Future application of hair follicle stem cells: capable in differentiation into sweat gland cells / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Sweat glands (SGs) can not regenerate after complete destruction in the severe skin injury, so it is important to find a ideal stem cell source in order to regenerate functional SGs. Hair follicle stem cells (HFSCs) possess the obvious properties of the adult stem cells, which are multipotent and easily accessible. In this research, we attempted to direct the HFSCs suffered from the sweat gland cells (SGCs) special differentiation by a co-operative coculture system in vitro.</p><p><b>METHODS</b>The designed co-culture microenvironment in the transwell was consist of two critial factors: heat shocked SGCs and dermis-like mesenchymal tissue, which appeared independently in the two control groups; after induction, the purified induced SGC-like cells were transplanted into the full-thickness scalded wounds of the nude mice, after 4 weeks, the reconstructed SG-like structures were identified by immunohistochemical and immunofluorescence analysis.</p><p><b>RESULTS</b>A part of HFSCs in experimental group finally expressed SGCs phenotypes, by contrast, the control group 1 which just containing dermis-like mesenchymal tissue failed and the control group 2 consisted of heat shocked SGCs was in a poor efficiency; by immunofluorescence staining and flow cytometry analysis, the expression of HFSCs special biomarkers was down regulated, instead of the positive efficiency of SGCs special antigens increased; besides, the induced SGCs displayed a high expression of ectodysplasin A (EDA) and ectodysplasin A receptor (EDAR) genes and proteins; after cell transplantation, the youngest SG-like structures formed and be positive in SGCs special antigens, which never happened in untreated wounds (P < 0.05).</p><p><b>CONCLUSION</b>The HFSCs are multipotential and capable in differentiating into SGCs which promise a potential stem cells reservoir for future use; our special co-culture microenvironment is promising for HFSCs differentiating; the induced SGCs are functional and could work well in the regeneration of SGs.</p>

Transplantation of human bone marrow-derived mesenchymal stem cells transfected with ectodysplasin for regeneration of sweat glands / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Patients with severe full-thickness burn injury suffer from their inability to maintain body temperature through perspiration because the complete destructed sweat glands can not be regenerated. Bone marrow-derived mesenchymal stem cells (BM-MSCs) represent an ideal stem-cell source for cell therapy because of their easy purification and multipotency. In this study, we attempted to induce human BM-MSCs to differentiate into sweat gland cells for sweat gland regeneration through ectodysplasin (EDA) gene transfection.</p><p><b>METHODS</b>The dynamic expression of EDA and EDA receptor (EDAR) were firstly observed in the sweat gland formation during embryological development. After transfection with EDA expression vector, human BM-MSCs were transplanted into the injured areas of burn animal models. The regeneration of sweat glands was identified by perspiration test and immunohistochemical analysis.</p><p><b>RESULTS</b>Endogenous expression of EDA and EDAR correlated with sweat gland development in human fetal skin. After EDA transfection, BM-MSC acquired a sweat-gland-cell phenotype, evidenced by their expression of sweat gland markers by flow cytometry analysis. Immunohistochemical staining revealed a markedly contribution of EDA-transfected BM-MSCs to the regeneration of sweat glands in the scalded paws. Positive rate for perspiration test for the paws treated with EDA-transfected BM-MSCs was significantly higher than those treated with BM-MSCs or EDA expression vector (P < 0.05).</p><p><b>CONCLUSIONS</b>Our results confirmed the important role of EDA in the development of sweat gland. BM-MSCs transfected with EDA significantly improved the sweat-gland regeneration. This study suggests the potential application of EDA-modified MSCs for the repair and regeneration of injured skin and its appendages.</p>

A new way for isolation and cultivation of sweat gland ductal cells from human split-thickness skin in vitro / 中华外科杂志

<p><b>OBJECTIVE</b>To explore a new method of isolation and culture of eccrine sweat gland ductal cells from human split-thickness skin graft in vitro.</p><p><b>METHODS</b>Human split-thickness skin graft which was presented by volunteer (n = 10) was digested with type II collagenase, and then sweat gland duct were isolated from the split-thickness skin graft, primary cultures were incubated at 37 degrees C in humidified atmosphere of 5% CO2, 95% O2. The cultured eccrine sweat gland ductal cells were identified by analysis CEA, CK8, CK18, CK19 antigens expression with flow cytometry, RT-PCR and Western Blot, and by detecting the electrophysiology with whole cell patch clamp technology.</p><p><b>RESULTS</b>The isolated eccrine sweat gland ductal cells could grow by adhering to the wall, proliferate in vitro after 48 h of adhering to the wall, and confluens after 2 - 4 weeks of adhering to the wall. The FACs analysis showed the expression of CEA was (90.26 +/- 1.12)%, (89.70 +/- 1.43)%, and CK8 was (94.41 +/- 1.84)%, (93.65 +/- 1.63)% in primary cultured sweat gland ductal cells and primary cultured eccrine sweat gland cells, respectively, and there is no significant difference between the two groups (P > 0.05). Immunocytochemistry staining showed CEA, CK8, CK18, CK19 was positive in sweat gland duct cells, RT-PCR revealed that CEA, CK8, CK18 and CK19 gene expression in sweat gland ductal cells, and Western Blot analysis showed the expression of CEA brand, CK8 brand, CK18 brand, and CK19 brand in sweat gland ductal cells, patch clamp indicated that this cells has distinct amiloride sensitive Na(+) channels.</p><p><b>CONCLUSIONS</b>The cultured human eccrine sweat gland duct cells in vitro display the markers and biological characteristics of sweat gland epithelial lineage, and this method of digest the split-thickness skin graft to get the sweat gland duct cells is better than classical dissect sweat gland under dissect microscope.</p>

Promotive effect of adipose-derived stem cells on the wound model of human epidermal keratinocytes in vitro / 中华外科杂志

<p><b>OBJECTIVE</b>To investigate the migrating effect of adipose derived stem cells (ADSCs) on the wound model of human epidermal keratinocyte (HEKa).</p><p><b>METHODS</b>Rat ADSCs (rADSCs) were isolated and cultured (n = 10), rADSCs were direct co-cultured with HEKa cells in experiment group (experimental group, n = 10). In the control groups, rADSCs were indirect co-cultured with HEKa cells in transwell chamber (indirect group, n = 8), or HEKa was cultured alone (single group, n = 8). Then confluent HEKa cells were scraped to establish a wound model under invert microscope. After scraped 24, 48, and 72 h, cell numbers of which migrated across the edge of the wound was measured, the rate of wound healing was calculated by using SigmaScan Pro 5 software, and the proliferating effect of rADSCs on HEKa were examined by incorporation of [(3)H] thymidine.</p><p><b>RESULTS</b>The cells migrated across the edge of wound after 24 hours in experimental group, indirect group, and single group were (9.2 + or - 0.2), (5.0 + or - 0.3), (4.2 + or - 0.3), and were (58.5 + or - 0.4), (26.5 + or - 0.3), (20.7 + or - 0.5) 48 hours after, and were (125.8 + or - 0.4), (43.0 + or - 0.5), (35.6 + or - 0.5) cells/HP 72 hours after, respectively; the numbers were all significantly higher in experimental group than those in control groups (P < 0.05). The rates of wound healing after scraped 72 hours were 61.0% + or - 3.0%, 35.0% + or - 2.5% and 32.0 + or - 2.1%, the outcome in experimental group was significantly better than in the control groups (P < 0.05). And the thymidine feeding displayed the proliferation of HEKa in the three groups were (1440 + or - 210), (1050 + or - 280) and (1130 + or - 390) cpm/10(5) cell, and there was significant difference between the experimental and the control groups (P < 0.05).</p><p><b>CONCLUSIONS</b>The rADSCs can promote the migration of HEKa by direct contact with it.</p>

Chemotactic effects of burn rat serum on mesenchymal stem cells derived from different sources / 中华烧伤杂志

<p><b>OBJECTIVE</b>To isolate and culture mesenchymal stem cells ( MSC) from different sources and to investigate the chemotactic effects of burn rat serum on MSC derived from different sources.</p><p><b>METHODS</b>Seventy-two Wistar rats were randomly divided into burn( n = 36, with 30% TBSA full-thickness burns on the back) and sham burn (n = 36, without burns) groups. Bone marrow and peripheral blood of the rats in both groups were collected to isolate and culture MSC. Ratio of MSC, growth speed and cell morphology were observed with inverted microscope. Effects of different serum ( fetal bovine serum, normal rat serum and burn rat serum) on chemotaxis of MSC derived from different sources and their migration ability were subsequently examined with a transwell system. Results MSC were obtained from bone marrow of the rats in both groups. MSC were successfully obtained from bone marrow of all burn rats(100% , P <0.05) , but only from peripheral blood of 7 burn rat(58% ) , and no MSCs were obtained from peripheral blood of 12 rats in sham group( P <0.05). There was small amount of adherent cells 24 hrs after culture, and fusiform shaped adherent cells were sporadically observed in scattered distribution 2-3 days later with inverted microscope. There was no obvious difference in the cell morphology between the 2 groups. In the sham group, the number of MSC migrating to the lower surface of transwell after burn serum treatment [ (94 Il ) cells/ high power field] was significantly greater than that after the treatment with normal rat serum and fetal bovine serum [ (37 +/- 6) , (38 +/- 11) cells/high power field , P <0.01 ] , while no difference in migration ability was found after normal serum treatment compared with that after fetal bovine serum treatment ( P >0. 05). The migration rate of MSCs which were derived from bone marrow in sham group was obviously lower than those derived from bone marrow and peripheral blood from burn rats ( P <0. 05 or 0. 01). Though some difference of the migration ability existed between MSC derived from bone marrow and peripheral blood, there was no statistically significant difference ( P >0. 05).</p><p><b>CONCLUSION</b>MSC can be isolated and cultured from bone marrow and peripheral blood of burn rat, but not from peripheral blood of normal rat. Burn rat serum has a stronger chemotactic effect on MSC. Moreover, the migration ability of MSC derived from burn rat is stronger than that of MSC derived from normal rat.</p>

Identification and cell phenotype transdifferentiation of adipose-derived stem cells / 中华整形外科杂志

<p><b>OBJECTIVE</b>To investigate the transdifferentiation of the ADSCs to epidermal cells.</p><p><b>METHODS</b>ADSCs were isolated and cultured from rat adipose tissue by digestion of enzyme. ADSCs was identified by immunocytochemistry and flow cytometry. ADSCs were divided into four groups in order to induce: the condition medium (containing 30% superior of homogenizing rat skin in 10% FBS/DMEM) group, 7 days; 10% FBS/DMEM with EGF (20 ng/ml) group, 7 days; the condition medium for 4 days and then 10% FBS/DEME instead of the condition medium for 3 days group; 10% FBS/DMEM for 7 days group (control group). Cytokeratin 19 and cytokeratin 10 expressions in ADSCs were detected by flow cytometry.</p><p><b>RESULTS</b>(1) The results of immunocytochemistry showed that ADSCs were positive for CD49d and negative for CD106, CD34, CD19, CD10. The results of flow cytometry showed ADSCs were positive for CD49d and CD44. (2) The CK19 expression of ADSCs was 45.32% in the condition medium group, 26.58% in the condition medium with EGF group, 23.37% in te condition medium for 4 days and then 10% FBS/DMEM instead of the condition medium for 3 days gropu and 18.53% in control group, P <0.01. The CK10 expression of ADSCs was 43.56% in the condition medium group, 25.54% in the condition medium with EGF group, 18.20% in the condition medium for 4 days and then 10% FBS/DMEM instead of the condition medium for 3 days group and 2.46% in control group, P < 0.01.</p><p><b>CONCLUSIONS</b>The superior of homogenizing rat skin can induce CK19 and CK10 expressing in ADSCs, and thereby demonstrating ADSCs can differentiate to epidermal cell phenotype in vitro.</p>

Analysis of differentially expressed genes in fetal skin of scarless and scar-forming periods of gestational rats / 中华创伤杂志(英文版)

<p><b>OBJECTIVE</b>To study the differences of gene expression between earlier gestational skin and later gestational skin of rats with the aids of single primer amplification (SPA) and high-density oligonucleotide DNA array to understand the molecular mechanism of scarless healing.</p><p><b>METHODS</b>Total RNAs were isolated from fetal rat skin of the scarless (E15) and scar-forming (E18) periods of gestation (term = 21.5 days). The RNAs from earlier gestational skin (EGS) and later gestational skin (LGS) were both reversely transcribed to cDNAs, then labeled with the incorporation of fluorescent dCTP for preparing the hybridization probes by SPA method. The mixed probes were then hybridized to the oligonucleotide DNA arrays which contained 5,705 probes representing 5,705 rat genes. After highly stringent washing, these DNA arrays were scanned for fluorescent signals to display the differentially expressed genes between the 2 groups of skin.</p><p><b>RESULTS</b>Among 5,705 rat genes, there were 53 genes (0.93 percent) with differentially expressed levels between EGS and LGS groups, 27 genes, including fibroblast growth factor 2 (FGF2) and follistatin were up-regulated (0.47%) and 26 genes were down-regulated (0.46%) in fetal skin during scarless period versus scar-forming period. Higher expressions of FGF2 and follistatin in EGS than those in LGS were also revealed by RT-PCR method.</p><p><b>CONCLUSIONS</b>High-density oligonucleotide DNA array provided a powerful tool for investigating differential gene expression in earlier and later gestational fetal skins. This technology validates that the mechanism of fetal scarless healing is very complicate and the change of many gene expressions is associated with fetal scarless healing.</p>

Relationship between the proliferation and immunoinduction of epithelial cells and the destruction of hair follicles and sebaceous glands in keloids / 中华整形外科杂志

<p><b>OBJECTIVE</b>To investigate the influence of biological behavior of epithelial cells on the hair follicles and sebaceous glands (HFSG) structure in keloids (K).</p><p><b>METHODS</b>The expression of intercellular adhesion molecule (ICAM)-1, D-related human leucocyte antigen (HLA-DR), and cytokeratin (CK) 14 on epithelial cells and the amount of activity T-lymphocytes were detected in specimens of keloid edge and normal skin with immunohistochemical and histological methods.</p><p><b>RESULTS</b>In comparison with normal skin specimens, epithelial cells were proliferated in K-HFSG presented structural aberration and disintegrate or abnormally to form solid-epithelial island-like structure, and the density of HSFG with hyperplasia and the ageing scar in keloids was apparently decreased. They strongly expressed ICAM-1, HLA-DR, and CK14 in the epithelial cells, there were many immunologic cells which expressed CD4, CD45RO, and interferon (IFN)-gamma around the K-HFSG. The expressed level of epithelial cells was positively correlated with the density of immunologic cells nearby K-HFSG.</p><p><b>CONCLUSION</b>It could be concluded that the reactivity with hyperplasia and immunoinduction of epithelial cells might be associated with the destruction of the some HFSG structure in the keloids.</p>

Ontogeny of expression of basic fibroblast growth factor and its receptors in human fetal skin / 中华创伤杂志(英文版)

<p><b>OBJECTIVE</b>To investigate the expression characteristics of basic fibroblast growth factor (bFGF) and its receptors, flg (FGFR1) and bek (FGFR2), in fetal skin at different gestational ages underlying the relevance of these 3 proteins to skin development and the mechanisms underlying the phenotypic transition from scarless to scar-forming healing.</p><p><b>METHODS</b>Eighteen specimens of fetal skin biopsies of human embryo were obtained from spontaneous abortions at different gestational ages of 13-32 weeks. Gene expression of bFGF, bek and flg was examined with reverse transcription-polymerase chain reaction (RT-PCR). The dynamic expression and distribution of these 3 proteins were detected with streptavidin peroxidase (SP) immunohistochemical staining method.</p><p><b>RESULTS</b>In the early gestational fetal skin, genes of bFGF and flg were strongly expressed and more protein contents of these 2 proteins were found as compared with the genes at late gestation fetal skin (2.446+/-0.116 and 2.066+/-0.152 versus 2.157+/-0.101 and 1.818+/-0.086, respectively, P<0.05). On the contrary, the levels of gene expression and protein content of bek were not differently expressed in the early gestational fetal skin versus the late ones. Protein particles of bFGF were mainly distributed in the epidermal cells and some fibroblasts. Bek was mainly located in the cell membrane and cytoplasm of epidermal cells while flg protein was principally located in the epidermal cells, endothelial cells and some fibroblasts.</p><p><b>CONCLUSIONS</b>The endogenous bFGF and their receptors might be involved in the cutaneous development at fetal stage. The differently expressing levels of bFGF and flg during gestation may be related to scarless or scar-forming repair during gestation.</p>

Investigation of mesenchymal-epithelial transdifferentiation in the morphogenesis mechanism of embryonic epidermic cells / 中华外科杂志

<p><b>OBJECTIVE</b>To study the relationship between the morphologic mechanism of human embryonic epidermic cells and mesenchymal-epithelial transformation (MET) and its modulation factor.</p><p><b>METHODS</b>Morphological occurrence of epidermis was detected with histologic methods in earlier period [estimated gestational age (EGA) 6-14 weeks] human embryonic skin samples. At the same time, the characteristic expression and their distribution markers of mesenchymal cells [vimentin and alpha-smooth muscle actin (alpha-SMA)], embryonic specific epidermic protein CK8&18, specific protein of epidermic stem cell CK19, transforming growth factor-beta1) (TGF-beta1) and its receptor (TGFbetaRI) in embryonic epidermis were examined with immunohistochemistry and indirect-immunofluorescent doble-labelling method.</p><p><b>RESULTS</b>During EAG 6-8 weeks, ectodermal cells containing Vim+/alpha-SMA(-) were found to transform into epidermal stem cells with CK8&18+/CK19+. In ectodermal cells, protein expression density of TGFbetaRI was moderate (+ +), while positive signal of TGFbeta1 was weak (+/-). After EGA10 weeks, epidermal cells showed typical morphological characteristics.</p><p><b>CONCLUSIONS</b>At EGA 6-8 weeks, human embryonic skin epidermal cells began to form through MET, in which the signal pathway mediated by TGFbetaRI might play important roles, but the role of TGFbeta1 need to be further studied.</p>

Morphological and distribution characteristics of sweat glands in hypertrophic scar and their possible effects on sweat gland regeneration / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>In hypertrophic scar tissue, no sweet gland and hair follicle exist usually because of the dermal and epidermal damage in extensive thermal skin injury, thus imparing regulation of body temperature. This study was designed to reveal the morphological and distributional characteristics of the sweat glands in normal skin and hypertrophic scar obtained from children and adults, and to study the possible interfering effects of the scar on regeneration of the sweat gland after burn injury.</p><p><b>METHODS</b>Biopsies of hypertrophic scar were taken from four children (4 - 10 years) and four adults (35 - 51 years). Normal, uninjured full-thickness skin adjacent to the scar of each patient was used as control. Keratin 19 (K19) was used as the marker for epidermal stem cells and secretory portion of the sweat glands, and keratin 14 (K14) for the tube portion, respectively. Immunohistochemical and histological evaluations were performed.</p><p><b>RESULTS</b>Histological and immunohistochemical staining of skin tissue sections from both the children and adults showed K19 positive cells in the basement membrane of epidermis of normal skin. These cells were seen only single layer and arranged regularly. The secretory or duct portion of the eccrine sweat glands was situated in the dermis and epidermal layer. However, in the scar tissue, K19 positive cells were scant in the basal layer, and the anatomic location of the secretory portion of sweat glands changed. They were located between the border of the scar and reticular layer of the dermis. These secretory portions of sweat glands were expanded and were organized irregularly. But a few K14 positive cells were scattered in the scar tissues in cyclic form.</p><p><b>CONCLUSIONS</b>There are some residual sweat glands in scar tissues, in which the regeneration process of active sweat glands is present. Possibly the sweat glands could regenerate from adult epidermal stem cells or residual sweat glands in the wound bed after burn injury.</p>

Study on the potentiation of bone marrow mesenchymal stem cells involved in sebaceous duct formation / 中华外科杂志

<p><b>OBJECTIVE</b>To investigate the feasibility of differentiation of bone marrow mesenchymal stem cells (MSCs) into skin appandence.</p><p><b>METHODS</b>Porcine MSCs were isolated from porcine marrow and grown in vitro. After labeling with BrdU, MSCs were engrafted to porcine skin. At 1, 2, 4 weeks after the transplantation, immunohistochemical examinations were carried out to detect the positive staining of BrdU and cytokeratin.</p><p><b>RESULTS</b>A few sebaceous duct cells, which expressed cytokeratin, were also BrdU positive, and these cells were considered may to be transplanted MSCs-derived cells.</p><p><b>CONCLUSION</b>Porcine MSCs might have the potential to differentiated into sebaceous duct cells in skin.</p>

Relationship of overexpression of angiogenesis factors and their receptors with invasive growth of keloid / 中华整形外科杂志

<p><b>OBJECTIVE</b>To study the relationship between invasive growth and the angiogenesis factors and their receptors in keloid.</p><p><b>METHODS</b>Biopsies from 17 keloid (Ke) were divided into atrophy group (Ke-A, n = 9), proliferating group (Ke-P, n = 13), infiltrating group (Ke-I, n = 9), normal skin around Ke (Ke-N, n = 10) and normal skin (NS, n = 10). The histology, immunohistochemistry and computerized imaging analysis were used for the study. The levels of basic fibroblast growth factor (bFGF) and its receptor-Flg, vascular endothelial growth factor (VEGF) and VEGF/KDR complex (11B5), and platelet derived growth factor (PDGF-A) and its receptor-PDGFR-alpha, and alpha-smooth muscle actin (alpha-SMA) were determined in specimens with immuneohistochemical staining.</p><p><b>RESULTS</b>In all 5 groups, bFGF, Flg, VEGF, 11B5, PDGF-A, and PDGFR-alpha were all expressed in fibroblasts (Fb), monocyte-phagocytes, vascular endothelial cells, adventitial cells, epidermal (cells and epithelial cells in appendage. The intensities of staining ranked as follows: Ke-I > Ke-N approximately equal to Ke-P > Ke-A approximately equal to NS, Flg > hFGF approximately equal to PDGFR-alpha > PDGF-A approximately equal to 11B5 > VEGF (P < 0.05 to approximately 0.01). 11B5 and VEGF were expressed (intensively in alpha-SMA positive myofibroblasts only in Ke-I group. The histological observation showed hyperplasia of endothelial cells and obliteration of microvessels.</p><p><b>CONCLUSION</b>The invasive growth of keloid may be related to the overexpression of angiogenesis factors and their receptors. The abnormal expression of 11B5 in myofibroblasts may be one of the important factors associated with tumor-like growth feature in the invasive parts sites of keloid. The results suggest that inhibition of these biological activities would be of significance in clinical therapy.</p>

Gene expression of extracellular-signal regulated protein kinase 5 and their MAPKK in fetal skin hypertrophic scars / 中华整形外科杂志

<p><b>OBJECTIVE</b>To explore the change of gene expression of extracellular-signal regulated protein kinase 5 (ERK5) and its upstream signaling molecule (MEK5) in fetal skin of differentially developmental stages and hypertrophic scars.</p><p><b>METHODS</b>After morphological characteristics of skin of different developmental stages and hypertrophic scars were detected with pathological methods, gene expression of ERK5 and MEK5 was examined with reverse transcription-polymerase chain reaction analysis (RT-PCR).</p><p><b>RESULTS</b>In early gestational fetal skin, genes of ERK5 and MEK5 were strongly expressed, while in late gestational skin and children skin, the expression of ERK5 and MEK5 was apparently decreased (P < 0.05). In normal skin, the level of gene expression of ERK5 was lower. In proliferative hypertrophic scars, mRNA content of this gene was apparently increased. In mature scars, the content of this gene transcript was 3.2 times the normal skin. In contrast, the levels of MEK5 transcript in normal skin and hypertrophic scars of various phases showed no substantial changes (P > 0.05).</p><p><b>CONCLUSION</b>ERKS medicating signaling pathway might be involved in regulating cutaneous development at the embryonic stage and determining cutaneous structure ad function. The increase of gene transcription of ERK5 and MEK5 in younger fetal skin might be a reason for rapid proliferation of the skin cells and scraless healing of skin. The activation of ERK5 gene expression in hypertrophic scars versus normal skin might be one of the mechanisms controlling the formation of hypertrophic scars, in which the role of MEK5 needed to be further studied.</p>

Relationship between epithelial-immunologic cells transdifferentiation and pseudoepitheliomatous granuloma lesion / 中华外科杂志

<p><b>OBJECTIVE</b>Inappropriate treatment at early stage of wound could result in the formation of pseudoepitheliomatous granuloma (PEG). The correlation of abnormal transdifferentiation of epithelial cells to immunologic cells and the occurrence of PEG lesion was investigated.</p><p><b>METHODS</b>Morphological change of epithelial tissue was observed with histopathology in 11 specimens of PEG lesions and 6 specimens of normal skins from PEG edge (PEG-N) from 11 patients with damaged skin. The expression characteristics and distribution of pan-cytokeratin (CKp), IV type collagen, laminin (LM), epithelial cadherin (E-Cad), beta-catenin (beta-Cat), focal adhesion kinase (FAK), stem cell factor (SCF) and its receptor-c-Kit, proliferating cell nuclear antigen(PCNA), and cluster of differentiation-14 (CD14), CD68 and mast cell tryptase (MCT) in PEG were detected with the immunohistochemical and the indirect immunofluorescent double-staining.</p><p><b>RESULTS</b>In comparison with PEG-N, epithelial tissue take on squamous metaplasia, and stroma was infiltrated with intensive microvessels and inflammatory cells in the PEG lesion. Poor epithelial basal layer constitution, basal polarization, and migration of basal cells to stroma could be observed. In the ultrastructure, the loose intercellular junction of basal cells and the increased nucleus/cytoplasm ratio and intercellular space could be observed, neonatal monocytoid cells and macrophages and mast cells as a exuviate-like manner brooded from cytoplasm of original epithelial cells and basement membrane. protein expression of CKp and E-Cad by basal cells was significantly decreased, and the IV type collagen and LM protein could not be found in basement membrane of identical locus. By contrast, the immunoreactivity of beta-Cat and FAK was apparently increased. In addition, CD14(+) monocytes, CD68(+) macrophages, MCT(+) mast cells and CD68(+)/MCT(+) cells with various size, and these cells of stronger immuno-staining of SCF, c-Kit and PCNA antigen could be found in epithelial tissue and stroma.</p><p><b>CONCLUSION</b>Epithelial cells in PEG related to wound are characteristized by transdifferentiation of epithelial cells to immunologic cells, wich may be associated with local infectious and inflammatory reaction, ultimately resulting in enhancement the ratio of beta-Cat/E-Cad signal and activation SCF-c-Kit signal pathway. The phenomena of transdifferentiation epithelial cells in the PEG lesion will help to recognize of the neoplatic immune and trauma repair mechanism.</p>

The regulation mechanisms of MMP-1,2 and TIMP-1,2 on wound healing after partial thickness scald / 中华外科杂志

<p><b>OBJECTIVE</b>To observe the changes of matrix metalloproteinase-1,2/tissue inhibitor of metalloproteinase-1,2 (MMP-1,2 and TIMP-1,2) in granulation tissue after 30% TBSA deeper partial thickness scald, and explore the regulation mechanism of MMP-2/TIMP-2 during wound healing.</p><p><b>METHODS</b>150 male Wistar rats were randomly divided into 5 groups as follows: (1) normal control (n = 6); (2) injured control group (n = 36): which is subdivided into postburn 3 h, 6 h, 1 d, 3 d, 7 d and 14 d groups, respectively; (3) BDM group (n = 36): intravenous injected of 400 mg 2,3-butanedione monoxime in each rat was done after anesthesia; (4) H7 group (n = 36): Each rat was intravenous injected of 0.2 mg 1-5-isoquinolinyl-sulfony-2-methylpiperazine after anesthesia; (5) anti-c-fos group (n = 36): Each rat was intravenous injected of 5 microg c-fos monoclony antibody after anesthesia. The immunohistochemistry staining technique and the reverse transcription polymerase chain reaction (RT-PCR) were used for detecting.</p><p><b>RESULTS</b>The expression of c-fos mRNA and protein was increased from 3 to 6 hours post-burn, and then decreased. The expression of MMP-1,2/TIMP-1,2 was delayed to 3 days post-burn compared with the expression of c-fos mRNA and protein. Treatment with BDM induced to raise c-fos mRNA and protein expression. The expression of MMP-1,2/TIMP-1,2 was also increased accordingly. However, following treatment with H7 inhibited the expression of c-fos mRNA and protein, MMP-1,2/TIMP-1,2 proteins expression decreased. Exogenous c-fos antibody could inhibit endogenous c-fos protein expression and the expression of MMP-1/TIMP-1,2 decreased, but MMP-2 has no notable changes.</p><p><b>CONCLUSIONS</b>The expression of MMP-1,2 and TIMP-1,2 has closely relation protein kinases activated signaling pathways. The expression changes of MMP-1 and TIMP-1/TIMP-2 depend on c-fos expression. Oncogenes play an important role in the change process of wound matrix degradation and remodeling.</p>

Effects of basic fibroblast growth factor on the matrix metalloproteinase-2,7 and its tissue inhibitor on deep partial thickness burn wounds in rats / 中华烧伤杂志

<p><b>OBJECTIVE</b>To observe the changes in matrix metalloproteinase-2,7 (MMP-2,7) and tissue inhibitor of metalloproteinase-2 (TIMP-2) in deep partial thickness burn during the process of wound healing, and the effects of bFGF on wound healing.</p><p><b>METHODS</b>The rats inflicted by 30% TBSA deep partial thickness burn were randomly divided into simple scald and bFGF treatment groups. Biopsies from wound skin were harvested at 3 and 6PBHs and 1, 3, 7, 14 PBDs for the detection of the epithelialization rate and collagen content. The above indices were also detected in the skin of another 6 normal rats as normal control.</p><p><b>RESULTS</b>(1) The epithelialization rate in bFGF treatment group was higher than that in simple scald group during 3PBH to 14 PBD. (2) The collagen contents in both bFGF treatment group and simple scald group were continually decreased during 3 PBH to 3 PBD, and increased from 7 to 14 PBD, but still lower than that in normal control (P < 0.05). (3) The expression of MMP-2,7 and TIMP-2 in simple scald group enhanced from 1 to 14 PBD, and peaked on 7 PBD. (4) The expression of MMP-2,7 in bFGF treatment group was similar to that in simple scald group from 3 to 6 PBH, while the expressions of MMP-2,7 and TIMP-2 was higher than those in simple scald group from 1 to 14 PBD.</p><p><b>CONCLUSION</b>The collagen deposition would be affected by the activities of extracellular matrix in scald wound in rats. Changes in MMP-2,7 and TIMP-2 expressions were an important process of wound repair, which was closely related to the acceleration of wound healing by the application of bFGF.</p>

A comparative study of EGFR and FGFR-2 expression in fetal and adult skin / 中华整形外科杂志

<p><b>OBJECTIVE</b>To observe the expression characteristics of EGFR and FGFR-2 in normal skin from fetal and adult, and attempted to probe the molecule mechanism of fetal scarless healing.</p><p><b>METHODS</b>The skin samples of fetal and adult were taken from abortive fetus of obstetrics unit and donor site of plastic operation patients in our burn unit, respectively. EGFR and FGFR-2 were used as the biochemical markers for reparative cells. Immunohistochemistry staining technique was employed to determine the expressive levels of different epithelial cells markers.</p><p><b>RESULTS</b>There were EGFR and FGFR-2 antibody positive cells in normal skin from fetal and adult, but the expressive levels of EGFR and FGFR-2 protein had apparent difference, with the time of fetation increasing, the EGFR and FGFR-2 positive expression rate became stronger gradually. The number of FGFR-2 antibody positive cells found in adult skin was much more than that in fetal skin.</p><p><b>CONCLUSION</b>There were the inherent differences of EGFR and FGFR-2 antibody immunohistochemistry staining in cells of adult and fetal skin. which may be an essential facet of fetal scarless healing.</p>

Study on the location and the expression characteristics of epidermal stem cells in normal adult skin and scar tissue / 中华烧伤杂志

<p><b>OBJECTIVE</b>To investigate the different location and the expression characteristics of epidermal stem cells in normal adult skin and scar tissue.</p><p><b>METHODS</b>Skin tissue specimens were harvested from the corresponding sites from 6 healthy volunteers and from scar tissue of 6 patients 1 year after major deep burn. beta1 integrin and keratin 19 (K19) were employed as the biochemical markers for stem cells and transit amplifying cells identification and keratin 14 (K14) and keratin 10 (K10) as markers for post-mitotic cells and terminally differentiated cells respectively. Integrin and keratin were determined by Elivision two-step immunohistochemistry.</p><p><b>RESULTS</b>The expression of beta1 integrin and the K19 positive cell count in the epithelial basal layers of scar tissue were evidently decreased and weakened than those in normal adult healthy skin. Furthermore, the positive cells expressing K14 in epidermis of scar tissue were only located in 2 - 3 layers of basal epidermis, and their number was much less than that in normal adult skin. Whereas the cells positively expressing K10 were distributed wider in area than that in normal healthy skin. The epidermal stem cells and transit amplifying cells in scar epidermis were much less in number than that in normal skin. The differentiation process of scar epidermal stem cells was different from that of normal skin. And the proportions of post-mitotic cells and terminally differentiated cells were abnormal.</p><p><b>CONCLUSION</b>The results indicated that the self-renewal ability of the scar epidermis was decreased, and the differentiation process of it was in disorder, which may be a reason for the abnormality of structure and function of the epidermis in scar, and a reason for the decreased ability of wound healing of scar tissue.</p>

An experimental study on the differentiation of bone marrow mesenchymal stem cells into vascular endothelial cells / 中华烧伤杂志

<p><b>OBJECTIVE</b>To investigate the feasibility of differentiation of bone marrow mesenchymal stem cells (MSCs) into vascular endothelial cells and the mechanism of its involvement in wound healing.</p><p><b>METHODS</b>Porcine MSCs were harvested from porcine marrow, and they were isolated and purified by density gradient centrifugation. After being cultured and amplified in vitro, the MSCs were labelled with BrdU (5-bromodeoxy-uridine). Full skin loss wound was created on the back of the mini-swine whose bone marrow was obtained. The labelled MSCs with fibrin glue as the vector were regrafted back to the donor animal wound. The wound tissue specimens were harvested at 2, 4, 6, 8 and 12 post-operation weeks and were immunohistochemically stained by BrdU and factor VIII (FVIII) for comparative study.</p><p><b>RESULTS</b>Most BrdU positive cells aggregated around small blood vessels in the granulation tissue of the wounds. Only individual vascular endothelial cells were BrdU positive. There was FVIII expression in the cytoplasm of BrdU positive cells.</p><p><b>CONCLUSION</b>MSCs were closely correlated with the formation of small blood vessels in granulation tissue during wound healing process. The porcine MSCs possessed the potential to differentiate into vascular entoehelial cells and to participate in wound healing under the micro-enviroment of the wound.</p>