Activation of Steroid and Xenobiotic Receptor (SXR, NR1I2) and Its Orthologs in Laboratory Toxicologic, and Genome Model Species

Background: Nuclear receptor subfamily 1, group I, member 2 (NR1I2), commonly known as steroid xenobiotic receptor (SXR) in humans, is a key ligand-dependent transcription factor responsible for the regulation of xenobiotic, steroid, and bile acid metabolism. The ligand-binding domain is principally responsible for species-specific activation of NR1I2 in response to xenobiotic exposure. Objectives: Our objective in this study was to create a common framework for screening NR1I2 orthologs from a variety of model species against environmentally relevant xenobiotics and to evaluate the results in light of using the species as predictors of xenobiotic disposition and for assessment of environmental health risk. Methods: Sixteen chimeric fusion plasmid vectors expressing the Gal4 DNA-binding domain and species-specific NR1I2 ligand-binding domain were screened for activation against a spectrum of 27 xenobiotic compounds using a standardized cotransfection receptor activation assay. Results: NR1I2 orthologs were activated by various ligands in a dose-dependent manner. Closely related species show broadly similar patterns of activation; however, considerable variation to individual compounds exists, even among species varying in only a few amino acid residues. Conclusions: Interspecies variation in NR1I2 activation by various ligands can be screened through the use of in vitro NR1I2 activation assays and should be taken into account when choosing appropriate animal models for assessing environmental health risk

The presentation, diagnosis and management of non-traumatic wrist pain: an evaluation of current practice in secondary care in the UK NHS

AbstractObjectivesThe study aims were to assess the burden of non-traumatic wrist pain in terms of numbers of referrals to secondary care, and to characterise how patients present, are diagnosed and are managed in secondary care in the United Kingdom National Health Service.MethodsTen consecutive patients presenting with non-traumatic wrist pain were identified retrospectively at each of 16 participating hospitals and data was extracted for twelve months following the initial referral.ResultsThe 160 patients consisted of 100 females and 60 males with a median age of 49, accounting for approximately 13% of all new hand/wrist referrals. The dominant wrist was affected in 60% of cases and the mean symptom duration was 13.3 months. Diagnoses were grouped into: osteoarthritis (OA) (31%), tendinopathy (13%), ganglion (14%), ulnar sided pain (17%) and other (25%). The OA group was significantly older than other groups, while other groups contained a predominance of females.The non-surgical interventions in decreasing frequency of usage were: steroid injections (39%), physiotherapy (32%), splint (31%) and analgesics (12%). Of those who underwent surgery, all patients had previously received non-surgical treatment, however 42% had undergone only one non-surgical intervention.ConclusionNon-traumatic wrist pain represents a significant burden to secondary care both in terms of new patient referrals and in terms of investigation, follow up and treatment. Those presenting with osteoarthritis are more likely to be older and male, while those presenting with other diagnoses are more likely to be younger and female

Sequencing three crocodilian genomes to illuminate the evolution of archosaurs and amniotes

The International Crocodilian Genomes Working Group (ICGWG) will sequence and assemble the American alligator (Alligator mississippiensis), saltwater crocodile (Crocodylus porosus) and Indian gharial (Gavialis gangeticus) genomes. The status of these projects and our planned analyses are described

The effects of testosterone exposure on urogenital tract androgen receptor distribution in immature eastern fence lizards

Androgens are important to the development and maintenance of reproductive structures and functions; however, less is known about the importance of androgen signaling in females compared to males. The distribution of androgen receptors (ARs) in mammalian urogenital systems has been characterized but relatively little is known about AR expression in non-mammalian vertebrates. In this study, AR was immunolocalized in the urogenital tracts of juvenile eastern fence lizards, Sceloporus undulatus. A subset of lizards received testosterone implants to understand and examine the effects of elevated testosterone (T) on AR expression. Our results showed control males had moderate AR-positive staining within the epididymal epithelium and diffuse staining throughout the testes, with minor staining surrounding the sexual segments of the kidney. Control females revealed a similar intensity of staining within their oviduct and mesonephric ducts. Elevated T in lizards that received implants led to hypertrophied reproductive tracts and increased AR expression throughout most of the urogenital system. As compared to control males, males that received exogenous T revealed darker AR staining in the testis, spermatic ducts, and the sexual segments of the kidney. Androgen receptor staining was more intense and concentrated in the nuclei of the epithelia of the oviducts and ductus deferentia of T-treated females in comparison to controls. Our results demonstrate that the urogenital tracts of both sexes are androgen sensitive, and the tissues that responded the most dramatically to elevated T were the same tissues that had the most intense AR staining. Our results also suggest that exposure to elevated T concentrations leads to upregulation of AR expression and increased nuclear translocation. Although oviducts of T-treated females exhibited hypertrophy relative to untreated females and were AR positive, it is unclear if this was through direct actions of testosterone or the aromatization of androgens to estrogens

OOCYTE AND FOLLICULAR DEVELOPMENT IN IMMATURE EASTERN FENCE LIZARDS

The early stages of oogenesis and follicular organization occur in utero in mammals. In contrast, the early stages of oogenesis and follicular organization in reptiles occur during postnatal development. The increased accessibility to individuals undergoing these processes and the relatively large, yolk-laden oocytes make reptiles, such as lizards, an especially useful model for studying reproductive development. Seasonal cycles of oocyte and follicular maturation have been described in adults of numerous species of lizards; however, few studies have examined these processes in immature lizards. In this study, immature female Eastern Fence Lizards (Sceloporus undulatus) were collected monthly over a 2-year-period. The gonads and reproductive tracts were photographed in situ at 1–5x magnification for examination at the gross anatomical level. Left ovaries were removed, embedded in paraffin wax, sectioned on a rotary microtome, and stained using Masson’s trichrome. Ovarian and follicular stages were assigned to each lizard based on previously established histological criteria. Relationships between body size, capture date, and ovarian and follicular developmental stage were evaluated. Ovarian stage was assigned based on the most advanced follicle. Most ovaries were comprised of follicles of varying stages. As expected, there was a positive relationship between body size and ovarian stage; however, the relationship between time of year and ovarian stage is more complex when considering immature lizards of different sizes. Larger, yet still immature, lizards have a greater proportion of follicles at an advanced stage, in comparison to smaller individuals. Previous studies and our unpublished observations suggest that female Sceloporus undulatus obtain reproductive maturity within 1-2 years, depending on the timing of hatching and growth rates. Our data suggests that lizards that do not attain sexual maturity in their first year exhibit a prolonged period of pubescence, as indicated by advanced stages of previtellogenic follicles

POSTHATCHING DEVELOPMENT OF THE REPRODUCTIVE ANATOMY IN MALE EASTERN FENCE LIZARD (SCELOPORUS UNDULATUS)

Recent observations from our lab suggest that the timing of fundamental events in the development of male lizard reproductive anatomy differs from mammals and birds. Masculinization of the embryonic urogenital tract occurs during embryonic development in mammals and birds but has not been thoroughly examined in reptiles. In this study we described the post-hatching development and maturation of the reproductive system in male Eastern Fence Lizards. Male lizards ranging from neonates to small adults were captured monthly by hand or with a noose pole in the Oconee National Forest, GA, USA over a span of two years. Testes and spermatic ducts were photographed at 1–5X magnification to obtain standardized morphological measures. The left testis and left reproductive tract were removed, sectioned, and stained with Masson’s trichrome to study the microanatomical changes associated with maturation. In small hatchlings, the mesonephric kidney and its associated duct are still present, as they have not yet differentiated to the epididymis and ductus deferens. The mesonephros gradually condenses anteriorly as the mesonephric tubules give rise to efferent ducts and the mesonephric duct differentiates to epididymal duct and ductus deferens. The seminiferous tubules of recently hatched lizards are populated by spermatogonia and Sertoli cells, with no distinct lumen. A positive relationship exists between testis size and spermatogenic stage in larger animals; however, testis size and spermatogenic activity were also affected by time of year. Male lizards approaching maturity show evidence of active spermatogenesis and secretions from the sexual segment of the kidney. Our results indicate masculinization of the urogenital tract in Eastern Fence Lizards occurs after hatching, and maturation of the reproductive system is influenced by the combination of size and season

Environmental contaminants activate human and polar bear (Ursus maritimus) pregnane X receptors (PXR, NR1I2) differently

Background Many persistent organic pollutants (POPs) accumulate readily in polar bears because of their position as apex predators in Arctic food webs. The pregnane X receptor (PXR, formally NR1I2, here proposed to be named promiscuous xenobiotic receptor) is a xenobiotic sensor that is directly involved in metabolizing pathways of a wide range of environmental contaminants. Objectives In the present study, we comparably assess the ability of 51 selected pharmaceuticals, pesticides and emerging contaminants to activate PXRs from polar bears and humans using an in vitro luciferase reporter gene assay. Results We found that polar bear PXR is activated by a wide range of our test compounds (68%) but has a slightly more narrow ligand specificity than human PXR that was activated by 86% of the 51 test compounds. The majority of the agonists identified (70%) produces a stronger induction of the reporter gene via human PXR than via polar bear PXR, however with some notable and environmentally relevant exceptions. Conclusions Due to the observed differences in activation of polar bear and human PXRs, exposure of each species to environmental agents is likely to induce biotransformation differently in the two species. Bioinformatics analyses and structural modeling studies suggest that amino acids that are not part of the ligand-binding domain and do not interact with the ligand can modulate receptor activation

Influences of Sex, Incubation Temperature, and Environmental Quality on Gonadal Estrogen and Androgen Receptor Messenger RNA Expression in Juvenile American Alligators (Alligator mississippiensis)1

Gonadal steroid hormone receptors play a vital role in transforming ligand signals into gene expression. We have shown previously that gonads from wild-caught juvenile alligators express greater levels of estrogen receptor 1 (ESR1) than estrogen receptor 2 (ESR2). Furthermore, sexually dimorphic ESR2 mRNA expression (female > male) observed in animals from the reference site (Lake Woodruff, FL, USA) was lost in alligators from the contaminated Lake Apopka (FL, USA). We postulated that environmental contaminant exposure could influence gonadal steroid hormone receptor expression. Here, we address questions regarding gonadal estrogen and androgen receptor (AR) mRNA expression in 1-yr-old, laboratory-raised alligators. What are relative expression levels within gonads? Do these levels vary between sexes or incubation temperatures? Can contaminant exposure change these levels? We observed a similar pattern of expression (ESR1 > AR > ESR2) in ovary and testis. However, both incubation temperature and environment modulated expression. Males incubated at 33.5°C expressed greater AR levels than females incubated at 30°C; dimorphic expression was not observed in animals incubated at 32°C. Compared to Lake Woodruff alligators, Lake Apopka animals of both sexes showed lesser ESR2 mRNA expression levels. Employing cluster analyses, we integrated these receptor expression patterns with those of steroidogenic factors. Elevated ESR2 and CYP19A1 expressions were diagnostic of alligator ovary, whereas elevated HSD3B1, CYP11A1, and CYP17A1 expressions were indicative of testis. In contrast, AR, ESR1, and NR5A1 showed variable expressions that were not entirely associated with sex. These findings demonstrate that the mRNA expression of receptors required for steroid hormone signaling are modified by exposure to environmental factors, including temperature and contaminants