Background: Sex steroid hormones and their receptors are important in female sexual function. The aim of this study was to investigate the expression and distribution of estrogen receptor (ER)α, ERβ, G-protein-coupled ER-1 (GPER), androgen receptor (AR), progesterone receptor (PR)A, PRB and connective tissue growth factor (CTGF) in the vaginal wall among women who had been treated for cervical cancer with radiotherapy. Material and methods: We included cervical cancer survivors treated with radiotherapy and premenopausal control women of the same age scheduled for benign gynecological surgery. We analyzed the expression and distribution of sex steroid hormone receptors and CTGF in biopsies from the vaginal wall, by real-time PCR and immunohistochemistry (IHC). Serum samples were analyzed for hormone levels and radiation dose at biopsy site were calculated and correlated to levels of the sex steroid hormone receptors. Results: In the cervical cancer survivors (n = 34), we found a lower expression of ERα at both mRNA and protein levels, compared to the control women (n = 37). In the survivors with high radiation dose at biopsy site, the immunostaining of ERα and AR was lower in the epithelium and the stroma, compared to survivors with minimal radiation dose. The later group showed expression of ERα comparable to the control women. The cancer survivors were sufficiently substituted with systemic estradiol with no difference in the serum estradiol levels compared to control women. Conclusions: We found that external radiation reduces the ERα and AR protein expression in the vaginal mucosa, indicating that the vaginal changes in irradiated cervical cancer survivors and the lack of response to hormonal treatment could be due to the decreases in sex steroid hormone receptor expression.