In cases of rectal cancer surgery, patients at high risk of anastomotic leakage often receive a defunctioning stoma (DS). However, its role in postoperative anastomotic strictures (AS) remains unclear. This study aimed to investigate the correlation between DS and AS and outcomes of transanal endoscopic microsurgery (TEM) in treating rectal AS. This retrospective study was conducted from January 2019 to September 2021 and included patients who underwent rectal cancer surgery. A 1:1 ratio was used for propensity score matching (PSM). Univariate analyses were performed to identify statistically significant variables, and multivariate analyses were conducted to determine the factors affecting AS. This study included 383 patients. The results of the univariate analysis suggested that surgery time (HR 4.597, 95% CI 1.563-13.525, P=0.006), postoperative anastomotic leakage (HR 11.830, 95% CI 3.773-37.094, P<0.001), and DS (HR 15.475, 95% CI 6.042-39.641, P<0.001) were significantly associated with AS. In the multivariate analysis, postoperative anastomotic leakage (HR 7.596, 95% CI 1.987-29.044, P= 0.003) and DS (HR 11.252, 95% CI 4.113-30.779, P<0.001) were identified as significant risk factors for AS. After matching, the univariate analysis revealed that postoperative anastomotic leakage (HR 8.333, 95% CI 1.541-45.052, P= 0.014) and DS (HR 9.965, 95% CI 2.200-45.142, P= 0.003) were associated with AS. The multivariate analysis indicated that postoperative anastomotic leakage (HR 14.549, 95% CI 1.765-119.913, P= 0.013) and DS (HR 12.450, 95% CI 2.418-64.108, P= 0.003) were significant risk factors for AS. This study provides evidence that DS is independently associated with AS, and postoperative anastomotic leakage increases the risk of AS. Furthermore, this study suggests that TEM could be a valuable treatment option for AS.