Patient experiences of their current asthma care and their views toward providing support for patients with asthma in community pharmacy: A Qualitative study.

Background: An estimated 300 million people live with asthma globally. In England, a significant percentage live with poorly controlled asthma symptoms. Community pharmacists might be able to play a role in filling gaps in asthma care as they have the expertise and are in regular contact with patients with long term conditions. This study described patients' experiences of the management of their asthma in the general physician (GP) practice and community pharmacy settings and explored patients' views on providing support for them in community pharmacy. Method: This is a descriptive qualitative study. Thirteen adult asthma patients were recruited from a GP practice in the Northwest of England. Semi-structured qualitative interviews were conducted face-to-face or by telephone. The interviews were recorded, transcribed and analysed using a thematic analysis approach. Ethics approval was obtained before the study commenced and all participants gave informed written consent to participate. Results: We identified challenges in the current asthma care provided to patients with asthma including lack of continuity of care, inability to book an appointment and other experienced differences in the quality of asthma care provided to them and/or access to annual asthma reviews across different GP practices. Additionally, there is lack of awareness of services provided in community pharmacy. These challenges along with having comorbidities alongside asthma may negatively affect asthma patients' engagement with their asthma appointments and their behaviour toward their asthma. Conclusions: Patients showed trust in community pharmacists same as other HCPs to support them with their asthma care. Patients thought that being provided with regular asthma care including reviews in community pharmacy might be a suitable approach to respond to patients' needs and preferences in terms of their asthma management because of ease of access to community pharmacy. Pharmacists could be involved in the provision of community pharmacy-based asthma interventions that involve more than inhaler technique education. Further research should focus on developing structured approaches for asthma patient education that can be implemented consistently in the context of community pharmacy in England.

Loss of LPAR6 and CAB39L dysregulates the basal-to-luminal urothelial differentiation program, contributing to bladder carcinogenesis.

We describe a strategy that combines histologic and molecular mapping that permits interrogation of the chronology of changes associated with cancer development on a whole-organ scale. Using this approach, we present the sequence of alterations around RB1 in the development of bladder cancer. We show that RB1 is not involved in initial expansion of the preneoplastic clone. Instead, we found a set of contiguous genes that we term "forerunner" genes whose silencing is associated with the development of plaque-like field effects initiating carcinogenesis. Specifically, we identified five candidate forerunner genes (ITM2B, LPAR6, MLNR, CAB39L, and ARL11) mapping near RB1. Two of these genes, LPAR6 and CAB39L, are preferentially downregulated in the luminal and basal subtypes of bladder cancer, respectively. Their loss of function dysregulates urothelial differentiation, sensitizing the urothelium to N-butyl-N-(4-hydroxybutyl)nitrosamine-induced cancers, which recapitulate the luminal and basal subtypes of human bladder cancer.

A transgenic bacterial artificial chromosome approach to identify regulatory regions that direct Amhr2 and Osterix expression in Müllerian duct mesenchyme.

A transgenic mouse approach using bacterial artificial chromosomes (BAC) was used to identify regulatory regions that direct Müllerian duct expression for Amhr2 and Osterix (Osx, also known as Sp7). Amhr2 encodes the receptor that mediates anti-Müllerian hormone (AMH) signaling for Müllerian duct regression in male embryos. Amhr2 is expressed in the Müllerian duct mesenchyme of both male and female embryos. A ∼147-kb BAC clone containing the Amhr2 locus was used to generate transgenic mice. The transgene was able to rescue the block in Müllerian duct regression of Amhr2-null males, suggesting that the BAC clone contains regulatory sequences active in male embryos. Osx is expressed in the developing skeleton of male and female embryos but is also an AMH-induced gene that is expressed in the Müllerian duct mesenchyme exclusively in male embryos. Osx-Cre transgenic mice were previously generated using a ∼204-kb BAC clone. Crosses of Osx-Cre mice to Cre-dependent lacZ reporter mice resulted in reporter expression in the developing skeleton and in the Müllerian duct mesenchyme of male but not female embryos. Osx-Cherry transgenic mice were previously generated using a 39-kb genomic region surrounding the Osx locus. Osx-Cherry embryos expressed red fluorescence in the developing skeleton and Müllerian duct mesenchyme of male but not female embryos. In addition, female Osx-Cherry embryos ectopically expressing human AMH from an Mt1-AMH transgene activated red fluorescence in the Müllerian duct mesenchyme. These results suggest that the 39-kb region used to generate Osx-Cherry contains male-specific Müllerian duct mesenchyme regulatory sequences that are responsive to AMH signaling. These BAC transgenic mouse approaches identify two distinct regions that direct Müllerian duct mesenchyme expression and contribute fundamental knowledge to define a gene regulatory network for sex differentiation.

Distal-less homeobox genes Dlx5/6 regulate Müllerian duct regression.

Dlx5 and Dlx6 encode distal-less homeodomain transcription factors that are present in the genome as a linked pair at a single locus. Dlx5 and Dlx6 have redundant roles in craniofacial, skeletal, and uterine development. Previously, we performed a transcriptome comparison for anti-Müllerian hormone (AMH)-induced genes expressed in the Müllerian duct mesenchyme of male and female mouse embryos. In that study, we found that Dlx5 transcripts were nearly seven-fold higher in males compared to females and Dlx6 transcripts were found only in males, suggesting they may be AMH-induced genes. Therefore, we investigated the role of Dlx5 and Dlx6 during AMH-induced Müllerian duct regression. We found that Dlx5 was detected in the male Müllerian duct mesenchyme from E14.5 to E16.5. In contrast, in female embryos Dlx5 was detected in the Müllerian duct epithelium. Dlx6 expression in Müllerian duct mesenchyme was restricted to males. Dlx6 expression was not detected in female Müllerian duct mesenchyme or epithelium. Genetic experiments showed that AMH signaling is necessary for Dlx5 and Dlx6 expression. Müllerian duct regression was variable in Dlx5 homozygous mutant males at E16.5, ranging from regression like controls to a block in Müllerian duct regression. In E16.5 Dlx6 homozygous mutants, Müllerian duct tissue persisted primarily in the region adjacent to the testes. In Dlx5-6 double homozygous mutant males Müllerian duct regression was also found to be incomplete but more severe than either single mutant. These studies suggest that Dlx5 and Dlx6 act redundantly to mediate AMH-induced Müllerian duct regression during male differentiation.

Chronic Estrus Disrupts Uterine Gland Development and Homeostasis.

Female mice homozygous for an engineered Gnrhr E90K mutation have reduced gonadotropin-releasing hormone signaling, leading to infertility. Their ovaries have numerous antral follicles but no corpora lutea, indicating a block to ovulation. These mutants have high levels of circulating estradiol and low progesterone, indicating a state of persistent estrus. This mouse model provided a unique opportunity to examine the lack of cyclic levels of ovarian hormones on uterine gland biology. Although uterine gland development appeared similar to controls during prepubertal development, it was compromised during adolescence in the mutants. By age 20 weeks, uterine gland development was comparable to controls, but pathologies, including cribriform glandular structures, were observed. Induction of ovulations by periodic human chorionic gonadotropin treatment did not rescue postpubertal uterine gland development. Interestingly, progesterone receptor knockout mice, which lack progesterone signaling, also have defects in postpubertal uterine gland development. However, progesterone treatment did not rescue postpubertal uterine gland development. These studies indicate that chronically elevated levels of estradiol with low progesterone and therefore an absence of cyclic ovarian hormone secretion disrupts postpubertal uterine gland development and homeostasis.

Spoken propositional idea density, a measure to help second language English speaking students: A multicentre cohort study.

PURPOSE: Idea density has been shown to influence comprehension time for text in various populations. This study aims to explore the influence of spoken idea density on attainment in young, healthy subjects using demographic characteristics. METHODS: Students watched two online lectures and answered 10 multiple choice questions on them. Students received one more idea dense (MID) and one less idea dense (LID) lecture on two different subjects. RESULTS: Seventy-five students completed the study achieving a higher median score after a less idea-dense lecture (LID = 7(3), MID = 6(3), p = 0.04). Artificial neural network models revealed the first language as the main predictor of exam performance. The odds ratio (OR) of obtaining ≥70% after a more idea-dense lecture was six-time higher for the first language versus second language English speakers (OR = 5.963, 95% CI 1.080-32.911, p = 0.041). The odds ratio was not significant when receiving a less dense lecture (OR = 2.298, 95% CI 0.635-8.315, p = 0.205). Second-language speakers benefited from receiving a lower idea density, achieving a 10.8% score increase from high to low density, versus a 3.2% increase obtained by first language speakers. CONCLUSIONS: The propositional idea density of lectures directly influences students' comprehension, and disproportionately for second language speakers; revealing the possibility of reduced spoken idea density in levelling the attainment differential between first and second language speakers.

The management of asthma in adult patients in the community pharmacy setting: Literature review.

BACKGROUND: Asthma poses a public health concern, with an estimated 235 million people currently living with the condition globally. The provision of evidence-based, patient-centred services for adult asthma patients in community pharmacy which involves collaboration across the multidisciplinary team could improve their asthma control. OBJECTIVES: A literature review was conducted to examine the evidence of asthma management in community pharmacy setting. METHODS: Five databases were searched to identify relevant articles published before February 2021. Screening of the potential studies was performed to remove articles that did not comply with the inclusion criteria. Relevant data from all included studies was collected using a data extraction form to ensure consistency throughout the review. RESULTS: Twenty studies were included in the review; all were conducted in community pharmacy settings in the period of 2001-2020, in different countries. The studies included randomised controlled trials, controlled trials and observational studies. Several successful community pharmacy-based services that were provided to asthma patients to improve their asthma management were highlighted in this review. These interventions consisted of one or more components and included: patient education, inhaler technique improvement, patient counselling, self-management plans, development and provision of asthma action plans and referral to other health care practitioners. None of the studies involved medication or dosage changes by community pharmacy. CONCLUSIONS: The evidence discussed in this review showed that community pharmacists are well-placed to deliver services to asthma patients and many studies were conducted in the community pharmacy to improve asthma control in adult patients. However, further research could be conducted to explore further opportunities for community pharmacy to enhance asthma patients management of their condition.

CreLite: An optogenetically controlled Cre/loxP system using red light.

BACKGROUND: Precise manipulation of gene expression with temporal and spatial control is essential for functional analysis and determining cell lineage relationships in complex biological systems. The cyclic recombinase (Cre)-loxP system is commonly used for gene manipulation at desired times and places. However, specificity is dependent on the availability of tissue- or cell-specific regulatory elements used in combination with Cre. Here, we present CreLite, an optogenetically controlled Cre system using red light in developing zebrafish embryos. RESULTS: Cre activity is disabled by splitting Cre and fusing with the Arabidopsis thaliana red light-inducible binding partners, PhyB and PIF6. Upon red light illumination, the PhyB-CreC and PIF6-CreN fusion proteins come together in the presence of the cofactor phycocyanobilin (PCB) to restore Cre activity. Red light exposure of zebrafish embryos harboring a Cre-dependent multicolor fluorescent protein reporter injected with CreLite mRNAs and PCB resulted in Cre activity as measured by the generation of multispectral cell labeling in several different tissues. CONCLUSIONS: Our data show that CreLite can be used for gene manipulations in whole embryos or small groups of cells at different developmental stages, and suggests CreLite may also be useful for temporal and spatial control of gene expression in cell culture, ex vivo organ culture, and other animal models.

Stakeholder perceptions of a new model of care for medication supply at hospital discharge.

OBJECTIVES: Problems relating to patients' medication are common during hospital discharge, often resulting in unnecessary harm to patients and even hospital readmission. To overcome these issues and improve patient discharge, an evidence-based approach was used to develop an innovative model of care for the supply of medication at hospital discharge. The model increases pharmacy involvement, uses community pharmacies in the supply process and encourages patient follow-up after discharge. This study aimed to determine if the proposed new model of care was considered feasible and acceptable by the target population. METHODS: This formative evaluation involved qualitative interviews and focus groups to explore the opinions of stakeholders involved in delivery or use of the new model of care. A range of stakeholders participated, including a variety of hospital and community-based healthcare professionals as well as expert patients and carers. Thematic analysis of the data was undertaken. RESULTS: The study provided a holistic overview of stakeholder perceptions of the new model of care. Overall, stakeholders were enthusiastic about the new model. Three themes emerged during analysis, providing an informative review of the new model. Themes included: impact, resources required and ensuring safety and quality of the new model of care. Potential issues were identified within each theme along with ideas for overcoming these issues. CONCLUSIONS: The new model of care appears to be a positive step towards improving patient discharge from hospital. Future work will involve a pilot of the new model of care using the study findings to assist with implementation.

AMH and AMHR2 mutations: A spectrum of reproductive phenotypes across vertebrate species.

Anti-Müllerian hormone (AMH) is a member of the Transforming Growth Factor-ß family of secreted signaling proteins. AMH is expressed in Sertoli cells of the fetal and adult testes and granulosa cells of the postnatal ovary. AMH is required for the regression of the Müllerian ducts in mammalian fetuses during male differentiation. AMH signals through its Type II receptor, AMHR2. AMHR2 is expressed in mesenchyme adjacent to the Müllerian ducts, and in Sertoli, Leydig, and granulosa cells. Although AMH and AMHR2 genes have been identified in numerous vertebrate species, spontaneous or engineered mutations or variants have been found or created in only a few mammals and teleost fishes. AMH or AMHR2 mutations in mammals lead to the development of Persistent Müllerian Duct Syndrome (PMDS), a recessive condition in which affected males are fully virilized but retain Müllerian duct-derived tissues, including a uterus and oviducts, and in human and dog, undescended testes. Amh mutant female mice had accelerated ovarian primordial follicle recruitment, suggesting a role for AMH in regulating germ cells. amh and amhr2 mutations have also been experimentally generated in various teleost fishes. Depending on the fish species, loss of AMH signaling results in infertility, germ cell tumors, or male-to-female sex reversal. Here we compare the spectrum of phenotypes caused by AMH and AMHR2 mutations in a variety of vertebrate species. There are both common and unique phenotypes between species, highlighting the range of biological processes regulated by AMH signaling.

The islet-expressed Lhx1 transcription factor interacts with Islet-1 and contributes to glucose homeostasis.

The LIM-homeodomain (LIM-HD) transcription factor Islet-1 (Isl1) interacts with the LIM domain-binding protein 1 (Ldb1) coregulator to control expression of key pancreatic ß-cell genes. However, Ldb1 also has Isl1-independent effects, supporting that another LIM-HD factor interacts with Ldb1 to impact ß-cell development and/or function. LIM homeobox 1 (Lhx1) is an Isl1-related LIM-HD transcription factor that appears to be expressed in the developing mouse pancreas and in adult islets. However, roles for this factor in the pancreas are unknown. This study aimed to determine Lhx1 interactions and elucidate gene regulatory and physiological roles in the pancreas. Co-immunoprecipitation using ß-cell extracts demonstrated an interaction between Lhx1 and Isl1, and thus we hypothesized that Lhx1 and Isl1 regulate similar target genes. To test this, we employed siRNA-mediated Lhx1 knockdown in ß-cell lines and discovered reduced Glp1R mRNA. Chromatin immunoprecipitation revealed Lhx1 occupancy at a domain also known to be occupied by Isl1 and Ldb1. Through development of a pancreas-wide knockout mouse model ( Lhx1∆Panc), we demonstrate that aged Lhx1∆Panc mice have elevated fasting blood glucose levels, altered intraperitoneal and oral glucose tolerance, and significantly upregulated glucagon, somatostatin, pancreatic polypeptide, MafB, and Arx islet mRNAs. Additionally, Lhx1∆Panc mice exhibit significantly reduced Glp1R, an mRNA encoding the insulinotropic receptor for glucagon-like peptide 1 along with a concomitant dampened Glp1 response and mild glucose intolerance in mice challenged with oral glucose. These data are the first to reveal that the Lhx1 transcription factor contributes to normal glucose homeostasis and Glp1 responses.

Assessing uptake of antimicrobials by Zea mays L. and prevalence of antimicrobial resistance genes in manure-fertilized soil.

Manure-borne antimicrobials and antimicrobial resistance genes (ARGs) are of environmental concern due to their potential to be transferred into the food-web via plant-uptake. In this study, Zea mays L. seeds were grown in three different soil conditions: soil without dairy manure, dairy manure-amended soil, and antimicrobial spiked dairy manure-amended soil, to investigate the potential uptake of antimicrobials and ARGs present in manure. The antimicrobial spiked manure consisted of dairy manure fortified with 1 mg/Kg of each individual antimicrobial compounds belonging to the sulfonamide and tetracycline classes. Samples of the Zea mays L. plants were harvested over the course of three weeks to determine potential uptake of antimicrobials from soil to plant shoots, and to compare prevalence of ARGs in manure amended soils and plant tissue. Antimicrobial analysis was performed using liquid chromatography with tandem mass spectrometry (LC-MS/MS), and ARGs (sul1, tetO, and OXA-1) were analyzed using quantitative polymerase chain reaction (qPCR). The study found that both tetracycline and sulfamerazine antimicrobials bioaccumulated in the Zea mays L., reaching concentrations of nearly 3000 ng/g and 1260 ng/g, respectively. Tetracycline residues predominated in the soil, while sulfonamides had mainly bioaccumulated in Zea mays L. tissue. The greatest average uptake factor within the Zea mays L. tissue was 8 for tetracyclines and 110 for sulfonamides indicating larger bioaccumulation of sulfonamides. Additionally, three ARGs (sul1, tetO, and OXA-1) were detected in the soil, only after manure application. However, ARGs were not detected in any of the plant samples.

Osterix functions downstream of anti-Müllerian hormone signaling to regulate Müllerian duct regression.

In mammals, the developing reproductive tract primordium of male and female fetuses consists of the Wolffian duct and the Müllerian duct (MD), two epithelial tube pairs surrounded by mesenchyme. During male development, mesenchyme-epithelia interactions mediate MD regression to prevent its development into a uterus, oviduct, and upper vagina. It is well established that transforming growth factor-ß family member anti-Müllerian hormone (AMH) secreted from the fetal testis and its type 1 and 2 receptors expressed in MD mesenchyme regulate MD regression. However, little is known about the molecular network regulating downstream actions of AMH signaling. To identify potential AMH-induced genes and regulatory networks controlling MD regression in a global nonbiased manner, we examined transcriptome differences in MD mesenchyme between males (AMH signaling on) and females (AMH signaling off) by RNA-seq analysis of purified fetal MD mesenchymal cells. This analysis found 82 genes up-regulated in males during MD regression and identified Osterix (Osx)/Sp7, a key transcriptional regulator of osteoblast differentiation and bone formation, as a downstream effector of AMH signaling during MD regression. Osx/OSX was expressed in a male-specific pattern in MD mesenchyme during MD regression. OSX expression was lost in mutant males without AMH signaling. In addition, transgenic mice ectopically expressing human AMH in females induced a male pattern of Osx expression. Together, these results indicate that AMH signaling is necessary and sufficient for Osx expression in the MD mesenchyme. In addition, MD regression was delayed in Osx-null males, identifying Osx as a factor that regulates MD regression.

Hospital patient discharge process: an evaluation.

OBJECTIVES: Medication discrepancies for patients after discharge from hospital are well documented. They have been shown to cause unnecessary harm to patients and can result in hospital readmission. To improve patient discharge, the current process of discharging patients from hospital (the discharge process) needs evaluating to determine where and why medication issues occur. This study aimed to identify and evaluate the discharge process used in a range of acute National Health Service hospitals across the North West of England. METHODS: This qualitative study involved semi-structured telephone interviews with 13 chief pharmacists or an appropriately nominated member of the hospital pharmacy team. Thematic analysis of the transcribed interview data was performed. Data analysis revealed eight main themes which all impacted on the discharge process. RESULTS: The study was successful in identifying the discharge process across the range of hospitals as well as key issues and examples of good practice. The hospitals involved in the study were found to have similar discharge processes with issues common to all. One significant finding was a lack of patient involvement in the discharge process. CONCLUSIONS: To improve the patient discharge process, innovative solutions are required to overcome the current issues. In future work, the study findings will be used to develop a new model of care for patient discharge from hospital.

UK hospital patient discharge: the patient perspective.

OBJECTIVES: Hospital discharge is a complex process that can result in errors and delays for patients, particularly around the supply of medicines and communication of information. To improve patient discharge, patient perspectives of the discharge service must be explored to determine where patients feel problems arise. This study aimed to explore inpatient perceptions and experiences of the current discharge process. METHODS: This study involved questionnaires with patients at a large city-centre teaching hospital. RESULTS: A total of 104 inpatients participated, 60% (n=62) were male with an average age of 55 (age range 19-93). Participants were from a range of medical, surgical and admissions wards. The majority, 71% of respondents (n=74), took regular medicines, with 65% (n=48) taking five or more medicines daily. Most patients, 89% (n=87), were satisfied with their hospital discharge but felt that it took too long. The perceived main cause of delay was waiting for medicines. Other highlighted issues included lack of counselling by pharmacists and the need for more patient involvement throughout the discharge process. CONCLUSIONS: This study showed that certain aspects of the discharge process need improving to provide safe, quality care for patients and improve patient experience of discharge. The findings from this study will inform the development of a new model of care for patient discharge from hospital.

Molecular genetics of Müllerian duct formation, regression and differentiation.

The Müllerian duct (MD) forms the female reproductive tract (FRT) consisting of the oviducts, uterus, cervix, and upper vagina. FRT function is vital to fertility, providing the site of fertilization, embryo implantation and fetal development. Developmental defects in the formation and diseases of the FRT, including cancer and endometriosis, are prevalent in humans and can result in infertility and death. Furthermore, because the MDs are initially formed regardless of genotypic sex, mesenchymal to epithelial signaling is required in males to mediate MD regression and prevents the development of MD-derived organs. In males, defects in MD regression result in the retention of FRT organs and have been described in several human syndromes. Although to date not reported in humans, ectopic activation of MD regression signaling components in females can result in aplasia of the FRT. Clearly, MD development is important to human health; however, the molecular mechanisms remain largely undetermined. Molecular genetics studies of human diseases and mouse models have provided new insights into molecular signaling during MD development, regression and differentiation. This review will provide an overview of MD development and important genes and signaling mechanisms involved.

Cell-specific actions of a human LHX3 gene enhancer during pituitary and spinal cord development.

The LIM class of homeodomain protein 3 (LHX3) transcription factor is essential for pituitary gland and nervous system development in mammals. In humans, mutations in the LHX3 gene underlie complex pediatric syndromes featuring deficits in anterior pituitary hormones and defects in the nervous system. The mechanisms that control temporal and spatial expression of the LHX3 gene are poorly understood. The proximal promoters of the human LHX3 gene are insufficient to guide expression in vivo and downstream elements including a conserved enhancer region appear to play a role in tissue-specific expression in the pituitary and nervous system. Here we characterized the activity of this downstream enhancer region in regulating gene expression at the cellular level during development. Human LHX3 enhancer-driven Cre reporter transgenic mice were generated to facilitate studies of enhancer actions. The downstream LHX3 enhancer primarily guides gene transcription in α-glycoprotein subunit -expressing cells secreting the TSHß, LHß, or FSHß hormones and expressing the GATA2 and steroidogenic factor 1 transcription factors. In the developing nervous system, the enhancer serves as a targeting module active in V2a interneurons. These results demonstrate that the downstream LHX3 enhancer is important in specific endocrine and neural cell types but also indicate that additional regulatory elements are likely involved in LHX3 gene expression. Furthermore, these studies revealed significant gonadotrope cell heterogeneity during pituitary development, providing insights into the cellular physiology of this key reproductive regulatory cell. The human LHX3 enhancer-driven Cre reporter transgenic mice also provide a valuable tool for further developmental studies of cell determination and differentiation in the pituitary and nervous system.

Passion in the performing arts: clarifying active occupational participation.

UNLABELLED: Active participation in daily occupations is a vital part of everyday life, social participation and healthy life long human development; however, enablers of active participation are not well understood. Passion, a strong tendency towards an activity that a person finds meaningful and spends a lot of time doing, is a potential enabler. Accordingly, it is important to understand how an individual's passion for a specific occupation plays out across the occupational life course. OBJECTIVE: To explore the experience of passion across the life course of older adults involved in the performing arts. PARTICIPANTS: Seven older adults involved in, or retired from, the performing arts, who consider themselves passionate about their occupation. METHODS: A constructivist grounded theory approach was used to explore, through interviews with older adults, passion for performing arts across the life course. RESULTS: Emerging themes supported development of an initial theoretical framework explicating active participation and passion. It centers on passion as an enabler of occupational participation through different modes, and suggests barriers to that enablement process. CONCLUSIONS: Findings suggest that passion has an important role in continued active participation in an occupation; however, barriers, such as social and financial, can derail the pursuit of a passionate occupation.

A distal modular enhancer complex acts to control pituitary- and nervous system-specific expression of the LHX3 regulatory gene.

Lin-11, Isl-1, and Mec-3 (LIM)-homeodomain (HD)-class transcription factors are critical for many aspects of mammalian organogenesis. Of these, LHX3 is essential for pituitary gland and nervous system development. Pediatric patients with mutations in coding regions of the LHX3 gene have complex syndromes, including combined pituitary hormone deficiency and nervous system defects resulting in symptoms such as dwarfism, thyroid insufficiency, infertility, and developmental delay. The pathways underlying early pituitary development are poorly understood, and the mechanisms by which the LHX3 gene is regulated in vivo are not known. Using bioinformatic and transgenic mouse approaches, we show that multiple conserved enhancers downstream of the human LHX3 gene direct expression to the developing pituitary and spinal cord in a pattern consistent with endogenous LHX3 expression. Several transferable cis elements can individually guide nervous system expression. However, a single 180-bp minimal enhancer is sufficient to confer specific expression in the developing pituitary. Within this sequence, tandem binding sites recognized by the islet-1 (ISL1) LIM-HD protein are essential for enhancer activity in the pituitary and spine, and a pituitary homeobox 1 (PITX1) bicoid class HD element is required for spatial patterning in the developing pituitary. This study establishes ISL1 as a novel transcriptional regulator of LHX3 and describes a potential mechanism for regulation by PITX1. Moreover, these studies suggest models for analyses of the transcriptional pathways coordinating the expression of other LIM-HD genes and provide tools for the molecular analysis and genetic counseling of pediatric patients with combined pituitary hormone deficiency.

LHX3 and LHX4 transcription factors in pituitary development and disease.

The LHX3 and LHX4 LIM-homeodomain proteins are regulatory transcription factors that play overlapping but distinct functions during the establishment of the specialized cells of the mammalian pituitary gland and the nervous system. Recent studies have identified a variety of mutations in the LHX3 and LHX4 genes in patients with combined pituitary hormone deficiency diseases. These patients have complex and variable syndromes involving short stature, metabolic disorders, reproductive system deficits, and nervous system developmental abnormalities. The short stature secondary to growth hormone deficiency is a key feature of the disorders associated with these gene mutations and responds well to supplementation with recombinant growth hormone. Overall, the frequency of mutations in the LHX3 and LHX4 genes in patients with combined pituitary hormone deficiency is low. Mutations in other regulatory genes such as HESX1, PROP1, PIT1 / POU1F1, and GLI2 have been shown to be additional causes of pituitary hormone deficiency, but overall, the etiology of many cases of hypopituitarism is not understood. Further investigation is therefore required to identify other genes, both primary regulatory genes and those with modifier functions, which contribute to pituitary development and function.