Expression Of Apoptotic Proteins Bcl-2 Research Articles

Laminarin Alleviates the Ischemia/Reperfusion Injury in PC12 Cells via Regulation of PTEN/PI3K/AKT Pathway

Objective. To investigate the protective effect of laminarin on PC12 cells damaged by oxygen glucose deprivation/reoxygenation (OGD/R) and its molecular mechanism. Methods. PC12 cells in the logarithmic phase were randomly divided into the control group, OGD/R group, and OGD/R+laminarin (0.5, 2.5, and 5 μg/ml) group. CCK-8 activity assay kit was used to detect cell viability. ELISA kit was performed to examine the levels of proinflammatory factors (TNF-α, IL-1β, and IL-6) and oxidative stress markers (ROS, LDH, and MPO). In addition, flow cytometry was employed to determine cell cycle and apoptosis. The expression of cell proliferation-related proteins (PCNA and Ki67), apoptosis-related proteins (Bcl-2, Bax, and Caspase-3), and PTEN/PI3K/AKT pathway-related proteins was evaluated by Western blot. Results. Compared with the control group, the cell viability was decreased significantly in the OGD/R group. CCK-8 results showed that laminarin could attenuate the damage of PC12 cell viability induced by OGD/R in a concentration-dependent manner. Meanwhile, the highest concentration of 5 μg/ml laminarin could significantly promote the viability of PC12 cells and the expression of PCNA and Ki67 than the OGD/R group. Additionally, ELISA assays showed that laminarin significantly inhibited the expression of proinflammatory factors (TNF-α, IL-1β, and IL-6) and the levels of oxidative stress markers (ROS, LDH, and MPO). Flow cytometry results demonstrated that laminarin promoted the cell cycle. And laminin upregulated the expression of apoptotic protein Bcl-2, while downregulated the expression of apoptotic proteins Bax and Caspase-3. Finally, laminarin significantly suppressed the expression of PTEN and facilitated the expression of PI3K and p-AKT compared to the OGD/R group. Conclusion. Laminarin could alleviate the OGD/R-induced PC12 cell neuronal injury via promoting cell activity and cycle and inhibiting inflammation, oxidative stress, and apoptosis. The mechanism may be related to the downregulation of PTEN protein and the activation of the PI3K/AKT pathway.

Rehabilitation training improves cognitive disorder after cerebrovascular accident by improving BDNF Bcl-2 and Bax expressions in regulating the JMK pathway.

To explore the effect of rehabilitation training on cognitive impairment after cerebrovascular accident and its potential mechanism. 100 patients of cerebrovascular accident treated in our hospital from August 2018 to August 2019 were selected as the subjects, and 50 patients with physical examination were selected as healthy control group. The patients with cerebrovascular accident were randomly divided into control group (50 patients) and research group (50 patients). The patients in the control group were given routine medication, the patients in research group were given rehabilitation training on the basis of routine drug therapy. The blood samples were collected on admission and 6 months after admission to detect the molecular markers related to inflammation, nerve cell nutrition and function and apoptosis in the serum. The cognitive function was evaluated by scales. We established a rat cerebral ischemia model, compared the differences in the evasive latency, serum CRP, BNDF, Bcl-2, BAX, Glu, NE levels and BNDF, TrkB, pTrkB, JNK levels in hippocampus, amygdala, and prefrontal tissue between model rats after rehabilitation training and model rats without rehabilitation training. On admission, there were no significant differences in the scores of Barthel index (BI), Fugl-Meyer motor function scale (FM), Montreal cognitive assessment scale (MoCA) and mini-mental state examination (MMSE) (p>0.05). 6 months later, the above scores and BNDF, Bcl-2, and norepinephrine were significantly higher in the research group (p<0.05), while CRP, Bax, 5-HT and glutamate in the research group were significantly lower than those in the control group (p<0.05). Rehabilitation training can improve the motor function, mental state and cognitive level of patients, reduce the levels of neurotoxic factors, pro-inflammatory factors and pro-apoptotic factors, and improve the levels of inhibiting apoptotic factors, neurotrophic factors and neurotransmitters. In animal experiments, rehabilitation training can increase BDNF and its activated receptors in hippocampus, amygdala and prefrontal lobe of rats, and decrease JNK of apoptotic protein, suggesting that rehabilitation training may regulate the expression of apoptotic proteins Bcl-2 and Bax by upregulating BDNF and its receptors and acting on JNK pathway, thereby inhibiting cell apoptosis and improving cognitive impairment after cerebrovascular accident.

Interference with the Expression of miR-93 Inhibits the Proliferation and Migration and Promotes Apoptosis of Thyroid Tumor Cells Through Targeting Large Tumor Suppressor Molecule 2

Thyroid cancer (TC) is a common malignant tumor of the endocrine system. Currently, it is an urgent problem to be solved to explore the relevant molecular targets and molecular mechanisms for the occurrence and development of TC. miR-93 plays a role in regulating tumors in multiple cancers, but its expression in TC and its effect on proliferation,migration and apoptosis of TC cells have not been reported so far. In this paper, RT-qPCR was used to detect the expression of miR-93 and LATS2. Cell transfection techniques was used to interfere with or overexpress miR-93 and interfere with the expression of LATS2, and RT-qPCR was used to detect transfection efficiency. CCK-8 technique and clone formation assay were used to detect cell survival. Transwell and wound healing assay was used to detect cell migration and invasion, and western blot was used to detect the expression of invasion and migration related proteins MMP2 and MMP9. The apoptosis rate was detected by flow cytometry, and the expression of apoptotic proteins bcl-2, Bax, caspase3/9 and pro-caspase-3/9 was detected by western blot. The results showed that miR-93 was significantly increased in TC cell lines, especially in TPC1 cells. Interference with the expression of miR-93 inhibited proliferation, invasion, and migration, and promoted the apoptosis of TPC1 cells. In addition, we also found that interfering with the expression of miR-93 can promote the expression level of LATS2 in TPC1 cells. Interference with the expression of LATS2 can inhibit the effect on the proliferation, invasion, migration and apoptosis of TPC1 cells after miR-93 interference. In conclusion, interference with the expression of miR-93 inhibits the proliferation and migration and promotes apoptosis of TC cells through targeting LATS2.

Interference with miR-210 Alleviated Renal Injury in Septic Rats by Inhibiting JAK-STAT Pathway.

Pediatric sepsis has become the leading cause of death in pediatric intensive care units (PICU). The regulation of target genes may be the key to the treatment of pediatric sepsis. The expression of miR-210 in rat serum was detected by RT-qPCR. The serum BUN, Scr, and CysC were detected by an automatic biochemical analyzer. The expression of inflammatory factors was detected by ELISA. The apoptosis level of the cells was detected by TUNEL staining. The expression of apoptotic proteins Bcl2, Bax, Cleaved caspase3, caspase3, and JAK/STAT3 pathway-related proteins were detected by western blot. The expression of miR-210 was abnormally elevated in sepsis pups. Interfering with the expression of miR-210 in rats could reduce the degree of renal injury and inhibit the inflammatory response in sepsis pups. In addition, interference with miR-210 could inhibit the apoptosis level of renal tissue cells, and the expression of apoptosis-related proteins was also significantly decreased. During this process, we found that after interfering with the expression of miR-210, the expression of the JAK/STAT pathway was inhibited. Then, pathway agonist SC-39100 can reverse the inhibitory effects of interfering with miR-210 on renal tissue damage, inflammatory response, and apoptosis. Interference with miR-210 alleviated renal injury in septic rats by inhibiting JAK-STAT pathway.

RETRACTED ARTICLE: Synthesis, Crystal Structure, and Anti-Gastric Cancer Activity of a Heterometallic Coordination Polymer Based on Flexible 6,6-Dithiodinicotinic Acid

A new bimetallic coordination polymer [In2Na(H2O)2(ddca)3](NO3)·4H2O (1, H2ddca = 6,6′-dithiodinicotinic acid) is produced via a hydrothermal reaction of In(NO3)3, H2ddca with NaOH as the pH modulator. Various characterization methods, such as thermogravimetric analysis (TGA), elemental analysis (EA), single crystal and powder X-ray diffraction measurements, are used. Furthermore, a process of sonochemistry with oleic acid at 453 K is used to produce nanoparticles of 1 (nano-1). In terms of the biological function of the synthesized complex, the real-time reverse transcription-polymerase chain reaction is carried out to explore the relative mRNA expression of miR-181 in MGC-803, BGC-823, and SGC-7901 gastric cancer cell lines, and the western blot is used to explore the protein expression of apoptotic protein Bcl-2.

Synthesis, crystal structure, and anti-gastric cancer activity of a heterometallic coordination polymer based on flexible 6,6′-dithiodinicotinic acid

A new bimetallic coordination polymer [In2Na(H2O)2(ddca)3](NO3)×4H2O (1, H2ddca = 6,6'-dithiodinicotinic acid) is produced via a hydrothermal reaction of In(NO3)3, H2ddca with NaOH as the pH modulator. Various characterization methods, such as thermogravimetric analysis (TGA), elemental analysis (EA), single crystal and powder X-ray diffraction measurements, are used. Furthermore, a process of sonochemistry with oleic acid at 453 K is used to produce nanoparticles of 1 (nano-1). In terms of the biological function of the synthesized complex, the real-time reverse transcription-polymerase chain reaction is carried out to explore the relative mRNA expression of miR-181 in MGC-803, BGC-823, and SGC-7901 gastric cancer cell lines, and the western blot is used to explore the protein expression of apoptotic protein Bcl-2.

Overexpressions of sentrin-specific protease-1 is associated with the invasion and metastasis of human hepatocellular carcinoma

Objective To investigate the expressions of sentrin specific protease-1 (SENP-1) in hepatocellular carcinoma (HCC), and explore its correlations with tumor metastasis. Methods The expression of SENP-1 and hypoxia inducible factor-1α (HIF-1α) were assessed by polymerase chain reaction (PCR)and immunohistochemical staining in human hepatocellular carcinoma, edge tissues and non-cancer tissues, microvascular density (MVD)was determined. The expressions of SENP-1 in HCC cell lines MHCC97H, MHCC97L and HepG2 were assessed by reverse transcriptase-PCR (RT-PCR), quantitative real-time quantitative PCR (Real-time PCR) and Western blotting. The plasmid SENP1-small interfering RNA (siRNA) conducted was used to transfected into MHCC97H cells. Western blotting analysis was identified for the effect of transfection. The expression of cysteinyl aspartate specific proteinase-8 (Caspase-8), B-cell lymphoma-2 (bcl-2), HIF-1α and vascular endothelial growth factor (VEGF) protein in hepatoma cells before and after transfection was detected by Western blotting. The invasion and metastasis of hepatoma cells were detected by wound assay and Transwell invasion assay. Results Compared with the hepatocellular carcinoma tissues and non-cancer tissues, the expression of SENP-1 and HIF-1α in tumor-surrounding tissues were much higher. Compared with immortalized liver cell line, the HCC cell lines showed higher expression of SENP1 mRNA and protein, the expressions of SENP-1 was positive correlation with the metastasis ability of HCC cell lines. After transfection of SENP1-siRNA plasmid into MHCC97H cells, the expression of apoptosis promoting protein Caspase-8 was increased, and the expression of apoptotic protein bcl-2 was inhibited. Scratch healing assay revealed that the scuffing distance of hepatoma cells after transfection for 24 h was significantly increased compared with non-treatment group. Transwel1 assay showed that the number of hepatoma cells that migrated through the matrigel after transfection for 24 h was significantly decreased compared with non-treatment group (P=0.000). Conclusion The high expressions of SENP-1 in HCC might play an important role in tumor metastasis. The mechanism may be related to the over expression of SENP-1, which promotes liver cancer cells to adapt to hypoxic microenvironment and inhibit cell apoptosis. Key words: Sentrin specific peptidase 1; Hepatocellular carcinoma; Metastasis

Studies of the effects and mechanisms of ginsenoside Re and Rk3 on myelosuppression induced by cyclophosphamide

BackgroundGinsenoside Re (Re) is one of the major components of Panax ginseng Meyer. Ginsenoside Rk3 (Rk3) is a secondary metabolite of Re. The aim of this study was to investigate and compare the effects and underlying mechanisms of Re and Rk3 on cyclophosphamide-induced myelosuppression. MethodsThe mice myelosuppression model was established by intraperitoneal (i.p.) injection of cyclophosphamide. Peripheral blood cells, bone marrow nucleated cells, and colony yield of hematopoietic progenitor cells in vitro were counted. The levels of erythropoietin, thrombopoietin, and granulocyte macrophage colony-stimulating factor in plasma were measured by enzyme-linked immunosorbent assay. Bone marrow cell cycle was performed by flow cytometry. The expression of apoptotic protein bcl-2, bax, and caspase-3 was detected by Western blotting. ResultsBoth Re and Rk3 could improve peripheral blood cells, bone marrow nucleated cell counts, thymus index, and spleen index. Furthermore, they could enhance the yield of colonies cultured in vitro and make the levels of granulocyte macrophage colony-stimulating factor, erythropoietin, and thrombopoietin normal, reduce the ratio of G0/G1 phase cells, and increase the proliferation index. Finally, Re and Rk3 could upregulate the expression of bcl-2, whereas they could downregulate the expression of bax and caspase-3. ConclusionRe and Rk3 could improve the hematopoietic function of myelosuppressed mice. The effect of Rk3 was superior to that of Re at any dose. Regulating the levels of cytokines, promoting cells enter the normal cell cycle, regulating the balance of bcl-2/bax, and inhibiting the expression of caspase-3 may be the effects of Re and Rk3 on myelosuppression.

Indian Ginseng (Withania somnifera) supplementation ameliorates oxidative stress and mitochondrial dysfunctions in experimental model of stroke.

Stroke is an increasingly prevalent clinical condition and second leading cause of death globally. The present study evaluated the therapeutic potential of Indian Ginseng, also known as Withania somnifera (WS), supplementation on middle cerebral artery occlusion (MCAO) induced mitochondrial dysfunctions in experimental model of ischemic stroke. Stroke was induced in animals by occluding the middle cerebral artery, followed by reperfusion injury. Ischemia reperfusion injury resulted in increased oxidative stress indicated by increased reactive oxygen species and protein carbonyl levels; compromised antioxidant system; in terms of reduced superoxide dismutase and catalase activity, along with reduction in GSH levels and the redox ratio, impaired mitochondrial functions and enhanced expression of apoptosis markers. Ischemia reperfusion injury induced mitochondrial dysfunctions in terms of (i) reduced activity of the mitochondrial respiratory chain enzymes, (ii) reduced histochemical staining of complex-II and IV, (iii) reduced in-gel activity of mitochondrial complex-I to V, (iv) mitochondrial structural changes in terms of increased mitochondrial swelling, reduced mitochondrial membrane potential and ultrastructural changes. Additionally, an increase in the activity of caspase-3 and caspase-9 was also observed, along with altered expression of apoptotic proteins Bcl-2 and Bax in MCAO animals. MCAO animals also showed significant impairment in cognitive functions assessed using Y maze test. WS pre-supplementation, on the other hand ameliorated MCAO induced oxidative stress, mitochondrial dysfunctions, apoptosis and cognitive impairments. The results show protective effect of WS pre-supplementation in ischemic stroke and are suggestive of its potential application in stroke management.

Ranolazine improves oxidative stress and mitochondrial function in the atrium of acetylcholine-CaCl2 induced atrial fibrillation rats

AimsRanolazine, an antianginal agent used for chronic stable angina treatment, was demonstrated to be effective in atrial fibrillation (AF) treatment. The aim of this study was to explore the molecular mechanisms of its anti-AF effects. Main methodsAF rat model was established using acetylcholine (ACh)-CaCl2 injection for 7days followed by ACh infusion into the heart. Prior to ACh infusion, ranolazine at 10.7mg/kg/0.5ml was injected into vein and followed by 0.56mg/kg/min infusion. Blood pressure and electrocardiogram were monitored during the infusion. Histological changes of atrial tissue were observed after H&E staining. Activities and protein expression of NADPH oxidase-4, xanthine oxidase, glutathione peroxidase and superoxide dismutase were examined using commercial assay kits and Western botting, respectively. Mitochondrial functions were evaluated through membrane potential, ATP production, activities of complex I and III and reactive oxygen species production. Apoptosis was measured using TUNEL staining. Protein expression of apoptotic proteins Bcl-2, Bax and cleaved-caspase 3 and Akt/mTOR signaling proteins were detected using Western blotting. Key findingsResults demonstrated that ranolazine attenuated AF in ACh-CaCl2-exposed rats. In addition, ranolazine restored mitochondrial function, suppressed oxidative stress, and inhibited atrial cells apoptosis. Furthermore, the activated Akt/mTOR signaling pathway induced by AF was further activated by ranolazine. SignificanceThe present study confirms the effects of ranolazine on AF rats induced by ACh-CaCl2, and provides evidence that the anti-AF effects are associated with the restoration of mitochondrial function and activation of the Akt/mTOR signaling pathway in atrial tissue.

Antitumor Effects of Ginkgolic Acid in Human Cancer Cell Occur via Cell Cycle Arrest and Decrease the Bcl-2/Bax Ratio to Induce Apoptosis

Background: Ginkgolic acids (GAs), extracted from the seed coat of Ginkgo biloba L. Our previous study has shown that GA monomer could inhibit the growth of Hep-2 significantly and induce the fragmentation of the chromosomal DNA. To further assess the antitumor potential and turn it into a candidate new antitumor drug, the antitumor mechanism of GA was investigated. Method: The cytotoxicity and antitumor effect of GA monomer were assayed by MTT colorimetric assay with nontumorogenic MC-3T3-E1 as well as tumorogenic Hep-2 and Tac8113 cell lines. The effect of GA monomer on the proliferation of tumor cell lines was analyzed with MTT colorimetric and CFSE labeled assay. Cell cycle distribution and measurement of the percentage of apoptotic cells were performed by flow cytometry following stained with propidium iodide, annexin V-FITC. The expression of apoptotic proteins Bcl-2, Bax and caspase-3 was analyzed with Western blot. Result: GA only inhibited the growth of tumorogenic cell lines in a both dose- and time-dependent manner. Tumor cells were treated with GA for 72 h, 70.53 ± 4.54% Hep-2 and 63.5 ± 7.2% Tca8113 cells were retarded at GO/G1 phase, and the percentage of apoptosis was 40.4 ± 1.58 and 38.4 ± 1.7%, respectively. GA-treated activated caspase-3 downregulated the expression of anti-apoptotic Bcl-2 protein and upregulated the expression of pro-apoptotic Bax protein, eventually leading to a decrease in the Bcl-2/Bax ratio in tumor cells. Conclusions: The antitumor action of GA was due to inhibiting the proliferation in a manner of inhibiting division, retarding the progress of cell cycle and inducing apoptosis, making GA a candidate as new antitumor drug.

Progress in studies of huperzine A, a natural cholinesterase inhibitor from Chinese herbal medicine1

Huperzine A (HupA), a novel alkaloid isolated from the Chinese herb Huperzia serrata, is a potent, highly specific and reversible inhibitor of acetylcholinesterase(AChE). Compared with tacrine, donepezil, and rivastigmine, HupA has better penetration through the blood-brain barrier, higher oral bioavailability, and longer duration of AChE inhibitory action. HupA has been found to improve cognitive deficits in a broad range of animal models. HupA possesses the ability to protect cells against hydrogen peroxide, beta-amyloid protein (or peptide), glutamate, ischemia and staurosporine-induced cytotoxicity and apoptosis. These protective effects are related to its ability to attenuate oxidative stress, regulate the expression of apoptotic proteins Bcl-2, Bax, P53, and caspase-3, protect mitochondria, upregulate nerve growth factor and its receptors, and interfere with amyloid precursor protein metabolism. Antagonizing effects of HupA on N-methyl-D-aspartate receptors and potassium currents may also contribute to its neuroprotection as well. Pharmacokinetic studies in rodents, canines, and healthy human volunteers indicated that HupA was absorbed rapidly, distributed widely in the body, and eliminated at a moderate rate with the property of slow and prolonged release after oral administration. Animal and clinical safety tests showed that HupA had no unexpected toxicity, particularly the dose-limiting hepatotoxicity induced by tacrine. The phase IV clinical trials in China have demonstrated that HupA significantly improved memory deficits in elderly people with benign senescent forgetfulness, and patients with Alzheimer disease and vascular dementia, with minimal peripheral cholinergic side effects and no unexpected toxicity. HupA can also be used as a protective agent against organophosphate intoxication.

Diazene JK-279 induces apoptosis-like cell death in human cervical carcinoma cells

Diazene N-phenyl-2-(2-pyridinyl)diazenecarboxamide (JK-279) is a newly synthesized compound, cytotoxic for several tumor cell lines and their drug-resistant sublines. In human cervical carcinoma cells (HeLa), this compound reduced intracellular glutathione content and increased sensitivity to cisplatin. The aim of the present study was to elucidate the molecular mechanisms involved in the cytotoxic effect of diazene JK-279 on HeLa cells. Cytotoxicity was determined by the MTT method. Flow cytometry analysis showed that diazene JK-279 induces G 2/M phase arrest, mediated by the increase in p21 expression, and accompanied by an alteration in the expression of survivin. The highest concentration of JK-279 altered nuclear morphology in intact cells, showing “apoptosis-like” features. No cleavage of procaspase-3, procaspase-9 and PARP, or altered expression of apoptotic proteins Bcl-2 and Bax were detected. At the same time, PS externalization and internucleosomal DNA cleavage were observed. Partial necrosis was detected as well. Our results demonstrate that cytotoxicity of diazene JK-279 is mostly the consequence of caspase-independent cell death, which is in some aspects “apoptosis-like”. Taking into account the multiplicity of mechanisms used by cancer cells to prevent apoptosis, the drugs (like diazene JK-279) that would activate alternative cell death pathways could provide a useful tool for new types of cancer therapy.