Bax inhibitor 1 inhibits vascular calcification in mice by activating optic atrophy 1 expression / 南方医科大学学报

OBJECTIVE@#To investigate the effects of Bax inhibitor 1 (BI- 1) and optic atrophy protein 1 (OPA1) on vascular calcification (VC).@*METHODS@#Mouse models of VC were established in ApoE-deficient (ApoE-/-) diabetic mice by high-fat diet feeding for 12 weeks followed by intraperitoneal injections with Nε-carboxymethyl-lysine for 16 weeks. ApoE-/- mice (control group), ApoE-/- diabetic mice (VC group), ApoE-/- diabetic mice with BI-1 overexpression (VC + BI-1TG group), and ApoE-/- diabetic mice with BI-1 overexpression and OPA1 knockout (VC+BI-1TG+OPA1-/- group) were obtained for examination of the degree of aortic calcification using von Kossa staining. The changes in calcium content in the aorta were analyzed using ELISA. The expressions of Runt-related transcription factor 2 (RUNX2) and bone morphogenetic protein 2 (BMP-2) were detected using immunohistochemistry, and the expression of cleaved caspase-3 was determined using Western blotting. Cultured mouse aortic smooth muscle cells were treated with 10 mmol/L β-glycerophosphate for 14 days to induce calcification, and the changes in BI-1 and OPA1 protein expressions were examined using Western blotting and cell apoptosis was detected using TUNEL staining.@*RESULTS@#ApoE-/- mice with VC showed significantly decreased expressions of BI-1 and OPA1 proteins in the aorta (P=0.0044) with obviously increased calcium deposition and expressions of RUNX2, BMP-2 and cleaved caspase-3 (P= 0.0041). Overexpression of BI-1 significantly promoted OPA1 protein expression and reduced calcium deposition and expressions of RUNX2, BMP-2 and cleaved caspase-3 (P=0.0006). OPA1 knockdown significantly increased calcium deposition and expressions of RUNX2, BMP-2 and cleaved caspase-3 in the aorta (P=0.0007).@*CONCLUSION@#BI-1 inhibits VC possibly by promoting the expression of OPA1, reducing calcium deposition and inhibiting osteogenic differentiation and apoptosis of the vascular smooth muscle cells.

Cloning and expression analysis of Bax inhibitor-1 gene from medicinal fungus Polyporus umbellatus / 中草药

Objective: To clone the Bax inhibitor-1 (BI-1) gene from Polyporus umbellatus and perform the bioinformatics and expression mode analysis. Methods: To clone the full-length cDNA of BI-1 using RACE technology. The characteristics of physiochemical properties, conserved domains, and transmembrane domain of the predicted BI-1 protein were determined using bioinformatic tools. Results: The full-length cDNA of BI-1 gene was 1 091 bp in length and encoded a 334-aa protein with a molecular weight of 36170 and an isoelectric point (pI) of 10.51. The polypeptide chain was a hydrophobin with five hydrophobic regions. The PuBI-1 belonged to basidiomycete group according to the phylogenetic analysis. Real time quantitative PCR (RT-qPCR) analysis revealed that the transcription level of PuBI-1 gene was significantly higher in the beginning and developing stages of sclerotial formation with 3.8 and 7.497 fold over those in the mycelium, but the transcripts decreased sharply with the sclerotial development. Conclusion: Molecular characterization and expression patten of PuBI-1 gene will be useful for the further functional determination of the gene during the development of P. umbellatus sclerotium.

The effect of RNA interference induced by inhibition of hTERT and Bi-1 gene expression / 重庆医学

Objective In this study ,we constructed a series of recombinant plasmids carriers expressing shRNA targeting hTERT and Bi‐1 gene .These recombinant plasmids carriers were transfected into CNE‐2Z cell lines using Lip and continuously in‐duced the expression of shRNAs .Furthermore ,the shRNAs caused the degradation of mRNAs homologous in sequence with the target genes ,which lead to a sequence‐specific gene silencing .Methods The CNE‐2Z cells was divided into untreated group ,pEG‐FP‐N1 group and pEGFP‐N1/Lip group .Flow cytometry(FCM ) was applied to determine the transfection efficiency .The changes of hTERT and Bi‐1 gene expression were detected by Real‐time RT‐PCR and Western blotting .Results The best transfection effi‐ciency between plasmid and Lip was 2 .5 μg plasmid and 6 .25μL Lip .Conclusion We constructed several shRNA recombinant eu‐karyotic expression plasmids successfully .The recombinant plasmid can inhibit the expression of hTERT and Bi‐1 gene specifically and effectively .

Effects of Chronic Mild Stress in Female Bax Inhibitor-1-Gene Knockout Mice

OBJECTIVE: The anti-apoptotic protein Bax inhibitor-1 (BI-1) is a regulator of apoptosis linked to endoplasmic reticulum (ER) stress, and BI-1-/- mice exhibit increased sensitivity to tissue damage. The purpose of this study was to investigate the role of BI-1 in the pathogenesis of chronic mild stress (CMS)-induced depression-like behaviors in BI-1-/- mice. METHODS: We delivered CMS for 2 or 6 weeks in BI-1-knockout and wild-type mice. Control groups of BI-1-knockout and wild-type mice were left undisturbed. The measured parameters were sucrose consumption at weeks 1, 2, 3, 4, 5, and 6, spontaneous locomotion, and a forced swimming test (FST) at weeks 2 and 6. RESULTS: Significant decreases in sucrose consumption and increases in immobility time in the FST were observed in both stress groups compared with the non-stress groups. Interestingly, at week 2, but not at week 6, BI-1-/--stress mice showed less sucrose intake and greater immobility time than did BI-1+/+-stress mice. CONCLUSION: These results suggest that BI-1 may play role in protecting against the depressogenic effects of CMS in the short term, but not in the long term. Further study is required to deepen understanding of the role of BI-1 in protecting against depression.

Ovarian serous cystadenocarcinoma cell apoptosis induced by inhibition of Bax inhibitor-1(bi-1) gene expression with RNA interference / 中国病理生理杂志

AIM: To study the inhibition of the Bax inhibitor-1(bi-1) gene expression induced with RNA interference in HO8910PM and HO8910 cell lines in addition to the apoptotic induction.METHODS: After transfection of recombinant plasmid which expresses bi-1shRNA into HO8910PM and HO8910 cells,bi-1 mRNA and protein levels were detected by reverse transcription-polymerase chain reaction(RT-PCR) and Western blotting.Apoptotic HO8910PM and HO8910 cells were detected by Hoechst 33258/PI fluorescent staining and flow cytometry.RESULTS: Compared with the control group,the expressions of bi-1 gene at mRNA and protein levels were declined evidently in the cells transfected with four candidate siRNA plasmids(P