The regulatory role and related mechanisms of macrophages in wound healing / 中华烧伤杂志

Wound healing is a complex process under precise regulation, including multiple stages such as inflammation, anti-inflammatory, and regeneration. Macrophages play an important regulatory role in the differentiated process of wound healing due to their obvious plasticity. If macrophages fail to express specific functions in a timely manner, it will affect the healing function of tissues and lead to pathological tissue healing. Therefore, it is of great significance to understand the different functions of different types of macrophages and to regulate them specifically in different stages of wound healing to promote the healing and regeneration of wound tissue. In this paper, we illustrate the different functions of macrophages in the wound and their basic mechanisms, according to the basic process of wound healing, and emphasize the strategies of macrophage regulation that may be applied to clinical treatment in the future.

Effects of P311 on the angiogenesis ability of human microvascular endothelial cell 1 in vitro and its molecular mechanism / 中华烧伤杂志

Objective: To explore the effects of P311 on the angiogenesis ability of human microvascular endothelial cell 1 (HMEC-1) in vitro and the potential molecular mechanism. Methods: The experimental research method was used. HMEC-1 was collected and divided into P311 adenovirus group and empty adenovirus group according to the random number table (the same grouping method below), which were transfected correspondingly for 48 h. The cell proliferation activity was detected using the cell counting kit 8 on 1, 3, and 5 days of culture. The residual scratch area of cells at post scratch hour 6 and 11 was detected by scratch test, and the percentage of the residual scratch area was calculated. The blood vessel formation of cells at 8 h of culture was observed by angiogenesis experiment in vitro, and the number of nodes and total length of the tubular structure were measured. The protein expressions of vascular endothelial growth factor receptor 2 (VEGFR2), phosphorylated VEGFR2 (p-VEGFR2), extracellular signal-regulated kinase 1/2 (ERK1/2), and phosphorylated ERK1/2 (p-ERK1/2) in cells were detected by Western blotting. HMEC-1 was collected and divided into P311 adenovirus+small interfering RNA (siRNA) negative control group, empty adenovirus+siRNA negative control group, P311 adenovirus+siRNA-VEGFR2 group, and empty adenovirus+siRNA-VEGFG2 group, which were treated correspondingly. The protein expressions of VEGFR2, p-VEGFR2, ERK1/2, and p-ERK1/2 in cells were detected by Western blotting at 24 h of transfection. The blood vessel formation of cells at 24 h of transfection was observed by angiogenesis experiment in vitro, and the number of nodes and total length of the tubular structure were measured. HMEC-1 was collected and divided into P311 adenovirus+dimethylsulfoxide (DMSO) group, empty adenovirus+DMSO group, P311 adenovirus+ERK1/2 inhibitor group, and empty adenovirus+ERK1/2 inhibitor group, which were treated correspondingly. The protein expressions of ERK1/2 and p-ERK1/2 in cells were detected by Western blotting at 2 h of treatment. The blood vessel formation of cells at 2 h of treatment was observed by angiogenesis experiment in vitro, and the number of nodes and total length of the tubular structure were measured. The sample number at each time point in each group was 6. Data were statistically analyzed with independent sample t test, analysis of variance for repeated measurement, one-way analysis of variance, and least significant difference test. Results: Compared with that of empty adenovirus group, the proliferation activity of cells in P311 adenovirus group did not show significant difference on 1, 3, and 5 days of culture (with t values of -0.23, -1.30, and -1.52, respectively, P>0.05). The residual scratch area percentages of cells in P311 adenovirus group were significantly reduced at post scratch hour 6 and 11 compared with those of empty adenovirus group (with t values of -2.47 and -2.62, respectively, P<0.05). At 8 h of culture, compared with those of empty adenovirus group, the number of nodes and total length of the tubular structure of cells in P311 adenovirus group were significantly increased (with t values of 4.49 and 4.78, respectively, P<0.01). At 48 h of transfection, compared with those of empty adenovirus group, the protein expressions of VEGFR2 and ERK1/2 of cells in P311 adenovirus group showed no obvious changes (P>0.05), and the protein expressions of p-VEGFR2 and p-ERK1/2 of cells in P311 adenovirus group were significantly increased (with t values of 17.27 and 16.08, P<0.01). At 24 h of transfection, the protein expressions of p-VEGFR2 and p-ERK1/2 of cells in P311 adenovirus+siRNA negative control group were significantly higher than those in empty adenovirus+siRNA negative control group (P<0.01). The protein expressions of VEGFR2, p-VEGFR2, and p-ERK1/2 of cells in P311 adenovirus+siRNA negative control group were significantly higher than those in P311 adenovirus+siRNA-VEGFR2 group (P<0.01). The protein expressions of VEGFR2 and p-ERK1/2 of cells in empty adenovirus+siRNA negative control group were significantly higher than those in empty adenovirus+siRNA-VEGFR2 group (P<0.05 or P<0.01). At 24 h of transfection, the number of nodes of the tubular structure in cells of P311 adenovirus+siRNA negative control group was 720±62, which was significantly more than 428±38 in empty adenovirus+siRNA negative control group and 364±57 in P311 adenovirus+siRNA-VEGFR2 group (with P values both <0.01). The total length of the tubular structure of cells in P311 adenovirus+siRNA negative control group was (21 241±1 139) μm, which was significantly longer than (17 005±1 156) μm in empty adenovirus+siRNA negative control group and (13 494±2 465) μm in P311 adenovirus+siRNA-VEGFR2 group (with P values both <0.01). The number of nodes of the tubular structure in cells of empty adenovirus+siRNA negative control group was significantly more than 310±75 in empty adenovirus+siRNA-VEGFR2 group (P<0.01), and the total length of the tubular structure of cells in empty adenovirus+siRNA negative control group was significantly longer than (11 600±2 776) μm in empty adenovirus+siRNA-VEGFR2 group (P<0.01). At 2 h of treatment, the protein expression of p-ERK1/2 of cells in P311 adenovirus+DMSO group was significantly higher than that in empty adenovirus+DMSO group and P311 adenovirus+ERK1/2 inhibitor group (with P values both <0.01), and the protein expression of p-ERK1/2 of cells in empty adenovirus+DMSO group was significantly higher than that in empty adenovirus+ERK1/2 inhibitor group (P<0.05). At 2 h of treatment, the number of nodes of the tubular structure in cells of P311 adenovirus+DMSO group was 726±72, which was significantly more than 421±39 in empty adenovirus+DMSO group and 365±41 in P311 adenovirus+ERK1/2 inhibitor group (with P values both <0.01). The total length of the tubular structure of cells in P311 adenovirus+DMSO group was (20 318±1 433) μm, which was significantly longer than (16 846±1 464) μm in empty adenovirus+DMSO group and (15 114±1 950) μm in P311 adenovirus+ERK1/2 inhibitor group (with P values both <0.01). The number of nodes of the tubular structure in cells of empty adenovirus+DMSO group was significantly more than 317±67 in empty adenovirus+ERK1/2 inhibitor group (P<0.01), and the total length of the tubular structure of cells in empty adenovirus+DMSO group was significantly longer than (13 188±2 306) μm in empty adenovirus+ERK1/2 inhibitor group (P<0.01). Conclusions: P311 can enhance the angiogenesis ability of HMEC-1 by activating the VEGFR2/ERK1/2 signaling pathway.

Regulatory role and related mechanism of skin gamma-delta T cell subsets in wound re-epithelialization / 中华烧伤杂志

Re-epithelialization is one of the core links that determines the healing process of skin wounds. The proliferation and differentiation of epidermal stem cells to form new epidermal tissue is the histological basis of re-epithelialization, and the smooth progress of the cell differentiation process of epidermal stem cells-precursor cells-terminal cells is the cytological basis for the continuous formation of new epidermal tissue. The proliferation of stem cells and their differentiation into precursor cells are the determinants of the proliferative potential of newly formed epidermal tissue, while the expansion and differentiation of precursor cells into terminal cells are key factors determining the rate of new epidermal tissue formation. The tissue microenvironment plays a key regulatory role in the process of wound re-epithelialization, and cell growth factor and inflammatory mediators are the two main components of tissue microenvironment, which play regulatory role in different aspects of proliferation and differentiation of epidermal stem cells, jointly promoting the smooth progress of wound re-epithelialization As an important part of skin immune system, the subsets of gamma-delta (γδ) T cells play crucial role in dynamically shaping early wound microenvironment via secreting different cell growth factors and inflammatory factors. From the prospective of immune microenvironment of wound, this paper discusses the role of skin γδ T cells in maintaining the balance of stem cell proliferation and differentiation and regulating wound re-epithelialization, providing a new direction for the prevention and treatment of refractory wound.

Process of Hypertrophic Scar Formation: Expression of Eukaryotic Initiation Factor 6 / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Hypertrophic scar is one of the most common complications and often causes the disfigurement or deformity in burn or trauma patients. Therapeutic methods on hypertrophic scar treatment have limitations due to the poor understanding of mechanisms of hypertrophic scar formation. To throw light on the molecular mechanism of hypertrophic scar formation will definitely improve the outcome of the treatment. This study aimed to illustrate the negative role of eukaryotic initiation factor 6 (eIF6) in the process of human hypertrophic scar formation, and provide a possible indicator of hypertrophic scar treatment and a potential target molecule for hypertrophic scar.</p><p><b>METHODS</b>In the present study, we investigated the protein expression of eIF6 in the human hypertrophic scar of different periods by immunohistochemistry and Western blot analysis.</p><p><b>RESULTS</b>In the hypertrophic scar tissue, eIF6 expression was significantly decreased and absent in the basal layer of epidermis in the early period, and increased slowly and began to appear in the basal layer of epidermis by the scar formation time.</p><p><b>CONCLUSIONS</b>This study confirmed that eIF6 expression was significantly related to the development of hypertrophic scar, and the eIF6 may be a target molecule for hypertrophic scar control or could be an indicator of the outcomes for other treatment modalities.</p>

Effects of antisense p38 α mitogen-activated protein kinase on myocardial cells exposed to hypoxia and burn serum / 中华烧伤杂志

<p><b>OBJECTIVE</b>To study the effects of antisense p38α mitogen-activated protein kinase (hereinafter referred to as p38α) on myocardial cells exposed to hypoxia and burn serum.</p><p><b>METHODS</b>Thirty adult SD rats were inflicted with 40% TBSA full-thickness burn on the back to obtain burn serum. The myocardial cells were isolated from 80 neonatal SD rats and cultured, then they were divided into 4 groups according to the random number table: normal control group (N, ordinary culture without any treatment), hypoxia+burn serum group (HB, exposed to hypoxia after being treated with 10% burn rat serum), hypoxia+burn serum+infection group (HBI, exposed to hypoxia and 10% burn rat serum after being infected with antisense p38α gene-carrying adenovirus), hypoxia+burn serum+empty vector infection group (exposed to hypoxia and 10% burn rat serum after being infected with adenovirus empty vector). At post hypoxia hour (PHH) 1, 3, 6, and 12, mRNA and protein expression levels of p38α in the latter 3 groups were determined by RT-PCR and Western blotting, cell viability was determined by methylthianolyldiphenyl-tetrazolium bromide assay, and lactate dehydrogenase (LDH) activity was assayed at the same time point. At PHH 1, 6, and 12, apoptosis rate of myocardial cells was assessed by annexin V staining method. The indexes of group N were determined with the methods mentioned-above. Three wells were set at each time point in each group. Data were processed with one-way analysis of variance and LSD- t test.</p><p><b>RESULTS</b>(1) At PHH 1, 3, and 6, the p38α mRNA level was higher in group HB than in group N and group HBI (with t values from 2.725 to 4.375, P values all below 0.05). (2) At PHH 1, 3, and 6, the p38α protein level was higher in group HB than those in group N and group HBI (with t values from 5.351 to 7.981, P values all below 0.01). (3) At PHH 3, 6, and 12, the cell viability in group HB (0.115 ± 0.007, 0.104 ± 0.006, 0.094 ± 0.005) was lower than that in group N (0.141 ± 0.014) and group HBI (0.136 ± 0.009, 0.124 ± 0.010, 0.112 ± 0.007, with t values from 2.357 to 6.812, P values all below 0.05). (4) The LDH activity was up-regulated in group HB as compared with that in group N and group HBI at each time point (with t values from 22.753 to 201.273, P values all below 0.01). (5) At PHH 1, 6, and 12, the apoptosis rate of myocardial cells in group HB [(5.4 ± 0.7)%, (8.7 ± 1.1)%, (13.6 ± 1.7)%] was higher than that of group N [(3.1 ± 0.3)%] and group HBI [(4.3 ± 0.5)%, (5.1 ± 0.7)%, (7.2 ± 0.9)%, with t values from 2.345 to 9.700, P < 0.05 or P < 0.01].</p><p><b>CONCLUSIONS</b>Antisense p38α can protect the myocardial cells from the injury of hypoxia and burn serum.</p>

Study of xenotransplantation of fetal pig skin precursor tissue / 中华烧伤杂志

<p><b>OBJECTIVE</b>To select the optimal pregnancy time window of embryonic pig skin precursor tissue for xenotransplantation and study its ability in wound repair.</p><p><b>METHODS</b>Skin precursor tissues were obtained from pig fetus of fetal age of 35, 42, 56, 70 days, and were minced into microskin and transplanted to dorsal wounds of BALB/c nude mice, then they were covered with residual skin after plastic surgery of patients or adult pig skin (white). The characteristics of growth and development were observed after transplantation. Pathological examination was performed on 6 and 12 post operation weeks respectively to observe the tissue structure and tumorigenicity.</p><p><b>RESULTS</b>Skin precursor tissues from fetal pig survived and developed after transplantation, and the microskin fused. New tissue area from skin precursor tissues with fetal age of 42 days was (47 +/- 6) mm2, which was higher than that of 35 days (18 +/- 8 mm2), 56 days (31 +/- 12 mm2), 70 days (20 +/- 8 mm2, P < 0.05). The skin precursor developed into "intact skin" with hair, sebaceous glands and sweat glands, and melanocytes were also detected in epidermis. The newly-grown skin tissue included epidermal and dermal layer, and obvious dermal papillae. Teratoma was not found after transplantation in skin precursor tissue with fetal age of 56, 70 days.</p><p><b>CONCLUSION</b>Fetal pig skin precursor tissue with fetal age of 56 days can be used to repair wound as xenotransplantation.</p>

Blockage of allergic rhinitis in mice with recombinant protein of cytotoxic T lymphocyte associated antigen-4 extracellular domain / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>To explore a new immunotherapy against allergic rhinitis.</p><p><b>METHODS</b>The recombinant protein of CTLA4 extracellular domain was obtained through construction of CTLA4-yeast expression system. The allergic rhinitis in mice was induced by sensitizing and challenging with ovalbumin (OVA). The allergic rhinitis related symptoms and the morphological changes in nasal mucosa were compared between the allergic rhinitis group and the CTLA4 extracellular domain group treated with CTLA4 extracellular domain before each challenge by ways of intraperitoneal injection.</p><p><b>RESULTS</b>CTLA4 extracellular domain with a molecular weight of 28 000, which was confirmed by Western blot, could be generated through CTLA4-yeast expression system. The purified CTLA4 extracellular domain could inhibit T cells proliferation in mixed lymphocyte reaction with a inhibitory rate of 95.4%. The mice in allergic rhinitis group appeared typical allergic rhinitis symptoms after OVA challenge, such as rhinorrhea and sneeze. Meanwhile the nasal pathological studies showed edema and congestion in mucosa tissue and local influx of inflammatory cells. Whereas in CTLA4 extracellular domain group, the nasal symptoms were rarely observed, and the pathological change in nasal mucosa was significantly abated.</p><p><b>CONCLUSIONS</b>The protein of CTLA4 extracellular domain could prevent the allergic rhinitis in mice. The underlying mechanism of which might be the inhibition of the T cell activation.</p>

An experimental study on the role of indirect antigen presentation pathway on the immunogenetic of epidermal cells / 中华烧伤杂志

<p><b>OBJECTIVE</b>To explore the role of indirect antigen presentation pathway on the immunogenecity of epidermal cells.</p><p><b>METHODS</b>Human epidermal cells (HEC), allogeneic human peripheral blood lymphocytes (PBL) and mononuclear cells (PBM, including monocytes) were isolated and cultured in vitro. HECs were transfected by human-originated CTLA4Ig-adenovirus vector. The CTLA4Ig expression was observed. Allogeneic PBLs or PBMs were added to the transfected and non-transfected HECs with simple cultured PBLs and PBMs as the control. The proliferation of PBL and PBM was determined by (3)H-TdR incooperation.</p><p><b>RESULTS</b>HECs could be successfully transfected by CTLA4Ig-adenovirus vector and expressed corresponding proteins. The non-transfected HECs could stimulate slight proliferation of allogeneic PBLs (P < 0.05) and stimulate remarkable proliferation of PBMs (including monocytes) (P < 0.05). The proliferation reaction of PBLs and PBMs decreased significantly (P < 0.05) after being stimulated by HEC which was modulated by CTLA4Ig genes.</p><p><b>CONCLUSION</b>Indirect antigen presentation pathway might play important roles in the HEC immunogenicity which could be evidently inhibited by CTLA4Ig.</p>