Studies On The Anti-bacterial Diarrhea And Molecule Mechanism Of Magnolol And Honokiol

In the modern intensive livestock cultivation, Diarrhea is one of the main disease which hinders the rapid growth of the livestock and affects on their health condition. While adding antibiotics into the feed is the main measure taken to livestock’s diarrhea prevention, it is vulnerable to the problems of the drug-resistance and drug residual caused by pathogenic microorganism. Thereby, in the research of livestock and poultry nutrition and immunology, it is of significant direction to seek out the safe, efficient and no residue plant active ingredients as the substitution of antibiotics which may turn out to a significant research study. Because of the limitation on the approach and method of the research, it has not have made breakthroughs in the application of the prescribed or compound Chinese herbal medicine for the livestock’s diarrhea prevention. The magnolol and honokiol contained in the herbal medicine have pronounced effects on anti-diarrhea, yet the mechanism of action is not quite clear.In this paper, we constitute a enterotoxigenic Escherichia coli bacterial diarrhea model, measuring HE staining and apoptosis, anti-inflammatory, antioxidant, and calcium-related gene expression levels and other aspects to elaborate on the anti-diarrhea and its possible molecular mechanism of Magnolol and honokiol, which provides vigorous scientific evidence on exploiting new type green additive possessing the function of anti-diarrhea and the high-level utilization of the resources of Magnolia officinais. Experimental study results are as follows:1The way to make up Enterotoxigenic Escherichia coli bacterial diarrhea model and the screening of the doseIt builded an Enterotoxigenic Escherichia coli bacterial diarrhea model, basing on the experimental subjects of kunming male mice, via the studies of different injected way of four bacterias of O78:K80,O141:K99,O20:K101,O149:K88,different concentraitons of bacterias, Whether or not counteract gastric acid in the former of injecting bacteria, different times of injecting bacteria, Whether or not treat with antibiotics, Whether or not enhance toxicity of bacterias, tests of different ages of mice in pathogenicity, tests of different adaptation times of mouse in pathogenicity, tests of different fasting times of mice in pathogenicity. At the same time,on these basis, it sifted appropriate doses of magnolol and honokiol in the research of the anti-diarrheal function of magnolol and honokiol by the studies of different doses of magnolol and honokiol.The results showed that afeasible method of creating Enterotoxigenic Escherichia coli bacterial diarrhea model were like these choosing Kunming male mice whose weight were between25±2g and were fed under the SPF conditions, with room temperature between25±2℃, light12hours, libitum drinking asepsis water for three days at our institution. After that, we injected the mice of its lumber with E.coli E44813whose dosage and consistency are0.02mL·g-1,3.29X109CFU-mL"’ respectively before6hours fasting. Also it got three different dosages (high, middle and low)of Magnolol and Honokiol by500mg·(kg·bw)-1of high,300mg·(kg·bw)-1of middle and100mg·(kg·bw)-1of low of drug treatment group in researching the anti-diarrheal function of them.2The effects and mechanisms of anti-inflammatory on mouse intestines of Magnolol and honokiolThe article clarified the effects of anti-diarrhea and the possible anti-diarrhea molecular mechanisms of Magnolol and honokiol,on the basis of researches of intestinal anti-inflammatory,by using real-time quantitative PCR technologies via the studies of genes expression of inflammatory factors including TNF-a,INF-y,IL-6, IL-10,COX2,the genes expression of iNOS, nNOS,eNOS and the concentration of NO,TNOS,iNOS, the genes expression of oxidation resistance factors including ECSOD,MNSOD,CU/ZNSOD,GSHPX,CAT. The results showed that the KM mice which stimulated by enterotoxigenic performance in obvious diarrhea symptom with severe inflammation symptom, the concentration of NO in the diarrhea treatment group were significantly higher than the normal group (P<0.01), the activity of iNOS enzyme was more active and the gene expression of iNOS was increased hardly, with the activity of TNOS enzyme passive and the genes expression of eNOS, nNOS lower, the concentration of NO and the activity of iNOS enzyme in the drug treatment groups is lower,with the activity of TNOS increasing, the gene expression of iNOS was decreased hardly than the normal group, wihle the gene expression of eNOS,nNOS were increased slowly than the normal group.At the same time, drug treatment groups does not significantly affect on COX-2、DAO, but obviously on TNF-α、INF-r、IL-6、IL-10.So it illuminated magnolol and magnolol had cure role on Enterotoxigenic Escherichia coli bacterial diarrhea in a short period of time, after the anti-inflammatory functions.3Effects and mechanisms of restorations in diarrhea damage on mouse intestines by Magnolol and honokiol.The tests mainly researched the effects and mechanisms of restorations in diarrhea damage of mouse intestines by Magnolol and honokiol after the HE staining and apoptosis tests in mouse jejunum and ileum.The results showed, Magnolol and Honokiol can repair the damage of intestinal in mice, reduce intestinal villi loss, keep a complete villus epithelial cells, reduce inflammatory cells, promote hair elongation, increase the number of goblet cells, and deepen the depth of crypt. So it can prove that Magnolol and Honokiol has a main role of the therapeutic effect in the ileum and jejunum to the E.coli diarrhea, which were to relate to maintaining the integrity of the intestinal mucosal structure, protect mucous membranes against damaging and promoting the role of mucosal repair. It indicated that honokiol and magnolol can inhibit apoptosis on diarrhea, via the test results from apoptosis, which the brown apoptotic cells increased significantly in the diarrhea group by contrast in normal group and each drug treatment group of less.4The effects on genes expression of calcium ion channels in intestinal canal by Magnolol and honokiolUsing real time quantitative PCR technology, the article clarifys the anti-diarrheal effects and its possible anti-diarrheal molecular mechanism of magnolol and honokiol based on the research of genes expression of calcium ion channels in intestine. It mainly investigated the effects of genes expression of calcium ion channels’albumen which includes RyR1,IP3R1, IP3R2, and IP3R3,calmodulin which includes CaMI, CaMKII-α, and CaMKII-β, calcium-activated potassium Channels which includes SKI, SK2, SK3and SK4, as well as calcium-dependent potassium ion channels which includes ALP, Kcnma, Kcnmβ1, Kcnmβ2, Kcnmβ3and Kcnmβ4in intestine. The results showed that honokiol and magnolol had significant effects on genes expression of calcium ion channels’albumen, calmodulin,calcium-activated potassium Channels,and calcium-dependent potassium ion channels in intestine.Low concentrat-ion of magnolol and high concentration of honokiol have significant effects on increasing genes expression of calcium ion channels’albumen and calmodulin. While high concentration of magnolol and low concentration of honokiol have distinct inhibitory action from droping genes expression of calcium-activated potassium channels, with a trend that the higher concentration of drug in treatment group will has greater markedness. Meanwhile different dose of magnolol and honokiol have a distinct role in genes expression of calcium-dependent potassium ion hannels without dose-dependent.Magnolol and honokiol,as a Ca2+antagonist,can antagonize the Na+and Ca+to transport to the cell inside,also can antagonize with the calmodulin, all of that are to promote the protein activity of calcium ion channels’albumen and calmodulin with their genes expression increasd to inhibit secretory diarrhea.At the meantime magnolol and honokiol can effectively inhibit genes expression of calcium-activated potassium channels via controlling the activity of potassium ion channels and receptor manipulation to calcium channels.Honokiol and magnolol, as a paralysor of calcium channel, had bidirectional effects on the calcium-dependent potassium ion channels in interdicting the activity of voltage-dependent calcium channels. The two medicaments can lead genes expression of calcium-dependent potassium ion channels including Kcnma, Kcnmβ1, Kcnmβ2and Kcnm(33to decrease in varying degrees, while the genes expression of calcium-dependent potassium ion channels including Kcnmβ4and ALP were increasedThe effects of magnolol, honokiol, and interaction of magnolol with honokiol,on the relative calcium channels,are inconsistent, bidirectional, inhibitive and facilitated. Overall, magnolol and honokiol have effects on a series of physiological to regulate or improve diarrhea, which can regulate the concentration of Ca2+-dependent ion and the activation of opening and closing in activated-ion channel, activate the activation of the protein kinase, control genes expression.