RESUMO
We review the literature on the little-known roles of specific CaMKs in regulating endocrine functions of the pineal gland, the pituitary gland, and the hypothalamus. Melatonin activates hippocampal CaMKII, which then influences dendritogenesis. In the pituitary gland, the signal pathways activated by the CaMK in lower vertebrates, such as fishes, differ from those of mammals. In the teleost anterior pituitary, the activation of CaMKII induces the expression of somatolactin by glucagon b. In rats and humans, CaMKIVs have been associated with gonadotropes and thyrotropes and CaMKII with several types of human tumor cells and with a specific signaling pathway. Neuropeptides such as vasopressin and endothelin are also involved in the CaMKII signaling chain, as is the CaMKIIδ isoform which participates in generating the circadian rhythms of the suprachiasmatic nucleus. What arises from this review is that most of the hypothalamic CaMKs are involved in activities of the endocrine brain. Furthermore, among the CaMKs, type II occurs with the highest frequency followed by CaMKIV and CaMKI.
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Organoids (ORGs) are increasingly used as models of cerebral cortical development. Here, we compared transcriptome and cellular phenotypes between telencephalic ORGs and monolayers (MONs) generated in parallel from three biologically distinct induced pluripotent stem cell (iPSC) lines. Multiple readouts revealed increased proliferation in MONs, which was caused by increased integrin signaling. MONs also exhibited altered radial glia (RG) polarity and suppression of Notch signaling, as well as impaired generation of intermediate progenitors, outer RG, and cortical neurons, which were all partially reversed by reaggregation of dissociated cells. Network analyses revealed co-clustering of cell adhesion, Notch-related transcripts and their transcriptional regulators in a module strongly downregulated in MONs. The data suggest that ORGs, with respect to MONs, initiate more efficient Notch signaling in ventricular RG owing to preserved cell adhesion, resulting in subsequent generation of intermediate progenitors and outer RG, in a sequence that recapitulates the cortical ontogenetic process.
Assuntos
Adesão Celular , Córtex Cerebral/metabolismo , Células Ependimogliais/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Neurogênese , Organoides/metabolismo , Transcriptoma , Diferenciação Celular , Córtex Cerebral/citologia , Células Ependimogliais/citologia , Regulação da Expressão Gênica no Desenvolvimento , Redes Reguladoras de Genes , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Integrinas/metabolismo , Masculino , Neurônios/citologia , Neurônios/metabolismo , Técnicas de Cultura de Órgãos/métodos , Organoides/citologia , Proteoma , RNA-Seq , Receptores Notch/metabolismo , Transdução de SinaisRESUMO
Superoxide dismutase is widespread in the human body, including skin and its appendages. Here, we focus on human skin copper/zinc superoxide dismutase, the enzyme that protects skin and its appendages against reactive oxygen species. Human skin copper/zinc superoxide dismutase resides in the cytoplasm of keratinocytes, where up to 90% of cellular reactive oxygen species is produced. Factors other than cell type, such as gender, age and diseased state influence its location in skin tissues. We review current knowledge of skin copper/zinc superoxide dismutase including recent studies in an attempt to contribute to solving the question of its remaining unexplained functions. The research described here may be applicable to pathologies associated with oxidative stress. However, recent studies on copper/zinc superoxide dismutase in yeast reveal that its predominant function may be in signaling pathways rather than in scavenging superoxide ions. If confirmed in the skin, novel approaches might be developed to unravel the enzyme's remaining mysteries.
RESUMO
The localization of copper and zinc-superoxide dismutase in normal and neoplastic human skin was examined with immunochemical techniques. Skin samples were taken from males and females of different ages, UV exposed and non-exposed areas and basal-/spino-cellular carcinomas. The enzyme was localized diffusely in the cytoplasm and was also found in the nuclei of epidermal cells, endothelial cells and other dermis cell types. The dismutase content in the epidermis was higher in males than females, UV-exposed than non-exposed and young than old people. In the tumors, the enzyme content of the superficial epidermal layers was higher than in the deep tumoral epithelial cells. These data suggest that the localization of Cu, Zn-SOD in skin tissues reflects the gender and age of the subject, the cell types and their normal or diseased state. Further studies based on the investigation of systemic changes of this enzyme in physiological and pathological epidermis could provide additional information on tumor cell generation.
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The existence of both calcium-binding proteins (CBPs) and neuropeptides in the retina and brain of various species of vertebrates and invertebrates is well documented. Octopus retina is particularly interesting because it represents a case of convergent evolution. The aim of this study was to characterize the distribution of two CBPs, calretinin and calbindin, in Octopus retina using morphology, in situ hybridization, immunocytochemistry and Western blot. Calretinin-like immunoreactivity was found in the photoreceptor cells, but unexpectedly also in the supporting cells. In situ hybridization and Western blot analysis confirmed these results. No immunoreactivity was found for calbindin. Two neuropeptides, Substance P and calcitonin gene-related peptide (CGRP), as well as neurofilament protein and glial fibrillary acidic protein were also localized in the Octopus retina by immunocytochemistry. Our work provides new insights about calcium-binding proteins and neuropeptide distribution in Octopus retina and suggests a functional role for calretinin, a highly conserved protein, in visual signal transduction of cephalopods.
Assuntos
Calbindina 2/metabolismo , Calbindinas/metabolismo , Octopodiformes/metabolismo , Retina/metabolismo , Animais , Encéfalo/metabolismo , Imuno-Histoquímica/métodos , Hibridização In Situ/métodos , Neuropeptídeos/metabolismo , Retina/patologia , Células Ganglionares da Retina/metabolismo , Células Ganglionares da Retina/patologiaRESUMO
The main functions of the testis are sex hormone and sperm cell production. Steroidogenesis occurs in the Leydig interstitial cells and spermatogenesis in the seminiferous tubules. Male gonad morphogenesis is a finely orchestrated process, mainly coordinated by hormones, whose actions can significantly affect post-pubertal testicular function. Calcium is a key intracellular messenger, which regulates many signal transduction pathways, and is also implicated in steroidogenesis. Calcium homeostasis and signaling rely on many calcium-binding proteins including calretinin, of the "EF-hand" protein family. Calretinin is a highly conserved protein mainly expressed in the nervous system but also detected in rat and human adult and fetal testis as well as in pathological conditions. Calretinin expression in the fetal testis, however, has not been thoroughly analyzed probably owing to limited availability and paucity of tissues. Here, we examined by immunocytochemistry the expression of calretinin in human fetal testis specimens, obtained from natural and therapeutic abortions, at various developmental ages. We found that calretinin-immunoreactive Leydig cells were visible throughout the timeframe studied (14th-27th week). Immunoreactivity was also observed in Sertoli cells and in the germ cells of the immature seminiferous tubules. Overall our data indicate that calretinin expression parallels the decline in Leydig cell number, suggesting that its presence is indeed correlated to their steroidogenic activity. They also suggest that the intratubular positivity of calretinin could be linked to the ability of Sertoli cells to produce locally acting hormones contributing to the histodifferentiation of the male genital tract. J. Cell. Physiol. 232: 1872-1878, 2017. © 2016 Wiley Periodicals, Inc.
Assuntos
Calbindina 2/metabolismo , Feto/metabolismo , Testículo/embriologia , Testículo/metabolismo , Criança , Epididimo/citologia , Epididimo/metabolismo , Receptor alfa de Estrogênio/metabolismo , Feto/citologia , Idade Gestacional , Humanos , Imuno-Histoquímica , Células Intersticiais do Testículo/citologia , Células Intersticiais do Testículo/metabolismo , Masculino , Puberdade , Testículo/citologiaRESUMO
It has already been reported that cannabinoids are neuroprotective agents against excitotoxicity in vitro and increase after acute brain damage in vivo. This background prompted us to study the localization and expression of the calcium -binding protein calretinin in a condition similar to Alzheimer disease and its possible relationship with cannabinoids and their supposed protective role. We carried out quantitative analysis of the transient changes in calretinin expression shown by hybridochemistry within neuronal cell populations in the hippocampus of a beta amyloid-treated rat model of Alzheimer's disease and their correlation with endocannabinoid increase. Calretinin expression increases throughout the first week after cortical amyloid-beta peptide injection, and then decreases towards normal levels in the rat hippocampus during the following weeks, indicating that decreased calretinin gene expression may be associated with either increase of endocannabinoids or VDM11-induced accumulation of endocannabinoids. In contrast, SR1, an antagonist, which limits the cannabinoid effect by selective binding to the cannabinoid receptor CB1, up-regulates calretinin expression with respect to non-treated rats. This could mean that the SR1 endocannabinoid-blocking action through CB1 receptors, that are normally stimulated by endocannabinoids to inhibit calcium increase, might cause a higher calretinin expression. This would allow us to speculate on a possible reverse relationship between endocannabinoid and calretinin levels in the hippocampal calcium-homeostasis balance.
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Peptídeos beta-Amiloides/toxicidade , Calbindina 2/metabolismo , Endocanabinoides/metabolismo , Hipocampo/metabolismo , Fragmentos de Peptídeos/toxicidade , Doença de Alzheimer/metabolismo , Animais , Modelos Animais de Doenças , Expressão Gênica , Hipocampo/efeitos dos fármacos , Masculino , RNA Mensageiro/metabolismo , Distribuição Aleatória , Ratos Wistar , Receptor CB1 de Canabinoide/metabolismo , Fatores de TempoRESUMO
BACKGROUND: Uncoupling protein 3 (ucp3) is a member of the mitochondrial anion carrier superfamily of proteins uncoupling mitochondrial respiration. In this study, we investigated the effects of ucp3 genetic deletion on mitochondrial function and cell survival under low oxygen conditions in vitro and in vivo. METHODS AND RESULTS: To test the effects of ucp3 deletion in vitro, murine embryonic fibroblasts and adult cardiomyocytes were isolated from wild-type (WT, n=67) and ucp3 knockout mice (ucp3(-/-), n=70). To test the effects of ucp3 genetic deletion in vivo, myocardial infarction (MI) was induced by permanent coronary artery ligation in WT and ucp3(-/-) mice. Compared with WT, ucp3(-/-) murine embryonic fibroblasts and cardiomyocytes exhibited mitochondrial dysfunction and increased mitochondrial reactive oxygen species generation and apoptotic cell death under hypoxic conditions in vitro (terminal deoxynucleotidyl transferase-dUTP nick end labeling-positive nuclei: WT hypoxia, 70.3 ± 1.2%; ucp3(-/-) hypoxia, 85.3 ± 0.9%; P<0.05). After MI, despite similar areas at risk in the 2 groups, ucp3(-/-) hearts demonstrated a significantly larger infarct size compared with WT (infarct area/area at risk: WT, 48.2 ± 3.7%; ucp3(-/-), 65.0 ± 2.9%; P<0.05). Eight weeks after MI, cardiac function was significantly decreased in ucp3(-/-) mice compared with WT (fractional shortening: WT MI, 42.7 ± 3.1%; ucp3(-/-) MI, 24.4 ± 2.9; P<0.05), and this was associated with heightened apoptotic cell death (terminal deoxynucleotidyl transferase-dUTP nick end labeling-positive nuclei: WT MI, 0.7 ± 0.04%; ucp3(-/-) MI, 1.1 ± 0.09%, P<0.05). CONCLUSIONS: Our data indicate that ucp3 levels regulate reactive oxygen species levels and cell survival during hypoxia, modulating infarct size in the ischemic heart.
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Apoptose/genética , Deleção de Genes , Insuficiência Cardíaca/etiologia , Canais Iônicos/genética , Proteínas Mitocondriais/genética , Isquemia Miocárdica/complicações , Animais , Células Cultivadas , Feminino , Insuficiência Cardíaca/genética , Insuficiência Cardíaca/metabolismo , Masculino , Camundongos , Isquemia Miocárdica/genética , Isquemia Miocárdica/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteína Desacopladora 3RESUMO
BACKGROUND AND PURPOSE: Propionyl-l-carnitine (pLc) exerts protective effects in different experimental models of ischemia-reperfusion (I/R). The aim of the present study was to assess the effects of intravenously and topically applied pLc on microvascular permeability increase induced by I/R in the hamster cheek pouch preparation. METHODS: The hamster cheek pouch microcirculation was visualized by fluorescence microscopy. Microvascular permeability, leukocyte adhesion to venular walls, perfused capillary length, and capillary red blood cell velocity (V(RBC)) were evaluated by computer-assisted methods. E-selectin expression was assessed by in vitro analysis. Lipid peroxidation and reactive oxygen species (ROS) formation were determined by thiobarbituric acid-reactive substances (TBARS) and 2'-7'-dichlorofluorescein (DCF), respectively. RESULTS: In control animals, I/R caused a significant increase in permeability and in the leukocyte adhesion in venules. Capillary perfusion and V(RBC) decreased. TBARS levels and DCF fluorescence significantly increased compared with baseline. Intravenously infused pLc dose-dependently prevented leakage and leukocyte adhesion, preserved capillary perfusion, and induced vasodilation at the end of reperfusion, while ROS concentration decreased. Inhibition of nitric oxide synthase prior to pLc caused vasoconstriction and partially blunted the pLc-induced protective effects; inhibition of the endothelium-derived hyperpolarizing factor (EDHF) abolished pLc effects. Topical application of pLc on cheek pouch membrane produced the same effects as observed with intravenous administration. pLc decreased the E-selectin expression. CONCLUSIONS: pLc prevents microvascular changes induced by I/R injury. The reduction of permeability increase could be mainly due to EDHF release induce vasodilatation together with NO. The reduction of E-selectin expression prevents leukocyte adhesion and permeability increase.
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The main regulator of neovascularization is Vascular Endothelial Growth Factor (VEGF). We recently demonstrated that QK, a de novo engineered VEGF mimicking peptide, shares in vitro the same biological properties of VEGF, inducing capillary formation and organization. On these grounds, the aim of this study is to evaluate in vivo the effects of this small peptide. Therefore, on Wistar Kyoto rats, we evaluated vasomotor responses to VEGF and QK in common carotid rings. Also, we assessed the effects of QK in three different models of angiogenesis: ischemic hindlimb, wound healing and Matrigel plugs. QK and VEGF present similar endothelium-dependent vasodilatation. Moreover, the ability of QK to induce neovascularization was confirmed us by digital angiographies, dyed beads dilution and histological analysis in the ischemic hindlimb as well as by histology in wounds and Matrigel plugs. Our findings show the proangiogenic properties of QK, suggesting that also in vivo this peptide resembles the full VEGF protein. These data open to new fields of investigation on the mechanisms of activation of VEGF receptors, offering clinical implications for treatment of pathophysiological conditions such as chronic ischemia.
Assuntos
Artéria Carótida Primitiva/fisiologia , Membro Posterior/irrigação sanguínea , Neovascularização Fisiológica/fisiologia , Peptídeos/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Cicatrização/fisiologia , Animais , Materiais Biocompatíveis/metabolismo , Biomimética , Capilares/efeitos dos fármacos , Colágeno/metabolismo , Combinação de Medicamentos , Células Endoteliais/metabolismo , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Endotélio Vascular/fisiopatologia , Membro Posterior/metabolismo , Membro Posterior/fisiopatologia , Peptídeos e Proteínas de Sinalização Intercelular , Isquemia/fisiopatologia , Laminina/metabolismo , Masculino , Neovascularização Fisiológica/efeitos dos fármacos , Peptídeos/química , Peptídeos/efeitos dos fármacos , Peptídeos/farmacologia , Engenharia de Proteínas , Proteoglicanas/metabolismo , Ratos , Ratos Endogâmicos WKY , Fator A de Crescimento do Endotélio Vascular/farmacologiaRESUMO
G-protein-coupled receptor (GPCR) kinases, GRKs, are known as serine/threonine kinases that regulate GPCR signaling, but recent findings propose functions for these kinases besides receptor desensitization. Indeed, GRK5 can translocate to the nucleus by means of a nuclear localization sequence, suggesting that this kinase regulates transcription events in the nucleus. To evaluate the effect of GRK5-IkappaB alpha interaction on NFkappaB signaling, we induced the overexpression and the knockdown of GRK5 in cell cultures. GRK5 overexpression causes nuclear accumulation of IkappaB alpha, leading to the inhibition of NFkappaB transcriptional activity. Opposite results are achieved by GRK5 knockdown through siRNA. A physical interaction between GRK5 and IkappaB alpha, rather than phosphorylative events, appears as the underlying mechanism. We identify the regulator of gene protein signaling homology domain of GRK5 (RH) and the N-terminal domain of IkappaB alpha as the regions involved in such interaction. To confirm the biological relevance of this mechanism of regulation for NFkappaB, we evaluated the effects of GRK5-RH on NFkappaB-dependent phenotypes. In particular, GRK5-RH overexpression impairs apoptosis protection and cytokine production in vitro and inflammation and tissue regeneration in vivo. Our results reveal an unexpected role for GRK5 in the regulation of NFkappaB transcription activity. Placing these findings in perspective, this mechanism may represent a therapeutic target for all those conditions involving excessive NFkappaB activity.
Assuntos
Núcleo Celular/metabolismo , Quinase 5 de Receptor Acoplado a Proteína G/metabolismo , Proteínas I-kappa B/metabolismo , NF-kappa B/genética , Transcrição Gênica , Adenoviridae , Animais , Apoptose/efeitos dos fármacos , Bovinos , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Núcleo Celular/efeitos dos fármacos , Células Endoteliais/citologia , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/enzimologia , Quinase 5 de Receptor Acoplado a Proteína G/química , Humanos , Lipopolissacarídeos/farmacologia , Inibidor de NF-kappaB alfa , Neovascularização Fisiológica/efeitos dos fármacos , Ligação Proteica/efeitos dos fármacos , Mapeamento de Interação de Proteínas , Estrutura Terciária de Proteína , Ratos , Regeneração/efeitos dos fármacos , Transcrição Gênica/efeitos dos fármacos , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Microbial products, including lipopolysaccharide (LPS), an agonist of Toll-like receptor 4 (TLR4), regulate the lifespan of dendritic cells (DCs) by largely undefined mechanisms. Here, we identify a role for calcium-calmodulin-dependent kinase IV (CaMKIV) in this survival program. The pharmacologic inhibition of CaMKs as well as ectopic expression of kinase-inactive CaMKIV decrease the viability of monocyte-derived DCs exposed to bacterial LPS. The defect in TLR4 signaling includes a failure to accumulate the phosphorylated form of the cAMP response element-binding protein (pCREB), Bcl-2, and Bcl-xL. CaMKIV null mice have a decreased number of DCs in lymphoid tissues and fail to accumulate mature DCs in spleen on in vivo exposure to LPS. Although isolated Camk4-/- DCs are able to acquire the phenotype typical of mature cells and release normal amounts of cytokines in response to LPS, they fail to accumulate pCREB, Bcl-2, and Bcl-xL and therefore do not survive. The transgenic expression of Bcl-2 in CaMKIV null mice results in full recovery of DC survival in response to LPS. These results reveal a novel link between TLR4 and a calcium-dependent signaling cascade comprising CaMKIV-CREB-Bcl-2 that is essential for DC survival.
Assuntos
Sinalização do Cálcio/fisiologia , Proteína Quinase Tipo 4 Dependente de Cálcio-Calmodulina/imunologia , Proteína Quinase Tipo 4 Dependente de Cálcio-Calmodulina/metabolismo , Células Dendríticas/metabolismo , Monócitos/metabolismo , Receptor 4 Toll-Like/metabolismo , Animais , Proteína de Ligação a CREB/genética , Proteína de Ligação a CREB/imunologia , Proteína de Ligação a CREB/metabolismo , Sinalização do Cálcio/efeitos dos fármacos , Proteína Quinase Tipo 4 Dependente de Cálcio-Calmodulina/genética , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Sobrevivência Celular/imunologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/imunologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Células Dendríticas/citologia , Células Dendríticas/imunologia , Humanos , Lipopolissacarídeos/imunologia , Lipopolissacarídeos/metabolismo , Lipopolissacarídeos/farmacologia , Camundongos , Camundongos Knockout , Monócitos/citologia , Monócitos/imunologia , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/imunologia , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2 , Baço/citologia , Baço/imunologia , Baço/metabolismo , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/imunologia , Proteína bcl-X/genética , Proteína bcl-X/imunologia , Proteína bcl-X/metabolismoRESUMO
AIMS: We investigated whether exercise training could promote angiogenesis and improve blood perfusion and left ventricular (LV) remodelling of the post-myocardial infarction (MI) failing heart. We also explored the contribution of ameliorated beta-adrenergic receptor signalling and function on the overall improvement of cardiac contractility reserve induced by exercise. METHODS AND RESULTS: Adult Wistar male rats were randomly assigned to one of four experimental groups. Sham-operated and post-MI heart failure (HF) rats were housed under sedentary conditions or assigned to 10-weeks of a treadmill exercise protocol. At 4 weeks after MI, sedentary HF rats showed LV eccentric hypertrophy, marked increase of LV diameters associated with severely impaired fractional shortening (14 +/- 5%), increased LV end diastolic pressure (20.9 +/- 2.6 mmHg), and pulmonary congestion. In addition, cardiac contractile responses to adrenergic stimulation were significantly blunted. In trained HF rats, exercise was able to (i) reactivate the cardiac vascular endothelial growth factor pathway with a concurrent enhancement of myocardial angiogenesis, (ii) significantly increase myocardial perfusion and coronary reserve, (iii) reduce cardiac diameters, and (iv) improve LV contractility in response to adrenergic stimulation. This latter finding was also associated with a significant improvement of cardiac beta-adrenergic receptor downregulation and desensitization. CONCLUSIONS: Our data indicate that exercise favourably affects angiogenesis and improves LV remodelling and contractility reserve in a rat model of severe chronic HF.
Assuntos
Vasos Coronários/fisiopatologia , Insuficiência Cardíaca/fisiopatologia , Infarto do Miocárdio/complicações , Miocárdio/metabolismo , Neovascularização Fisiológica , Esforço Físico , Receptores Adrenérgicos beta/metabolismo , Transdução de Sinais , Agonistas Adrenérgicos beta/farmacologia , Animais , Circulação Coronária , Vasos Coronários/efeitos dos fármacos , Vasos Coronários/metabolismo , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Insuficiência Cardíaca/diagnóstico por imagem , Insuficiência Cardíaca/etiologia , Insuficiência Cardíaca/metabolismo , Isoproterenol/farmacologia , Masculino , Contração Miocárdica , Infarto do Miocárdio/diagnóstico por imagem , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/fisiopatologia , Miocárdio/enzimologia , Miocárdio/patologia , Neovascularização Fisiológica/efeitos dos fármacos , Óxido Nítrico Sintase Tipo II/metabolismo , Óxido Nítrico Sintase Tipo III , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Ratos Wistar , Receptores Adrenérgicos beta/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Fatores de Tempo , Ultrassonografia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Função Ventricular Esquerda , Remodelação VentricularRESUMO
Downregulation of hypoxia-inducible factor 1 (HIF-1) and vascular endothelial growth factor (VEGF) are shown to be involved in age-dependent impairment of angiogenesis. In this study, we explore whether prior exercise is able to affect these molecular patterns favorably and to enhance neoangiogenesis in old Wistar rats with hind-limb ischemia. At day 7 after surgery, HIF-1alpha and VEGF expression increased in the ischemic muscle of trained animals. Exercise increased capillary density and limb perfusion as revealed by histologic, angiographic, and dyed bead techniques. Furthermore, exercise capacity and limb trophism have significantly improved in trained aged rats. In these animals, the reduction of VEGF serum levels has reflected the comprehensive improvement in local ischemia evoked by exercise. In conclusion, prior exercise represents a valid tool to counteract age-related molecular alterations resulting in impaired angiogenesis in response to ischemia.
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Envelhecimento/fisiologia , Regulação para Baixo/fisiologia , Membro Posterior/irrigação sanguínea , Isquemia , Neovascularização Fisiológica/fisiologia , Condicionamento Físico Animal/fisiologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Análise de Variância , Animais , Velocidade do Fluxo Sanguíneo , Western Blotting , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Masculino , Ratos , Ratos WistarRESUMO
The distribution of calretinin containing neurons examined by in situ hybridization mapping was compared with that obtained by immunocytochemistry in the brain of octopus. Results revealed a close correspondence between the two types of investigations. Western blot analysis disclosed a 29 kDa protein immunostained with anti-calretinin antibody. Calretinin containing neurons were localized mainly in the cortex of octopus lobes, including the vertical, frontal, basal, buccal, palliovisceral, pedal and branchial, with variations of staining intensity and density of immunoreactive cells. The amacrine cells surrounding calretinin containing neuronal bodies of the cortex were also labeled unlike the glial cells. The close correspondence of blotting analysis, immunocytochemistry and in situ hybridization indicates with no doubt that calretinin, like other calcium-binding proteins previously studied, is also present in the nervous system of cephalopods. Furthermore, although recent findings localize calretinin also in endocrine glands, the presence of this calcium-binding protein in the brain of octopus indicates that calretinin appeared early in the phylogeny as a neuronal protein already in invertebrates.
Assuntos
Encéfalo/anatomia & histologia , Encéfalo/metabolismo , Neurônios/metabolismo , Octopodiformes/anatomia & histologia , Octopodiformes/metabolismo , Proteína G de Ligação ao Cálcio S100/metabolismo , Células Amácrinas/citologia , Células Amácrinas/metabolismo , Animais , Mapeamento Encefálico , Calbindina 2 , Cálcio/metabolismo , Sistema Endócrino/citologia , Sistema Endócrino/metabolismo , Evolução Molecular , Gânglios dos Invertebrados/anatomia & histologia , Gânglios dos Invertebrados/metabolismo , Imuno-Histoquímica , Hibridização In Situ , Filogenia , RNA Mensageiro/metabolismo , Proteína G de Ligação ao Cálcio S100/genéticaRESUMO
Beta2-adrenergic receptors (beta2ARs) are widely expressed, although their physiological relevance in many tissues is not yet fully understood. In vascular endothelial cells, they regulate NO release and vessel tone. Here we provide novel evidence that beta2ARs can regulate neoangiogenesis in response to chronic ischemia. We used in vivo adenoviral-mediated gene transfer of the human beta2AR to the endothelium of the rat femoral artery and increased beta2AR signaling resulting in ameliorated angiographic blood flow and hindlimb perfusion after chronic ischemia. Histological analysis confirmed that beta2AR overexpression also produced benefits on capillary density. The same maneuver partially rescued impaired angiogenesis in spontaneously hypertensive rats (SHR), whereas gene delivery of the G-protein-coupling defective mutant Ile164 beta2AR failed to provide ameliorations. Stimulation of endogenous and overexpressed beta2AR on endothelial cells in vitro was found to regulate cell number by inducing proliferation and [3H]-thymidine incorporation through means of extracellular receptor-activated kinase and vascular endothelial growth factor. The beta2AR also has novel effects on endothelial cell number through stimulation of proapoptosis and antiapoptosis pathways involving p38 mitogen-activated protein kinase and PI3-kinase/Akt activation. Therefore, beta2ARs play a critical role in endothelial cell proliferation and function including revascularization, suggesting a novel and physiologically relevant role in neoangiogenesis in response to ischemia.
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Células Endoteliais/fisiologia , Isquemia/fisiopatologia , Neovascularização Fisiológica , Receptores Adrenérgicos beta 2/fisiologia , Animais , Apoptose , Proliferação de Células , Células Cultivadas , Células Endoteliais/patologia , MAP Quinases Reguladas por Sinal Extracelular/fisiologia , Terapia Genética , Humanos , Hipertensão/fisiopatologia , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Receptores Adrenérgicos beta 2/análise , Receptores Adrenérgicos beta 2/genética , Transdução de Sinais , Proteínas Quinases p38 Ativadas por Mitógeno/fisiologiaRESUMO
OBJECTIVE: To study the cell effects of raloxifene on uterine and leiomyoma tissue in postmenopausal women. DESIGN: Prospective, randomized, double-blind, placebo-controlled study. SETTING: Department of Obstetrics and Gynecology, University "Magna Graecia" of Catanzaro, Italy. PATIENT(S): Forty postmenopausal women affected by uterine leiomyomas and selected for hysterectomy. INTERVENTION(S): Treatment for three cycles of 28 days with raloxifene at a dose of 180 mg/day orally (raloxifene group) or placebo tablets (3 tablets/day orally) (placebo group). MAIN OUTCOME MEASURE(S): Uterine and leiomyoma dimensions were measured in each subject at entry and before surgery. On leiomyomas and homologous myometrium the proliferating cell nuclear antigen (PCNA)-positive cells/total cells (PCNA/TC) and the Bcl-2-positive cells/Bax-positive cells (Bcl-2/Bax) ratios (%), as proliferation and apoptotic indexes, respectively, were measured. RESULT(S): After treatment, uterine and leiomyoma sizes were significantly changed in comparison with baseline and the placebo group. PCNA/TC and Bcl-2/Bax ratios were significantly higher in leiomyomas than in homologous myometrium. A significant difference was detected in PCNA/TC between the myometrium of the raloxifene and control groups, whereas no difference was observed in the Bcl-2/Bax ratio. A significant difference in PCNA/TC and Bcl-2/Bax ratios was detected in leiomyoma tissue between the raloxifene group and controls. CONCLUSION(S): In postmenopausal women, raloxifene administration reduces uterine leiomyomas by exerting a cell antiproliferative and proapoptotic action.
Assuntos
Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Leiomioma/tratamento farmacológico , Pós-Menopausa/efeitos dos fármacos , Cloridrato de Raloxifeno/uso terapêutico , Neoplasias Uterinas/tratamento farmacológico , Apoptose/fisiologia , Método Duplo-Cego , Feminino , Inibidores do Crescimento/farmacologia , Inibidores do Crescimento/uso terapêutico , Humanos , Leiomioma/metabolismo , Leiomioma/patologia , Pessoa de Meia-Idade , Pós-Menopausa/metabolismo , Cloridrato de Raloxifeno/farmacologia , Neoplasias Uterinas/metabolismo , Neoplasias Uterinas/patologiaRESUMO
Calcium binding proteins such as calbindin and calretinin have been studied in the pituitary gland, but information on them is still incomplete. To investigate the localization, distribution and role of calbindin in the pituitary, several antibodies to calbindin and to other pituitary markers, such as calretinin and tyrosine-hydroxylase, have been used in male, female and lactating rats. Calbindin has not been localised to a specific endocrine cell population unlike calretinin in the thyrotrophs. There was occasional localization in somatotrophs, thyrotrophs and luteotrophs, but not in corticotrophs or lactotrophs. However, there are sex differences in the expression of this protein as the number of calbindin-immunoreactive cells is higher in the male than in the female pituitary. Furthermore, the number of calbindin containing cells, not lactotrophs, increases in lactating rats and decreases after removal of the pups. It is concluded that calbindin expression may be altered by physiological and endocrine events such lactation, even though it is still unclear why the protein is not related to a specific cell type. The simultaneous use of monoclonal and polyclonal antisera to calbindin revealed that the rabbit antibody recognizes nuclear and cytoplasmic calbindin, while the monoclonal one binds only to the cytoplasmic calbindin. The suggestion is that calbindin may have a secondary role that is not simply to bind calcium.
Assuntos
Lactação/metabolismo , Hipófise/química , Hipófise/metabolismo , Proteína G de Ligação ao Cálcio S100/análise , Proteína G de Ligação ao Cálcio S100/biossíntese , Caracteres Sexuais , Animais , Calbindinas , Feminino , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
Angiogenesis is a tightly regulated process, both during development and adult life. Animal models with mutations in the genes coding for placental growth factor (PlGF), a member of vascular endothelial growth factor (VEGF) family, or the tyrosine kinase domain of the PlGF receptor (Flt-1) have revealed differences between normal physiological angiogenesis and pathological angiogenesis associated with conditions such as tumor growth, arthritis and atherosclerosis. In the present paper, we investigated the potential role of PlGF in regulating physiological angiogenesis by analyzing vascular changes in heart and skeletal muscles of wild-type and Plgf-/- mice following prolonged and sustained physical training. Sedentary Plgf-/- mice showed a reduced capillary density in both heart and skeletal muscles as compared to wild-type mice (P < 0.05). However, after a 6-week training period, heart/body weight ratio, citrate synthase activity, vessel density and capillary/myocyte ratio were significantly increased in both wild-type and Plgf-/- mice (all P < 0.05). At the same time intercapillary distance was significantly reduced. Finally, acute exercise was not associated with any change in PlGF protein level in the skeletal muscle. Our results demonstrate that PlGF is not necessary for exercise-training-induced angiogenesis. We thus suggest that the role of PlGF is confined to the selective regulation of angiogenesis only under pathological conditions.
Assuntos
Neovascularização Fisiológica/fisiologia , Esforço Físico/fisiologia , Proteínas da Gravidez/fisiologia , Animais , Capilares/fisiologia , Citrato (si)-Sintase/análise , Citrato (si)-Sintase/metabolismo , Vasos Coronários/fisiologia , Coração/fisiologia , Masculino , Camundongos , Músculo Esquelético/irrigação sanguínea , Músculo Esquelético/química , Músculo Esquelético/metabolismo , Mutação/genética , Miocárdio/química , Miocárdio/metabolismo , Neovascularização Fisiológica/genética , Condicionamento Físico Animal , Fator de Crescimento Placentário , Proteínas da Gravidez/análise , Proteínas da Gravidez/genética , Fator A de Crescimento do Endotélio Vascular/análise , Fator A de Crescimento do Endotélio Vascular/fisiologiaRESUMO
Recent studies using biotechnological methods have achieved significant advances in our knowledge of molecular mechanisms underlying pituitary gland development and the differentiation of pituitary cytotypes. A large number of neuropeptides have been reported in the adult pituitary gland as well as in the central and peripheral nervous system. The early presence of neuropeptides during pituitary development is reviewed here. Neuromedin U (NmU), galanin and the polypeptide 7B2 have been localised to different endocrine cells of the gland. Their expression seems to be manifold even though it is temporally and spatially regulated. There is now firm immunocytochemical evidence that neuropeptides are present during morphogenesis of the pituitary and can be present simultaneously with all pituitary hormones.