Interacciones epitelio-mesenquimáticas en el desarrollo testicular / Epithelial-mesenchymal transitions in the development of testis

RESUMEN: La espermatogénesis es un proceso continuo que se inicia durante el desarrollo embriofetal. Las relaciones auto, para y yuxtacrinas indican la interdependencia de las células intersticiales (de Leydig) con las células peritubulares (lamina propia) y células sustentaculares (de Sertoli). Ciertos morfógenos son fundamentales en este proceso. Las células sustentaculares son capaces de regular la diferenciación y función de las células peritubulares e intersticiales a través de la producción de IGF1, TGFA, TGFB y DHH. Las células peritubulares son capaces de producir P-Mod-S, regulando la diferenciación de las células sustentaculares, y a través de FGF2 y FGF9 modulan las transiciones epitelio-mesenquimática entre células sustentaculares y mesonefros. También remodelan la membrana basal del condón testicular y regulan la diferenciación y función de las células intersticiales por medio de IGF1, TGFA y TGFB. Las células intersticiales son las reponsables de la producción de testosterona e INSL3, influyendo en la diferenciación sexual masculina. Se plantea que provienen de células mesenquimales del epitelio celómico y mesonefros. Sin embargo, otros autores proponen su origen a partir de células de la cresta neural. Estas influyen a través de mecanismos paracrinos en la proliferación de las células sutentaculares por medio de activina A, teniendo como resultado la expansión del cordón testicular. Las interacciones entre las distintas poblaciones celulares a través de morfógenos inducen una transición epitelio-mesénquima fundamental en la formación y diferenciación de la gónada masculina.

SUMMARY: Spermatogenesis is a continuous process which starts during the embryo-fetal development. Auto, para and juxtacrine relations indicate the interdependence of the interstitial cells (Leydig) with the peritubular cells (lamina propria) and sustentacular cells (Sertoli). Certain morphogens are fundamental in this process. Sustentacular cells are able to regulate differentiation and function and peritubular interstitial cells through production of IGF1, TGFA, TGFB and DHH. Peritubular cells are able to produce P-Mod-S regulating differentiation sustentacular cells and through FGF2 and FGF9 modulate epithelial-mesenchymal transitions between sustentacular cells and mesonephros. They also remodel the basal membrane of the testicular condom and regulate the differentiation and function of the interstitial cells by means of IGF1, TGFA and TGFB. Interstitial cells are responsible for the production of testosterone and INSL3, influencing male sexual differentiation. It is suggested that they come from mesenchymal cells of the coelomic epithelium and mesonephros. However, other authors propose their origin from cells of the neural crest. These influence through paracrine mechanisms proliferation sutentaculares cells by activin A, resulting in the expansion of cord testicular. The interactions between the different cell populations through morphogens induce a fundamental epithelial-mesenchymal transition in the formation and differentiation of the male gonad.

Inductive role of sustentacular cells (Sertoli cells) conditioned medium on bone marrow derived mesenchymal stem cells / Papel inductivo del medio acondicionado de células sustentaculares (células de Sertoli) en las células madre mesenquimales derivadas de la médula ósea

RESUMEN: Las células madre de la línea germinal masculina son factores clave para la espermatogénesis masculina y la fertilidad. Las células sustentaculares (células de Sertoli) como células somáticas juegan un papel fundamental en la creación de un microambiente esencial para la auto-renovación y diferenciación de las células de la línea germinal masculina. Las células madre mesenquimales son reconocidas como células auto-renovables y multipotentes capaces de diferenciarse en múltiples tipos de células. La generación de células germinales masculinas a partir de células madre mesenquimales puede proporcionar un método terapéutico para tratar la infertilidad masculina. En este estudio, las células mesenquimales derivadas de la médula ósea (BMMSCs) se recuperaron de la médula ósea de ratones de 6-8 semanas de edad del Instituto de Investigación Médico Naval (NMRI). En el estudio se aislaron las células sustentaculares y se enrriquecieron usando placas revestidas con lectina. Se obtuvo el medio de condición celular después de diferentes intervalos de tiempo. Posteriormente se cultivaron las BMMSC con diferentes concentraciones de SCCM y medio de Eagle modificado por Dulbecco (DMEM) en diversos momentos. Se evaluaron marcadores específicos de células de línea germinal usando la reacción en cadena de polimerasa transcriptasa inversa (RT-PCR) e inmunocitoquímica. Los resultados mostraron que las BMMSCs cultivadas con SCCM durante 48h exhibieron transcritos específicos de línea germinal (Mvh, Iid4, piwil2) (p <0,05) y marcadores (Mvh, Scp3). Nuestros resultados indican que el cultivo de BMMSCs con SCCM puede conducir a la diferenciación efectiva de BMMSCs en células germinales y proporcionar una estrategia de tratamiento para la infertilidad masculina.

SUMMARY: Male germ line stem cells are key factors for male spermatogenesis and fertility. Sustentacular cells (Sertoli cells) as somatic cells play a pivotal role in creating essential microenvironment for the self-renewal and differentiation of the male germ line cells. Mesenchymal stem cells are recognized as self-renewing and multipotent cells able to differentiate into multiple cell types. The generation of male germ cells from mesenchymal stem cells may provide a therapeutic method to treat male infertility. In this study, Bone marrow derived mesenchymal cells (BMMSCs) were retrieved from the bone marrow of 6-8-week old Naval Medical Research Institute (NMRI) mice. Sustentacular cells (Sertoli cells) were isolated and made rich using lectin coated plates. Sustentacular cell condition medium (SCCM) was collected after different time intervals. Then the BMMSCs were cultured with different concentration of SCCM and Dulbecco's Modified Eagle's medium (DMEM) at various times. Specific markers of Germ line cells were evaluated by using Reverse transcriptase polymerase chain reaction (RT-PCR) and immunocytochemistry. The results showed that BMMSCs cultured with SCCM for 48h exhibited germ line specific transcripts (Mvh, Iid4, piwil2) (p< 0.05) and markers (Mvh, Scp3). Our findings represent that culturing BMMSCs with SCCM may lead to effective differentiation of BMMSCs into germline cells and provide a treatment strategy for male infertility.

The quantification of testicular cells during the postnatal development in two Caviomorph rodents: the guinea pig (Cavia porcellus) and the cutia (Dasyprocta agouti)

ABSTRACT The germinative, Sertoli and Leydig cells of two caviomorph rodents (Cavia porcellus and Dasyprocta agouti) were counted as well as the estimation of the total volume of the testis and the total volume of seminiferous tubules and interstitium in prepubertal, pubertal and adult animals. The number of spermatogonia, spermatocytes and spermatids cells increased during the pubertal phase in both rodents, notably the spermatid cells. The spermatocyte and spermatid slightly decreased in the adult of both rodents, but the increment in spermatogonia cells number was seen, mainly in cutias. The number of Sertoli cells increased in pubertal rodents, but in the adult the number reduced. Substantial number of Leydig cells was counted in pubertal and adult guinea pigs. In cutias, the number of Leydig cells increased in pubertal phase and decline in adults. The design-based stereological method has proven to be unbiased and reliable to be applied in reproduction studies.

Cobertura de la estrategia preventiva de cáncer de vesícula biliar en Chile: Resultados de la Encuesta Nacional de Salud 2009-2010 / Coverage of the gallbladder cancer prevention strategy in Chile: results from the 2009-2010 National Health Survey

Background: In Chile, gallbladder cancer (GBC) is one of the most important causes of death and gallstone disease (GSD) is its main risk factor. Abdominal ultrasonography (AU) is used for the diagnosis of GSD and cholecystectomy is used to prevent it. Aim: To estimate GSD prevalence in the general population and to assess the diagnostic and therapeutic coverage of GSD as a preventive strategy for GBC in Chile. Material and Methods: A standardized digestive symptoms questionnaire of the 2009-2010 Chilean National Health Survey was answered by 5412 adults over 15 years old. Self-reports of AU, GBD and cholecystectomies were recorded. Results: The prevalence of biliary-type pain was 7.1%. During the last five years, the prevalence of AU was 16%. GSD was reported in 20% of these tests and 84% of them were asymptomatic. The prevalence of AU was significantly lower in Araucanía region and among people with less than 12 years of education. Life cholecystectomy prevalence was 11% and reached 40% in people aged over 60 years. Women accounted for 75% of total cholecystectomies. Twenty-one percent of individuals who referred biliary-type pain, were studied with an AU. Only 60% of people with GSD confirmed by AU underwent a cholecystectomy. Conclusions: GSD affects at least 27% of the Chilean adult population. Important deficits and inequities in GSD diagnostic and therapeutic coverage were identified.

Differential expression and localization of ADAM10 and ADAM17 during rat spermatogenesis suggest a role in germ cell differentiation

BACKGROUND: Extracellular metolloproteases have been implied in different process such as cell death, differentiation and migration. Membrane-bound metalloproteases of the ADAM family shed the extracellular domain of many cytokines and receptor controlling auto and para/juxtacrine cell signaling in different tissues. ADAM17 and ADAM10 are two members of this family surface metalloproteases involved in germ cell apoptosis during the first wave of spermatogenesis in the rat, but they have other signaling functions in somatic tissues. RESULTS: In an attempt to further study these two enzymes, we describe the presence and localization in adult male rats. Results showed that both enzymes are detected in germ and Sertoli cells during all the stages of spermatogenesis. Interestingly their protein levels and cell surface localization in adult rats were stage-specific, suggesting activation of these enzymes at particular events of rat spermatogenesis. CONCLUSIONS: Therefore, these results show that ADAM10 and ADAM17 protein levels and subcellular (cell surface) localization are regulated during rat spermatogenesis.

Mechanism of Humoral and Cellular Immune Modulation Provided by Porcine Sertoli Cells

The understanding of main mechanisms that determine the ability of immune privilege related to Sertoli cells (SCs) will provide clues for promoting a local tolerogenic environment. In this study, we evaluated the property of humoral and cellular immune response modulation provided by porcine SCs. Porcine SCs were resistant to human antibody and complement-mediated formation of the membrane attack complex (38.41+/-2.77% vs. 55.02+/-5.44%, p=0.027) and cell lysis (42.95+/-1.75% vs. 87.99 +/-2.25%, p<0.001) compared to immortalized aortic endothelial cells, suggesting that porcine SCs are able to escape cellular lysis associated with complement activation by producing one or more immunoprotective factors that may be capable of inhibiting membrane attack complex formation. On the other hand, porcine SCs and their culture supernatant suppressed the up-regulation of CD40 expression (p<0.05) on DCs in the presence of LPS stimulation. These novel findings, as we know, suggest that immune modulatory effects of porcine SCs in the presence of other antigen can be obtained from the first step of antigen presentation. These might open optimistic perspectives for the use of porcine SCs in tolerance induction eliminating the need for chronic immunosuppressive drugs.

Morfogênese do testículo de embriões e fetos de vacas da raça nelore (Bos taurus indicus) / Morfogenesis of Testis From Embryos and Fetuses of Nelore Breed Cows (Bos taurus Indicus)

Este estudo teve como objetivo acompanhar o processo de desenvolvimento testicular desde a fase indiferenciada até sua completa formação. Embriões e fetos de vacas da raça nelore (Bos taurus indicus) foram obtidos em frigoríficos próximos à cidade de Uberlândia, Minas Gerais. As gônadas dos fetos e os embriões foram fixados em bouin e processados para microscópica óptica convencional. A gônada foi observada primeiramente em um embrião de 1,0 cm de comprimento. Em embriões com 2,5 cm a presença da albugínea permite a identificação do sexo. A espessura média da albugínea variou de 29,08 a 558,46 mm. Gradativamente, observou-se aumento da vascularização da albugínea e do parênquima. O mediastino encontrava-se localizado centralmente. Houve uma diminuição no espaço ocupado pelos cordões testiculares de 63,71 para 41,99% do volume total dos testículos. O seu diâmetro variou de 31,68 a 48,80 mm. O diâmetro das células germinativas (e dos seus núcleos) foi de 12,27(6,65) a 16,95 (14,21) mm, respectivamente. A quantidade de células germinativas por corte transversal de cordão diminuiu de um máximo de 2,80 para 0,76. O número total de células germinativas foi de 16 no princípio da colonização da gônada para 18,32 x 106 no final do estudo. O número de células de Sertoli por corte transversal de cordão variou de 10,00 a 16,25. Os resultados obtidos mostram que a origem e a formação dos testículos nos embriões e fetos de vacas da raça Nelore (Bos taurus indicus) ocorre de forma muito semelhante ao do que é descrito para Bos taurus taurus.

The aim of this study was to accompany the process of testicular development from the non-differentiable phase to its complete formation. Embryos and fetuses of Nelore breed cows (Bos Taurus indicus) were obtained in slaughterhouses near the Uberlandia city, Minas Gerais. The gonads and the embryos were fixed in Bouin’s fixative and afterwards processed for conventional optical microscopy. The gonadal was observed firstly in a 1.0 cm long embryo. In 2.5 cm long embryos the presence of the albuginea allows the sex identification. The mean thickness of the albuginea ranged from 29.08 to 558.45 mm. Gradually increase of vascularization of the albuginea and parenchyma is observed. The mediastinum is located centrally. There was a decrease in the space occupied by the testicular cords, from 63.71 to 41.99% of the total testes volume. Its diameter ranged from 31.68 to 48.80 mm. The diameter of germinal cells (and their nuclei) was from 12.27 (6.65) to 16.95 914.21) mm. The quantity of germinal cells by cross section of cord decreased from a maximum of 2.80 to 0.76. The total number of germinal cells was from 16 at the beginning of colonization of the gonad to 18.32 x 106 at the end of the study. The number of Sertoli’s cells by cross section of cord ranged from 10.00 to 16.25. The results obtained show that the origin and formation of testes in embryos and fetuses from Nelore breed cows (Bos taurus indicus) does occur in a very similar way to what is described for Bos taurus taurus.

Sertoli cell proliferation during the post hatching period in domestic fowl

There has been no study aimed at directly determiningof the periods of Sertoli cell proliferation in birds evendomestic fowl. The aims of this study were to observe thecessation of post-hatching mitotic proliferation of Sertolicells in domestic fowl, and to determine the volumedensity of Sertoli and germ cells during this period. Atotal of 50 Leghorn chicks were used in this study. Thetestes sections of the animals were immunostained withBrdU to observe the proliferation of cells from one to 10weeks of age. The volume density of the Sertoli and germcells were determined using the standard point countingmethod. The volume density of the germ cell nuclei wasinitially less than that of the Sertoli cells but the volumedensity converged by week 6, and remained relativelyconstant until the commencement of meiosis. Clearlabeling of Sertoli and germ cells was observed from week1 to week 7. The only those cells still labeled after 8 weekswere germ cells, indicating that Sertoli cell proliferationhad ceased. Therefore, it is recommended that anyresearch into the testes of domestic fowl should considerthe cessation of Sertoli cell proliferation by approximately8 weeks.