Jacek Kieżun

Seasonal changes in the expression of the androgen receptor in the testes of the domestic goose (Anser anser f. domestica)

It is generally acknowledged that seasonal fluctuations in the morphology and function of bird testes are primarily regulated by seasonal changes in circulating concentrations of testosterone (T) which mediates its action via the androgen receptor (AR). However, it has not yet been elucidated whether gonadal sensitivity to androgens also varies across the bird reproductive cycle. In order to answer the above question, this study makes the first ever attempt to account for the gonadal expression of the AR gene and protein in relation to circulating and testicular T concentrations in the gonads of male birds during the reproductive cycle. The experimental model used in this study was the domestic goose, Anser anser f. domestica, a species with three distinct phases of the annual reproductive cycle: the breeding season in March, the non-breeding season in July and the sexual reactivation phase in November. The plasma and testicular T concentrations were highest in the breeding season, followed by a dramatic decline in the non-breeding season with a successive rise in the sexual reactivation phase. Interestingly, we observed the divergent effect of season on AR mRNA and protein expression. Whereas the AR gene expression showed a nearly inverse relationship with T levels, the seasonal variations in AR protein levels primarily reflected the differences in T concentrations. The results of our study also indicated that regardless of the examined phase of the season, an abundance of AR protein was found only in the nuclei of Leydig and Sertoli cells and myoid cells. The above supports the observation that somatic cells are the targets for androgen action in bird testes. Summarizing, this study revealed that seasonal variations in sensitivity to androgens in the gonads of male birds are reflected in variations in the availability of their cognate receptors. Furthermore, a different pattern of seasonal expression of the AR gene and protein suggests that the AR system is subject to complex regulation that includes both steroid-dependent and steroid-independent factors.

Estradiol concentration and the expression of estrogen receptors in the testes of the domestic goose (Anser anser f. domestica) during the annual reproductive cycle.

Seasonal fluctuations in the activity of bird testes are regulated by a complex mechanism where androgens play a key role. Until recently, the role played by estrogens in males has been significantly underestimated. However, there is growing evidence that the proper functioning of the testes is associated with optimal estradiol (E2) concentration in both the plasma and testes of many mammalian species. Estrogens are gradually emerging as very important players in hormonal regulation of reproductive processes in male mammals. Despite the previously mentioned, it should be noted that estrogenic action is limited by the availability of specific receptors--estrogen receptor alpha (ERα) and estrogen receptor beta (ERβ). Interestingly, there is a general scarcity of information concerning the estrogen responsive system in the testes of male birds, which is of particular interest in exploring the phenomenon of seasonality of reproduction. To address this question, we have investigated for the first time the simultaneous expression of testicular ERα and ERβ genes and proteins with the accompanying plasma and testicular E2 concentrations during the annual reproductive cycle of male bird. The research model was the domestic goose (Anser anser f. domestica), a species whose annual reproductive cycle can be divided into 3 distinct phases characterized by changes in testicular activity. It has been revealed that the stable plasma E2 profile did not correspond to changing intratesticular E2 profile throughout the experiment. The expression of ERα and ERβ genes and proteins was detected in gander testes and it fluctuated on a seasonal basis with lower level in breeding and sexual reactivation stages and higher level during the nonbreeding stage. Our results demonstrated changes in testicular sensitivity to estrogens in male domestic goose during the annual reproductive cycle. The seasonal pattern of estrogen receptors (ERs) expression was analyzed against the hormonal background and a potential mechanism of ERs regulation in bird testes was proposed. The present study revealed seasonal variations in the estrogen responsive system, but further research is needed to fully explore the role of estrogens in the reproductive tract of male birds.

Colorectal cancer patients exhibit increased levels of galanin in serum and colon tissues

Galanin (GAL) is a 30-amino acid neuropeptide that is expressed in both the central and peripheral nervous system, including the enteric nervous system (ENS). Increased GAL concentrations have been identified in the blood of colorectal cancer (CRC) patients. The aim of the present study was to assess whether sections of the colon wall containing ENS plexuses or CRC tumor are associated with increased GAL concentrations. Blood samples were collected from 68 CRC patients and 39 healthy volunteers. In addition, samples of CRC tumors and colon wall tissue in close proximity to and distant from the neoplastic tissue were obtained from 22 CRC patients. The GAL concentration of sera and tissue homogenates obtained from three sections of the colon wall (mucosa with submucosa, muscularis externa and CRC tumor) was analyzed by ELISA. The localization of GAL was evaluated using immunohistochemistry and morphometry was used to measure the distribution of GAL-immunoreactive (GAL-Ir) myenteric plexuses in the vicinity of cancer invasion and in sections of the colon wall distant from the tumor. The GAL serum concentration of CRC patients was 2.4 times higher than that of the control group. The GAL concentration was highest in the homogenates of neoplastic tissue and mucosa obtained from the control (distant) section of the intestinal wall, followed by that in the mucosa and muscularis externa proximal to the tumor. The lowest GAL concentrations were identified within the muscular layer of the colon wall located distant from the tumor. Strong GAL immunoreactivity was identified in the cancer cells, intestinal epithelium and the submucosal and myenteric plexuses. Morphometric analysis revealed that the GAL-Ir myenteric plexuses in the vicinity of tumor infiltration were significantly smaller in size than those in the intact section of the large intestine. Furthermore, no associations were identified between the clinicopathological characteristics of CRC patients and GAL serum and tissue concentration. The increased GAL serum concentrations observed in CRC patients in comparison to healthy controls may result from GAL secretion by CRC tumors, however, other sources of GAL cannot be excluded. The atrophy of myenteric plexuses within close proximity to the tumor may affect the colon function of CRC patients. In conclusion, investigation into the presence of GAL in the colon wall and serum of CRC patients revealed that serum and tissue GAL levels may present useful potential biomarkers in CRC patients.

Expression of the androgen receptor in the testes and the concentrations of gonadotropins and sex steroid hormones in male turkeys (Meleagris gallopavo) during growth and development.

Androgens, including testosterone (T) and androstenedione (A4), are essential for puberty, fertility and sexual functions. The biological activity of those hormones is mediated via the androgen receptor (AR). The regulation of androgen action in birds is poorly understood. Therefore, the present study analysed mRNA and protein expression of AR in the testes, plasma concentrations of the luteinizing hormone (LH), follicle-stimulating hormone (FSH), T, A4 and oestradiol (E2), as well as the levels of T, A4 and E2 in testicular homogenates of male turkeys (Meleagris gallopavo) at the age of 4, 8, 12, 16, 20, 24 and 28weeks. Plasma concentrations of LH and FSH, as well as plasma and testicular levels of T and A4 began to increase at 20weeks of age. The lowest plasma levels of E2 were noted at 20weeks relative to other growth stages. The 20th week of life seems to be the key phase in the development of the reproductive system of turkeys. The AR protein was found in the nuclei of testicular cells in all examined growth stages. Higher expression of AR protein in the testes beginning at 20weeks of age was accompanied by high plasma concentrations of LH and high plasma and testicular levels of androgens. This relationship seems to be necessary to regulate male sexual function.

PLAGL1 protein is differentially expressed in the nephron segments and collecting ducts in human kidney

INTRODUCTION PLAGL1 (pleiomorphic adenoma gene-like 1) is a C2H2-type zinc finger transcription factor associated with the regulation of cell growth and development. Although PLAGL1 expression in kidney was assessed by biochemical methods, the exact localization of the PLAGL1 protein in human kidney has not yet been described. MATERIAL AND METHODS Macroscopically unchanged specimens of kidney tissue were collected from 39 patients undergoing nephrectomy due to renal cell carcinoma. H & E staining of paraffin sections was used to assess histology of the kidney whereas immunohistochemistry was used to localize PLAGL1 protein in kidney compartments. In addition, database sequences search for putative PLAGL1 binding sites among the kidney-related genes was performed. RESULTS PLAGL1 staining intensity differed depending on the kidney compartment. Strong PLAGL1 immunoreactivity was found in thick ascending limbs of Henles loop, distal tubules and collecting ducts, whereas PLAGL1 expression in proximal tubules and renal corpuscles (including podocytes) was moderate and weak, respectively. By the in sillico screening of promoter sequences for PLAGL1 specific DNA-binding sites GGG-GCCCC we designated 43 candidate genes for PLAGL1-regulated genes. Analysis of their functional annotations identified three significantly over-represented gene sets: inositol phosphate metabolic processes (GO), endocrine and other factor-regulated calcium reabsorption (KEGG) and calcium signaling pathways (KEGG). CONCLUSION Differences in the renal expression of PLAGL1 suggest that this protein may be involved in the regulation of several cellular pathways both as transcriptional factor and coactivator/corepressor of other tran-scription factors reflecting its role in the cell type-specific control of gene expression.

Concentrations of the adrenocorticotropic hormone, corticosterone and sex steroid hormones and the expression of the androgen receptor in the pituitary and adrenal glands of male turkeys (Meleagris gallopavo) during growth and development

Androgens take part in the regulation of puberty and promote growth and development. They play their biological role by binding to a specific androgen receptor (AR). The aim of this study was to evaluate the expression of AR mRNA and protein in the pituitary and adrenal glands, to localize AR protein in luteinizing hormone (LH)-producing pituitary and adrenocortical cells, to determine plasma concentrations of adrenocorticotropic hormone (ACTH) and corticosterone and the concentrations of corticosterone, testosterone (T), androstenedione (A4) and oestradiol (E2) in the adrenal glands of male turkeys at the age of 4, 8, 12, 16, 20, 24 and 28weeks. The concentrations of hormones and the expression of AR varied during development. The expression of AR mRNA and protein in pituitary increased during the growth. The increase of AR mRNA levels in pituitary occurred earlier than increase of AR protein. The percentage of pituitary cells expressing ARs in the population of LH-secreting cells increased in week 20. It suggests that AR expression in LH-producing pituitary cells is determined by the phase of development. The drop in adrenal AR mRNA and protein expression was accompanied by an increase in the concentrations of adrenal androgens. Those results could point to the presence of a compensatory mechanism that enables turkeys to avoid the potentially detrimental effects of high androgen concentrations. Our results will expand our knowledge of the role of steroids in the development of the reproductive system of turkeys from the first month of age until maturity.

The Immunoexpression of YAP1 and LATS1 Proteins in Clear Cell Renal Cell Carcinoma: Impact on Patients’ Survival

The aim of the study was to determine by immunohistochemistry cellular localization and immunoreactivity levels of YAP1 and LATS1 proteins in paired sections of tumor and unchanged renal tissues of 54 clear cell renal cell carcinoma (ccRCC) patients. Associations between clinical-pathological and overall survival (OS; median follow-up was 40.6 months) data of patients and YAP1 and LATS1 immunoreactivity were analyzed by uni- and multivariate Cox regression model and log-rank test. YAP1 immunoreactivity was found in the nuclei of tumor cells in 64.8% of ccRCC patients, whereas only 24.1% of tumors revealed cytoplasmic YAP1 expression. LATS1 immunoexpression was observed only in the cytoplasm of tumor cells in 59.3% of patients. LATS1 immunoreactivity in cancer cells negatively correlated with the size of primary tumor. The overall YAP1 immunoreactivity did not correlate with clinical-pathological data of patients. However, the subgroup of ccRCC patients who presented with cytoplasmic YAP1 immunoexpression had significantly shorter OS (median = 26.8 months) than patients without cytoplasmic YAP1 expression (median undefined). Multivariate Cox analysis revealed that increased cytoplasmic YAP1 (HR = 4.53) and decreased LATS1 immunoreactivity levels (HR = 0.90) were associated with worse prognosis, being independent prognostic factors. These results suggest that YAP1 and LATS1 can be considered as new prognostic factors in ccRCC.