Expression and functional role of hepatocyte growth factor receptor (C-MET) during postnatal rat testis development

The met protooncogene encodes the hepatocyte growth factor receptor (HGFR, c-met). C-met, a tyrosine kinase receptor protein, is widely expressed in different cell types including the male reproductive tract. As we recently demonstrated, both c-met messenger RNA and protein are expressed in prebuberal rat testis. The aim of this work was to detect the expression of c-met during postnatal testis development and to study its functional role. Our findings show that in total rat testis c-met is expressed during postnatal life until the sexual maturation of the animals. To evaluate the receptor expression in the different cell types in the testis, homogeneous cell populations of Sertoli and peritubular myoid cells were isolated from the seminiferous tubules of 10- and 35-day-old animals. c-met gene is expressed in myoid cells at the ages considered and its expression decreases with increasing age. By contrast, in Sertoli cells c-met expression is first detectable at 25 days of life and its expression increases with the increasing age being well evident at 35 days of age. C-met protein was detected by immunocytochemistry and its expression correlates with gene expression. The receptor is functionally active because HGF administration induces morphological changes in myoid cells and in c-met-expressing Sertoli cells. As a consequence of HGF addition, Sertoli cells cultured on reconstituted basement membrane reorganize into cord-like structures that resemble testicular seminiferous cords. The data here reported demonstrate for the first time that in Sertoli cells c-met expression is developmentally regulated being present and functionally active in postpuberal Sertoli cells. Given that c-met expression persists in myoid cells during postnatal testis development and that in Sertoli cells its expression correlates over time with germ cell differentiation and lumen formation, we conclude that the c-met/HGF system is involved in testis development and function.     

Nerve growth factor is required for early follicular development in the mammalian ovary

Nerve growth factor (NGF) epitomizes a family of proteins known as the neurotrophins (NTs), which are required for the survival and differentiation of neurons within both the central and peripheral nervous system. Synthesis of NGF in tissues innervated by the peripheral nervous system is consistent with its function as a target-derived trophic factor. However, the presence of low- and high-affinity NGF receptors in the gonads suggests another function for the NTs within the reproductive endocrine system. We now report that NGF is required for the growth of primordial ovarian follicles, a process known to occur independently of pituitary gonadotropins. Both the NT receptor p75(NTR) and the NGF tyrosine kinase receptor trkA were found to be expressed in the ovaries of infantile normal mice and mice carrying a null mutation of the NGF gene. The ovaries from homozygote NGF-null (-/-) mutant animals, analyzed after completion of ovarian histogenesis, exhibited a markedly reduced population of primary and secondary follicles in the presence of normal serum gonadotropin levels, and an increased number of oocytes that failed to be incorporated into a follicular structure. Assessment of mitogenic activity using two complementary proliferation markers revealed a conspicuous reduction in somatic cell proliferation in the ovaries of NGF-deficient mice. These results suggest that the delay in follicular growth observed in NGF(-/-) mice may be related to the loss of a proliferative signal provided by NGF to the nonneural endocrine component of the ovary.     

Essential role for Galpha13 in endothelial cells during embryonic development

Toward identifying the roles of protease-activated receptor-1 (PAR1) and other G protein-coupled receptors important for vascular development, we investigated the role of Galpha13 in endothelial cells in the mouse embryo. LacZ inserted into Galpha13 exon 1 was highly expressed in endothelial cells at midgestation. Endothelial-specific Galpha13 knockout embryos died at embryonic days 9.5-11.5 and resembled the PAR1 knockout. Restoration of Galpha13 expression in endothelial cells by use of a Tie2 promoter-driven Galpha13 transgene rescued development of endothelial-specific Galpha13 knockout embryos as well the embryonic day 9.5 vascular phenotype in Galpha13 conventional knockouts; transgene-positive Galpha13-/- embryos developed for several days beyond their transgene-negative Galpha13-/- littermates and then manifested a previously uncharacterized phenotype that included intracranial bleeding and exencephaly. Taken together, our results suggest a critical role for Galpha13 in endothelial cells during vascular development, place Galpha13 as a candidate mediator of PAR1 signaling in this process, and reveal roles for Galpha13 in other cell types in the mammalian embryo.     

Expression and functional role of hepatocyte growth factor and its receptor (c-met) during fetal mouse testis development

The hepatocyte growth factor (HGF) is a pleiotropic cytokine able to regulate different cellular functions. HGF action is mediated by its receptor, c-met, a glycoprotein with tyrosine kinase activity. We previously demonstrated that c-met is expressed in the newly formed seminiferous cords of the mice embryonic testes and that HGF acts as a morphogenetic factor. In this paper, we report that at 15.5 days post-coitum (dpc) c-met is expressed in the testicular cords, whereas at 18.5 dpc c-met expression is almost exclusively localized in the interstitial tissue of the testis in particular in the fetal Leydig cells. In addition, we demonstrate that HGF gene is expressed during the fetal period of testis development, heavily detectable in the interstitial compartment of 18.5 dpc testes. Interestingly, HGF is not expressed in the Leydig cells that, as above reported, express the HGF receptor. Looking for the functional role of HGF on Leydig cells, we evaluated the amount of testosterone secreted by testes isolated from 18.5 dpc embryos and cultured in the presence of HGF. The results of the in vitro organ culture show that, at this age, HGF increases the amount of testosterone secreted in the culture medium. On the contrary, HGF does not modulate the amount of testosterone secreted by testes isolated from 15.5 dpc embryos. In conclusion, we report that HGF is produced in the interstitial compartment of the developing testis but not by the Leydig cells. Conversely, the HGF receptor c-met is expressed in the Leydig cells and HGF modulates Leydig cell function during the late period of prenatal development.     

A role for the transcription intermediary factor 2 in zebrafish myelopoiesis

OBJECTIVE: TIF2 is fused with MOZ in the inv(8)(p11q13) acute myeloid leukemia. TIF2, member of the p160 family, is a histone acetyl transferase (HAT). Deletion of p160 genes were performed in mice. Some observations suggest that p160 family members may perform overlapping functions in mice. Therefore, we decided to choose the zebrafish model to study TIF2. The aim of this study was to characterize the role of this HAT during embryonic development. MATERIAL AND METHODS: We use antisense, morpholino-modified oligomers to transiently knockdown tif2 gene, thus determining whether TIF2 plays a role in zebrafish early development. RESULTS: We show that tif2 is involved in embryogenesis and in primitive hematopoiesis. tif2-knockdown zebrafish embryos are smaller than controls, they demonstrate shorter tails, they display notochord deformation and they exhibit U-shaped tail somites. A synthetic RNA encoding human TIF2 rescues the tif2-knockdown phenotype. Analysis of fli1 expression by whole-mount in situ hybridization indicates normal angioblast specification, but altered localization of intersomitic vessels. The posterior intermediate cell mass, in which a part of primitive hematopoiesis occurs, is altered in tif2 morphants and whole-mount in situ hybridization analyses of l-plastin and mpx expression suggest a specific inhibition of granulocytic and macrophagic differentiation at late stages. CONCLUSION: These data indicate an important role for TIF2 in zebrafish primitive myelopoiesis.     

A role for cAMP-dependent protein kinase in early embryonic divisions.

The cAMP-dependent protein kinase (PKA) pathway affects cell cycle progression in "cycling" Xenopus egg extracts. The concentration of free PKA catalytic subunit oscillates during the cell cycle with a peak at the mitosis-interphase transition and a minimum at the onset of mitosis. Inhibition of endogenous PKA in interphase hastens the onset of mitosis. Stimulation of PKA induces interphase arrest, preventing the activation of the M-phase-promoting factor. PKA does not block the accumulation of cyclin or its binding to p34cdc2, but the resultant complex lacks kinase activity and p34cdc2 remains tyrosine-phosphorylated. PKA appears to stimulate an okadaic acid-sensitive serine/threonine phosphatase that acts upon cdc25. In this way PKA could downregulate the p34cdc2 tyrosine phosphatase activity of cdc25 and consequently block the activation of the M-phase-promoting factor.     

A role for corticotropin-releasing factor in functional gastrointestinal disorders

Functional gastrointestinal disorders (FGIDs), which include irritable bowel syndrome (IBS), encompass a heterogeneous group of diseases identified by chronic or recurrent symptom-based diagnostic criteria. Psychosocial factors are key components in the outcome of clinical manifestations of IBS symptoms. Anxiogenic and endocrine responses to stress are mediated by the corticotropin-releasing factor (CRF)-CRF₁ receptor pathway. Preclinical studies show that activation of the CRF₁ receptor by exogenous CRF or stress recapitulates many functional symptoms of IBS diarrhea-predominant patients as related to anxiogenic/hypervigilant behavior, autonomic nervous system alterations, induction of diarrhea, visceral hyperalgesia, enhanced colonic motility, mucus secretion, increased permeability, bacterial translocation, and mast cell activation, which are all alleviated by selective CRF₁ receptor antagonists. Clinical studies also support that CRF administration can induce IBS-like symptoms in healthy subjects and heighten colonic sensitivity in IBS patients. Yet to be ascertained is whether CRF₁ receptor antagonists hold promise as a new therapy in IBS treatment.     

Morphological changes in the ovary of newly hatched chickens treated with chorionic gonadotropin during embryonic development

The present study describes morphological changes produced in the ovary of newly hatched chickens by treatment with human chorionic gonadotropin (hCG). Morphometric measurements of the volume of steroidogenic cells and structures of the ovarian medulla were performed in controls and experimental chickens treated with hCG (1.0 IU/embryo) at 13, 15, and 17 days of embryonic development. After hCG treatment, the volume of interstitial cell cords increased, and modifications in poorly differentiated cells neighbouring the interstitial cells were also observed. Other changes obtained after hCG treatment were an increment in the development of the lacunar system and blood capillaries, as well as a reduction in the number of germ cells in the stroma of the ovarian medulla.     

A functional role for inducible costimulator (ICOS) in atherosclerosis

BACKGROUND: Lymphocytes appear to influence atherosclerosis by altering cytokine production. Whereas primary lymphocyte activation requires T cell receptor ligation, costimulatory signals also appear requisite for generation of a functional T cell response. Inducible costimulator (ICOS) is a newly discovered T cell molecule with a dual role in immune mediated disorders. Herein, we tested the importance of ICOS in atherosclerosis. METHODS AND RESULTS: Atherosclerotic plaques from ApoE-KO mice were studied immunohistochemically for the presence and localization of ICOS and its receptors and its expression in splenocytes. ApoE-KO mice were immunized with human ICOS/Fc-chimera or non-fused Fc and either provided a chow diet for 6 weeks, or a high fat diet for 8 weeks. ICOS and its ligand were abundantly expressed within plaques from ApoE-KO mice: Spleen cells from atherosclerotic mice exhibited lowered constitutive expression of ICOS yet priming with oxLDL enhanced ICOS expression dose-dependently. In mice induced to develop fatty streaks and to generate ICOS blocking antibodies, early atherosclerosis was increased by approximately 77% whereas upon inducing more advanced lesions, the increase in plaque area upon ICOS blockade group was approximately 36%. IFN-gamma secretion by oxLDL-primed splenocytes in ICOS-immunized mice increased whereas IL-10 secretion diminished as compared to control animals. A similar trend in cytokine production was evident in the lesion by immunohistochemistry. CONCLUSION: ICOS appears as an influential costimulatory pathway in atherosclerosis that may play a protective rather that a proatherogenic role.     

A novel role for cardiac neural crest in heart development

It is well known that cardiac neural crest participates in development of the cardiac outflow septation and patterning of the great arteries. Less well known is that ablation of the cardiac neural crest leads to a primary myocardial dysfunction. Recent data suggests that the myocardial dysfunction occurs because of the absence of an interaction of neural crest and pharyngeal endoderm to alter signaling from the endoderm. Continuation of an FGF-like signal from the endoderm past a precise time in development appears to be detrimental to myocardial maturation.