RESUMO
Exposure to heavy metals may cause the overproduction of reactive oxygen species, generating oxidative stress and consequently, various harms to human health. The soil surrounding the Ventanas Industrial Complex, in Puchuncaví and Quintero municipal districts on the central Chilean coast, contains heavy metal concentrations (As, Cu, Pb, Zn, among others) that far exceed the maximum permissible levels established by Italian soil standards (used as a reference). This study aimed to investigate the potential association between heavy metal exposure in humans and the levels of oxidative stress biomarkers in inhabitants of these locations. We took blood samples from 140 adults living in sites with high concentrations of heavy metals in the soil and compared them with blood samples from 140 adults living in areas with normal heavy metal concentrations. We assessed lipid peroxidation, damage to genetic material, and Total Antioxidant Capacity in these blood samples. Our results indicate an association between oxidative damage and heavy metal exposure, where the inhabitants living in exposed areas have a higher level of DNA damage compared with those living in control areas. Given that DNA damage is one of the main factors in carcinogenesis, these results are of interest, both for public health and for public policies aimed at limiting human exposure to environmental pollution.
Assuntos
Metais Pesados , Poluentes do Solo , Adulto , Humanos , Chile , Monitoramento Ambiental/métodos , Metais Pesados/toxicidade , Metais Pesados/análise , Estresse Oxidativo , Solo , Poluentes do Solo/toxicidade , Poluentes do Solo/análise , Medição de Risco , ChinaRESUMO
Tissue regeneration capabilities vary significantly throughout an organism's lifespan. For example, mammals can fully regenerate until they reach specific developmental stages, after which they can only repair the tissue without restoring its original architecture and function. The high regenerative potential of fetal stages has been attributed to various factors, such as stem cells, the immune system, specific growth factors, and the presence of extracellular matrix molecules upon damage. To better understand the local differences between regenerative and reparative tissues, we conducted a comparative analysis of skin derived from mice at regenerative and reparative stages. Our findings show that both types of skin differ in their molecular composition, structure, and functionality. We observed a significant increase in cellular density, nucleic acid content, neutral lipid density, Collagen III, and glycosaminoglycans in regenerative skin compared with reparative skin. Additionally, regenerative skin had significantly higher porosity, metabolic activity, water absorption capacity, and elasticity than reparative skin. Finally, our results also revealed significant differences in lipid distribution, extracellular matrix pore size, and proteoglycans between the two groups. This study provides comprehensive data on the molecular and structural clues that enable full tissue regeneration in fetal stages, which could aid in developing new biomaterials and strategies for tissue engineering and regeneration.
Assuntos
Colágeno , Cicatrização , Camundongos , Animais , Materiais Biocompatíveis , Mamíferos , LipídeosRESUMO
BACKGROUND: Liver transplantation has been demonstrated to be the best treatment for several liver diseases, while grafts are limited. This has caused an increase in waiting lists, making it necessary to find ways to expand the number of organs available for transplantation. Normothermic perfusion (NMP) of liver grafts has been established as an alternative to static cold storage (SCS), but only a small number of perfusion machines are commercially available. METHODS: Using a customized ex situ machine perfusion, we compared the results between ex situ NMP and SCS preservation in a porcine liver transplant model. RESULTS: During NMP, lactate concentrations were 80% lower after the 3-h perfusion period, compared with SCS. Bile production had a 2.5-fold increase during the NMP period. After transplantation, aspartate transaminase (AST) and alanine transaminase (ALT) levels were 35% less in the NMP group, compared to the SCS group. In pathologic analyses of grafts after transplant, tissue oxidation did not change between groups, but the ischemia-reperfusion injury score was lower in the NMP group. CONCLUSION: NMP reduced hepatocellular damage and ischemia-reperfusion injury when compared to SCS using a customized perfusion machine. This could be an alternative for low-income countries to include machine perfusion in their therapeutic options.
Assuntos
Transplante de Fígado , Traumatismo por Reperfusão , Suínos , Animais , Transplante de Fígado/efeitos adversos , Transplante de Fígado/métodos , Preservação de Órgãos/métodos , Perfusão/métodos , Traumatismo por Reperfusão/etiologia , Traumatismo por Reperfusão/prevenção & controle , Traumatismo por Reperfusão/patologia , Bile , Fígado/cirurgia , Fígado/patologiaRESUMO
Introduction: During pregnancy, arterial hypertension may impair placental function, which is critical for a healthy baby's growth. Important proteins during placentation are known to be targets for O-linked ß-N-acetylglucosamine modification (O-GlcNAcylation), and abnormal protein O-GlcNAcylation has been linked to pathological conditions such as hypertension. However, it is unclear how protein O-GlcNAcylation affects placental function and fetal growth throughout pregnancy during hypertension. Methods: To investigate this question, female Wistar and spontaneously hypertensive rats (SHR) were mated with male Wistar rats, and after pregnancy confirmation by vaginal smear, rats were divided into groups of 14, 17, and 20 days of pregnancy (DOPs). On the 14th, 17th, and 20th DOP, rats were euthanized, fetal parameters were measured, and placentas were collected for western blot, immunohistochemical, and morphological analyses. Results: SHR presented a higher blood pressure than the Wistar rats (p=0.001). Across all DOPs, SHR showed reduced fetal weight and an increase in small-for-gestational-age fetuses. While near-term placentas were heavier in SHR (p=0.006), placental efficiency decreased at 17 (p=0.01) and 20 DOPs (p<0.0001) in this group. Morphological analysis revealed reduced junctional zone area and labyrinth vasculature changes on SHR placentas in all DOPs. O-GlcNAc protein expression was lower in placentas from SHR compared with Wistar at 14, 17, and 20 DOPs. Decreased expression of O-GlcNAc transferase (p=0.01) and O-GlcNAcase (p=0.002) enzymes was found at 14 DOPs in SHR. Immunohistochemistry showed reduced placental O-GlcNAc content in both the junctional zone and labyrinth of the placentas from SHR. Periodic acid-Schiff analysis showed decreased glycogen cell content in the placentas from SHR at 14, 17, and 20 DOPs. Moreover, glucose transporter 1 expression was decreased in placentas from SHR in all DOPs. Conclusions: These findings suggest that decreased protein O-GlcNAcylation caused by insufficient placental nutritional apport contributes to placental dysfunction during hypertensive pregnancy, impairing fetal growth.
Assuntos
Hipertensão , Placenta , Feminino , Gravidez , Ratos , Masculino , Animais , Placenta/metabolismo , Ratos Wistar , Ratos Endogâmicos SHR , Placentação , NutrientesRESUMO
It is broadly described that almost every step of the regeneration process requires proper levels of oxygen supply; however, due to the vascular disruption in wounds, oxygen availability is reduced, being detrimental to the regeneration process. Therefore, the development of novel biomaterials combined with improved clinical procedures to promote wound oxygenation is an active field of research in regenerative medicine. This case report derives from a cohort of patients enrolled in a previously published ongoing phase I clinical trial (NCT03960164), to assess safety of photosynthetic scaffolds for the treatment of full skin defects. Here, we present a 56 year old patient, with a scar contracture in the cubital fossa, which impaired the elbow extension significantly affecting her quality of life. As part of the treatment, the scar contracture was removed, and the full-thickness wound generated was surgically covered with a photosynthetic scaffold for dermal regeneration, which was illuminated to promote local oxygen production. Then, in a second procedure, an autograft was implanted on top of the scaffold and the patient's progress was followed for up to 17 months. Successful outcome of the whole procedure was measured as improvement in functionality, clinical appearance, and self-perception of the treated area. This case report underscores the long-term safety and applicability of photosynthetic scaffolds for dermal regeneration and their stable compatibility with other surgical procedures such as autograft application. Moreover, this report also shows the ability to further improve the clinical outcome of this procedure by means of dermal vacuum massage therapy and, more importantly, shows an overall long-term improvement in patient´s quality of life, supporting the translation of photosynthetic therapies into human patients.
RESUMO
As hypoxic tumors show resistance to several clinical treatments, photosynthetic microorganisms have been recently suggested as a promising safe alternative for oxygenating the tumor microenvironment. The relationship between organisms and the effect microalgae have on tumors is still largely unknown, evidencing the need for a simple yet representative model for studying photosynthetic tumor oxygenation in a reproducible manner. Here, we present a 3D photosynthetic tumor model composed of human melanoma cells and the microalgae Chlamydomonas reinhardtii, both seeded into a collagen scaffold, which allows for the simultaneous study of both cell types. This work focuses on the biocompatibility and cellular interactions of the two cell types, as well as the study of photosynthetic oxygenation of the tumor cells. It is shown that both cell types are biocompatible with one another at cell culture conditions and that a 10:1 ratio of microalgae to cells meets the metabolic requirement of the tumor cells, producing over twice the required amount of oxygen. This 3D tumor model provides an easy-to-use in vitro resource for analyzing the effects of photosynthetically produced oxygen on a tumor microenvironment, thus opening various potential research avenues.
Assuntos
Chlamydomonas reinhardtii , Microalgas , Neoplasias , Humanos , Microalgas/metabolismo , Fotossíntese , Chlamydomonas reinhardtii/metabolismo , Oxigênio/metabolismo , Comunicação Celular , Microambiente TumoralRESUMO
Antiphospholipid syndrome (APS) is an autoimmune disorder characterized by pregnancy morbidity or thrombosis and persistent antiphospholipid antibodies (aPL) that bind to the endothelium and induce endothelial activation, which is evidenced by the expression of adhesion molecules and the production of reactive oxygen species (ROS) and subsequent endothelial dysfunction marked by a decrease in the synthesis and release of nitric oxide (NO). These endothelial alterations are the key components for the development of severe pathological processes in APS. Patients with APS can be grouped according to the presence of other autoimmune diseases (secondary APS), thrombosis alone (thrombotic APS), pregnancy morbidity (obstetric APS), and refractoriness to conventional treatment regimens (refractory APS). Typically, patients with severe and refractory obstetric APS exhibit thrombosis and are classified as those having primary or secondary APS. The elucidation of the mechanisms underlying these alterations according to the different groups of patients with APS could help establish new therapies, particularly necessary for severe and refractory cases. Therefore, this study aimed to evaluate the differences in endothelial activation and dysfunction induced by aPL between patients with refractory obstetric APS and other APS clinical manifestations. Human umbilical vein endothelial cells (HUVECs) were stimulated with polyclonal immunoglobulin-G (IgG) from different groups of patients n = 21), including those with primary (VTI) and secondary thrombotic APS (VTII) and refractory primary (RI+), refractory secondary (RII+), and non-refractory primary (NR+) obstetric APS. All of them with thrombosis. The expression of adhesion molecules; the production of ROS, NO, vascular endothelial growth factor (VEGF), and endothelin-1; and the generation of microparticles were used to evaluate endothelial activation and dysfunction. VTI IgG induced the expression of adhesion molecules and the generation of microparticles and VEGF. RI+ IgG induced the expression of adhesion molecules and decreased NO production. RII+ IgG increased the production of microparticles, ROS, and endothelin-1 and reduced NO release. NR+ IgG increased the production of microparticles and endothelin-1 and decreased the production of VEGF and NO. These findings reveal differences in endothelial activation and dysfunction among groups of patients with APS, which should be considered in future studies to evaluate new therapies, especially in refractory cases.
RESUMO
Insufficient oxygen supply represents a relevant issue in several fields of human physiology and medicine. It has been suggested that the implantation of photosynthetic cells can provide oxygen to tissues in the absence of a vascular supply. This approach has been demonstrated to be successful in several in vitro and in vivo models; however, no data is available about their safety in human patients. Here, an early phase-1 clinical trial (ClinicalTrials.gov identifier: NCT03960164, https://clinicaltrials.gov/ct2/show/NCT03960164) is presented to evaluate the safety and feasibility of implanting photosynthetic scaffolds for dermal regeneration in eight patients with full-thickness skin wounds. Overall, this trial shows that the presence of the photosynthetic microalgae Chlamydomonas reinhardtii in the implanted scaffolds did not trigger any deleterious local or systemic immune responses in a 90 days follow-up, allowing full tissue regeneration in humans. The results presented here represent the first attempt to treat patients with photosynthetic cells, supporting the translation of photosynthetic therapies into clinics. Clinical Trial Registration: www.clinicaltrials.gov/ct2/show/NCT03960164, identifier: NCT03960164.
RESUMO
Estrogenic steroids and adenosine A2A receptors promote the wound healing and angiogenesis processes. However, so far, it is unclear whether estrogen may regulate the expression and pro-angiogenic activity of A2A receptors. Using in vivo analyses, we showed that female wild type (WT) mice have a more rapid wound healing process than female or male A2A-deficient mice (A2AKO) mice. We also found that pulmonary endothelial cells (mPEC) isolated from female WT mice showed higher expression of A2A receptor than mPEC from male WT mice. mPEC from female WT mice were more sensitive to A2A-mediated pro-angiogenic response, suggesting an ER and A2A crosstalk, which was confirmed using cells isolated from A2AKO. In those female cells, 17ß-estradiol potentiated A2A-mediated cell proliferation, an effect that was inhibited by selective antagonists of estrogen receptors (ER), ERα, and ERß. Therefore, estrogen regulates the expression and/or pro-angiogenic activity of A2A adenosine receptors, likely involving activation of ERα and ERß receptors. Sexual dimorphism in wound healing observed in the A2AKO mice process reinforces the functional crosstalk between ER and A2A receptors.
Assuntos
Estradiol/farmacologia , Receptor alfa de Estrogênio/genética , Receptor beta de Estrogênio/genética , Neovascularização Fisiológica/efeitos dos fármacos , Receptor A2A de Adenosina/genética , Ferimentos Penetrantes/genética , Adenosina/análogos & derivados , Adenosina/farmacologia , Animais , Proliferação de Células/efeitos dos fármacos , Células Endoteliais/citologia , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Receptor alfa de Estrogênio/antagonistas & inibidores , Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/antagonistas & inibidores , Receptor beta de Estrogênio/metabolismo , Feminino , Regulação da Expressão Gênica , Pulmão/citologia , Pulmão/metabolismo , Masculino , Camundongos , Camundongos Knockout , Neovascularização Fisiológica/genética , Fenetilaminas/farmacologia , Pirazóis/farmacologia , Pirimidinas/farmacologia , Receptor Cross-Talk , Receptor A2A de Adenosina/metabolismo , Fatores Sexuais , Transdução de Sinais , Cicatrização/efeitos dos fármacos , Cicatrização/genética , Ferimentos Penetrantes/tratamento farmacológico , Ferimentos Penetrantes/metabolismo , Ferimentos Penetrantes/patologiaRESUMO
The placenta is a transitory organ, located between the mother and the foetus, which supports intrauterine life. This organ has nutritional, endocrine and immunologic functions to support foetal development. Several factors are related to the correct functioning of the placenta including foetal and maternal blood flow, appropriate nutrients, expression and function of receptors and transporters, and the morphology of the placenta itself. Placental morphology is crucial for understanding the pathophysiology of the organ as represents the physical structure where nutrient exchange occurs. In pathologies of pregnancy such as diabetes mellitus in humans and animal models, several changes in the placental morphology occur, related mainly with placental size, hypervascularization, higher branching capillaries of the villi and increased glycogen deposits among others. Gestational diabetes mellitus is associated with modifications in the structure of the human placenta including changes in the surface area and volume, as well as histological changes including an increased volume of intervillous space and terminal villi, syncytiotrophoblast number, fibrinoid areas, and glycogen deposits. These modifications may result in functional changes in this organ thus limiting the wellbeing of the developing foetus. This review gives an overview of recurrent morphological changes at macroscopic and histological levels seen in the placenta from gestational diabetes in humans and animal models. This article is part of a Special Issue entitled: Membrane Transporters and Receptors in Pregnancy Metabolic Complications edited by Luis Sobrevia.
Assuntos
Diabetes Gestacional/metabolismo , Placenta/metabolismo , Placenta/patologia , Animais , Diabetes Mellitus Experimental , Feminino , Desenvolvimento Fetal , Humanos , Gravidez , RoedoresRESUMO
Successful placentation is a key event for fetal development, which commences following embryo implantation into the uterine wall, eliciting decidualization, placentation, and remodeling of blood vessels to provide physiological exchange between embryo-fetus and mother. Several signaling pathways are recruited to modulate such important processes and specific proteins that regulate placental function are a target for the glycosylation with O-linked ß-N-acetylglucosamine (O-GlcNAc), or O-GlcNAcylation. This is a reversible post-translational modification on nuclear and cytoplasmic proteins, mainly controlled by O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). O-GlcNAcylation has been implicated as a modulator of proteins, both in physiological and pathological conditions and, more recently, O-GlcNAc has also been shown to be an important modulator in placental tissue. In this mini-review, the interplay between O-GlcNAcylation of proteins and placental function will be addressed, discussing the possible implications of this post-translational modification through placental development and pregnancy.
RESUMO
Introduction: Fetal growth restriction may be the consequence of maternal, fetal, or placental factors. The insulin-like growth factors (IGFs) are major determinants of fetal growth, and are expressed in the mother, fetus and placenta in most species. Previously we reported higher placental protein content of IGF-I, IGF-IR, and AKT in small (SGA) compared with those from appropriate for gestational age (AGA) placentas. The protein Klotho, has been reported in placenta and may regulate IGF-I activity. In this study we determined Klotho gene expression and protein immunostaining in term (T-SGA y T-AGA) and preterm (PT-SGA y PT-AGA) human placentas. In addition, we assessed the effect of Klotho on the IGF-IR and AKT activation induced by IGF-I. Methods: Placentas (n = 1 17) from 32 T-SGA (birth weight (BW) = -1.74 ± 0.08 SDS), 37 T-AGA (BW = 0.12 ± 0.12 SDS), 20 PT-SGA (BW = -2.08 ± 0.14 SDS), and 28 PT-AGA (BW = -0.43 ± 0.13 SDS) newborns were collected. mRNA expression by RT-PCR in the chorionic (CP) and basal (BP) plates of the placentas, and the presence of Klotho was evaluated by immunohistochemistry (integral optical density, IOD). In addition, we developed placental explants that were incubated with IGF-I in the presence or absence of Klotho. Results: We found a lower mRNA expression and protein immunoreactivity of Klotho in the CP of SGA (term and preterm) compared with AGA placentas. We also observed a significant reduction in IGF-IR tyrosine activation induced by IGF-I 10 nM when preincubated with 2.0 nM of Klotho (2.4 ± 0.5 arbitrary units vs. 1.3 ± 0.3 AU), and similar results we observed on AKT and ERK42/44 activation. Conclusion: We describe for the first time that Klotho mRNA and protein varies according to fetal growth and gestational age. In addition, Klotho appears to down-regulate the activation induced by IGF-I on IGF-IR and AKT, suggesting that Klotho may be regulating IGF-I activity in human placentas according to intrauterine fetal growth.
RESUMO
PROBLEM: Women with antiphospholipid antibodies (aPLs) present a risk of pregnancy morbidity (PM), vascular thrombosis (VT), or both (PM/VT). aPLs affect trophoblast function, and the aim of this study was to determine the modulation of this aPL-induced damage by different drugs. METHOD OF STUDY: IgG was obtained from women with PM and PM/VT positive to aPLs. Binding of IgG to trophoblastic cells, proliferation, mitochondrial membrane integrity, and trophoblast invasion were assessed. The effect of enoxaparin, aspirin, and aspirin-triggered lipoxin (ATL) were evaluated as well as signal transducer and activator of transcription 3 (STAT3) phosphorylation. RESULTS: IgG from women with aPLs strongly binds to trophoblastic cells. Integrity of mitochondrial membrane was reduced, and proliferation was increased by IgG-PM/VT. Both IgG-PM and IgG-PM/VT decreased trophoblast invasion, which was restored by enoxaparin, aspirin, and ATL. IgG-PM triggered reduction in STAT3 phosphorylation. CONCLUSION: Some drugs used to prevent aPL-induced PM modulated the alteration of trophoblast function.
Assuntos
Anticorpos Antifosfolipídeos/imunologia , Síndrome Antifosfolipídica/complicações , Complicações na Gravidez/imunologia , Trofoblastos/imunologia , Adulto , Anticoagulantes/farmacologia , Síndrome Antifosfolipídica/imunologia , Aspirina/farmacologia , Western Blotting , Feminino , Citometria de Fluxo , Humanos , Imunoglobulina G/imunologia , Imuno-Histoquímica , Lipoxinas/farmacologia , Gravidez , Trofoblastos/efeitos dos fármacos , Trofoblastos/patologiaRESUMO
OBJECTIVES: Human tissues are usually studied using a series of two-dimensional visualizations of in vivo or cutout specimens. However, there is no precise anatomical description of some of the processes of human fetal development. The purpose of our study is to develop a quantitative description of the normal axial skeleton by means of high-resolution three-dimensional magnetic resonance (MR) images, collected from six normal 20-week-old human fetuses fixed in formaldehyde. METHODS: Fetuses were collected after spontaneous abortion and subsequently fixed with formalin. They were imaged using a 1.5 T MR scanner with an isotropic spatial resolution of 200 µm. The correct tissue discrimination between ossified and cartilaginous bones was confirmed by comparing the images achieved by MR scans and computerized axial tomographies. The vertebral column was segmented out from each image using a specially developed semi-automatic algorithm. RESULTS: Vertebral body dimensions and inter-vertebral distances were larger in the lumbar region, in agreement with the beginning of the ossification process from the thoracolumbar region toward the sacral and cephalic ends. CONCLUSION: In this article, we demonstrate the feasibility of using MR images to study the ossification process in formalin-fixed fetal tissues. A quantitative description of the ossification centers of vertebral bodies and arches is presented.
Assuntos
Feto Abortado/diagnóstico por imagem , Osso e Ossos/diagnóstico por imagem , Osso e Ossos/embriologia , Imageamento por Ressonância Magnética , Osteogênese/fisiologia , Segundo Trimestre da Gravidez , Feto Abortado/anatomia & histologia , Feto Abortado/efeitos dos fármacos , Feto Abortado/embriologia , Aborto Espontâneo/diagnóstico por imagem , Padronização Corporal/fisiologia , Densidade Óssea/fisiologia , Osso e Ossos/anatomia & histologia , Osso e Ossos/fisiologia , Feminino , Fêmur/anatomia & histologia , Fêmur/diagnóstico por imagem , Formaldeído/farmacologia , Idade Gestacional , Humanos , Imageamento por Ressonância Magnética/métodos , Masculino , Tamanho do Órgão , Gravidez , RadiografiaRESUMO
CONTEXT: In humans, IGF-I and -II have an important role in pre- and postnatal growth. The IGFs circulate in plasma principally as a ternary complex with the IGF binding protein-3 and an acid-labile subunit (ALS), which increases their half life. OBJECTIVES: The objectives of the study were to determine whether the human placenta expresses the mRNA and protein for ALS and to evaluate any possible differences in the mRNA and protein for ALS in placentas from small (SGA) and appropriate (AGA) or gestational age newborns. SUBJECTS/METHODS: We studied the placentas from 47 AGA and 42 SGA pregnancies. IGF-I, IGF-II, IGF binding protein-3, and ALS placental mRNA and protein contents were determined in both the basal and the chorionic plates of the placenta. RESULTS: We observed that the human placenta expresses the gene and protein for ALS. The ALS mRNA in SGA was higher compared with AGA placentas (0.15 ± 0.01 vs. 0.12 ± 0.01 arbitrary units, respectively, P < 0.05). In addition, the ALS protein content in SGA (31.7 ± 3.3 pmol/g) was higher compared with AGA (22.1 ± 2.3 pmol/g, P < 0.05) placentas. CONCLUSION: We describe that the human placenta expresses the mRNA and the protein for ALS, and we observed an increase in ALS mRNA expression and protein content in SGA compared with AGA placentas.
Assuntos
Peso ao Nascer/genética , Proteínas de Transporte/metabolismo , Glicoproteínas/metabolismo , Placenta/metabolismo , Western Blotting , Proteínas de Transporte/genética , Feminino , Expressão Gênica , Idade Gestacional , Glicoproteínas/genética , Humanos , Imuno-Histoquímica , Recém-Nascido , Proteína 3 de Ligação a Fator de Crescimento Semelhante à Insulina/genética , Proteína 3 de Ligação a Fator de Crescimento Semelhante à Insulina/metabolismo , Fator de Crescimento Insulin-Like I/genética , Fator de Crescimento Insulin-Like I/metabolismo , Fator de Crescimento Insulin-Like II/genética , Fator de Crescimento Insulin-Like II/metabolismo , Gravidez , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Estatísticas não ParamétricasRESUMO
INTRODUCTION: Erectile dysfunction is considered an early clinical manifestation of vascular disease and an independent risk factor for cardiovascular events associated with endothelial dysfunction and increased levels of pro-inflammatory cytokines. Tumor necrosis factor-alpha (TNF-alpha), a pro-inflammatory cytokine, suppresses endothelial nitric oxide synthase (eNOS) expression. AIM: Considering that nitric oxide (NO) is of critical importance in penile erection, we hypothesized that blockade of TNF-alpha actions would increase cavernosal smooth muscle relaxation. METHODS: In vitro organ bath studies were used to measure cavernosal reactivity in wild type and TNF-alpha knockout (TNF-alpha KO) mice and NOS expression was evaluated by western blot. In addition, spontaneous erections (in vivo) were evaluated by videomonitoring the animals (30 minutes). Collagen and elastin expression were evaluated by Masson trichrome and Verhoff-van Gieson stain reaction, respectively. MAIN OUTCOME MEASURES: Corpora cavernosa from TNF-alpha KO mice exhibited increased NO-dependent relaxation, which was associated with increased eNOS and neuronal NOS (nNOS) cavernosal expression. RESULTS: Cavernosal strips from TNF-alpha KO mice displayed increased endothelium-dependent (97.4 +/- 5.3 vs. CONTROL: 76.3 +/- 6.3, %) and nonadrenergic-noncholinergic (93.3 +/- 3.0 vs. CONTROL: 67.5 +/- 16.0; 16 Hz) relaxation compared to control animals. These responses were associated with increased protein expression of eNOS and nNOS (P < 0.05). Sympathetic-mediated (0.69 +/- 0.16 vs. CONTROL: 1.22 +/- 0.22; 16 Hz) as well as phenylephrine-induced contractile responses (1.6 +/- 0.1 vs. CONTROL: 2.5 +/- 0.1, mN) were attenuated in cavernosal strips from TNF-alpha KO mice. Additionally, corpora cavernosa from TNF-alpha KO mice displayed increased collagen and elastin expression. In vivo experiments demonstrated that TNF-alpha KO mice display increased number of spontaneous erections. CONCLUSION: Corpora cavernosa from TNF-alpha KO mice display alterations that favor penile tumescence, indicating that TNF-alpha plays a detrimental role in erectile function. A key role for TNF-alpha in mediating endothelial dysfunction in ED is markedly relevant since we now have access to anti-TNF-alpha therapies.
Assuntos
Disfunção Erétil/imunologia , Disfunção Erétil/terapia , Músculo Liso/imunologia , Fator de Necrose Tumoral alfa/imunologia , Vasodilatação/fisiologia , Animais , Colágeno/metabolismo , Elastina/metabolismo , Endotélio Vascular/imunologia , Endotélio Vascular/metabolismo , Endotélio Vascular/patologia , Disfunção Erétil/metabolismo , Masculino , Camundongos , Camundongos Knockout , Músculo Liso/metabolismo , Músculo Liso/patologia , Óxido Nítrico Sintase/metabolismo , PênisRESUMO
Primordial germ cells (PGCs) are the progenitor cells of the vertebrate germ line. These cells originate outside of the embryo and, through separation, migration, and colonization, arrive at the genital ridge, contributing to gonad development. Diverse extracellular matrix molecules are present along the PGC migratory pathway, permitting or inhibiting PGC displacement. Collagens and tenascin form the substratum for in vitro migration of neural crest cells and PGCs. However, little is known about the expression and distribution of these molecules during in situ PGC migration. Using immunohistochemistry, we identified tenascin-C and types I, III, and V collagen along the mouse PGC migration pathway. These molecules were spatiotemporally expressed in basement membranes of hindgut, coelomic epithelia, and mesonephric tubules and mesenchyme throughout the study. Our results complement previous data from our laboratory and contribute to building comprehension of the composition of the mouse PGC migratory pathway extracellular matrix, thereby enhancing understanding of the process.
Assuntos
Movimento Celular/fisiologia , Colágeno/metabolismo , Células Germinativas/citologia , Gônadas/embriologia , Tenascina/metabolismo , Animais , Colágeno/classificação , Matriz Extracelular/metabolismo , Feminino , Células Germinativas/metabolismo , Idade Gestacional , Gônadas/citologia , Técnicas Imunoenzimáticas , Masculino , Camundongos , Camundongos Endogâmicos , Organogênese , GravidezRESUMO
Primordial germ cells are an embryonic cell line that give rise to gametes in vertebrates. They originate outside the embryo proper and migrate by a well-defined route to the genital ridges. Proteoglycans and glycosaminoglycans have distinctive properties that affect many of the characteristics of the extracellular microenvironment of migratory pathways in a variety of developmental systems. The purpose of this work was to identify the proteoglycans and glycosaminoglycans that are spatially and temporally expressed in the migratory pathway of primordial germ cells. We showed that the expression of proteoglycans and glycosaminoglycans in the primordial germ cells migratory pathway changes according to the different phases of the migratory process. Some molecules such as chondroitin-0-sulfate, decorin, and biglycan are present only in certain phases of the migratory process of primordial germ cells. Heparan sulfate, chondroitin-6-sulfate, versican, perlecan, and syndecan-4, although exhibiting some variation in expression were detected during all phases of the migratory process. Our results indicate that the successive steps of primordial germ cell migration require a coordinated expression of proteoglycans and glycosaminoglycans, that should be present in appropriate levels and in specific areas of the embryo, and that the sequential expression of these extracellular matrix molecules is under a genetic program that appears to be common to a variety of cell types during embryonic development.
Assuntos
Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Células Germinativas/citologia , Células Germinativas/metabolismo , Glicosaminoglicanos/biossíntese , Proteoglicanas/biossíntese , Animais , Movimento Celular , Embrião de Mamíferos , Proteínas da Matriz Extracelular/biossíntese , Proteínas da Matriz Extracelular/fisiologia , Glicosaminoglicanos/classificação , Glicosaminoglicanos/fisiologia , Camundongos , Proteoglicanas/classificação , Proteoglicanas/fisiologia , Fatores de TempoRESUMO
In the present report we followed the distribution of hyaluronan during the phases of separation, migration, and colonization of the primordial germ cell migratory process. Hyaluronan was detected by the use of two cytochemical methods: (1) ruthenium hexammine trichloride (RHT) associated with enzymatic treatment with hyaluronate lyase and (2) a binding specific probe for hyaluronan. After RHT treatment the proteoglycans and/or glycosaminoglycans were observed as a meshwork formed by electron-dense granules connected by thin filaments. After enzymatic digestion, no filaments could be detected in the migratory pathway. Quantitative analysis showed a close correlation between cell migration and the concentration of RHT-positive filaments. It was also shown that high amounts of hyaluronan were expressed in the separation phase and migration phases whereas during the colonization phase the amount of hyaluronan was clearly diminished. This study showed that the presence of primordial germ cells in each compartment of the migratory pathway was always accompanied by a high expression of hyaluronan. These results indicate that hyaluronan is an important molecule in the migratory process, providing the primordial germ cells with a hydrated environment that facilitates their movement toward the genital ridges.