Anticancer effect and apoptosis induction of gambogic acid in human gastric cancer line BGC-823

AIM: To investigate the anticancer effect of a traditional Chinese medicine gambogic acid (GA) in human gastric cancer line BGC-823 and further study the mechanism of apoptosis induction of GA.METHODS: Low differential human gastric cancer line BGC-823 were treated with GA at different doses and different times, the inhibitory rates were detected by MTT assay. Apoptosis induced by GA in BGC-823 cells was observed by Annexin-V/PI doubling staining flow cytometry assay. And T/C (%) was chosen to detect the inhibition of GA on human gastric adenocarcinoma BGC-823 nude mice xenografts. Apoptosis on nude mice xenografts was observed by Annexin-V/PI doubling staining flow cytometry assay and DNA fragmentation assay. To further determine the molecular mechanism of apoptosis induced by GA, the changes on the expression of bcl-2 and bax genes were detected by RT-PCR.RESULTS: After incubation with GA, low differential human gastric cancer line BGC-823 was dramatically inhibited in a dose-dependent manner. After these cells were exposedto GA for 24, 48 and 72 h, the IC50 value was 1.02±0.05, 1.41±0.20 and 1.14±0.19 μmol/L, respectively. Apoptosis in BGC-823 cells induced by GA was observed by AnnexinV/PI doubling staining flow cytometry assay. The apoptotic population of BGC-823 cells was about 12.96% and 24.58%, respectively, when cells were incubated with 1.2 μmol/L GA for 48 and 72 h. T/C (%) of human gastric carcinoma adenocarcinoma BGC-823 nude mice xenografts was 44.3, when the nude mice were treated with GA (8 mg/kg). Meanwhile, apoptosis induced by GA was observed in human gastric carcinoma adenocarcinoma BGC-823 nude mice xenografts. The increase of bax gene and the decrease of bc1-2 gene expressions were found by RT-PCR.CONCLUSION: The inhibition of GA on human gastric cancer line BGC-823 was confirmed. This effect connects with the inducing apoptosis in BGC-823 cells and the molecular mechanism might be related to the reduction of expression of apoptosis-regulated gene bcl-2, and the improvement of the expression of apoptosis-regulated gene bax. The result was also confirmed in vivo.

Tissue engineering of blood vessels with endothelial cells differentiated from mouse embryonic stem cells

Endothelial cells (TEC3 cells) derived from mouse embryonic stem (ES) cells were used as seed cells to construct blood vessels. Tissue engineered blood vessels were made by seeding 8 × l06 smooth muscle cells (SMCs) obtained from rabbit arteries onto a sheet of nonwoven polyglycolic acid (PGA) fibers, which was used as a biodegradable polymer scaffold. After being cultured in DMEM medium for 7 days in vitro, SMCs grew well on the PGA fibers, and the cell-PGA sheet was then wrapped around a silicon tube, and implanted subcutaneously into nude mice. After 6～8 weeks, the silicon tube was replaced with another silicon tube in smaller diameter, and then the TEC3 cells (endothelial cells differentiated from mouse ES cells) were injected inside the engineered vessel tube as the test group. In the control group only culture medium was injected. Five days later, the engineered vessels were harvested for gross observation, histological and immunohistochemical analysis. The preliminary results demonstrated that the SMC-PGA construct could form a tubular structure in 6～8 weeks and PGA fibers were completely degraded. Histological and immunohistochemical analysis of the newly formed tissue revealed a typical blood vessel structure, including a lining of endothelial cells (ECs) on the lumimal surface and the presence of SMC and collagen in the wall. No EC lining was found in the tubes of control group. Therefore, the ECs differentiated from mouse ES cells can serve as seed cells for endothelium lining in tissue engineered blood vessels.

EFFECTS OF INTEGRIN ALPHAⅡb^(R995A) MUTATION ON RECEPTOR AFFINITY AND pp125 (FAK) PHOSPHORYLATION

Objective To investigate the role of cytoplasmic domain of integrin alphaⅡb in platelet signal transduction. Methods Binding capacity of integrin alphaⅡb R995A to antibody platelet activation complex-1 (PAC-1) and pp125 focal adhesion kinase (FAK) phosphorylation of cells were detected by flow cytometry, immune precipitation, and Western blotting. Results Without activation, wild-type alphaⅡbbeta3 Chinese hamster ovary (CHO) cells failed to bind to PAC-1, but mutant chimera alphaⅡb R995A beta3 CHO cells were able to bind with PAC-1. Furthermore, phosphorylation of pp125 (FAK) in wild-type alphaⅡbbeta3 CHO cells occured only when cells were adhered to fibrinogen, but could not be detected in bovine serum albumin suspension. However in the mutant chimera group, it could be detected in both conditions. Conclusion The mutation in integrin alphaⅡb R995A alters its affinity state as a receptor, thus also mediating cytoplasmic signal transduction leading to the phosphorylation of pp125 (FAK) without ligand binding.

Determination of Surface pKa of Pure Mercaptoacetic Acid and 2-Mercaptobenzothiazole Mixed Monolayers by Impedance Titration

Interfacial proton transfer reactions of pure mercaptoacetic acid (MA) and 2-mercaptobenzothiazole (Mbz) mixed self-assembled monolayers (SAMs) have been studied using a.c. impedance titration method. The charge-transfer resistance (Rct,) is measured with the monolayer composition and the ionic strength of pH solution. The surface pKa can be obtained by the plots of Rct and pH, the reasons of shifts of surface pKa are also explained.

The Stabilization Effect of Glutaraldehyde on the Spirulina platensis Phycobilisomes

The spectral properties of the glutaraldehyde-treated phycobilisomes were studied. The results showed that glutaraldehyde was effective in preventing phycobilisomes from dilution- induced dissociation and preserving the intra-phycobilisomes energy transfer.

Effect of rituximab combined with CVAD regimen on serum VEGF and β2-MG levels in patients with primary gastrointestinal B-cell lymphoma

Objective:To investigate the clinical effect of rituximab combined with CVAD regimen in patients with primary gastrointestinal B-cell lymphoma and serum vascular endothelial growth factor (VEGF) andβ2 microglobulin (β2-MG) The impact of the level.Methods:Eighty-four patients with primary gastrointestinal B-cell lymphoma treated from May 2014 to December 2015 were enrolled. Based on the random number table, all the patients were divided into a control group (n=42) and an observation group (n=42). The control group was treated with CVAD. The observation group was treated with rituximab on the basis of the control group. The effect of the patients was evaluated after 3 courses of treatment. The patients were followed up for 3 years after treatment. US RECIST 1.1 was used to evaluate the short-term efficacy on the patients;VEGF, TNF receptor-associated factor 6 (TRAF6) and B-cell lymphoma factor-6 (Bcl-6) levels were measured by enzyme-linked immunosorbent assay;β2-MG level test was implemented to compare the short-term efficacy, biochemical indicators, incidence of toxic side effects and long-term survival rate of the two groups. Results: The short-term efficacy rate of the observation group was 76.19%, which was higher than that of the control group (50.00%) (P<0.05). The levels of VEGF, TRAF6, Bcl-6, andβ2-MG were lower in the observation group after 3 courses of treatment than that in the control group (P<0.05);there was no significant difference in the incidence of neutropenia, gastrointestinal reactions, sepsis, infection, infusion-related reactions and cardiovascular events between the observation group and the control group (P>0.05);The 1-year long-term survival rate after treatment was not statistically significant (P>0.05). The long-term survival rate of the observation group was higher than that of the control group at 2 and 3 years after treatment (P<0.05).Conclusion: The combination of rituximab and CVAD in patients with primary gastrointestinal B-cell lymphoma can improve short-term efficacy, lower VEGF andβ2-MG levels, and lower incidence of side effects. It can improve the long-term survival rate of patients and is worthy of promotion and application.

Lipopolysaccharide Could Be Internalized into Human Peripheral Blood Mononuclear Cells and Elicits TNF-α Release,but not via the Pathway of Toll-Like Receptor 4 on the Cell Surface

Lipopolysaccharide(LPS),the principal component of the outer membrane of Gram-negative bacteria,stimulates various cell types to release numerous proinflammatory mediators such as TNF-α,IL-6 and IL-12,which may damage cells and lead to organ injury,even sepsis and septic shock.Toll-like receptor 4(TLR4) has been identified as the receptor involved in the recognition of LPS,but the role of LPS uptake in activating signal transduction remains controversial.In the present study,TNF-α was used as a marker of macrophages/ monocytes activated by LPS,and CQ was used as an inhibitor of endosome mature in order to definitude what stage of the signal transduction elicited by LPS was interrupted.We found that there indeed existed internalization of LPS and internalization partially participated in LPS signaling since CQ inhibited cytokine release,and decreased accumulation of FITC-LPS in hPBMCs.In contrast,anti-hTLR4 antibody could decrease cytokine release,but had no inhibition on accumulation of FITC-LPS.This result revealed that inhibition of cytokine release was related to reduction of FITC-LPS accumulation in the cells.But TLR4 on the cell surface couldn't participate in internalization of LPS.Thus,LPS signaling and internalization couldn't be viewed as mutually independent processes.Cellular & Molecular Immunology.2004;1(5):373-377.

Association of HLA-DQ with Idiopathic Dilated Cardiomyopathy in a Northern Chinese Han Population

Autoimmune mechanisms are likely involved in the pathogenesis of idiopathic dilated cardiomyopathy (IDC) and components of MHC may serve as markers for the propensity to develop immune-mediated myocardial damage.This study was conducted to investigate the possible association between HLA-DQA1,-DQB1 alleles and IDC in Han population from northern China by using PCR-based sequence-specific primer (PCR-SSP) technique for HLA genotyping.Among 68 unrelated IDC patients,4 probands of IDC pedigrees and 100 healthy controls,we found that the alleles of HLA-DQA1*0501 and HLA-DQB1*0303 conferred susceptibility to IDC while HLA-DQA1*0201 and HLA-DQB1*0502,*0504 alleles were in negative association with IDC.The serine at position 57 (SER^(57)) in the exon of HLA-DQB1*0502 and *0504 was confirmed in our experiment as a marker for resistance to IDC.The results suggest that HLA-DQ polymorphism may be involved in the pathogenesis of IDC.Cellular & Molecular Immunology.2004;1 (4):311-314.