细胞增殖率及计算单药的ICARTS、Ara-C、DNR均呈浓度依赖性抑制MV4-1l细胞增殖(体外研究显示ARTS+DNR+Ara-C、ARTS+Ara-C方案协同抑制MV4-11细胞增殖和诱导MV4-11细胞凋亡作用均强于传统方案Ara-C+DNR，其作用机制可能是通过下调Caspase-3表达发挥作用,对Caspase-9、DR4、DR5的表达无影响.

This study aims to investigate the effect of ethoxysanguinarine(Eth) on cisplatin(DDP)-resistant human gastric cancer cells and decipher the underlying mechanism. The human gastric cancer cell line SGC7901 and the DDP-resistant cell line SGC7901/DDP were used as the cell models. Western blot was employed to determine the expression levels of multidrug resistance-related proteins, and methyl thiazolyl tetrazolium(MTT) assay to detect the proliferation of SGC7901 and SGC7901/DDP cells exposed to DDP. After treatment with different concentrations of Eth, the proliferation of SGC7901 and SGC7901/DDP cells was detected by MTT assay, trypan blue exclusion assay, colony formation assay, and high-content imaging and analysis system. The apoptosis of SGC7901/DDP cells was detected by flow cytometry with Annexin V-FITC/PI staining. GFP-LC3 transfection was carried out to detect the effect of Eth on the autophagy of SGC7901/DDP cells. The expression levels of the multidrug resistance-related protein P-glycoprotein(P-gp), the apoptosis-related proteins [caspase-9, caspase-3, and poly(ADP-ribose) polymerase(PARP)], the autophagy-related protein light chain 3-Ⅱ(LC3-Ⅱ), the key effectors [mammalian target of rapamycin(mTOR), 70 kDa ribosomal protein S6 kinase(P70 S6 K), and 4 E binding protein 1(4 E-BP1)] of the mammalian target of rapamycin complex 1(mTORC1) signaling pathway, cancerous inhibitor of protein phosphatase 2A(CIP2A), and protein kinase B(Akt) were measured by Western blot. The mRNA level of CIP2A in the SGC7901/DDP cells exposed to Eth for 24 h was analyzed by RT-qPCR. After SGC7901/DDP cells were transfected with CIP2A expression vector pcDNA3.1-HA-CIP2A and treated with different concentrations of Eth, MTT assay was used to determine the prolife-ration of SGC7901/DDP cells and Western blot to detect the expression levels of related proteins. The interaction sites of Eth and CIP2A were predicted by molecular docking. The affinity between Eth and CIP2A was determined by drug affinity responsive target stability(DARTS) assay. The pharmacokinetic properties and drug-like activity of Eth were predicted by SwissADME. The results indicated that SGC7901/DDP cells were more sensitive to Eth than SGC7901 cells. Eth significantly inhibited proliferation and colony formation and changed the morphology, roundness, and area of SGC7901/DDP cells. Eth treatment caused the nucleus shrinking and significantly increased the apoptosis rate of the cells. Furthermore, Eth down-regulated the expression of caspase-9 and caspase-3 precursors and promoted the cleavage of PARP, which suggested that Eth induced the apoptosis of SGC7901/DDP cells. The GFP-LC3 in Eth-treated cells showed speckled aggregation. The up-regulated expression of LC3-Ⅱ by Eth indicated that Eth activated the autophagy of SGC7901/DDP cells. Eth down-regulated the expression of P-gp, the phosphorylation of mTOR, P70 S6K, and 4E-BP1, the expression of CIP2A, and the phosphorylation of Akt. Additionally, it increased the activity of PP2A, and had no significant effect on the expression of CIP2A in SGC7901/DDP cells. CIP2A overexpression antagonized the inhibition of cell proliferation and the activation of autophagy by Eth. Molecular docking suggested that Eth bound to CIP2A. The results of DARTS assay further proved the above binding effect. Eth has potential drug-like activity. The above results demonstrated that Eth inhibited the proliferation, induced the apoptosis, and activated the autophagy of SGC7901/DDP cells by targeting CIP2A and then down-regulating PP2A/mTORC1 signaling pathway. This study provided a new target for the treatment of cisplatin-resistant gastric cancer.

Acquired and primary resistance to epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) is still the bottleneck of clinical treatment of advanced non-small cell lung cancer (NSCLC). STE029 is a novel anticancer drug which consists of 3-hydroxy-3-methylglutarylcoenzyme A reductase (HMGCR) inhibitor and novel cancer cell membrane targeting molecular. This study aimed to investigate the reversal mechanism of EGFR-TKI resistance by STE029 in lung adenocarcinoma.CCK8 test was used to test the cell viability and survival rate of EGFR mutated PC9 cell (Gefitinib sensitive), PC9/BB4 cell (acquired Gefitinib resistant), and EGFR wild type A549 cell after treatment of STE029, Gefitinib or combination of both. EdU test was applied to detect changes in cell cycle and Hoechst 33258 was applied to detect apoptosis rate in overcoming the EGFR-TKI resistance. The activity of EGFR/PI3K/Akt, cell cycle and apoptosis signal pathways were examined. In vivo, nude mice were exposed to STE029, Gefitinib and STE029+Gefitinib for 5 wk. And the the tumor volume was measured and tumor weight was obtained on the last day.(1) PC9 cells was highly sensitive to Gefitinib, while PC9/BB4 and A549 cell showed significant resistance to Gefitinib treatment; (2) STE029+Gefitinib treatment could significantly decrease the 50% inhibitory concentrarion (IC₅₀) of Gefitinib in PC9, PC9/BB4 and A549 cells (P0.05, respectively); (3) In PC9 and PC9/BB4 cells, STE029+Gefitinib can block cell cycle and inhibit cell proliferation (P0.001), while there was no significant difference in apoptosis rate among three drug intervention groups (P0.05); However, apoptosis rate was increased in STE029+Gefitinib group in A549 cell (P0.01), while no significance detected in cell proliferation (P0.05). (4) In PC9 and PC9/BB4 cells, the combination of STE029 and Gefitinib could downregulate p-EGFR, p-Akt, p-Cyclin D1 and Cyclin D1 (P0.001), and upregulate the expression of GSK-3β (P0.001), and the expression of cleaved caspase-8, caspase-8 cleaved caspase-9, caspase-9 showed no difference among groups (P0.05). In A549 cells, the combination of STE029 and Gefitinib could downregulate p-Akt (P0.001) and upregulate cleaved caspase-8 and cleaved caspase-9 (P0.001); (5)In vivo, the combination of STE029 and Gefitinib effectively inhibited tumor development and progression compared to STE029 alone or Gefitinib alone, with significant difference (P0.05) in PC9 and PC9/BB4 xenografted tumor.STE029 could sensitize Gefitinib by inhibiting EGFR/PI3K/Akt pathway, blocking the tumor cell cycle and proliferation and inducing apoptosis through caspase-8 and caspase-9 dependent pathway. STE029 deserves further investigations in overcoming EGFR-TKI resistance in lung cancer.【中文题目：STE029逆转肺腺癌EGFR-TKI耐药 及其机制研究】 【中文摘要：背景与目的 表皮生长因子受体酪氨酸激酶抑制剂（epidermal growth factor receptor-tyrosine kinase inhibitor, EGFR-TKI）获得性耐药和原发性耐药至今仍是临床治疗晚期非小细胞肺癌（non-small cell lung cancer, NSCLC）的瓶颈。STE029是一种同时具有羟甲基戊二酸单酰辅酶A还原酶（3-hydroxy-3-methylglutarylcoenzyme A reductase, HMGCR）抑制剂抗肿瘤作用和肿瘤特异性细胞膜靶向功能的新型抗肿瘤药物。本研究旨在探讨STE029逆转肺腺癌EGFR-TKI的耐药机制。方法 STE029、吉非替尼单药及联合用药分别处理PC9、PC9/BB4、A549细胞，检测各细胞株的活性变化、细胞增殖、细胞凋亡，鉴定EGFR/PI3K/Akt信号通路、细胞周期及凋亡相关蛋白的表达；同时观察STE029、吉非替尼单药及联合用药对PC9、PC9/BB4裸鼠皮下移植瘤生长的影响。结果 ①PC9细胞为EGFR突变的敏感细胞，PC9/BB4细胞为EGFR突变的获得性耐药细胞，A549细胞为非EGFR突变的耐药细胞；②STE029联合吉非替尼作用于PC9、PC9/BB4、A549细胞后，吉非替尼的半数抑制浓度（50% inhibitory concentration, IC₅₀）值较对照组均显著降低（P0.05）；③STE029联合吉非替尼可抑制PC9、PC9/BB4细胞的增殖（P0.001），诱导A549细胞凋亡增加（P0.01）；④在PC9、PC9/BB4细胞中，联合用药组较单药组p-EGFR、p-Akt的表达明显下调（P0.000,1），GSK-3β表达升高（P0.001），其下游p-Cyclin D1及Cyclin D1表达明显降低（P0.001），cleaved caspase-8、caspase-8、cleaved caspase-9、caspase-9的表达在各给药组间未见明显差异（P0.05）；在A549细胞中，联合用药组p-Akt表达明显下调（P0.001），cleaved caspase-8、cleaved caspase-9表达均升高（P0.001），而GSK-3β、p-Cyclin D1、Cyclin D1、caspase-8、caspase-9的蛋白表达在各给药组间则无明显差异（P0.05）；⑤裸鼠体内，联合用药组PC9皮下移植瘤的生长明显受到抑制，差异有明显统计学意义（P0.01）；联合用药组PC9/BB4皮下移植瘤的生长速率亦明显降低（P0.05）。结论 STE029可在体内外明显增加非EGFR-T790M突变耐药的人肺腺癌细胞对吉非替尼的敏感性，其机制可能与STE029通过EGFR/PI3K/Akt信号通路调节GSK-3β、Cyclin D1的表达、阻滞细胞增殖、诱导细胞凋亡有关。 】 【中文关键词：STE029；EGFR-TKI；耐药；细胞凋亡；细胞周期；肺腺癌】.

OBJECTIVE: To investigate the effects of matrine combined with LY294002 on proliferation, apoptosis and cell cycle of human myeloid leukemia K562 cells and explore the underlying mechanism. METHODS: The effects of different concentrations of matrine alone and in combination with LY294002 on the proliferation of K562 cells were examined with CCK-8 assay. The changes in morphology of K562 cells were observed following treatment for 48 h with 0.4 g/L matrine and 10 μmol/L Y294002, either alone or in combination, and cell apoptosis was detected using flow cytometry with annexin V-FITC/PI double labeling; the changes in cell cycle was detected with PI labeling. Western blotting was performed to examine the effect of matrine combined with LY294002 on expressions of p-mTOR, p-PI3K, Akt, p-Akt, cyclinD1, Bcl-2 and caspase-9 in the cells. RESULTS: Treatment with different concentrations of matrine, both alone and in combination with LY294002, inhibited the proliferation of K562 cells in a time- and concentration-dependent manner. Compared with matrine treatment alone, the combined treatment caused more obvious morphological changes of the cells, significantly increased cell apoptosis (P < 0.01), and induced cell cycle arrest in G(0)/G(1) (P < 0.01). Western blotting showed that the protein expression levels of p-mTOR, cyclinD1, p-PI3K, p-Akt and Bcl-2 in K562 cells increased while the expression level of caspase-9 decreased significantly after the combined treatment (P < 0.01). CONCLUSION: Matrine combined with LY294002 produces a synergistic inhibitory effect on K562 cells possibly by down-regulating the p-Akt expression in PI3K/Akt signaling pathway, reducing the expressions of p-mTOR, cyclinD1 and Bcl-2, and increasing the expression of caspase-9.

To investigate the therapeutic effect of acupuncture stimulation combined with administration of"Gushen Zhuyu Tang"(decoction for consolidating kidney to eliminate blood stasis, DCKEBS) in the treatment of lumbar disc herniation (LDH) patients.A total of 147 patients with LDH were randomly divided into DCKEBS, acupuncture and DCKEBS+acupuncture groups (After the treatment, the VAS and M-JOA scores, contents of serum TNF-α, MMP-2, Caspase-3 and Caspase-9 were significantly decreased (Acupuncture combined with DCKEBS can relieve pain, improve lumbar muscle strength and lumbar movement function, and reduce serum TNF-α, MMP-2, Caspase-3 and Caspase-9 levels in LDH patients.

Neuroglial apoptosis and neuroinflammation play an important role in epileptogenesis. The aim of this study is to evaluate neuronal and glial apoptosis in association with neuroinflammation in brain epileptic focus and inflammatory changes in blood in patients with focal drug-resistant epilepsy (DRE). Pathological changes in the temporal lobe in epilepsy (histology, transmission electron microscopy), levels of apoptotic and neuroinflammatory proteins: active caspase-3 (immunohistochemistry), full-length form caspase-3, caspase-9, FAS, FAS-L, NF-kB, TNF-α, p53 (Western blot), and cytokine levels in blood: IL-1β, IL-2, IL-4, IL-7, TNF-α, etc. (multiplex analysis) were studied. In the present work, ultrastructural and immunohistochemical apoptotic signs were found in neurons and oligodendrocytes in the temporal lobe of DRE patients. Levels of proinflammatory cytokines that play a role in apoptosis (TNF-α, FAS, NF-kB) were increased. The blood concentration of IL-4, IL-7, TNF-α is increased and IL-2 is reduced. Oligodendroglial apoptosis has been shown to play an important role in DRE pathogenesis and to explain demyelination. Thus, a comprehensive analysis of revealed changes in the blood and brain in DRE patients showed the neuroinflammation in the epileptic focus, which was combined with the development of apoptosis of glial cells and neurons. This creates conditions for the development of drug resistance and the epilepsy progression.

Legumain is a lysosomal cysteine protease with strict specificity for cleaving after asparagine residues. By sequence comparison, legumain belongs to MEROPS clan CD of the cysteine proteases, which indicates its structural and mechanistic relation to caspases. Contrasting caspases, legumain harbors a pH-dependent ligase activity in addition to the protease activity. Although we already have a significant body of knowledge on the catalytic activities of legumain, many mechanistic details are still elusive. In this study, we provide evidence that extended active site residues and substrate conformation are steering legumain activities. Biochemical experiments and bioinformatics analysis showed that the catalytic Cys189 and His148 residues are regulated by sterically close Glu190, Ser215 and Asn42 residues. While Glu190 serves as an activity brake, Ser215 and Asn42 have a favorable effect on legumain protease activity. Mutagenesis studies using caspase-9 as model enzyme additionally showed that a similar Glu190 activity brake is also implemented in the caspases. Furthermore, we show that the substrate’s conformational flexibility determines whether it will be hydrolyzed or ligated by legumain. The functional understanding of the extended active site residues and of substrate prerequisites will allow us to engineer proteases with increased enzymatic activity and better ligase substrates, with relevance for biotechnological applications.

Atractylodes is the dry root of atractylodes macrocephala koidz and has been commonly used as a traditional Chinese medicine (TCM). Atractylenolide III, a main component of atractylodes, has displayed significant effects on anti-inflammation and anticancer. However, the effects of atractylenolide III on growth inhibition and apoptosis induction in colon cancer remain unclear. The results showed that atractylenolide III significantly inhibited the cell growth and induce cellular apoptosis in HCT-116 cells in a concentration dependence manner in vitro. Mechanistic studies further showed that atractylenolide III could regulate the Bax/Bcl-2 apoptotic signaling pathway through promoting the expression of proapoptotic related gene/proteins Bax, caspase-9 and caspase-3 but inhibiting the expression of antiapoptotic related gene/protein Bcl-2 in HCT-116 cells. Furthermore, atractylenolide III also significantly inhibited the tumor growth of HCT-116 tumor xenografts bearing in nude mice through inducing apoptosis by upregulation of the expressions of Bax, cleaved caspase-3 and p53 but downregulation of the expressions of Bcl-2 in HCT-116 tumor tissues in vivo. The studies may provide the scientific rationale for the understanding of the anticancer effect of atractylenolide III. Therefore, atractylenolide III may have the potential to be developed as a promising novel anticancer agent for the treatment of colorectal cancer clinically.

Effective cancer therapy can be achieved by using nano-drug delivery systems which provide a targeted drug delivery strategy by overcoming the drawbacks of conventional treatments like chemotherapy and radiation. ZnO nanoparticles are a potent anticancer agent that causes tumor cell destruction with the targeted drug delivery. In this present study, green synthesis of ZnO nanoparticles has been done using the plant Solanum nigrum. The synthesized ZnO nanoparticles were studied by the characterization techniques like UV-visible spectroscopy, SEM, TEM, DLS, zeta potential, FTIR, and XRD. The synthesized ZnO nanoparticles of Solanum nigrum exhibited a significant anticancer activity against HeLa cell lines through the apoptotic pathway. The cytotoxicity of ZnO nanoparticles was assessed using MTT assay, wound healing assay, DAPI staining, and acridine orange and ethidium bromide double staining. The expression patterns of β-catenin, p53, caspase-3, and caspase-9 were analyzed using reverse transcriptase-PCR. The results obtained from the study indicate that the ZnO nanoparticles of Solanum nigrum possess a dose-dependent cytotoxic effect against HeLa cell lines through the inhibition of β-catenin and increasing the levels of p53, caspase-3, and caspase-9.

Stress‐induced apoptosis is mediated primarily through the intrinsic pathway that involves caspase‐9. We previously reported that in caspase‐9‐deficient cells, a protein complex containing ATG5 and Fas‐associated death domain (FADD) facilitated caspase‐8 activation and cell death in response to endoplasmic reticulum (ER) stress. Here, we investigated whether this complex could be activated by other forms of cell stress. We show that diverse stress stimuli, including etoposide, brefeldin A and paclitaxel, as well as heat stress and gamma‐irradiation, caused formation of a complex containing ATG5‐ATG12, FADD and caspase‐8 leading to activation of downstream caspases in caspase‐9‐deficient cells. We termed this complex the ‘stressosome’. However, in these cells, only ER stress and heat shock led to stressosome‐dependent cell death. Using in silico molecular modelling, we propose the structure of the stressosome complex, with FADD acting as an adaptor protein, interacting with pro‐caspase‐8 through their respective death effector domains (DEDs) and interacting with ATG5‐ATG12 through its death domain (DD). This suggests that the complex could be regulated by cellular FADD‐like interleukin‐1β‐converting enzyme–inhibitory protein (cFLIPL), which was confirmed experimentally. This study provides strong evidence for an alternative mechanism of caspase‐8 activation involving the stressosome complex.

The stability of IFN-γ as a therapeutic protein can play a key role on its anticancer effects. Herein, we explored the thermodynamic parameters and conformational stability of IFN-γ in the presence of calycosin, the main active compound of Radix astragali, by different biophysical and theoretical analysis. Afterwards, the improved anticancer effects of IFN-γ-calycosin interaction relative to IFN-γ alone were assessed on hepatocellular carcinoma (HepG2) cell line by MTT and caspase assays. ITC data indicated that upon interaction of calycosin with IFN-γ the binding and thermodynamic parameters were as follows: Kd = 1.9 μM, ΔG° = −32.45 kJ/mol, ΔH° = −11.91 kJ/mol, and TΔS° = 20.54 kJ/mol. ANS/synchronous fluorescence, CD and UV–Vis spectroscopy studies indicated that the interaction between calycosin and IFN-γ caused the folding of the IFN-γ backbone in to a more packed structure with enhanced α-helix content and higher melting temperature (Tm) value. The spectroscopic outcomes were then verified by molecular docking and molecular dynamic analysis. It was also shown that after incubation of the IFN-γ samples at 50 °C for 60 min in the presence of calycosin (5 μM), the IFN-γ-calycosin system showed a significant antiproliferative effects against hepatocellular carcinoma (HepG2) cells through caspase-9/3 activation and this anticancer effect was more pronounced than free IFN-γ. This data may provide useful information about the development of IFN-γ-based therapeutic platforms.

Objectives Tonkinensine B, a novel compound with cytisine–pterocarpan skeleton isolated from the root of Sophora tonkinensis Gagnep, was reported to have a significant antitumor effect. The effect and intrinsic mechanism of tonkinensine B on tumour need to be further investigated. Methods With the help of cell cytotoxicity, the effect of tonkinensine B on MDA-MB-231 cells was investigated. By observing mitochondrial function changes, the intrinsic mechanism was further studied. The levels of key apoptosis-associated proteins Bcl-2, Bax, caspase-9, caspase-3 and AKT in MDA-MB-231 cells were analysed to determine whether tonkinensine B caused apoptosis via the mitochondrial pathway. Key findings After treated with tonkinensine B, MDA-MB-231 cells multiplication was repressed, and the decreased mitochondrial membrane potential, loss of ATP synthesis and elevated ROS generation were detected. Furthermore, the proportions of Bax/Bcl-2, cleaved caspase-3 and caspase-9 proteins production were up-regulated, indicating that tonkinensine B acted on intrinsic mitochondrial-mediated apoptosis pathway. In addition, tonkinensine B also reduced phosphorylation levels of AKT, and thus the activation of apoptosis might likewise be correlated with the inhibition of the PI3K/AKT pathway. Conclusions Tonkinensine B may be a hopeful candidate for human triple-negative breast cancer, and further structural optimization is expected to improve its anti-tumour activity.

Thyroid cancer (TC) was the most frequent thyroid malignant tumour, accounting for about 1% of all malignant tumours. Some long non‐coding RNAs (lncRNAs) have been reported to exert essential tumour promotion effects, while caspase‐9 (CASP9) gene could play a promotive role in the cell apoptosis in TC. However, whether they have a specific effect on TC remains unclear. Hence, this study aims to explore the relationship between LINC00607 and CASP9, and its effect in TC. LINC00607 expression in the TC tissues and cell lines was determined. Then, we explored the combination effect between a LINC00607 and a methylation inhibitor 5‐Aza‐dc in doxorubicin‐resistant ARO cells using colony formation assay, flow cytometry, WST‐1 and EdU assay, as well as in vivo tumour growth assay. Besides, the dual‐luciferase reporter gene assay, RIP, ChIP, methylation‐specific PCR and BSP method were employed to detect the relationship between LINC00607 and CASP9 and its methylation. LINC00607 expression was up‐regulated in the doxorubicin‐resistant TC cell lines and tissues and negatively correlated to the poor prognosis of TC patients. Knockdown of LINC00607 suppressed doxorubicin resistance, proliferation and colony formation, and promoted cell apoptosis of TC cells in vitro, as well as suppressed tumour growth in vivo, whereas LINC00607 overexpression was observed to exercise the opposite effects. Notably, it was also revealed that LINC00607 down‐regulated the CASP9 expression by promoting CASP9 promoter methylation. In conclusion, LINC00607 could inhibit the apoptosis and augment the doxorubicin resistance of TC cells by decreasing CASP9 expression, which might provide a novel therapeutic target for TC treatment.

Esophageal cancer is one of the most common malignancies which induces cancer-related death. Cancer metastasis and recurrence are the main obstacle faced in esophageal cancer treatment. β-Asarone has been shown to act as an anti-cancer reagent in various cancer types. However, the anti-cancer activities of β-Asarone in esophageal cancer have not been shown. In the current study, we show that β-Asarone suppressed the proliferation of esophageal squamous cancer cells (ESCC) in both dose- and time-dependent manners. Moreover, β-Asarone treatment increases activated caspase 3, caspase 9, and cleaved poly ADP-ribose polymerase, and induces apoptosis in ESCC. Additionally, β-Asarone also suppresses epithelial-mesenchymal transition (EMT) and the invasive and migratory abilities in ESCC. Interestingly, β-Asarone suppresses TGF-β/Smad signaling by inhibition of TGF-β-induced phosphorylation of Smad2 and Smad3. Importantly, we show that inhibition of TGF-β/Smad signaling activation is critical for β-Asarone-suppressed EMT. Our data revealed a novel role of β-Asarone which targets invasive properties by inhibiting TGF-β/Smad signaling activation in ESCC. Our study suggests the potential application of β-Asarone to reduce cancer metastasis and recurrence in esophageal cancer treatment.

The influence of cannabidiol (CBD) on brain development is inadequately understood. Since CBD is considered a non-intoxicating drug, it has attracted great interest concerning its potential medical applicability, including in pregnant women and children. Here, we elucidated the response of perinatal rat cortical neurons and astrocytes to CBD at submicromolar (0.1, 0.5, 1, 5 µM) concentrations attainable in humans. The effect of CBD was concentration- and time-dependent and cell-specific. In neurons, 0.1 µM CBD induced an early and transient change in mitochondrial membrane potential (ΔΨm), ATP depletion, and caspase-8 activation, followed by rapid ATP recovery and progressive activation of caspase-9 and caspase-3/7, resulting in early apoptotic cell death with reduction and shortening of dendrites, cell shrinkage, and chromatin condensation. The decrease in neuronal viability, ATP depletion, and caspase activation due to CBD exposure was prevented by transient receptor potential vanilloid 1 (TRPV1) antagonist. In astrocytes, 0.5 µM CBD caused an immediate short-term dysregulation of ΔΨm, followed by ATP depletion with transient activation of caspase-8 and progressive activation of caspase-9 and caspase-3/7, leading to early apoptosis and subsequent necroptosis. In astrocytes, both TRPV1 and cannabinoid receptor 1 (CB1) antagonists protected viability and prevented apoptosis. Given that CBD is a non-intoxicating drug, our results clearly show that this is not the case during critical periods of brain development when it can significantly interfere with the endogenous cannabinoid system.

Marine microalgae are receiving great interest as sustainable sources of bioactive metabolites for health, nutrition and personal care. In the present study, a bioassay-guided screening allowed identifying an enriched fraction from SPE separation of the methanolic extract of the marine diatom Thalassiosira rotula with a chemically heterogeneous composition of cytotoxic molecules, including PUFAs, the terpene phytol, the carotenoid fucoxanthin and the phytosterol 24-methylene cholesterol (24-MChol). In particular, this latter was the object of deep investigation aimed to gain insight into the mechanisms of action activated in two tumour cell models recognised as resistant to chemical treatments, the breast MCF7 and the lung A549 cell lines. The results of our studies revealed that 24-MChol, in line with the most studied β-sitosterol (β-SIT), showed cytotoxic activity in a 3–30 µM range of concentration involving the induction of apoptosis and cell cycle arrest, although differences emerged between the two sterols and the two cancer systems when specific targets were investigated (caspase-3, caspase-9, FAS and TRAIL).

Oxidative stress contributes to major complications of obesity. This study intended to identify whether orlistat could mitigate myocardial damage in obese animal models. The tested rats were divided into two groups and fed either with normal chow (n = 6 per group) or with a high-fat diet (HFD) for 6 weeks to induce obesity (n = 12 per group). Obese rats were further subjected to treatment either with distilled water (OB group) or orlistat 10 mg/kg/day (OB + OR group). Key indices of oxidative stress, inflammation, and apoptosis were assessed using an immunohistochemical-based technique and real-time PCR. The OB group showed significant increases of oxidative stress markers (TBARs and PCO), with significant decreases of anti-oxidant markers (Nrf2, SOD, CAT, and GPx). Furthermore, mRNA expression of pro-inflammatory markers (TNF-α and NF-κβ) and pro-apoptosis markers (Bax, Caspase-3, Caspase-8, and Caspase-9) were significantly upregulated in the OB group. Obese rats developed pathological changes of myocardial damages as evidenced by the presence of myocardial hypertrophy and inflammatory cells infiltration. Orlistat dampened the progression of myocardial damage in obese rats by ameliorating the oxidative stress, and by inhibiting NF-κβ pathway and caspase-dependent cell apoptosis. Our study proposed that orlistat could potentially mitigate oxidative stress-linked myocardial damage by mitigating inflammation and apoptosis, thus rationalizing its medical usage.

The objective of this study was to examine the influences of glycinin for growth and intestinal structural integrity related to oxidative damage, apoptosis and tight junction of juvenile hybrid yellow catfish (

Furanocoumarins, the secondary metabolites of plants, are considered to be natural insecticides and fungicides because they prevent the invasion of plant pathogenic microorganisms and the predation of herbivorous insects. In this study, novel 2-arylfuranocoumarin derivatives were designed to synthesize by condensation, esterification, bromination, and Wittig reaction. The results showed an excellent photosensitive activity of 2-thiophenylfuranocoumarin (I34). Cell Counting Kit-8 detected that I34 could inhibit the proliferation of

A Protopanaxatriol class Glucopyranosides Consisting Anti- Cancer Formulation with Malignant Tumor Suppressive Features via Inhibiting HiF & Cyclin D2 & MMP-9 Expression and Elevating Caspase 3&9 Activity. MIT Bioceutical Innovations Articles by Erdal Can Alkoclar & Metehan Yeşil

Background Lymphocytes are a group of white blood cells with a variety of roles their integrity is crucial for the body’s immune responses. Cadmium, a heavy metal and environmental pollutant, is known as a toxicant to exert its adverse effects on some sort of cells including blood cells. Research Design In this study, human lymphocytes were divided into 3 groups: (1) lymphocytes at 0-h, (2) lymphocytes at 24 h (control), (3) lymphocytes treated with cadmium chloride (15 μM). Lymphocyte viability and plasma membrane integrity were assessed in these groups. In addition, the occurrence of apoptosis was investigated by assessment of nucleus diameter and flow cytometry. Activation of caspase-3 was also detected by immunocytochemistry. Results Result showed that lymphocyte’s viability and plasma membrane integrity decreased in lymphocytes treated with cadmium as compared with the control group. Decreased nucleus diameter and result of flow cytometry demonstrated cadmium-induced apoptosis in human lymphocytes. Furthermore, lymphocytes treated with cadmium displayed intensely activated caspase-3 immunoreactivity in their cytoplasm. Conclusion In conclusion, cadmium not only negatively effect on viability and plasma membrane, but also induces caspase-dependent apoptosis in human lymphocytes.

Caspases (CASPs) are the main executors of the apoptotic process. Studies to date have shown the role of caspase-8 (CASP8) and caspase-9 (CASP9) in carcinogenesis. Therefore, the aim of this study was to investigate the associations between CASP9-rs4233532, CASP9-rs4646018, and CASP8- rs1045485 gene polymorphisms and non-Hodgkin lymphoma (NHL) susceptibility in an Iranian population-based study. Moreover, it was examined whether such the genotype of these polymorphisms is related with clinicopathological characteristics of NHL.175 patients with NHL and 175 age- and sex-matched controls were enrolled in this study. We determined the genotypes of single nucleotide polymorphisms (SNPs) from CASP genes with Tetra ARMS-PCR (Amplification refractory mutation system) method.Statistically significant association were observed between CASP9-rs4646018 and increased risk of NHL under codominant CC, codominant TC, and dominant TC+CC genetic models. Our results showed that the A allele of CASP8-rs1045485 was a protective factor for NHL and GArs1045485 genotype significantly reduced risk of NHL. In contrast, CASP9- rs4233532 was not linked to NHL susceptibility. No relationship was detected between CASP8-rs1045485 and CASP9-rs4233532 and NHL clinicopathological characteristics, however genetic variation in CASP9-rs4646018 was associated with histology, treatment and radio therapy of NHL.Our study presented that the CASP8- rs1045485 and CASP9-rs4646018 polymorphisms could affect the risk of NHL in Iranian populations which was the first report to show the significant relationship between rs1045485, rs4646018 polymorphisms and NHL susceptibility. Replication large-scale case-control studies in different ethnicities are warranted to verify these findings.

Previous studies revealed that melatonin ameliorated acute renal injury induced by cisplatin, but the mechanisms remain unclear. Peroxidase proliferative receptor α (PPARα) is considered the major regulator of fatty acid oxidation (FAO), which is an important source of energy for renal tubular epithelial cells. In this study, the aim was to investigate the role of melatonin in cisplatin-induced NRK-52E (rat renal tubular epithelial cell line) cell damage and the underlying mechanisms. We established a cisplatin-stimulated NRK-52E model in vitro. We assessed the levels of apoptotic proteins, including caspase-3, caspase-9, and B-cell lymphoma 2-associated X protein (Bax), as well as PPARα and FAO-related genes (Acadm, Acat1, Acsm2, Acsm3, PGC-1α, Pecr, Bdh2, and Echs1). Furthermore, we detected the effects of miR-21 and PPARα antagonist on the above indicators. We found that melatonin reduced the protein expression levels of caspase-3, caspase-9, and Bax, and increased the expression levels of the PPARα gene and protein and PPARα activity, as well as FAO-related genes, in NRK-52E cells. However, miR-21 mimics and PPARα antagonists partially antagonized the above effects of melatonin. Our data indicated that melatonin could alleviate cisplatin-induced cell damage through the upregulation of PPARα/FAO.

Retinal neurovascular injuries are a leading cause of vision loss in young adults presenting unmet therapeutic needs. Neurovascular injuries damage homeostatic communication between endothelial, pericyte, glial, and neuronal cells through signaling pathways that remain to be established. To understand the mechanisms that contribute to neuronal death, we use a mouse model of retinal vein occlusion (RVO). Using this model, we previously discovered that after vascular damage, there was non-apoptotic activation of endothelial caspase-9 (EC Casp9); knock-out of EC Casp9 led to a decrease in retinal edema, capillary ischemia, and neuronal death. In this study, we aimed to explore the role of EC Casp9 in vision loss and inflammation. We found that EC Casp9 is implicated in contrast sensitivity decline, induction of inflammatory cytokines, and glial reactivity. One of the noted glial changes was increased levels of astroglial cl-caspase-6, which we found to be activated cell intrinsically by astroglial caspase-9 (Astro Casp9). Lastly, we discovered that Astro Casp9 contributes to capillary ischemia and contrast sensitivity decline after RVO (P-RVO). These findings reveal specific endothelial and astroglial non-apoptotic caspase-9 roles in inflammation and neurovascular injury respectively; and concomitant relevancy to contrast sensitivity decline.

Rutin, also called quercetin-3-rhamnosyl glucoside, is a natural flavonol glycoside present in many plants. Rutin is used to treat various diseases, such as inflammation, diabetes, and cancer. For polymeric biomaterials, triethylene glycol dimethacrylate (TEGDMA) is the most commonly used monomer and serves as a restorative resin, a dentin bonding agent and sealant, and a bone cement component. Overall, TEGDMA induces various toxic effects in macrophages, including cytotoxicity, apoptosis, and genotoxicity. The aim of this study was to investigate the protective mechanism of rutin in alleviating TEGDMA-induced toxicity in RAW264.7 macrophages. After treatment with rutin, we assessed the cell viability and apoptosis of TEGDMA-induced RAW264.7 macrophages using an methylthiazol tetrazolium (MTT) assay and Annexin V-FITC/propidium iodide assay, respectively. Subsequently, we assessed the level of genotoxicity using comet and micronucleus assays, assessed the cysteinyla aspartate specific proteinases (caspases) and antioxidant enzyme (AOE) activity using commercial kits, and evaluated the generation of reactive oxygen species (ROS) using a dichlorodihydrofluorescein diacetate (DCFH-DA) assay. We evaluated the expression of heme oxygenase (HO)-1, the expression of nuclear factor erythroid 2 related factor (Nrf-2), and phosphorylation of AMP activated protein kinase (AMPK) using the Western blot assay. The results indicated that rutin substantially reduced the level of cytotoxicity, apoptosis, and genotoxicity of TEGDMA-induced RAW264.7 macrophages. Rutin also blocked the activity of caspase-3, caspase-8, and caspase-9 in TEGDMA-stimulated RAW264.7 macrophages. In addition, it decreased TEGDMA-induced ROS generation and AOE deactivation in macrophages. Finally, we found that TEGDMA-inhibited slightly the HO-1 expression, Nrf-2 expression, and AMPK phosphorylation would be revered by rutin. In addition, the HO-1 expression, Nrf-2 expression, and AMPK phosphorylation was enhanced by rutin. These findings indicate that rutin suppresses TEGDMA-induced caspase-mediated toxic effects through ROS generation and antioxidative system deactivation through the Nrf-2/AMPK pathway. Therefore, rutin has the potential to serve as a novel antitoxicity agent for TEGDMA in RAW264.7 macrophages.

Nickel (Ni) is ubiquitous in the environment and evidence has suggested that Ni can cause ocular surface inflammation, especially in fine particulate matter and personal products. Continuous daily exposure to Ni-containing dust may adversely impact the human cornea, whereas the underlying mechanism of this phenomenon remains not fully understood. Here, human corneal epithelial cells (HCEC) were employed to analyze the toxicity of Ni via detections of cell morphology, cell viability, reactive oxygen species production, cell apoptosis rate, and apoptotic gene expression levels after exposure for 24 h to uncover the damage of Ni to the cornea. A concentration-dependent inhibition of HCECs’ viability and growth was observed. In particular, Ni at 100 μM significantly decreased cell viability to 76%, and many cells displayed an abnormal shape and even induced oxidative damage of HCEC by increasing ROS to 1.2 times, and further led to higher apoptosis (24%), evidenced by up-regulation of apoptotic genes Caspase-8, Caspase-9, NF-κB, IL-1β, and Caspase-3, posing a risk of dry eye. Our study suggested that Ni induces apoptosis of HCEC through oxidative damage. Therefore, Ni pollution should be comprehensively considered in health risks or toxic effects on the ocular surface.

Glioblastoma multiforme (GBM) is the most common and malignant brain tumor. Temozolomide (TMZ) is the first-line chemotherapeutic drug for treating GBM. However, drug resistance is still a challenging issue in GBM therapy. Our preliminary results showed upregulation of androgen receptor (AR) gene expression in human GBM tissues. This study was designed to evaluate the effects of enzalutamide, a specific inhibitor of the AR, on killing drug-resistant and -sensitive glioblastoma cells and the possible mechanisms. Data mining from The Cancer Genome Atlas (TCGA) database revealed upregulation of AR messenger (m)RNA and protein expressions in human GBM tissues, especially in male patients, compared to normal human brains. In addition, expressions of AR mRNA and protein in human TMZ-sensitive U87 MG and -resistant U87 MG-R glioblastoma cells were elevated compared to normal human astrocytes. Exposure of human U87 MG and U87 MG-R cells to enzalutamide concentration- and time-dependently decreased cell viability. As to the mechanism, enzalutamide killed these two types of glioblastoma cells via an apoptotic mechanism. Specifically, exposure to enzalutamide augmented enzyme activities of caspase-9 rather than those of caspase-8. Moreover, enzalutamide successively triggered an elevation in levels of the proapoptotic Bax protein, a reduction in the mitochondrial membrane potential, release of cytochrome c, cascade activation of caspases-3 and -6, DNA fragmentation, and cell apoptosis in human TMZ-sensitive and -resistant glioblastoma cells. Pretreatment with Z-VEID-FMK, an inhibitor of caspase-6, caused significant attenuations in enzalutamide-induced morphological shrinkage, DNA damage, and apoptotic death. Taken together, this study showed that enzalutamide could significantly induce apoptotic insults to human drug-resistant and -sensitive glioblastoma cells via an intrinsic Bax-mitochondrion-cytochrome c-caspase cascade activation pathway. Enzalutamide has the potential to be a drug candidate for treating GBM by targeting the AR signaling axis.

Ilimaquinone (IQ), a metabolite found in marine sponges, has been reported to have a number of biological properties, including potential anticancer activity against colon cancer. However, no clear understanding of the precise mechanism involved is known. The aim of this study was to examine the molecular mechanism by which IQ acts on HCT-116 cells. The anticancer activity of IQ was investigated by means of a cell viability assay followed by the determination of induction of apoptosis by means of the use of acridine orange–ethidium bromide (AO/EB) staining, Annexin V/PI double staining, DNA fragmentation assays, and TUNEL assays. The mitochondrial membrane potential (ΔΨm) was detected using the JC-1 staining technique, and the apoptosis-associated proteins were analyzed using real-time qRT-PCR. A molecular docking study of IQ with apoptosis-associated proteins was also conducted in order to assess the interaction between IQ and them. Our results suggest that IQ significantly suppressed the viability of HCT-116 cells in a dose-dependent manner. Fluorescent microscopy, flow cytometry, DNA fragmentation and the TUNEL assay in treated cells demonstrated apoptotic death mode. As an additional confirmation of apoptosis, the increased level of caspase-3 and caspase-9 expression and the downregulation of Bcl-2 and mitochondrial dysfunction were observed in HCT-116 cells after treatment with IQ, which was accompanied by a decrease in mitochondrial membrane potential (ΔΨm). Overall, the results of our studies demonstrate that IQ could trigger mitochondria-mediated apoptosis as demonstrated by a decrease in ΔΨm, activation of caspase-9/-3, damage of DNA and a decrease in the proportion of Bcl-2 through the mitochondrial-mediated apoptosis pathway.

Seaweeds are a great source of compounds with cytotoxic properties with the potential to be used as anticancer agents. This study evaluated the cytotoxic and proteasome inhibitory activities of 12

To investigate the mechanism of the effect of acupuncture and moxibustion on improving liver injury by observing the changes of cysteine protease (Caspase) associated with hepatocyte apoptosis based on cisplatin (DDP) induced liver injury model mice.Forty KM mice were randomly divided into control group, model group, acupuncture group and moxibustion group, with 10 mice in each group. The liver injury model was replicated by intraperitoneal injection of DDP. In the acupuncture group and the moxibustion group, acupuncture and moxibustion were performed at"Dazhui"(GV14), and bilateral "Ganshu"(BL18), "Shenshu"(BL23), and "Zusanli"(ST36), respectively, once per day for 5 d. General condition of mice in each group were observed;The activities of AST, ALT and GLDH in mice serum were detected by biochemical method. ELISA and Western blot assay were used to detect Caspase-3, Caspase-8 and Caspase-9 contents and protein expression in the liver tissues of each group of mice, respectively.Compared with the control group, the general condition of the mice in the model group was poorer, and the Caspase-3, Caspase-8 and Caspase-9 contents and protein expressions in liver tissues and the activities of AST, ALT and GLDH in serum were increased (Acupuncture and moxibustion can reduce liver injury due to DDP chemotherapy by modulating the expression of apoptotic factors Caspase-3, Caspase-8 and Caspase-9 in liver tissues of DDP model mice and improving liver function, which may be one of the mechanisms of the effect of acupuncture and moxibustion to ameliorates liver injury after DDP chemotherapy.

Diabetic nephropathy is one of the major diabetic complications which has become the major cause of end-stage renal disease. It has been demonstrated that apoptosis induced by hyperglycemia is a critical factor in the pathophysiology of diabetic nephropathy. Taurine is a semi-essential amino acid in mammals and has been shown to be a potent endogenous antioxidant. The protective effect of taurine against apoptosis in diabetic kidney deserves to be explored. In the present study, mRNA expression of cysteinyl aspartate-specific proteinase-3 (caspase-3) and caspase-9 was examined, and the activity of caspase-3 was also detected as the marker of apoptosis. The expression of Bax and Bcl-2 was detected by Western blot. In addition, the level of total Akt and phosphorylated Akt (p-Akt) was measured. We found that caspase-3 and caspase-9 mRNA expression was decreased in diabetic kidney, which was recovered by taurine treatment. The activity of caspase-3 was increased in diabetic kidney, while the increased activity was significantly attenuated after taurine treatment. We also found that the expressions of Bax and Bcl-2 were disturbed in diabetic kidney, which were reversed by taurine treatment. The decrease of the p-Akt level was also prevented by taurine treatment. These results indicated that taurine-ameliorated apoptosis in diabetic kidney may be through activating of the Akt signaling pathway.

The family of caspases is known to mediate many cellular pathways beyond cell death, including cell differentiation, axonal pathfinding, and proliferation. Since the identification of the family of cell death proteases, there has been a search for tools to identify and expand the function of specific family members in development, health, and disease states. However, many of the currently commercially available caspase tools that are widely used are not specific for the targeted caspase. In this report, we delineate the approach we have used to identify, validate, and target caspase-9 in the nervous system using a novel inhibitor and genetic approaches with immunohistochemical read-outs. Specifically, we used the retinal neuronal tissue as a model to identify and validate the presence and function of caspases. This approach enables the interrogation of cell-type specific apoptotic and non-apoptotic caspase-9 functions and can be applied to other complex tissues and caspases of interest. Understanding the functions of caspases can help to expand current knowledge in cell biology, and can also be advantageous to identify potential therapeutic targets due to their involvement in disease.

Fluoronitrile gas (C

Objective To explore the effect and mechanism of paeoniflorin (PAE) on apoptosis of fibroblast synovial cells derived from rheumatoid arthritis tissues. Methods Rheumatoid arthritis fibroblast-like synovial cells (RA-FLSs) were cultured under different concentrations of PAE (20, 40, 80 μmol/L). Survival rate of cells was determined at different incubation time-points (24, 48, 72 hours) by MTT assay. Flow cytometry was performed to determine apoptosis rate of cells. Cells were transfected with 3 small interfering RNAs(siRNAs) fragments of long non-coding RNA metastasis-associated lung adenocarcinoma transcript 1(lncRNA MALAT1) for 48 hours. Real-time quantitative PCR was performed to determine if MALAT1 knockdown was successful. Cells were assigned into control group, PAE group, si-NC group, si-MALAT1 group and PAE combined with si-MALAT1 group. Cell apoptosis rate was determined by flow cytometry after knockdown of MALAT1 expression. mRNA expression levels of Wnt1, β-catenin, caspase-3, caspase-9, B-cell lymphoma 2(Bcl2) and Bcl2-associated X protein (BAX) were determined using real-time quantitative PCR. Western blot analysis was performed to determine protein expression levels of Wnt1, β-catenin, caspase-3, caspase-9, Bcl2 and BAX in each group. Results The findings showed that incubation of RA-FLSs with PAE at the tested concentration decreased their viability in a time-dependent manner. Expression level of lncRNA MALAT1 was lower in RA-FLSs group compared with the level in NC-FLSs group. Analysis showed that apoptosis rate was higher in cells treated with PAE. si-MALAT1 group had the lowest apoptosis rate, whereas the PAE combined with si-MALAT1 group showed a significantly higher apoptosis rate compared with si-MALAT1 group. mRNA levels of Bcl2, Wnt1 and β-catenin were lower in PAE group compared with the level in control group, whereas mRNA expression levels of caspase-3, caspase-9 and BAX were higher in PAE group compared with the levels in control group. Expression level of Bcl2 was significantly higher in si-MALAT1 group compared with the level in the control group. Expression level of BAX, caspase-3 and caspase-9 were higher in PAE combined with si-MALAT1 group compared with the level in the control group. Conclusion PAE promotes apoptosis and inhibits the Wnt1/β-catenin pathway in RA-FLSs by upregulating expression of lncRNA MALAT1.

Due to their promising effects, gold(III) complexes recently drew increasing attention in the design of new metal-based anticancer therapeutics. Two gold(III) complexes, square-planar [Au(DPP)Cl2]+ - Complex 1 and distorted square-pyramidal [Au(DMP)Cl3] - Complex 2 (where DPP=4,7-diphenyl-1,10-phenanthroline and DMP=2,9-dimethyl-1,10-phenanthroline) were previously synthetized, described and approved as complexes with pronounced cytotoxic effects on colorectal HCT-116 and breast MDA-MB-231 cancer cells. This study investigated the type of cell death by AO/EB double staining, and identification of possible targets responsible for their cytotoxicity, monitored by immunofluorescence and qPCR methods. Both complexes induced apoptosis in all applied concentrations. In the HCT-116 cells apoptosis was activated by external apoptotic pathway, via increase of Fas receptor protein expression and Caspase 8 gene expression. Also, the mitochondrial pathway was triggered by affecting the Bcl-2 members of regulatory proteins and increased caspase 9 protein expression. In MDA-MB-231 cells, apoptosis was initiated from the mitochondria, due to disbalance between expressions of pro- and anti-apoptotic Bcl-2 family members and caspase 9 activation. Complex 1 shows better activity compared to Complex 2, which is in accordance with its structural characteristics. The results deal weighty data about proapoptotic activity of gold(III) complexes and highlighted potential targets for cancer therapy.

Farrerol is an herbal compound extracted from rhododendron. Here, our study is to investigate biological effects of farrerol on lung adenocarcinoma (LAC) cells. Human LAC cell lines and xenograft mouse model were utilized to define the effects of farrerol on tumor growth. Our findings indicated that farrerol significantly reduced LAC cell viability as well as the colony-forming capacity. Flow cytometry analysis demonstrated that farrerol contributed to cell apoptosis and G0/G1 phase cell cycle arrest. Mechanistically, farrerol treatment upregulated proapoptotic molecules (Bak, Bid, cleaved caspase-3 and cleaved caspase-9) and senescence markers (p16 and p2), but downregulated antiapoptosis genes (Bcl-2 and Bcl-XL) and cell cycle-associated genes (CyclinD1 and CDK4); meanwhile, the phosphorylation of retinoblastoma (Rb) protein was attenuated upon pretreatment of LAC cells with farrerol in comparison to untreated control. Further studies indicated that farrerol elevated reactive oxygen species levels, activating mitochondrial apoptotic pathway and causing cell apoptosis. However, exposure to farrerol did not result in significant apoptosis in normal lung epithelial cells, suggesting a tumor-specific effect of farrerol on LAC cells. In animal model, farrerol showed a significant inhibitory effect on LAC xenograft tumor growth. And gene expressions in tumor tissues, as mentioned above, were in line with the in vitro results. Taken together, these results suggested that farrerol caused LAC cell apoptosis by activating mitochondrial apoptotic pathway, whereas farrerol treatment had no notable effect on normal lung epithelial cells. Farrerol might be an effective therapeutic drug for LAC.

Chemotherapeutics remain the first choice for advanced gastric cancers (GCs). However, drug resistance and unavoidable severe toxicity lead to chemotherapy failure and poor prognosis. Long noncoding RNAs (lncRNAs) play critical roles in tumor progression in many cancers, including GC. Here, through RNA screening, an apoptotic protease-activating factor 1 (APAF1)-binding lncRNA (ABL) that is significantly elevated in cancerous GC tissues and an independent prognostic factor for GC patients is identified. Moreover, ABL overexpression inhibits GC cell apoptosis and promotes GC cell survival and multidrug resistance in GC xenograft and organoid models. Mechanistically, ABL directly binds to the RNA-binding protein IGF2BP1 via its KH1/2 domain, and then IGF2BP1 further recognizes the METTL3-mediated m6A modification on ABL, which maintains ABL stability. In addition, ABL can bind to the WD1/WD2 domain of APAF1, which competitively prevent cytochrome c from interacting with APAF1, blocking apoptosome assembly and caspase-9/3 activation; these events lead to resistance to cell death in GC cells. Intriguingly, targeting ABL using encapsulated liposomal siRNA can significantly enhance the sensitivity of GC cells to chemotherapy. Collectively, the results suggest that ABL can be a potential prognostic biomarker and therapeutic target in GC.