Search

Introduction. The aim of this study was to determine estrone (E1), 17β-estradiol (E2) and progesterone (P4) concentrations in processed milk with different fat contents and to compare the concentrations of these hormones in commercial ultrahigh temperature (UHT) processed milk and commercial pasteurized milk. Materials and Methods. Commercial milks with different fat contents (UHT 0.5 %, UHT 1.5 %, UHT 3.5 % and pasteurized 3.5 % (10 samples of each type of milk)) were purchased in local stores. E1, E2 and P4 concentrations were determined by commercial ELISA kits. Results and Conclusions. E1 concentrations were below the limit of detection (15 pg mL-1) in all milks except in two UHT 3.5 % (out of 10) and two pasteurized 3.5 % (out of 10) milk samples. Mean E2 and P4 concentrations in UHT 3.5 % milk (25.37 ± 1.15 pg mL-1 and 10.76 ± 0.43 ng mL-1, respectively) were significantly higher than in UHT 0.5 % milk (19.38 ± 0.79 pg mL-1 and 7.06 ± 0.26 ng mL-1, respectively). Significant positive correlations were determined between hormone concentrations and milk fat contents. Relatively high E2 and P4 concentrations indicate that the bulk of milk in the commercial milks examined originated from pregnant cows.

Cartilage is an avascular tissue with a limited rate of oxygen and nutrient diffusion, resulting in its inability to heal spontaneously. Articular cartilage defects eventually lead to osteoarthritis (OA), the endpoint of progressive destruction of cartilage. In companion animals, OA is the most common joint disease, and many pain management and surgical attempts have been made to find an appropriate treatment. Pain management of OA is usually the first choice of OA therapy, which is often managed with nonsteroidal anti-inflammatory drugs (NSAIDs). To avoid known negative side effects of NSAIDs, other approaches are being considered, such as the use of anti-nerve growth factor monoclonal antibodies (anti-NGF mAB), hyaluronic acid (HA), platelet-rich plasma (PRP), and mesenchymal stem cells (MSCs). The latter is increasingly being recognized as effective in reducing or even eliminating pain and lameness associated with OA. However, the in vivo mechanisms of MSC action do not relate to their differentiation potential, but rather to their immunomodulatory functions. Achieving actual regeneration of cartilage to prevent OA from developing or even revert already existing OA condition has not yet been achieved. Several techniques have been tried to overcome cartilage's inability to regenerate, from osteochondral transplantation, autologous chondrocyte implantation (ACI), and matrix-induced ACI (MACI). Combinatory use of MSCs unique features and biomaterials is also being investigated with the aim to as much as possible recapitulate the native microenvironment of the cartilage, yet so far none of the methods have produced reliable and truly effective results. Although OA, for now, remains an incurable disease, novel techniques are being developed, rendering hope for the future accomplishment of actual cartilage regeneration. The aim of this chapter is firstly to summarize known and developing pain management options for OA, secondly to present surgical attempts to regenerate articular cartilage, and finally to present the attempts to improve existing regenerative treatment options using mesenchymal stem cells, with the vision for the possible use of developing strategies in veterinary medicine., (© 2022. Springer Nature Switzerland AG.)

Remarkable immunomodulatory abilities of mesenchymal stem cells, also called multipotent mesenchymal stromal cells or medicinal signaling cells (MSCs), have entailed significant advances in veterinary regenerative medicine in recent years. Despite positive outcomes from MSC therapies in various diseases in dogs and cats, differences in MSC characteristics between small animal veterinary patients are not well-known. We performed a comparative study of cells' surface marker expression, viability, proliferation, and differentiation capacity of adipose-derived MSCs (ADMSCs) from dogs and domestic cats. The same growth media and methods were used to isolate, characterize, and culture canine and feline ADMSCs. Adipose tissue was collected from 11 dogs and 8 cats of both sexes. The expression of surface markers CD44, CD90, and CD34 was detected by flow cytometry. Viability at passage 3 was measured with the hemocytometer and compared to the viability measured by flow cytometry after 1 day of handling. The proliferation potential of MSCs was measured by calculating cell doubling and cell doubling time from second to eighth passage. Differentiation potential was determined at early and late passages by inducing cells toward adipogenic, osteogenic, and chondrogenic differentiation using commercial media. Our study shows that the percentage of CD44 + CD90 + and CD34 -/- cells is higher in cells from dogs than in cells from cats. The viability of cells measured by two different methods at passage 3 differed between the species, and finally, canine ADMSCs possess greater proliferation and differentiation potential in comparison to the feline ADMSCs., Competing Interests: GM is a partial owner of Animacel Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Voga, Kovač and Majdic.)

COVID-19 morbidity and mortality have significant gender disparities, with higher prevalence and mortality in men. SARS-CoV-2 enters the lungs through the ACE2 enzyme, a member of the renin-angiotensin system (RAS). Although there are no data for the lung, the expressions of RAS components in other tissues are modulated by sex hormones, androgens, and estrogens. However, there are no data on sex-specific differences in ACE2 expression. If there is a sex difference in the expression of ACE2 in the lung, this could theoretically explain the gender disparity in COVID-19 disease. More importantly, although modulation of ACE2 will certainly not provide a cure for the COVID-19 disease, modulation of ACE2 by sex hormone modulators, if they affect the expression of ACE2, could potentially be developed into a supportive therapy for COVID-19 patients., (Copyright © 2020 Majdic.)

Regenerative medicine is a branch of medicine that develops methods to grow, repair, or replace damaged or diseased cells, organs or tissues. It has gained significant momentum in recent years. Stem cells are undifferentiated cells with the capability to self-renew and differentiate into tissue cells with specialized functions. Stem cell therapies are therefore used to overcome the body's inability to regenerate damaged tissues and metabolic processes after acute or chronic insult. The concept of stem cell therapy was first introduced in 1991 by Caplan, who proposed that massive differentiation of cells into the desired tissue could be achieved by isolation, cultivation, and expansion of stem cells in in vitro conditions. Among different stem cell types, mesenchymal stem cells (MSC) currently seem to be the most suitable for therapeutic purposes, based on their simple isolation and culturing techniques, and lack of ethical issues regarding their usage. Because of their remarkable immunomodulatory abilities, MSCs are increasingly gaining recognition in veterinary medicine. Developments are primarily driven by the limitations of current treatment options for various medical problems in different animal species. MSCs represent a possible therapeutic option for many animal diseases, such as orthopedic, orodental and digestive tract diseases, liver, renal, cardiac, respiratory, neuromuscular, dermal, olfactory, and reproductive system diseases. Although we are progressively gaining an understanding of MSC behavior and their mechanisms of action, some of the issues considering their use for therapy are yet to be resolved. The aim of this review is first to summarize the current knowledge and stress out major issues in stem cell based therapies in veterinary medicine and, secondly, to present results of clinical usage of stem cells in veterinary patients., (Copyright © 2020 Voga, Adamic, Vengust and Majdic.)

Under appropriate culture conditions, mesenchymal stem cells (MSC), also called more properly multipotent mesenchymal stromal cells (MMSC), can be induced toward differentiation into different cell lineages. In order to guide stem cell fate within an environment resembling the stem cell niche, different biomaterials are being developed. In the present study, we used silk fibroin (SF) as a biomaterial supporting the growth of MMSC and studied its effect on chondrogenesis of canine adipose-derived MMSC (cADMMSC). Adipose tissue was collected from nine privately owned dogs. MMSC were cultured on SF films and SF scaffolds in a standard cell culture medium. Cell morphology was evaluated by scanning electron microscopy (SEM). Chondrogenic differentiation was evaluated by alcian blue staining and mRNA expression of collagen type 1, collagen type 2, Sox9, and Aggrecan genes. cADMMSC cultured on SF films and SF scaffolds stained positive using alcian blue. SEM images revealed nodule-like structures with matrix vesicles and fibers resembling chondrogenic nodules. Gene expression of chondrogenic markers Sox9 and Aggrecan were statistically significantly upregulated in cADMMSC cultured on SF films in comparison to negative control cADMMSC. This result suggests that chondrogenesis of cADMMSC could occur when cells were grown on SF films in a standard cell culture medium without specific culture conditions, which were previously considered necessary for induction of chondrogenic differentiation., Competing Interests: Declaration of conflicting interests: The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Female sexual behavior is a complex process regulated by multiple brain circuits and influenced by sex steroid hormones acting in the brain. Several regions in the hypothalamus have been implicated in the regulation of female sexual behavior although a complete circuitry involved in female sexual behavior is not understood. Fez family zinc finger 1 (Fezf1) gene is a brain specific gene that has been mostly studied in the context of olfactory development, although in a recent study, FEZF1 has been identified as one of the genes responsible for the development of Kallman syndrome. In the present study, we utilized shRNA approach to downregulate Fezf1 in the ventromedial nucleus of the hypothalamus (VMN) with the aim to explore the role of this gene. Adult female mice were stereotaxically injected with lentiviral vectors encoding shRNA against Fezf1 gene. Mice injected with shRNA against Fezf1 had significantly reduced female sexual behavior, presumably due to the downregulation of estrogen receptor alpha (ERα), as the number of ERα-immunoreactive cells in the VMN of Fezf1 mice was significantly lower in comparison to controls. However, no effect on body weight or physical activity was observed in mice with downregulated Fezf1, suggesting that the role of Fezf1 in the VMN is limited to the regulation of sexual behavior., Significance Statement: Fezf1 gene has been identified in the present study as a regulator of female sexual behavior in mice. Regulation of the female sexual behavior could be through the regulation of estrogen receptor alpha expression in the ventromedial nucleus of the hypothalamus, as the expression of this receptor was reduced in mice with downregulated Fezf1. As expression of Fezf1 is very specific in the brain, this gene could present a potential target for the development of novel drugs regulating hypoactive sexual desire disorder in women, if similar function of FEZF1 will be confirmed in humans., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Sex differences are consistently reported in different visuospatial tasks with men usually performing better in mental rotation tests while women are better on tests for memory of object locations. In the present study, we investigated sex differences in solving jigsaw puzzles in children. In total 22 boys and 24 girls were tested using custom build tablet application representing a jigsaw puzzle consisting of 25 pieces and featuring three different pictures. Girls outperformed boys in solving jigsaw puzzles regardless of the picture. Girls were faster than boys in solving the puzzle, made less incorrect moves with the pieces of the puzzle, and spent less time moving the pieces around the tablet. It appears that the strategy of solving the jigsaw puzzle was the main factor affecting differences in success, as girls tend to solve the puzzle more systematically while boys performed more trial and error attempts, thus having more incorrect moves with the puzzle pieces. Results of this study suggest a very robust sex difference in solving the jigsaw puzzle with girls outperforming boys by a large margin.

Women and men, female and male animals and cells are biologically different, and acknowledgement of this fact is critical to advancing medicine. However, incorporating concepts of sex-specific analysis in basic research is largely neglected, introducing bias into translational findings, clinical concepts and drug development. Research funding agencies recently approached these issues but implementation of policy changes in the scientific community is still limited, probably due to deficits in concepts, knowledge and proper methodology. This expert review is based on the EUGenMed project (www.eugenmed.eu) developing a roadmap for implementing sex and gender in biomedical and health research. For sake of clarity and conciseness, examples are mainly taken from the cardiovascular field that may serve as a paradigm for others, since a significant amount of knowledge how sex and oestrogen determine the manifestation of many cardiovascular diseases (CVD) has been accumulated. As main concepts for implementation of sex in basic research, the study of primary cell and animals of both sexes, the study of the influence of genetic vs. hormonal factors and the analysis of sex chromosomes and sex specific statistics in genome wide association studies (GWAS) are discussed. The review also discusses methodological issues, and analyses strength, weaknesses, opportunities and threats in implementing sex-sensitive aspects into basic research., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2017. For permissions, please email: journals.permissions@oup.com.)

Although considerable progress has been made in our understanding of brain function, many questions remain unanswered. The ultimate goal of studying the brain is to understand the connection between brain structure and function and behavioural outcomes. Since sex differences in brain morphology were first observed, subsequent studies suggest different functional organization of the male and female brains in humans. Sex and gender have been identified as being a significant factor in understanding human physiology, health and disease, and the biological differences between the sexes is not limited to the gonads and secondary sexual characteristics, but also affects the structure and, more crucially, the function of the brain and other organs. Significant variability in brain structures between individuals, in addition to between the sexes, is factor that complicates the study of sex differences in the brain. In this review, we explore the current understanding of sex differences in the brain, mostly focusing on preclinical animal studies., (© 2016 The Author(s). published by Portland Press Limited on behalf of the Biochemical Society.)

The molecular mechanisms that underlie spleen development and congenital asplenia, a condition linked to increased risk of overwhelming infections, remain largely unknown. The transcription factor TLX1 controls cell fate specification and organ expansion during spleen development, and Tlx1 deletion causes asplenia in mice. Deregulation of TLX1 expression has recently been proposed in the pathogenesis of congenital asplenia in patients carrying mutations of the gene-encoding transcription factor SF-1. Herein, we have shown that TLX1-dependent regulation of retinoic acid (RA) metabolism is critical for spleen organogenesis. In a murine model, loss of Tlx1 during formation of the splenic anlage increased RA signaling by regulating several genes involved in RA metabolism. Uncontrolled RA activity resulted in premature differentiation of mesenchymal cells and reduced vasculogenesis of the splenic primordium. Pharmacological inhibition of RA signaling in Tlx1-deficient animals partially rescued the spleen defect. Finally, spleen growth was impaired in mice lacking either cytochrome P450 26B1 (Cyp26b1), which results in excess RA, or retinol dehydrogenase 10 (Rdh10), which results in RA deficiency. Together, these findings establish TLX1 as a critical regulator of RA metabolism and provide mechanistic insights into the molecular determinants of human congenital asplenia.

Steroidogenic factor 1 (SF-1), officially designated NR5A1, is essential for gonadal and adrenal development and for the normal structure of the ventromedial hypothalamus (VMH), as demonstrated by SF-1 knockout mice (SF-1 KO), but much less is known about the possible effects of haploinsufficiency of the SF-1 gene. In the present study, maternal behavior in SF-1 KO heterozygous mice was evaluated. Behavioral tests revealed that SF-1 KO heterozygous females have impaired maternal behavior. In comparison to wild-type (WT) females, SF-1 KO heterozygous females retrieved significantly fewer pups into their nests, latency to retrieve and crouch over the pups was longer, and their nests were lower quality. As suggested by previous studies full dosage of SF-1 gene is needed for appropriate stress response and expression of brain-derived neurotrophic factor (BDNF) in the brain, and this might present a mechanism through which maternal behavior in SF-1 KO heterozygous females is impaired.

Kisspeptin, a regulator of reproductive function and puberty in mammals, is expressed in the rostral (anteroventral) periventricular nucleus (AVPV) and arcuate nucleus (Arc), and its expression is at least partially regulated by estradiol in rodents. The aim of the present study was to determine contributions of genetic factors and gonadal steroid hormones to the sexual differentiation of kisspeptin-immunoreactive (kisspeptin-ir) cell populations in the AVPV and Arc during postnatal development using agonadal steroidogenic factor 1 (SF-1) knockout (KO) mice. To examine the effects of gonadal hormones on pubertal development of kisspeptin neurons, SF-1 KO mice were treated with estradiol benzoate (EB) from postnatal day (P)25 to P36, and their brains were examined at P36. No sex differences were observed in the SF-1 KO mice during postnatal development and after treatment with EB - which failed to increase the number of kisspeptin-ir cells at P36 to the levels found in wild-type (WT) control females. This suggests that specific time periods of estradiol actions or other factors are needed for sexual differentiation of the pattern of immunoreactive kisspeptin in the AVPV. Kisspeptin immunoreactivity in the Arc was significantly higher in gonadally intact WT and SF-1 KO females than in male mice at P36 during puberty. Further, in WT and SF-1 KO females, but not in males, adult levels were reached at P36. This suggests that maturation of the kisspeptin system in the Arc differs between sexes and is regulated by gonad-independent mechanisms., (© 2015 S. Karger AG, Basel.)

The presence of steroidogenic enzymes in the brain suggests de novo synthesis of steroid hormones in the brain. The current study was designed to determine the developmental profiles of cytochrome p450 aromatase (cyp19), 17β-hydroxysteroid dehydrogenase (17β-HSD), 5α-reductase type I and 3α-hydroxysteroid dehydrogenase (3α-HSD) mRNA expression levels in the fetal mouse brain and potential influence of peripheral steroids, and the steroidogenic factor 1 (SF-1) gene on their expression. Brains were collected from WT and SF-1 knockout male and female fetuses at embryonic (E) days E12, E14, E16, and E18. Quantitative PCR analyses revealed age related increases in the expression levels of 17β-HSD and 5α-reductase. Differences between genotypes in the expression levels of 17β-HSD and 5α-reductase were detected on E14, with reduced levels of expression in SF-1 KO males and females for 17β-HSD and only between females for 5α-reductase. Expression of 3α-HSD mRNA did not differ significantly between sexes, age groups or genotypes with the exception of SF-1 KO males, which had an unexplained increase in mRNA for this enzyme on day E18. Expression of cyp19 was at the limit of detection and could not be analyzed effectively. There were no sex differences and, with the exception of small difference on E14 for 17β-HSD and 5α-reductase, no differences between genotypes. The results suggest that gonadal steroids do not influence the production of neurosteroids in the fetal brain, nor does SF-1 play a major role in the regulation of steroidogenic enzyme expression in the brain.

Exposure to stress during puberty can lead to long-term behavioral alterations in adult rodents coincident with sex steroid hormone-dependent brain remodeling and reorganization. Social isolation is a stress for social animals like mice, but little is known about the effects of such stress during adolescence on later reproductive behaviors. The present study examined sexual behavior of ovariectomized, estradiol and progesterone primed female mice that were individually housed from 25 days of age until testing at approximately 95 days, or individually housed from day 25 until day 60 (during puberty), followed by housing in social groups. Mice in these isolated groups were compared to females that were group housed throughout the experiment. Receptive sexual behaviors of females and behaviors of stimulus males were recorded. Females housed in social groups displayed greater levels of receptive behaviors in comparison to both socially isolated groups. Namely, social females had higher lordosis quotients (LQs) and more often displayed stronger lordosis postures in comparison to isolated females. No differences between female groups were observed in stimulus male sexual behavior suggesting that female "attractiveness" was not affected by their social isolation. Females housed in social groups had fewer cells containing immunoreactive estrogen receptor (ER) α in the anteroventral periventricular nucleus (AVPV) and in the ventromedial nucleus of the hypothalamus (VMH) than both isolated groups. These results suggest that isolation during adolescence affects female sexual behavior and re-socialization for 1 month in adulthood is insufficient to rescue lordosis behavior from the effects of social isolation during the pubertal period.

Sex steroid hormones secreted by gonads influence development and expression of many behaviors including parental behaviors. The capacity to display many behaviors develops under the influence of sex steroid hormones; it begins with gonadal differentiation and lasts through puberty. The timing of gonadectomy may have important and long lasting effects on the organization and activation of neural circuits regulating the expression of different behaviors. The present study investigated the importance of exposure to endogenous gonadal steroid hormones during pubertal period/adolescence on parental behavior in adult mice. Male and female WT mice were gonadectomized either before puberty (25 days of age) or after puberty (60 days of age) and tested for parental behavior with and without estradiol benzoate (EB) replacement in adulthood. Additional groups of mice were gonadectomized at P25 and supplemented with estradiol (females) or testosterone (males) during puberty. Female mice gonadectomized after puberty or gonadectomized before puberty and supplemented with estradiol during puberty, displayed better pup directed parental behaviors in comparison to mice gonadectomized at 25 days of age regardless of treatment with estradiol in adulthood. However, mice treated with EB in adulthood displayed better non-pup directed nest building behavior than when they were tested without EB treatment regardless of sex and time of gonadectomy. To examine whether the sensitivity to sex steroid hormones was altered due to differences in time without gonads prior to the testing, mice were also tested for female sex behavior and there were no differences between mice gonadectomized at P25 or P60, although this could not completely rule out the possibility that parental behavior is more sensitive to prolonged absence of steroid hormones than female sex behavior. These results suggest that the absence of gonads and thereby the absence of appropriate gonadal steroid hormones during puberty/adolescence may have a profound effect on pup directed parental behaviors in adult mice., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Neurons of the paraventricular nucleus of the hypothalamus (PVN) regulate the hypothalamic- pituitary-adrenal (HPA) axis and the autonomic nervous system. Females lacking functional GABA(B) receptors because of a genetic disruption of the R1 subunit have altered cellular characteristics in and around the PVN at birth. The genetic disruption precluded appropriate assessments of physiology or behavior in adulthood. The current study was conducted to test the long term impact of a temporally restricting pharmacological blockade of the GABA(B) receptor to a 7-day critical period (E11-E17) during embryonic development. Experiments tested the role of GABA(B) receptor signaling in fetal development of the PVN and later adult capacities for adult stress related behaviors and physiology. In organotypic slices containing fetal PVN, there was a female specific, 52% increase in cell movement speeds with GABA(B) receptor antagonist treatment that was consistent with a sex-dependent lateral displacement of cells in vivo following 7 days of fetal exposure to GABA(B) receptor antagonist. Anxiety-like and depression-like behaviors, open-field activity, and HPA mediated responses to restraint stress were measured in adult offspring of mothers treated with GABA(B) receptor antagonist. Embryonic exposure to GABA(B) receptor antagonist resulted in reduced HPA axis activation following restraint stress and reduced depression-like behaviors. There was also increased anxiety-like behavior selectively in females and hyperactivity in males. A sex dependent response to disruptions of GABA(B) receptor signaling was identified for PVN formation and key aspects of physiology and behavior. These changes correspond to sex specific prevalence in similar human disorders, namely anxiety disorders and hyperactivity.

Prion protein (Prp(c)) is involved in the etiology of prion neurodegenerative diseases in mammals. The biological functions of Prp(c) are still largely unknown despite many studies in recent years. Different studies have shown impairment in locomotion, emotional/social behaviors, sleep disorders and memory impairment in mice lacking the prion gene Prnp (Prnp(-/-)) but its exact functions in the brain are still unclear. In the present study, Zurich I Prnp(-/-) and their littermate wild type (WT) control male mice were behaviorally characterized for offensive aggressive behavior in a resident-intruder paradigm with the aim to establish the possible function of Prp(c) in the regulation of offensive aggressive behavior. Prnp(-/-) mice showed reduced latencies to the first attack and bite, higher percentage of mice biting and higher frequencies of attacks of stimulus males. These results show that Prnp(-/-) mice exhibit altered aggressive behavior in comparison to their WT controls and therefore suggest that lack of the Prnp either directly or indirectly affects brain circuitry responsible for the regulation of offensive aggressive behavior., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Decline in semen quality in humans and increased incidence of male reproductive problems could be caused by different factors, including pesticides that could mimic or block the action of endogenous hormones. If the decline in semen quality is real, and environmental chemicals are at least partially responsible for this decline, similar changes should be observed in animals that live in close connection with humans and are exposed to similar levels of pollutants. In the present study, the semen quality of bulls in the last 30 years was examined with respect to the year of birth. Furthermore, semen quality results were correlated to the total pesticide use in a limited geographical area. The results indicate a notable decrease in both ejaculate volume and total number of spermatozoa in ejaculates of bulls born in the late 1970s, while after that (until 2006) there was no obvious downward or upward trend either in ejaculate volume or in the total number of sperm cells. The amount of pesticides released into the environment increased about twofold in the given period, and linear regression analysis revealed a strong and statistically significant correlation between the amount of pesticides used and the total number of spermatozoa in the ejaculate.

Female receptivity including the immobile hormone-dependent lordosis posture is essential for successful reproduction in rodents. It is well documented that lordosis is organized during the perinatal period when the actions of androgens decrease the males' ability to display this behavior in adulthood. Conversely the absence of androgens, and the presence of low levels of prepubertal estrogens, preserve circuitry that regulates this behavior in females. The current study set out to determine whether sex chromosomal genes are involved in the differentiation of this behavior. An agonadal mouse model was used to test this hypothesis. The SF-1 gene (Nr5a1) is required for development of gonads and adrenal glands, and knockout mice are consequently not exposed to endogenous gonadal steroids. Thus contributions of sex chromosome genes can be disassociated from the actions of estrogens. Use of this model reveals a direct genetic contribution from sex chromosomes in the display of lordosis and other female-typical sexual behavior patterns. It is likely that the concentrations of gonadal steroids present during normal male development modify the actions of sex chromosome genes on the potential to display female sexual behavior., (Copyright © 2012 Elsevier Inc. All rights reserved.)

The cytochrome P450 (CYP) genes Cyp51, Cyp11a1, Cyp17a1, Cyb11b1, Cyp11b2 and Cyp21a1 are involved in the adrenal production of corticosteroids, whose circulating levels are circadian. cAMP signaling plays an important role in adrenal steroidogenesis. By using cAMP responsive element modulator (Crem) knockout mice, we show that CREM isoforms contribute to circadian expression of steroidogenic CYPs in the mouse adrenal gland. Most striking was the CREM-dependent hypomethylation of the Cyp17a1 promoter at zeitgeber time 12, which resulted in higher Cyp17a1 mRNA and protein expression in the knockout adrenal glands. The data indicate that products of the Crem gene control the epigenetic repression of Cyp17 in mouse adrenal glands., (© 2011 The Authors Journal compilation © 2011 FEBS.)

The essential role of the Crem gene in normal sperm development is widely accepted and is confirmed by azoospermia in male mice lacking the Crem gene. The exact number of genes affected by Crem absence is not known, however a large difference has been observed recently between the estimated number of differentially expressed genes found in Crem knock-out (KO) mice compared to the number of gene loci bound by CREM. We therefore re-examined global gene expression in male mice lacking the Crem gene using whole genome transcriptome analysis with Affymetrix microarrays and compared the lists of differentially expressed genes from Crem-/- mice to a dataset of genes where binding of CREM was determined by Chip-seq. We determined the global effect of CREM on spermatogenesis as well as distinguished between primary and secondary effects of the CREM absence. We demonstrated that the absence of Crem deregulates over 4700 genes in KO testis. Among them are 101 genes associated with spermatogenesis 41 of which are bound by CREM and are deregulated in Crem KO testis. Absence of several of these genes in mouse models has proven their importance for normal spermatogenesis and male fertility. Our study showed that the absence of Crem plays a more important role on different aspects of spermatogenesis as estimated previously, with its impact ranging from apoptosis induction to deregulation of major circadian clock genes, steroidogenesis and the cell-cell junction dynamics. Several new genes important for normal spermatogenesis and fertility are down-regulated in KO testis and are therefore possible novel targets of CREM.

The ventromedial nucleus of the hypothalamus (VMH) is a key cell group in the medial-basal hypothalamus that participates in the regulation of energy balance. Previous studies have shown that the cellular organization of the VMH is altered in mice with a disruption of the steroidogenic factor-1 (NR5a1) gene (SF-1 KO mice). The present study examined orexigenic/anorexigenic peptides (neuropeptide Y (NPY), agouti-related peptide (AgRP) and cocaine- and amphetamine-regulated transcript (CART)) and neural connections to and from the VMH in SF1 KO mice. NeuroVue tracing and Golgi staining were used to evaluate connections between the preoptic area (POA) and VMH and the orientation of dendrites in the VMH, respectively. Results of this study reveal changes in the cytoarchitecture of the region of the VMH with respect to the distribution of immunoreactive NPY, AgRP and CART. In WT mice projections from the POA normally surround the VMH while in SF-1 KO mice, projections from the POA stream through the region that would otherwise be VMH. Golgi impregnation of the region revealed fewer dendrites with ventrolateral orientations and in general, more variable dendritic orientations in SF-1 KO mice providing additional evidence that the connectivity of cells in the region is likely altered due to the cellular rearrangements consequent to disruption of the NR5a1 gene. In conclusion, this study greatly extends the data showing that the morphology of the regions containing the VMH is disrupted in SF-1 KO mice and suggests that changes in the location of cells or fibers containing NPY, AgRP and CART may, in part, account for changes in body weight homeostasis in these mice., (Copyright © 2011 Elsevier Inc. All rights reserved.)