Quercetin induces morphological and proliferative changes of rat's uteri under estrogen and progesterone influences

Abstract

This study investigated the effect of 10 or 100 mg/kg/day quercetin on the uterus of ovariectomized adult female rats receiving sex-steroid replacement regime mimicking changes in hormonal profiles during the reproductive cycle. Following seven days of treatment with estrogen and progesterone with or without quercetin, uteri were harvested for histological and proliferative cell nuclear antigen (PCNA) protein and mRNA expression and PCNA protein distribution analyses. Our findings indicated that co-administration of 10 mg/kg/day quercetin with estrogen and progesterone caused a significant decrease in the size of uterine lumen and epithelial heights with lower PCNA protein and mRNA expression as compared to estrogen plus progesterone-only treatment (P < 0.05). Concomitant treatment with estrogen and progesterone with 100 mg/kg/day quercetin resulted in a marked increase in the number of glands with increased PCNA protein and mRNA expression. Significantly higher PCNA distribution was observed in the stroma and glands as compared to estrogen plus progesterone-only treatment (P < 0.05). In conclusion, at 10 mg/kg/day, quercetin affects uterine morphology but not proliferation, however at 100 mg/kg/day, quercetin induced significant stromal and glandular proliferation which could predispose the uterus towards neoplastic development.

Keywords: PCNA; Quercetin; proliferation; uterus.

Figure 1

Low magnification image of the uterus in different treatment groups. A lumen in (E+P)+10Q is relatively smaller than E+P and (E+P)+100Q group. A significantly high number of glands could be seen in the (E+P)+100Q group. E+P = estrogen plus progesterone, (E+P)+10Q = estrogen plus progesterone with 10 mg/kg/day quercetin, (E+P)+100Q = estrogen plus progesterone with 100 mg/kg/day quercetin. L, lumen. scale bar = 100 µm

Figure 2

PCNA mRNA fold changes in different treatment group. A marked increase in PCNA mRNA expression was observed following concomitant treatment of estrogen plus progesterone with 100 mg/kg/day quercetin. Meanwhile, significant decrease in PCNA mRNA expression was also noted following co-administration of 10 mg/kg/day quercetin with estrogen plus progesterone. C = vehicle, E+P = estrogen plus progesterone, (E+P)+10Q = estrogen plus progesterone with 10 mg/kg/day quercetin, (E+P)+100Q = estrogen plus progesterone with 100 mg/kg/day quercetin. *P < 0.05 as compared to C. †P < 0.05 as compared to E+P.

Figure 3

PCNA protein expression in different treatment groups. A significant increase in PCNA protein expression was noted following concomitant treatment of estrogen and progesterone with 100 mg/kg/day quercetin. Meanwhile, significantly lower expression of PCNA protein was noted following co-administration of 10 mg/kg/day quercetin with estrogen plus progesterone. C = vehicle, E+P = estrogen plus progesterone, (E+P) +10Q = estrogen plus progesterone with 10 mg/kg/day quercetin, (E+P)+100Q = estrogen plus progesterone with 100 mg/kg/day quercetin. *P < 0.05 as compared to C. †P < 0.05 as compared to E+P.

Figure 4

PCNA distribution in the glands of different treatment groups. A-D: FTIC with DAPI staining and E: FTIC staining alone for PCNA. Fluorescence signal could be seen arising from the glands. The highest intensity could be seen arising from (E+P)+100Q group (D & E images). L, lumen; G, glands; S, stroma; M, myometrium. A = control, B = estrogen plus progesterone, C = estrogen plus progesterone with 10 mg/kg/day quercetin, D & E = estrogen plus progesterone with 100 mg/kg/day quercetin.

Figure 5

PCNA distribution in the stroma in different treatment groups. (A-D) FTIC with DAPI staining and (E) FTIC staining alone for PCNA. Fluorescence signal could be seen arising from the stroma. The highest intensity could be seen arising from (E+P)+100Q group (D & E images). L, lumen; G, glands; S, stroma; E, epithelium. A = control, B = estrogen plus progesterone, C = estrogen plus progesterone with 10 mg/kg/day quercetin, D & E = estrogen plus progesterone with 100 mg/kg/day quercetin.

Figure 6

Mean fluorescence intensity in different treatment group. The highest fluorescence intensity in both (A) glands and (B) stroma could be seen in the group which received estrogen plus progesterone in the presence of 100 mg/kg/day quercetin. The intensity in the group receiving 10 mg/kg/day quercetin was significantly lesser than estrogen plus progesterone-only treatment group. C = vehicle, E+P = estrogen plus progesterone, (E+P)+10Q = estrogen plus progesterone with 10 mg/kg/day quercetin, (E+P)+100Q = estrogen plus progesterone with 100 mg/kg/day quercetin. *P < 0.05 as compared to C. †P < 0.05 as compared to E+P.

Figure 7

PCNA immunoperoxidase-stained cell nuclei. High number nuclei staining could be seen in the group which received estrogen plus progesterone-only treatment and estrogen plus progesterone in the presence of 100 mg/kg/day quercetin, with the latter has a relatively higher number of peroxidase reaction in the nuclei than the former. Stainings were mostly seen in the stroma. In E+P plus 100 mg/kg/day group, stainings were also seen in the glands. A = control, B = estrogen plus progesterone, C = estrogen plus progesterone with 10 mg/kg/day quercetin, D & E = estrogen plus progesterone with 100 mg/kg/day quercetin.