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Background: There is a strong interest in designing new scaffolds for their potential application in tissue engineering and regenerative medicine. The incorporation of functionalization molecules can lead to the enhancement of scaffold properties, resulting in variations in scaffold compatibility. Therefore, the efficacy of the therapy could be compromised by the foreign body reaction triggered after implantation. Methods: In this study, the biocompatibilities of three scaffolds made from an alginate-chitosan combination and functionalized with gold nanoparticles (AuNp) and alginate-coated gold nanoparticles (AuNp + Alg) were evaluated in a subcutaneous implantation model in Wistar rats. Scaffolds and surrounding tissue were collected at 4-, 7- and 25-day postimplantation and processed for histological analysis and quantification of the expression of genes involved in angiogenesis, macrophage profile, and proinflammatory (IL-1ß and TNFα) and anti-inflammatory (IL-4 and IL-10) cytokines. Results: Histological analysis showed a characteristic foreign body response that resolved 25 days postimplantation. The intensity of the reaction assessed through capsule thickness was similar among groups. Functionalizing the device with AuNp and AuNp + Alg decreased the expression of markers associated with cell death by apoptosis and polymorphonuclear leukocyte recruitment, suggesting increased compatibility with the host tissue. Similarly, the formation of many foreign body giant cells was prevented. Finally, an increased detection of alpha smooth muscle actin was observed, showing the angiogenic properties of the elaborated scaffolds. Conclusion: Our results show that the proposed scaffolds have improved biocompatibility and exhibit promising potential as biomaterials for elaborating tissue engineering constructs.
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Hyperglycemia during gestation can disrupt fetal heart development and increase postnatal cardiovascular disease risk. It is therefore imperative to identify early biomarkers of hyperglycemia during gestation-induced fetal heart damage and elucidate the underlying molecular pathomechanisms. Clinical investigations of diabetic adults with heart dysfunction and transgenic mouse studies have revealed that overexpression or increased expression of TNNI3K, a heart-specific kinase that binds troponin cardiac I, may contribute to abnormal cardiac remodeling, ventricular hypertrophy, and heart failure. Optimal heart function also depends on the precise organization of contractile and excitable tissues conferred by intercellular occlusive, adherent, and communicating junctions. The current study evaluated changes in embryonic heart development and the expression levels of sarcomeric proteins (troponin I, desmin, and TNNI3K), junctional proteins, glucose transporter-1, and Ki-67 under fetal hyperglycemia. Stage 22HH Gallus domesticus embryos were randomly divided into two groups: a hyperglycemia (HG) group, in which individual embryos were injected with 30 mmol/L glucose solution every 24 h for 10 days, and a no-treatment (NT) control group, in which individual embryos were injected with physiological saline every 24 h for 10 days (stage 36HH). Embryonic blood glucose, height, and weight, as well as heart size, were measured periodically during treatment, followed by histopathological analysis and estimation of sarcomeric and junctional protein expression by western blotting and immunostaining. Hyperglycemic embryos demonstrated delayed heart maturation, with histopathological analysis revealing reduced left and right ventricular wall thickness (-39% and -35% vs. NT). Immunoexpression levels of TNNI3K and troponin 1 increased (by 37% and 39%, respectively), and desmin immunofluorescence reduced (by 23%). Embryo-fetal hyperglycemia may trigger an increase in the expression levels of TNNI3K and troponin I, as well as dysfunction of occlusive and adherent junctions, ultimately inducing abnormal cardiac remodeling.
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Natural biopolymer scaffolds and conductive nanomaterials have been widely used in cardiac tissue engineering; however, there are still challenges in the scaffold fabrication, which include enhancing nutrient delivery, biocompatibility and properties that favor the growth, maturation and functionality of the generated tissue for therapeutic application. In the present work, different scaffolds prepared with sodium alginate and chitosan (alginate/chitosan) were fabricated with and without the addition of metal nanoparticles and how their fabrication affects cardiomyocyte growth was evaluated. The scaffolds (hydrogels) were dried by freeze drying using calcium gluconate as a crosslinking agent, and two types of metal nanoparticles were incorporated, gold (AuNp) and gold plus sodium alginate (AuNp+Alg). A physicochemical characterization of the scaffolds was carried out by swelling, degradation, permeability and infrared spectroscopy studies. The results show that the scaffolds obtained were highly porous (>90%) and hydrophilic, with swelling percentages of around 3000% and permeability of the order of 1 × 10−8 m2. In addition, the scaffolds proposed favored adhesion and spheroid formation, with cardiac markers expression such as tropomyosin, troponin I and cardiac myosin. The incorporation of AuNp+Alg increased cardiac protein expression and cell proliferation, thus demonstrating their potential use in cardiac tissue engineering.
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The damage to the gastrointestinal mucosa induced by ischemia/reperfusion (I/R) is closely related to high mortality in critically ill patients, which is attributable, in part, to the lack of an early method of diagnosis to show the degree of ischemia-induced injury in this type of patients. Electrical Impedance Spectroscopy (EIS) has been shown to be a tool to early diagnose gastric mucosal damage induced by ischemia. A therapeutic alternative to reduce this type of injury is melatonin (MT), which has gastroprotective effects in I/R models. In this work, the effect of treatment with MT on the electrical properties of gastric tissue, biomarkers of inflammatory (iNOS and COX-2), proliferation, and apoptotic process under I/R conditions in male Wistar rats was evaluated through EIS, histological and immunohistochemical analysis. Treatment with MT prevents gastric mucosa damage, causing a decrease in gastric impedance parameters related to the inflammatory process and cellular damage. This suggests that EIS could be used as a tool to diagnose and monitor the evolution of gastric mucosal injury, as well as in the recovery process in critically ill patients.
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Melatonina , Traumatismo por Reperfusão , Gastropatias , Animais , Biomarcadores , Estado Terminal , Impedância Elétrica , Mucosa Gástrica/patologia , Isquemia/patologia , Masculino , Melatonina/farmacologia , Melatonina/uso terapêutico , Ratos , Ratos Wistar , Reperfusão , Traumatismo por Reperfusão/patologia , Gastropatias/patologiaRESUMO
Jatropha neopauciflora is an endemic species of Mexico. Its latex is used to treat wounds, scarring, oral infections, and loose teeth. To date, there are no studies that validate at a morphological level a wound-healing use in diabetes. The present research aimed to evaluate the wound-healing capacity of the latex of J. neopauciflora in the skin of healthy and streptozotocin-induced diabetic mice. Also, a chemical analysis of the latex through molecular exclusion chromatography and HPLC were performed. Male mice (Mus musculus) of 7-week-old CD1 strain were used. Groups of healthy and diabetic mice were formed. A longitudinal cut of 1 cm was performed on the depilated skin. All treatments were topically applied to the wound area twice a day for ten days. At the end of the experiments, the skin sections were obtained from the wound area and stained with Hematoxylin-Eosin. Then we counted the number of active fibroblasts in all the experimental groups. In normal mice, the latex accelerated the wound-healing process and decreased the number of active fibroblasts, similarly to Recoveron. In diabetic mice, the latex and Recoveron increased the number of active fibroblasts. In normal and diabetic mice, a thin and orderly epidermis was observed. Molecular exclusion chromatography exhibited 58 fractions, 14 of which were subjected to HPLC, to detect catechin, a flavonoid with antioxidant, antimicrobial, and anti-inflammatory properties. J. neopauciflora latex can be useful for wound treatment in patients with diabetes mellitus because it accelerates and promotes the wound-healing process.
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Diabetes Mellitus Experimental , Jatropha , Animais , Diabetes Mellitus Experimental/tratamento farmacológico , Humanos , Látex , Camundongos , Pele , CicatrizaçãoRESUMO
In Central and South American traditional medicine, people use Cecropia obtusifolia Bertol (Cecropiaceae) for the treatment of diabetes mellitus. However, its hypoglycemic action mechanism at pancreatic and liver level has been poorly explored. The present research aimed to establish the influence of the aqueous extract of C. obtusifolia, standardized in its content of chlorogenic acid, on insulin secretion in RINm5F cells and over the liver carbohydrates and lipids metabolism, and to determine concomitantly its hepatoprotective effect on mice with streptozotocin-induced diabetes. In RINm5F cells, concentrations 5, 50, 100, and 200 µg/mL of aqueous extract of C. obtusifolia were used to determine [Ca2+]i and insulin secretion. In an acute study, the extract was administered at doses of 500 mg/kg. In another test (subacute), the extract was daily administrated to diabetic mice (200 mg/kg/day) for 30 days. Blood glucose levels and other biochemical parameters were determined, and a liver histological analysis was performed. In RINm5F cells, C. obtusifolia increased [Ca2+]i and insulin secretion, whereas in diabetic mice exhibited acute and subacute hypoglycemic effects. Daily administration of C. obtusifolia to diabetic mice also increased liver glycogen storage and glycogen synthase levels, without apparent changes in gluconeogenesis. Besides, it increased peroxisome proliferator-activated receptor-α (PPAR-α) and long-chain-fatty-acid-CoA ligase 1 (ACSL-1) expression and reduced triglycerides, transaminases (alanine aminotransferase and aspartate aminotransferase), and collagen fibers, modifying anti-inflammatory (adiponectin and interleukin-10) and inflammatory (tumor necrosis factor-α) cytokines in serum. Therefore, the hypoglycemic effect of C. obtusifolia implicates a dual action, promoting insulin secretion, liver glycogen accumulation, and hepatoprotection by decreasing collagen fibers and inflammatory markers, whereas it improves lipid metabolism, due in part to PPAR-α.
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Cecropia/química , Diabetes Mellitus Experimental , Hipoglicemiantes/uso terapêutico , Extratos Vegetais/uso terapêutico , Animais , Glicemia , Diabetes Mellitus Experimental/tratamento farmacológico , Fígado/efeitos dos fármacos , Camundongos , Fitoterapia , Substâncias Protetoras/uso terapêuticoRESUMO
Damage to the gastrointestinal mucosa caused by ischemia - reperfusion is a significant clinical problem associated with various physiopathological conditions. Our group has conducted various studies in patients in critical conditions and in animal models to identify early damage to the gastric mucosa under ischemia using impedance spectroscopy. It is important to perform a quantitative histopathological analysis which can be linked to changes in impedance of the gastric mucosa under conditions of ischemia and I/R. AIM: To propose a tissue lesion index which considers pathological alterations inherent to the inflammatory process and cell damage which may be directly related to changes in impedance under conditions of ischemia and I/R. METHODS: The animals were randomly distributed into 4 groups: control, ischemia (30 min), and I/R (30 and 60 min). Qualitative histopathological analysis was performed; the vascular area, glandular lumen area, the number of damaged cells, and the depth of the erosion were also quantified to obtain a scale to propose a tissue lesion index (TLI). RESULTS: Under ischemic conditions, histopathological analysis showed edema and necrosis in epithelial cells, and vascular congestion. In I/R (30 and 60 min) conditions, areas of epithelial erosion were generated. Damage was classified based on the TLI. A TLI threshold of 3 showed a predictive value of tissue lesion. CONCLUSION: The proposed gastric lesion index allows us to objectively quantify and classify damage to the gastric mucosa produced by I/R.
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Células Epiteliais/patologia , Mucosa Gástrica/patologia , Traumatismo por Reperfusão/patologia , Animais , Modelos Animais de Doenças , Edema/patologia , Masculino , Necrose , Ratos WistarRESUMO
Gastrointestinal ischemia/reperfusion (I/R) generates pathological alterations that could lead to death. Early ischemic damage markers could be used to guide therapy and improve outcomes. AIM: To relate hypoxia-inducible factor 1α (HIF-1α) activation and inducible nitric oxide synthase (iNOS) expression to gastric impedance changes due to I/R damage. METHODS: Experimental animals were randomly distributed into 3 groups: control, ischemia (30 min) and I/R (60 min). Gastric ischemia was generated by celiac artery clamping for 30 min, and then blood flow was restored for 60 min. Impedance spectra and biopsies of the glandular portion were obtained for histological and immunohistochemical analyses. Immunodetection of both HIF-1α and iNOS was performed. RESULTS: Under ischemia and I/R conditions, there was an increase (p<0.05) in the impedance parameters. Histologically, under ischemic conditions, edema and necrosis were observed in epithelium and significant vascular congestion. In I/R condition, alterations of the glandular and luminal integrity were found, which generated areas of epithelial erosion. Immunohistochemical analysis of HIF-1α revealed an increase (p<0.01) in the number of immunoreactive cells in the ischemia (35.7±13.9) and I/R (119.9±18.8) conditions compared to the control (0.8±1.2). Immunodetection of iNOS showed an increase (p<0.01) in the number of cells expressing iNOS under the ischemia (5.4±2.9) and I/R conditions (27.4±11.3) was observed compared to the control (0.4±0.8). CONCLUSION: Early changes in impedance in response to I/R is related to histopathological changes, the nuclear stabilization and translocation of HIF-1α as well as expression of iNOS.
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Mucosa Gástrica/enzimologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Imuno-Histoquímica , Óxido Nítrico Sintase Tipo II/metabolismo , Traumatismo por Reperfusão/enzimologia , Gastropatias/enzimologia , Transporte Ativo do Núcleo Celular , Animais , Biópsia , Modelos Animais de Doenças , Edema/enzimologia , Edema/patologia , Impedância Elétrica , Mucosa Gástrica/patologia , Masculino , Necrose , Estabilidade Proteica , Ratos Wistar , Traumatismo por Reperfusão/patologia , Gastropatias/patologia , Fatores de TempoRESUMO
The parasympathetic nervous system has a crucial role in immunomodulation of the vagus nerve, its structure provides a pathogen detection system, and a negative feedback to the immune system after the pathogenic agent has been eliminated. Amebiasis is a disease caused by the protozoan parasite Entamoeba histolytica, considered the third leading cause of death in the world. The rats are used as a natural resistance model to amoebic liver infection. The aim of this study is to analyze the interaction of Entamoeba histolytica with neutrophils, macrophages, and NK cells in livers of intact and vagotomized rats. Six groups were studied (n = 4): Intact (I), Intact + amoeba (IA), Sham (S), Sham + amoeba (SA), Vagotomized (V) and Vagotomized + amoeba (VA). Animals were sacrificed at 8 h post-inoculation of E. histolytica. Then, livers were obtained and fixed in 4% paraformaldehyde. Tissue liver slides were stained with H-E, PAS and Masson. The best development time for E. histolytica infection was at 8 h. Amoeba was identified with a monoclonal anti-220 kDa E. histolytica lectin. Neutrophils (N) were identified with rabbit anti-human neutrophil myeloperoxidase, macrophages (Mɸ) with anti-CD68 antibody and NK cells (NK) with anti-NK. Stomachs weight and liver glycogen were higher in V. Collagen increased in VA, whereas vascular and neutrophilic areas were decreased. There were fewer N, Mɸ, NK around the amoeba in the following order IA > SA > VA (p < 0.05 between IA and VA). In conclusion, these results suggest that the absence of parasympathetic innervation affects the participation of neutrophils, macrophages and NK cells in the innate immune response, apparently by parasympathetic inhibition on the cellular functions and probably for participation in sympathetic activity.
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Entamoeba histolytica/imunologia , Imunidade Inata/fisiologia , Abscesso Hepático Amebiano/imunologia , Nervo Vago/fisiologia , Animais , Colágeno/metabolismo , Imunofluorescência , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/parasitologia , Cinética , Fígado/imunologia , Fígado/parasitologia , Fígado/patologia , Fígado/ultraestrutura , Macrófagos/imunologia , Macrófagos/parasitologia , Masculino , Camundongos , Microscopia Eletrônica de Transmissão , Neutrófilos/imunologia , Neutrófilos/parasitologia , Coelhos , Ratos , Ratos Wistar , Vagotomia , Nervo Vago/cirurgiaRESUMO
All organs of the immune system are innervated and almost all neurotransmitter receptors are present on immune cells. We studied the effects of sympathetic innervation in the development of amebic liver abscess (ALA) in rats. Our results showed that lack of sympathetic innervation promote a decrease in size of ALA. We found scarce amoebas, increased the number of neutrophils and a few collagen fibers surrounding the abscess, meanwhile in control group, we observed abscesses areas with typical necrosis including trophozoites and neutrophils. Macrophages were differentially distributed surrounding abscess area in control and vehicle groups, but equally located in and outside of the abscesses in sympathectomized rat. No significant differences were observed on NK cells in analysed groups. In cytokines quantification studies, we observed down-expression of IFN-g and TNF-a, moreover, we found overexpression of IL-10 in sympathectomized and ALA group. In conclusion, our results suggest that elimination of sympathetic nerve fibers in a model rat of amebic liver abscess induces reduction of the innate immune response and presence of amebas through the liver at seven days post-inoculation.
Todos los órganos del sistema inmune están inervados y casi todos los receptores para neurotransmisores están presentes en las células de la respuesta inmune. Nosotros estudiamos el efecto de la inervación simpática en el desarrollo del Absceso Hepático Amebiano (AHA) en ratas. Nuestros resultados muestran que la inervación simpática promueve una disminución en el tamaño del AHA. Nosotros encontramos áreas fibróticas bien definidas con algunas amibas, mayor número de neutrófilos y pocas fibras de colágena rodeando el área de daño, mientras que en el grupo control, nosotros observamos áreas con necrosis, trofozoítos y pocos neutrófilos en el área fibrótica. Los macrófagos se observaron distribuidos en el área fibrótica en los animales simpatectomizados, mientras que en los controles encontramos a los macrófagos distribuidos en la periferia del absceso. No se encontró diferencia significativa en la distribución y cantidad de células NK. En el estudio de citocinas nosotros observamos una disminución de IFN-g y TNF-a y un incremento de IL-10 en animales simpatectomizados. En conclusión, nuestros resultados sugieren que la eliminación de las fibras del sistema nervioso simpático en el modelo de AHA en rata, reduce la respuesta inmune innata y persisten amebas en el tejido dañados a los 7 días post-inoculación.