RESUMO
The bacterial cellulose membrane (CM) is a promising biomaterial due to its easy applicability and moist environment. Moreover, nanoscale silver compounds (AgNO3) are synthesized and incorporated into CMs to provide these biomaterials with antimicrobial activity for wound healing. This study aimed to evaluate the cell viability of CM incorporated with nanoscale silver compounds, determine the minimum inhibitory concentration (MIC) for Escherichia coli and Staphylococcus aureus, and its use on in vivo skin lesions. Wistar rats were divided according to treatment: untreated, CM (cellulose membrane), and AgCM (CM incorporated with silver nanoparticles). The euthanasia was performed on the 2nd, 7th, 14th, and 21st days to assess inflammation (myeloperoxidase-neutrophils, N-acetylglucosaminidase-macrophage, IL-1ß, IL-10), oxidative stress (NO-nitric oxide, DCF-H2O2), oxidative damage (carbonyl: membrane's damage; sulfhydryl: membrane's integrity), antioxidants (superoxide dismutase; glutathione), angiogenesis, tissue formation (collagen, TGF-ß1, smooth muscle α-actin, small decorin, and biglycan proteoglycans). The use of AgCM did not show toxicity, but antibacterial effect in vitro. Moreover, in vivo, AgCM provided balanced oxidative action, modulated the inflammatory profile due to the reduction of IL-1ß level and increase in IL-10 level, in addition to increased angiogenesis and collagen formation. The results suggest the use of silver nanoparticles (AgCM) enhanced the CM properties by providing antibacterial properties, modulation the inflammatory phase, and consequently promotes the healing of skin lesions, which can be used clinically to treat injuries.
Assuntos
Interleucina-10 , Nanopartículas Metálicas , Ratos , Animais , Interleucina-10/farmacologia , Prata/farmacologia , Celulose , Peróxido de Hidrogênio/farmacologia , Ratos Wistar , Cicatrização , Antibacterianos/farmacologia , Bactérias , Colágeno/farmacologia , Modelos AnimaisRESUMO
This study aimed to investigate the application of low-intensity electrostimulation (ES) and electromagnetic stimulation (EM) associated with bioactive glass (BG) or allogeneic grafts (BB) in bone regeneration. A cell viability test on osteoblasts (UMR-106) was performed in the presence of BB and BG grafts associated with ES (10 µA/5 min) and EM (500 Hz/2 min). Critical defects (25 mm2 ) in calvaria were generated in male Wistar rats, and bone regeneration was evaluated on the 30th, 60th, and 120th days after surgery. Cell proliferation increased with the application of ES in both grafts and after EM with BG. Bone remodeling was more effective using the allogeneic graft in both therapies, with increased angiogenesis, osteoblast proliferation, and OPN expression in the BB + EM group. A higher number of osteoblasts and osteoclasts, and an increase in bone sialoprotein, Runx-2, and Opn gene expression were found in the BB + ES group. The BG graft associated with EM therapy had an increased proliferation of osteoblasts and increased expression of Runx-2 and Opn. Groups that had BG and ES therapy had increased numbers of osteoblasts, osteoclasts, and increased OPN expression. The expression of voltage-gated calcium channels increased in groups with ES, while calmodulin expression increased in therapies without grafting. ES and EM therapies favored the repair of bone defects upon grafting by improving angiogenesis, osteogenic gene expression, and tissue reorganization. Despite activating different pathways, both therapies increased the intracellular concentrations of calmodulin, leading to cell proliferation and bone regeneration.
Assuntos
Regeneração Óssea , Vidro , Aloenxertos , Animais , Diferenciação Celular , Masculino , Osteoblastos/metabolismo , Osteogênese , Ratos , Ratos Wistar , Transdução de SinaisRESUMO
CONTEXT: The quantification of total collagen is of major importance in a wide range of research areas, including the study of cutaneous wound healing and new drugs trials. OBJECTIVE: The total collagen content in skin biopsies was compared by biochemical hydroxyproline assay and by two computer-aided histomorphometric analyses of histological sections. MATERIALS AND METHODS: Two methods were used to evaluate collagen formation: the hydroxyproline assay, as the gold standard and histomorphometric image analysis of the filled areas by corresponding stained collagen fibres, using picrosirius and Gomori's trichrome staining. The image analyses were determined by digital densitometry recognition using computer-aided ImageJ software. One-way ANOVA, simple linear regression and ANCOVA were applied for the statistical analysis and correlation. RESULTS: In a simple linear regression analysis carried out on the 14th day period after the induction of skin injury, three techniques, picrosirius red (F = 33.57, p = 0.00), Gomori's trichrome (F = 81.61, p = 0.00) and hydroxyproline content (F = 16.85, p = 0.00) were able to detect collagen production. After scale adjustment, there were no significant differences among either the slopes (F = 1.17, p = 0.32) or the intercepts (F = 0.69, p = 0.51) of the estimated regression lines. It seems that a highly significant correlation exists between the histomorphometrical analysis and hydroxyproline assay. DISCUSSION AND CONCLUSION: The morphometric analysis proved to be adequate and can be used as a simple, rapid, low-cost technology for evaluating total collagen in cutaneous wound specimens, compared with the gold standard hydroxyproline assay.
Assuntos
Colágeno/análise , Pele/lesões , Cicatrização/fisiologia , Animais , Hidroxiprolina/análise , Processamento de Imagem Assistida por Computador , Masculino , Ratos , Ratos Wistar , Pele/químicaRESUMO
Hyaluronidases are enzymes that degrade hyaluronan an important constituent of the extracellular matrix. They have been used as a spreading agent, improving the absorption of drugs and facilitating the subcutaneous infusion of fluids. Here, we investigated the influence of bovine testes hyaluronidase (HYAL) during cutaneous wound healing in in vitro and in vivo assays. We demonstrated in the wound scratch assay that HYAL increased the migration and proliferation of fibroblasts in vitro at low concentration, e.g. 0.1 U HYAL enhanced the cell number by 20%. HYAL presented faster and higher reepithelialization in in vivo full-thickness excisional wounds generated on adult Wistar rats back skin already in the early phase at 2nd day post operatory compared to vehicle-control group. Wound closured area observed in the 16 U and 32 U HYAL treated rats reached 38% and 46% compared to 19% in the controls, respectively. Histological and biochemical analyses supported the clinical observations and showed that HYAL treated wounds exhibited increased granulation tissue, diminished edema formation and regulated the inflammatory response by modulating the release of pro and anti-inflammatory cytokines, growth factor and eicosanoids mediators. Moreover, HYAL increased gene expression of peroxisome proliferator-activated receptors (PPAR) γ and PPAR ß/δ, the collagen content in the early stages of healing processes as well as angiogenesis. Altogether these data revealed that HYAL accelerates wound healing processes and might be beneficial for treating wound disorders.