Analysis Of The Difference Of T Cell And Fibroblast Expression In The Development Of Colorectal Carcinoma With Different Syndromes Of Cold And Heat

Colorectal Cancer(CRC)is the third leading cancer diagnosed worldwide and the second leading cause of cancer-related death.The main reasons for the high mortality rate are the late diagnosis and the high recurrence rate after treatment,with 70% of patients being diagnosed only when the tumor has already metastasized to distant organs,it is usually associated with low penetrance susceptibility genes and specific polymorphisms that are regulated by environmental or other genetic factors,and co-inheritance increases the risk of cancer.However,in the past cancer research,medical scientists always focused on the tumor cells themselves,but neglected the role of tumor microenvironment in the generation,proliferation and metastasis of tumor cells,and the decisive role in the tolerance to various therapies.Therefore,the theory of tumor microenvironment is a turning point for the change of the direction of modern medicine in the understanding of tumor,and a study of the interaction between tumor and its host,the remarkable feature of TCM treatment based on syndrome differentiation is to intervene and regulate the microenvironment of tumor tissue by multi-target effect.The concept of "Syndrome" in traditional Chinese medicine involves many dimensions of western medicine.With the development of complexity science,the "Uncertainty principle" in the organism has been revealed.The state description of human body as a whole can only adopt the mode of traditional Chinese medicine,starting with the investigation of human behavior function on the whole level.The introduction of modern medical testing methods and data analysis technology,based on traditional Chinese medicine,to establish a holistic level of precision medicine system,not only feasible and it is bound to be the best way for precision medicine to achieve its long-term goals.The core aim of TCM tumor treatment based on syndrome differentiation is to improve microenvironment through intervention.Among them,CRC tumors can manipulate the tumor microenvironment(TME),promote cancer growth by modulating immune infiltration,and generate an immunosuppressive and inflammatory environment.In this way,the TME plays a key role in regulating the plasticity of tumor cells and providing mechanisms for immune escape.“Cold and heat” is an important component of“Syndrome differentiation of the eight cardinal principles”.Its content mainly depends on the nature of pathogenic factors and the rise and fall of yin and yang in the body.It plays an important role in the clinical diagnosis and treatment of traditional Chinese medicine,it provides important reference for clinical treatment.This study is about the difference of the development pattern of colorectal cancer in rats with different syndromes of cold and heat,and the further study of the mechanism from the point of view of T cells and fibroblasts.ObjectiveThe purpose of this study was to explore the influence of TCM syndromes on the difference of occurrence and development patterns of colorectal cancer from the perspective of T cell and fibroblast markers,and to provide a new basis for the immune stratification diagnosis and treatment of cancer.MethodsThe rats were randomly divided into four groups.Normal group: rats were given normal diet and pure water(10 ml/kg).The Rats in DMH group were subcutaneously injected with 35,30,25,20 mg/kg DMH from the sixth week,once a week for 12 weeks.In cold model group(DMH + ice water),the rats were given DMH and 0°C ice water(10ml/kg)three times a day until the end of the experiment.Hot model group(DMH +capsaicin): DMH was given as the model group,and 20% ethanol(10 ml/kg)was given once a day for the first week,in the second week,30% ethanol solution(9 mg/kg,10ml/kg)of capsaicin was given once a day until the end of the experiment.In this study,general signs,food intake,body weight,colon appearance length and energy metabolism enzymes were used to judge whether the model was established successfully.then did hematoxylin and eosin staining on sections of colon tissue to measure histopathology levels.FAP,FSP-1,PDGFR β,PD-1,PD-L1,HMGB1,Tim-3,CD4 and CD8 were detected by immunohistochemistry and western blotting.Results1.Establishment and evaluation of rat model of colorectal carcinoma with different syndromes of cold and heatAccording to the standard of cold-heat model,the general signs,food intake,body weight,appearance length of colon and energy metabolism enzymes of rats were evaluated.He staining showed that the colon was divided into four stages: dysplasia,colorectal adenoma,colorectal intramucosal carcinoma and colorectal invasive carcinoma.According to the results,the malignant degree of the cold model in the later stage of cancer was higher,and the morphological observation and length detection showed that the cold model was the shortest,which indicated that the fibrosis was the most serious.2.Expression of immune cells in colorectal carcinoma with different syndromes of cold and heatThe protein levels of PD-1,PD-L1,HMGB1,Tim-3,CD4 and CD8 were detected by IHC and WB,the results showed that the protein levels of PD-1,PD-L1,HMGB1 and Tim-3 in cold group and heat group were higher than those in normal group and model group at 17 and 24 weeks,and the protein levels of PD-1,PD-L1,HMGB1 and Tim-3 in cold group were higher than those in heat group,there was statistical significance(P <0.05).The expression levels of CD4 and CD8 in the cold and heat model groups were significantly lower than those in the normal group and the model group,and the expression levels of CD4 and CD8 in the cold model group were significantly lower than those in the heat model group(P < 0.05).In addition,PD-1,PD-L1,HMGB1,Tim-3,CD4 and CD8 protein levels were not significantly different among the four groups at week 5.3.Expression of fibroblast markers in colorectal carcinoma with different syndromes of cold and heatThe expression levels of FAP,FSP-1 and PDGFR β proteins were detected by IHC and WB.The results showed that the expression levels of FAP,FSP-1 and PDGFR βproteins in the cold and hot groups were higher than those in the normal group and the model group at the 17 th and 24 th week,the expression levels of FAP,FSP-1 and PDGFRβ in cold group were higher than those in hot group(P < 0.05).In addition,there was no significant difference in FAP,FSP-1 and PDGFR β protein expression between cold and hot model at the 5th week.ConclusionThis study demonstrated the influence of different syndromes of cold and heat on the occurrence and development of tumor immune microenvironment in CRC rats by establishing rat model,by means of immunohistochemistry and WB experiments,we compared the molecular differences of the expression of the immune cells and fibroblasts in different syndromes of cold and heat,and concluded that the immunogenicity of tumor in cold syndrome was the worst compared with that in normal and heat syndrome rats,the malignant degree was the highest,and the ability of migration,invasion and proliferation of tumor cells was increased.