| ObjectiveColorectal cancer(CRC)is a common malignant tumor of the digestive tract in clinical practice,and its incidence is increasing year by year.In clinical treatment of CRC,surgical operation is the main treatment,supplemented by chemotherapy,radiotherapy and traditional Chinese medicine treatment.And the operation is the most extensive and the curative effect is more accurate.In recent years,it.has been found that CRC surgery will further destroy the patient's intestinal microecology,and its structure and abundance will change significantly,leading to a series of complications that affect the patient's quality of life?condition restore and prognosis.Clinically,it is often combined with traditional Chinese medicine to treat patients after CRC,which can reduce the relevant clinical symptoms of patients after surgery.Moreover,the study found that TCM treatment can regulate the structure and abundance of intestinal flora,and intestinal flora can also act on the metabolism of Chinese medicine.After clinical observation and literature research,it is found that many patients with CRC belong to the syndrome of yang deficiency of the spleen and kidney.Traditional Chinese medicine by warming the kidney and invigorating the spleen has a prominent clinical effect.To this end,this study intends to investigate the changes in related clinical symptoms of patients with spleen and kidney yang deficiency syndrome after CRC intervention by traditional Chinese medicine,and to use the second-generation high-throughput sequencing technology to taxonomically analyze the gut microbial structure of patients before and after the intervention,and compared with the gut microbial structure of normal healthy people in order to find the changes in the intestinal microecology of patients with spleen and kidney yang deficiency after CRC,while improving the relevant symptoms of traditional Chinese medicine.The intestinal microecology of the patient further shows that the efficacy of TCM intervention is related to the intestinal microecology.And try to find the beneficial bacteria,harmful bacteria and marker bacteria related to CRC treatment in the intestinal flora,so as to provide new ideas and new methods for the research of anti-tumor of Chinese medicine.MethodsIn this study,a total of 40 cases of patients with spleen-kidney Yang deficiency syndrome after CRC who were treated in the patient department and inpatient department of Dongfang Hospital of Beijing University of Chinese Medicine from January 2018 to December 2020 were included in the prospective control trial.Traditional Chinese medicine(TCM)was used for continuous intervention for 2 weeks,and the total score of related symptoms before and after TCM intervention was evaluated by TCM symptom score method.Compare the changes of TCM symptoms before and after TCM intervention in patients.40 patients before and after medication feces samples and 10 normal healthy person's stool specimens(as a baseline study),and then extract the stool samples of microbial total DNA target fragment PCR amplification,build Miseq library,Miseq computer sequencing.Splicing PE reads obtained by MiSeqPE 300 sequencing,according to overlap relationship.Filtering and quality control for sequence quality at the same time,to distinguish the samples after cluster and the analysis of species taxonomy.Intestinal microbial information of healthy volunteers,patients with spleen-kidney-yang deficiency syndrome after CRC and patients with spleen-kidney-yang deficiency syndrome after CRC after TCM intervention were detected respectively,and then a series of in-depth statistical and visual analysis of community structure were performed.For example,Alpha diversity analysis,Beta diversity analysis,species composition analysis at various classification levels,multivariate statistical analysis and correlation analysis were used to compare the changes in intestinal flora structure of each sample,so as to explain the special changes in intestinal flora of patients with spleen-kidney-yang deficiency syndrome after CRC operation,and prove the efficacy of TCM in the intervention of patients with spleen-kidney-yang deficiency syndrome after colorectal cancer operation and the relationship between intestinal flora and TCM.And try to find the beneficial bacteria,harmful bacteria and marker bacteria in the intestinal flora related to CRC treatment.Results1.Forty patients with spleen and kidney yang deficiency syndrome after CRC surgery,2 cases were cured,16 cases were effective,22 cases were effective,and 0 cases were ineffective.The rate of Chinese Medicine symptom relief is 100.The total scores of Chinese medicine symptoms before,1 week after intervention and 2 weeks after intervention were 14.3±1.5,9.6±1.2,5.8±0.9.Three time points are compared in pairs,All the differences were statistically significant(P<0.05).Through the intervention,All patients did not have adverse drug reactions.This means that after the intervention of Chinese medicine,the related symptoms of the spleen and kidney yang deficiency syndrome after CRC surgery were obviously relieved.2.The results of classification and identification showed that the structure of intestinal microecology in patients after CRC surgery with spleen and kidney yang deficiency syndrome before and after Chinese medicine intervention was similar,but there were some differences.Chinese medicine intervention relieved the patients' related symptoms and increased the diversity of intestinal microbes,which were richer than before intervention.3.The results of Alpha diversity analysis showed that the richness and diversity of intestinal microbial community in normal healthy people were higher than those in patients with spleen and kidney yang deficiency syndrome after CRC surgery.After Chinese medicine intervention,the richness and diversity of intestinal microbial community in patients with spleen and kidney yang deficiency syndrome after CRC surgery increased to a certain extent,making it close to normal healthy people.4.The results of Beta diversity analysis showed that intestinal flora of patients with spleen and kidney yang deficiency after CRC surgery was significantly different from normal healthy people and in a state of imbalance,while the microecology of intestinal flora was improved and the difference of intestinal flora was smaller after Chinese medicine intervention.This means that after Chinese medicine intervention,patients after CRC surgery can improve their intestinal microenvironment..5.The composition and relative abundance of intestinal flora in patients with spleen and kidney Yang deficiency syndrome after CRC surgery were not significantly(P<0.05)reduced compared with those in normal healthy subjects.Species of p_Proteobacteria?c Gammaproteobacteria?c Bacilli?o Enterobacteriales?o Lactobacillales?f_enterobacteriaceae,especially g_Klebsiella?g_Escherichia and g_Lachnoclostridiumwere significantly higher than those of normal healthy subjects(P<0.05).Compared with the normal healthy population,both before and after Chinese medicine intervention,the differences of c_Gammaproteobacteria?o_Enterobacteriales?f_Enterobacteriaceae and g_Escherichia in the intestinal flora of CRC patients were statistically significant.It can be seen that the imbalance of intestinal flora in patients with spleen-kidney Yang deficiency after CRC surgery is mainly caused by the imbalance of symbiotic bacteria and pathogenic bacteria,abnormal increase of pathogenic bacteria.This may be related to the symptoms of spleen-kidney Yang deficiency patients after CRC surgery.6.The structure of intestinal flora of patients with spleen-kidney Yang deficiency syndrome after CRC surgery changed at the level of phylum,class,order,family and genus before and after TCM intervention.After TCM intervention,the composition and relative abundance of c_gammaproteobacteria?c_Bacilli?o Enterobacterales?o_Lactobacillales?f enterobacteriaceae?f Enterococcaceae,especially g_Lachnoclostridium?g_Escherichia?g_Klebsiella and g_Enterococcusin of patients with spleen-kidney Yang deficiency syndrome after CRC surgery were significantly decreased(P<0.05).Species of f_Prevotellaceae,especially g_Faecalibacterium and g_Rombotusia were significantly higher than those of normal healthy subjects(P<0.05).Therefore,Chinese medicine intervention can inhibit the growth of harmful bacteria in intestinal microorganism of patients with spleen-kidney Yang deficiency syndrome after CRC surgery,promote the growth of beneficial bacteria.This may be related to the improvement of the symptoms of patients with spleen-kidney Yang deficiency syndrome after CRC surgery by TCM.7.In this study,through composition and abundance detection,it was found that some specific bacteria existed in different groups.These species can be used as indicators to monitor disease activity and treatment responsiveness at different stages of treatment.The intestinal flora markers of patients with spleen-kidney Yang deficiency syndrome after CRC surgery were o Lactobacillales and g_Klebsiella;the marker bacteria of patients after TCM intervention were o Lactobacillales?g_Klebsiella;the marker bacteria of the normal healthy group were g_Megamonas?g_Faecalibacterium?g_Subdoligranulum?g_Roseburia?g_Alistipes?g_Sutterella.8.Within the genus level of the top 20 absolute abundences of all samples of human intestinal microbes,this study found that there was no antagonistic bacterial community,while there was synergistic interaction between g_Prevotella 9 and g_Megamonas.ConclusionThis study confirmed the CRC spleen kidney Yang deficiency syndrome in patients with postoperative intestinal flora structure compared with normal healthy people was in a state of disorder,the main problem of CRC spleen kidney Yang deficiency syndrome in patients with postoperative intestinal dysbacteriosis is the imbalance of symbiotic bacteria and pathogenic bacteria,and the abnormal increase of pathogenic bacteria,in this may associated with CRC spleen kidney Yang deficiency patients postoperative symptoms have a certain correlation.TCM intervention can improve the clinical symptoms of postoperative patients.Meanwhile,it was also found that Chinese medicine intervention could inhibit the growth of harmful bacteria and promote the growth of beneficial bacteria,which mainly increased the relative abundance of f Prevotellaceae.However,the relative richness of f Enterobacteriaceae and f_Enterococcaceae decreased,which improved the intestinal microecology of patients with spleen-kidney Yang deficiency syndrome after CRC,and increased the richness and diversity of intestinal microbial community.It is helpful to restore the intestinal flora structure of patients after CRC operation to a normal and healthy state,which may be related to the improvement of related symptoms of patients with spleen-kidney Yang deficiency syndrome after CRC by TCM intervention.In addition,the intestinal flora of patients with spleen and kidney Yang deficiency syndrome after CRC surgery and normal healthy subjects before and after TCM intervention all had their own specific marker bacteria.This suggests that the mechanism of TCM intervention in improving the clinical symptoms of patients with spleen-kidney Yang deficiency syndrome after CRC may be related to the improvement of intestinal microecological environment of patients. |
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