Primary cilia disappear in rat podocytes during glomerular development

Most tubular epithelial cell types express primary cilia, and mutations of primary-cilium-associated proteins are well known to cause several kinds of cystic renal disease. However, until now, it has been unclear whether mammalian podocytes express primary cilia in vivo. In this study, we determined whether primary cilia are present in the podocytes of rat immature and mature glomeruli by means of transmission electron microscopy of serial ultrathin sections. In immature glomeruli of fetal rats, podocytes express the primary cilia with high percentages at the S-shaped body (88 ± 5%, n = 3), capillary loop (95 ± 4%, n =  4), and maturing glomerulus (76 ± 13%, n = 5) stages. The percentage of ciliated podocytes was significantly lower at the maturing glomerulus stage than at the former two stages. In mature glomeruli of adult rats, ciliated podocytes were not found at all (0 ± 0%, n = 11). These findings indicate that the primary cilia gradually disappear in rat podocytes during glomerular development. Since glomerular filtration rate increases during development, the primary cilia on the podocytes are subjected to a stronger bending force. Thus, the disappearance of the primary cilia presumably prevents the entry of excessive calcium-ions via the cilium-associated polycystin complexes and the disturbance of intracellular signaling cascades in mature podocytes

Evaluation of a vaccine's DNA against Photobacterium damselae subsp.. piscicida. Study of immune response in the Senegal sole (Solea senegalensis kaup)

Photobacterium damselae subsp. piscicida is the aetiological agent of pasteurellosis, a bacterial disease that affects seriously soles grown in captivity provoking significant mortalities. Until moment, long-term effective and oral protective vaccines to prevent pasteurellosis in sole are not commercially available. In this work, the HSP60 gene from P. d. piscicida was used to prepare a DNA vaccine (pPDPHSP60) that was employed to determine the antibacterial immune response elicited by DNA vaccination in sole. Expression of pPDPHSP60 was confirmed both in transfected cells and in vivo using gut of orally vaccinated sole by RT-PCR analysis. Expression profiles of genes involved in the innate immune system were determined in spleen from orally vaccinated sole using an OpenArray real time PCR approach. Results revealed that oral vaccination induced coordinately the expression of genes involved in the response against bacterial infection. This study has been co-funded by project AQUAGENET (SOE2/P1/ E287) program INTERREG IVB SUDOE, and by project RTA2009-00066-00-00 from the Instituto Nacional de Investigaci&oacute;n y Tecnolog&iacute;a Agraria y Alimentaria (INIA, Spain), and FEDER (EU).www.juntadeandalucia.es/agriculturaypesca/ifapa/aquagenetPhotobacterium damselae subsp. piscicida es el agente etiol&oacute;gico de la pasteurelosis, una enfermedad bacteriana que afecta gravemente a los lenguados criados en cautividad produciendo importantes mortalidades. Hasta el momento, no existen vacunas efectivas comercialmente disponibles con una larga protecci&oacute;n y por v&iacute;a oral para prevenir la pasteurelosis. En este trabajo, el gen HSP60 de P. d. piscicida se utiliz&oacute; para construir una vacuna de ADN (pPDPHSP60) y se us&oacute; para determinar la respuesta inmune antibacteriana provocada por la vacunaci&oacute;n en lenguado. La expresi&oacute;n in vitro e in vivo de pPDPHSP60 fue confirmada mediante RT-PCR, tanto en c&eacute;lulas transfectadas como en intestino de los individuos vacunados oralmente. Los perfiles de expresi&oacute;n de los genes implicados en el sistema inmune se determinaron en bazo de los lenguados vacunados oralmente mediante la t&eacute;cnica de OpenArray PCR en tiempo real. Los resultados demostraron que la expresi&oacute;n de los genes implicados en la respuesta contra infecciones bacterianas aumentaba significativamente de forma coordinada. Este estudio ha sido cofinanciado por el proyecto AQUAGENET (SOE2/P1/ E287) programa INTERREG IVB SUDOE, y por el proyecto RTA2009-00066-00-00 para el Instituto Nacional de Investigaci&oacute;n y Tecnolog&iacute;a Agraria y Alimentaria (INIA, Spain), and FEDER (EU). www.juntadeandalucia.es/agriculturaypesca/ifapa/aquagenet</p

Involution of seminiferous tubules in aged hamsters: an ultrastructural, immunohistochemical and quantitative morphological study

In this study, we examined the age-related changes on morphometric parameters and ultrastructure of seminiferous tubules, and on the expression of extracellular matrix proteins in lamina propria of Syrian hamsters. A significant decrease in the percentage of normal tubules and an increase in the percentage of hypospermatogenic and arrested maturation tubules was observed with aging. Aged animals showed a decrease in tubular diameter, tubular lumen, seminiferous epithelium volume and total tubular volume. However, the total length of seminiferous tubules was significantly increased with aging. The most important ultrastructural changes with aging were the thickening of the lamina propria, the presence of diverse abnormalities in the spermiogenesis process, degeneration of germ cells, and vacuolization and flattening of Sertoli cells showing abundant lipofucsin droplets and residual bodies. Laminin immunoreactivity was found along the lamina propria of seminiferous tubules both in young and aged animals. Fibronectin immunoreactivity was found along the lamina propria and blood vessels. Both laminin and fibronectin total volume of immunostaining per testis was increased in aged hamsters. In conclusion, the agerelated changes in seminiferous tubules of hamster include: a decrease in tubular width and an increase in tubular length; widening of the lamina propria caused by a more extensive connective matrix between the peritubular cells and the basal membrane; and a strong disarrangement of the seminiferous epithelium, including germ cell degeneration and important alterations in both spermiogenesis and Sertoli cell structure

19. reunion bienal de la Sociedad EspaNola de Microscopia Electronica

Centro de Informacion y Documentacion Cientifica (CINDOC). C/Joaquin Costa, 22. 28002 Madrid. SPAIN / CINDOC - Centro de Informaciòn y Documentaciòn CientìficaSIGLEESSpai

Melanosome transfer, photoreception and toxicity assays in melanophores

Many animals such as fish and frogs have developed the ability to change colour of their skin to adapt to the environment or to signal to other individuals. This ability is due to specialised skin cells called melanophores. Melanophores contain thousands of melanosomes, small membrane-enclosed organelles containing the black or brown pigment melanin. The melanosomes can aggregate to the cell centre rendering the cells pale or disperse throughout the cell to become dark. The intracellular transport of melanosomes is regulated by neuronal or hormonal external stimuli. Fast colour change is achieved by aggregation/dispersion of melanosomes but long-term colour change can also be achieved by melanosome transfer to surrounding skin cells. An amphibian immortalized melanophore cell line was used from the African claw frog, Xenopus laevis to study transfer of melanosomes to co-cultured fibroblasts. Melanosome transfer was observed and up regulated by the hormone α-MSH . The transfer was quantified using light-, fluorescence and electron microscopy. A new and powerful method for transfer experiments was developed. Fluorescent semiconductor nanocrystals, qdots, were used in combination with flow cytometry. The qdots were taken up by the cultured Xenopus laevis melanophores, localised to the melanosomes and transferred to co-cultured fibroblasts. The method is a step towards enabling large scale analysis of pigment transfer. Xenopus laevis melanophores can be cultivated in 96-well culture plates which allow quantification of aggregation or dispersion in a fast and reproductive way. Glyphosate containing herbicides, i.e. Roundup, are commonly used in the world, but some toxic effects have been found on amphibians in vivo and human and mouse cells in vitro. To learn more about potential effects on intracellular transport and the cytoskeleton in animal Roundup, glyphosate, glyphosateisopropylamine and isopropylamine were tested on the transport of melanosomes to the cell centre by spectrophotmetry and by fluorescence microscopy on microtubules and actin filaments. All tested compounds inhibited the aggregation and affected the morphology of the cytoskeleton. The effect was found to be pH dependent. Amphibian melanophores can be regulated directly by light via a melanopsin receptor. Photoreception was found in cultured early embryos of the zebrafish Danio rerio. Light induced dispersion of the melanophores was contrast to what is found at adults when light causes aggregation of the melanosomes due to signals from the CNS. At least one subclass of melanopsin was detected in the zebrafish retinal pigment epithelial cells