Objective:To explore the relationship between microsatellite instability (MSI) and Ki-67 expression level and the clinicopathological features of colorectal cancer, and investigate their impact for prognosis, so as to provide reference for prognostic judgment of colorectal cancer.Methods:The data of 183 patients who underwent radical colorectal cancer surgery and were diagnosed pathologically in the Department of General Surgery, the First Affiliated Hospital of Xinxiang Medical University from January 2017 to December 2019 were retrospectively analysed, including 101 males (55.2%)and 82 females(44.8%), ranged from 20 to 86 years and the mean age was(60.27±13.13)years. According to the results of mismatch repair protein immunohistochemical staining, the patients were divided into MSI-H group ( n=32) and MSI-L/MSS group ( n=151). According to the results of Ki-67 antigen immunohistochemical staining, the patients were divided into low Ki-67 expression group (<82.5%, n=136) and high Ki-67 expression group (≥82.5%, n=47) , among which 62 cases (78.5%) with low Ki-67 expression and 17 cases (21.5%) with high Ki-67 expression were in patients with Ⅲ+ Ⅳ stage colorectal cancer. The data of clinicopathological features, disease-free survival, and overall survival were collected and analyzed. The cotegorical variables were presented as n(%), and the comparisons between groups were performed using Chi-square test or Fisher′s exact test. The multivariate Logistic regression model was used to estimate the correlation between microsatellite instability and Ki-67 expression level and clinicopathologic characteristics of colorectal cancer. Kaplan-Meier survival curve and COX proportional hazards regression model were used to analyze the correlation between microsatellite instability and Ki-67 expression level and disease-free survival and overall survival. Results:Single factor analysis showed that the differences in gender ( χ2=4.37, P=0.037), tumor site ( χ2=26.40, P<0.001), tumor maximum diameter ( χ2=11.12, P=0.001) and nerve invasion ( χ2=5.53, P=0.019) between MSI-H group and MSI-L/MSS group were statistically significant. Multivariate Logistic regression model analysis showed that only gender ( OR=3.013, 95% CI: 1.183-7.672, P=0.021), tumor location ( OR=0.167, 95% CI: 0.067-0.419, P<0.001) and nerve invasion ( OR=0.202, 95% CI: 0.042-0.968, P=0.045) were independently correlative factors for MSI status. In Ⅲ+ Ⅳ stage colorectal cancers, the difference in tumor site between low Ki-67 expression group and high Ki-67 expression group was statistically significant( χ2=3.91, P=0.048). Multivariate Cox proportional hazards regression model analysis revealed that high Ki-67 expression ( HR=0.301, 95% CI: 0.118-0.768, P=0.012; HR=0.275, 95% CI: 0.083-0.912, P=0.035) and MSI-H ( HR=0.072, 95% CI: 0.010-0.525, P=0.009; HR=0.122, 95% CI: 0.017-0.900, P=0.039) were independently protective factors for disease-free survival and overall survival. Conclusions:MSI-H colorectal cancer is common in males, right-sided colonic cancers and non-neuroinvasive patients. In stage Ⅲ+ Ⅳ colorectal cancer, the expression level of Ki-67 in right-sided colonic cancer was lower than in left-sided colorectal cancer. Patients with MSI-H and high Ki-67 expressive colorectal cancer had longer disease-free survival, longer overall survival and better prognosis.