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ObjectiveTo establish a rat model of diabetic wound by feeding on a high-fat and high-sugar diet combined with intraperitoneal injection of streptozotocin （STZ） and surgical preparation of full-thickness skin defects， observe the effect of cinnamaldehyde on the wound healing of diabetes rats， and explore the therapeutic mechanism of cinnamaldehyde in improving wound healing of diabetes rats based on the PTEN-induced putative kinase （PINK1）/Parkin pathway-mediated mitochondrial autophagy. MethodForty-eight male SD rats were randomly divided into blank group （n=12） and diabetes group （n=36）. The diabetes group was further randomly divided into model group， cinnamaldehyde group， and Beifuxin group， with 12 rats in each group. The blank group and the model group received routine disinfection with physiological saline after creating the wounds， while the cinnamaldehyde group received topical application of polyethylene glycol 400 （PEG 400） gel containing 4 μmol·L-1 cinnamaldehyde， and the Beifuxin group received topical application of Beifuxin gel. Dressings were changed once daily. The wound healing rate of each group was observed. On the 7th and 14th days after intervention， the wound tissues of the rats were collected. Hematoxylin and eosin （HE） staining was performed to observe the pathological changes in the local tissues. Immunohistochemistry （IHC） was used to detect the expression of interleukin-6 （IL-6）， tumor necrosis factor-α （TNF-α）， vascular endothelial growth factor （VEGF）， and collagen fibers. Immunofluorescence （IF） and Real-time polymerase chain reaction （Real-time PCR） were used to detect the protein， and mRNA expression of PINK1， Parkin， microtubule-associated protein 1 light chain 3 Ⅱ （LC3 Ⅱ）. ResultAfter intraperitoneal injection of STZ， compared with the blank group， the random blood glucose values of rats in the diabetic group increased significantly （P<0.01）， all higher than 16.7 mmol·L-1， and persistently hyperglycemic for some time after modeling. Compared with the blank group， the model group showed poor growth and healing of granulation tissue in the wounds， and the wound healing rate decreased （P<0.01）. On the 7th day after intervention， the blank group had squamous epithelial coverage on the wounds. Compared with the blank group， the model group only had a small amount of scab at the wound edges， with a large number of infiltrating inflammatory cells in the wounds. The protein expression levels of IL-6 and TNF-α in the tissues increased （P<0.01）， and the protein and mRNA levels of PINK1， Parkin， and LC3Ⅱ decreased （P<0.01）. On the 14th day after the intervention， the granulation tissue in the wounds of the blank group was mature and well-healed. Compared with the blank group， the model group still had infiltrating inflammatory cells and red blood cell exudation. The protein expression levels of VEGF and collagen fibers in the tissues decreased （P<0.01）， and the protein and mRNA expression levels of PINK1， Parkin， and LC3Ⅱ increased （P<0.01）. Compared with the model group， the cinnamaldehyde group and the Beifuxin group showed better wound healing， with increased wound healing rates （P<0.01）. On the 7th day after intervention， the protein expression levels of IL-6 and TNF-α in the tissues decreased （P<0.01）， and the protein and mRNA expression levels of PINK1， Parkin， and LC3Ⅱ increased （P<0.01）. On the 14th day after intervention， the protein expression levels of VEGF and collagen fibers in the tissues increased （P<0.01）， and the protein and mRNA expression levels of PINK1， Parkin， and LC3Ⅱ decreased （P<0.01）. ConclusionCinnamaldehyde can promote the wound healing of diabetes rats by increasing the wound healing rate， reducing the levels of inflammatory factors IL-6 and TNF-α， and increasing the levels of VEGF and collagen fibers. Its mechanism may be related to the regulation of the PINK1/Parkin signaling pathway， activation of mitochondrial autophagy， inhibition of inflammatory responses， and promotion of angiogenesis and collagen synthesis， thereby promoting the wound healing of diabetes rats.
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Objective:To investigate the effect of Astragaloside Ⅳ-mediated Endothelial progenitor cells derived exosomes (EPC-Exos) on the biological function of EPC-Exos damaged by high glicose.Methods:EPCs from human umbilical cord blood were isolated and cultured in vitro. the EPC-Exos secreted by EPCs were extracted by ultracentrifugation combined with ultrafiltration, and identified by specific markers CD9, CD63 and CD81, respectively. After the cells were cultured for 24 hours with AS-IV at 100 mg/L and PBS at the same volume, the morphological characteristics of EPC-Exos were observed by transmission electron microscope. Human endothelial cells were isolated, cultured and identified in vitro. The identified endothelial cells were pretreated with 30 mmol/L glucose for 120 h and randomly divided into experimental group and control group, at the same time set the normal group. The cells were cultured for 24 hours, the effects of EPC-Exos on proliferation, adhesion, migration and angiogenesis of endothelial cells damaged by high glucose were observed by using cell counting kit-8 (CCK-8) Cell Proliferation Assay Kit, cell scratch test, adhesion assay and in vitro angiogenesis assay by Matrigel. Results:Compared with the normal group, the proliferation, migration, adhesion and tubulogenesis of human endothelial cells in the control group were significantly lower ( t=24.35, 6.80, 10.65, 9.62, P<0.05). Compared with the control group, the proliferation, adhesion, migration and tubulogenesis of human endothelial cells in the experimental group were significantly enhanced ( t=30.68, 5.99, 5.40, 8.25, P<0.05). Conclusions:EPC-Exos mediated by AS-Ⅳ can significantly improve the biological function of human endothelial cells damaged by high glucose and has the potential to modulate endothelial neovascularization in diabetic rats.
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Objective To prepare a new-type acellular dermal matrix (ADM) and research on its relevant performance,which would provide theoretical evidence for clinical application.Methods Skin of Bama suckling pig was taken as resource of skin,and technologies of physics,chemistry and biology were selected to prepare new-type ADM.To detect the external structure,physical and chemical property as well as biological property of the prepared new-type ADM,hematoxylin-eosin (HE) staining observation,scanning electron microscope observation,amino acid analysis,material porosity and hydrophilicity test,tensile strength and in vitro degradation experiment,cytotoxicity test,and animal experiment have been conducted.Results New-type ADM cells have been thoroughly removed and dermal matrix remains intact with collagen content of 95.55％,connective three-dimensional pore structure,(85.03 ± O.99) ％ of porosity,(24.56 ± 0.57) ° of contact angle implying new-type ADM was hydrophilic substance,(5.48 ± 0.44) Pa of tensile strength implying its moderate level of pulling force,in vitro degradation period reduced to (28.7 ± O.76) h,and ＞75％ relative growth rate (RGR).Cells grew and proliferated on new-type ADM and could be replaced by original tissue after degradation.Conclusions New-type ADM have overcome disadvantages of traditional preparation method in sabotaging dermal matrix structure and incompletely removing cells from matrix,which is qualified with higher level of collagen content and porosity.With improved biological property,greatly reduced inflammation immunoreactions,and accelerated degradation rate,new-type ADM is of higher level of clinical application value.
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Objective To explore the effectiveness and safety of tiotropium bromide inhalation agent with tulobuterol patch in the treatment of C and D level of COPD in stable period .Methods According to the digital table,255 cases of C and D level in patients with stable COPD were randomly divided into the study group and control group,the patients in study group received inhalation of tiotropium bromide dry powder 18μg/times,one time every day,and give tulobuterol patch(2mg/paste),one time every day.The control group received inhalation of tiotropium bromide dry powder 18μg/times,one time every day.The changes of lung function were observed before and after treatment,the clinical symptom score and inhaled short acting beta 2 agonists used,6min walk test,times of acute exacerbation condition.Results The patients in the two groups after treatment ,pulmonary function ,clinical symptoms score,inhaled short acting beta 2 agonists used,6min walk test,times of acute exacerbation compared with statistical significance(all P0.05).Conclusion Tiotropium bromide inhalation agent with tulobuterol patch can improve clinical symptoms and pulmonary function in patients with C and D level part of the stable phase of COPD .