Dexamethasone affects human fetal adrenal steroidogenesis and subsequent ACTH response in an ex vivo culture model.

Introduction: Administration of dexamethasone (DEX) has been used experimentally to suppress androgenization of external genitalia in 46,XX fetuses with congenital adrenal hyperplasia. Despite this, the prenatal biological mechanism-of-action of DEX on fetal development is not known. This study aimed to examine direct effects of DEX on human fetal adrenal (HFA) steroidogenic activity including possible effects on the subsequent response to ACTH-stimulation. Methods: Human fetal adrenal (HFA) tissue from 30 fetuses (1st trimester) were cultured ex vivo with A) DEX (10 µm) for 14 days, or B) DEX (10 µm) for 10 days followed by ACTH (1 nM) for 4 days. DEX-mediated effects on HFA morphology, viability, and apoptosis (immunohistochemistry), gene expression (quantitative PCR), and steroid hormone secretion (LC-MS/MS) were investigated. Results: DEX-treatment caused decreased androstenedione (p<0.05) and increased cortisol (p<0.01) secretion suggesting that direct effects on the adrenal gland may contribute to the negative feedback on the hypothalamic-pituitary-adrenal axis in vivo. An altered response to ACTH stimulation in HFA pre-treated with DEX included increased androgen (p<0.05) and reduced cortisol production (p<0.05), supporting clinical observations of a temporary decreased ACTH-response following prenatal DEX-treatment. Additionally, the secretion of corticosterone was decreased (p<0.0001) following ACTH-stimulation in the initially DEX-treated HFAs. Discussion: The observed effects suggest that prenatal DEX-treatment can cause direct effects on HFA steroidogenesis and in the subsequent response to ACTH-stimulation. This may indicate a requirement for careful monitoring of adrenal function in prenatally DEX-treated neonates, with particular focus on their mineralocorticoid levels.

Signs of a Glucose- and Insulin-Independent Gut-Bone Axis and Aberrant Bone Homeostasis in Type 1 Diabetes.

CONTEXT: Gut hormones seem to play an important role in postprandial bone turnover, which also may be affected by postprandial plasma glucose excursions and insulin secretion. OBJECTIVE: To investigate the effect of an oral glucose tolerance test (OGTT) and an isoglycemic intravenous glucose infusion (IIGI) on bone resorption and formation markers in individuals with type 1 diabetes and healthy controls. METHODS: This observational case-control study, conducted at the Center for Clinical Metabolic Research, Gentofte Hospital, Hellerup, Denmark, included 9 individuals with C-peptide negative type 1 diabetes and 8 healthy controls matched for gender, age, and body mass index. Subjects underwent an OGTT and a subsequent IIGI. We analyzed changes in bone resorption assessed by measurements of carboxy-terminal type I collagen crosslinks (CTX) and in bone formation as assessed by procollagen type I N-terminal propeptide (PINP) concentrations. RESULTS: Baseline CTX and PINP levels were similar in the 2 groups. Both groups exhibited significantly greater suppression of CTX during OGTT than IIGI. PINP levels were unaffected by OGTT and IIGI, respectively, in healthy controls. Participants with type 1 diabetes displayed impaired suppression of CTX-assessed bone resorption and inappropriate suppression of PINP-assessed bone formation during OGTT. CONCLUSION: Our data suggest the existence of a gut-bone axis reducing bone resorption in response to oral glucose independently of plasma glucose excursions and insulin secretion. Subjects with type 1 diabetes showed impaired suppression of bone resorption and reduced bone formation during OGTT, which may allude to the reduced bone mineral density and increased fracture risk characterizing these individuals.

Using Wearable Technology to Quantify Physical Activity Recovery: Secondary Report From the AFTER (App-Facilitated Tele-Rehabilitation) Program for COVID-19 Survivors Randomized Study.

BACKGROUND: Knowledge on physical activity recovery after COVID-19 survival is limited. The AFTER (App-Facilitated Tele-Rehabilitation) program for COVID-19 survivors randomized participants, following hospital discharge, to either education and unstructured physical activity or a telerehabilitation program. Step count data were collected as a secondary outcome, and we found no significant differences in total step count trajectories between groups at 6 weeks. Further step count data were not analyzed. OBJECTIVE: The purpose of this analysis was to examine step count trajectories and correlates among all participants (combined into a single group) across the 12-week study period. METHODS: Linear mixed models with random effects were used to model daily steps over the number of study days. Models with 0, 1, and 2 inflection points were considered, and the final model was selected based on the highest log-likelihood value. RESULTS: Participants included 44 adults (41 with available Fitbit [Fitbit LLC] data). Initially, step counts increased by an average of 930 (95% CI 547-1312; P<.001) steps per week, culminating in an average daily step count of 7658 (95% CI 6257-9059; P<.001) at the end of week 3. During the remaining 9 weeks of the study, weekly step counts increased by an average of 67 (95% CI -30 to 163; P<.001) steps per week, resulting in a final estimate of 8258 (95% CI 6933-9584; P<.001) steps. CONCLUSIONS: Participants showed a marked improvement in daily step counts during the first 3 weeks of the study, followed by more gradual improvement in the remaining 9 weeks. Physical activity data and step count recovery trajectories may be considered surrogates for physiological recovery, although further research is needed to examine this relationship. TRIAL REGISTRATION: ClinicalTrials.gov NCT04663945; https://tinyurl.com/2p969ced.

Identification of a window of androgen sensitivity for somatic cell function in human fetal testis cultured ex vivo.

BACKGROUND: Reduced androgen action during early fetal development has been suggested as the origin of reproductive disorders comprised within the testicular dysgenesis syndrome (TDS). This hypothesis has been supported by studies in rats demonstrating that normal male development and adult reproductive function depend on sufficient androgen exposure during a sensitive fetal period, called the masculinization programming window (MPW). The main aim of this study was therefore to examine the effects of manipulating androgen production during different timepoints during early human fetal testis development to identify the existence and timing of a possible window of androgen sensitivity resembling the MPW in rats. METHODS: The effects of experimentally reduced androgen exposure during different periods of human fetal testis development and function were examined using an established and validated human ex vivo tissue culture model. The androgen production was reduced by treatment with ketoconazole and validated by treatment with flutamide which blocks the androgen receptor. Testicular hormone production ex vivo was measured by liquid chromatography-tandem mass spectrometry or ELISA assays, and selected protein markers were assessed by immunohistochemistry. RESULTS: Ketoconazole reduced androgen production in testes from gestational weeks (GW) 7-21, which were subsequently divided into four age groups: GW 7-10, 10-12, 12-16 and 16-21. Additionally, reduced secretion of testicular hormones INSL3, AMH and Inhibin B was observed, but only in the age groups GW 7-10 and 10-12, while a decrease in the total density of germ cells and OCT4+ gonocytes was found in the GW 7-10 age group. Flutamide treatment in specimens aged GW 7-12 did not alter androgen production, but the secretion of INSL3, AMH and Inhibin B was reduced, and a reduced number of pre-spermatogonia was observed. CONCLUSIONS: This study showed that reduced androgen action during early development affects the function and density of several cell types in the human fetal testis, with similar effects observed after ketoconazole and flutamide treatment. The effects were only observed within the GW 7-14 period-thereby indicating the presence of a window of androgen sensitivity in the human fetal testis.

Safety, feasibility and initial efficacy of an app-facilitated telerehabilitation (AFTER) programme for COVID-19 survivors: a pilot randomised study.

OBJECTIVES: Determine the safety, feasibility and initial efficacy of a multicomponent telerehabilitation programme for COVID-19 survivors. DESIGN: Pilot randomised feasibility study. SETTING: In-home telerehabilitation. PARTICIPANTS: 44 participants (21 female, mean age 52 years) discharged home following hospitalisation with COVID-19 (with and without intensive care unit (ICU) stay). INTERVENTIONS: Participants were block randomised 2:1 to receive 12 individual biobehaviourally informed, app-facilitated, multicomponent telerehabilitation sessions with a licenced physical therapist (n=29) or to a control group (n=15) consisting of education on exercise and COVID-19 recovery trajectory, physical activity and vitals monitoring, and weekly check-ins with study staff. Interventions were 100% remote and occurred over 12 weeks. PRIMARY AND SECONDARY OUTCOME MEASURES: The primary outcome was feasibility, including safety and session adherence. Secondary outcomes included preliminary efficacy outcomes including tests of function and balance; patient-reported outcome measures; a cognitive assessment; and average daily step count. The 30 s chair stand test was the main secondary (efficacy) outcome. RESULTS: No adverse events (AEs) occurred during testing or in telerehabilitation sessions; 38% (11/29) of the intervention group compared with 60% (9/15) of the control group experienced an AE (p=0.21), most of which were minor, over the course of the 12-week study. 27 of 29 participants (93%; 95% CI 77% to 99%) receiving the intervention attended ≥75% of sessions. Both groups demonstrated clinically meaningful improvement in secondary outcomes with no statistically significant differences between groups. CONCLUSION: Fully remote telerehabilitation was safe, feasible, had high adherence for COVID-19 recovery, and may apply to other medically complex patients including those with barriers to access care. This pilot study was designed to evaluate feasibility; further efficacy evaluation is needed. TRIAL REGISTRATION NUMBER: NCT04663945.

Prospective telehealth analysis of functional performance, frailty, quality of life, and mental health after COVID-19 hospitalization.

BACKGROUND: COVID-19 is a global pandemic with poorly understood long-term consequences. Determining the trajectory of recovery following COVID-19 hospitalization is critical for prioritizing care, allocating resources, facilitating prognosis, and informing rehabilitation. The purpose of this study was to prospectively evaluate recovery following COVID-19 hospitalization. METHODS: Participants age 18 years or older who were hospitalized for ≥24 h due to COVID-19 completed phone/video call virtual assessments (including the 10-time chair rise test) and survey forms at three time points (2-6, 12, and 18 weeks) after hospital discharge. Univariate logistic and linear regression models assessed the associations of the outcomes with primary predictors (categorical age, sex, race/ethnicity group, and categorical pre-hospitalization frailty) at baseline; the same were used to assess differences in change from week 2-6 (continuous outcomes) or outcome persistence/worsening (categorical) at last contact. RESULTS: One hundred nine adults (age 53.0 [standard deviation 13.1]; 53% female) participated including 43 (39%) age 60 or greater; 59% identified as an ethnic and/or racial minority. Over 18 weeks, the mean time to complete the 10-time chair rise test decreased (i.e., improved) by 6.0 s (95% CI: 4.1, 7.9 s; p < 0.001); this change did not differ by pre-hospital frailty, race/ethnicity group, or sex, but those age ≥ 60 had greater improvement. At weeks 2-6, 67% of participants reported a worse Clinical Frailty Scale category compared to their pre-hospitalization level, whereas 42% reported a worse frailty score at 18 weeks. Participants who did not return to pre-hospitalization levels were more likely to be female, younger, and report a pre-hospitalization category of 'very fit' or 'well'. CONCLUSIONS: We found that functional performance improved from weeks 2-6 to 18 weeks of follow-up; that incident clinical frailty developed in some individuals following COVID-19; and that age, sex, race/ethnicity, and pre-hospitalization frailty status may impact recovery from COVID-19. Notably, individuals age 60 and older were more likely than those under age 45 years to return to their pre-hospitalization status and to make greater improvements in functional performance. The results of the present study provide insight into the trajectory of recovery among a representative cohort of individuals hospitalized due to COVID-19.

The effects of selected inhibitors on human fetal adrenal steroidogenesis differs under basal and ACTH-stimulated conditions.

BACKGROUND: Disordered fetal adrenal steroidogenesis can cause marked clinical effects including virilization of female fetuses. In postnatal life, adrenal disorders can be life-threatening due to the risk of adrenal crisis and must be carefully managed. However, testing explicit adrenal steroidogenic inhibitory effects of therapeutic drugs is challenging due to species-specific characteristics, and particularly the impact of adrenocorticotropic hormone (ACTH) stimulation on drugs targeting steroidogenesis has not previously been examined in human adrenal tissue. Therefore, this study aimed to examine the effects of selected steroidogenic inhibitors on human fetal adrenal (HFA) steroid hormone production under basal and ACTH-stimulated conditions. METHODS: This study used an established HFA ex vivo culture model to examine treatment effects in 78 adrenals from 50 human fetuses (gestational weeks 8-12). Inhibitors were selected to affect enzymes critical for different steps in classic adrenal steroidogenic pathways, including CYP17A1 (Abiraterone acetate), CYP11B1/2 (Osilodrostat), and a suggested CYP21A2 inhibitor (Efavirenz). Treatment effects were examined under basal and ACTH-stimulated conditions in tissue from the same fetus and determined by quantifying the secretion of adrenal steroids in the culture media using liquid chromatography-tandem mass spectrometry. Statistical analysis was performed on ln-transformed data using one-way ANOVA for repeated measures followed by Tukey's multiple comparisons test. RESULTS: Treatment with Abiraterone acetate and Osilodrostat resulted in potent inhibition of CYP17A1 and CYP11B1/2, respectively, while treatment with Efavirenz reduced testosterone secretion under basal conditions. ACTH-stimulation affected the inhibitory effects of all investigated drugs. Thus, treatment effects of Abiraterone acetate were more pronounced under stimulated conditions, while Efavirenz treatment caused a non-specific inhibition on steroidogenesis. ACTH-stimulation prevented the Osilodrostat-mediated CYP11B1 inhibition observed under basal conditions. CONCLUSIONS: Our results show that the effects of steroidogenic inhibitors differ under basal and ACTH-stimulated conditions in the HFA ex vivo culture model. This could suggest that in vivo effects of therapeutic drugs targeting steroidogenesis may vary in conditions where patients have suppressed or high ACTH levels, respectively. This study further demonstrates that ex vivo cultured HFAs can be used to evaluate steroidogenic inhibitors and thereby provide novel information about the local effects of existing and emerging drugs that targets steroidogenesis.

A 3D system to model human pancreas development and its reference single-cell transcriptome atlas identify signaling pathways required for progenitor expansion.

Human organogenesis remains relatively unexplored for ethical and practical reasons. Here, we report the establishment of a single-cell transcriptome atlas of the human fetal pancreas between 7 and 10 post-conceptional weeks of development. To interrogate cell-cell interactions, we describe InterCom, an R-Package we developed for identifying receptor-ligand pairs and their downstream effects. We further report the establishment of a human pancreas culture system starting from fetal tissue or human pluripotent stem cells, enabling the long-term maintenance of pancreas progenitors in a minimal, defined medium in three-dimensions. Benchmarking the cells produced in 2-dimensions and those expanded in 3-dimensions to fetal tissue identifies that progenitors expanded in 3-dimensions are transcriptionally closer to the fetal pancreas. We further demonstrate the potential of this system as a screening platform and identify the importance of the EGF and FGF pathways controlling human pancreas progenitor expansion.

Establishment of a Novel Human Fetal Adrenal Culture Model that Supports de Novo and Manipulated Steroidogenesis.

CONTEXT: Disorders affecting adrenal steroidogenesis promote an imbalance in the normally tightly controlled secretion of mineralocorticoids, glucocorticoids, and androgens. This may lead to differences/disorders of sex development in the fetus, as seen in virilized girls with congenital adrenal hyperplasia (CAH). Despite the important endocrine function of human fetal adrenals, neither normal nor dysregulated adrenal steroidogenesis is understood in detail. OBJECTIVE: Due to significant differences in adrenal steroidogenesis between human and model species (except higher primates), we aimed to establish a human fetal adrenal model that enables examination of both de novo and manipulated adrenal steroidogenesis. DESIGN AND SETTING: Human adrenal tissue from 54 1st trimester fetuses were cultured ex vivo as intact tissue fragments for 7 or 14 days. MAIN OUTCOME MEASURES: Model validation included examination of postculture tissue morphology, viability, apoptosis, and quantification of steroid hormones secreted to the culture media measured by liquid chromatography-tandem mass spectrometry. RESULTS: The culture approach maintained cell viability, preserved cell populations of all fetal adrenal zones, and recapitulated de novo adrenal steroidogenesis based on continued secretion of steroidogenic intermediates, glucocorticoids, and androgens. Adrenocorticotropic hormone and ketoconazole treatment of ex vivo cultured human fetal adrenal tissue resulted in the stimulation of steroidogenesis and inhibition of androgen secretion, respectively, demonstrating a treatment-specific response. CONCLUSIONS: Together, these data indicate that ex vivo culture of human fetal adrenal tissue constitutes a novel approach to investigate local effects of pharmaceutical exposures or emerging therapeutic options targeting imbalanced steroidogenesis in adrenal disorders, including CAH.

Secretion of parathyroid hormone may be coupled to insulin secretion in humans.

OBJECTIVE: Parathyroid hormone (PTH) is a key hormone in regulation of calcium homeostasis and its secretion is regulated by calcium. Secretion of PTH is attenuated during intake of nutrients, but the underlying mechanism(s) are unknown. We hypothesized that insulin acts as an acute regulator of PTH secretion. METHODS: Intact PTH was measured in plasma from patients with T1D and matched healthy individuals during 4-h oral glucose tolerance tests (OGTT) and isoglycemic i.v. glucose infusions on 2 separate days. In addition, expression of insulin receptors on surgical specimens of parathyroid glands was assessed by immunochemistry (IHC) and quantitative PCR (qPCR). RESULTS: The inhibition of PTH secretion was more pronounced in healthy individuals compared to patients with T1D during an OGTT (decrementalAUC0-240min: -5256 ± 3954 min × ng/L and -2408 ± 1435 min × ng/L, P = 0.030). Insulin levels correlated significantly and inversely with PTH levels, also after adjusting for levels of several gut hormones and BMI (P = 0.002). Expression of insulin receptors in human parathyroid glands was detected by both IHC and qPCR. CONCLUSION: Our study suggests that insulin may act as an acute regulator of PTH secretion in humans.

Endometrial scratch injury with office hysteroscopy before IVF/ICSI: A randomised controlled trial.

OBJECTIVE: Endometrial scratch injury (ESI) has been proposed to improve endometrial receptivity and thereby increase implantation rates in assisted reproductive technology (ART) treatment. ESI has been widely incorporated into clinical practice despite inconclusive evidence of its effect on reproductive outcomes. We aimed to assess pregnancy and live birth rates in subfertile women receiving ESI before IVF treatment in comparison to controls. STUDY DESIGN: This was a randomised controlled trial (RCT) with no blinding of participants, investigators or health care personnel. Women in ART treatment were allocated to either office hysteroscopy with ESI (ESI group) or no intervention (control group). In total 184 women in IVF/ICSI treatment with minimum one previous failed IVF/ICSI cycle, were included in the final analysis. The primary outcome was positive serum hCG (s-hCG). Secondary outcomes were ongoing pregnancy and live birth rate. Only per-protocol analyses were performed as all patients included at one centre had to be excluded. The trial is registered at ClinicalTrials.gov, NCT01743391. RESULTS: Our results showed a non-significant increase in positive s-hCG (OR 1.23, 95 % CI (0.65-2.33)), ongoing pregnancy (OR 1.52, 95 % CI (0.73-3.17)), and live birth rates (OR 1.69, 95 % CI (0.78-3.64)) per randomised woman between the ESI and the control group. CONCLUSION: We observed no significant differences in positive s-hCG or other reproductive outcomes in the ESI vs. the control group. While the crude estimates of positive reproductive outcomes were higher in the ESI group, statistical significance was not reached, and the study was not powered to show smaller differences. However, data from this study will be re-evaluated in the context of an individual participant data meta-analysis (IPD-MA) of RCTs on ESI.

Tracing the origin of adult intestinal stem cells.

Adult intestinal stem cells are located at the bottom of crypts of Lieberkühn, where they express markers such as LGR51,2 and fuel the constant replenishment of the intestinal epithelium1. Although fetal LGR5-expressing cells can give rise to adult intestinal stem cells3,4, it remains unclear whether this population in the patterned epithelium represents unique intestinal stem-cell precursors. Here we show, using unbiased quantitative lineage-tracing approaches, biophysical modelling and intestinal transplantation, that all cells of the mouse intestinal epithelium-irrespective of their location and pattern of LGR5 expression in the fetal gut tube-contribute actively to the adult intestinal stem cell pool. Using 3D imaging, we find that during fetal development the villus undergoes gross remodelling and fission. This brings epithelial cells from the non-proliferative villus into the proliferative intervillus region, which enables them to contribute to the adult stem-cell niche. Our results demonstrate that large-scale remodelling of the intestinal wall and cell-fate specification are closely linked. Moreover, these findings provide a direct link between the observed plasticity and cellular reprogramming of differentiating cells in adult tissues following damage5-9, revealing that stem-cell identity is an induced rather than a hardwired property.

Targeted Rhesus immunoglobulin for RhD-negative women undergoing an induced abortion: A clinical pilot study.

INTRODUCTION: The aim of this clinical pilot study was to examine the accuracy of noninvasive fetal RHD genotyping in early pregnancy (8+0  to 11+6  weeks) and to clarify whether targeted administration of Rhesus immunoglobulin (RhIg) is possible for women undergoing an induced abortion such that unnecessary injections can be avoided. The study examines the correlation between gestational age and the amount of cell-free fetal DNA in maternal plasma, the fetal fraction of DNA and whether transportation time or body mass index affects these parameters. MATERIAL AND METHODS: Fifty-two RhD-negative women undergoing a surgically induced abortion were included. A maternal blood sample was collected prior to the abortion and a tissue sample was collected from the placental part of the abortion material after the intervention. Fetal RhD type was determined by PCR analysis of cell-free fetal DNA extracted from maternal plasma and on DNA from the tissue sample, with the latter providing a reference standard. Copies of RHD/mL were determined on RHD-positive samples and the fetal fraction of DNA was calculated. RESULTS: We demonstrated complete concordance between results from plasma and tissue, with 31 RhD-positive and 21 RhD-negative samples, corresponding to 40% being RhD-negative, specificity 100% [95% confidence interval (CI) 88.8-100] and sensitivity 100% (95% CI 83.9-100). We found no significant correlation between gestational age and the amount or the fraction of cell-free fetal DNA in maternal plasma, nor did we find that transportation time or BMI significantly affected these factors in this setup. CONCLUSIONS: Fetal RHD genotyping can be accurately performed from the 8th week of gestation and unnecessary injections of RhIg can be avoided for women undergoing an induced abortion. A larger study is needed to determine a more accurate sensitivity for the analysis early in pregnancy.

Characterization of Human Adrenal Steroidogenesis During Fetal Development.

CONTEXT: The endocrine function of human fetal adrenals (HFAs) is activated already during first trimester, but adrenal steroidogenesis during fetal life is not well characterized. OBJECTIVE: This study aimed to investigate HFA steroidogenesis by analyzing adrenal glands from first and second trimesters. DESIGN AND SETTING: Male and female HFA from gestational weeks (GWs) 8 to 19 were examined, including a total of 101 samples from 83 fetuses. MAIN OUTCOME MEASURE(S): Expression level of steroidogenic genes and protein expression/localization were determined by quantitative PCR and immunohistochemistry, respectively, and intra-adrenal steroid levels were quantified by LC-MS/MS. RESULTS: Transcriptional levels of StAR, CYP11A1, CYP17A1, CYP21A2, CYP11B1/2, and SULT2A1 were significantly higher in second trimester compared to first trimester (P < 0.05), whereas expression levels of 3ß-HSD2 and ARK1C3 were unaltered between GWs 8 and 19. All investigated steroidogenic proteins were expressed in a distinct pattern throughout the investigated period, with most enzymes expressed primarily in the fetal zone, except 3ß-HSD1/2, which was expressed mainly in the definitive zone. Abundant steroidogenic enzyme expression was reflected in overall high intra-adrenal tissue concentrations of mineralocorticoids, glucocorticoids, and androgens; cortisol was the most abundant (1071 to 2723 ng/g tissue), and testosterone levels were the lowest (2 to 14 ng/g tissue). CONCLUSIONS: The expression profiles of HFA steroidogenic enzymes are distinct from first to second trimester, with no major differences between male and female samples. Intra-adrenal steroid hormone concentrations confirm that cortisol is produced throughout first and second trimesters, suggesting continued regulation of the hypothalamus-pituitary-adrenal axis during this entire period.

Nodal Signaling Regulates Germ Cell Development and Establishment of Seminiferous Cords in the Human Fetal Testis.

Disruption of human fetal testis development is widely accepted to underlie testicular germ cell cancer (TGCC) origin and additional disorders within testicular dysgenesis syndrome (TDS). However, the mechanisms for the development of testicular dysgenesis in humans are unclear. We used ex vivo culture and xenograft approaches to investigate the importance of Nodal and Activin signaling in human fetal testis development. Inhibition of Nodal, and to some extent Activin, signaling disrupted seminiferous cord formation, abolished AMH expression, reduced androgen secretion, and decreased gonocyte numbers. Subsequent xenografting of testicular tissue rescued the disruptive effects on seminiferous cords and somatic cells but not germ cell effects. Stimulation of Nodal signaling increased the number of germ cells expressing pluripotency factors, and these persisted after xenografting. Our findings suggest a key role for Nodal signaling in the regulation of gonocyte differentiation and early human testis development with implications for the understanding of TGCC and TDS origin.

Determinants of Fasting Hyperglucagonemia in Patients with Type 2 Diabetes and Nondiabetic Control Subjects.

BACKGROUND: Fasting hyperglucagonemia can be detrimental to glucose metabolism in patients with type 2 diabetes (T2D) and may contribute to metabolic disturbances in obese and/or prediabetic subjects. However, the mechanisms underlying fasting hyperglucagonemia remain elusive. METHODS: We evaluated the interrelationship between fasting hyperglucagonemia and demographic and biochemical parameters in 106 patients with T2D (31% female, age: 57 ± 9 years [mean ± standard deviation; body mass index (BMI): 30.1 ± 4.4 kg/m2; fasting plasma glucose (FPG): 9.61 ± 2.39 mM; hemoglobin A1c (HbA1c): 57.1 ± 13.1 mmol/mol] and 163 nondiabetic control subjects (29% female; age: 45 ± 17 years; BMI: 25.8 ± 4.1 kg/m2; FPG: 5.2 ± 0.4 mM; and HbA1c: 35.4 ± 3.8 mmol/mol). Multiple linear regression analysis was applied using a stepwise approach with fasting plasma glucagon as dependent parameter and BMI, waist-to-hip ratio (WHR), blood pressure, hemoglobin A1c, FPG, and insulin concentrations as independent parameters. RESULTS: Fasting plasma glucagon concentrations were significantly higher among patients with T2D (13.5 ± 6.3 vs. 8.5 ± 3.8 mM, P < 0.001) together with HbA1c (P < 0.001), FPG (P < 0.001), and insulin (84.9 ± 56.4 vs. 57.7 ± 35.3 mM, P < 0.001). When adjusted for T2D, HbA1c and insulin were significantly positive determinants for fasting plasma glucagon concentrations. Furthermore, WHR comprised a significant positive determinant. CONCLUSIONS: We confirm that fasting plasma glucagon concentrations are abnormally high in patients with T2D, and show that fasting plasma glucagon concentrations are influenced by WHR (in addition to glycemic control and fasting plasma insulin concentrations), which may point to visceral fat deposition as an important determinant of increased fasting plasma glucagon concentrations.

[Myoma with an intra-abdominal bleeding].

I this case report a 37-year-old nulliparous woman was admitted for acute onset of abdominal pain. CT scan showed a homogeneous tumour related to the internal genitalia and extravasation of contrast material, but the site of bleeding was not identifiable. Ultrasonography revealed leiomyoma and haemoperitoneum, and emergency laparoscopy was performed. There was an ongoing venous bleeding from two subserosal myomas. Myomectomy was done, and 588 g tissue was removed with a benign histology. Spontaneous rupture of a vessel overlying a uterine myoma has been documented in the literature, though it is extremely rare.

Sharp compared with blunt fascial incision at cesarean delivery: a randomized controlled trial with each case as her own control.

OBJECTIVE: To compare patient preference for either sharp incision with scissors or blunt manual cleavage of the fascia at cesarean delivery in a randomized controlled trial in which each woman was her own control. STUDY DESIGN: Women undergoing primary cesarean delivery (n=34) were randomized to side distribution of sharp or blunt incision of the fascia (sharp right and blunt left or blunt right and sharp left) and followed three months postoperatively. The primary outcome was patient preference for the right or left side of the scar 3 months postoperatively and modeled by polytomous logistic regression. The secondary outcome was difference in pain between the two sides measured on a 0.0-10.0 numerical rating scale at 1, 3, and 7 days and 1 and 3 months postoperatively. Pain scores were analyzed with a Wilcoxon signed rank test. RESULTS: 28 cases were analyzed and no significant difference was found in preference after three months. Nine women preferred the sharp (32%, 95% CI 16-52%) and 7 the blunt side (25%, 95% CI 11-45%) (P=0.804). Pain scores did not differ significantly between the two sides at any time postoperatively either at rest or during mobilization. CONCLUSION: No significant difference was found in patient preference with regard to sharp or blunt incision of the fascia, nor was there a significant difference in postoperative pain scores. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov: www.clinicaltrials.org;NCT01297725.

Postprandial gut hormone responses and glucose metabolism in cholecystectomized patients.

Preclinical studies suggest that gallbladder emptying, via bile acid-induced activation of the G protein-coupled receptor TGR5 in intestinal L cells, may play a significant role in the secretion of the incretin hormone glucagon-like peptide-1 (GLP-1) and, hence, postprandial glucose homeostasis. We examined the secretion of gut hormones in cholecystectomized subjects to test the hypothesis that gallbladder emptying potentiates postprandial release of GLP-1. Ten cholecystectomized subjects and 10 healthy, age-, gender-, and body mass index-matched control subjects received a standardized fat-rich liquid meal (2,200 kJ). Basal and postprandial plasma concentrations of glucose, insulin, C-peptide, glucagon, GLP-1, glucose-dependent insulinotropic polypeptide (GIP), glucagon-like peptide-2 (GLP-2), cholecystokinin (CCK), and gastrin were measured. Furthermore, gastric emptying and duodenal and serum bile acids were measured. We found similar basal glucose concentrations in the two groups, whereas cholecystectomized subjects had elevated postprandial glucose excursions. Cholecystectomized subjects had reduced postprandial concentrations of duodenal bile acids, but preserved postprandial plasma GLP-1 responses, compared with control subjects. Also, cholecystectomized patients exhibited augmented fasting glucagon. Basal plasma CCK concentrations were lower and peak concentrations were higher in cholecystectomized patients. The concentrations of GIP, GLP-2, and gastrin were similar in the two groups. In conclusion, cholecystectomized subjects had preserved postprandial GLP-1 responses in spite of decreased duodenal bile delivery, suggesting that gallbladder emptying is not a prerequisite for GLP-1 release. Cholecystectomized patients demonstrated a slight deterioration of postprandial glycemic control, probably because of metabolic changes unrelated to incretin secretion.

Glucagon-like peptide-1 (GLP-1) receptor agonism or DPP-4 inhibition does not accelerate neoplasia in carcinogen treated mice.

INTRODUCTION: Glucagon-like peptide-1 (GLP-1) and glucagon-like peptide-2 (GLP-2) are secreted in parallel from the intestinal endocrine cells after nutrient intake. GLP-1 is an incretin hormone and analogues are available for the treatment of type 2 diabetes mellitus (T2DM). GLP-2 is an intestinal growth hormone and is shown to promote growth of colonic adenomas in carcinogen treated mice. Both peptides are degraded by dipeptidyl peptidase-4 (DPP-4) into inactive metabolites. DPP-4 inhibitors are therefore also in use for treatment of T2DM. It is possible that DPP-4 inhibition by enhancing the exposure of endogenous GLP-2 to the intestinal epithelia also might mediate growth and promote neoplasia. We investigated the intestinal growth effect of the GLP-1 receptor agonists (GLP-1 RAs) (liraglutide and exenatide) and DPP-4 inhibition (sitagliptin) in healthy mice. We also investigated the potential tumour promoting effect of liraglutide and sitaglitin in the colon of carcinogen treated mice. We used GLP-2 as a positive control. METHODS: For the growth study we treated healthy CD1 mice with liraglutide (300 µg×2), exenatide (12.5 µg×2) or vehicle subcutaneously and sitagliptin (8mg×2) or water by oral gavage for 10 or 30 days. We measured intestinal weight, cross sectional area, villus height and crypt depth. For the tumour study we treated carcinogen treated mice (1,2 dimethylhydrazine 21 mg/kg/week for 12 weeks) with liraglutide (300 µg×2), Gly2-GLP-2 (25 µg×2) or vehicle subcutaneously and sitagliptin (8 mg×2) or water by oral gavage for 45 days. We counted aberrant crypt foci (ACF), mucin depleted foci (MDF) and adenomas in the colon. Using COS-7 cells transfected with a GLP-2 receptor, we tested if liraglutide or exenatide could activate the receptor. RESULTS: In the 10 days experiment the relative small intestinal weight was increased with 56% in the liraglutide group (p<0.001) and 26% in the exenatide group (p<01) compared with vehicle treated mice. After 30 days of treatment, liraglutide did also increase the colonic weight (p<0.01). By morphometry the growth pattern mimicked that of GLP-2. Sitagliptin treatment had only a minor effect. In the carcinogen treated mice we found no increase of ACF in any of the groups, the numbers of MDF and adenomas after liraglutide and sitagliptin treatments were similar to their respective control groups. Neither liraglutide nor exenatide stimulated cAMP release from GLP-2 receptor transfected cells. CONCLUSION: Both GLP-1 analogues were potent growth stimulators of the healthy mouse intestine. No agonism was found for GLP-1 RAs at the GLP-2 receptor. Despite of the growth effect, liraglutide did not promote dysplasia in the colon. Sitagliptin did not show any tumour promoting effects, and non considerable growth effects.