RESUMO
Ex situ lung perfusion (ESLP) is used for organ reconditioning, repair, and re-evaluation prior to transplantation. Since valid preclinical animal models are required for translationally relevant studies, we developed a 17 mL low-volume ESLP for double- and single-lung application that enables cost-effective optimal compliance "reduction" of the 3R principles of animal research. In single-lung mode, ten Fischer344 and Lewis rat lungs were subjected to ESLP and static cold storage using STEEN or PerfadexPlus. Key perfusion parameters, thermal lung imaging, blood gas analysis (BGA), colloid oncotic pressure (COP), lung weight gain, histological work-up, and cytokine analysis were performed. Significant differences between perfusion solutions but not between the rat strains were detected. Most relevant perfusion parameters confirmed valid ESLP with homogeneous lung perfusion, evidenced by uniform lung surface temperature. BGA showed temperature-dependent metabolic activities with differences depending on perfusion solution composition. COP is not decisive for pulmonary oedema and associated weight gain, but possibly rather observed chemokine profile and dextran sensitivity of rats. Histological examination confirmed intact lung architecture without infarcts or hemorrhages due to optimal organ procurement and single-lung application protocol using our in-house-designed ESLP system.
Assuntos
Pulmão , Perfusão , Ratos Endogâmicos F344 , Ratos Endogâmicos Lew , Animais , Ratos , Perfusão/métodos , Pulmão/fisiologia , Preservação de Órgãos/métodos , Transplante de Pulmão/métodos , Modelos Animais , Masculino , Experimentação AnimalRESUMO
PURPOSE: This study aimed to introduce and evaluate two new microvascular anastomosis techniques compared to the conventional method in a rat renal transplant model. METHODS: Using a Fisher-to-Lewis rat kidney transplantation model, the renal artery anastomosis was performed using the interrupted (I) suture technique, Y-shaped continuous (Y) suture technique, and anterior-interrupted and posterior-continuous (I-C) suture technique. The rats were then divided into three groups: I group, Y group, and I-C group. Parameters such as arterial anastomosis time, warm ischemia time, seven-day survival rate of the rats, and vessel histopathology were assessed. RESULTS: The mean arterial anastomosis time, blood leakage scores, and warm ischemia time were significantly reduced in groups Y and I-C compared to group I. Moreover, the seven-day survival rate was significantly higher in the I-C group compared to the other two groups. Arterial histopathology demonstrated vessel wall recovery without damage in all three groups, suggesting the safety of both Y and I-C techniques. CONCLUSIONS: The anterior-interrupted and posterior-continuous suture method is particularly beneficial for small artery reconstruction in organ transplantation.
Assuntos
Anastomose Cirúrgica , Transplante de Rim , Ratos Endogâmicos Lew , Artéria Renal , Técnicas de Sutura , Animais , Transplante de Rim/métodos , Anastomose Cirúrgica/métodos , Masculino , Artéria Renal/cirurgia , Ratos Endogâmicos F344 , Fatores de Tempo , Ratos , Modelos Animais , Isquemia Quente/métodos , Reprodutibilidade dos Testes , Sobrevivência de EnxertoRESUMO
BACKGROUND: Ischemia-reperfusion injury (IRI) is a phenomenon that affects transplant survival. The aim of our study was to examine the effects of IRI in isogenic and allogeneic muscle and skin transplantation models exposed to prolonged warm ischemia. METHODS: Forty-eight Lewis rats and 16 Brown-Norway rats were used to create four groups: Isogenic Inguinal Flap Transplantation (IST), Isogenic Gastrocnemius Muscle Flap Transplantation (IMT), Allogeneic Inguinal Flap Transplantation (AST), and Allogeneic Gastrocnemius Muscle Flap Transplantation (AMT). Malonyldialdehyde (MDA) and superoxide dismutase (SOD) levels were measured on postoperative days 1, 7, 21, 35, 63, 100, and 120 in all groups. Donor-specific chimerism (DSC) in peripheral blood was evaluated in the allogeneic groups on postoperative days 7, 21, 35, 63, 100, and 120. The microRNA-21 and microRNA-205 levels were evaluated on postoperative days 1, 7, and 120 in all groups. At the end of the study, a histopathological examination was performed. RESULTS: A statistically significant difference was found between the groups in terms of MDA and SOD levels. DSC was detected in the AMT group. A significant increase in microRNA-205 was observed, especially in the AMT group. There was no significant difference in the number of functional muscle units between the muscle transplantation groups. CONCLUSION: The presence of DSC in the AMT group and the lack of a significant difference in the number of functional muscle units in the IMT and AMT groups are noteworthy findings.
Assuntos
Ratos Endogâmicos Lew , Traumatismo por Reperfusão , Transplante de Pele , Animais , Traumatismo por Reperfusão/patologia , Ratos , Masculino , Músculo Esquelético/irrigação sanguínea , Retalhos Cirúrgicos/irrigação sanguínea , Modelos Animais de Doenças , Transplante Homólogo , Ratos Endogâmicos BNRESUMO
Application of polyethylene glycol (PEG) to a peripheral nerve injury at the time of primary neurorrhaphy is thought to prevent Wallerian degeneration via direct axolemma fusion. The molecular mechanisms of nerve fusion and recovery are unclear. Our study tested the hypothesis that PEG alters gene expression in neural and muscular environments as part of its restorative properties. Lewis rats underwent unilateral sciatic nerve transection with immediate primary repair. Subjects were randomly assigned to receive either PEG treatment or standard repair at the time of neurorrhaphy. Samples of sciatic nerve distal to the injury and tibialis muscle at the site of innervation were harvested at 24 hours and 4 weeks postoperatively. Total RNA sequencing and subsequent bioinformatics analyses were used to identify significant differences in differentially expressed genes (DEGs) and their related biological pathways (p<0.05) in PEG-treated subjects compared to non-PEG controls. No significant DEGs were identified in PEG-treated sciatic nerve compared to controls after 24 hours, but 1,480 DEGs were identified in PEG-treated tibialis compared to controls. At 4 weeks, 918 DEGs were identified in PEG-treated sciatic nerve, whereas only 3 DEGs remained in PEG-treated tibialis compared to controls. DEGs in sciatic were mostly upregulated (79%) and enriched in pathways present during nervous system development and growth, whereas DEGs in muscle were mostly downregulated (77%) and related to inflammation and tissue repair. Our findings indicate that PEG application during primary neurorrhaphy leads to significant differential gene regulation in the neural and muscular environment that is associated with improved functional recovery in animals treated with PEG compared to sham non-PEG controls. A detailed understanding of key molecules underlying PEG function in recovery after peripheral nerve repair may facilitate amplification of PEG effects through systemic or focal treatments at the time of neurotmesis.
Assuntos
Músculo Esquelético , Traumatismos dos Nervos Periféricos , Polietilenoglicóis , Ratos Endogâmicos Lew , Nervo Isquiático , Animais , Ratos , Nervo Isquiático/lesões , Traumatismos dos Nervos Periféricos/genética , Polietilenoglicóis/farmacologia , Músculo Esquelético/metabolismo , Músculo Esquelético/inervação , Músculo Esquelético/efeitos dos fármacos , Modelos Animais de Doenças , Análise de Sequência de RNA , Regeneração Nervosa/efeitos dos fármacos , Regeneração Nervosa/genética , Masculino , Regulação da Expressão Gênica/efeitos dos fármacos , Perfilação da Expressão GênicaRESUMO
BACKGROUND: Rheumatic heart disease (RHD) is an autoimmune disease caused by recurrent infections of Group A streptococcus (GAS), ultimately leading to inflammation and the fibrosis of heart valves. Recent studies have highlighted the crucial role of C-C chemokine receptor type 2-positive (CCR2+) macrophages in autoimmune diseases and tissue fibrosis. However, the specific involvement of CCR2+ macrophages in RHD remains unclear. METHODS: This study established an RHD rat model using inactivated GAS and complete Freund's adjuvant, demonstrating a correlation between CCR2+ macrophages and fibrosis in the mitral valves of these rats. RESULTS: Intraperitoneal injection of the CCR2 antagonist Rs-504393 significantly reduced macrophage infiltration, inflammation, and fibrosis in valve tissues of RHD rats compared to the solvent-treated group . Existing evidence suggests that C-C motif chemokine ligand 2 (CCL2) acts as the primary recruiting factor for CCR2+ cells. To validate this, human monocytic leukemia cells (THP-1) were cultured in vitro to assess the impact of recombinant CCL2 protein on macrophages. CCL2 exhibited pro-inflammatory effects similar to lipopolysaccharide (LPS), promoting M1 polarization in macrophages. Moreover, the combined effect of LPS and CCL2 was more potent than either alone. Knocking down CCR2 expression in THP-1 cells using small interfering RNA suppressed the pro-inflammatory response and M1 polarization induced by CCL2. CONCLUSIONS: The findings from this study indicate that CCR2+ macrophages are pivotal in the valvular remodeling process of RHD. Targeting the CCL2/CCR2 signaling pathway may therefore represent a promising therapeutic strategy to alleviate valve fibrosis in RHD.
Assuntos
Inflamação , Macrófagos , Receptores CCR2 , Cardiopatia Reumática , Animais , Humanos , Masculino , Ratos , Quimiocina CCL2/metabolismo , Quimiocina CCL2/genética , Modelos Animais de Doenças , Ácido Eicosapentaenoico/análogos & derivados , Fibrose , Valvas Cardíacas/patologia , Inflamação/metabolismo , Macrófagos/metabolismo , Macrófagos/imunologia , Ratos Endogâmicos Lew , Receptores CCR2/metabolismo , Receptores CCR2/genética , Cardiopatia Reumática/imunologia , Cardiopatia Reumática/microbiologia , Cardiopatia Reumática/metabolismo , Cardiopatia Reumática/patologia , Infecções Estreptocócicas/imunologia , Infecções Estreptocócicas/microbiologia , Infecções Estreptocócicas/metabolismo , Streptococcus pyogenes , Células THP-1RESUMO
BACKGROUND: Normothermic ex vivo liver perfusion (NEVLP) is an exciting strategy to preserve livers prior to transplant, however, the effects of NEVLP on the phenotype of tissue-resident immune cells is largely unknown. The presence of tissue-resident memory T cells (TRM) in the liver may protect against acute rejection and decrease allograft dysfunction. Therefore, we investigated the effects of NEVLP on liver TRMs and assessed the ability of anti-inflammatory cytokines to reduce TRM activation during NEVLP. METHODS: Rat livers underwent NEVLP with or without the addition of IL-10 and TGF-ß. Naïve and cold storage livers served as controls. Following preservation, TRM T cell gene expression profiles were assessed through single cell RNA sequencing (scRNA-seq). Differential gene expression analysis was performed with Wilcoxon rank sum test to identify differentially expressed genes (DEGs) associated with a specific treatment group. Using the online Database for Annotation, Visualization and Integrated Discovery (DAVID), gene set enrichment was then conducted with Fisher's exact test on DEGs to highlight differentially regulated pathways and functional terms associated with treatment groups. RESULTS: Through scRNA-seq analysis, an atlas of liver-resident memory T cell subsets was created for all livers. TRM T cells could be identified in all livers, and through scRNA-seq, DEG was identified with Wilcoxon rank sum test at FDR < 0.05. Based on the gene set enrichment analysis of DEGs using Fisher's exact test, NEVLP is associated with downregulation of multiple gene enrichment pathways associated with surface proteins. Furthermore, NEVLP with anti-inflammatory cytokines was associated with down regulation of 52 genes in TRM T cells when compared to NEVLP alone (FDR <0.05), most of which are pro-inflammatory. CONCLUSION: This is the first study to create an atlas of liver TRM T cells in the rat liver undergoing NEVLP and demonstrate the effects of NEVLP on liver TRM T cells at the single cell gene expression level.
Assuntos
Transplante de Fígado , Fígado , Perfusão , Análise de Sequência de RNA , Análise de Célula Única , Animais , Ratos , Fígado/imunologia , Fígado/metabolismo , Masculino , Preservação de Órgãos/métodos , Ratos Endogâmicos Lew , Células T de Memória/imunologia , Células T de Memória/metabolismo , Rejeição de Enxerto/imunologia , Rejeição de Enxerto/genéticaRESUMO
Conventional rodent neuromyelitis optica spectrum disorder (NMOSD) models using patient-derived immunoglobulin G (IgG) are potentially affected by the differences between the human and rodent aquaporin-4 (AQP4) extracellular domains (ECDs). We hypothesized that the humanization of AQP4 ECDs would make the rodent model lesions closer to human NMOSD pathology. Humanized-AQP4-expressing (hAQP4) rats were generated using genome-editing technology, and the human AQP4-specific monoclonal antibody (mAb) or six patient-derived IgGs were introduced intraperitoneally into hAQP4 rats and wild-type Lewis (WT) rats after immunization with myelin basic protein and complete Freund's adjuvant. Human AQP4-specific mAb induced astrocyte loss lesions specifically in hAQP4 rats. The patient-derived IgGs also induced NMOSD-like tissue-destructive lesions with AQP4 loss, demyelination, axonal swelling, complement deposition, and marked neutrophil and macrophage/microglia infiltration in hAQP4 rats; however, the difference in AQP4 loss lesion size and infiltrating cells was not significant between hAQP4 and WT rats. The patient-derived IgGs bound to both human and rat AQP4 M23, suggesting their binding to the shared region of human and rat AQP4 ECDs. Anti-AQP4 titers positively correlated with AQP4 loss lesion size and neutrophil and macrophage/microglia infiltration. Considering that patient-derived IgGs vary in binding sites and affinities and some of them may not bind to rodent AQP4, our hAQP4 rat is expected to reproduce NMOSD-like pathology more accurately than WT rats.
Assuntos
Aquaporina 4 , Modelos Animais de Doenças , Edição de Genes , Imunoglobulina G , Neuromielite Óptica , Ratos Endogâmicos Lew , Animais , Aquaporina 4/genética , Aquaporina 4/metabolismo , Neuromielite Óptica/genética , Neuromielite Óptica/patologia , Neuromielite Óptica/metabolismo , Ratos , Humanos , Anticorpos Monoclonais , Feminino , Astrócitos/metabolismo , Astrócitos/patologia , Ratos TransgênicosRESUMO
Recently, therapies utilizing extracellular vesicles (EVs) derived from mesenchymal stromal/stem cells (MSCs) have begun to show promise in clinical trials. However, EV therapeutic potential varies with MSC tissue source and in vitro expansion through passaging. To find the optimal MSC source for clinically translatable EV-derived therapies, this study aims to compare the angiogenic and immunomodulatory potentials and the protein and miRNA cargo compositions of EVs isolated from the two most common clinical sources of adult MSCs, bone marrow and adipose tissue, across different passage numbers. Primary bone marrow-derived MSCs (BMSCs) and adipose-derived MSCs (ASCs) were isolated from adult female Lewis rats and expanded in vitro to the indicated passage numbers (P2, P4, and P8). EVs were isolated from the culture medium of P2, P4, and P8 BMSCs and ASCs and characterized for EV size, number, surface markers, protein content, and morphology. EVs isolated from different tissue sources showed different EV yields per cell, EV sizes, and protein yield per EV. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses of proteomics data and miRNA seq data identified key proteins and pathways associated with differences between BMSC-EVs and ASC-EVs, as well as differences due to passage number. In vitro tube formation assays employing human umbilical vein endothelial cells suggested that both tissue source and passage number had significant effects on the angiogenic capacity of EVs. With or without lipopolysaccharide (LPS) stimulation, EVs more significantly impacted expression of M2-macrophage genes (IL-10, Arg1, TGFß) than M1-macrophage genes (IL-6, NOS2, TNFα). By correlating the proteomics analyses with the miRNA seq analysis and differences observed in our in vitro immunomodulatory, angiogenic, and proliferation assays, this study highlights the trade-offs that may be necessary in selecting the optimal MSC source for development of clinical EV therapies.
Assuntos
Vesículas Extracelulares , Células-Tronco Mesenquimais , MicroRNAs , Ratos Endogâmicos Lew , Vesículas Extracelulares/metabolismo , Células-Tronco Mesenquimais/metabolismo , MicroRNAs/metabolismo , MicroRNAs/genética , Animais , Feminino , Ratos , Tecido Adiposo/metabolismo , Tecido Adiposo/citologia , Neovascularização Fisiológica , Imunomodulação , Humanos , Células Cultivadas , Proliferação de Células , Células da Medula Óssea/metabolismoRESUMO
BACKGROUND: Dihydroartemisinin (DHA), a derivative and active metabolite of artemisinin, possesses various immunomodulatory properties. However, its role in myasthenia gravis (MG) has not been clearly explored. Here, we investigated the role of DHA in experimental autoimmune myasthenia gravis (EAMG) and its potential mechanisms. METHODS: The AChR97-116 peptide-induced EAMG model was established in Lewis rats and treated with DHA. Flow cytometry was used to assess the release of Th cell subsets and Treg cells, and 16S rRNA gene amplicon sequence analysis was applied to explore the relationship between the changes in the intestinal flora after DHA treatment. In addition, network pharmacology and molecular docking were utilized to explore the potential mechanism of DHA against EAMG, which was further validated in the rat model by immunohistochemical and RT-qPCR for further validation. RESULTS: In this study, we demonstrate that oral administration of DHA ameliorated clinical symptoms in rat models of EAMG, decreased the expression level of Th1 and Th17 cells, and increased the expression level of Treg cells. In addition, 16S rRNA gene amplicon sequence analysis showed that DHA restored gut microbiota dysbiosis in EAMG rats by decreasing Ruminococcus abundance and increasing the abundance of Clostridium, Bifidobacterium, and Allobaculum. Using network pharmacology, 103 potential targets of DHA related to MG were identified, and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that PI3K-AKT signaling pathway was related to the treatment of DHA on EAMG. Meanwhile, molecular docking verified that DHA has good binding affinity to AKT1, CASP3, EGFR, and IGF1. Immunohistochemical staining showed that DHA treatment significantly inhibited the phosphorylated expression of AKT and PI3K in the spleen tissues of EAMG rats. In EAMG rats, RT-qPCR results also showed that DHA reduced the mRNA expression levels of PI3K and AKT1. CONCLUSIONS: DHA ameliorated EAMG by inhibiting the PI3K-AKT signaling pathway, regulating CD4+ T cells and modulating gut microbiota, providing a novel therapeutic approach for the treatment of MG.
Assuntos
Artemisininas , Microbioma Gastrointestinal , Simulação de Acoplamento Molecular , Miastenia Gravis Autoimune Experimental , Ratos Endogâmicos Lew , Animais , Microbioma Gastrointestinal/efeitos dos fármacos , Artemisininas/farmacologia , Artemisininas/uso terapêutico , Ratos , Miastenia Gravis Autoimune Experimental/tratamento farmacológico , Miastenia Gravis Autoimune Experimental/imunologia , Feminino , Disbiose/tratamento farmacológico , Disbiose/imunologia , Transdução de Sinais/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/efeitos dos fármacos , Fosfatidilinositol 3-Quinases/metabolismoRESUMO
OBJECTIVES: For nerve injuries, not amendable to tensionless epineural coaptation of the nerve, autografts are the preferred treatment. Although absorbable sutures are not recommended for nerve repair, there is no evidence that non-absorbable sutures are superior to absorbable sutures. This study aims to assess the effectiveness of non-absorbable monofilament nylon sutures, absorbable monofilament vicryl sutures, and fibrin glue when used for nerve grafting. METHODS: Lewis rats (N = 32) were subjected to a sciatic nerve transection and randomly assigned to a group: graft with Nylon, graft with Vicryl, graft with Fibrin Glue, or no graft. Motor function, sensory function, and thermal pain were assessed during a 12-week recovery period, and immunohistochemistry was used to assess macrophage response. RESULTS: At 12 weeks, the Vicryl and Nylon groups had significantly larger ankle angles at to lift off, which is a measure of motor function, compared to injured controls (p < 0.05). Grafted rats displayed no difference in thermal response but hypersensitivity to mechanical stimuli compared to the uninjured hindlimb. The Nylon, Vicryl, and Fibrin Glue groups all had significantly less atrophy of the gastrocnemius muscle compared to injured controls (p < 0.0001). In the Fibrin Glue group, 3/9 grafts did not incorporate. The Nylon group had significantly less (p = 0.0004) axon growth surrounding the suture holes compared to the Vicryl group. There were no differences in the axon counts, motor neurons, or sensory neurons between all grafted rats. CONCLUSIONS: These results demonstrate that vicryl sutures work just as well as nylon for nerve recovery after injury and grafting.
Assuntos
Adesivo Tecidual de Fibrina , Nylons , Poliglactina 910 , Ratos Endogâmicos Lew , Animais , Adesivo Tecidual de Fibrina/farmacologia , Ratos , Nervo Isquiático/lesões , Regeneração Nervosa/fisiologia , Regeneração Nervosa/efeitos dos fármacos , Modelos Animais de Doenças , Suturas , Adesivos Teciduais/farmacologia , Recuperação de Função Fisiológica/fisiologia , Masculino , FemininoRESUMO
Introduction: Maternal synbiotic supplementation during pregnancy and lactation can significantly influence the immune system. Prebiotics and probiotics have a positive impact on the immune system by preventing or ameliorating among others intestinal disorders. This study focused on the immunomodulatory effects of B. breve M-16V and short chain galacto-oligosaccharides (scGOS)/long chain fructo-oligosachairdes (lcFOS), including systemic and mucosal compartments and milk composition. Methods: Lewis rats were orally administered with the synbiotic or vehicle during pregnancy (21 days) and lactation (21 days). At the weaning day, small intestine (SI), mammary gland (MG), adipose tissue, milk, mesenteric lymph nodes (MLN), salivary gland (SG), feces and cecal content were collected from the mothers. Results: The immunoglobulinome profile showed increased IgG2c in plasma and milk, as well as elevated sIgA in feces at weaning. The supplementation improved lipid metabolism through enhanced brown adipose tissue activity and reinforced the intestinal barrier by increasing the expression of Muc3, Cldn4, and Ocln. The higher production of short chain fatty acids in the cecum and increased Bifidobacterium counts suggest a potential positive impact on the gastrointestinal tract. Discussion: These findings indicate that maternal synbiotic supplementation during gestation and lactation improves their immunological status and improved milk composition.
Assuntos
Bifidobacterium breve , Lactação , Leite , Oligossacarídeos , Animais , Feminino , Gravidez , Bifidobacterium breve/imunologia , Leite/imunologia , Leite/química , Ratos , Ratos Endogâmicos Lew , Suplementos Nutricionais , Simbióticos/administração & dosagem , Probióticos/administração & dosagem , Probióticos/farmacologiaRESUMO
Despite significant efforts toward improving therapy for septic shock, mortality remains high. Applying veno-arterial (V-A) extracorporeal membrane oxygenation (ECMO) in this context remains controversial. Since the cannulation of the femoral artery for V-A ECMO return leads to lower body hyperoxia, this study investigated the impact of V-A ECMO therapy on the intestinal and hepatic microcirculation during septic shock in a rodent model. Thirty male Lewis rats were randomly assigned to receive V-A ECMO therapy with low (60 mL/kg/min) or high (90 mL/kg/min) blood flow or a sham procedure. Hemodynamic data were collected through a pressure-volume catheter in the left ventricle and a catheter in the lateral tail artery. Septic shock was induced by intravenous administration of lipopolysaccharide (1 mg/kg). The rats received lung-protective ventilation during V-A ECMO therapy. The hepatic and intestinal microcirculation was measured by micro-lightguide spectrophotometry after median laparotomy for two hours. Systemic and pulmonary inflammation was detected via enzyme-linked immunosorbent assays (ELISA) of the plasma and bronchoalveolar lavage (BAL), respectively, measuring tumor necrosis factor-alpha (TNF-α), interleukins 6 (IL-6) and 10 (IL-10), and C-X-C motif ligands 2 (CXCL2) and 5 (CXCL5). Oxygen saturation and relative hemoglobin concentration were reduced in the hepatic and intestinal microcirculation during V-A ECMO therapy, independent of the blood flow rate. Further, rats treated with V-A ECMO therapy also presented elevated systolic, diastolic, and mean arterial blood pressure and increased stroke volume, cardiac output, and left ventricular end-diastolic volume. However, left ventricular end-diastolic pressure was only elevated during high-flow V-A ECMO therapy. Blood gas analysis revealed a dilutional anemia during V-A ECMO therapy. ELISA analysis showed an elevated plasma CXCL2 concentration only during high-flow V-A ECMO therapy and elevated BAL CXCL2 and CXCL5 concentrations only during low-flow V-A ECMO therapy. Rats undergoing V-A ECMO therapy exhibited impaired microcirculation of the intestine and liver during septic shock despite increased blood pressure and cardiac output. Increased pulmonary inflammation was detected only during low-flow V-A ECMO therapy in septic shock.
Assuntos
Modelos Animais de Doenças , Oxigenação por Membrana Extracorpórea , Intestinos , Fígado , Microcirculação , Ratos Endogâmicos Lew , Choque Séptico , Animais , Oxigenação por Membrana Extracorpórea/métodos , Masculino , Ratos , Choque Séptico/terapia , Choque Séptico/fisiopatologia , Choque Séptico/metabolismo , Fígado/metabolismo , Fígado/irrigação sanguínea , Intestinos/irrigação sanguínea , Pneumonia/terapia , Pneumonia/metabolismo , Pneumonia/fisiopatologia , Hemodinâmica , Fator de Necrose Tumoral alfa/metabolismo , Fator de Necrose Tumoral alfa/sangueRESUMO
Coronary artery bypass surgery can result in endothelial dysfunction due to ischemia/reperfusion (IR) injury. Previous studies have demonstrated that DuraGraft helps maintain endothelial integrity of saphenous vein grafts during ischemic conditions. In this study, we investigated the potential of DuraGraft to mitigate endothelial dysfunction in arterial grafts after IR injury using an aortic transplantation model. Lewis rats (n = 7-9/group) were divided in three groups. Aortic arches from the control group were prepared and rings were immediately placed in organ baths, while the aortic arches of IR and IR + DuraGraft rats were preserved in saline or DuraGraft, respectively, for 1 h before being transplanted heterotopically. After 1 h after reperfusion, the grafts were explanted, rings were prepared, and mounted in organ baths. Our results demonstrated that the maximum endothelium-dependent vasorelaxation to acetylcholine was significantly impaired in the IR group compared to the control group, but DuraGraft improved it (control: 89 ± 2%; IR: 24 ± 1%; IR + DuraGraft: 48 ± 1%, p < 0.05). Immunohistochemical analysis revealed decreased intercellular adhesion molecule-1, 4-hydroxy-2-nonenal, caspase-3 and caspase-8 expression, while endothelial cell adhesion molecule-1 immunoreactivity was increased in the IR + DuraGraft grafts compared to the IR-group. DuraGraft mitigates endothelial dysfunction following IR injury in a rat bypass model. Its protective effect may be attributed, at least in part, to its ability to reduce the inflammatory response, oxidative stress, and apoptosis.
Assuntos
Endotélio Vascular , Ratos Endogâmicos Lew , Traumatismo por Reperfusão , Animais , Ratos , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Traumatismo por Reperfusão/metabolismo , Masculino , Ponte de Artéria Coronária/métodos , Ponte de Artéria Coronária/efeitos adversos , Estresse Oxidativo/efeitos dos fármacos , Molécula 1 de Adesão Intercelular/metabolismo , Modelos Animais de Doenças , Aldeídos/metabolismo , Aldeídos/farmacologia , Caspase 3/metabolismo , Vasodilatação/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Acetilcolina/farmacologiaRESUMO
Cement mediated peri-implantitis accounts for 1.9-75% of dental implant failures associated with peri-implant diseases. This study evaluated the biological impact of dental cements on osseointegrated implants using Lewis rats. Twenty-two rats were distributed into 6 groups: negative control (NC) soft diet (SD), and hard diet (HD); positive control SD and HD (n = 3); Implant + bio-ceramic Cement (BC) SD and HD which included contralateral Sham sites (n = 5). Titanium implants were placed on either side of the maxillae and allowed to heal for 14 days. Later, both sides of experimental groups underwent a re-entry surgery to simulate clinical cementation. The right side received 0.60 mg of BC. At 14 days post cement application, maxillae were harvested for clinical, microtomographic, and histological evaluations. Clinical and microtomographic evaluations indicated evidence of extensive inflammation and circumferential bone resorption around BC implants in comparison to NC. Histology revealed cement particles surrounded by inflammatory infiltrate in the implant area accompanied by biofilm for SD groups. Both sides of BC indicated intensive bone resorption accompanied by signs of osteolysis when compared to NC. Cemented groups depicted significantly lower bone to implant contact when compared to NC. In conclusion, residual cement extravasation negatively impacted osseointegrated implants after re-entry surgeries.
Assuntos
Cimentos Dentários , Implantes Dentários , Peri-Implantite , Microtomografia por Raio-X , Animais , Ratos , Implantes Dentários/efeitos adversos , Peri-Implantite/patologia , Peri-Implantite/etiologia , Masculino , Ratos Endogâmicos Lew , Osseointegração , Titânio/efeitos adversos , Modelos Animais de Doenças , Maxila/cirurgiaRESUMO
Segmental grafts from living donors have advantages over grafts from deceased donors when used for small intestine transplantation. However, storage time for small intestine grafts can be extremely short and optimal graft preservation conditions for short-term storage remain undetermined. Secreted factors from mesenchymal stem cells (MSCs) that allow direct activation of preserved small intestine grafts. Freshly excised Luc-Tg LEW rat tissues were incubated in preservation solutions containing MSC-conditioned medium (MSC-CM). Preserved Luc-Tg rat-derived grafts were then transplanted to wild-type recipients, after which survival, injury score, and tight junction protein expression were examined. Luminance for each graft was determined using in vivo imaging. The findings indicated that 30-100 and 3-10 kDa fractions of MSC-CM have superior activating effects for small intestine preservation. Expression of the tight-junction proteins claudin-3, and zonula occludens-1 preserved for 24 h in University of Wisconsin (UW) solution containing MSC-CM with 50-100 kDa, as shown by immunostaining, also indicated effectiveness. Reflecting the improved graft preservation, MSC-CM preloading of grafts increased survival rate from 0% to 87%. This is the first report of successful transplantation of small intestine grafts preserved for more than 24 h using a rodent model to evaluate graft preservation conditions that mimic clinical conditions.
Assuntos
Intestino Delgado , Células-Tronco Mesenquimais , Preservação de Órgãos , Ratos Endogâmicos Lew , Animais , Intestino Delgado/transplante , Ratos , Preservação de Órgãos/métodos , Masculino , Soluções para Preservação de Órgãos , Sobrevivência de Enxerto , Meios de Cultivo Condicionados , Proteína da Zônula de Oclusão-1/metabolismo , Claudina-3/metabolismo , Ratos Transgênicos , Glutationa , Rafinose , Alopurinol , Insulina , AdenosinaRESUMO
Doxorubicin (DOX) is a common and highly effective chemotherapeutic. However, its use is limited by cardiotoxic effects and the lack of methods to detect these at early time points. In the present study, we evaluated if [64Cu]Cu-NODAGA-E[(cRGDyK)]2 positron emission tomography-computed tomography ([64Cu]Cu-RGD PET/CT) could detect cardiotoxicity in a rat model of DOX-induced heart failure. Male Lewis rats were divided into two groups and treated with either a cumulative dose of 15 mg/kg of DOX or left untreated. Cardiac anatomy and function were assessed using magnetic resonance imaging at baseline and in week 8. [64Cu]Cu-RGD PET/CT scans were performed in week 4. DOX treatment led to a decline in pump function as well as an increase in cardiac and thymic uptake of [64Cu]Cu-RGD. In addition, DOX altered cardiac gene expression, led to infiltration of immune cells, reduced endothelial content, and increased interstitial fibrosis. Furthermore, concentrations of inflammatory plasma proteins were increased in the DOX group. In conclusion, DOX treatment resulted in the development of cardiotoxicity and heart failure, which could be detected using [64Cu]Cu-RGD PET/CT at early time points. [64Cu]Cu-RGD uptake in the myocardial septum and thymus predicted a low left ventricular ejection fraction in week 8.
Assuntos
Radioisótopos de Cobre , Modelos Animais de Doenças , Doxorrubicina , Insuficiência Cardíaca , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada , Ratos Endogâmicos Lew , Animais , Doxorrubicina/efeitos adversos , Doxorrubicina/toxicidade , Ratos , Insuficiência Cardíaca/induzido quimicamente , Insuficiência Cardíaca/diagnóstico por imagem , Masculino , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada/métodos , Compostos Heterocíclicos com 1 Anel , Cardiotoxicidade/etiologia , Acetatos/química , Imageamento por Ressonância Magnética/métodos , Tomografia por Emissão de Pósitrons/métodos , Coração/efeitos dos fármacos , Coração/diagnóstico por imagem , Compostos RadiofarmacêuticosRESUMO
BACKGROUND: Full-thickness skin defects often are managed with split-thickness skin grafting. The wound healing process, including formation of new vessels during the healing of skin grafts, is complex. OBJECTIVE: To evaluate the microcirculatory changes in the treated tissue after skin grafting to analyze perfusion dynamics during the wound healing process. MATERIALS AND METHODS: Fourteen full-thickness skin defects were created on the back of 14 adult male Lewis rats. All wounds were treated with autologous split-thickness skin grafts. The perfusion dynamics were assessed for 84 days with an O2C device that combines a laser light to determine blood flow and white light to determine postcapillary SO2 and the rHb. RESULTS: Blood flow increased for 50 days after grafting. SO2 decreased in superficial skin layers (depth of 2 mm) and increased in deep skin layers (depth of 8 mm) during the entire observation period. The rHb increased until day 10 in superficial layers and until day 20 in deep tissue layers. CONCLUSION: The microcirculatory changes reflect the different phases of wound healing. Long after the skin transplants were macroscopically healed, alterations in microcirculation were still detected. These alterations were caused by the long-lasting changes in tissue metabolism due to the formation, conversion, and degradation of the dermal matrix and vessels during wound healing and scar formation.
Assuntos
Microcirculação , Ratos Endogâmicos Lew , Transplante de Pele , Pele , Cicatrização , Animais , Cicatrização/fisiologia , Microcirculação/fisiologia , Transplante de Pele/métodos , Ratos , Masculino , Pele/irrigação sanguínea , Modelos Animais de DoençasRESUMO
Periodontitis is a chronic inflammatory disease that affects the periodontal tissues. Although it is associated with various systemic diseases, the impact of periodontitis on kidney transplantation (KT) outcomes, particularly allograft rejection, remains unclear. This study investigated the effect of periodontitis on transplant immunity, specifically examining Porphyromonas gingivalis-derived lipopolysaccharide (LPS-PG). In vitro experiments revealed that LPS-PG increased regulatory T cells (Tregs) in Lewis rat spleen cells. In a mixed lymphocyte reaction assay, concentrations of interferon-γ, indicative of alloreactivity, were lower than in controls when LPS-PG was added to the culture and when LPS-PG-administered Lewis rat spleen cells were used as responders. In a rat KT model, LPS-PG administration to recipients promoted mild tubulitis and low serum creatinine and blood urea nitrogen levels 5 days post-KT compared with PBS-administered controls. Furthermore, LPS-PG-administered recipients had an elevated Treg proportion in their peripheral blood and spleen cells, and increased infiltrating Tregs in kidney allografts, compared with controls. The elevated Treg proportion in peripheral blood and spleen cells had a significant negative correlation with serum creatinine, suggesting elevated Tregs modulated allograft rejection. These findings suggest that periodontitis might modulate alloimmune reactivity through LPS-PG and Tregs, offering insights to refine immunosuppressive strategies for KT recipients.
Assuntos
Rejeição de Enxerto , Transplante de Rim , Lipopolissacarídeos , Porphyromonas gingivalis , Ratos Endogâmicos Lew , Linfócitos T Reguladores , Animais , Porphyromonas gingivalis/imunologia , Transplante de Rim/efeitos adversos , Ratos , Linfócitos T Reguladores/imunologia , Masculino , Rejeição de Enxerto/imunologia , Aloenxertos , Periodontite/imunologia , Periodontite/microbiologia , Modelos Animais de Doenças , Baço/imunologiaRESUMO
Maternal breast milk plays a key role in providing newborns with passive immunity and stimulating the maturation of an infant's immune system, protecting them from many diseases. It is known that diet can influence the immune system of lactating mothers and the composition of their breast milk. The aim of this study was to establish if a supplementation during the gestation and lactation of Lewis rats with extra virgin olive oil (EVOO), due to the high proportion of antioxidant components in its composition, has an impact on the mother's immune system and on the breast milk's immune composition. For this, 10 mL/kg of either EVOO, refined oil (control oil) or water (REF group) were orally administered once a day to rats during gestation and lactation periods. Immunoglobulin (Ig) concentrations and gene expressions of immune molecules were quantified in several compartments of the mothers. The EVOO group showed higher IgA levels in both the breast milk and the mammary glands than the REF group. In addition, the gene expression of IgA in mammary glands was also boosted by EVOO consumption. Overall, EVOO supplementation during gestation and lactation is safe and does not negatively affect the mother's immune system while improving breast milk immune composition by increasing the presence of IgA, which could be critical for an offspring's immune health.
Assuntos
Lactação , Azeite de Oliva , Ratos Endogâmicos Lew , Animais , Feminino , Gravidez , Ratos , Fenômenos Fisiológicos da Nutrição Materna , Imunoglobulina A/metabolismo , Imunoglobulina A/análise , Sistema Imunitário/efeitos dos fármacos , Suplementos Nutricionais , Glândulas Mamárias Animais/imunologia , Glândulas Mamárias Animais/metabolismo , Leite/química , Leite/imunologia , Leite Humano/química , Leite Humano/imunologiaRESUMO
Volumetric muscle loss (VML) injury causes irreversible deficits in muscle mass and function, often resulting in permanent disability. The current standard of care is physical therapy, but it is limited in mitigating functional deficits. We have previously optimized a rehabilitation technique using electrically stimulated eccentric contraction training (EST) that improved muscle mass, strength, and size in VML-injured rats. A biosponge scaffold composed of extracellular matrix proteins has previously enhanced muscle function postVML. This study aimed to determine whether combining a regenerative therapy (i.e., biosponge) with a novel rehabilitation technique (i.e., EST) could enhance recovery in a rat model of VML. A VML defect was created by removing ~20% of muscle mass from the tibialis anterior muscle in adult male Lewis rats. Experimental groups included VML-injured rats treated with biosponge with EST or biosponge alone (n = 6/group). EST was implemented 2 weeks postinjury at 150 Hz and was continued for 4 weeks. A linear increase in eccentric torque over 4 weeks showed the adaptability of the VML-injured muscle to EST. Combining biosponge with EST improved peak isometric torque by ~52% compared with biosponge treatment alone at 6 weeks postinjury. Application of EST increased MyoD gene expression and the percentage of large (>2000 µm2) type 2B myofibers but reduced fibrotic tissue deposition in VML-injured muscles. Together, these changes may provide the basis for improved torque production. This study demonstrates the potential for combined regenerative and rehabilitative therapy to improve muscle recovery following VML.