Alicja Kamińska

Chlorinated biphenyls effect on estrogen-related receptor expression, steroid secretion, mitochondria ultrastructure but not on mitochondrial membrane potential in Leydig cells

To characterize polychlorinated biphenyls (PCBs) action on Leydig cells, PCBs congeners, low-chlorinated (delor 103; d103) and high-chlorinated ones (delor 106; d106) were selected. The cells were treated according to PCBs dose (d103 or d106 0.2xa0ng/ml in low doses:, or 2xa0ng/ml in high doses) and type (d103u2009+u2009d106 in low doses or 103u2009+u2009106 in high doses). After 24xa0h treatment with PCBs, a distinct increase in estrogen-related receptors (ERRs type α, β and γ) expression was revealed. However, the dose- and type-dependent PCBs effect was mostly exerted on ERRα expression. A similar increase in ERRs expression was demonstrated by estradiol but not testosterone, which was without an effect on ERRs. PCBs caused no decrease in the membrane potential status of Leydig cells (either in dose or type schedule) but had severe effects on the mitochondria number and structure. Moreover, PCBs markedly increased calcium (Ca2+) concentration and sex steroid secretion (both androgens and estrogens were elevated). These findings suggest a similar estrogenic action of PCBs congeners (d103 and d106) on Leydig cell function. We report dose- and type-specific effects of PCBs only on Leydig cell ERRs expression. Both delors showed common effects on the mitochondria ultrastructural and functional status. Based on our results, ERRα seems to be the most sensitive to hormonal modulation. The increases in Ca2+ and sex steroid secretion may be due to the activation of ERRs by PCBs binding and/or direct effect of PCBs on ERRs mRNA/protein expression. Nevertheless, to confirm the existence of possible relationships between ERRs signaling (including PCBs as ligands) and mitochondria function in Leydig cells, further intensive studies are needed.

Primary and tumor mouse Leydig cells exposed to polychlorinated naphthalenes mixture: Effect on estrogen related-receptors expression, intracellular calcium level and sex hormones secretion.

We report the effects of polychlorinated napthalanes (PCNs) on the mRNA expression of estrogen-related receptors (ERRs) α, β and γ, calcium (Ca2+) concentration, and sex steroid secretion in mouse primary and tumor Leydig cells. The cells were exposed to a mixture of PCNs (10nM) alone or in combination with one of sex steroid receptor antagonists; 182,780 (ICI; 10μM); hydroxyflutamide (HF; 10(-4)M) and G-coupled estrogen receptor antagonist (G15; 10nM) respectively. The expression of mRNAs and protein for ERRα, β, and γ was detected in primary and tumor Leydig cells. The expression of ERRs was always lower in primary Leydig cells. Exposure of Leydig cells to PCNs significantly increased the expression of ERRs mRNA irrespective of the cell type. Concomitantly, an increased concentration of Ca2+ and sex steroids was revealed in exposed cells. After ICI, HF or G15 was added no changes in expression of ERRs was found. In Leydig cells changes in ERRs expression at mRNA level are clearly linked to changes in Ca2+ level and steroid secretion. Estrogen and androgen receptors are not involved in PCNs action in Leydig cells. The effect of PCNs on mouse Leydig cells is independent on the cell of origin (primary or tumor).

Flutamide induces alterations in the cell-cell junction ultrastructure and reduces the expression of Cx43 at the blood-testis barrier with no disturbance in the rat seminiferous tubule morphology

BackgroundPresent study was designed to establish a causal connection between changes in the cell-cell junction protein expression at the blood-testis barrier and alterations in the adult rat testis histology following an anti-androgen flutamide exposure. Particular emphasis was placed on the basal ectoplasmic specialization (ES) in the seminiferous epithelium and expression of gap junction protein, connexin 43 (Cx43).MethodsFlutamide (50xa0mg/kg body weight) was administered to male rats daily from 82 to 88 postnatal day. Testes from 90-day-old control and flutamide-exposed rats were used for all analyses. Testis morphology was analyzed using light and electron microscopy. Gene and protein expressions were analyzed by real-time RT-PCR and Western blotting, respectively, protein distribution by immunohistochemistry, and steroid hormone concentrations by radioimmunoassay.ResultsSeminiferous epithelium of both groups of rats displayed normal histology without any loss of germ cells. In accord, no difference in the apoptosis and proliferation level was found between control and treated groups. As shown by examination of semi-thin and ultrathin sections, cell surface occupied by the basal ES connecting neighboring Sertoli cells and the number of gap and tight junctions coexisting with the basal ES were apparently reduced in flutamide-treated rats. Moreover, the appearance of unconventional circular ES suggests enhanced internalization and degradation of the basal ES. These changes were accompanied by decreased Cx43 and ZO-1 expression (pu2009<u20090.01) and a loss of linear distribution of these proteins at the region of the blood-testis barrier. On the other hand, Cx43 expression in the interstitial tissue of flutamide-treated rats increased (pu2009<u20090.01), which could be associated with Leydig cell hypertrophy. Concomitantly, both intratesticular testosterone and estradiol concentrations were elevated (pu2009<u20090.01), but testosterone to estradiol ratio decreased significantly (pu2009<u20090.05) in flutamide-treated rats compared to the controls.ConclusionsShort-term treatment with flutamide applied to adult rats exerts its primary effect on the basal ES, coexisting junctional complexes and their constituent proteins Cx43 and ZO-1, without any apparent morphological alterations in the seminiferous epithelium. In the interstitial compartment, however, short-term exposure leads to both histological and functional changes of the Leydig cells.

Anti-androgen 2-hydroxyflutamide modulates cadherin, catenin and androgen receptor phosphorylation in androgen-sensitive LNCaP and androgen-independent PC3 prostate cancer cell lines acting via PI3K/Akt and MAPK/ERK1/2 pathways

This study aimed to investigate rapid effect of anti-androgen 2-hydroxyflutamide (HF) on cadherin/catenin complex and androgen receptor (AR) phosphorylation in prostate cancer cell lines. In addition, a role of PI3K/Akt and MAPK/ERK1/2 pathways in mediating these effects was explored. We have demonstrated that in androgen-sensitive LNCaP cells HF induced rapid increase of E-cadherin phosphorylation at Ser 838/840 (p<0.05) in MAPK/ERK1/2-dependent manner, whereas phosphorylation of β-catenin at Tyr 654 was unchanged. Concomitantly, the reduction of the level of AR phosphorylated at Ser210/213 was found (p<0.01). In androgen-independent PC3 cells HF decreased Tyr 860 N-cadherin and Tyr 645 β-catenin phosphorylation (p<0.01), acting via both MAPK/ERK1/2 and PI3K/Akt pathways. Further, we evidenced that MAPK/ERK1/2 and PI3K/Akt pathways were differentially influenced by HF in LNCaP and PC3 cells. In LNCaP cells, both Akt (p<0.01) and ERK1/2 (p<0.001) phosphorylation were negatively regulated and this effect was mediated by Raf-1 (p<0.05). In contrast, in PC3 cells HF stimulated Akt (p<0.001) and ERK1/2 (p<0.001) activation, but had no effect on the crosstalk between PI3K/Akt and MEK/ERK1/2 pathways at the Raf-1 kinase level. Our findings expand the role of anti-androgen into non-genomic signaling, creating a link between anti-androgen action and phosphorylation of adherens junction proteins in prostate cancer cells.

Bisphenol A and dibutyl phthalate affect the expression of juxtacrine signaling factors in rat testis

The study was designed to examine the effects of model plastic derived compounds, bisphenol A (BPA) and dibutyl phthalate (DBP), on juxtacrine communication in adult rat testis, by evaluating the expression of Notch pathway components. Testicular explant were exposed inxa0vitro to BPA (5u202f×u202f10-6u202fM, 2.5u202f×u202f10-5u202fM, 5u202f×u202f10-5u202fM) or DBP (10-6u202fM, 10-5u202fM, 10-4u202fM) for 24u202fh. To determine the expression of Notch1, Dll4, Hey1, Hes1 and Hey5 real-time RT-PCR was used. Protein levels and localization of NOTCH1 receptor, its ligand DLL4 as well as HEY1, HES1 and HEY5 factors were detected by western blot analysis and immunohistochemistry, respectively. Upregulation of Notch1, Dll4 and Hey1 at the mRNA and protein level was demonstrated in testis explants after BPA and DBP treatment (pu202f<u202f0.05; pu202f<u202f0.01; pu202f<u202f0.001). Hes5 expression decreased after BPA (pu202f<u202f0.05; pu202f<u202f0.01; pu202f<u202f0.001), whereas Hes1 expression was not altered by either BPA or DBP. Tested chemicals altered immunoexpression of activated NOTCH1, DLL4, HEY1 and HES5 both in seminiferous epithelium and interstitial tissue, exerting differential effects on particular cell types. In conclusion, BPA and DBP affect Notch signaling pathway in rat testis, which indicates that juxtacrine communication is a potential target for the action of plastic derived compounds in male gonad.

Regulation of steroidogenic function of mouse Leydig cells : G-coupled membrane estrogen receptor and peroxisome proliferator-activated receptor partnership

We tested whether G-coupled membrane estrogen receptor (GPER) and peroxisome proliferator activated receptor (PPAR) partnership exists and whether this interaction regulates mouse Leydig cell function. Mature and aged mice were treated with the antagonist of GPER (G-15; 50 μg/kg b.w). Leydig cells (MA-10) were treated with G-15 (10 nM) alone or in combination with peroxisome proliferator-activated receptor α or γ antagonists, respectively (PPARα, 10 μM; PPARγ, 10 μM). GPER blockage affected testis steroidogenic status via changes in lutropin and cholesterol levels as well as protein expression alterations of the lutropin receptor, acute steroidogenesis activating protein, translocator protein, and protein kinase A in mouse Leydig cells both in vivo and in vitro. Inactivation of both GPER and PPAR in vitro revealed expressional modulation of other steroidogenesis-controlling molecules acting on various steps of lipid homeostasis e.g. cytochrome P450scc, perilipin, hormone sensitive lipase, and 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase. Concomitantly, microscopic analysis of cells treated with antagonists showed changes in morphology, migration competences and cytoskeleton structure. In the above processes, the action of GPER and PPARα was regulated through the PI3K/Akt pathway, while PPARγ was mediated by the Ras/Raf pathway. In addition, GPER and PPARs specifically controlled individual signaling proteins. For the first time, we report here the importance of GPER-PPARα and -PPARγ neopartnership in maintenance of Leydig cell morpho-functional status.

Differences in aromatase expression and steroid hormone concentrations in the reproductive tissues of male domestic turkeys (Meleagris gallopavo) with white and yellow semen

ABSTRACT 1. To show hormonal differences between male turkeys with yellow semen syndrome (YSS) and white, normal semen (WNS), the expression of aromatase, oestrogen receptor α (ERα), and oestrogen receptor β (ERβ) as well as testosterone and oestradiol concentrations in YSS and WNS testes, epididymis, and ductus deferens were examined. 2. To measure gene expression levels of aromatase and oestrogen receptors (ERs), three complementary techniques (real-time PCR, Western blot, and immunohistochemistry) were used, whereas steroid hormone levels were determined radio-immunologically. 3. Upregulation of aromatase and ERα mRNAs in YSS testes (P < 0.05; P < 0.01), epididymis (P < 0.001; P < 0.001), and ductus deferens (P < 0.05; P < 0.01) compared to those of WNS tissues was detected. Significant increases in the levels of aromatase and ERα proteins were detected in YSS testes (P < 0.001; P < 0.05), epididymis (P < 0.001; P < 0.001), and ductus deferens (P < 0.001; P < 0.05). The expression of ERβ mRNA and protein level was upregulated in the testes (P < 0.05; P < 0.01) and epididymis (P < 0.001; P < 0.01) but not in ductus deferens where it was downregulated (P < 0.01; P < 0.01). Increased intensity of immunoreactive proteins in YSS versus WNS reproductive tissues corroborated gene expression results. 4. Testosterone concentration diminished in YSS epididymis (P < 0.05) and ductus deferens (P < 0.05), but not in the testes, remaining at high level (P < 0.05) compared to WNS values. Concomitantly, increased oestradiol concentration was found in YSS testes (P < 0.05) and epididymis (P < 0.05) but decreased in the ductus deferens (P < 0.05). 5. From the published literature, this study is the first to demonstrate the ability for androgen aromatisation in the turkey reproductive tissues and to show the cellular targets for locally produced oestrogens. The data suggested that the androgen/oestrogen ratio is a mechanistic basis for amplification of differences between turkeys with white and yellow semen and that these results can have a relevance in applied sciences to widen the knowledge on domestic bird reproduction.

Stężenie homocysteiny w surowicy krwi i styl życia katolików zamieszkujących teren Polski Południowej

Introduction: Public health specialists and nurses are interested in identifying the elements of lifestyle that prevent disease in specific human populations, including religious groups. Aim: The term lifestyle Catholic compound with the level of homocysteine in blood serum, as a risk factor for cardiovascular disease (CVD). Material and Methods: The cross-sectional study was conducted among 134 Catholics. Use of a questionnaire of own authorship, the International Physical Activity Questionnaire, Inventory Behavioral Health (HBI), Scale Experienced Stress PSS-10, laboratory tests (homocysteine), Test Fagerstrom Nicotine Dependence. Results: In more than half of Catholics diagnosed increased homocysteine levels. Knowledge of about halves of the respondents on CVD was fully satisfactory. Selected elements of life style Catholics, like physical activity, addictions, perceived stress, health behaviors in the HBI, psychosocial factors and diet deviate from the recommendations relevant to the prevention of CVD and were not significantly linked to the performance level of homocysteine in the blood serum, with the exception of a significant relationship between its proper level and the consumption of milk, whole wheat flakes, pasta and wheat groats and also not smoking cigarettes. Conclusions: Public health professionals and nurses should develop culturally specific educational interventions, taking into the life style and homocysteine levelaccount cardiovascular risk factors among followers of different religions.

Stereotypy i postawy mieszkańców Małopolski wobec przedstawicieli wybranych kultur

Wstep. W Polsce kontakty z osobami mowiącymi innym jezykiem, o innym kolorze skory, innego wyznania, nalezącymi do innego kregu kulturowego, mającymi inne obyczaje i wierzenia, znacznie sie nasilily po wejściu Polski do rodziny krajow Unii Europejskiej. Cel. Celem pracy bylo poznanie opinii mieszkancow Malopolski na temat stereotypow, uprzedzen i dyskryminacji oraz ich postaw wobec osob z odmiennych obszarow kulturowych. Material i metody. Badania przeprowadzono wśrod 120 osob zamieszkujących teren poludniowej Malopolski. Do przeprowadzenia badania wykorzystano kwestionariusz ankiety wlasnego autorstwa oraz Skale Bogardusa, jako narzedzie badawcze wystandaryzowane. Do analizy statystycznej uzyto testu U Manna-Whitneya, Spearmana. Wyniki. Wiekszośc osob badanych miala kontakt ze stereotypami na temat mniejszości kulturowych. Wszyscy ankietowani potrafili wymienic kilka stereotypow o Romach, muzulmanach oraz Żydach. Najwiekszy dystans ankietowani wykazali wobec mniejszości romskiej, najmniejszy w stosunku do mniejszości muzulmanskiej. Wnioski. Malopolska jest regionem, ktory zamieszkany jest przez wiele osob nalezących do mniejszości kulturowych i narodowościowych. Codzienny kontakt z tymi osobami powinien sprowadzac sie do zrozumienia ich kultury, tradycji i zachowan, a nie do dyskryminacji, szykanowania, odtrącania. Bardzo wazne jest, aby zapobiegac potencjalnym konfliktom spolecznym, do ktorych moze dochodzic, jeśli bedziemy kierowac sie uprzedzeniami, ktore najcześciej wynikają ze stereotypow. Problemy Pielegniarstwa 2015; 23 (1): 27–33

Sytuacje trudne dla studentów pielęgniarstwa w czasie pierwszych zajęć praktycznych w szpitalu

Wstep. Studenci pielegniarstwa w trakcie realizacji pierwszych zajec praktycznych, mają kontakt z nowym środowiskiem, podejmują nowe dzialania, co wiąze sie z wystepowaniem sytuacji trudnych. Cel. Celem pracy byla analiza sytuacji trudnych z jakimi spotykają sie studenci pielegniarstwa w czasie realizacji pierwszych zajec praktycznych. Material i metody. Badaniami objeto 105 studentow studiow licencjackich na kierunku pielegniarstwo UJ CM w Krakowie. Zastosowano metode sondazu diagnostycznego i szacowania. Wykorzystano Skale Uogolnionej Wlasnej Skuteczności autorstwa Schwarzera i Jerusalem, w adaptacji Juczynskiego (GSES) i kwestionariusz ankiety wlasnej konstrukcji. Analiza statystyczna zostala przeprowadzona testem chi2, za poziom istotności przyjeto p < 0,05. Wyniki. Prawie polowa badanych studentow pielegniarstwa przyznala, ze w czasie realizacji pierwszych zajec praktycznych w szpitalu bardzo czesto doświadczala sytuacji trudnych. Sytuacji takich istotnie rzadziej doznawali studenci mlodsi wiekiem (p = 0,00219; chi2 = 11,6908). Najcześciej sytuacje trudne dotyczyly kontaktu z pacjentem. Ich przyczyną byly: brak doświadczenia (47,5%), brak wiedzy (18,8%) oraz brak wsparcia (15%). Najcześciej stosowanym sposobem radzenia sobie z takimi sytuacjami bylo poszukiwanie wsparcia emocjonalnego (49,2%). Wnioski. Wiedza na temat wystepowania sytuacji znaczących w czasie realizacji pierwszych zajec praktycznych dla studentow, poza celami poznawczymi, moze rowniez spelniac funkcje prewencyjne. Problemy Pielegniarstwa 2015; 23 (2): 160–165