GnRH-a-Induced Perimenopausal Rat Modeling and Black Cohosh Preparations’ Effect on Rat’s Reproductive Endocrine

BackgroundEndometriosis (EMS) is an estrogen-dependent disease, which easily recurs after operation. Gonadotropin-releasing hormone agonist (GnRH-a), an estrogen-inhibiting drug, can effectively inhibit the secretion of gonadotropin by pituitary gland, so as to significantly decrease the ovarian hormone level and facilitate the atrophy of ectopic endometrium, playing a positive role in preventing postoperative recurrence. The application of GnRH-a can lead to the secondary low estrogen symptoms, namely the perimenopausal symptoms, and is a main reason for patients to give up further treatment. The add-back therapy based on sex hormones can well address the perimenopausal symptoms, but long-term use of hormones may cause the recurrence of EMS, as well as liver function damage, venous embolism, breast cancer and other risks, which has long been a heated topic in the industry. Therefore, it is necessary to find effective and safe anti-additive drugs soon. Studies at home and abroad show that, as a plant extract, isopropanolic extract of cimicifuga racemosa (ICR) can well relieve the perimenopausal symptoms caused by natural menopause. Some studies have preliminarily confirmed that black cohosh preparations can antagonize perimenopausal symptoms of EMS patients treated with GnRH-a after operation.ObjectiveTo establish a rat model of perimenopausal symptoms induced by GnRH-a injection, for the purposes of laying a foundation for further research and preliminarily exploring the effect of black cohosh preparations on reproductive endocrine of the rat model.MethodThe rat model of perimenopausal symptoms was established by GnRH-a injection, and normal saline (NS injection) was used as the control. The rats were randomly divided into four groups according to different modeling methods and drug intervention schemes. GnRH-a injection + normal saline intervention group (GnRH-a + NS), normal saline injection control + normal saline intervention group (NS + NS), GnRH-a injection + estradiol intervention group (GnRH-a + E2), and GnRH-a injection + black cohosh preparations intervention group (GnRH-a + ICR). After modelling was assessed to be successful with the vaginal smear method, the corresponding drugs were given for intervention for 28d. In the process of rat modeling and drug intervention, the skin temperature and anus temperature of the rat tails were measured every other day, the body weights of the rats were measured every other day, and the dosage was adjusted according to the body weight. After the intervention was over, the serum sex hormone level, the uterine weight, the uterine index, and the endometrial histomorphology changes, as well as the ovarian weight, the ovarian index, and the morphological changes of ovarian tissues of each group were measured.Results(1) The vaginal cell smears of the control group (NS + NS) showed estrous cycle changes, while other model rats had no estrous cycle of vaginal cells. (2) The body weight gains of the GnRH-a + NS, GnRH-a + E2 and GnRH-a + ICR groups were significantly higher than that of the NS + NS control group. The intervention with E2 and ICR could delay the weight gain trend of rats induced by GnRH-A. (3) After GnRH-a injection, the temperature of the tail and anus of rats showed an overall upward trend, and the intervention with E2 and ICR could effectively improve such temperature change. (4) The E2, FSH, and LH levels in the GnRH-a + NS, GnRH-a + E2, and GnRH-a + ICR groups were significantly lower than those in the NS + NS group (P < 0.01). The E2 level was significantly higher and the LH level was significantly lower in the GnRH-a + E2 group than those in the GnRH-a + NS and GnRH-a + ICR groups (P < 0.05). Compared with those of the GnRH-a + NS and GnRH-a + ICR groups, the FSH level of the GnRH-a + E2 group showed a slight downward trend, but the difference was not statistically significant (P > 0.05). There was no significant difference in the levels of sex hormones between the GnRH-a + NS group and GnRH-a + ICR group (P > 0.05). (5) Compared with those of the NS + NS group, the uterine weight and uterine index of the GnRH-a + NS, GnRH-a + E2 and GnRH-a + ICR groups significantly decreased (P < 0.01). In a comparison between the groups, the uterine weight and uterine index in the GnRH-a + NS and GnRH-a + ICR groups were significantly lower than those in the GnRH-a + E2 group (P < 0.01). There was a statistical difference in the uterine weight and uterine index between the GnRH-a + NS group and GnRH-a + ICR group (P > 0.05). (6) Compared with those of the NS + NS group, the ovarian weight and ovarian index of the GnRH-a + NS, GnRH-a + E2 and GnRH-a + ICR groups significantly decreased (P < 0.01). There was no statistical difference in the ovarian weight and ovarian index among the GnRH-a + E2, GnRH-a + NS and GnRH-a + ICR groups (P > 0.05). (7) Compared with those in the NS + NS group, the number of primordial follicles increased significantly, while the number of growing follicles and mature follicles decreased significantly in the GnRH-a + NS, GnRH-a + E2, and GnRH-a + ICR groups (P < 0.01), but there was a statistical difference in the total number of follicles among the four groups (P > 0.05).ConclusionsThe GnRH-a injection could achieve the desired effect. The animal model successfully achieved a significant decrease in the E2, FSH, and LH levels in rats, and could cause the rats to have rising body surface temperature similar to hot flashes in the perimenopausal period. The intervention with E2 and ICR could effectively relieve such “perimenopausal symptoms”, and ICR had no obvious effect on the serum sex hormone level in rats.

Uterine volume measurement as a determinant in route of hysterectomy

Background: Objectives of the study were to determine the role of uterine volume rather than uterine length in assessing the route of hysterectomy; to estimate the cut-off of uterine volume for route of hysterectomy; and to correlate uterine volume measured preoperatively by ultrasound with post-operative uterine weight.Methods: This was a prospective observational study including a total of 101 women who underwent hysterectomies (vaginal, laparascopic assisted vaginal hysterectomy (LAVH), total laparascopic hysterectomy (TLH), abdominal) in a period of 2 years 2 months from July 2018 to August 2020 in Mehta Multispeciality Hospital, Chetpet, Chennai. Uterine size was measured by clinical examination. Ease of the procedure with various uterine volume and routes of hysterectomy were studied.Results: 51 (50.49%) underwent vaginal route (including laparascopic hysterectomy), 50 (49.50%) underwent abdominal hysterectomy. Mean uterine volume leading to removal vaginally was 168.09±139.28 cc whereas 309.12±182.47 cc for abdominal hysterectomy (p=0.001) which was statistically significant. vaginal hysterectomy was done without difficulty up to 300 cc. Postoperative complications were less with vaginal hysterectomy compared to abdominal hysterectomy was statistically significant (p=0.0001).Uterine volume measured pre operatively by ultrasound showed positive correlation (r=0.82) with post-operative uterine weight proved that uterine volume measurements was superior to the clinical estimate of uterine size in assessing the route of hysterectomy.Conclusions: Uterine volume on ultrasonography (USG) can be a good predictor in deciding whether hysterectomy via vaginal route is possible.

Operative Complications and Outcomes Comparing Small and Large Uterine Weight in Case of Laparoscopic Hysterectomy for a Benign Indication

Study Objective: This study was performed to evaluate the association between uterine weight and operative outcomes in women undergoing laparoscopic hysterectomy for a benign indication.Methods: This is a secondary analysis of a randomized trial with data collected prospectively and retrospectively. The data of 159 women undergoing laparoscopic hysterectomy for a benign indication were analyzed. Women were divided in two groups according to the postoperative uterine weight: small uterus group (<250 grams) and large uterus group (≥250 grams). Operative complications were compared between the two groups. Operative outcomes (need for uterine morcellation, operative duration, estimated blood loss), postoperative pain, and hospital length of stay were also analyzed.Main Results: Operative complications were not significantly different between the two groups (37% in the large uterus group versus 41% in the small uterus group). Operative outcomes showed a significantly increased use of uterine morcellation in the large uterus group (61% in the large uterus group versus 10% in the small uterus group). The operative duration was 150 min in the small uterus group and 176 min in the large uterus group, which corresponds to an increase of 17% in the large uterus group. The mean pain score on the day of surgery was identical in both groups (VAS pain score 5), but significantly in favor of the large uterus group on day 1 postoperatively (VAS pain score 4 in the small uterus group and 3 in the large uterus group). There was no statistical difference between groups in the mean hospital stay (62 ± 37 hours in the small uterus group versus 54 ± 21 hours in the large uterus group). In terms of surgical indication, the small uterus group comprised more patients with endometriosis/adenomyosis (36%) and the large uterus group more patients with leiomyoma (93%).Conclusion: The results from this study show that, even if a large uterine weight is associated with increased uterine morcellation requirement and operative duration, a laparoscopic approach is safe and does not increase operative complications nor pain and/or length of hospital stay in women undergoing hysterectomy for a benign indication.

Clinical Risk Factors for Uterine Cervical Elongation among Women with Pelvic Organ Prolapse

Background: Cervical elongation is commonly associated with pelvic organ prolapse (POP). It was an identified risk for recurrent prolapse after hysteropexy, requiring additional surgeries. The aim of the study is to investigate the risk factors for uterine cervical elongation among women with POP. Methods: In this single-center retrospective cohort study, women who underwent vaginal total hysterectomy for POP between 2014 and 2016 were collected. The cervical and total uterine lengths were measured by pathologists, while the ratio of cervical length to total uterine length were calculated. The cervical elongation is defined as corpus/cervix ratio ≤ 1.5. Results: A total of 133 patients were enrolled in this study. Among these patients, 43 women had cervical elongation and 90 women had normal length of uterine cervix. We found that age > 65 years old (67.4% vs. 42.2%, p = 0.007), total vaginal length ≥ 9.5 cm (65.1% vs. 45.6%, p = 0.035), uterine weight < 51 gm (72.1% vs. 52.2%, p = 0.03), and Pelvic Organ Prolapse Distress Inventory 6 (POPDI-6) ≥ 12 (30.2% vs. 14.4%, p = 0.032) were associated with the risk of cervical elongation. There were no significant differences on preoperative urodynamic parameters in the two groups. Conclusion: The patient age > 65 years old, the total vaginal length of POP-Q system ≥ 9.5 cm, uterine weight < 51 g, and POPDI-6 ≥ 12 are independent risk factors of cervical elongation in women with POP. For women scheduled for pelvic reconstructive hysteropexy, concomitant cervical amputation should be considered.

Accuracy of Preoperative Pelvic Ultrasound in Predicting Uterine Weight Across Varying Pathologies

Objectives: The purpose of this study is to evaluate the accuracy of preoperative pelvic sonography in predicting the weight of a non-gravid uterus at the time of hysterectomy. Additionally, the study seeks to examine the effect of varying uterine pathologies on the accuracy of these predictions. Methods: This was a retrospective comparative study in a tertiary care hospital with a total of 318 patients who underwent pelvic ultrasound, with reported uterine volumes, prior to undergoing hysterectomy in 2018. Estimated uterine weights from preoperative pelvic ultrasounds were compared to actual uterine weights from post-hysterectomy pathology reports. A Bland-Altman plot was constructed to determine intermethod agreement with 95% limits of agreement. Simple linear regression analysis was used to correlate estimated and actual uterine weights. Multivariable linear regression was performed, allowing for subgrouping by uterine pathology. Results: While a strong positive correlation was identified between the two measurements (r = .93, p <.01), the Bland-Altman analysis showed that actual uterine weight ranged from 0.51 to 1.99 times the magnitude predicted by preoperative ultrasound. These wide deviations in intermethod agreement were similar amongst the various uterine pathologies. Multivariable linear regression analysis demonstrated the difference in estimated and actual uterine weight was 2 to 3.5 times larger for patients with endometrial pathology, adenomyosis, and leiomyomata as compared to normal uteri (p < .03). Conclusion: This study shows that in 95% of cases, actual uterine weight ranged from half to twice as large as estimated by preoperative pelvic ultrasound. Uterine pathology had a significant impact on the reliability of ultrasound for estimation of uterine weight. In gynecologic practices where preoperative estimations of uterine weight are instrumental in choosing a surgical approach for hysterectomy, reliance on pelvic ultrasound as a means of predicting uterine weight should be carefully considered.

Risk Factors for Perioperative Blood Transfusion in Patients Undergoing Hysterectomy for Benign Disease in a Teaching Institution

Purpose: The purpose is to identify risk factors for perioperative blood transfusion in patients undergoing hysterectomy for benign disease. Methods: This study is a retrospective chart review including all the patients who underwent hysterectomy for benign disease between January 1st 2018 and December 31st 2019. Patients who received perioperative blood transfusion were identified and compared to those who did not. The following risk factors for blood transfusion were analyzed: route of hysterectomy, BMI, presence of adhesions, history of cesarean section, uterine weight. Descriptive statistics was used to analyze the data. Results: A total of 517 patients were identified and included in the study. Forty-seven patients (9.09 %) received a perioperative blood transfusion. The abdominal hysterectomy route (TAH) was a significant risk factor for receiving blood transfusion (p=0.012). Other identified risk factors for blood transfusion included: Body mass index above 33.0 (p=0.002), and uterine weight (p=0.002). There was no association between the presence of pelvic adhesions (p=0.91) or a personal history of cesarean section (p=0.89) and receiving perioperative blood transfusion. When analyzing only the patients who underwent TLH, the presence of pelvic adhesion was found as a risk factor for perioperative blood transfusion (p=0.024) Conclusion: The abdominal hysterectomy route, the presence of a large uterus, and obesity are risk factors for receiving a blood transfusion. Early identification of the patient at risk of requiring perioperative blood transfusion provides better patient counseling and surgical preparation.

Placenta Morphological and Amniotic Fluid Biochemical Changes Associated with Mid-Gestation Single and Twin Pregnancies in Red Sokoto Goats

Fetal fluids and placentas, which are important in feto-maternal homeostasis, could be affected by the number of developing fetuses. Therefore, placenta morphological and amniotic fluid biochemical changes associated with single and twin pregnancies in Red Sokoto goats were studied. Using Richardson’s formula, 2.1 × [CRL (cm) + 17], 12 intact gravid uteri (7 single-pregnant and 5 twin-pregnant) from mid-gestation [≈70-100 days gestational age (dGA)] pregnant goats were purposively selected. Feto-maternal biometrics [gravid uterine weight (GUW), total placental fluid volume (TPFV), average placentome diameter (APD), mean crown-rump length (MCRL), mean gestational age (MGA), mean fetal weight (MFW), total placentome number (TPN), and average interplacentomal distance (AID)]; amniotic fluid electrolytes [sodium (Na), potassium (K), chloride (Cl), calcium (Ca) and phosphorus (P)]; total protein; glucose; liver markers (AST and ALT); kidney markers (urea and creatinine), and cortisol concentrations were measured. The histology of the placentomes and the interplacentomal areas was also studied. The twin-pregnant (TP) goats had significantly higher (P<0.05) GUW, TPFV, APD, amniotic fluid glucose and cortisol; and lower (P<0.05) Ca levels. The rest of the parameters assessed did not differ between the TP and single-pregnant (SP) groups. Placentomes and the interplacentomal areas from the TP group had more diffuse villous inter-digitations and thicker luminal endothelium, respectively. It was concluded that TP Red Sokoto goats regulated placentome morphology, calcium, glucose and cortisol levels in order to achieve optimal conditions for the dams and the fetuses.

Evaluation of C. pareira L stem and T. peruviana (pers.) K. Schum leaf for estrogenic activity

Context: Methanolic extract of C. pareira stem and T. peruviana leaves are capable of producing antifertility activity in female rat by increasing estrogen level and decreasing progesterone level. Aim: The aim of the present study is to C. pareira L stem and T. peruviana (Pers.) K. Schum leaf for estrogenic activity. Material and Method: Methanolic extract were made from C. pareira stem and T. peruviana leaves (after removal of cardiac glycoside) using cold maceration process. Extracts were tested for estrogenic activity on female immature rat (50-60 gm) at dose level of 250mg/kg. Obtained data were compared with control and standard (estrogen treated) group data. Statistical Analysis: Statistical analysis of all data was made with statistical packages (GraphPad Instat) and ANOVA followed by Dunnett’s ttest was used for statistical analysis. Result: Decrease in the absolute weight of the uterus and uterine weight to body weight ratio in CPS-Me and TPL-Me-G group treated were significant (p<0.05) as compared to control. Conclusion: In our earlier studies it was found that methanolic extract of C. pareira stem and T. peruviana leaves increased serum estrogen level and decreased progesterone level in pregnant rat. In the present study, these extracts decreased the absolute weight of the uterus and uterine weight to body weight ratio significantly (p<0.05). So, we can conclude that these two-extract possessed partial estrogenic activity.

Effects of Isoliquiritigenin on Bone Metabolism and Uterus in Ovariectomized Rats

Objectives Isoliquiritigenin (ILQ) is a phenolic compound found in licorice and is a popular dietary supplement. ILQ exhibits model-specific antioxidant, anti-inflammatory, anti-tumor, and estrogenic activities. Limited data suggest the potential of ILQ to prevent or treat osteoporosis. Therefore, this study evaluated the effects of short-duration treatment with ILQ on bone and uterine tissue in estrogen-deplete ovariectomized (ovx) rats. The uterus was important to evaluate because ILQ stimulates proliferation of MCF7 breast cancer cells through an estrogen receptor-dependent mechanism. Methods Six-week-old rats (ovx'd at 4 weeks of age) were fed diets containing 0, 100, 250 or 750 ppm ILQ (n = 5/treatment) for 1 week and sacrificed. Gene expression in femur and uterus, blood markers of global bone turnover, body composition, and uterine weight and epithelial cell height were determined. In addition, the effect of ILQ on in vitro differentiation of osteoclasts derived from bone marrow was assessed. Results Treatment resulted in a dose-dependent increase in serum ILQ with levels reaching 2.4 ± 0.2 mM in rats receiving the highest dose. ILQ did not alter serum levels of osteocalcin, a global marker of bone formation, or osteocalcin gene expression in femur. Additionally, there was little or no effect of ILQ on genes related to osteoblast differentiation or activity in femur. These largely null findings contrast with a reduction in serum CTX, a global marker of bone resorption, at all dose levels of ILQ. At the gene level, ILQ resulted in lower mRNA for genes related to osteoclast differentiation and function in femur, including Acp5 (tartrate resistant acid phosphatase), Timp2 and Mmp2, and suppressed osteoclast differentiation in vitro. ILQ had no effect on the ovx-induced increase in body weight. Ovx resulted in lower uterine weight. Treatment with ILQ at 750 ppm resulted in development of severe uterine epithelial cell hyperplasia in two of five animals. Conclusions ILQ supplementation led to reduced biochemical and gene expression markers of bone resorption in vivo and reduced osteoclast differentiation in vitro without increasing estrogen-dependent gene expression. However, the potential benefits must be weighed against potential detrimental off-target effects, including uterine hypertrophy. Funding Sources NIH [P50AT006268].