The Effect Of Polysaccharide Of Sijunzi Decoction On Immune Functions Improving By Gut Microbiota In Spleen Deficiency Mice

Objective:Sijunzi Decoction becomes a classic party for the treatment of spleen deficiency disease after repeated clinical validations. And polysaccharides are one of the most abundant and major effective components in Sijunzi Decoction bases on the modern medical methods researches. Polysaccharide of Sijunzi Decoction can affect the immune system by activate the gut local immune system.. However, the mechanism of modulate immune was not clear due to the restriction of polysaccharide itself. The human gut microbial communities could utilize the polysaccharide and were adaptive to change its structure and function to participate in maintaining the normal physiological states of human body. Therefore, the main purpose of this study was to replicate spleen deficiency model with different methods to select a suitable way for this study, and then to observe the immune function and intestinal flora in mice after the influence of polysaccharide of Sijunzi Decoction to investigate whether the polysaccharide of Sijunzi Decoction could improve the immune function in mice with spleen deficiency by regulating intestinal flora or not.Methods:1. Sulfuric acid-phenol method was used to measure polysaccharide of Sijunzi Decoction content.2. Senna water decoction, Senna water decoction with hunger disorder together and Senna water decoction with antibiotic by intragastric administrationa were tried to build spleen deficiency models; flowing cytometry was used to detect the proportion of peripheral blood leukocytes, thymus index, the number of Peyer’s patches were measeured and the proliferation ability of lymphocyte in mesenteric lymph nodes and spleen was measured by MTT, then comparing the changes of these immune functions in mice with spleen deficiency by different methods to determine a suitable method for the study.3. Traditional colony counting and fluorescence quantitative PCR method were used to detect the changes of intestinal flora to observe whether the intestinal flora disturbance or not in mice with spleen deficiency.4. Detecting the proportion of peripheral blood leukocytes the proportion of T and B lymphocytes in mesenteric lymph nodes by flow cytometry; thymus index, the number of Peyer’s patches and intestinal slgA content by ELISA and the proliferation ability of lymphocyte in mesenteric lymph nodes and spleen by MTT, and the change of gut bacteria such as bifidobacterium, lactobacillus, E.coli and bacteroides fragilis was measured by colony counting and fluorescence quantitative PCR in spleen deficiency mice after treated with polysaccharide of Sijunzi Decoction.5. Observing the effect of polysaccharide of Sijunzi Decoction on gut bacteria by culture in vitro.Results:1. The content of polysaccharide of Sijunzi Decoction was72.5%.2. All of three methods that used to make spleen deficience model were working, the thymus index, the number of Peyer’s patches, the ability of lymphocytes proliferation in spleen and mesenteric lymph node were significantly decreased in mice with spleen deficiency by intragastric administration with Senna water decoction and hunger disorder (P<0.05), and the mice recovered most slowly.3. The number of intestinal bifidobacterium, lactobacillus, escherichia coli, bacteroides fragilis were rising compared with the normal mice detecting by the two methods and the value of B/E by the colony counting was reduced.4. After treated with polysaccharide of Sijunzi decoction, spleen deficiency condition of the mice was recovered, and the thymus index, the number of Peyer’s patches and level of intestinal sIgA were increased (P<0.05), the proliferation ability of lymphocyte in mesenteric lymph nodes and spleen is upregulate, the ratio of peripheral blood leukocyte and T/B lymphocyte in MLN recovery to normal, the number of intestinal flora reduced, closed to normal level.5. Polysaccharide of Sijunzi decoction could promote the development of gut microbiota, and for probiotics, the influence was more significant.Conclusion: 1. Senna water decoction by intragastric administration with hunger disorder was the best method of spleen deficiency mice model2. Polysaccharide of Sijunzi decoction has regulatory effect to immune function and gut microbiota of mice with spleen deficiency.