Tumstatin (Tum) is a powerful angiostatin that inhibits proliferation and induces apoptosis of tumorous vascular endothelial cells. A nonpathogenic and anaerobic bacterium, Bifidobacterium longum (BL), selectively localizes to and proliferates in the hypoxia location within solid tumor. The aims of this study were to develop a novel delivery system for Tum using engineered Bifidobacterium and to investigate the inhibitory effect of Tum on tumor in mice. A vector that enabled the expression of Tum under the control of the pBBADs promoter of BL was constructed and transformed into BL NCC2705 by electroporation. The mouse colon carcinoma cells CT26 (1 × 10(7)/mL) were subcutaneously inserted in the left armpit of BALB/c mice. The tumor-bearing mice were treated with Tum-transformed BL, and green fluorescent protein (GFP)-transformed BL was used as a negative control. The microvessel density (MVD) in the transplanted tumor was determined, and terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate nick end labeling was used to detect apoptosis of vascular endothelial cells in transplanted tumor. The in vitro expression of Tum was examined in BL after l-arabinose induction. Bifidobacterium longum with pBBAD-Tum (BL-Tum) showed significant antitumor effect in tumor-bearing mice. The weight, volume, growth, and MVD, as well as the percentage of apoptotic vascular endothelial cells of transplanted tumors in the tumor-bearing mice treated with Tum-transformed BL were all significantly lower than those in the GFP negative control group. Intragastric administration, injection in tumor and vena caudalis injection of Tum-transformed BL exerted marked antitumor effects in tumor-bearing mice. This is the first demonstration of the utilization of Tum-transformed BL as a specific gene delivery system for treating tumor.

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Recent studies showed that probiotics could improve metabolic syndrome, making the identification of factors affecting metabolic control more important than ever.The mammalian sirtuin protein family has received much attention for its regulatory role, especially in various mitochondrial ATP, glucose, and lipid metabolic pathways.However, compared with the mammalian sirtuin protein family, the function of prokaryotic sir2 protein is much less known. We studied the effects of probiotics sir2 protein on cell energy metabolize pathway, which showed that deletion of Enterococcus faecalis sir2 inhibited the aerobic oxidation of bacteria and increased the bacterial fermentation. The study of EF-sir2 (sir2 protein of E. faecalis) role of molecular targets demonstrated that deacetylation of EF-sir2 was via Rho upregulating in E. faecalis.When transfected into HEK293T cells, EF-sir2 could significantly facilitate aerobic oxidation of glucose, enhance the respiration to generate more ATP, and cause upregulation of NRF1 target gene. Then, we found EF-sir2 could increase activity of PGC-1α by deacetylation and PGC-1α inhibition decreased the expression of NRF1 target gene. Finally, we demonstrated that EF-sir2 could significantly improve the metabolic index of mammalian cells through insulin resistanced model in vitro and metabolic syndrome rat model in vivo. Our results first revealed that prokaryotic sir2 genes affect the molecular mechanism of cellular metabolism and the regulatory of cell homeostasis in prokaryotic and mammalian cells, suggesting that EF-sir2 has a positive regulatory effect on metabolic disturbance and may be used for the prevention and treatment of pathological processes related to metabolic syndrome. KEYWORDS cellular energy, metabolic syndrome, NRF1, PGC-1α, probiotics, sir2-like gene * Shiyu Li and Zhengbin Fei contributed equally to this work.