Arnt2 nucleocytoplasmic translocation in the process of CGNs apoptosis induced by low potassium

AIM：To analyze the changes of Arnt2 subcellular localization in the process of cerebellar granule neurons (CGNs) apoptosis induced by low concentration of potassium.METHODS：Western blotting was used to detect the variation of Arnt2 proteins in nuclei extracts of CGNs in the process of apoptosis induced by low concentration of potassium.Laser scanning confocal microscopy (LSM) was used to detect the changes of Arnt2 subcellular localization.RESULTS：In nuclei extracts of CGNs,when treated with low concentration of potassium,Arnt2 protein was up-regulated obviously at 30 min and peaked at 1 h,returned to the level of normal control at 6 h.LSM analysis result showed that Arnt2,which located in normal CGNs nuclei,was translocated into cytoplasm after induced by low concentration of potassium gradually.At the late stage of apoptosis,Arnt2 proteins located in cytoplasm and overlapped with HSP60 completely,which was regarded as a protein of mitochondria localization.CONCLUSIONS：Arnt2 protein translocates from nuclei into cytoplasm (probably into mitochondria) during CGNs apoptosis evoked by low concentration of potassium;suggesting that Arnt2 is involved in apoptosis probably by transmitting apoptotic or survival signals evoked by potassium directly.     

Hormonal and neural regulation of intestinal function in pigs

At birth, the development of hormonal and neural systems which control gut functions is not finished in pigs. However, it seems to be much more advanced than that in rodents. The relative immaturity is manifested by dynamic changes in the expression of gut regulatory peptides, tissue hormones and their receptors during the early postnatal period. The structure and function of the vagal nerves and enteric nervous system change dynamically as well. The development of neural system is manifested by changes in neuron density, morphology, and distribution of transmitters and modulators. However, little information on this topic is available, particularly in pigs. Recently, the relationship between the development of enteric nervous system and pacemaker interstitial cells of Cajal was described. Colostrum and milk contain a large number of bioactive substances involving regulatory peptides, hormones and growth factors structurally identical to those produced endogenously. Thus colostrum and milk provide the neonate not only with energy but also with the means to control the development, in particular that of the gut as well as maturation of hormonal and neural systems controlling gut function. A number of newly discovered hormones, regulatory peptides and neurotransmitters such as leptin, ghrelin, orexin A and B, and other, will be discussed as well as regulation of gut mucosa function in terms of a balance between cell proliferation and programmed death (apoptosis and autophagy).     

Dengue virus type 2 induces apoptosis and expression of death receptor in hepatic veno-endotheliocyte ED25

AIM: To investigate apoptosis and the expression of death receptors of TRAIL,TNF and Fas on hepatic veno-endotheliocyte ED25 cell strain induced by dengue virus type 2(DV2).METHODS: Flow cytometric analysis was used to detect the number of apoptotic cells and the expression levels of TRAILR1-4 ，TNFR1-2，Fas on ED25 cells before/after DV2 infection.RESULTS: The numbers of apoptotic cells of ED25 increased after DV2 infection,there were only about 5.7%±1.2% of apoptotic cells before virus infection while there were approximately 27.3%±1.6% of apoptotic cells after virus infection.At the same time the expression level of Fas also increased,before virus infection about 44.3%±2.2% of ED25 cells expressed Fas while 63.0%±2.3% of ED25 cells expressed Fas after virus infection.CONCLUSION: DV2 infection can induce apoptosis of ED25 cells,and it suggests strongly that Fas/FasL may be involved in the apoptotic signal transduction.     

Advances in the mechanism of retina ischemia-reperfusion injury

Retinal ischemia-reperfusion injury (RIRI) is a common cause of visual impairment and blindness. At the cellular level, ischemic and reperfuion retinal injury consists of a self-reinforcing destructive cascade involving oxidative stress initiated by energy failure, inflammatory reaction, calcium influx, increased glutamatergic stimulation and neuronal depolarisation and apoptosis. A number of animal models and analytical techniques have been used to study the retinal ischemia-reperfusion injury, we now understand much better than ever before in the mechanism of RIRI, and an increase in the therapeutic strategies has been developed experimentally to attenuate the detrimental effects of retinal ischemia-reperfusion injury. Thus far, however, success in the laboratory has not been translated to the clinic. Given the increasing understanding of the events involved in ischemia-reperfusion neuronal injury, it is hoped that clinically effective treatments for retinal ischemia-reperfusion injury will soon be available.     

Expression of TGF-β1 and apoptosis in myocardium of diabetic rats

AIM: To study the pathophysiological mechanism of cardiomyopathy, the expression of TGF-β₁ and apoptosis in myocardium of diabetic rats were investigated. METHODS:The diabetes models were established by single intravenous injection of streptozotocin (50 mg/kg) in rats. By the method of immunochemistry, the expression of TGF-β₁ in the cardiomyocytes was detected as the index to evaluate the degree of fibrosis. The method of TUNEL was used to measure the cardiomyocyte apoptosis as the index to explore its importance in process of diabetic cardiomyopathy. RESULTS:① The weight of diabetic rats was apparently lower than that in the rats before the diabetic model was built (P<0.01), and the increase in weight in diabetic rats within three month was less than that in normal group. ② Compared with control group, the concentration of blood glucose was continually elevated during the experiment. ③ The expression of TGF-β₁ in the diabetic cardiac muscle was much more than that in normal group (P<0.01). ④ The apoptosis of myocardium measured by the method of TUNEL was apparent in the diabetic groups than that in normal one (P<0.01). However, no significance was detected in the different courses of diabetic groups. CONCLUSIONS:The apoptosis might play an important role in leading the diabetic cardiomyopathy to heart failure. The expression of TGF-β₁ in the myocardium of diabetic rats was more than that in normal and had an increasing trend in the procession of diabetic cardiomyopathy. TGF-β₁ might be a significant factor in diabetic myocardium fibrosis. Apoptosis might play an important role in the initial stage of diabetes, which promotes the diabetic cardiomyopathy to heart failure.     

Effect of low molecular weight heparin on hypoxia-induced apoptosis in primary cultured neonate rat cerebral cortical neurons

AIM:To investigate the mechanism by which low molecular weight heparin (LMWH) inhibits apoptosis of primary cultured cerebral cortical neurons caused by hypoxia. METHODS:The anti-apoptosis effect of LMWH on primary cultured neurons was observed by methods of the primary culture of cerebral neurons of postnatal rats in free-serum with neurobasal medium supplied with 2% B27 supplement, hypoxic culture of neurons, Hoechst 33342 staining and immunocytochemistry. RESULTS:At concentrations of 250-500 U/L, LMWH reduced apoptosis rate of cerebral cortical neurons induced by hypoxia (P<0.05) and apoptosis rate was lower in LMWH groups than that in BDNF (50 μg/L) group (P<0.05). LMWH (500 U/L) increased Bcl-2 protein expression (P<0.05) and decreased Bax protein expression (P<0.01) in the cerebral cortical neurons induced by hypoxia, the ratio of Bcl-2/Bax was improved (P<0.01). The ratio of Bcl-2/Bax in LMWH (500 U/L) group was higher than that in normal control group, BDNF group and apoptosis positive group (P<0.05). CONCLUSION:LMWH at concentrations of 250-500 U/L is able to prevent cerebral cortical neurons from apoptosis in primary culture under hypoxia. The effect of anti-apoptosis may be related to up-regulation of Bcl-2 protein expression, down-regulation of Bax-2 protein expression, and increase in the ratio of Bcl-2/Bax.     

The effect of Shenmai injection on cardiac myocyte apoptosis after hypoxia

AIM: To observe the effect of Shenmai injection, a chinese medicine, on apoptosis of cardiac myocytes after hypoxia. METHODS: Cardiac myocytes were separated from neonate rat heart and cultivated in vitro. Hypoxia condition was induced by mixture of 95%N2 and 5%CO2. Cells were exposed to hypoxia for 6 h or 12 h and treated with Shenmai injection (5 mL/L) from 24 h before hypoxia until the end of hypoxia. First, apoptosis was detected with Annexin V-FITC and PI staining by flowcytometry. Then, the activity of cardiac myocyte mitochondria was observed by MTT method. Mitochondria membrane potential and the activity of caspase 3,7 were also measured by laser scan microscopy and multi-detection microplate reader, respectively. RESULTS: The apoptotic cells became more and more with prolonged hypoxia. Shenmai injection enhanced mitochondria activity, kept membrane potential, inhibited the activation of caspase3,7 and then decreased apoptotic cells (P<0.01). CONCLUSION: Hypoxia induces apoptosis in cardiac myocytes by mitochondria pathway. Shenmai injection can decrease cardiomyocyte apoptosis after hypoxia, the mechanism is related to mitochondria membrane potential stabilization and caspase inhibition.     

Apoptosis induction in gastric carcinoma cells by celecoxib combined with adriamycin

AIM:To study the apoptosis induction of cyclooxygenase-2 (COX-2) inhibitor，celecoxib and adriamycin (ADM) on tumor apoptosis of gastric carcinoma MGC-803 cells, and to explore their possible molecular mechanism(s) and interactions.METHODS:The number of MGC-803 cells was observed by MTT assay. Tumor apoptosis was studied by fluorescence microscopy, flow cytometry (FCM), and DNA ladder. RESULTS:MGC-803 cell number was significantly decreased with increasing dose of ADM. Cells were accumulated in G₀/G₁ phase and the number of cells in S phase was decreased. ADM (5 mg/L) combined with celecoxib (25 μmol/L) markably inhibited the growth of MGC-803 cells. Significant morphological changes of typical apoptosis were observed after treatment with combined use of celecoxib and ADM. Compared with ADM or celecoxib alone, ADM plus celecoxib obviously enhanced the DNA ladder fragment revealed by agarose gel electrophoresis of DNA. After exposure to combined celecoxib and ADM treatment for 48 h, MGC-803 cells were accumulated in G₀/G₁ phase. There was a decrease in the number of cells in S phase as compared to celecoxib or ADM alone. CONCLUSION:Celecoxib and ADM appear to have synergistic effects for the apoptosis induction. This may be an important prospect for applying COX-2 inhibitors to assist chemical therapy of ADM in clinical use.     

Stat5b signaling pathway regulates the expression of Survivin and promotes apoptosis in human colon cancer cells

AIM: The purpose of the study was to examine colon cancer cell lines to determine whether Stat5b/Survivin plays an important role in the process of apoptosis in colon cancer cells. METHODS: Protein lysates were extracted from colon cancer cells. Human colon cancer cell line HT29 was transfected with Stat5b antisense oligonucleotide mediated by liposome. MTT assay was used to measure the proliferation. Flow cytometry was applied to analyze the cell cycle and apoptosis. EMSA was used to detect the activity of Stat5. Western blotting was applied to measure the expression of Stat5, p-Stat5, cyclin D1, Survivin, Bcl-2 and Bcl-x L. RESULTS: Targeting of Stat5 using antisense oligonucleotide against the translation site resulted in apoptosis and downregulaed the expressions of Stat5, p-Stat5, cyclin D1 and Survivin, but not Bcl-2 and Bcl-xL. CONCLUSION: Constitutive activation of Stat5 is associated with the carcinogenesis of colon cancer cells. Blocking of Stat5 signaling inhibits the expression of Survivin and induces apoptosis in colon cancer cells.