Controlled dual release of dihydrotestosterone and flutamide from polycaprolactone electrospun scaffolds accelerate burn wound healing.

Wound healing is a complex process involving multiple independent and overlapping sequential physiological mechanisms. In addition to cutaneous injury, a severe burn stimulates physiological derangements that induce a systemic hypermetabolic response resulting in impaired wound healing. Topical application of the anti-androgen drug, flutamide accelerates cutaneous wound healing, whereas paradoxically systemic dihydrotestosterone (DHT) improves burn wound healing. We developed and characterized a PCL scaffold that is capable of controlled release of androgen (DHT) and anti-androgen (F) individually or together. This study aims to investigate whether local modification of androgen actions has an impact on burn injury wound healing. In a full-thickness burn wound healing, mouse model, DHT/F-scaffold showed a significantly faster wound healing compared with F-scaffold or DHT-scaffold. Histology analysis confirmed that DHT/F-scaffold exhibited higher re-epithelization, cell proliferation, angiogenesis, and collagen deposition. Dual release of DHT and F from PCL scaffolds promoted cell proliferation of human keratinocytes and alters the keratinocyte cell cycle. Lastly, no adverse effects on androgen-dependent organs, spleen and liver were observed. In conclusion, we demonstrated DHT plus F load PCL scaffolds accelerated burn wound healing when loading alone did not. These findings point to a complex role of androgens in burn wound healing and open novel therapeutic avenues for treating severe burn patients.

Dihydrotestosterone (DHT) Enhances Wound Healing of Major Burn Injury by Accelerating Resolution of Inflammation in Mice.

Androgens have been known to inhibit cutaneous wound healing in men and male mice. However, in children with major burn injuries, a synthetic androgen was reported clinically to improve wound healing. The aim of this study is to investigate the role of dihydrotestosterone (DHT) as a new therapeutic approach in treating major burn injury. In the present study, mice received systemic androgen treatment post major burn injury. Wound healing rate and body weight were monitored over 21 days. The serum level of inflammatory cytokines/chemokines were measured using multiplex immunoassays. In addition, splenocyte enumeration was performed by flow cytometry. Healing phases of inflammation, re-epithelialization, cell proliferation and collagen deposition were also examined. In results, DHT treated mice lost less weight and displayed accelerated wound healing but has no impact on hypermetabolism. Mice, after burn injury, displayed acute systemic inflammatory responses over 21 days. DHT treatment shortened the systemic inflammatory response with reduced splenic weight and monocyte numbers on day 14 and 21. DHT treatment also reduced wound infiltrating macrophage numbers. In conclusion, DHT treatment facilitates local wound healing by accelerating the resolution of inflammation, but not through alterations of post-burn hypermetabolic response.

Macronutrient balance, reproductive function, and lifespan in aging mice.

In invertebrates, reproductive output and lifespan are profoundly impacted by dietary macronutrient balance, with these traits achieving their maxima on different diet compositions, giving the appearance of a resource-based tradeoff between reproduction and longevity. For the first time in a mammal, to our knowledge, we evaluate the effects of dietary protein (P), carbohydrate (C), fat (F), and energy (E) on lifespan and reproductive function in aging male and female mice. We show that, as in invertebrates, the balance of macronutrients has marked and largely opposing effects on reproductive and longevity outcomes. Mice were provided ad libitum access to one of 25 diets differing in P, C, F, and E content, with reproductive outcomes assessed at 15 months. An optimal balance of macronutrients exists for reproductive function, which, for most measures, differs from the diets that optimize lifespan, and this response differs with sex. Maximal longevity was achieved on diets containing a P:C ratio of 1:13 in males and 1:11 for females. Diets that optimized testes mass and epididymal sperm counts (indicators of gamete production) contained a higher P:C ratio (1:1) than those that maximized lifespan. In females, uterine mass (an indicator of estrogenic activity) was also greatest on high P:C diets (1:1) whereas ovarian follicle number was greatest on P:C 3:1 associated with high-F intakes. By contrast, estrous cycling was more likely in mice on lower P:C (1:8), and the number of corpora lutea, indicative of recent ovulations, was greatest on P:C similar to those supporting greatest longevity (1:11).

A simple, accurate and universal method for quantification of PCR.

BACKGROUND: Research into gene expression enables scientists to decipher the complex regulatory networks that control fundamental biological processes. Quantitative real-time PCR (qPCR) is a powerful and ubiquitous method for interrogation of gene expression. Accurate quantification is essential for correct interpretation of qPCR data. However, conventional relative and absolute quantification methodologies often give erroneous results or are laborious to perform. To overcome these failings, we developed an accurate, simple to use, universal calibrator, AccuCal. RESULTS: Herein, we show that AccuCal quantification can be used with either dye- or probe-based detection methods and is accurate over a dynamic range of ≥10(5) copies, for amplicons up to 500 base pairs (bp). By providing absolute quantification of all genes of interest, AccuCal exposes, and circumvents, the well-known biases of qPCR, thus allowing objective experimental conclusions to be drawn. CONCLUSION: We propose that AccuCal supersedes the traditional quantification methods of PCR.

Androgen actions in mouse wound healing: Minimal in vivo effects of local antiandrogen delivery.

The aims of this work were to define the role of androgens in female wound healing and to develop and characterize a novel wound dressing with antiandrogens. Androgens retard wound healing in males, but their role in female wound healing has not been established. To understand androgen receptor (AR)-mediated androgen actions in male and female wound healing, we utilized the global AR knockout (ARKO) mouse model, with a mutated AR deleting the second zinc finger to disrupt DNA binding and transcriptional activation. AR inactivation enhanced wound healing rate in males by increasing re-epithelialization and collagen deposition even when wound contraction was eliminated. Cell proliferation and migration in ARKO male fibroblasts was significantly increased compared with wild-type (WT) fibroblasts. However, ARKO females showed a similar healing rate compared to WT females. To exploit local antiandrogen effects in wound healing, while minimizing off-target systemic effects, we developed a novel electrospun polycaprolactone (PCL) scaffold wound dressing material for sustained local antiandrogen delivery. Using the antiandrogen hydroxyl flutamide (HF) at 1, 5, and 10 mg/mL in PCL scaffolds, controlled HF delivery over 21 days significantly enhanced in vitro cell proliferation of human dermal fibroblasts and human keratinocytes. HF-PCL scaffolds also promoted in vivo wound healing in mice compared with open wounds but not to PCL scaffolds.

Development and Characterization of Uterine Glandular Epithelium Specific Androgen Receptor Knockout Mouse Model.

While estrogen action is the major driver of uterine development, androgens acting via the androgen receptor (AR) may also promote uterine growth as suggested by uterine phenotypes in global AR knockout (ARKO) female mice. Because AR is expressed in uterine endometrial glands, we generated (Cre/loxP) uterine gland epithelium-specific ARKO (ugeARKO) to determine the role of endometrial gland-specific androgen actions. However, AR in uterine gland epithelium may not be required for normal uterine development and function because ugeARKO females had normal uterine development and fertility. To determine if exogenous androgens acting via AR can fully support uterine growth in the absence of estrogens, the ARKO and ugeARKO females were ovariectomized and treated with supraphysiological doses of testosterone or dihydrotestosterone (nonaromatizable androgen). Both dihydrotestosterone and testosterone supported full uterine regrowth in wild-type females while ARKO females had no regrowth (comparable to ovariectomized only). These findings suggest that androgens acting via AR can promote full uterine regrowth in the absence of estrogens. The ugeARKO had 50% regrowth when compared to intact uterine glands, and histomorphologically, both the endometrial and myometrial areas were significantly (P < 0.05) reduced, suggesting glandular epithelial AR located in the endometrium may indirectly modify myometrial development. Additionally, to confirm Cre function in endometrial glands, we generated uge-specific PTEN knockout mouse model. The ugePTEN knockout females developed severe endometrial hyperplasia and therefore present a novel model for future research.

The role of androgens in experimental rodent mammary carcinogenesis.

Breast cancer is currently the most frequent, fatal cancer of women in western countries. While estrogens have a widely understood involvement in breast cancer, a significant but not yet fully understood role for androgens has also been suggested. The principal androgen, testosterone, is the obligate steroidal precursor of estradiol, but can equally be metabolized into dihydrotestosterone, a more potent, pure androgen. Both androgens exert their distinctive biological effects via the androgen receptor, which is coexpressed with estrogen receptor alpha in 80 to 90% of breast cancers. The hormonal control of breast development and pathology has been examined experimentally through the use of animal models, notably mice and rats. This review summarizes the data from experimental rodent models on the effects of androgens in experimental breast cancer, aiming to address the importance of androgens and the androgen receptor in the origins and pathogenesis of breast cancers, as well as to discuss potential biomarker and therapeutic opportunities arising from novel insights based on the experimental research.

Glucocorticoid receptor in prostate epithelia is not required for corticosteroid-induced epithelial hyperproliferation in the mouse prostate.

BACKGROUND: Glucocorticoids are used as a last resort treatment for prostate cancer but the cell-specific glucocorticoid receptor (GR) mediated actions and the role of endogenous glucocorticoids in prostate are not understood. METHODS: We evaluated the influence of prostate epithelial GR mediated actions of glucocorticoids in prostate structural development by comparing the intact wild-type (WT) and prostate epithelia selective GR knockout (peGRKO) males at 8, 20, and 35 weeks of age. We also determined the cell-specific role of GR on corticosterone treatment induced prostate abnormalities by treating peGRKO and WT male mice with corticosterone depot pellets or placebo for 4 weeks. RESULTS: GR was not expressed in the epithelial cells of peGRKO prostate unlike WT but was expressed in stromal of both peGRKO and WT mice. Nevertheless, prostate weights, histological appearance, and secretory protein probasin expression in peGRKO were no different from WT. Despite lacking epithelial GR, the peGRKO prostate demonstrated corticosterone treatment induced hyperplasia similar to WT suggesting that stromal rather than epithelial GR mediates the hyperproliferative mouse prostate response to corticosterone. As circulating androgen levels were not affected by corticosterone treatment, this effect is likely to be mediated directly via prostate GR. CONCLUSIONS: Sustained administration of corticosterone induces prostate hyperplasia, which is mediated via GR expressed predominantly in the stroma. Thus GR mediated actions in the prostate may have significant cell-specific effects that could be utilized for more rational therapeutic approaches in prostate cancer treatment. This also illustrates the paracrine hormonal mechanisms in prostate pathophysiology.

Prostate epithelial AR inactivation leads to increased intraprostatic androgen synthesis.

BACKGROUND: Regulation of steroid synthesis within the prostate is not well understood. In this study, we examined androgen synthesis and metabolism in the mouse prostate. METHODS: Using LC-MSMS steroid assays, immunohistochemistry and real-time PCR we examined the role of prostate epithelial AR in regulating 5αR expression and subsequent androgen metabolism by analyzing natural differences in epithelial AR expression between lobes as well as in the prostate epithelial AR knockout (PEARKO) mouse model. Subsequently, the role of intraprostatic androgen metabolism and epithelial AR in the generation and progression of prostate epithelial pathology was examined using long-term exogenous testosterone (T) + estradiol (E2) exposure. RESULTS: Epithelial AR and 5αR2 expression as well as intraprostatic DHT followed the same lobe-specific pattern being lower in anterior than the other lobes (n = 6-8, P < 0.05). Lobe-specific 5αR2 expression was similar in PEARKO and wild-type (WT) prostate. However, PEARKO prostate had higher intraprostatic DHT content with significantly increased 5αR2 expression localized in abnormal epithelium. T + E2 treatment induced epithelial pathology was more common in PEARKO prostate compared to WT (20% vs. 2%), and was associated with increased 5αR2 expression (n = 6, P < 0.001). CONCLUSIONS: We suggest that androgen synthesis via 5αR2 expression is driven by its own product (DHT) acting on adjacent stromal cells in a paracrine loop leading to increased in situ androgen levels in the PEARKO prostate. This may form part of a feed-forward loop that promotes the development of epithelial pathology.

Targeted loss of androgen receptor signaling in murine granulosa cells of preantral and antral follicles causes female subfertility.

Ovarian granulosa cells display strong androgen receptor (AR) expression, suggesting a functional role for direct AR-mediated actions within developing mammalian follicles. By crossing AR-floxed and anti-Müllerian hormone (AMH)-Cre recombinase mice, we generated granulosa cell-specific androgen receptor knockout mice (GCARKO). Cre expression, assessed by lacZ activity, localized to 70%-100% of granulosa cells in most preantral to antral follicles, allowing for selected evaluation of granulosa cell AR-dependent actions during follicle development. Relative to wild-type (WT) females, GCARKO females were subfertile, producing a 24% reduction in the number of litters (P < 0.05) over 6 mo and an age-dependent decrease in total number of pups born, evident from 6 mo of age (P < 0.05). Follicle dynamics were altered in GCARKO ovaries at 3 mo of age, with a significant reduction in large preantral and small antral follicle numbers compared to WT ovaries (P < 0.05). Global premature follicle depletion was not observed, but increased follicular atresia was evident in GCARKO ovaries at 6 mo of age, with an 81% increase in unhealthy follicles and zona pellucida remnants (P < 0.01). Cumulus cell expansion was decreased (P < 0.01) and oocyte viability was diminished in GCARKO females, with a significant reduction in the percentage of oocytes fertilized after natural mating and, thus, in the rate of progression to the two-cell embryo stage (P < 0.05). In addition, compared with age-matched WT females, 6-mo-old GCARKO females exhibited significantly prolonged estrous cycles (P ≤ 0.05), suggesting altered hypothalamic-pituitary-gonadal feedback signaling. In conclusion, our findings revealed that selective loss of granulosa cell AR actions during preantral and antral stages of development leads to a premature reduction in female fecundity through reduced follicle health and oocyte viability.

Anterior prostate epithelial AR inactivation modifies estrogen receptor expression and increases estrogen sensitivity.

Androgens influence prostate growth and development, so androgen withdrawal can control progression of prostate diseases. Although estrogen treatment was originally used to induce androgen withdrawal, more recently direct estrogen effects on the prostate have been recognized, but the nature of androgen-estrogen interactions within the prostate remain poorly understood. To characterize androgen effects on estrogen sensitivity in the mouse prostate, we contrasted models of castration-induced androgen withdrawal in the prostate stromal and epithelial compartments with a prostate epithelial androgen receptor (AR) knockout (PEARKO) mouse model of selective epithelial AR inactivation. Castration markedly increased prostate epithelial estrogen receptor (ER)α immunoreactivity compared with very low ERα expression in intact males. Similarly, strong basal and luminal ERα expression was detected in PEARKO prostate of intact males, suggesting that epithelial AR activity regulated epithelial ERα expression. ERß was strongly expressed in intact, castrated, and PEARKO prostate. However, strong clusters of epithelial ERß positivity coincided with epithelial stratification in PEARKO prostate. In vivo estrogen sensitivity was increased in PEARKO males, with greater estradiol-induced prostate growth and epithelial proliferation leading to squamous metaplasia, featuring markedly increased epithelial proliferation, thickening, and keratinization compared with littermate controls. Our results suggest that ERα expression in the prostate epithelial cells is regulated by local, epithelia-specific, androgen-dependent mechanisms, and this imbalance in the AR- and ER-mediated signaling sensitizes the mature prostate to exogenous estrogens.

Long-term corticosterone treatment induced lobe-specific pathology in mouse prostate.

BACKGROUND: Glucocorticoids influence prostate development and pathology, yet the underlying mechanisms including possible direct glucocorticoid effect on the prostate are not well characterized. METHODS: We evaluated the expression of the glucocorticoid receptor (GR) together with the effects of supraphysiological glucocorticoid (corticosterone) on mouse prostate morphology and epithelial proliferation. Mature male mice were treated by weekly subdermal implantation of depot pellets containing either 1.5 mg corticosterone or placebo providing steady-state release for 4 weeks. RESULTS: Corticosterone treatment significantly increased dorsolateral and anterior prostate weights as well as prostate epithelial cell proliferation while epithelial apoptosis remained low upon corticosterone treatment. Histological analysis of the anterior lobe demonstrated abnormal, highly disorganized luminal epithelium with frequent formation of bridge-like structures lined by continuous layer of basal cells not observed following placebo treatment. Molecular analysis revealed corticosterone-induced increase in expression of stromal growth factor Fgf10 which, together with prominent stromal GR expression, suggest that glucocorticoid modify stromal-to-epithelial signaling in the mouse prostate. The mitogenic effects were prostate specific and not mediated by systemic effects on testosterone production suggesting that corticosterone effects were primarily mediated via prostate GR expression. CONCLUSION: These data demonstrate that murine prostate is significantly and directly influenced by corticosterone treatment via aberrant stromal-to-epithelial growth factor signaling.

The contradictory role of androgens in cutaneous and major burn wound healing.

Wound healing is a complex process involving four overlapping phases: haemostasis, inflammation, cell recruitment and matrix remodeling. In mouse models, surgical, pharmacological and genetic approaches targeting androgen actions in skin have shown that androgens increase interleukin-6 and tumor necrosis factor-α production and reduce wound re-epithelization and matrix deposition, retarding cutaneous wound healing. Similarly, clinical studies have shown that cutaneous wound healing is slower in men compared to women. However, in major burn injury, which triggers not only local wound-healing processes but also systemic hypermetabolism, the role of androgens is poorly understood. Recent studies have claimed that a synthetic androgen, oxandrolone, increases protein synthesis, improves lean body mass and shortens length of hospital stay. However, the possible mechanisms by which oxandrolone regulates major burn injury have not been reported. In this review, we summarize the current findings on the roles of androgens in cutaneous and major burn wound healing, as well as androgens as a potential therapeutic treatment option for patients with major burn injuries.

Severe subfertility in mice with androgen receptor inactivation in sex accessory organs but not in testis.

Androgen action on sex accessory organs influences rodent fertility, but the mechanisms remain unclear and investigation is difficult without the ability to restrict androgen action in specific tissues. We used Cre-LoxP technology to generate male mice with prostate epithelial-specific androgen receptor deficiency (denoted PEARKO). In addition to prostate, these males have reduced androgen action due to tissue-selective androgen receptor inactivation in seminal vesicle, epididymis, and vas deferens, whereas the testis is unaffected. We find that fertility of PEARKO males was severely reduced, compared with littermates with prominent defects in copulatory plug formation, which were smaller, softer, and more friable than controls. Despite normal testis sperm production, sperm numbers were reduced in caput but increased in cauda epididymis, suggesting alterations in sperm epididymal transit kinetics associated with increased rate of spontaneous acrosome reaction and abnormal flagellar morphology in PEARKO cauda epididymal sperm. Whereas the quantitative in vitro fertilizing ability of PEARKO epididymal sperm was normal, fewer fertilized oocytes were flushed from the oviducts of females after natural mating with PEARKO males. These data show that sperm formed in mice with impaired androgen action restricted to accessory glands and epididymis are quantitatively normal in number and in vitro fertilizing function but that severe in vivo subfertility reflects other functions related to sperm transport and survival in female reproductive tract that determine fertility in vivo.

Disruption of prostate epithelial androgen receptor impedes prostate lobe-specific growth and function.

Prostate development and maturation requires stromal-epithelial interactions and androgen action via the androgen receptor (AR) within these compartments. However, the specific roles of epithelial and stromal AR in postnatal prostate differentiation are unclear. We used Cre-LoxP technology to determine the prostate phenotype in mice with epithelial-selective genetic inactivation of the AR leaving the stromal AR functionally intact. We find that prostate development abolished in mice globally lacking a functional AR can be rescued by restricting the AR knockout to the postnatal prostate epithelium. We show that, at 8 wk of age, prostate epithelial AR knockout (PEARKO) mice exhibit prostate development with normal branching morphogenesis but lobe-specific decrease in prostate weight and hindered structural and functional differentiation of the mature prostate epithelium. No change was observed in PEARKO testis weight or serum testosterone compared with littermate controls. The most striking change was increased proliferation and abnormal lesions of epithelial cells predominantly in the anterior lobe of PEARKO mice. These findings highlight the vital role of stromal AR in postnatal prostate growth and structural differentiation and emphasize the requirement of epithelial AR in maintaining functional differentiation and restraining proliferation of epithelial cells in a lobe-specific manner. This unique PEARKO mouse provides a new paradigm with which to define the molecular mechanisms of the androgen signaling in mature prostate lobes in vivo and provides insight into the identification of better targets for treatment of prostate cancer and hyperplasia.

Differences in acute toxicity syndromes of 2,3,7,8-tetrachlorodibenzo-p-dioxin and 1,2,3,4,7,8-hexachlorodibenzo-p-dioxin in rats.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is the most potent congener of polychlorinated dibenzo-p-dioxins. The potency of 1,2,3,4,7,8-hexachlorodibenzo-p-dioxin (HxCDD) is only 10% of that of TCDD for typical aryl hydrocarbon receptor (AHR)-mediated effects. Acute lethality, macroscopic effects, and liver toxicity of TCDD and HxCDD were compared in male rats of the strain Han/Wistar (Kuopio; H/W), and of the lines A and B. The latter two rat lines originate from crossbreeding of H/W and Long-Evans (Turku/AB) rats. H/W and line A rats are highly resistant to acute toxicity of TCDD due to an altered AHR, while line B rats are moderately resistant due to H/W-type alleles of another, yet unidentified gene contributing to TCDD resistance ("gene B"). The rats received 200-10,000 microg/kg of either TCDD or HxCDD intragastrically and were monitored for 46 days. In all rats, the highest dose of HxCDD (10,000 microg/kg) reduced body weight more effectively than an identical dose of TCDD. Only HxCDD (10,000 microg/kg) caused gastrointestinal hemorrhage, pale (fatty) livers and death by day 15 in H/W and line A rats. In line B rats, HxCDD caused pronounced hepatic fatty degeneration, whereas TCDD induced hepatic accumulation of biliverdin and its derivatives. Both congeners induced sinusoidal distension in liver. In H/W and line A rats, the estimated LD(50) values were >10,000 microg/kg and 2000-10,000 microg/kg for TCDD and HxCDD, respectively; for line B rats they were 480 microg/kg and 1000-2000 microg/kg, respectively. Thus, HxCDD was more potent than TCDD in inducing acute mortality in H/W and line A rats, contrary to what is predicted by toxic equivalency factor (TEF) values. In line B, the expected rank order of potencies prevailed. These findings suggest that in addition to the canonical AHR-mediated toxic pathways, HxCDD possesses an AHR-independent mechanism of toxicity, whose main manifestations are rapid body weight loss, mortality, fatty liver and gastrointestinal hemorrhage.

Androgen action in female reproductive physiology.

PURPOSE OF REVIEW: In the last decade, it has been proven that androgens acting via the androgen receptor (AR) play an important role in the regulation of female reproductive function. However, the specific site of action and the precise pathways involved remain to be fully elucidated. This review aims to combine findings from emerging basic research to provide new insights into the roles of AR-mediated actions, and the mechanisms involved, in normal ovarian, uterine, and mammary gland function. RECENT FINDINGS: Our understanding of the specific roles of androgens in females has been hindered as females with complete androgen insensitivity cannot be generated by natural breeding, and interpretation of results from pharmacological studies has led to confusion as some androgens can be converted into estrogens, which can mediate actions via estrogen receptors. However, with the creation of global and cell-specific female AR knockout mouse models by Cre-LoxP technology, and the use of aromatizable and nonaromatizable androgens, novel roles for androgens in the regulation of female reproductive physiology have been revealed. SUMMARY: AR-mediated mechanisms play important roles in mediating normal ovarian, uterine, and mammary gland function and there is hope that further elucidation of the role of androgens in female reproductive physiology may translate into the development of novel, evidence-based, and targeted treatment for androgen-associated conditions.

Glandular epithelial AR inactivation enhances PTEN deletion-induced uterine pathology.

Phosphatase and tensin homolog (PTEN) deletion induces uterine pathology, whereas androgen actions via androgen receptor (AR) support uterine growth and therefore may modify uterine cancer risk. We hypothesized that the androgen actions mediated via uterine glandular epithelial AR could modify PTEN deletion-induced uterine pathology. To test our hypothesis, we developed uterine glandular epithelium-specific PTEN and/or AR knockout mouse models comparing the uterine pathology among wild-type (WT), glandular epithelium-specific AR inactivation (ugeARKO), PTEN deletion (ugePTENKO), and the combined PTEN and AR knockout (ugePTENARKO) female mice. The double knockout restricted to glandular epithelium showed that AR inactivation enhanced PTEN deletion-induced uterine pathology with development of intraepithelial neoplasia by 20 weeks of age. In ugePTENARKO, 6/10 (60%) developed intraepithelial neoplasia, whereas 3/10 (30%) developed only glandular hyperplasia in ugePTENKO uterus. No uterine pathology was observed in WT (n=8) and ugeARKO (n=7) uteri. Uterine weight was significantly (P=0.002) increased in ugePTENARKO (374±97 mg (mean±s.e.)) compared with WT (97±6 mg), ugeARKO (94±12 mg), and ugePTENKO (205±33 mg). Estrogen receptor alpha (ERα) and P-AKT expression was modified by uterine pathology but did not differ between ugePTENKO and ugePTENARKO, suggesting that its expressions are not directly affected by androgens. However, progesterone receptor (PR) expression was reduced in ugePTENARKO compared to ugePTENKO uterus, suggesting that PR expression could be regulated by glandular epithelial AR inactivation. In conclusion, glandular epithelial AR inactivation (with persistent stromal AR action) enhanced PTEN deletion-induced uterine pathology possibly by downregulating PR expression in the uterus.

Effects of in utero and lactational TCDD exposure on bone development in differentially sensitive rat lines.

2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is a notorious model compound of highly toxic environmental pollutants, polychlorinated dibenzo-p-dioxins (PCDDs). Their toxic effects are mediated via cytosolic aryl hydrocarbon receptor (AHR). We studied the effects of several dose levels of TCDD on developing rat bone after maternal exposure at different times of gestation and lactation in three differentially sensitive rat lines. Rat lines A, B, and C differ in their sensitivity to TCDD due to mutated AHR (Ahr(hw)) in line A and another TCDD-resistance allele (B(hw)) in line B. Line C rats have no resistance alleles. Offspring were analyzed for bone mineral density and geometry by peripheral quantitative computed tomography (pQCT) and for bone biomechanics by three-point bending at mid-diaphysis of tibia and femur and by axial loading at femoral neck. TCDD treatment resulted in bone defects, mainly in offspring of the most sensitive line C at a maternal dose of 1 microg/kg. They included decreased bone length, cross-sectional area of cortex, and bone mineral density. Mechanical testing revealed significantly reduced bending breaking force and stiffness of tibia, femur, and femoral neck. The effects were exposure time-dependent, and earlier exposure caused more severe defects. Gestational exposure alone was not sufficient, but lactational exposure was required to cause the bone defects. Most of the defects were recovered at the age of 1 year. The results indicate that dioxins affect developing bone by interfering with bone growth and mechanical strength and that the effects are mainly reversible. The dioxin-resistance alleles, Ahr(hw) and B(hw) increase the resistance to these defects.