The effects of low ambient temperature on steroidogenesis and mitochondrial functions in the testes of wild ground squirrels (Spermophilus dauricus).

Seasonal reproduction is a widely used breeding strategy in wildlife, especially vertebrates inhabiting temperate regions. Generally, ambient temperature is considered a significant factor influencing the reproductive status of animals. In the present study, wild ground squirrels (Spermophilus dauricus), typical seasonal breeders, were used as an animal model to investigate the mechanism behind the impact of low ambient temperature on testicular function. To simulate the winter environment of wild ground squirrels, we lowered the temperature gradient in the rearing environment to 4 °C. At sampling, the body surface temperature of the squirrels reared under normal ambient temperature (22 °C, NAT group) and the low ambient temperature (4 °C, LAT group) were 31.5 °C and 22.8 °C, respectively. Subsequently, we conducted immunohistochemical assays, qPCR, and enzyme-linked immunosorbent assays (ELISA) to examine the variations in testicular functions, as well as the dynamics and functions of mitochondria, in the squirrels of NAT and LAT groups. As a result, the levels of positive immunostaining for PCNA, P21, and P27 were significantly lower in the testes of LAT group, while the levels of immunostaining for Cleaved Caspase-3 and TUNEL were significantly higher. In addition, the low-temperature treatment reduced the expression level of steroidogenesis-related genes, including LHR, FSHR, GATA-4, P450scc, and P450arom, and decreased the testosterone concentration. Moreover, markers of mitochondrial fission and fusion, DRP1 and MFN2, respectively, were increased in the testes of LAT group. Additionally, the mRNA level of SOD1 was notably higher in the testes of LAT group. In conclusion, the low ambient temperature inhibited spermatogenesis, steroidogenesis, as well as mitochondrial dynamics and functions in the testes of wild ground squirrels.

Regulation and Role of Adiponectin Secretion in Rat Ovarian Granulosa Cells.

Adiponectin is an important adipokine involved in glucose and lipid metabolism, but its secretion and potential role in regulating glucose utilization during ovarian development remains unclear. This study aims to investigate the mechanism and effects of follicle-stimulating hormones (FSHs) on adiponectin secretion and its following impact on glucose transport in the granulosa cells of rat ovaries. A range of experimental techniques were utilized to test our research, including immunoblotting, immunohistochemistry, immunofluorescence, ELISA, histological staining, real-time quantitative PCR, and transcriptome analysis. The immunohistochemistry results indicated that adiponectin was primarily located in the granulosa cells of rat ovaries. In primary granulosa cells cultured in vitro, both Western blot and immunofluorescence assays demonstrated that FSH significantly induced adiponectin secretion within 2 h of incubation, primarily via the PKA signaling pathway rather than the PI3K/AKT pathway. Concurrently, the addition of the AdipoR1/AdipoR2 dual agonist AdipoRon to the culture medium significantly stimulated the protein expression of GLUT1 in rat granulosa cells, resulting in enhanced glucose absorption. Consistent with these in vitro findings, rats injected with eCG (which shares structural and functional similarities with FSH) exhibited significantly increased adiponectin levels in both the ovaries and blood. Moreover, there was a notable elevation in mRNA and protein levels of AdipoRs and GLUTs following eCG administration. Transcriptomic analysis further revealed a positive correlation between the expression of the intraovarian adiponectin system and glucose transporter. The present study represents a novel investigation, demonstrating that FSH stimulates adiponectin secretion in ovarian granulosa cells through the PKA signaling pathway. This mechanism potentially influences glucose transport (GLUT1) and utilization within the ovaries.

The association between several autophagy-related genes and their prognostic values in hepatocellular carcinoma: a study on the foundation of TCGA, GEPIA and HPA databases.

BACKGROUND: The purpose of this study was to investigate the relationship between the expression of autophagy-related genes and prognosis in hepatocellular carcinoma (HCC). METHODS AND RESULTS: We selected three autophagy-related genes (ATG3, ATG7, and ATG9A) from gene expression data of liver cancer patients in The Cancer Genome Atlas (TCGA) database by Kaplan-Meier survival analysis, univariate and multivariate Cox regression analysis, and Gene Set Enrichment Analysis (GSEA). Human Protein Atlas (HPA) and Gene Expression Profiling Interactive Analysis (GEPIA) databases were applied to testify the credibility of our results. The expression levels of ATG3, ATG7, and ATG9A were verified by real-time quantitative PCR (RT-qPCR) in normal liver cells (L02) and three HCC cell lines (HepG2, Hep3b, and Li-7). Data analysis results from TCGA showed high ATG3, ATG7, ATG9A expression in HCC tumor tissues. Kaplan-Meier survival analysis showed that the survival rate of the high expression group of ATG3, ATG7, and ATG9A was all significantly lower than the low expression group. GSEA analysis showed that many signaling pathways (such as the regulation of autophagy, glycine serine and threonine metabolism, pathways in cancer, mitogen-activated protein kinase (MAPK) signaling pathway, mammalian target of rapamycin (mTOR) signaling pathway, as well as P53 signaling pathway) were differentially enriched in HCCs with ATG3, ATG7, and ATG9A expression. GEPIA and RT-qPCR also identified that the mRNA expression level of ATG3, ATG7, and ATG9A in normal liver cells were significantly lower than in HCC cells. High protein expression of ATG3, ATG7, and ATG9A was displayed in HCCs from the HPA database. CONCLUSIONS: The ATG3, ATG7, ATG9A might be utilized as prognostic biomarkers for liver cancer.

Seasonal expression of extracellular signal regulated kinases in the colon of wild ground squirrels (Spermophilus dauricus).

BACKGROUND: The purpose of the experiment was to explore the localization and seasonal expression of extracellular signal regulated kinase (ERK) in the colonic tissue of wild ground squirrels (Spermophilus dauricus). METHODS AND RESULTS: Hematoxylin-eosin staining, immunohistochemistry, real-time quantitative PCR and Western blotting were used in this experiment. The histological results showed that the diameter of the colon lumen enlarged and the number of glandular cells increased in the non-breeding season. It was found in the immunochemical results that both ERK1/2 and pERK1/2 were expressed in the cytoplasm of goblet cells and intestinal epithelial cells, while pERK1/2 was also expressed in the nucleus of them. The immune localization of both was more obvious in the non-breeding season, especially in intestinal epithelial cells. Real-time quantitative PCR and Western blotting showed that ERK1/2 and pERK1/2 were seasonally highly expressed in the non-breeding season. CONCLUSIONS: The expression of ERK1/2 and pERK1/2 was seasonal changes and had significant increases in the non-breeding season. This study revealed that ERK1/2 had potential roles in the colon to the adaptation of seasonal changes in wild ground squirrels.

PTEN Inhibition Ameliorates Muscle Degeneration and Improves Muscle Function in a Mouse Model of Duchenne Muscular Dystrophy.

Duchenne muscular dystrophy (DMD) is caused by a mutation of the muscle membrane protein dystrophin and characterized by severe degeneration of myofibers, progressive muscle wasting, loss of mobility, and, ultimately, cardiorespiratory failure and premature death. Currently there is no cure for DMD. Herein, we report that skeletal muscle-specific knockout (KO) of the phosphatase and tensin homolog (Pten) gene in an animal model of DMD (mdx mice) alleviates myofiber degeneration and restores muscle function without increasing tumor incidence. Specifically, Pten KO normalizes myofiber size and prevents muscular atrophy, and it improves grip strength and exercise performance in mdx mice. Pten KO also reduces fibrosis and inflammation, and it ameliorates muscle pathology in mdx mice. Unbiased RNA sequencing reveals that Pten KO upregulates extracellular matrix and basement membrane components positively correlated with wound healing and suppresses negative regulators of wound healing and lipid biosynthesis, thus improving the integrity of muscle basement membrane at the ultrastructural level. Importantly, pharmacological inhibition of PTEN similarly ameliorates muscle pathology and improves muscle integrity and function in mdx mice. Our findings provide evidence that PTEN inhibition may represent a potential therapeutic strategy to restore muscle function in DMD.

Seasonal Change in Adiponectin Associated with Ovarian Morphology and Function in Wild Ground Squirrels (Citellus dauricus Brandt).

The goal of this study is to explore the relationship between altered circulating adiponectin concentration, ovarian tissue morphology, ovarian steroidogenesis, and sex hormone production in ovaries of wild ground squirrels. The ovarian mass differed significantly during the breeding and non-breeding seasons, and the circulating estradiol and progesterone concentrations were significantly higher in the breeding season, while the circulating adiponectin level was significantly lower. The expression levels of gonadotropin receptors (FSHR and LHR) and steroidogenic enzymes (StAR, P450scc, P450arom, and 3ß-HSD) were significantly higher during the breeding season. Comparing the ovarian transcriptome data of wild ground squirrels between the two periods, we found that some differentially expressed genes were enriched for ovarian steroidogenesis and the adipocytokine signaling pathway, which correlated with our present results. Notably, the MAPK signaling pathway was also enriched and its related genes (Erk1, p38 Mapk, Jnk) were up-regulated by qPCR during the non-breeding season. These findings suggested that adiponectin may be involved in the regulation of seasonal changes in the ovarian function of wild ground squirrels, possibly by acting on the MAPK signaling pathway to regulate sex steroidogenesis in the ovaries.

Seasonal changes of mitochondrial autophagy and oxidative response in the testis of the wild ground squirrels (Spermophilus dauricus).

Mitochondria are the main organelles for mammalian energy metabolism and have been implicated in the regulation of germ cell maintenance and spermatogenesis. However, little is known about the changes in the mitochondria of the testis of seasonal breeders. Here, we characterized the seasonal changes in the mitochondria in the testis of the wild ground squirrels. Increased testicle weight, seminiferous tubule diameter, and sperm count were observed in the wild ground squirrels at the breeding season. RNA-seq analysis of the wild ground squirrel testes revealed that mitochondrial-related genes were expressed differentially between the breeding and nonbreeding seasons. Immunohistochemical staining showed that key mitophagy factors including PINK1, MFN2, and PARKIN were highly expressed in various cell types of testis during the breeding season. In addition, the abundance and enzymatic activities of mitochondrial-localized antioxidative enzymes superoxide dismutase 2 (SOD2) and Catalase were decreased in the testis during the breeding season, suggesting a tightly controlled redox balance at least partially facilitated by mitophagy during the seasonal breeding. Taken together, our study reveals that mitochondrial autophagy and oxidative stress may be implicated in the seasonal reproductive recrudescence of the wild ground squirrels, which deepens our understanding of the mitochondrial regulation of seasonal reproductivity in wildlife and provides new insights into the development of potential therapeutic interventions of male infertility.

The seasonal profile of proliferation and apoptosis in the prostate gland of the wild ground squirrel (Spermophilus dauricus).

The seasonal cycle of growth and regression in the prostate gland of wild ground squirrel provide a unique research model to understand the morphological changes of prostate glands. Our previous studies showed that the local production of dihydrotestosterone could affect the morphology and function of the prostate gland in either an autocrine or paracrine manner. In the present study, we attempted to gain more insight into this process by investigating the expression of key factors implicated in cell proliferation, apoptosis, and the cell cycle, including mechanistic target of rapamycin (mTOR), cyclin-D2, p21, p27 and retinoblastoma 1 (pRB). Morphological and histological observations confirmed that the prostate increased significantly in both size and weight during the breeding season. Positive immunostaining for proliferating cell nuclear antigen (PCNA) was mainly localized to the prostate epithelial cells during the breeding season, which is significantly higher in the prostate gland during the breeding season (2470 ± 81/mm2) than that in the nonbreeding season (324 ± 54/mm2). However, there was no significant difference in the prostate gland when compared between the breeding and nonbreeding seasons, with regards to TUNEL staining. Moreover, cell cycle regulators were mainly localized to the epithelial cells, including mTOR, cyclin-D2, p21, p27 and pRB. the immunostaining of mTOR and cyclin D2 were stronger during the breeding season, whereas the immunostaining of p27 and pRB were stronger during the nonbreeding season. The mRNA expression levels of mTOR, cyclin D2, and PCNA, were higher during the breeding season while those of p27 and p21 were higher during the nonbreeding season. Collectively, this study profiled the distinct expression pattern of key cell cycle regulators throughout the breeding and nonbreeding seasons. Collectively, these factors may play important roles in regulating the seasonal growth and regression of the prostatic epithelium in the wild ground squirrel.

Seasonal expressions of prostaglandin E synthases and receptors in the prostate of the wild ground squirrel (Spermophilus dauricus).

The prostate gland is a male accessory reproductive gland, whose vitality and function are under tight regulation of different hormones. Prostaglandins E2 (PGE2) is one of the major products generated by the actions of cyclooxygenases (COX) and prostaglandin E synthase (PTGES) on arachidonic acid, and is involved in a number of physiological and pathological processes. In this study, we investigated the seasonal immunolocalizations and expressions of COX-1, COX-2 and PTGES, as well as PGE2 receptors (PTGERs) subtypes 1-4 (EP1, EP2, EP3, EP4) in the prostate of the wild ground squirrel. Histological examination observed enlarged prostatic lumens in the breeding season and significantly shrunken lumens in the nonbreeding season. COX-1, COX-2, PTGES and PTGERs were mainly localized in epithelial and stromal cells in the breeding and nonbreeding seasons. The mRNA expression levels of Cox-1, Cox-2, Ptges, Ptger2 (encoding EP2) and Ptger4 (encoding EP4) were higher in the prostate of the breeding season than in the nonbreeding season. The relative mRNA levels of Cox-1, Cox-2, Ptges, Ptger2 and Ptger4 were positively correlated with prostatic weights. In addition, both the prostatic and plasma concentrations of PGE2 were significantly higher in the breeding season compared to the nonbreeding season. These results suggested that PGE2 synthesis and signaling might play an important autocrine or paracrine role in the regulation of seasonal changes in the prostatic function of the wild ground squirrel.

Seasonal expressions of oxytocin and oxytocin receptor in the epididymides in the wild ground squirrels (Citellus Dauricus Brandt).

The aim of this study was to detect the seasonal expressions of oxytocin (OT), oxytocin receptor (OTR), extracellular signal-regulated kinase1 and 2 (ERK1/2) and phospho-ERK1/2 (pERK1/2) in the epididymis of the wild ground squirrels (Citellus Dauricus Brandt) during the breeding season and non-breeding season. Histological results showed that size, weight, cell number and lumen diameter of epididymis underwent acute seasonal changes, which were all peaked in the breeding season. Immunohistochemical results suggested that strong staining of OT, OTR, ERK1/2, and pERK1/2 were observed in the epithelial layer in the whole epididymis, along with intense OT and OTR signal in smooth muscle cell (Smc) in caudal epididymis in the breeding season. The protein expression levels of OTR, ERK1/2, and pERK1/2 in the epididymis were higher in the breeding season than those of the non-breeding season. Besides, hormone assay revealed that there was no significant serum concentration of OT in these two periods, while epididymal concentration showed higher value in the breeding season. In summary, the identified localization and local concentration of OT in the epididymis in the wild ground squirrel suggested that epididymis may assume as a source of OT, and OT could act via OTR to activate ERK1/2 signaling to regulate seasonal epididymal functions.

The role of the adiponectin system in acute fasting-impaired mouse ovaries.

Adiponectin (ADIPOQ, encoded by Adipoq) is an important white adipose-derived adipokine linked to energy homeostasis and reproductive function. This study aims to reveal the expression and role of the adiponectin system in the ovaries under acute malnutrition. In this study, 48-h food deprivation significantly inhibited ovarian growth by suppressing cell proliferation and inducing cell apoptosis in the ovaries of gonadotrophin-primed immature mice. It was also accompanied by significantly decelerated basic metabolism (glucose, triacylglycerol and cholesterol), varied steroid hormones (follicle-stimulating hormone, luteinizing hormone and estradiol) and vanishment of the peri-ovarian fat. It is noteworthy that after acute fasting, the adiponectin levels in ovaries rather than in blood were significantly elevated. Immunohistochemical study demonstrated that adiponectin and its receptors (ADIPOR1 and ADIPOR2) primarily appeared in ovarian somatic and/or germ cells, and their protein expressions were upregulated in the ovaries from fasted mice. Further in vitro study verified that ADIPOR1/2 agonist obviously inhibited follicle-stimulating hormone-induced oocyte meiotic resumption, while the antagonist significantly enhanced the percentage of oocyte maturation in the absence of follicle-stimulating hormone. Furthermore, the build up of peri-ovarian fat under physiological status in mice showed a positive correlation with both the hypertrophy of adipocytes and growth of ovaries. Taken together, these findings indicate that the upregulation of the adiponectin system disturbs the normal female reproductive function under the malnutrition status, and it may be associated with the loss of peri-ovarian fat depots.

Seasonal expressions of COX-1, COX-2 and EP4 in the uteri of the wild Daurian ground squirrels (Spermophilus dauricus).

Prostaglandins (PGs) play a pivotal role in uterine reproductive process including maternal recognition of pregnancy, cell proliferation, and myometrium contractions in mammals. In this study, we investigated the immunolocalizations and expression levels of Prostaglandin E2 synthases cyclo-oxygenase (COX)-1 and COX-2, as well as one of PGE2 receptor subtypes 4 (EP4) in the uteri of the wild Daurian ground squirrels (Spermophilus dauricus) during the breeding and non-breeding seasons. Histologically, the thickness of endometrium: myometrium ratio in the uteri of the breeding season was higher than that of the non-breeding season. The immunostainings of COX-1, COX-2 and EP4 were observed in stromal cells, glandular cells and myometrium cells in the breeding and non-breeding seasons. The protein and mRNA expression levels of COX-1, COX-2 and EP4 were higher in the uteri of the breeding season than those of in the non-breeding season. The mean mRNA levels of COX-1, COX-2 and EP4 were positively correlated with uterine weights. In addition, the PGE2 concentration of uterine tissues as well as plasma PGE2, 17ß-estradiol, progesterone, LH and FSH levels were also significantly higher in the breeding season compared to those of the non-breeding season. These results suggested that PGE2 might play an important autocrine or paracrine role in the regulation of seasonal changes in the uterine functions of the wild Daurian ground squirrels during the breeding and non-breeding seasons.

Seasonal expressions of luteinising hormone receptor, follicle-stimulating hormone receptor and prolactin receptor in the epididymis of the male wild ground squirrel (Spermophilus dauricus).

Luteinising hormone (LH), follicle-stimulating hormone (FSH) and prolactin (PRL) are pituitary-derived hormones and mediate their functions through LH receptor (LHR), FSH receptor (FSHR) and PRL receptor (PRLR) respectively. This study aimed to investigate the seasonal expression patterns of LHR, FSHR and PRLR in the epididymis of the male wild ground squirrel during the breeding and non-breeding seasons. Histologically, principal cells, basal cells, cilia and mature spermatozoa were found in the lumen of caput, corpus and cauda epididymidis in the breeding season, whereas in the non-breeding season, cilia and basal cells were rarely found and the epididymidal duct was devoid of spermatozoa. Immunohistochemical results showed that LHR, FSHR and PRLR were mainly present in the filamentous cytoplasm layer of epithelial cells of the caput, corpus and cauda epididymidis and FSHR and PRLR displayed stronger staining in the breeding season than in the non-breeding season. Furthermore, the mRNA and protein levels of FSHR and PRLR in all regions of epididymis as well as the levels of LHR in the caput and cauda epididymidis were higher during the breeding season. The protein levels of FSHR, LHR and PRLR were positively correlated with epididymal weight. Together, these results suggest that LHR, FSHR and PRLR may regulate epididymal functional changes in the male wild ground squirrel during its seasonal breeding cycle.

Seasonal expressions of growth hormone receptor, insulin-like growth factor 1 and insulin-like growth factor 1 receptor in the scented glands of the muskrats (Ondatra zibethicus).

The growth hormone (GH)/insulin-like growth factor-1 (IGF-1) system plays an important role in regulating the cellular growth and organ development. The present study investigated the seasonal expressions of growth hormone receptor (GHR), IGF-1 and insulin-like growth factor 1 receptor (IGF-1R) in the scented glands of the muskrats. Morphological changes in the scented glands of the muskrats were observed significantly between the breeding and non-breeding seasons. Immunohistochemically, the expressions of GH, GHR, IGF-1 and IGF-1R were found in glandular cells and epithelial cells of the scented glands in both seasons. The protein and mRNA expression levels of GHR, IGF-1 and IGF-1R in the scented glands during the breeding season were noticeably higher than those of the non-breeding season. In parallel, the levels of GH and IGF-1 in the sera and scented glands were remarkably higher during the breeding season. In addition, small RNA sequencing showed that the predicted targets of the significantly changed hsa-miR-5100 and mmu-miR-6937-5p might regulate the expressions of Ghr, Igf-1 or Igf-1r. These results suggested that the morphological changes in the scented glands of the muskrats during the different seasons might be related to the expression levels of GHR, IGF-1 and IGF-1R. Meanwhile, GHR/IGF-1 system might regulate the scented glandular functions via endocrine or autocrine/paracrine manners.

Seasonal expressions of androgen receptor, P450arom and estrogen receptors in the epididymis of the wild ground squirrel (Citellus dauricus Brandt).

The aim of this study was to investigate the seasonal expressions of androgen receptor (AR), estrogen receptors alpha and beta (ERα and ERß) and aromatase cytochrome P450 (P450arom) in the epididymis of the wild ground squirrel. Histologically, the epididymis was with larger duct diameter and cell population during the breeding season. AR was presented in the peritubular smooth muscle cells and epithelial cells in the whole epididymis with stronger staining in the breeding period. P450arom was intensely localized in epithelial cells and spermatozoa during the breeding season, absent in the non-breeding season and moderately stained in pre-hibernation. During the breeding season, ERα was intensely expressed in epithelial cytoplasm and/or nucleus, whereas in the non-breeding season and pre-hibernation, weaker staining signal was found in nucleus of epithelial cells. ERß was absent in the entire annual cycle by immunohistochemical and Real-time PCR detection. The mRNA levels of AR, P450arom and ERα were higher in the epididymis of the breeding season when compared to those of the non-breeding season and pre-hibernation. Taken together, these results suggest that epididymis of the wild ground squirrel is a primary target for androgen and estrogen, and the expression of P450arom represents that epididymis may be a potential source of estrogen.

The clinicopathological and prognostic significance of TP53 alteration in K27M mutated gliomas: an individual-participant data meta-analysis.

This study aimed to investigate the impact of TP53 alteration on survival and clinicopathological features of glioma patients with H3K27M mutations. An individual-participant-data (IPD) meta-analysis was performed to investigate the impact of TP53 alteration on survival and clinicopathological features of patients with H3K27M mutations. Three hundred thirty-one individual records from 12 eligible glioma studies involving the H3K27M mutation were finally included in our meta-analysis, and a pooled hazard ratio (HR) of 1.53 (95%CI, 1.10-2.11; P = 0.01) indicated that TP53 alterations were associated with a shorter overall survival. The pooled odds ratios (ORs) indicated that TP53 alterations were significantly associated with the age at diagnosis ≥ 7 years (OR = 1.97, 95%CI = 1.15-3.38, P = 0.01), the status of histone H3.3 mutations (OR = 9.15, 95%CI = 4.18-20.06, P < 0.00001), and high WHO grade histology (III + IV) (OR = 2.70, 95%CI = 1.33-5.48, P = 0.006). However, no association was found between TP53 alterations and gender or tumor location. This IPD meta-analysis suggests that TP53 alteration is a valuable predictor for the prognosis of patients with H3K27M mutated gliomas. TP53 alteration may be used for identifying a subset of patients who potentially benefit from targeted reactivation of TP53 activity.

Seasonal expression of 5α-reductases and androgen receptor in the prostate gland of the wild ground squirrel (Spermophilus dauricus).

The prostate gland is a male accessory reproductive gland that requires androgenic steroids for maintaining its vitality and function. The aim of the study was to investigate the localization and expressions of the androgen receptor (AR), SRD5A1 and SRD5A2 in the prostate glands of wild male ground squirrels during different reproductive periods by immunohistochemistry and real-time PCR. Gross mass of the prostate gland was higher in the breeding season than that of the non-breeding season. Histological observation showed that exocrine lumens in the prostate gland were enlarged in the breeding season and shrank in the non-breeding season. Immunohistochemical results demonstrated that AR was presented in both epithelial and stromal cells' nuclei during both the breeding and non-breeding seasons. Intriguingly, the positive staining of SRD5A1 and SRD5A2 was only found in the stromal cells. The mRNA expression of the three genes including Ar, Srd5a1 and Srd5a2 in the prostate gland was higher in the breeding season than those of the nonbreeding season. In addition, the circulating concentration of testosterone (T) and the concentration of dihydrotestosterone (DHT) in the prostate gland were also markedly higher in the breeding season than those of the nonbreeding season. These results suggest that expression levels of AR, SRD5A1 and SRD5A2, as well as DHT synthesis, might be correlated with seasonal changes in morphology and function of the prostate gland, indicating that DHT may serve as a pivotal regulator to affect the morphology and function of prostate gland via a autocrine/paracrine pathway in wild male ground squirrels.

Seasonal expression of P450arom and estrogen receptors in scented glands of muskrats (Ondatra zibethicus).

Male muskrats have one pair of scented glands that grow and involute annually. To investigate the annual changes in the scented gland, we measured the expressions of aromatase cytochrome P-450 (P450arom) and estrogen receptors (ERs) in the scented glands. P450arom was expressed in glandular cells and epithelial cells in the scented glands during the breeding season, and only in glandular cells during the nonbreeding season. ERα and ERß were also detected in different types of cells in the scented gland during the breeding and nonbreeding seasons. Both mRNA and protein levels of P450arom, ERα, and ERß were higher in the scented glandular tissues during the breeding season than those during the nonbreeding season. In addition, small RNA sequencing showed that the predicted targets of the significantly changed microRNAs might be the genes encoding P450arom and ERs. In conclusion, the seasonal changes in the expression of P450arom and ERs may be involved in the regulation of scented gland functions.

Seasonal expressions of follicle-stimulating hormone receptor and luteinizing hormone receptor in the scented gland of the male muskrat (Ondatra zibethicus).

Accumulating evidence has shown that follicle-stimulating hormone (FSH) and luteinizing hormone (LH) may influence the functions of nongonadal tissues in addition to their classic target gonads. Our previous studies revealed that the scented glands of male muskrats expressed prolactin receptor, steroidogenic enzymes, and inhibin/activin subunits. To further seek the evidence of the activities of pituitary gonadotropins in scented glands, we investigated the seasonal expression patterns of FSH receptor (FSHR) and LH/choriogonadotropin receptor (LHCGR). The weight and size of scented glands during the breeding season were significantly higher than those during the nonbreeding season. Immunohistochemical studies showed that FSHR was present in the serous cells of scented glands, whereas LHCGR was present in the interstitial cells. The protein and mRNA expression levels of FSHR and LHCGR were significantly higher in the scented glands during the breeding season than those during the nonbreeding season. Importantly, the levels of circulating FSH and LH were remarkably higher during the breeding season. Taken together, these results suggested that gonadotropins may affect the function of muskrat scented gland via the locally expressed receptors in a season-dependent manner.

Proliferation and apoptosis processes in the seasonal testicular development of the wild Daurian ground squirrel (Citellus dauricus Brandt, 1844).

The aim of the present study was to elucidate the regulatory role of cell proliferation and apoptosis in testicular development of wild Daurian ground squirrels during the breeding season (April), the non-breeding season (June) and before hibernation (September). Gross mass and hormonal analysis showed that the testis:body mass ratio and plasma testosterone concentration fluctuated seasonally, with a peak in April and lowest values in June. Similarly, spermatogenesis was fully developed in April but suppressed in June and September. Testicular decellularisation and vacuolisation was seen during the transition from the breeding to the non-breeding season. Furthermore, testicular levels of proliferating cell nuclear antigen, cyclin D2 and caspase-3 protein were significantly increased in June and September. Intriguingly, positive terminal deoxyribonucleotidyl transferase-mediated dUTP-digoxigenin nick end-labelling staining and nuclear translocation of caspase-3 in testicular germ cells appeared only during the prehibernation period, whereas accumulation of cyclin D2 in spermatocyte nuclei occurred in September. These findings demonstrate, for the first time, that both cell proliferation and apoptosis are stimulated during the prehibernation period, indicating that a hormonal-regulated balance of testicular germ cell proliferation and apoptosis may play a pivotal role in preparing for testicular recrudescence of wild Daurian ground squirrels.