Mental health impact of multiple sexually minoritized and gender expansive stressors among LGBTQ+ young adults: a latent class analysis.

AIMS: In the United States, lesbian, gay, bisexual, transgender, queer, intersex, asexual and other sexually minoritized and gender expansive (LGBTQ+) young adults are at increased risk for experiencing mental health inequities, including anxiety, depression and psychological distress-related challenges associated with their sexual and gender identities. LGBTQ+ young adults may have unique experiences of sexual and gender minority-related vulnerability because of LGBTQ+-related minority stress and stressors, such as heterosexism, family rejection, identity concealment and internalized homophobia. Identifying and understanding specific LGBTQ+-related minority stress experiences and their complex roles in contributing to mental health burden among LGBTQ+ young adults could inform public health efforts to eliminate mental health inequities experienced by LGBTQ+ young adults. Therefore, this study sought to form empirically based risk profiles (i.e., latent classes) of LGBTQ+ young adults based on their experiences with familial heterosexist experiences, LGBTQ+-related family rejection, internalized LGBTQ+-phobia and LGBTQ+ identity concealment, and then identify associations of derived classes with psychological distress. METHODS: We recruited and enrolled participants using nonprobability, cross-sectional online survey data collected between May and August 2020 (N = 482). We used a three-step latent class analysis (LCA) approach to identify unique classes of response patterns to LGBTQ+-related minority stressor subscale items (i.e., familial heterosexist experiences, LGBTQ+-related family rejection, internalized LGBTQ+-phobia and LGBTQ+ identity concealment), and multinomial logistic regression to characterize the associations between the derived classes and psychological distress. RESULTS: Five distinct latent classes emerged from the LCA: (1) low minority stress, (2) LGBTQ+ identity concealment, (3) family rejection, (4) moderate minority stress and (5) high minority stress. Participants who were classified in the high and moderate minority stress classes were more likely to suffer from moderate and severe psychological distress compared to those classified in the low minority stress class. Additionally, relative to those in the low minority stress class, participants who were classified in the LGBTQ+ identity concealment group were more likely to suffer from severe psychological distress. CONCLUSION: Familial heterosexist experiences, LGBTQ+-related family rejection, internalized LGBTQ+-phobia and LGBTQ+ identity concealment are four constructs that have been extensively examined as predictors for mental health outcomes among LGBTQ+ persons, and our study is among the first to reveal nuanced gradients of these stressors. Additionally, we found that more severe endorsement of minority stress was associated with greater psychological distress. Given our study results and the previously established negative mental health impacts of minority stressors among LGBTQ+ young adults, findings from our study can inform research, practice, and policy reform and development that could prevent and reduce mental health inequities among LGBTQ+ young adults.

Optimisation of geometric aspect ratio of thin film transistors for low-cost flexible CMOS inverters and its practical implementation.

A low-cost, flexible processor is essential to realise affordable flexible electronic systems and transform everyday objects into smart-objects. Thin film transistors (TFTs) based on metal-oxides (or organics) are ideal candidates as they can be manufactured at low processing temperatures and low-cost per unit area, unlike traditional silicon devices. The development of complementary metal-oxide-semiconductor (CMOS) technology based on these materials remains challenging due to differences in performance between n- and p-type TFTs. Existing geometric rules typically compensate the lower mobility of the metal-oxide p-type TFT by scaling up the width-to-length (W/L) ratio but fail to take into account the significant off-state leakage current. Here we propose the concept of an optimal geometric aspect ratio which maximises the inverter efficiency represented by the average switching current divided by the static currents. This universal method is especially useful for the design of low-power CMOS inverters based on metal-oxides, where the large off-current of the p-type TFT dominates the static power consumption of the inverter. We model the inverter efficiency and noise margins of metal-oxide CMOS inverters with different geometric aspect ratios and compare the performance to different inverter configurations. The modelling results are verified experimentally by fabricating CMOS inverter configurations consisting of n-type indium-silicon-oxide (ISO) TFTs and p-type tin monoxide (SnO) TFTs. Notably, our results show that reducing W/L of metal-oxide p-type TFTs increases the inverter efficiency while reducing the area compared to simply scaling up W/L inversely with mobility. We anticipate this work provides a straightforward method to geometrically optimise flexible CMOS inverters, which will remain relevant even as the performance of TFTs continues to evolve.

Diagnosis and treatment of confirmed and suspected primary hyperparathyroidism in equids: 17 cases (1999-2016).

BACKGROUND: Primary hyperparathyroidism is uncommon in equids. OBJECTIVES: To describe the diagnostic findings and efficacy of treatment in equids with primary hyperparathyroidism. STUDY DESIGN: Retrospective case series describing 16 horses and one mule. METHODS: Cases were identified by retrospective review of records at Cornell University and via an ACVIM listserv query. Inclusion criteria were an equid with hypercalcemia, normal renal function and high parathyroid hormone (PTH) or histopathological diagnosis of a parathyroid adenoma. Equids with normal PTH and PTH-related protein (PTHrP) in the face of hypercalcemia were included as suspect cases. RESULTS: The most common presenting complaints were weight loss (12/17) and hypercalcemia (10/17). PTH was above reference range in 12/17 cases. Suspected parathyroid tumours were localised in 12/14 equids imaged using ultrasonography alone (2/3), technetium 99m Tc sestamibi scintigraphy alone (1/1) or both modalities (9/10). Three horses did not have imaging performed. Surgical exploration successfully excised tumours in six of 10 cases. Five were located at the thoracic inlet, and surgery resulted in complete cure. One tumour was excised from the thyroid lobe, and the horse remained hypercalcemic. Four other cases explored surgically, four treated medically and three that were not treated also remained hypercalcemic. MAIN LIMITATIONS: The small study size prohibited statistical analysis. CONCLUSIONS: Parathyroid adenomas in equids can be successfully localised with ultrasonography and scintigraphy. Surgical excision appears more likely to be successful for single gland disease at the thoracic inlet.

Response of hen pre-recruitment ovarian follicles to follicle stimulating hormone, in vivo.

In laying hens, pre-recruitment ovarian follicles (1-8 mm diameter) are arranged as a continuum of size and predicted maturity. Cyclic recruitment of a pre-recruitment follicle to the preovulatory stage begins, in part, by the ability of the granulosa cell (GC) layer to initiate responsiveness to follicle stimulating hormone- (FSH-) induced cyclic adenosine monophosphate. The objective of this study was to determine if increased circulating concentrations of FSH during the ovulatory cycle increase the number of recruited follicles, in a dose-dependent manner. Equine chorionic gonadotropin (eCG) was initially tested due to its FSH-like properties and long half-life. Laying hens were injected, i.m., with 0 or 100 IU eCG, and ovaries were collected 29 h later. Recruited follicles were initially identified based on incorporation of yellow yolk and a weight of 250-900 mg. Recruitment was subsequently confirmed by both incubating the GC layer for 3 h with recombinant human (rh) FSH to establish FSH-responsiveness and quantifying P450 side-chain cleavage enzyme (CYP11A1) mRNA. Additional hens were injected with 0, 30, 75, and 300 IU eCG to establish a dose-response. Because eCG exhibits some luteinizing hormone activity, FSH-induced recruitment was evaluated by injecting 0.1, 0.33, 0.66, 1 or 3.3 µg rhFSH. Ovaries were collected 29 h post-injection, and expression of CYP11A1 mRNA was quantitated in GCs from recruited and pre-recruitment follicles. One hundred IU eCG induced recruitment of 2-8 follicles compared to a single follicle in control hens. In contrast to pre-recruitment follicles, incubated GC from eCG-recruited follicles had initiated differentiation, indicated by increased CYP11A1 and rhFSH-induced STAR mRNA and progesterone. Equine CG and rhFSH each increased the number of recruited follicles in a dose-dependent manner. Further, CYP11A1 mRNA was significantly increased in GC layers from recruited, compared to non-recruited, follicles. We conclude that FSH-responsiveness within the GC layer of each pre-recruitment follicle increases with follicle size, and propose that this establishes the order of daily follicle recruitment.

Follicle dynamics and granulosa cell differentiation in the turkey hen ovary.

Similar to the domestic hen ovary, entry of a follicle into the preovulatory hierarchy in the turkey hen represents a process in which a single follicle initiates rapid growth and final maturation prior to ovulation. Published data derived from the laying hen support the proposal that differentiation of the follicle granulosa cell (GC) layer begins coincident with entry into the rapid growth phase and is characterized by the initial capacity for follicle stimulating hormone (FSH)-mediated cell signaling. The present studies were conducted with photostimulated B.U.T. Big 6 turkey hens to compare follicle dynamics and cellular mechanisms to those in the laying hen. The measurement and weights of turkey ovarian follicles greater than 1 mm in diameter revealed a discrete size hierarchy that was maintained throughout follicle development. GC layers collected from the single follicle initiating rapid growth (at the 11 to 13 mm stage of development) and incubated, in vitro, for 3 h with recombinant human (rh) FSH (10 ng/mL) responded with significantly increased steroidogenic acute regulatory protein (STAR) mRNA expression and progesterone production. The same treatment induced minimal STAR expression and no significant progesterone accumulation in GCs from 8 to 9 mm follicles (prior to the rapid growth phase). By comparison, dispersed GCs from 8 to 9 mm follicles pre-cultured for 18 h followed by a 3 h challenge with rhFSH resulted in significantly increased STAR expression plus progesterone production. Significantly, such cultured GCs pretreated for 15 min with transforming growth factor alpha (TGFα; 10 ng/mL) completely prevented both rhFSH-induced STAR expression and progesterone production. Culture of GCs from 8 to 9 mm follicles for 21 h with Bone Morphogenetic Protein 6 (BMP6) increased both cholesterol side-chain cleavage enzyme (CYP11A1) and FSH receptor mRNA (FSHR) expression. BMP6 also enhanced rhFSH-induced STAR expression, and this effect was blocked by TGFα. Collectively, these results support a conservation of mechanisms that maintain a hierarchy of follicles throughout development plus initiate FSH-responsiveness and GC differentiation as the recruited follicle enters the rapid growth phase in these closely related species.

Evaluation of digital cryotherapy using a commercially available sleeve style ice boot in healthy horses and horses receiving i.v. endotoxin.

BACKGROUND: Continuous digital cryotherapy experimentally prevents development and reduces severity of sepsis-associated laminitis. A sleeve style ice boot where ice is in direct contact with the skin, and water drains from the boot is being used clinically for distal limb cryotherapy. The degree of cooling achieved by this boot is unknown. OBJECTIVES: Evaluate skin and lamellar cooling after application of the ice sleeve in healthy horses, and the same horses during an endotoxaemia model. STUDY DESIGN: Prospective study, crossover design. METHODS: In eight healthy horses thermocouples were inserted into dorsal lamellae of both front feet, and under skin on both metacarpi. One forelimb received cryotherapy using sleeve style ice boot, with contralateral limb as control. Temperature was recorded on data logging devices at 5 min intervals during each cryotherapy session. Day 1: temperature data was collected for healthy horses. Day 2: data was collected for the same horses during i.v. administration of endotoxin. RESULTS: In healthy and endotoxaemic horses, the sleeve style ice boot significantly decreased mean skin (7.2°C and 5.8°C respectively) and lamellar (10.8°C and 9.6°C respectively) temperatures compared with control limbs (P<0.001). Skin and lamellar temperatures in endotoxaemic horses undergoing cryotherapy were significantly colder than in healthy horses (P = 0.01). MAIN LIMITATIONS: Order of treatment not randomised. CONCLUSIONS: The boot caused significant decreases in lamellar temperatures compared with untreated control limbs in all horses. Endotoxaemic horses had significantly colder lamellae and skin than healthy horses. This study is the first to show that a sleeve style boot, where ice does not cover the hoof, can cause significant decreases in lamellar temperatures through cooling of blood as it travels to the foot.

Borrelia burgdorferi Infection and Lyme Disease in North American Horses: A Consensus Statement.

Borrelia burgdorferi infection is common in horses living in Lyme endemic areas and the geographic range for exposure is increasing. Morbidity after B. burgdorferi infection in horses is unknown. Documented, naturally occurring syndromes attributed to B. burgdorferi infection in horses include neuroborreliosis, uveitis, and cutaneous pseudolymphoma. Although other clinical signs such as lameness and stiffness are reported in horses, these are often not well documented. Diagnosis of Lyme disease is based on exposure to B. burgdorferi, cytology or histopathology of infected fluid or tissue and antigen detection. Treatment of Lyme disease in horses is similar to treatment of humans or small animals but treatment success might not be the same because of species differences in antimicrobial bioavailability and duration of infection before initiation of treatment. There are no approved equine label Lyme vaccines but there is strong evidence that proper vaccination could prevent infection in horses.

Pneumatosis intestinalis in an SIV-infected rhesus macaque (Macaca mulatta).

Pneumatosis intestinalis (PI) has been described as an incidental finding in domestic animals and humans where it is associated with human immunodeficiency virus infection among other comorbidities. This report describes emphysematous changes consistent with PI in a simian immunodeficiency virus (SIV)-infected rhesus macaque (Macaca mulatta).

Parelaphostrongylus tenuis Cerebrospinal Nematodiasis in a Horse with Cervical Scoliosis and Meningomyelitis.

There are reports of horses with acute onset acquired cervical scoliosis and cutaneous analgesia. The underlying dorsal gray column myelitis that produces these neurologic signs has been only presumptively attributed to migration of Parelaphostrongylus tenuis within the spinal cord. Despite previous confirmation brain by polymerase chain reaction testing, of P. tenuis within the brain of horses by polymerase chain reaction testing, genetic testing has failed to definitively identify the presence of this parasite in cases of equine myelitis. This case report provides molecular confirmation via polymerase chain reaction of P. tenuis within the cervical spinal cord of a horse with scoliosis and cutaneous analgesia.

Fatal pulmonary acariasis in an aged indoor rhesus macaque (Macaca mulatta).

Pulmonary acariasis is a sporadic, incidental finding in colony-raised rhesus macaques (Macaca mulatta). Prophylactic treatment in indoor-raised and indoor-housed macaques is not routine due to low prevalence, lack of clinical significance, and potential risk of toxicosis. This case is an unusually severe infestation of Pneumonyssus simicola in an indoor-housed rhesus macaque, which ultimately resulted in this animal's death.

Third-Degree Atrioventricular Block and Collapse Associated with Eosinophilic Myocarditis in a Horse.

Third-degree atrioventricular block (AVB) and primary inflammatory myocarditis are uncommon findings in horses. The horse of this report presented for collapse at rest and was found to have multiple cardiac arrhythmias, most notably 3rd-degree AVB. The horse was subsequently diagnosed with eosinophilic myocarditis on necropsy, a rare form of myocarditis not previously reported in horses. Despite extensive testing, an etiologic agent could not be identified, illustrating the difficulty in identifying a specific cause of myocarditis in horses.

Presumptive Iatrogenic Microcystin-Associated Liver Failure and Encephalopathy in a Holsteiner Gelding.

An 8-year-old Holsteiner gelding was presented for evaluation of anorexia, obtundation, icterus, and mild colic signs of 48 hours duration. History, physical examination, and initial diagnostics were suggestive of hepatic disease and encephalopathy. Microcystin toxicosis was suspected based on historical administration of a cyanobacteria supplement, associated serum biochemistry abnormalities, and characteristic histopathological changes. Microcystin contamination was confirmed in both supplement containers fed to the horse. Fulminant hepatic failure and encephalopathy progressed resulting in euthanasia. Necropsy findings were consistent with microcystin induced liver failure.

Retrospective Evaluation of Horses Diagnosed with Neuroborreliosis on Postmortem Examination: 16 Cases (2004-2015).

BACKGROUND: Equine neuroborreliosis (NB), Lyme disease, is difficult to diagnose and has limited description in the literature. OBJECTIVE: Provide a detailed description of clinical signs, diagnostic, and pathologic findings of horses with NB. ANIMALS: Sixteen horses with histologically confirmed NB. METHODS: Retrospective review of medical records at the University of Pennsylvania and via an ACVIM listserv query with inclusion criteria requiring possible exposure to Borrelia burgdorferi and histologic findings consistent with previous reports of NB without evidence of other disease. RESULTS: Sixteen horses were identified, 12 of which had additional evidence of NB. Clinical signs were variable including muscle atrophy or weight loss (12), cranial nerve deficits (11), ataxia (10), changes in behavior (9), dysphagia (7), fasciculations (6), neck stiffness (6), episodic respiratory distress (5), uveitis (5), fever (2), joint effusion (2), and cardiac arrhythmias (1). Serologic analysis was positive for B. burgdorferi infection in 6/13 cases tested. CSF abnormalities were present in 8/13 cases tested, including xanthochromia (4/13), increased total protein (5/13; median: 91 mg/dL, range: 25-219 mg/dL), and a neutrophilic (6/13) or lymphocytic (2/13) pleocytosis (median: 25 nucleated cells/µL, range: 0-922 nucleated cells/µL). PCR on CSF for B. burgdorferi was negative in the 7 cases that were tested. CONCLUSION AND CLINICAL IMPORTANCE: Diagnosis of equine NB is challenging due to variable clinical presentation and lack of sensitive and specific diagnostic tests. Negative serology and normal CSF analysis do not exclude the diagnosis of NB.

Accuracy of a Mouse Bioassay for the Diagnosis of Botulism in Horses.

BACKGROUND: The laboratory diagnosis of botulism in horses traditionally has relied upon the mouse bioassay (MBA). The accuracy of this test for the diagnosis of botulism in horses is unknown. HYPOTHESIS/OBJECTIVES: Our goal was to determine the sensitivity, specificity, positive predictive value, and negative predictive value of the MBA on laboratory-processed fecal and gastrointestinal samples for foals and adult horses. ANIMALS: Cases included all horses with a final clinical diagnosis of botulism that were admitted between 1986 and 2011 and had MBA testing performed. Controls included horses without botulism that were admitted during the same time period and had MBA testing performed. METHODS: Retrospective study. Horses suspected of having botulism had fecal or (less commonly) gastrointestinal content samples tested using MBA. For every hospitalized botulism suspect, control samples were obtained from ≥1 additional hospitalized horses not suspected to have botulism. RESULTS: One hundred and twenty-nine adult horses and 253 adult controls were identified. Overall sensitivity of the MBA was only 32% but specificity was 97%. Forty-three foal cases and 21 foal controls were evaluated; sensitivity of the MBA was 53% and specificity was 100%. Positive predictive value was substantially higher (100% for foals and 89% for adults) than negative predictive value (51% for foals and 67% for adults). CONCLUSIONS AND CLINICAL IMPORTANCE: Mouse bioassay has low sensitivity but high specificity for the diagnosis of botulism in horses. Positive results are highly suggestive of botulism but negative results do not exclude the diagnosis. Unaffected horses and foals rarely shed C. botulinum in their feces.

Transfusion in adults: 10-year survival of red cell, plasma and platelet recipients following transfusion.

OBJECTIVE: To determine the long-term survival of adult recipients (>16 years) transfused with red blood cells (RBC), platelets (PLT) and fresh frozen plasma (FFP) in England and Wales. STUDY DESIGN AND METHODS: The EASTR study (Epidemiology and Survival of Transfusion Recipients) was a national multi-centre epidemiological study with cross-sectional sampling from 29 representative hospitals in England supplied by NHS Blood and Transplant (NHSBT). Three separate groups of RBC (n = 9142), FFP (n = 4232) and PLT (3584) recipients were sampled over 1 year (1 October 2001-30 September 2002), with prospective survival monitoring for 10 years. This study presents the data for adult recipients (>16 years of age). RESULTS: The median age interquartile range (IQR) of adult transfusion recipients was RBC 70 (54-79), FFP 66 (51-76), PLT 62 (48-72). The 10-year survival for adult RBC, FFP and PLT recipients was highest for RBC recipients at 36% confidence interval (CI 35-37%, n = 8675), compared with 30% for both FFP (CI 29-32%, n = 3849) and PLT (CI 28-30%, n = 3110) recipients. In all groups, post-transfusion survival decreased with age, and a risk-adjusted analysis showed that reason for transfusion, transfusion type (surgical or medical) and cancer diagnosis (presence or absence) were all significantly associated with survival. Older patients with cancer receiving a medical rather than surgical transfusion had the highest hazard of death. CONCLUSION: This study shows that survival following transfusion in England is broadly similar to that reported in other wealthy nations. More than 70% of recipients die within 10 years of transfusion, but long-term survival is common in younger patients (>80% 10-year survival in RBC recipients aged 16-39 years).

Vascular endothelial growth factor and angiopoietins during hen ovarian follicle development.

Growth and maturation of ovarian follicles in the hen (Gallus gallus) requires a network of blood vessels that increases in complexity during development. The present studies investigate expression of vascular endothelial growth factor A (VEGF), angiopoietin1 (ANGPT1) and ANGPT2 mRNAs together with their associated receptors (VEGFR and TIE2, respectively) during maturation. Elevated expression of VEGF and its receptors is associated with healthy, compared to atretic, follicles. Levels of VEGF significantly increase, while antagonistic ANGPT2 decrease, in granulosa cells (GC) at follicle selection. By comparison, levels of VEGF, VEGFR1, VEGFR2, ANGPT1, ANGPT2 and TIE2 within the theca layer do not change (P>0.05) relative to developmental stages surrounding follicle selection (6-8mm versus 9-12mm follicles). Prior to selection, treatment with transforming growth factor ß1 (TGFß1) significantly increases levels of VEGF in undifferentiated GC from prehierarchal (6-8mm) follicles and actively differentiating GC from selected (9-12 and F4) follicles. Moreover, subsequent to selection follicle stimulating hormone (FSH) increases VEGF expression in GC from 9 to 12mm follicles, and eventually luteinizing hormone (LH) promotes VEGF expression in GC from more mature preovulatory follicles. It is concluded that prior to follicle selection VEGF expression is regulated by autocrine and paracrine actions of TGFß1 (but not FSH), and that a comparatively limited extent of vasculature is sufficient to maintain prehierarchal follicles in a viable and undifferentiated state. At follicle selection, FSH- and subsequently LH-induced VEGF production within the GC layer enhance angiogenesis within the theca layer, which facilitates the rapid growth of preovulatory follicles via enhanced incorporation of yellow yolk.

Transfusion in children: epidemiology and 10-year survival of transfusion recipients.

OBJECTIVE: To describe the epidemiology of blood transfusion in children: including the incidence of transfusion, the diagnoses leading to transfusion, donor exposure (DE) and post-transfusion survival. STUDY DESIGN AND METHODS: The Epidemiology and Survival of Transfusion Recipients (EASTR) Study was a multi-centre epidemiological study with prospective survival monitoring. Cross-sectional sampling of adult and paediatric transfusion recipients in 29 hospitals was used to select three separate cohorts of red cell (RBC), platelet (PLT) and fresh frozen plasma (FFP) recipients between October 2001 and September 2002. This paper presents the analysis of results for children <16 years. RESULTS: Children <16 years comprised 449 (5%) of the RBC, 362 (9%) of the FFP and 452 (13%) of the PLT recipients. In children 54% of RBC, 63% FFP and 45% PLT recipients were under 1 year of age and 57% RBC, 60% FFP and 52% PLT were male. Median (IQR) DEduring the study year was 3(2-8); 5(2-13) and 11(6-21) in the RBC, FFP and PLT cohorts, respectively. A total of 20% of RBC, 31% of FFP and 54% of PLT recipients had been exposed to >10 donors. Perinatal conditions were the commonest indication for transfusion in the RBC (36%) and FFP (44%) cohorts and comprised 31% of the PLT cohort. Medical conditions (48%), predominantly malignancy (33%), were the most frequent indication in the PLT cohort. The 10 year (95% CI) survival rates were 81% (77-85%), 72% (67-76%) and 71% (66-75%)for RBC, FFP and PLT cohorts, respectively. CONCLUSIONS: Around half of paediatric transfusion recipients are under 1 year of age. Exposure to components from multiple donors is common. At least 70% of paediatric recipients are long survivors and are at risk for late complications of transfusion.

Equine Protozoal Myeloencephalitis: An Updated Consensus Statement with a Focus on Parasite Biology, Diagnosis, Treatment, and Prevention.

Equine protozoal myeloencephalitis (EPM) remains an important neurologic disease of horses. There are no pathognomonic clinical signs for the disease. Affected horses can have focal or multifocal central nervous system (CNS) disease. EPM can be difficult to diagnose antemortem. It is caused by either of 2 parasites, Sarcocystis neurona and Neospora hughesi, with much less known about N. hughesi. Although risk factors such as transport stress and breed and age correlations have been identified, biologic factors such as genetic predispositions of individual animals, and parasite-specific factors such as strain differences in virulence, remain largely undetermined. This consensus statement update presents current published knowledge of the parasite biology, host immune response, disease pathogenesis, epidemiology, and risk factors. Importantly, the statement provides recommendations for EPM diagnosis, treatment, and prevention.

Granulosa cell responsiveness to follicle stimulating hormone during early growth of hen ovarian follicles.

In the laying hen ovary, the cyclic recruitment of a follicle represents a process in which a single follicle is selected to enter the rapid growth phase and undergo final maturation prior to ovulation. Published data support the proposal that final differentiation of the granulosa cell (GC) layer commences at the time of follicle selection. This process is characterized by the enhanced capacity for FSH-induced cell signaling via the protein kinase A/cyclic adenosine monophosphate (cAMP) pathway. One consequence of such signaling within the GC layer is the initial capacity for steroidogenesis (predominantly progesterone production) mediated by increased expression of mRNA encoding steroidogenic acute regulatory protein (STAR) and the cholesterol side-chain cleavage enzyme (CYP11A). Prior to selection, the GC layer remains minimally responsive to a 3 h challenge with FSH (10 ng/mL), in vitro, compared to that from the most recently selected 9- to 12-mm follicle. By comparison, when the duration of the cell culture prior to FSH challenge is increased to 18 h, GCs collected from 1- to 2-mm, 3- to 5-mm, and 6- to 8-mm follicles respond to a 3 h FSH challenge by increasing STAR expression and progesterone production, with the greatest response from GCs collected from 6- to 8-mm follicles. Culture with Bone Morphogenetic Protein 6 (BMP6) enhances both CYP11A expression and FSH responsiveness at each stage of development, with the greatest response again occurring in GCs from 6- to 8-mm follicles. Significantly, factors that activate mitogen activated protein kinase (MAPK) or protein kinase C (PKC) signaling prevent the ability of prolonged culture or culture with BMP6 to induce FSH-responsiveness and the initiation of GC differentiation at each stage of development. Collectively, these results provide further support for the hypothesis that prior to follicle selection, inhibitory cell signaling (e.g., MAPK, PKC) maintains the GC layer in an undifferentiated state in follicles of all sizes, even in the presence of a differentiation-promoting signal (BMP6). The process by which the GC layer from the single 6- to 8-mm follicle selected each ovulatory cycle ultimately escapes inhibitory signaling to initiate FSH-responsiveness remains to be established.