结肠炎相关结直肠癌小鼠模型的中医证候动态探究OA

Objective To establish a dynamic evaluation system for traditional Chinese medicine(TCM)syndromes in a mouse model of colitis-associated colorectal cancer(CAC),and to explore the immunological mechanism underlying the"inflammation-cancer transformation"process,thereby providing a reference for integrated disease-syndrome research.Methods A CAC model was induced in 32 SPF C57BL/6J Nifdc mice using Azoxymethane/Dextran Sulfate Sodium(DSS).The mice were divided into control(n=8)and model groups(n=24).Based on the DSS cycle,three dynamic time windows were established at weeks 4,7,and 10 post-modeling.The model group was further subdivided into three subgroups(n=8 each):the cycle 1,2,and 3 model groups.Macroscopic indicators,including body weight,food and water intake,fecal water content,and Disease Activity Index(DAI),were monitored.Infrared thermography(core/surface temperature)and tongue image red-green-blue parameters were analyzed to assess dynamic syndrome changes.Colon tumor burden,tumor multiplicity,and colon length were recorded.Histopathological examination was performed to evaluate the degree of inflammation and dysplasia.Immunofluorescence was used to quantify M1/M2 macrophage surface markers CD86 and CD206.Enzyme-linked immunosorbent assay was employed to measure M1-associated cytokines(IL-12,TNF-α,and CXCL9)and M2-associated cytokines(IL-10,VEGF,and CCL17)to investigate the immunological mechanisms of"inflammation-cancer transformation.Results The cycle 1 model group was characterized by spleen-deficiency and phlegm-dampness syndrome.Mice exhibited decreased food and water intake,lethargy,huddling,and other spleen deficiency symptoms.Fecal water content was significantly elevated(P＜0.01),whereas tumor load and number were low.Colonic tissue showed mild structural abnormalities with partial inflammatory infiltration.Compared to the control group,the cycle 1 model group showed significantly increased positive rates of CD86 and CD206,as well as elevated levels of M1 and M2-type cytokines(P＜0.05 or P＜0.01).The cycle 2 model group was characterized by dampness-heat in the spleen syndrome.Mice showed peak food and water intake,alongside heat signs like mental excitement,irritability,hyperactivity,yellow urine,and hematochezia.Fecal water content decreased significantly compared to that of the cycle 1 model group(P＜0.01).The core body temperature,surface temperature,and tongue color red-value significantly increased,reaching their peaks(P＜0.05 or P＜0.01),indicating intense heat-toxicity.Colonic tissue exhibited moderate abnormalities with inflammatory cell infiltration and a few tubular adenomas.Compared to the cycle 1 model group,the cycle 2 model group demonstrated a significantly increased CD86 positive rate and elevated M1-type cytokines(P＜0.05 or P＜0.01),but no significant difference in the CD206 positive rate or M2-type cytokine levels.The cycle 3 model group was classified as damp-heat stasis toxin syndrome.Mice appeared emaciated and listless,with the lowest body weight and food and water intake.The mice exhibited dull fur,easy hair loss,and purulent bloody stools.Fecal water content peaked and was significantly higher than that of all other groups(P＜0.05 or P＜0.01).Concurrently,tumor burden,multiplicity,and DAI scores increased and were significantly higher than those in the cycle 1 and 2 model groups(P＜0.05 or P＜0.01).This was accompanied by a decline in core and surface body temperature,and an increase in tongue color green and blue values,presenting a cyanotic tongue characteristic(P＜0.05 or P＜0.01),suggesting the pathogenesis of prolonged heat transforming into stasis,with phlegm and heat combining to form stasis-toxin.Colonic tissue showed severe abnormalities with intense inflammatory cell infiltration and numerous tubular adenomas.Furthermore,compared to the cycle 1 and 2 model groups,the cycle 3 model group showed a significant increase in the CD206 positive rate and M2-type cytokine levels(P＜0.01).Conclusion The TCM syndromes in CAC follow a dynamic evolutionary pattern:"spleen deficiency and phlegm-dampness → dampness-heat in the spleen meridian → damp-heat stasis toxin."This syndromic evolution was synchronized with an imbalance in M1/M2 macrophages.The spleen deficiency and phlegm-dampness stage may correspond to an initial state of impaired immune surveillance;the dampness-heat stage was associated with enhanced inflammatory response dominated by M1 polarization;and the damp-heat stasis toxin stage was associated with an immunosuppressive microenvironment mediated by M2 polarization.These findings provide a novel theoretical model for elucidating the biological basis of"disease-syndrome"relationships in TCM,warranting further validation.