Expression Of And Related Research P38MAPK And INF-γ In The Disseminated Candidiasis Mice Kidneys

Objective: p38MAPK was a sub-class of the mitogen-activated proteinkinase(MAPK)signal transduction pathway. Many factors, such as hormones,G-protein coupled receptors and the osmotic pressure could activate p38signaling pathway which played an important role in the physiological andpathological processes of inflammation, cell stress, apoptosis and the growth.Recent studies showed that the MAPK signal transduction pathway played animportant role in the regulation of innate and specific immune response.Interferon-γ (IFN-γ) was an immune cytokine, the level of IFN-γ levels mayreflect the immune status of the body cells. Candida albicans was the mostcommon nosocomial infection pathogens. Research showed that when theorganism infected Candida albicans, cellular immunity against Candidaalbicans infection was dominant. Recent studies had shown that, p38MAPKcould promote the expression of IFN-γ which has not been reported in thecandidiasis.In this study, we established mice model of disseminated Candidasis,and detected the expression of p38MAPK and IFN-γ in different time in micekidney by immunohistochemical staining methods, and explored the role ofp38MAPK and IFN-γ in host protective mechanism against disseminatedcandidiasis, so as to provide new methods for the clinical treatment ofdisseminated candidiasis.Methods:1Strains of Candida albicans saved from the Second Hospital of HebeiMedical University.2Established mice model of disseminated candidiasis：90Kunming mice,male and female, were purchased from the Animal Breeding Center of HebeiMedical University, mice were feed in the general environment in the Experimental Center of the Second Hospital of Hebei Medical University, andgived the full price of feed feeding. After adapting to the environment, themice were divided into A, B, and C group by weight randomly, n=30. Themice of group A was the experimental group, each mouse was injectedcyclophosphamide200mg/kg by ip form1^th to4^th day, in the5^th day, eachmouse was injected1ml suspension of Candida albicans for1～5×10⁶cfu/ml by ip. Group B was cyclophosphamide control group, each mousewas injected cyclophosphamide200mg/kg by ip form1^th to4^th day, in5^thday, by intraperitoneal injection of saline1ml; group C was blank controlgroup: each mouse was injected saline200mg/kg by ip form1^thto5^thday.At experiment6^th day,9^th day,20^th day, mice were broken neck to observeabdominal viscera in general situation.3Identification of the disseminated candidiasis infection of mice kidneys：thekidneys of mice were removed under sterile conditions, sterile normal salineseveral times, after the grinding of the side, inoculated in cultured onSabouraud medium.4Histopathological examination and immunohistochemical;another part of therenal tissue were fixed with10%formalin solution for routine pathological HEand PAS staining to be observed pathological changes and Candida albicansinfections in mice kidney tissue. We detect the expression of p38MAPK andIFN-γ by immunohistochemical methold with chemical Streptomycesantibiotic protein-peroxisome connection method (SP).Result:1Mice model of disseminated candidasis infection: during the experiment, atotal of10mice died, bacuse of multiple organ hemorrhage, edema byimmunosuppressant cyclophosphamide,owing to disseminated infection byCandida albicans, a total of11mice died,a total of15mice died byunexplained reasons.54mice were included in the trial. The mice of group Afrom the experiment4^thday, mice spiritual and appetite were poor, had dullhair, activities were not good. In experiment6^thday, the mice organ wereedema, kidney had a little white pus points. In experiment9^thday, by the anatomy of mice, kidney had white pus point generated in abdominal cavity.In20^thday, kidney had a lot of white pus point generation, part of the organwere congestion, necrosis. Group B mice from the experiment4^thday,spiritual and appetite were poor, dull hair, activities were not good. In theexperimental of6^thday, mice abdominal internal organs were edema andcongestive. In experiment9^thday, after dissection, kidney were part of theedema, hyperemia, necrosis. In experiment20^thday, by autopsy, the micekidney were enlarged, edema. The mice of group C spirit and appetite did notchange, hair gloss, activities normal. In the experimental sections6^thday,9^thday,20^thday, the mice abdominal organs showed normal performancerespectively.2Fungal culture and identification in mice kidney tissue of disseminatedCandida albicans infections: on the6^thday,9^thday,20^thday, the kidneystissue of group A mice were cultured in Sabouraud medium for72hours.There were white colonies growth which were identified as Candidaalbicans by serum culture method with chlamydospore and pseuohypha.3Histopathological examination: the mice of group A, in the experiment6^thday, glomerular and renal tubule have complete structure, no hyphae andspores were seen by PAS stain purple. In9^thday, the structure of glomerularand renal tubule were not complete, nuclear condensation, hyphae and sporeswere seen by PAS stain purple. In20^thday, parts of renal tissue were necrosis,,hyphae and spores were seen by PAS stain purple. Group B mice, inexperiment,6^thday,9^thday and20^thday, renal cell degeneration. In group Cmice showed normal performance.4The expression of p38MAPK at the protein level: the mouse kidney tissuewere stained by immunehistochemical staining methold. The expression ofp38MAPK mainly in the cytoplasmic, positive cells had cytoplasmstained brown,immunohistochemical staining results showed that there werevisible positive cells in each group. Group A: In experiment6^thday,9^thdayand20^thday, OD values were0.1727±0.0089,0.1799±0.0102,0.1638±0.0079respectively, the expression trend was increasing in early phage and decreasing in late phage, P <0.05, the difference was statistically significant.Group B: In experiment6^thday,9^thday,20^thday, OD values were0.1697±0.0074,0.1763±0.0149,0.1618±0.0043respectively, the expression trendwas increasing in early phage and decreasing in late phage, P>0.05, thedifference was not statistically significant. Group C: In experiment6^thday,9^thday,20^thday, OD values were0.1594±0.0033,0.1659±0.0056,0.1609±0.0097respectively, P>0.05, the difference was not statistically significant. Inexperiment6^thday, OD value of group A, B and C was gradualy decreasing, P<0.05, the difference was statistically significant. In experiment9^thday, ODvalues in group A, B and Cdecreased gradually, P>0.05, the difference wasnot statistically significant. In experiment20^thday, OD values in group A, Band C decreased gradually, P>0.05, the difference was not statisticallysignificant. Repeated measures design ANOVA: there was significantdifference between the groups at different time points, P <0.05the differencewas statistically significant. Further pairwise comparisons: there wassignificant difference between group A and group C, group B and group C, P<0.05.5The expression of IFN-γ at the protein level: the mouse kidney tissue werestained by immunehistochemical staining methold. The expression of IFN-γmainly in the cytoplasmic, positive cells had cytoplasm stained brown;immunohistochemical staining results showed that there were visible positivecells in each group. Group A: In experiment6^thday,9^thday and20^thday, ODvalues were0.1967±0.1236,0.2108±0.0183,0.1705±0.0171respectively,the expression trend was increasing in early phage and decreasing in latephage, P <0.05, the difference was statistically significant. Group B: Inexperiment6^thday,9^thday,20^thday, OD values were0.1725±0.0179,0.1843±0.0226,0.1650±0.0090respectively, the expression trend was increasing inearly phage and decreasing in late phage, P>0.05, the difference was notstatistically significant. Group C: In experiment6^thday,9^thday,20^thday, ODvalues were0.1588±0.0080,0.1841±0.0132,0.1629±0.0080respectively,P <0.05, the difference was not statistically significant. In experiment,6^thday, OD value of group A, B and C was gradually decreasing, P <0.05, thedifference was statistically significant. In experiment9^thday, OD values ingroup A, B and C decreased gradually, P>0.05, the difference was notstatistically significant. In experiment20^thday, OD values in group A, B andC decreased gradually, P>0.05, the difference was not statistically significant.Repeated measures design ANOVA: there was significant difference betweenthe groups at different time points, P <0.05the difference was statisticallysignificant. Further pairwise comparisons: there was significantdifference between group A and group C, group A and group B, P <0.05.6the expression of p38MAPK and IFN-γ in experimental group in theexperiment6^thday,9^thday,20^thday had no linear correlation, r=0.00,0.1761,0.2408, respectively, P>0.05, the difference was not statistically significant.Conclusion:1Expression of p38MAPK in the disseminated candidiasis infected mousekidney rised in early stages of infection, downward trend followed thetime.2Expression of IFN-γ in the disseminated candidiasis infected mouse kidneywas rising in early stages of infection, in the late stages appeared downwardtrend.3The downward trend in late stage of infection may be one of the reasons forCandida albicans invasive infection in kidney in expression of p38MAPK andIFN-γ.4The expression of IFN-γ and p38MAPK was not linear relation indisseminated candidiasis mouse kidney. The expression of IFN-γ may not beactivated through P38MAPK signaling pathway.