Studies On Antihypertensive Effect And Mechanism Of Recombinant Lactobacillus Plantarum Expressing Angiotensin Converting Enzyme Inhibitory Peptide

Hypertension is known to be the most impotant risk factor for cardiovascular disease. Because of its high incidence, hypertension has been thought as the first murderer to the patients nowadays and could be a threat to the health of human beings. Currently, people are constantly developing new drugs for the treatment of hypertension, but these drugs often have pain, fever, nausea, abdominal distension, dry cough and other side effects. So, it is very necessary to look for a safe material which is having the biological activity and could fall the blood pressure effectively to avoid the side effects emerging in the traditional medicines. While compared to the synthetic drugs some peptides from natural protein have overcome the shortcoming of low activity and have demonstrated these advantages of safety, validity and low side effects. With the rapid development of genetic engineering techniques, in oder to overcome the difficulties of direct extraction, high costs and other issues of chemical, recombinant DNA technology has advantages to express antihypertensive peptides. In this study, ACEIP gene was cloned into the E.coli-Lactobacillus shuttle vector, was transformed into lactic acid, had high blood pressure activity of recombinant lactic acid bacteria, and to lay a foundation of further study on the lactic acid bacteria products development of lowering blood pressure.The specific studies are as follows:The angiotensin converting enzyme inhibitory peptides (ACEIP) from frame protein tuna and yellowfin sole frame protein both have the effect of treating hypertension, which were obtained by means of enzymatic hydrolysis. In this study, these peptides sequence data of ACEIP connected with arginine were used to synthesize complementary oligonucleotides, and then were recombined with the plasmid vector pSIP-409 which could express in both Escherichia coli and lactobacilli. The recombinant plasmid was transformed into E. coli DH5a and screened the positive recombinants. When it could successfully express in Escherichia coli, the plasmid was electrotransformed into the Lb. plantarum NC8. The expression of the recombinant protein was induced in Lb. plantarum NC8 using Sakacin-P, and detected by electrophoresis of SDS-PAGE. The SDS-PAGE showed that a specific protein band of about 8.6 kDa expressed in Lb. Plantarum NC8. The results showed that the ACEIP gene was expressed successfully in Lb. Plantarum NC8. Basis of it, the recombinant Lb. plantarum NC8 was named RLP.The antihypertensive effects of the recombinant Lb. plantarum NC8 were determined by measuring the blood pressure following oral administration of it on 12-week-old spontaneously hypertensive rats, and also haematology, blood biochemistry and NO, ET and Ang II levels have been detected. The results showed that antihypertensive effect in SHR revealed that oral administration of recombinant Lb. plantarum NC8 can decrease systolic blood pressure significantly (P<0.01), and after drug withdrawal, antihypertensive effect lasted two weeks. Furthermore, administration of recombinant Lb. plantarum NC8 leads to increased the NO level in heart, plasma and kidney, reduced ET and Angâ…¡levels, and there was no significant change in haematology and blood biochemistry except for lowering triglyceride level. Recombinant lactic acid bacteria could reduce blood pressure by reducing angiotensin levels, regulating of vascular endothelial function and relexing blood vessels.