Role of estrogen in controlling the genital microflora of female rats

Plate counts of viable bacteria recovered by lavage from rat vaginae demonstrated that the number of bacteria associated with the vaginal epithelium varied cyclically and that this pattern was abolished by ovariectomy. After ovariectomy, vaginal bacterial counts remained relatively stable at low levels. The estrogen 17beta-estradiol (1,3,5(10)-estratriene-3,17beta-diol cypionate) administered to ovariectomized rats caused a significant increase in vaginal bacterial counts on day 3 post-treatment. A similar effect was seen in non-ovariectomized rats, but a larger dose of estrogen antagonist may have been present in non-ovariectomized animals. Progesterone (4-pregnene-3,20-dione) given with estradiol diminished the effect of the estrogen on vaginal bacterial counts, but did not abolish it. Progesterone administered without estradiol had no detectable effect on vaginal bacterial counts. These findings suggested that the cyclic variation in bacterial content of rat vaginae could be explained primarily as the effect of the secretory pattern of ovarian estrogen.     

Regulation of the bacterial microflora of the vagina in cyclic female rats.

The bacterial microflora was examined in the vagina of cyclic female rats kept under normal laboratory conditions. Large variations occurred during the cycle with high numbers of bacteria (10(5)-10(8) per vagina) during proestrus and estrus and low numbers (10(1)-10(4) per vagina) during the diestrus period. Histological analysis of in situ vaginal tissue and transplanted vaginal tissue revealed an association of high bacterial numbers with the presence of large amounts of cellular debris in the vaginal lumen during the period of epithelial keratinization. Absence of phagocytosis in leucocytes at mestestrus suggested that leucocytes did not play an active role in reduction of bacterial numbers between estrus and metestrus. Accurate measurement of the pH in the vaginal lumen failed to reveal differences which could explain the reduction in bacterial numbers between estrus and metestrus. The cyclic changes in the bacterial population-consisting of species which are normally present in the intestinal flora-- seem to be controlled by cyclic changes in the amounts of cellular debris in the vaginal lumen.     

Spatial relationship of the genital microflora to the vaginal epithelium of femal rats: transmission electron microscopy.

Bacterial associations with vaginal epithelium were examined by transmission electron microscopy in castrate rats before and after estrogen treatment. Bacteria were associated with keratinized epithelium and appeared to be most abundant on exfoliated vaginal cells.     

Diabetes abolishes the vascular protective effects of estrogen in female rats

Estrogen is known to exert a protective effect against cardiovascular disease. However, women with diabetes have three times the risk as compared with age-matched non-diabetic women. Our previous study on aortic rings of ovariectomized (OVX) female rats treated with 17-beta-estradiol (E2) demonstrated that the beneficial effect of estrogen is related to the basal release of NO from endothelial cells. In the present study, in order to understand why estrogen protection is abolished in diabetes, we tested vascular responses in OVX, streptozotocin-diabetic female rats and their non-diabetic controls receiving or not E2 replacement. Concentration-response curves to norepinephrine (NE) showed attenuation of the contractile response in E2-treated diabetic, with respect to non-diabetic preparations. This response was further impaired in diabetic, E2-deprived rats. The basal release of NO, as evaluated by concentration-related responses to N(G)-methyl-L-arginine acetate in NE-precontracted aortic rings, was found to be impaired in E2-treated diabetic rats, no further effect being induced by E2 deprivation. The endothelium-dependent relaxation produced by carbachol did not change between groups, whereas the relaxation produced by histamine was enhanced by both diabetes and E2 deprivation. However, E2 treatment counteracted the response to histamine only in preparations from non-diabetic animals. Finally, the relaxation induced by sodium nitroprusside, an endothelium-independent relaxant agent, was comparable between groups. These findings suggest that the lack of protective effects of estrogen in diabetes may be mainly ascribed to the failure of estrogen to reverse the impaired basal release of NO and the abnormal relaxation to histamine, which are observed in the aorta of diabetic rats.     

Group B streptococci in the female genital tract.

Vaginal carriage rates of group B streptococci among 250 women attending a clinic for sexually transmitted diseases, 123 attending family planning clinics, and 110 in labour wages were 36.0%, 17-1% and 6.4% respectively. The presence of group B streptococci was not associated with a vaginal discharge or the use of oral contraceptives in the non-pregnant women, or with the isolation of Neisseria gonorrhoeae or Trichomonas vaginalis from the women attending the clinic for sexually transmitted diseases. Serotyping showed a predominance of types II and III in non-pregnant women and an overall incidence of non-typable strains of 14.8%. There was no relationship between serotype and antibacterial susceptibility.     

Effects of ovariectomy and estrogen on ischemia-reperfusion injury in hindlimbs of female rats.

The effects of estrogen and ovariectomy on indexes of muscle damage after 2 h of complete hindlimb ischemia and 2 h of reperfusion were investigated in female Sprague-Dawley rats. The rats were assigned to one of three experimental groups: ovariectomized with a 17beta-estradiol pellet implant (OE), ovariectomized with a placebo pellet implant (OP), or control with intact ovaries (R). It was hypothesized that following ischemia-reperfusion (I/R), muscle damage indexes [serum creatine kinase (CK) activity, calpain-like activity, inflammatory cell infiltration, and markers of lipid peroxidation (thiobarbituric-reactive substances)] would be lower in the OE and R rats compared with the OP rats due to the protective effects of estrogen. Serum CK activity following I/R was greater (P < 0.01) in the R rats vs. OP rats and similar in the OP and OE rats. Calpain-like activity was greatest in the R rats (P < 0.01) and similar in the OP and OE rats. Neutrophil infiltration was assessed using the myeloperoxidase (MPO) assay and immunohistochemical staining for CD43-positive (CD43+) cells. MPO activity was lower (P < 0.05) in the OE rats compared with any other group and similar in the OP and R rats. The number of CD43+ cells was greater (P < 0.01) in the OP rats compared with the OE and R rats and similar in the OE and R rats. The OE rats had lower (P < 0.05) thiobarbituric-reactive substance content following I/R compared with the R and OP rats. Indexes of muscle damage were consistently attenuated in the OE rats but not in the R rats. A 10-fold difference in serum estrogen content may mediate this. Surprisingly, serum CK activity and muscle calpain-like activity were lower (P < 0.05) in the OP rats compared with the R rats. Increases in serum insulin-like growth factor-1 content (P < 0.05) due to ovariectomy were hypothesized to account for this finding. Thus both ovariectomy and estrogen supplementation have differential effects on indexes of I/R muscle damage.     

Spatial relationship of the vaginal microflora to the vaginal epithelium of female rats: scanning electron microscopy.

The vaginal mucosa of ovariectomized female rats has been examined by scanning electron microscopy before and after estrogen [1,3,5(10)-estratriene-3,17beta-diol] treatment. Without estrogen stimulation vaginal colonization is minimal and the epithelium is characterized by a layer of epithelial cells covered with small microvillous-like projections. Progressive changes that were consistent with estrogenic cytoproliferative effects were seen after estrogen treatment. By post-treatment day 3 bacterial colonization was maximal and the epithelium was comprised of flat squamous cells that tended to become detached from the underlying tissue layers. Bacteria were seen in association with the intercellular borders of this tissue and occurred singly or as microcolonies. No distinct physical attachment structures were identified, although an amorphous extracellular material that may serve to attach the bacteria to the squamous epithelial cells was frequently seen.     

Role of estrogen in modulating EDHF-mediated dilations in the female rat middle cerebral artery

We tested the hypothesis that endothelium-derived hyperpolarizing factor (EDHF) plays a less dominant role in the female cerebrovasculature. The contribution of EDHF to the ATP-mediated dilation was determined in middle cerebral arteries (MCAs) isolated from male and female rats. Four groups of rats were tested: intact male (n = 12), intact female (n = 13), estrogen-treated ovariectomized female (n = 13), and vehicle-treated ovariectomized female (n = 20) rats. Maximal dilation to ATP was similar in all groups. However, in the presence of N(omega)-nitro-L-arginine methyl ester (L-NAME, 3 x 10(-5) M) and indomethacin (10(-5) M), the maximal dilation to ATP was significantly reduced in intact female (24 +/- 9%) and estrogen-treated ovariectomized female (29 +/- 9%) rats compared with intact male (95 +/- 4%) and vehicle-treated ovariectomized female (96 +/- 2%) rats. The ATP-mediated dilation in L-NAME- and indomethacin-treated MCAs isolated from male and ovariectomized female rats was inhibited by charybdotoxin (10(-7) M), an inhibitor of large-conductance Ca2+-sensitive K+ channels. We have defined EDHF as the L-NAME- and indomethacin-insensitive component of the ATP-mediated dilation. Our findings indicate that EDHF-mediated dilations are negligible in the female rat MCA; these dilations can be significantly enhanced after ovariectomy, suggesting that this effect is mediated by estrogen.     

Spatial relationship of the genital microflora to the vaginal epithelium of femal rats: transmission electron microscopy

Bacterial associations with vaginal epithelium were examined by transmission electron microscopy in castrate rats before and after estrogen treatment. Bacteria were associated with keratinized epithelium and appeared to be most abundant on exfoliated vaginal cells.     

The influence of estrogen on the hyperglycemic action of alloxan in female rats

The purposes of this study were to investigate (1) the relationship between estrogen and glucose levels in the circulation during an estrous cycle of the rat and (2) the effect of estrogen on the hyperglycemic action of alloxan. Three-month old Long Evans strain rats were used. The present study established for the first time the correlation between estrogen and glucose levels in the circulation during an estrous cycle. A significant (p less than 0.01) negative correlation was observed with the lowest glucose level at the proestrus stage where estrogen is at peak. The severity of hyperglycemia induced by alloxan was positively correlated with the circulating level of endogenous estrogen, but was negatively correlated with the fasting glucose level. A similar pattern was observed in ovariectomized female rats that were followed by estradiol implantation to achieve a constant level of circulating estrogen. It may be concluded from the present study that estrogen, under physiological conditions, participates in the regulation of glucose metabolism during various stages of the estrous cycle in the rats, and that glucose is likely able to protect the pancreatic beta cells of the animals against the hyperglycemic action of alloxan.     

Effect of estrogen on global myocardial ischemia-reperfusion injury in female rats

We investigated the effects of estrogen on global myocardial ischemia-reperfusion injury in rats that were ovariectomized (Ovx), sham-operated, or ovariectomized and then given 17beta-estradiol (E(2)beta) supplementation (Ovx+E(2)beta). Hearts were excised, cannulated, perfused with and then immersed in chilled (4 degrees C) cardioplegia solution for 30 min, and then retrogradely perfused with warm (37 degrees C), oxygenated Krebs-Henseleit bicarbonate buffer for 120 min. The coronary flow rate, first derivative of left ventricular pressure, and nitrite production were all significantly lower in Ovx than in sham-operated or Ovx+E(2)beta hearts. However, coronary flow rates or nitrate production were not consistently different throughout the entire reperfusion period. Ca(2+) accumulated more in Ovx rat hearts than in sham-operated or Ovx+E(2)beta hearts, and mitochondrial respiratory function was lower in Ovx hearts than in hearts from the other two groups. Marked interstitial edema and contraction bands were seen in hematoxylin-eosin-stained sections of Ovx rat hearts but not in hearts from either of the other groups. Hematoxylin-basic fuchsin-picric acid-stained sections revealed fewer viable myocytes in hearts from the Ovx group than from the sham or Ovx+E(2)beta group. Transmission electron microscopy demonstrated more severely damaged mitochondria and ultrastructural damage to myocytes in Ovx rat hearts. Our results indicate that estrogen plays a cardioprotective role in global myocardial ischemia-reperfusion injury in female rats.     

Influence of wheat bran on NMU-induced mammary tumor development, plasma estrogen levels and estrogen excretion in female rats.

In our animal experiments the hypothesis was tested that a high-fiber (HF) diet reduces tumor promotion by interruption of the enterohepatic circulation resulting in lowered estrogen exposure of the estrogen-sensitive tissue. In the first experiment the development of N-nitrosomethylurea (NMU) induced mammary tumors was investigated. One group of rats (HF) was fed a HF diet (11% fiber, based on wheat bran), the other group (LF) fed a low-fiber diet (0.5% fiber, based on white wheat flour). Tumor incidence (90 and 80%, respectively) and latency (121 and 128 days, respectively) were similar in the HF and LF groups. Compared to the LF group, HF rats had lower tumor weights (0.16 vs 0.55 g; P less than 0.01) and a slightly lower tumor multiplicity (1.8 vs 2.8 tumors per tumor-bearing rat). These differences were reduced after adjustment for body weight. In a second experiment rats, not treated with the carcinogen, were kept on the same HF and LF diets. From these rats 24-h urine and feces and orbital blood samples were collected for analysis of (un)conjugated estrogens. The excretion of both free and conjugated estrogens in fecal samples was about 3-fold higher in HF rats than in LF rats. During the basal period of the cycle urinary excretion of estrone was lower in HF rats (mean 9.7 ng/day) than in LF rats (mean 13.0 ng/day; P less than 0.05). It is concluded that wheat bran interrupts the enterohepatic circulation of estrogens, but plasma levels are not affected. Whether the development of mammary tumors is reduced by the introduction of specific components of wheat bran, or by a reduced body weight due to a lower (effective) energy intake remains to be determined.     

Effect of estrogen pretreatment on the intestinal cytochrome p-450 in female rats

The effect of estrogen hormones on the cytochrome P-45O-linked hydroxylation system of rat small intestine was investigated. Estrogens were found to increase the activity of the intestinal arylhydrocarbon hydroxylase as well as the level of cytochrome P-45O in female rats. There was essentially no difference in the increase of the concentration of cytochrome P-45O due to the duration of pretreatment.