Background: Endometrial carcinoma is the most common cancer of the female genitalia and its prevalence is 2% to 3% along the females' lifetime. This adenocarcinoma is diagnosed in the early stages because the patients become symptomatic early in the course of disease. The correlation of the lower uterine segment involvement (LUSI) with the recurrence, and the survival rates in patients with endometrial adenocarcinoma are always questionable. Confirming the prognostic significance of LUSI can fundamentally improve the current state of patients' surveillance. The current study aimed at investigating the association of lower uterine segment involvement with deep myometrial invasion in endometrial adenocarcinoma. Methods: In the current retrospective cohort study, 54 patients with stage I endometrial endometrioid adenocarcinoma who underwent surgery in Mirza-Koochak-Khan Hospital, Tehran, Iran, from 2004 to 2014 were divided into 2 groups according to the presence of LUSI. In the current cohort study, the data from the patients exposed to LUSI were obtained by questionnaires to measure deep myometrial invasion besides post-operative adjuvant radiotherapy. The median of follow-up period was 48 months for all of the patients, which started after their surgery, and the measurement period for variables were similar to those of the follow-up period, which was 48 months. They were compared regarding to age, tumor grade, depth of myometrial invasion, lymphovascular involvement, extra uterine diseases, the extent of lymphadenectomy, adjuvant therapy, recurrence rate, time, and location. The data analysis was conducted by the SPSS v.16 statistical software (Chicago, IL, USA) at the significance level of 5%. In the investigation, Chi-square, the Kolmogorov-Smirnov, t test, and the Mann-Whitney U tests were used. Results: Group 1 consisted of 13 patients with LUSI and group 2 had 41 patients without LUSI divided by a non-random sampling method. According to the results of the Mann-Whitney U test, there was a significant difference between the mean age of patients with LUSI and that of the ones without LUSI (P = 0.03). It showed that the mean age in the former group was significantly higher than that of the latter. A Chi-square test showed no significant association between the lymphovascular involvement and the presence of LUSI (P = 0.1). The Fisher exact test showed that patients with LUSI had significantly higher rate of radiotherapy after surgery rather than the ones without LUSI (P < 0.001). Conclusions: In conclusion, there was a significant association between the lower uterine segment involvements with deep myometrial invasion in the endometrial endometrioid adenocarcinoma. In other words, there was a significant difference in the depth of myometrial invasion between the groups, and the patients with LUSI had deeper myometrial invasion (the Mann-Whitney U test, P < 0.001).