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1.
Anticancer Res ; 41(9): 4313-4319, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34475051

RESUMO

BACKGROUND/AIM: In order to produce an animal model for oral mucositis induced by anticancer drugs, it is necessary to maintain an immunosuppressive state. We determined the optimal dose and frequency of 5-fluorouracil for a model mouse production. In addition, we used this model to investigate the effect of GGsTop® gelation on the therapeutic effect of oral mucositis. MATERIALS AND METHODS: Changes in body weight and white blood cell count were measured to determine the optimal dosing schedule. The therapeutic effect of GGsTop® gel using chitosan was evaluated by observing changes in the ulcer area for three weeks and measuring collagen and glutathione concentrations in oral mucosal tissue. RESULTS: The optimal dose and frequency of 5-fluorouracil were found to be 50 mg/kg every four days. It was revealed that the therapeutic effect of GGsTop® was enhanced by gelation. CONCLUSION: GGsTop® gel is suggested to be a promising formulation for the treatment of oral mucositis.


Assuntos
Aminobutiratos/administração & dosagem , Antimetabólitos Antineoplásicos/efeitos adversos , Fluoruracila/efeitos adversos , Leucócitos/efeitos dos fármacos , Organofosfonatos/administração & dosagem , Estomatite/tratamento farmacológico , Aminobutiratos/farmacologia , Animais , Antimetabólitos Antineoplásicos/administração & dosagem , Peso Corporal/efeitos dos fármacos , Colágeno/metabolismo , Modelos Animais de Doenças , Cálculos da Dosagem de Medicamento , Fluoruracila/administração & dosagem , Géis , Glutationa/metabolismo , Masculino , Camundongos , Organofosfonatos/farmacologia , Estomatite/induzido quimicamente , Estomatite/metabolismo
2.
Int J Mol Sci ; 22(16)2021 Aug 17.
Artigo em Inglês | MEDLINE | ID: mdl-34445538

RESUMO

Decellularized tissues are biocompatible materials that engraft well, but the age of their source has not been explored for clinical translation. Advanced glycation end products (AGEs) are chemical cross-links that accrue on skeletal muscle collagen in old age, stiffening the matrix and increasing inflammation. Whether decellularized biomaterials derived from aged muscle would suffer from increased AGE collagen cross-links is unknown. We characterized gastrocnemii of 1-, 2-, and 20-month-old C57BL/6J mice before and after decellularization to determine age-dependent changes to collagen stiffness and AGE cross-linking. Total and soluble collagen was measured to assess if age-dependent increases in collagen and cross-linking persisted in decellularized muscle matrix (DMM). Stiffness of aged DMM was determined using atomic force microscopy. AGE levels and the effect of an AGE cross-link breaker, ALT-711, were tested in DMM samples. Our results show that age-dependent increases in collagen amount, cross-linking, and general stiffness were observed in DMM. Notably, we measured increased AGE-specific cross-links within old muscle, and observed that old DMM retained AGE cross-links using ALT-711 to reduce AGE levels. In conclusion, deleterious age-dependent modifications to collagen are present in DMM from old muscle, implying that age matters when sourcing skeletal muscle extracellular matrix as a biomaterial.


Assuntos
Envelhecimento/metabolismo , Colágeno/metabolismo , Matriz Extracelular/metabolismo , Produtos Finais de Glicação Avançada/metabolismo , Músculo Esquelético/metabolismo , Envelhecimento/patologia , Animais , Matriz Extracelular/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Músculo Esquelético/patologia
3.
Int J Mol Sci ; 22(16)2021 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-34445106

RESUMO

Tissue decellularization is typically assessed through absorbance-based DNA quantification after tissue digestion. This method has several disadvantages, namely its destructive nature and inadequacy in experimental situations where tissue is scarce. Here, we present an image processing algorithm for quantitative analysis of DNA content in (de)cellularized tissues as a faster, simpler and more comprehensive alternative. Our method uses local entropy measurements of a phase contrast image to create a mask, which is then applied to corresponding nuclei labelled (UV) images to extract average fluorescence intensities as an estimate of DNA content. The method can be used on native or decellularized tissue to quantify DNA content, thus allowing quantitative assessment of decellularization procedures. We confirm that our new method yields results in line with those obtained using the standard DNA quantification method and that it is successful for both lung and heart tissues. We are also able to accurately obtain a timeline of decreasing DNA content with increased incubation time with a decellularizing agent. Finally, the identified masks can also be applied to additional fluorescence images of immunostained proteins such as collagen or elastin, thus allowing further image-based tissue characterization.


Assuntos
Engenharia Tecidual/métodos , Tecidos Suporte/química , Animais , Colágeno/metabolismo , DNA/metabolismo , Elastina/metabolismo , Matriz Extracelular/metabolismo , Coração/fisiopatologia , Pulmão/metabolismo
4.
Clin Immunol ; 230: 108813, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34333094

RESUMO

Cigarette smoking has been implicated in the pathogenesis of seropositive rheumatoid arthritis (RA), as well as RA-associated lung disease. Fibrotic interstitial lung disease as well as emphysema occur in RA and cause substantial morbidity. We used arthritis-susceptible HLA-DQ8 transgenic mice to generate RA-associated lung disease. Mice were exposed to cigarette smoke (CS) prior to induction of arthritis, and subsequently injected with a low dose of bleomycin intra-tracheally to induce lung injury. Exposure of arthritic mice to both CS and bleomycin led to a significant reduction in lung compliance consistent with development of diffuse lung disease. Morphologic evaluation of the lung demonstrated areas of emphysematous change and co-existent fibrosis, consistent with a combined pattern of fibrosis and emphysema. These changes were accompanied by inflammatory cell infiltration and upregulation of fibrosis-associated genes. This humanized mouse model can serve as a valuable research tool to understand the pathogenesis of RA associated lung disease.


Assuntos
Artrite Reumatoide/complicações , Doenças Pulmonares Intersticiais/etiologia , Animais , Artrite Reumatoide/etiologia , Bleomicina/toxicidade , Fumar Cigarros/efeitos adversos , Colágeno/metabolismo , Modelos Animais de Doenças , Feminino , Expressão Gênica , Humanos , Pulmão/metabolismo , Pulmão/patologia , Complacência Pulmonar/efeitos dos fármacos , Doenças Pulmonares Intersticiais/patologia , Masculino , Camundongos , Camundongos Transgênicos , Enfisema Pulmonar/etiologia , Fibrose Pulmonar/etiologia
5.
FASEB J ; 35(9): e21860, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34411340

RESUMO

Desminopathy is the most common intermediate filament disease in humans. The most frequent mutation causing desminopathy in patients is a R350P DES missense mutation. We have developed a rat model with an analogous mutation in R349P Des. To investigate the role of R349P Des in mechanical loading, we stimulated the sciatic nerve of wild-type littermates (WT) (n = 6) and animals carrying the mutation (MUT) (n = 6) causing a lengthening contraction of the dorsi flexor muscles. MUT animals showed signs of ongoing regeneration at baseline as indicated by a higher number of central nuclei (genotype: P < .0001). While stimulation did not impact central nuclei, we found an increased number of IgG positive fibers (membrane damage indicator) after eccentric contractions with both genotypes (stimulation: P < .01). Interestingly, WT animals displayed a more pronounced increase in IgG positive fibers with stimulation compared to MUT (interaction: P < .05). In addition to altered histology, molecular signaling on the protein level differed between WT and MUT. The membrane repair protein dysferlin decreased with eccentric loading in WT but increased in MUT (interaction: P < .05). The autophagic substrate p62 was increased in both genotypes with loading (stimulation: P < .05) but tended to be more elevated in WT (interaction: P = .05). Caspase 3 levels, a central regulator of apoptotic cell death, was increased with stimulation in both genotypes (stimulation: P < .01) but more so in WT animals (interaction: P < .0001). Overall, our data indicate that R349P Des rats have a lower susceptibility to structural muscle damage of the cytoskeleton and sarcolemma with acute eccentric loading.


Assuntos
Desmina/genética , Contração Muscular , Músculo Esquelético/lesões , Músculo Esquelético/metabolismo , Mutação , Doença Aguda , Animais , Apoptose , Doença Crônica , Colágeno/metabolismo , Modelos Animais de Doenças , Estimulação Elétrica , Feminino , Masculino , Fibras Musculares Esqueléticas/metabolismo , Fibras Musculares Esqueléticas/patologia , Músculo Esquelético/patologia , Ratos , Risco
6.
FASEB J ; 35(9): e21862, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34416035

RESUMO

Loss of muscle mass and strength after disuse followed by impaired muscle recovery commonly occurs with aging. Metformin (MET) and leucine (LEU) individually have shown positive effects in skeletal muscle during atrophy conditions but have not been evaluated in combination nor tested as a remedy to enhance muscle recovery following disuse atrophy in aging. The purpose of this study was to determine if a dual treatment of metformin and leucine (MET + LEU) would prevent disuse-induced atrophy and/or promote muscle recovery in aged mice and if these muscle responses correspond to changes in satellite cells and collagen remodeling. Aged mice (22-24 months) underwent 14 days of hindlimb unloading (HU) followed by 7 or 14 days of reloading (7 or 14 days RL). MET, LEU, or MET + LEU was administered via drinking water and were compared to Vehicle (standard drinking water) and ambulatory baseline. We observed that during HU, MET + LEU resolved whole body grip strength and soleus muscle specific force decrements caused by HU. Gastrocnemius satellite cell abundance was increased with MET + LEU treatment but did not alter muscle size during disuse or recovery conditions. Moreover, MET + LEU treatment alleviated gastrocnemius collagen accumulation caused by HU and increased collagen turnover during 7 and 14 days RL driven by a decrease in collagen IV content. Transcriptional pathway analysis revealed that MET + LEU altered muscle hallmark pathways related to inflammation and myogenesis during HU. Together, the dual treatment of MET and LEU was able to increase muscle function, satellite cell content, and reduce collagen accumulation, thus improving muscle quality during disuse and recovery in aging.


Assuntos
Envelhecimento , Colágeno/metabolismo , Leucina/uso terapêutico , Metformina/uso terapêutico , Músculo Esquelético/efeitos dos fármacos , Atrofia Muscular/prevenção & controle , Células Satélites de Músculo Esquelético/efeitos dos fármacos , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Peso Corporal/efeitos dos fármacos , Fibrose/tratamento farmacológico , Elevação dos Membros Posteriores , Imunoglobulina G/análise , Leucina/farmacologia , Masculino , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Metformina/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Desenvolvimento Muscular/efeitos dos fármacos , Fibras Musculares Esqueléticas/efeitos dos fármacos , Força Muscular/efeitos dos fármacos , Músculo Esquelético/citologia , Músculo Esquelético/patologia , Atrofia Muscular/patologia , Tamanho do Órgão/efeitos dos fármacos , RNA-Seq , Células Satélites de Músculo Esquelético/citologia , Células Satélites de Músculo Esquelético/patologia , Transdução de Sinais/efeitos dos fármacos
7.
Int Heart J ; 62(4): 891-899, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34334583

RESUMO

Long-chain noncoding RNA (lncRNA) is a new class of molecular regulators in heart development and disease. However, the role of specific lncRNA in cardiac fibrosis remains to be fully explored. This study aimed to investigate the role and potential mechanism of lncRNA MHRT in myocardial fibrosis after myocardial infarction (MI).Cardiac fibroblasts (CFs) were isolated from a mouse model of MI. The expression levels of MHRT and miR-3185 in the hearts of MI and CFs mice treated with transforming growth factor beta 1 (TGF-ß1) were analyzed by qRT-PCR. The collagen expression was assessed using qRT-PCR and Western blot. Cell proliferation was assessed by performing MTT and EdU assays. The direct interaction between lncRNA and miRNA was analyzed by luciferase assay, RNA-binding protein immunoprecipitation (RIP) assay, and RNA pull-down assay.The expression levels of MHRT were raised in MI and CFs mice treated with TGF-ß1. Overexpression of MHRT promoted collagen production and CF proliferation, while silencing of MHRT showed the opposite effect. MiR-3185 was a target gene of MHRT. In addition, overexpression of MHRT reduced the expression levels of miR-3185, and siMHRT reversed the inhibitory effect of TGF-ß1 on the expression of miR-3185. Overexpression of miR-3185 inhibited the upregulation of Col I and Col III induced by TGF-ß1.MHRT promoted cardiac fibrosis after MI through miR-3185 and increased myocardial collagen deposition and promoted myocardial fibrosis.


Assuntos
Infarto do Miocárdio/metabolismo , Miocárdio/patologia , RNA Longo não Codificante/metabolismo , Animais , Colágeno/metabolismo , Fibroblastos/metabolismo , Fibrose , Masculino , Camundongos Endogâmicos C57BL , Infarto do Miocárdio/patologia , Ratos Sprague-Dawley
8.
Int J Mol Sci ; 22(15)2021 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-34360950

RESUMO

The Bruch's membrane (BrM) is a five-layered extracellular matrix (ECM) that supports the retinal pigment epithelium (RPE). Normal age-related changes in the BrM may lead to RPE cell damage and ultimately to the onset and progression of age-related macular degeneration (AMD), which is the most common cause of visual loss among the elderly. A role for the complement system in AMD pathology has been established, but the disease mechanisms are poorly understood, which hampers the design of efficient therapies to treat millions of patients. In an effort to identify the mechanisms that lead from normal aging to pathology, we have developed a cell-based model using complement deficient human induced pluripotent stem cell (iPSC)-derived RPE cells cultured on an AMD-like ECM that mimics BrM. The data present evidence that changes in the ECM result in loss of differentiation and promote epithelial mesenchymal transition (EMT) of healthy RPE cells. This pathological process is mediated by complement activation and involves the formation of a randomly oriented collagen meshwork that drives the dedifferentiation of the RPE monolayer. Genetic ablation of complement component 3 has a protective effect against EMT but does not prevent the abnormal deposition of collagens. These findings offer new insights into the sequence of events that initiate AMD and may guide the design of efficient therapies to treat this disease with unmet medical needs.


Assuntos
Complemento C3/metabolismo , Transição Epitelial-Mesenquimal , Degeneração Macular/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Linhagem Celular , Colágeno/metabolismo , Ativação do Complemento , Complemento C3/genética , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Matriz Extracelular/metabolismo , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Epitélio Pigmentado da Retina/citologia
9.
Mol Med Rep ; 23(5)2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-34240225

RESUMO

Tracheal stenosis following injury cannot be effectively treated. The current study compared the protective effects of different anti­inflammatory drugs on tracheal stenosis and investigated their possible mechanisms. Rabbit tracheal stenosis models following injury were constructed and confirmed using hematoxylin and eosin (H&E) staining. A total of 30 rabbits were divided into the control (CON), penicillin (PEN), erythromycin (ERY), budesonide (BUD) and PEN + ERY + BUD groups (n=6). Stenotic tracheal tissue, serum and bronchoalveolar lavage fluid (BALF) were collected 10 days after continuous treatment. Pathological changes in the tracheas were observed by H&E staining. Histone deacetylase 2 (HDAC2) expression in tracheal tissues was detected by immunofluorescence. Immunohistochemistry was performed to detect collagen I (Col­I) and collagen III (Col­III) levels in tracheal tissues. Transforming growth factor ß1 (TGF­ß1), vascular endothelial growth factor (VEGF) and interleukin 8 (IL­8) levels in serum and BALF samples were determined using ELISA kits. Western blotting detected HDAC2, IL­8, TGF­ß1 and VEGF levels in tracheal tissues. H&E staining demonstrated that tracheal epithelial hyperplasia and fibroblast proliferation in the ERY and PEN + ERY + BUD groups markedly improved compared with the CON group. Furthermore, in tracheal tissues, HDAC2 expression was significantly increased and IL­8, TGF­ß1, VEGF, Col­I and Col­III levels were significantly decreased in the ERY and PEN + ERY + BUD groups compared with the CON group. Additionally, the results for the PEN + ERY + BUD were more significant compared with the ERY group. In serum and BALF samples, IL­8, TGF­ß1 and VEGF levels in the ERY and PEN + ERY + BUD groups were significantly lower compared with the CON group, with the results of the PEN + ERY + BUD group being more significant compared with the ERY group. There were no significant differences between the PEN, BUD and CON groups. ERY inhibited tracheal granulation tissue proliferation and improved tracheal stenosis following injury and synergistic effects with PEN and BUD further enhanced these protective effects. The mechanism may involve HDAC2 upregulation and inhibition of local airway and systemic inflammatory responses.


Assuntos
Anti-Inflamatórios/uso terapêutico , Budesonida/uso terapêutico , Eritromicina/uso terapêutico , Penicilinas/uso terapêutico , Substâncias Protetoras/uso terapêutico , Estenose Traqueal/metabolismo , Estenose Traqueal/prevenção & controle , Animais , Anti-Inflamatórios/farmacologia , Líquido da Lavagem Broncoalveolar/química , Budesonida/farmacologia , Colágeno/metabolismo , Modelos Animais de Doenças , Eritromicina/farmacologia , Tecido de Granulação/efeitos dos fármacos , Histona Desacetilase 2/genética , Histona Desacetilase 2/metabolismo , Hiperplasia/tratamento farmacológico , Hiperplasia/metabolismo , Interleucina-8/sangue , Interleucina-8/metabolismo , Penicilinas/farmacologia , Substâncias Protetoras/farmacologia , Coelhos , Traqueia/lesões , Traqueia/patologia , Estenose Traqueal/etiologia , Estenose Traqueal/patologia , Fator de Crescimento Transformador beta1/sangue , Fator de Crescimento Transformador beta1/metabolismo , Regulação para Cima/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/sangue , Fator A de Crescimento do Endotélio Vascular/metabolismo
10.
Int J Mol Sci ; 22(14)2021 Jul 13.
Artigo em Inglês | MEDLINE | ID: mdl-34299124

RESUMO

Lung fibrosis has specific computed tomography (CT) findings and represents a common finding in advanced COVID-19 pneumonia whose reversibility has been poorly investigated. The aim of this study was to quantify the extension of collagen deposition and aeration in postmortem cryobiopsies of critically ill COVID-19 patients and to describe the correlations with qualitative and quantitative analyses of lung CT. Postmortem transbronchial cryobiopsy samples were obtained, formalin fixed, paraffin embedded and stained with Sirius red to quantify collagen deposition, defining fibrotic samples as those with collagen deposition above 10%. Lung CT images were analyzed qualitatively with a radiographic score and quantitatively with computer-based analysis at the lobe level. Thirty samples from 10 patients with COVID-19 pneumonia deceased during invasive mechanical ventilation were included in this study. The median [interquartile range] percent collagen extension was 6.8% (4.6-16.2%). In fibrotic compared to nonfibrotic samples, the qualitative score was higher (260 (250-290) vs. 190 (120-270), p = 0.036) while the gas fraction was lower (0.46 (0.32-0.47) vs. 0.59 (0.37-0.68), p = 0.047). A radiographic score above 230 had 100% sensitivity (95% confidence interval, CI: 66.4% to 100%) and 66.7% specificity (95% CI: 41.0% to 92.3%) to detect fibrotic samples, while a gas fraction below 0.57 had 100% sensitivity (95% CI: 66.4% to 100%) and 57.1% specificity (95% CI: 26.3% to 88.0%). In COVID-19 pneumonia, qualitative and quantitative analyses of lung CT images have high sensitivity but moderate to low specificity to detect histopathological fibrosis. Pseudofibrotic CT findings do not always correspond to increased collagen deposition.


Assuntos
COVID-19/complicações , Colágeno/metabolismo , Fibrose Pulmonar/diagnóstico , SARS-CoV-2/isolamento & purificação , Tomografia Computadorizada por Raios X/métodos , Idoso , Autopsia , COVID-19/epidemiologia , COVID-19/virologia , Feminino , Humanos , Itália/epidemiologia , Masculino , Pessoa de Meia-Idade , Fibrose Pulmonar/diagnóstico por imagem , Fibrose Pulmonar/metabolismo , Fibrose Pulmonar/virologia , Estudos Retrospectivos
11.
Int J Mol Sci ; 22(14)2021 Jul 07.
Artigo em Inglês | MEDLINE | ID: mdl-34298935

RESUMO

In vitro evaluation of bone graft materials is generally performed by analyzing the interaction with osteoblasts or osteoblast precursors. In vitro bone models comprising different cell species can give specific first information on the performance of those materials. In the present study, a 3D co-culture model was established comprising primary human osteoblasts, osteoclasts and osteocytes. Osteocytes were differentiated from osteoblasts embedded in collagen gels and were cultivated with osteoblast and osteoclasts seeded in patterns on a porous membrane. This experimental setup allowed paracrine signaling as well as separation of the different cell types for final analysis. After 7 days of co-culture, the three cell species showed their typical morphology and gene expression of typical markers like ALPL, BSPII, BLGAP, E11, PHEX, MEPE, RANKL, ACP5, CAII and CTSK. Furthermore, relevant enzyme activities for osteoblasts (ALP) and osteoclasts (TRAP, CTSK, CAII) were detected. Osteoclasts in triple culture showed downregulated TRAP (ACP5) and CAII expression and decreased TRAP activity. ALP and BSPII expression of osteoblasts in triple culture were upregulated. The expression of the osteocyte marker E11 (PDPN) was unchanged; however, osteocalcin (BGLAP) expression was considerably downregulated both in osteoblasts and osteocytes in triple cultures compared to the respective single cultures.


Assuntos
Osso e Ossos/metabolismo , Osteoblastos/metabolismo , Osteoclastos/metabolismo , Osteócitos/metabolismo , Idoso , Diferenciação Celular/genética , Células Cultivadas , Técnicas de Cocultura , Colágeno/metabolismo , Regulação para Baixo/genética , Feminino , Expressão Gênica/genética , Humanos , Leucócitos Mononucleares/metabolismo , Masculino , Pessoa de Meia-Idade , Regulação para Cima/genética
12.
Toxicology ; 459: 152847, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34245815

RESUMO

Previous findings have confirmed that prenatal nicotine exposure (PNE) leads to retarded cartilage development in the fetal growth plate. It is characterized by insufficient matrix synthesis and decreased expression of matrix phenotype genes aggrecan (ACAN) and Col2A1 in the fetal growth plate chondrocytes; however, the specific molecular mechanism is yet unclear. This study intends to clarify the specific molecular mechanism of fetal osteochondral retardation caused by PNE through animal and cellular experiments. The present study demonstrated that in male offspring of the PNE group (the pregnant rats were subcutaneously administered nicotine 1.0 mg/kg twice per day (2.0 mg/kg.d) at GD11-20), the cartilage matrix of the fetal growth plate was lightly stained, the collagen was reduced, and expression of the matrix phenotype genes, ACAN and Col2A1, was significantly decreased. It was further found that PNE decreased histone acetylation (H3K9/H3K14) levels in the ACAN and Col2A1 promoter regions. Moreover, the expression of Snail and HDAC1/2 was increased in the PNE group. in vitro, the nicotine treatment at different concentrations elevated the expression of Snail/HDAC1/2 while decreasing the H3K9/H3K14 levels in the ACAN and Col2A1 promoter regions. Snail-siRNA transfection partially abolished the nicotine-induced increase in HDAC1/2 expression and decreased the histone acetylation levels in the ACAN and Col2A1 promoter regions. Trichostatin A (TSA) treatment partially reversed the nicotine-induced changes in downstream parameters. In summary, PNE-induced decreased cartilage matrix synthesis in the fetal growth plate of male offspring is effectuated by Snail/HDAC1/2-mediated decreased H3K9/H3K14 levels in the ACAN and Col2A1 promoter regions.


Assuntos
Retardo do Crescimento Fetal/induzido quimicamente , Histona Desacetilase 1/efeitos dos fármacos , Histona Desacetilase 1/metabolismo , Histona Desacetilase 2/efeitos dos fármacos , Histona Desacetilase 2/metabolismo , Nicotina/toxicidade , Agonistas Nicotínicos/toxicidade , Fatores de Transcrição da Família Snail/efeitos dos fármacos , Fatores de Transcrição da Família Snail/metabolismo , Agrecanas/metabolismo , Animais , Cartilagem/efeitos dos fármacos , Cartilagem/patologia , Condrócitos/efeitos dos fármacos , Condrócitos/patologia , Colágeno/metabolismo , Colágeno Tipo II/metabolismo , Feminino , Lâmina de Crescimento/efeitos dos fármacos , Masculino , Gravidez , Efeitos Tardios da Exposição Pré-Natal , Ratos , Ratos Wistar , Transfecção
13.
Cell Prolif ; 54(8): e13087, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34255393

RESUMO

OBJECTIVES: Histatin 1(Hst 1) has been proved to promote wound healing. However, there was no specific study on the regulation made by Hst 1 of fibroblasts in the process of wound healing. This research comprehensively studied the regulation of Hst 1 on the function of fibroblasts in the process of wound healing and preliminary mechanism about it. MATERIALS AND METHODS: The full-thickness skin wound model was made on the back of C57/BL6 mice. The wound healing, collagen deposition and fibroblast distribution were detected on days 3, 5 and 7 after injury. Fibroblast was cultured in vitro and stimulated with Hst 1, and then, their biological characteristics and functions were detected. RESULTS: Histatin 1 can effectively promote wound healing, improve collagen deposition during and after healing and increase the number and function of fibroblasts. After healing, the mechanical properties of the skin also improved. In vitro, the migration ability of fibroblasts stimulated by Hst 1 was significantly improved, and the fibroblasts transformed more into myofibroblasts, which improved the function of contraction and collagen secretion. In fibroblasts, mTOR signalling pathway can be activated by Hst 1. CONCLUSIONS: Histatin 1 can accelerate wound healing and improve the mechanical properties of healed skin by promoting the function of fibroblasts. The intermolecular mechanisms need to be further studied, and this study provides a direction about mTOR signalling pathway.


Assuntos
Histatinas/farmacologia , Cicatrização/efeitos dos fármacos , Animais , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Proliferação de Células , Colágeno/metabolismo , Módulo de Elasticidade , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/transplante , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Transdução de Sinais/efeitos dos fármacos , Pele/patologia , Serina-Treonina Quinases TOR/metabolismo
14.
J Photochem Photobiol B ; 222: 112256, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34330080

RESUMO

Photobiomodulation (PBM) is a promising medical treatment modality in the area of photodynamic therapy (PDT). In this study, we investigated the effect of combined therapy in a 3D microenvironment using aluminum chloride phthalocyanines (AlClPc) as the photosensitizing agent. Normal human fibroblast-containing collagen biomatrix was prepared and treated with an oil-in-water (o/a) AlClPc-loaded nanoemulsion (from 0.5 to 3.0 µM) and irradiated at a range of fluences (from 0.1 to 3.0 J/cm2) using a continuous-wave light-emitting diode (LED) irradiation system (660 nm). PBM at 1.2 J/cm2 and AlClPc/NE at 0.5 µM modified the fibroblast signaling response under 3D conditions, promoting collagen synthesis, ROS production, MMP-9 secretion, proliferation of the actin network, and facile myofibroblastic differentiation. PBM alone (at 1.2 J/cm2 and 0.3 J/cm2) had no significant effect on any of these parameters. The combined therapy affected myofibroblastic differentiation, inflammatory response, and extracellular matrix pliability, and should thus be examined further in subsequent studies considering that no side effects of PBM have been reported. Even though significant progress has been made in the field of phototherapy in recent years, it is necessary to further elucidate the detailed mechanisms underlying its effects already shown in 2D conditions to increase the acceptance of this beneficial and non-invasive therapeutic approach.


Assuntos
Cloreto de Alumínio/farmacologia , Fibroblastos/efeitos dos fármacos , Indóis/farmacologia , Luz , Compostos Organometálicos/farmacologia , Fármacos Fotossensibilizantes/farmacologia , Cloreto de Alumínio/química , Técnicas de Cultura de Células , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Células Cultivadas , Colágeno/metabolismo , Fibroblastos/citologia , Fibroblastos/metabolismo , Fibroblastos/efeitos da radiação , Humanos , Indóis/química , Metaloproteinase 9 da Matriz/metabolismo , Compostos Organometálicos/química , Fármacos Fotossensibilizantes/química , Espécies Reativas de Oxigênio/metabolismo
15.
Nat Chem Biol ; 17(8): 865-871, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34253910

RESUMO

Collagens are fibrous proteins that are integral to the strength and stability of connective tissues. During collagen maturation, lysyl oxidases (LOX) initiate the cross-linking of fibers, but abnormal LOX activity is associated with impaired tissue function as seen in fibrotic and malignant diseases. Visualizing and targeting this dynamic process in healthy and diseased tissue is important, but so far not feasible. Here we present a probe for the simultaneous monitoring and targeting of LOX-mediated collagen cross-linking that combines a LOX-activity sensor with a collagen peptide to chemoselectively target endogenous aldehydes generated by LOX. This synergistic probe becomes covalently anchored and lights up in vivo and in situ in response to LOX at the sites where cross-linking occurs, as demonstrated by staining of normal skin and cancer sections. We anticipate that our reactive collagen-based sensor will improve understanding of collagen remodeling and provide opportunities for the diagnosis of fibrotic and malignant diseases.


Assuntos
Colágeno/metabolismo , Reagentes para Ligações Cruzadas/metabolismo , Peptídeos/metabolismo , Proteína-Lisina 6-Oxidase/metabolismo , Aldeídos/química , Aldeídos/metabolismo , Animais , Colágeno/química , Reagentes para Ligações Cruzadas/química , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Estrutura Molecular , Peptídeos/química , Proteína-Lisina 6-Oxidase/química
16.
Molecules ; 26(12)2021 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-34200590

RESUMO

The aim of the presented research was to obtain reconstituted atelocollagen fibers after extraction from poultry cartilage using the pepsin-acidic method in order to remove telopeptides from the tropocollagen. Firstly, we examined the extraction of collagen from the cartilage extracellular matrix (ECM) after proteoglycans (PG) had been removed by the action of salts, i.e., NaCl or chaotropic MgCl2. Additionally, the effects of the salt type used for PG and hyaluronic acid removal on the properties of self-assembled fibers in solutions at pH 7.4 and freeze-dried matrices were investigated. The basic features of the obtained fibers were characterized, including thermal properties using scanning calorimetry, rheological properties using dynamic oscillatory rheometry, and the structure by scanning electron microscopy. The fibers obtained after PG removal with both analyzed types of salts had similar thermal denaturation characteristics. However, the fibers after PG removal with NaCl, in contrast to those obtained after MgCl2 treatment, showed different rheological properties during gelatinization and smaller diameter size. Moreover, the degree of fibrillogenesis of collagens after NaCl treatment was complete compared to that with MgCl2, which was only partial (70%). The structures of fibers after lyophilization were fundamentally different. The matrices obtained after NaCl pretreatment form regular scaffolds in contrast to the thin, surface structures of the cartilage matrix after proteoglycans removal using MgCl2.


Assuntos
Cartilagem/metabolismo , Galinhas/metabolismo , Colágeno/metabolismo , Proteoglicanas/metabolismo , Cloreto de Sódio/metabolismo , Animais , Matriz Extracelular/metabolismo , Ácido Hialurônico/metabolismo , Pepsina A/metabolismo
17.
Aging (Albany NY) ; 13(13): 17536-17547, 2021 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-34233296

RESUMO

Pathological manifestations in either heart or kidney impact the function of the other and form the basis for the development of cardiorenal syndrome. However, the mechanism or factors involved in such scenario are not completely elucidated. In our study, to find the correlation between late fetal gene expression in diabetic hearts and their influence on diabetic nephropathy, we created a rat model with cardiac specific overexpression of IGF-IIRα, which is an alternative splicing variant of IGFIIR, expressed in pathological hearts. In this study, transgenic rats over expressing cardiac specific IGF-IIRα and non-transgenic animal models established in SD rats were administered with single dose of streptozotocin (STZ, 55 mg/Kg) to induce Type I diabetes. The correlation between IGF-IIRα and kidney damages were further determined based on their intensity of damage in the kidneys. The results show that cardiac specific overexpression of IGF-IIRα elevates the diabetes associated inflammation and morphological changes in the kidneys. The diabetic transgenic rats showed advancement in the pathological features such a renal tubular damage, collagen accumulation and enhancement in STAT3 associated mechanism of renal fibrosis. The results therefore show that that IGF-IIRα expression in the heart during pathological condition may worsen symptoms of diabetic nephropathy in rats.


Assuntos
Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/patologia , Cardiomiopatias Diabéticas/genética , Cardiomiopatias Diabéticas/patologia , Nefropatias Diabéticas/genética , Nefropatias Diabéticas/patologia , Receptor IGF Tipo 2/genética , Animais , Apoptose/genética , Colágeno/metabolismo , Fibrose , Regulação da Expressão Gênica , Rim/patologia , Túbulos Renais/patologia , Masculino , Miocárdio/patologia , Ratos , Ratos Sprague-Dawley , Ratos Transgênicos , Fator de Transcrição STAT3/genética
18.
Int J Mol Sci ; 22(14)2021 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-34299243

RESUMO

(1) Background: The aim of the present study was the biocompatibility analysis of a novel xenogeneic vascular graft material (PAP) based on native collagen won from porcine aorta using the subcutaneous implantation model up to 120 days post implantationem. As a control, an already commercially available collagen-based vessel graft (XenoSure®) based on bovine pericardium was used. Another focus was to analyze the (ultra-) structure and the purification effort. (2) Methods: Established methodologies such as the histological material analysis and the conduct of the subcutaneous implantation model in Wistar rats were applied. Moreover, established methods combining histological, immunohistochemical, and histomorphometrical procedures were applied to analyze the tissue reactions to the vessel graft materials, including the induction of pro- and anti-inflammatory macrophages to test the immune response. (3) Results: The results showed that the PAP implants induced a special cellular infiltration and host tissue integration based on its three different parts based on the different layers of the donor tissue. Thereby, these material parts induced a vascularization pattern that branches to all parts of the graft and altogether a balanced immune tissue reaction in contrast to the control material. (4) Conclusions: PAP implants seemed to be advantageous in many aspects: (i) cellular infiltration and host tissue integration, (ii) vascularization pattern that branches to all parts of the graft, and (iii) balanced immune tissue reaction that can result in less scar tissue and enhanced integrative healing patterns. Moreover, the unique trans-implant vascularization can provide unprecedented anti-infection properties that can avoid material-related bacterial infections.


Assuntos
Prótese Vascular/veterinária , Transplante de Tecidos/métodos , Animais , Aorta/metabolismo , Aorta/transplante , Materiais Biocompatíveis/metabolismo , Bioprótese , Bovinos , Colágeno/metabolismo , Xenoenxertos/metabolismo , Xenoenxertos/fisiologia , Ratos , Ratos Wistar , Suínos/metabolismo , Imunologia de Transplantes/imunologia , Cicatrização/fisiologia
19.
FASEB J ; 35(8): e21779, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34314047

RESUMO

The incredible mechanical strength and durability of mature fibrous tissues and their extremely limited turnover and regenerative capacity underscores the importance of proper matrix assembly during early postnatal growth. In tissues with composite extracellular matrix (ECM) structures, such as the adult knee meniscus, fibrous (Collagen-I rich), and cartilaginous (Collagen-II, proteoglycan-rich) matrix components are regionally segregated to the outer and inner portions of the tissue, respectively. While this spatial variation in composition is appreciated to be functionally important for resisting complex mechanical loads associated with gait, the establishment of these specialized zones is poorly understood. To address this issue, the following study tracked the growth of the murine meniscus from its embryonic formation through its first month of growth, encompassing the critical time-window during which animals begin to ambulate and weight bear. Using histological analysis, region specific high-throughput qPCR, and Col-1, and Col-2 fluorescent reporter mice, we found that matrix and cellular features defining specific tissue zones were already present at birth, before continuous weight-bearing had occurred. These differences in meniscus zones were further refined with postnatal growth and maturation, resulting in specialization of mature tissue regions. Taken together, this work establishes a detailed timeline of the concurrent spatiotemporal changes that occur at both the cellular and matrix level throughout meniscus maturation. The findings of this study provide a framework for investigating the reciprocal feedback between cells and their evolving microenvironments during assembly of a mechanically robust fibrocartilage tissue, thus providing insight into mechanisms of tissue degeneration and effective regenerative strategies.


Assuntos
Cartilagem , Colágeno/metabolismo , Matriz Extracelular/metabolismo , Menisco , Animais , Cartilagem/embriologia , Cartilagem/crescimento & desenvolvimento , Cartilagem/metabolismo , Diferenciação Celular , Proliferação de Células , Menisco/embriologia , Menisco/crescimento & desenvolvimento , Menisco/metabolismo , Camundongos , Camundongos Transgênicos
20.
Biomed Pharmacother ; 138: 111537, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-34311535

RESUMO

Aging of the skin is a complicated bioprocess that is affected by constant exposure to ultraviolet irradiation. The application of herbal-based anti-aging creams is still the best choice for treatment. In the present study, Citrus sinensis L. fruit peels ethanolic extract (CSPE) was formulated into lipid nanoparticles (LNPs) anti-aging cream. Eight different formulations of CSEP-LNPs were prepared and optimized using 23 full factorial designs. In vivo antiaging effect of the best formula was tested in Swiss albino mice where photo-aging was induced by exposure to UV radiation. HPLC-QToF-MS/MS metabolic profiling of CSPE led to the identification of twenty-nine metabolites. CSPE was standardized to a hesperidin content of 15.53 ± 0.152 mg% using RP-HPLC. It was suggested that the optimized formulation (F7) had (245 nm) particle size, (91.065%) EE, and (91.385%) occlusive effect with a spherical and smooth surface. The visible appearance of UV-induced photoaging in mice was significantly improved after topical application on CSPE-NLC cream for 5 weeks, levels of collagen and SOD were significantly increased in CSPE- NLC group, while levels of PGE2, COX2, JNK, MDA, and elastin was reduced. Finally, The prepared anti-aging CSPE-NLC cream represents a safe, convenient, and promising skincare cosmetic product.


Assuntos
Anti-Inflamatórios/farmacologia , Antioxidantes/farmacologia , Citrus sinensis , Metaloproteinase 13 da Matriz/metabolismo , Extratos Vegetais/administração & dosagem , Envelhecimento da Pele/efeitos dos fármacos , Creme para a Pele/administração & dosagem , Pele/efeitos dos fármacos , Administração Cutânea , Animais , Anti-Inflamatórios/química , Anti-Inflamatórios/isolamento & purificação , Antioxidantes/química , Antioxidantes/isolamento & purificação , Citrus sinensis/química , Colágeno/metabolismo , Regulação para Baixo , Composição de Medicamentos , Feminino , Frutas , Lipídeos/química , Metaloproteinase 13 da Matriz/genética , Camundongos , Nanopartículas , Estresse Oxidativo/efeitos dos fármacos , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Pele/enzimologia , Pele/patologia , Pele/efeitos da radiação , Creme para a Pele/química , Creme para a Pele/isolamento & purificação , Superóxido Dismutase/metabolismo , Raios Ultravioleta
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