Effects of small interfering RNA inhibit Class Ⅰ phosphoinositide 3-kinase on human gastric cancer cells

AIM: To investigate the effects of small interfering RNA (siRNA)-mediated inhibition of Class?I?phosphoinositide 3-kinase (Class?I?PI3K) signal transduction on the proliferation, apoptosis, and autophagy of gastric cancer SGC7901 and MGC803 cells.METHODS: We constructed the recombinant replication adenovirus PI3K(I)-RNA interference (RNAi)-green fluorescent protein (GFP) and control adenovirus NC-RNAi-GFP, and infected it into human gastric cancer cells. MTT assay was used to determine the growth rate of the gastric cancer cells. Activation of autophagy was monitored with monodansylcadaverine (MDC) staining after adenovirus PI3K(I)-RNAi-GFP and control adenovirus NC-RNAi-GFP treatment. Immunofluorescence staining was used to detect the expression of microtubule-associated protein 1 light chain 3 (LC3). Mitochondrial membrane potential was measured using the fluorescent probe JC-1. The expression of autophagy was monitored with MDC, LC3 staining, and transmission electron microscopy. Western blotting was used to detect p53, Beclin-1, Bcl-2, and LC3 protein expression in the culture supernatant.RESULTS: The viability of gastric cancer cells was inhibited after siRNA targeting to the Class?I?PI3K blocked Class?I?PI3K signal pathway. MTT assays revealed that, after SGC7901 cancer cells were treated with adenovirus PI3K(I)-RNAi-GFP, the rate of inhibition reached 27.48% ± 2.71% at 24 h, 41.92% ± 2.02% at 48 h, and 50.85% ± 0.91% at 72 h. After MGC803 cancer cells were treated with adenovirus PI3K(I)-RNAi-GFP, the rate of inhibition reached 24.39% ± 0.93% at 24 h, 47.00% ± 0.87% at 48 h, and 70.30% ± 0.86% at 72 h (P < 0.05 compared to control group). It was determined that when 50 MOI, the transfection efficiency was 95% ± 2.4%. Adenovirus PI3K(I)-RNAi-GFP (50 MOI) induced mitochondrial dysfunction and activated cell apoptosis in SGC7901 cells, and the results described here prove that RNAi of Class?I?PI3K induced apoptosis in SGC7901 cells. The results showed that adenovirus PI3K(I)-RNAi-GFP transfection induced punctate distribution of LC3 immunoreactivity, indicating increased formation of autophagosomes. The results showed that the basal level of Beclin-1 and LC3 protein in SGC7901 cells was low. After incubating with adenovirus PI3K(I)-RNAi-GFP (50 MOI), Beclin-1, LC3, and p53 protein expression was significantly increased from 24 to 72 h. We also found that Bcl-2 protein expression down-regulated with the treatment of adenovirus PI3K(I)-RNAi-GFP (50 MOI). A number of isolated membranes, possibly derived from ribosome-free endoplasmic reticulum, were seen. These isolated membranes were elongated and curved to engulf a cytoplasmic fraction and organelles. We used transmission electron microscopy to identify ultrastructural changes in SGC7901 cells after adenovirus PI3K(I)-RNAi-GFP (50 MOI) treatment. Control cells showed a round shape and contained normal-looking organelles, nucleus, and chromatin, while adenovirus PI3K(I)-RNAi-GFP (50 MOI)-treated cells exhibited the typical signs of autophagy.CONCLUSION: After the Class?I?PI3K signaling pathway has been blocked by siRNA, the proliferation of cells was inhibited and the apoptosis of gastric cancer cells was enhanced.

A theoretical study of miRNA155-DNA methylation mediated mitokATP regulating mitochondrial autophagy in the improvement of myocardial ischemia-reperfusion injury by Qishen Yiqi Dropping Pills

In the state of acute myocardial ischemia,miRNA expression can regulate related genes and proteins,reduce myocardial cell damage,and thus play a protective role in the myocardium.However,the specific mechanism still needs to be further explored.Recent studies have found that the opening of the mitoKATP channel can regulate mitochondrial autophagy,and the initiation of miRNA-DNA methylation plays a regulatory role in inducing cell autophagy.The applicant research team previously found that Qishen Yiqi Dropping Pills could significantly improve myocardial ischemia by mediating MitokATP channels to regulate mitochondrial autophagy.,and animal experiments have confirmed that miR-155 plays a significant role in the aspect of autophagy regulates,inflammatory reaction and Vascular smooth muscle cell migration.Therefore,the applicant innovatively proposed that Qishen Yiqi Dropping Pills can regulate miRNA155-DNA methylation to mediate the opening of mitoKATP,thereby regulating mitochondrial autophagy and improving myocardial ischemia.In this paper,the association between mitochondrial autophagy and oxidative stress injury after myocardial ischemia was described,and the possible mechanism of Qisen Yiqi dropping pills regulating mitochondrial autophagy by regulating miRNA155-DNA methylation to mediate MitokATP to improve myocardial ischemia reperfusion injury was discussed,so as to provide theoretical ideas for related research.

Effects of Qishen Yiqi dripping pills-containing serum on KATP channel opening and PI3K/AKT signaling pathway in hypoxic/reoxygenated H9C2 cardiocytes

Objective:To investigate the effects of Qishen Yiqi dropping pills serum on KATP channel opening and PI3K/AKT signaling pathway of hypoxic/reoxygenated H9C2 cardiocytes.Methods:H9C2 cardiocytes cultured in vitro were randomly divided into five groups,A:H9C2 cell group B:H9C2 cells+H2O2 model group C:H9C2 cells+H2O2 model+Qishen Yiqi group D:H9C2 cells+H2O2 model+Qishen Yiqi+wort group E:H9C2 cells+H2O2 model+Qishenyiqi+5-HD group,the drug intervention is according to the corresponding conditions.CCK-8 method was used to detect the cell activity of each group;Western blot was used to detect the expression of AKT and P-Akt proteins in myocardial cells in each group.The current was recorded by the standard patch clamp whole cell recording method,and the current was collected and analyzed by Pclamp6.0 software.Results:CCK-8 test results showed that compared with group A,the activity of myocardial cells in group B was significantly decreased,and the difference was statistically significant(P<0.01);compared with group B,the difference in group C was statistically significant(P<0.01);compared with C,cardiomyocyte activity in D and E group were significantly decreased,and the difference was statistically significant(P<0.05);WB results showed that compared with A,p-Akt protein expression in B,C,D and E groups were significantly decreased,and the difference was statistically significant(P<0.01);compared with group B,p-Akt protein expression in C,D and E group were significantly increased,and the difference was statistically significant(P<0.01),but there was no significant difference in AKT expression among groups(P>0.05);The results of whole cell patch clamp experiment showed that the outward current of B was significantly increased compared with that of A,and the difference between groups was statistically significant(P<0.01);compared with group B,cardiomyocytes in group C further increased the outward current,and the difference between groups was statistically significant(P<0.01);compared with C,the current of D and E group were significantly decreased,with statistical significance between groups(P<0.01).Conclusion:QishenYiqi dropping pills can protect cardiomyocytes by activating p-Akt protein expression and KATP channel opening in H9C2 cardiomyocytes.

Update on establishment of TCM animal model of myocardial ischemia and Chinese medicine treatment

The prevention and treatment of myocardial ischemia is a hot and difficult point in clinical research and animal experimental research in the cardiovascular field.The combined disease and syndrome animal model with the disease characteristics of Western medicine and guided by the theory of Chinese medicineto replicate the"syndrome"of Chinese medicine on the model animals by using experimental methods is an effective tool for the research of Chinese medicine.This article summarized the methoding methods and evaluation index of the animal model combined disease of coronary heart disease myocardial ischemia with cold blood stasis,phlegm and blood stasis,qi deficiency and blood stasis syndrome,kidney deficiency and blood stasis,and concluded the treatment of Traditional Chinese medicine.At the same time,it analyzed the main deficiencies of myocardial ischemia syndrome combined with animal models to provide reference for the establishment and research of related animal models.

Research progress of long non-coding RNA in dilated cardiomyopathy

Dilated cardiomyopathy(DCM)is a type of composite cardiomyopathy characterized by left ventricular,right ventricular or double ventricular enlargement and diastolic dysfunction caused by genetic and non-genetic causes,with a high incidence,High mortality,gradual rejuvenation,etc.,are common types of cardiomyopathy.With the progressive development of the disease,it can cause arrhythmia,severe heart failure,thromboembolism,and even sudden death.Currently,there is no effective treatment plan,and the high mortality rate is a major problem in the treatment of cardiovascular diseases.Long-chain non-coding RNAs(lncRNAs)are a class of RNAs with a molecular mass of more than 200 bt and which are not involved or rarely involved in protein coding.Studies have shown that lncRNAs play an important role in the occurrence and development of DCM.Finding relevant clinical diagnostic markers and therapeutic targets is the current research focus in the field of DCM.This article reviews the mechanism and research progress of lncRNAs in the occurrence and development of DCM,and aims to provide a new direction for the prevention and treatment of DCM.

A new polyketide synthase nonribosomal peptide synthetase hybrid metabolite from plant endophytic fungus Periconia sp.

A new polyketide synthase-nonribosomal peptide synthetase （PKS-NRPS） hybrid metabolite, named pericoannosin B （1）, was isolated from endophytic fungus Periconia sp. F-31 of the medicinal plant Annona rnuricata. The structure and absolute configuration were elucidated by means of extensive spectroscopic analyses （HRMS, NMR, IR, and CD）.