Screening Of Compound Mongolian Medicine Treating Bovine Mastitis And Research Of Its Mechanism On Anti-inflammation And Immunoenhancement

Besides of the direct lose, such as the yield decease and milk quality degradation, treatment expense or even the cow culling, bovine mastitis also leads to indirect lose, such as inflammation after delivery, delayed empathema, ichoremia, brucellic disease, aftosa and diseases of such kind, because it depresses the immunity, by which bovine mastitis makes the largest lose in dairy cow raising industry. Antibiotics, the most popular treatment for mastitis, is losing its effectiveness, because its abusage which leads to the arising of the drug resistant strains and antibiotics residual. So a new and effective way to treat mastitis is needed to be explored. Traditional Mongolian medicine, one of the four most famous traditional Chinese ethnic medicines, includes peculiar medical theories and thousands of herbs with antiinflammatory effectiveness are recorded in its pharmacopeia which provides a possibility to explore a new method to treat bovine mastitis. This research is mainly to explore effective Monglolian compound medicines curing bovine mastitis and reveal their regulation action on inflammation and immunological responses.Four compound Mongolian medicines were developed through orthogonal design and invivo and invitro antibiosis experiment regarding to Staphylococcus aureus and Escherichia coli isolated from the milk of the mastitis infected cows. The other traditional Mongolian medicine was developed according purely to the traditional Monglian medicine theory. Given all the arguments of compound medicines'protection rates to the mixed bacteria infected mice, cost and the effects of the rumen microbes to the invitro antibiosis of the compound medicines, orthogonal designed compound Mongolian medicineⅠand the traditional compound Mongolian medicine were further studied on their regulation actions on inflammation and immunological reactions and clinical effectiveness.Pathological sections of the mice mammary glands infected by Staphylococcus aureus and Escherichia coli mixture showed that the two compound Mongolian medcines could effectively reduce the damage and promote recovery of the mammary glands. The experiment of the effect of compound Mongolian medicines on mice thymus and spleen index showed the two compound Mongolian medicines had no significant impact on normal mice but they could significantly elevate the thymus and spleen index of the pathogenic bacteria infected mice(P<0.05). The study of macrophage activity by carbon clearance method showed the two compound Mongolian medicines could significantly raise the activity of normal and model mice(P<0.05). The study of the red cell immunity by Guo method showed that the two compound Mongolian medicines could significantly raise the C3bRR rate(P<0.05), but they had no significant effect on ICR rate of the normal mice. But they could significantly raise the C3bRR rate and reduce the ICR rate of the pathogenic bacteria infected clinical mastitis model mice(P<0.01). The study of the activity of the NK cells in the spleen by MTT method showed that the two compound Mongolian medicines could significantly raise the activity of the spleen NK cells of both the normal and model mice(P<0.01). The study of the peripheral blood lymphocytes and their subgoup cells by the flow cytometry showed that the two compound Mongolian medicines had not significant effect on the percentage of CD3+ cell, CD4+ cell and CD19+ cell of the normal mice, but they could significantly raise the CD4+/CD8+(P<0.05). The two compound medcines could significantly raised the percentage of CD3+ cell, CD4+ cell and CD19+ cell and CD4+/CD8+ of the clinical mastitis model mice(P<0.01), but they had no significante effect on the percentage of CD8+ cell. The study of the enzyme and cytokines in clinical mastits model mice mammary gland showed that the two compound Mongolian medicines could significantly raise the IL-2 level and depress the elevation of IL-6 and TNF-α(P<0.01), and they could significantly reduce the NAGase concentration(P<0.01). The above study showed that the two compound Mongolian medicines could improve the nonspecific immunity by raising the macrophage activity; they could improve the specific immunity by raising the B cell rate, T cell rate and CD4+/CD8+. They could also effectively regulate the immunological reaction by adjusting the secretion content of IL-2, IL-6 and TNF-α.The study of the effect of two compound Mongolian medicines on capillary permeability rising induced by acetic acid in mice showed that they could effectively depress the permeability of capillar rasied by the acetic acid(P<0.01)and the traditional compound Mongolian medicine had a better result than the orthogonal compound Mongolian medicine. The study of the effect of the two compound Mongolian medicines on ear-swelling of mice induced by xylene and edema of rat hind paw induced by formaldehyde showed that the two compound Mongolian medicines could significantly depress the engorgement of the inflammation area(P<0.05)and they could also effectively depress the PGE2 secretion in the inflammation area(P<0.01). The study of the effects of the two compound Mongolian medicines on granuloma induced by planting cotton ball in mice showed that they could effectively reduce the amount of the granuloma(P<0.01). The study of the ICAM-1 and E-selectin content in the peripheral blood of the endotoxin syndroma mice induced by LPS injection through ELISA showed that they could significiantly depress the rising of ICAM-1 and E-selectin content(P<0.01), and the NO content (P<0.01)in the cell culture fluid of Human umbillical vein endothelial cell added with blood serum of LPS induced endotoxin syndroma mice was also significant reduced, which showed that the two Mongolian coumpound medicines could effectively reduce reduce the secretion of the PGE2, ICAM-1, E-selectin and NO content to regulate the inflammation reaction to minimize the damage to the tissue.The study of the nertraphilia cell phagocytic ability in the peripheral blood of subclinical mastitis cows through Staphylococci Phagocytosis showed that the two compound Mongolian medicines could significantly raised the nertraphilia cell phagocytic ability(P<0.01). The study of T cell rate and IgG content in the subclinical mastitis cow peripheral blood through flow cytometry and immuno-turbidity method respectively showed that the two Mongolian coumpound medicines could significantly raised theT cell rate(P<0.01)and IgG conten(tP<0.01)in the cow peripheral blood content, by which they could regulate the nonspecific and specific immunological reaction of the subclinical mastitis cows. So the somatics and the NAGase and LDH content in the milk of subclinical mastitis cows was reduced and the milk fat rate, protein rate, sugar rate and the total solid content were improved(P<0.01). The study of the clinical mastitis cows showed that the recovery rates of the traditional compound Mongolian medicine and the orthogonal designed compound Mongolian medicine was 84.62% and 71.43% respectively, and compared with that of the ciprofloxacin(85.71%), the result indicated that the two compound Mongolian medicines could effectively reduce the inflammation and recover the milk yield and quality.Along with the study, the invitro bacteriostasis in the development of antiinflammatory traditional ethnic medicine and the mutual reaction between the rumen microbes and traditional ethic medicines are primarily discussed. The invitro antibiosis experiments showed that the orthogonal compound Mongolian medicines had significant better antibiosis effectiveness than the traditional compound Mongolian medicine. While the invivo antibiosis experiment showed the total opposite results which indicate invivo antibiosis can not be used as an adequate condition for the development of anti- inflammatory traditional medicines. Five compound Mongolian medicines were fermented invitro by the rumen microbes, through the invitro antibiosis experiment, the products of which showed that the invitro antibiosis effectiveness of orthogonal compound Mongolian medicineⅡandⅢwere significantly degraded(P<0.05)by the rumen microbes which indicated that the mutual reaction of traditional medicines and the rumen microbes should be considered when the traditional compound Mongolian medicines were given orally to the ruminant.