Role of inflammation in benign gynecologic disorders: from pathogenesis to novel therapies†.

Emerging evidence supports the notion that inflammation fosters the development of common benign gynecologic disorders, including uterine leiomyoma, endometriosis, and adenomyosis. Numerous cytokines, chemokines, and growth and transcription factors have indisputable roles in the establishment and maintenance of benign gynecologic disorders by initiating complex cascades that promote proliferation, angiogenesis, and lesion progression. The interaction between inflammation and benign gynecologic disorders is orchestrated by a plethora of factors, including sex steroids, genetics, epigenetics, extracellular matrix, stem cells, cardiometabolic risk factors, diet, vitamin D, and the immune system. The role of inflammation in these disorders is not limited to local pathobiology but also extends to involve clinical sequelae that range from those confined to the reproductive tract, such as infertility and gynecologic malignancies, to systemic complications such as cardiovascular disease. Enhanced understanding of the intricate mechanisms of this association will introduce us to unvisited pathophysiological perspectives and guide future diagnostic and therapeutic implications aimed at reducing the burden of these disorders. Utilization of inflammatory markers, microRNA, and molecular imaging as diagnostic adjuncts may be valuable, noninvasive techniques for prompt detection of benign gynecologic disorders. Further, use of novel as well as previously established therapeutics, such as immunomodulators, hormonal treatments, cardiometabolic medications, and cyclooxygenase-2 and NF-κB inhibitors, can target inflammatory pathways involved in their pathogenesis. In this comprehensive review, we aim to dissect the existing literature on the role of inflammation in benign gynecologic disorders, including the proposed underlying mechanisms and complex interactions, its contribution to clinical sequelae, and the clinical implications this role entails.

Vascular biology of uterine fibroids: connecting fibroids and vascular disorders.

Fibroids are benign tumors caused by the proliferation of myometrial smooth muscle cells in the uterus that can lead to symptoms such as abdominal pain, constipation, urinary retention, and infertility. While traditionally thought of as a disease process intrinsic to the uterus, accumulating evidence suggests that fibroid growth may be linked with the systemic vasculature system, although cell-intrinsic factors are certainly of principal importance in their inception. Fibroids are associated with essential hypertension and preeclampsia, as well as atherosclerosis, for reasons that are becoming increasingly elucidated. Factors such as the renin-angiotensin-aldosterone system, estrogen, and endothelial dysfunction all likely play a role in fibroid pathogenesis. In this review, we lay out a framework for reconceptualizing fibroids as a systemic vascular disorder, and discuss how pharmaceutical agents and other interventions targeting the vasculature may aid in the novel treatment of fibroids.

Effect of endometrial mechanical stimulation in an unselected population undergoing in vitro fertilization: futility analysis of a double-blind randomized controlled trial.

PURPOSE: Implantation failure is a major limiting factor of successful in vitro fertilization (IVF). The objective of this study was to determine if endometrial mechanical stimulation (EMS) by endometrial biopsy in the luteal phase of the cycle prior to embryo transfer (ET) improves clinical outcomes in an unselected subfertile population. METHODS: Double-blind, randomized controlled trial of EMS versus sham biopsy and odds of clinical pregnancy after IVF and embryo transfer. Secondary outcomes included spontaneous miscarriage and live birth. RESULTS: One hundred women enrolled and were randomized from 2013 to 2017. Enrollment was terminated after futility analysis showed no difference in clinical pregnancy between EMS versus control, 47.2% vs 61.7% (OR 0.55, 95% CI 0.25-1.23, p = 0.15). There were no significant differences between women who underwent EMS and those who did not in terms of positive pregnancy test 54.7% vs 63.8% (OR 0.69, 95% CI 0.31-1.53, p = 0.36), miscarriage 7.5% vs 2.1% (OR 3.76 95% CI 0.41-34.85, p = 0.22), or live birth 43.4% vs 61.7% (OR 0.48 95% CI 0.21-1.06, p = 0.07). CONCLUSIONS: EMS in the luteal phase of the cycle preceding embryo transfer does not improve clinical outcomes in an unselected subfertile population and may result in a lower live birth rate. We caution the routine use of EMS in an unselected population.

Correction to: Effect of endometrial mechanical stimulation in an unselected population undergoing in vitro fertilization: futility analysis of a double-blind randomized controlled trial.

The original version of this article unfortunately contained mistakes. The complete list of corrections is given below.

Elevated uric acid and obesity-related cardiovascular disease risk factors among hypertensive youth.

BACKGROUND: Uric acid (UA) is associated with high blood pressure in adolescents and with left ventricular hypertrophy (LVH) and cardiovascular disease (CVD) in adults. We sought to determine if UA is independently associated with CVD risk factors and left ventricular mass (LVM) over time in hypertensive youth. METHODS: This was a 1-year prospective observational study of hypertensive children aged 3-19 years. Cross-sectional and longitudinal associations of serum UA with CVD risk factors and LVM were explored. RESULTS: Of the 49 children who completed both the baseline and 12-month assessments, at baseline the mean age was 13.8 years and mean UA was 5.5 mg/dL; 24% had elevated UA, 51% were overweight/obese and 39% had LVH. Measures of adiposity, low high-density lipoprotein cholesterol, high-sensitivity C-reactive protein, LVM and LVH were all significantly associated with elevated UA at baseline, but not with change over time. Each 1 mg/dL increase in baseline UA was associated with a 2.5 g/m(2.7) increase in the LVM index at follow-up (95% confidence interval 0.64, 4.39; p = 0.01); after adjustment for age, sex, race, body mass index z-score, change in UA, time, blood pressure and medication use, this association was no longer significant. CONCLUSIONS: Hypertensive children with elevated UA have a higher prevalence of obesity-related CVD risk factors. Among hypertensive children, UA may be a marker of adiposity and not an independent CVD risk factor.

Adipocyte coculture induces a pro-inflammatory, fibrotic, angiogenic, and proliferative microenvironment in uterine leiomyoma cells.

OBJECTIVE: Obesity and its consequences are among the biggest challenges facing the healthcare system. Uterine leiomyomas are the most common gynecologic tumors. The risk of leiomyoma increases with obesity, but the underlying mechanisms of this association remain unclear. The aim of the present study to determine the cellular and molecular mechanisms by which adipocyte contributes to both leiomyoma tumor initiation and promotion. METHODS: Primary myometrium and leiomyoma cells were isolated from patients who underwent a hysterectomy or myomectomy. Pro-inflammatory, fibrotic, and angiogenic factors were measured using a multiplex cytokine array in human primary and immortalized myometrial and leiomyoma cells cocultured with human adipocyte (SW872) cells, or in animal ELT3 leiomyoma cells cocultured with 3 T3-L1 adipocytes. The free fatty acids (FFAs) and fatty acid-binding protein 4 (FABP4) levels were measured using immunofluorescence assays. Other protein abundances were determined using western blots. The expression levels of TNF-α, MCP-1, phospho-NF-κB, TGFß3 and VEGF-A in lean and obese in different leiomyoma patients were determined by immunofluorescence staining. RESULTS: Adipocytes promote inflammation, fibrosis, and angiogenesis in uterine leiomyoma cells by upregulating associated factors, such as IL-1ß, TNF-α, MCP-1, GM-CSF, TGF-ßs, PLGF, VEGF, HB-EGF, G-CSF and FGF2. Coculture led to the transfer of FFAs and FABP4 from adipocytes to leiomyoma cells, suggesting that adipocytes may modulate metabolic activity in these tumor cells. Increased levels of FFA and FABP4 expressions were detected in obese leiomyoma tissue compared to lean. The adipocyte-leiomyoma cell interaction increased the phospho-NF-κB level, which plays a key role in inflammation, restructuring metabolic pathways, and angiogenesis. Obese leiomyoma patients expressed a higher amount of TNF-α, MCP-1, phospho-NF-κB, TGFß3 and VEGF-A than lean leiomyoma patients, consistent with in vitro findings. Furthermore, we found that adipocyte secretory factors enhance leiomyoma cell proliferation by increasing PCNA abundance. Finally, the inhibition of the inflammatory factors TNF-α, MCP-1, and NF-κB abrogated the adipocyte coculture-induced proliferation of leiomyoma cells. CONCLUSIONS: Adipocytes release inflammatory, fibrotic, and angiogenic factors, along with FFAs, which contribute to a tumor-friendly microenvironment that may promote leiomyoma growth and can represent a new target for leiomyoma prevention and treatment.

Differential response to hypoxia in leiomyoma and myometrial cells.

AIMS: Recent evidence suggests that repetitive hypoxia occurs during menstrual cycles due to vasoconstriction and myometrial contraction. It is unknown if hypoxia contributes to the development of uterine leiomyoma, the most common tumor of the female reproductive system. This study aims to characterize the response to hypoxia in leiomyoma and myometrial cells; and determine if an aberrant leiomyoma response to hypoxia may contribute to leiomyomatogenesis. MAIN METHODS: Primary and immortalized leiomyoma and myometrial cells were cultured under normoxic and hypoxic conditions. Expression levels of vascular endothelial growth factor-A (VEGF-A), adrenomedullin (ADM), endothelin-1 (ET-1), and hypoxia-inducible factor-1 alpha (HIF-1α) were measured by qRT-PCR, western blotting and ELISA. Cell proliferation was assessed using MTT assay and proliferating-cell-nuclear-antigen (PCNA) expression. KC7F2 (HIF-1α inhibitor) was used to examine the regulating mechanisms. KEY FINDINGS: As expected, hypoxia induced HIF-1α expression in both leiomyoma and myometrial cells. However, hypoxia induced VEGF-A, ET-1 and ADM expression and VEGF-A secretion into the culture media in leiomyoma but not myometrial cells. MTT assay and PCNA expression showed that hypoxia induces proliferation in leiomyoma, but not myometrial cells. HIF-1α inhibitor abrogated the hypoxia-induced VEGF-A, ET-1, ADM, and PCNA expression in leiomyoma cells. SIGNIFICANCE: This study suggests an aberrant leiomyoma cellular response to hypoxia compared to myometrium. This differential response to menstruation-related repetitive hypoxia episodes may lead to selective proliferation of hypoxia-adaptive leiomyoma cells and contribute to leiomyoma growth. Thus, in addition to adding to our understanding of leiomyoma pathobiology, the study proposes angiogenic factors as a potential leiomyoma therapeutic target.

Leptin induces leiomyoma cell proliferation and extracellular matrix deposition via JAK2/STAT3 and MAPK/ERK pathways.

OBJECTIVE: To investigate the molecular effects of leptin on uterine leiomyoma cells. DESIGN: Experimental study using in vitro culture of immortalized human leiomyoma (HuLM) cells. SETTING: Academic university center. PATIENT(S): Women with uterine fibroids who underwent a hysterectomy or myomectomy. INTERVENTION(S): Administration of human recombinant leptin to the media of cultured HuLM cells separately or in combination with pharmacologic Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) or mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) inhibitors. MAIN OUTCOME MEASURE(S): We examined HuLM tissues and cells for the expression of the leptin receptor, termed OB-R. Cellular proliferation was measured at 6, 24, and 48 hours using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide assay. Protein expression levels of proliferating cell nuclear antigen, collagen 1, phosphorylated STAT3/total STAT3, and phosphorylated ERK1/2 and total ERK1/2 were quantified using immunoblotting. Pharmacologic inhibitors were employed to further assess the role of the JAK2/STAT3 and MAPK/ERK pathways in the proliferative response. RESULT(S): The presence of OB-R was confirmed in clinical leiomyoma and myometrial tissue obtained from 3 separate human subjects using immunofluorescence staining, and the expression of OB-R in HuLM cells was identified using immunoblotting. There was no significant difference in the expression of the leptin receptor in the myometrium compared with that in the leiomyoma tissue. Leptin stimulated cell proliferation and extracellular matrix (ECM) deposition at 24 hours after treatment. Pretreatment with a JAK2/STAT3 inhibitor attenuated ECM deposition, and pretreatment with a MAPK/ERK inhibitor significantly decreased leptin's stimulatory effect on cell proliferation and ECM deposition. CONCLUSION(S): Leptin induces a proliferative response and ECM deposition in HuLM cells. These findings suggest that leptin, acting through the JAK2/STAT3 and MAPK/ERK pathways, is involved in the development of uterine leiomyomas, which may partly explain their increased incidence in obese women.

Hypoxia induces proliferation via NOX4-Mediated oxidative stress and TGF-ß3 signaling in uterine leiomyoma cells.

Uterine leiomyomas, the most common tumors of the female reproductive system, are known to have a hypoxic microenvironment. However, the role of such environment in leiomyoma pathobiology remains unknown. The objective was to determine the effects of hypoxia on leiomyoma cells, and the mechanisms. We found that hypoxia induces proliferation and inhibits apoptosis in human leiomyoma cells. This pro-proliferative effect was accompanied by an increase in reactive oxygen species (ROS) generation and the expression of NADPH oxidase 4 (NOX4). The specific NOX4 inhibitor GLX351322 abrogated this hypoxia-induced ROS generation, cellular proliferation, and apoptosis inhibition. To further investigate the mechanism of NOX4-mediated proliferation, we treated leiomyoma cells grown in normoxia with media from leiomyoma cells cultured under hypoxia. This resulted in increased ROS generation and NOX4 expression, suggesting the hypoxia-induced effects are mediated by an autocrine mechanism. We worked to identify the nature of this autocrine factor. We found that the expression of TGF-ß3 and its downstream signaling target pSmad3, are increased in hypoxic leiomyoma cells. To examine the hypothesis that TGF-ß3 is, at least, a part of this autocrine mechanism, we treated hypoxic leiomyoma cells with the HIF-1α inhibitor KC7F2 which we discovered to ameliorate the hypoxia-induced TGF-ß3 expression. Furthermore, pharmacologic inhibition with the TGF-ß/Smad inhibitor SB431542 reduced hypoxia-induced NOX4 expression and ROS generation and attenuated cell proliferation. Thus, we have identified a novel mechanism by which hypoxia induces proliferation in leiomyoma cells. This finding adds to our understanding of leiomyoma pathobiology and can help in identifying new therapeutic targets.

Trainee-Led Quality Improvement Project to Improve Fertility Preservation Counseling for Patients With Cancer.

PURPOSE: Oncofertility counseling regarding the reproductive risks associated with cancer therapy is essential for quality cancer care. We aimed to increase the rate of oncofertility counseling for patients of reproductive age (18-40 years) with cancer who were initiating systemic therapy at the Johns Hopkins Cancer Center from a baseline rate of 37% (25 of 68, June 2019-January 2020) to 70% by February 2021. METHODS: We formed an interprofessional, multidisciplinary team as part of the ASCO Quality Training Program. We obtained data from the electronic medical record and verified data with patients by phone. We surveyed patients, oncologists, and fertility specialists to identify barriers. After considering a prioritization matrix, we implemented Plan-Do-Study-Act (PDSA) cycles. RESULTS: We identified the following improvement opportunities: (1) oncologist self-reported lack of knowledge about counseling and local fertility preservation options and (2) lack of a standardized referral mechanism to fertility services. During the first PDSA cycle (February 2020-August 2020, disrupted by COVID-19), we introduced the initiative to increase oncofertility counseling at faculty meetings. From September 2020 to November 2020, we implemented a second PDSA cycle: (1) educating and presenting the initiative at Oncology Grand Rounds, (2) distributing informative pamphlets to oncologists and patients, and (3) implementing an electronic medical record order set. In the third PDSA cycle (December 2020-February 2021), we redesigned the order set to add information (eg, contact information for fertility coordinator) to the patient after-visit summary. Postimplementation (September 2020-February 2021), counseling rates increased from 37% to 81% (38 of 47). CONCLUSION: We demonstrate how a trainee-led, patient-centered initiative improved oncofertility care. Ongoing work focuses on ensuring sustainability and assessing the quality of counseling.

Diet and Nutrition in Gynecological Disorders: A Focus on Clinical Studies.

A healthy lifestyle and a balanced diet play a paramount role in promoting and maintaining homeostatic functions and preventing an array of chronic and debilitating diseases. Based upon observational and epidemiological investigations, it is clear that nutritional factors and dietary habits play a significant role in gynecological disease development, including uterine leiomyoma, endometriosis, polycystic ovary syndrome, and gynecological malignancies. Diets rich in fruits and vegetables, Mediterranean diets, green tea, vitamin D, and plant-derived natural compounds may have a long-term positive impact on gynecological diseases, while fats, red meat, alcohol, and coffee may contribute to their development. Data regarding the association between dietary habits and gynecological disorders are, at times, conflicting, with potential confounding factors, including food pollutants, reduced physical activity, ethnic background, and environmental factors limiting overall conclusions. This review provides a synopsis of the current clinical data and biological basis of the association between available dietary and nutritional data, along with their impact on the biology and pathophysiology of different gynecological disorders, as well as an outlook on future directions that will guide further investigational research.

Frozen-thawed embryo transfer: the potential importance of the corpus luteum in preventing obstetrical complications.

The use of frozen-thawed embryo transfer (FET) has increased over the past decade with improvements in technology and increasing live birth rates. FET facilitates elective single-embryo transfer, reduces ovarian hyperstimulation syndrome, optimizes endometrial receptivity, allows time for preimplantation genetics testing, and facilitates fertility preservation. FET cycles have been associated, however, with an increased risk of hypertensive disorders of pregnancy for reasons that are not clear. Recent evidence suggests that absence of the corpus luteum (CL) could be at least partly responsible for this increased risk. In a recent prospective cohort study, programmed FET cycles (no CL) were associated with higher rates of preeclampsia and preeclampsia with severe features compared with modified natural FET cycles. FET cycles are commonly performed in the context of a programmed cycle in which the endometrium is prepared with the use of exogenous E2 and P. In these cycles, ovulation is suppressed and therefore the CL is absent. The CL produces not only E2 and P, but also vasoactive products, such as relaxin and vascular endothelial growth factor, which are not replaced in a programmed FET cycle and which are hypothesized to be important for initial placentation. Emerging evidence has also revealed other adverse obstetrical and perinatal outcomes, including postpartum hemorrhage, macrosomia, and post-term birth specifically in programmed FET cycles compared with natural FET cycles. Despite the widespread use of FET, the optimal protocol with respect to live birth rate, maternal health, and perinatal outcomes has yet to be determined. Future practice regarding FET should be based on high-quality evidence, including rigorous controlled trials.

Maternal obesogenic diet induces endometrial hyperplasia, an early hallmark of endometrial cancer, in a diethylstilbestrol mouse model.

Thirty-eight percent of US adult women are obese, meaning that more children are now born of overweight and obese mothers, leading to an increase in predisposition to several adult onset diseases. To explore this phenomenon, we developed a maternal obesity animal model by feeding mice a diet composed of high fat/ high sugar (HF/HS) and assessed both maternal diet and offspring diet on the development of endometrial cancer (ECa). We show that maternal diet by itself did not lead to ECa initiation in wildtype offspring of the C57Bl/6J mouse strain. While offspring fed a HF/HS post-weaning diet resulted in poor metabolic health and decreased uterine weight (regardless of maternal diet), it did not lead to ECa. We also investigated the effects of the maternal obesogenic diet on ECa development in a Diethylstilbestrol (DES) carcinogenesis mouse model. All mice injected with DES had reproductive tract lesions including decreased number of glands, condensed and hyalinized endometrial stroma, and fibrosis and increased collagen deposition that in some mice extended into the myometrium resulting in extensive disruption and loss of the inner and outer muscular layers. Fifty percent of DES mice that were exposed to maternal HF/HS diet developed several features indicative of the initial stages of carcinogenesis including focal glandular and atypical endometrial hyperplasia versus 0% of their Chow counterparts. There was an increase in phospho-Akt expression in DES mice exposed to maternal HF/HS diet, a regulator of persistent proliferation in the endometrium, and no difference in total Akt, phospho-PTEN and total PTEN expression. In summary, maternal HF/HS diet exposure induces endometrial hyperplasia and other precancerous phenotypes in mice treated with DES. This study suggests that maternal obesity alone is not sufficient for the development of ECa, but has an additive effect in the presence of a secondary insult such as DES.

Chronodisruption: An untimely cause of preterm birth?

Circadian rhythms, endogenous and entrainable adaptations to 24-hour cycles of light and dark, influence almost all physiologic functions. Emerging evidence suggests that the disruption of normal circadian rhythms, termed chronodisruption, could affect a wide range of disease-related processes. In this review, we describe the molecular generation of circadian rhythms, the effects of chronodisruption on human health, the circadian timing of birth in multiple species, the possible effects of chronodisruption on preterm birth, and some of the open questions in this field.