The Research Of B Cell Subsets And Humoral Immune Response In BALB/c Mice Infected With Mycobacterium Tuberculosis

ObjectivesTuberculosis is still an important infectious diseases so far, and it can invadetissues and organs, in which pulmonary tuberculosis is most prevalent. The role ofhumoral immunity resisting Mycobacterium tuberculosis(M. Tuberculosis, Mtb) wasminimal in a large number of literature. However, since the late19th century, therewere reports that antibodies played a role in M. tuberculosis infection. This suggeststhat humoral immunity plays a role in M. tuberculosis infection, but the role is notclear, and it need to be further explore. This subject will analyze the immune status ofthe pathological changes in different tissues, and clarify the B cell responses to theimpact and role of M. tuberculosis infection by measuring serum antibodies in miceand B cell subsets surface molecule. At the same time, we further explore the role ofmeasured progress which the immune response to the anti-tuberculosis infection andtuberculosis.MethodsTaking BALB/c female mice about5-6weeks old age random divided DaliM. tuberculosis clinical isolates infected groups, H37Ra infected groups and normalsaline(NS) groups, and injecting intravenously106CFU per mouse. we established the mouse model infected M. tuberculosis (NS groups were injected with the sameamount of NS). when mice infected4weeks or8weeks, they were sacrificed. Thedetermination of lungs, livers and kidneys was used with pathological changescomparative analysis and bacterial analysis; we determined serum antibody types andlevels of the different stages infection of IgG, IgM and IgA of mice taken by ELISA;peritoneal cells were used to analyze differences of the early stage and late stage ofthe eluting B1cell subsets; spleen cells were used to analyze subsets of B cells(B1acells:.B220IntIgD-CD5+CD11+; B1b cells:B220IntIgD-CD5-CD11+; FO B cells:B220+CD23+CD21int; MZ B cells:B220+CD23-CD21hi).Results1. Pathological changes and tissue culture of the main organs: the media haveseen varying amounts of M. tuberculosis growth, and we can determine thesuccessful establishment of M. tuberculosis infection in mice models. What ismore, throughout the course of infection were cultured M. tuberculosis. The mainorgans of infected mice were weighed at late stage, and suggesting that the impactof isolates infection on the liver is greater than the NS group (P=0.000<0.05) andH37Ra infected group (P=0.004<0.05). The impact of isolates infection onpulmonary is greater than the NS group (P=0.000<0.05) and H37Ra infectedgroup (P=0.014<0.05). The impact of H37Ra infected group on lungs is greateraffecting than the NS group (P=0.000<0.05). Effects on the kidneys of H37Rainfected group (P=0.000<0.05) and isolate the infected group (P=0.000<0.05)were higher than the NS group. The impact of the spleen of isolates infectedgroup was higher than H37Ra infected group (P=0.022<0.05) and NS group(P=0.004<0.05). The spleen, kidney and liver of infected mice of H37Ra andMtb clinical isolates were enlargement significantly higher than the saline groupby visually observed, and red in color, and clinical isolates of Mtb infection wasenlargement significantly higher than H37Ra infection. We observed mice tissuesof lungs, livers, kidneys tissue by HE staining and found that three kinds oforganizations have different levels of congestion and edema, and the lung tissues were the most obvious with inflammatory cell infiltration, and the last werekidneys, three kinds of tissues were not typical pathological changes oftuberculosis, which consistent with the literature.2. Antibody levels were measured at different stages of infection:(1) IgM: IgMconcentrations in early stages of infection in the NS group was significantly lowerthan H37Ra infection group (P=0.021<0.05); IgM concentration in late stages ofinfection in the NS group was significantly lower than isolates infected group(P=0.019<0.05); comparing different stages of infection: non-infected group wasless than8W isolate infected group (P=0.022<0.05) and H37Ra infection4Wgroup (P=0.021<0.05).(2) IgG: IgG concentration in the NS group in lateinfection was less than isolates infection group(P=0.020<0.05), H37Rainfection group was less than isolate infected group (P=0.029<0.05); comparingdifferent stages of infection: uninfected group was less than8W isolates infectedgroup (P=0.023<0.05),4W infection group was less than8W infected group(P=0.044<0.05).(3) IgA: IgA concentrations in the saline group was less thanH37Ra infection group (P=0.03<0.05) and isolates infected group (P=0.013<0.05), while H37Ra infection group higher than isolates infected group(P=0.047<0.05); comparing different stages of infection: non-infected groupwas less than8W isolate infected group (P=0.013<0.05) and H37Ra infection8W group (0.031<0.05), isolates infected group at4W was higher than8Winfected group (P=0.015<0.05), H37Ra infection group at4W was higher than8W infected group (P=0.015<0.05).3. B cells and their subsets: The mice of H37Ra infected and Mtb clinicalisolates were splenic B cells (CD45R+) decreased, while lymph node B cells(CD19+) increased both early and late infection. The results of B1a cells(B220IntIgD-CD5+CD11+) ratios are inconsistent between H37Ra infected andMtb clinical isolates; while B1b cells (B220IntIgD-CD5-CD11+) were lower thanthe control group in the early and late stages of infection. FO B cells(B220+CD23+CD21int) were higher than the control group, MZ B cells (B220+CD23-CD21hi) were lower than the control group.Conclusion1This study successfully established a BALB/c mouse model of tuberculosisinfection. Until the late stage of infection, we can culture and isolate Mtb fromorgans. After tuberculosis infection in mice, although Mtb can causeinflammation of the major organs, but not cause pathological changes similar tohuman tuberculosis.2In this study, through measuring antibody at levels the different stages ofinfection showed, IgM increased only on early stages in tuberculous infection, butIgG, IgM and IgA levels were increased at late stages of infection, and increasedwith extended duration.3This study, through detection of subsets change of B cells,showed that TBinfection affected the distribution of the major subsets of mature B cells. B cellsin the peripheral immune organs reallocated in Mtb infection. The total B cellsreduced in the spleen, while elevated in lymph nodes; FO B cells increased, andMZ B cells were reduced; within the experimental time, B1b cells reduced, B1acells increased in the early stages of infection.