Texture of the skin graft, with special reference to the hydration state of the stratum corneum

The water contained in the superficial portion of the stratum corneum plays an important role in keeping the skin surface soft and smooth. In an attempt to study the true nature of the "texture" of grafted skin, in vivo water sorption-desorption tests were performed on grafts, on normal skin adjacent to grafts, and on normal skin surrounding the donor sites. The results showed that skin grafted to sites other than the face maintained a functional similarity to normal skin surrounding the donor site. However, in skin grafted to the face, the functions of the stratum corneum, as measured in terms of hydration state, increased, and this was associated with an improved water-holding capacity that probably results from the effect of abundant sebum and sweat excretion in adjacent skin. From these results it is suggested that the recipient site plays an important role in determining the difference in the function of the stratum corneum, i.e., the "texture" between grafts on the face and those on other sites.     

The Environment and the Microbial Ecology of Human Skin

Microbial flora of the skin of three human population groups representing different natural environments was examined quantitatively and qualitatively to determine whether environmental differences in temperature and humidity can influence the microbial flora of normal skin. Five anatomical skin sites - hands, back, axillae, groin, and feet - were sampled from 10 subjects working in a high-humidity, high-temperature environment, 10 subjects from a low-temperature, high-humidity environment, and 10 subjects working in a moderate-temperature and low-humidity environment. Bacterial populations were significantly larger from the back, axillae, and feet in individuals from the high-temperature and high-humidity environment as compared to the moderate-temperature, low-humidity environment. High humidity and low temperature had no significant effect on total populations, but this group showed a higher frequency of isolation of fungi, and gram-negative bacteria from the back and feet. Although there was an indication that increase in the environmental humidity could result in an increased frequency of isolation of gram-negative bacteria, there was no evidence that an increase in either temperature or humidity altered the relative proportions of gram-negative bacteria in the predominantly gram-positive microbial flora found on normal skin. It was concluded that, although climatic changes may cause fluctation in microbial populations from certain sites, they are not a major influence on the ecology of the microbial flora of normal skin in the natural environment. The variables introduced by studying individuals in their natural environment and the influence of these on the results are discussed.     

Effects of T cell frequency and graft size on transplant outcome in mice

The features that determine whether graft-reactive T lymphocytes develop into effector cells capable of mediating organ destruction are not well understood. To investigate potential factors involved in this process, we first confirmed that female recipient mice acutely rejected minor Ag-disparate male skin, but not heart transplants. Despite this difference in outcome, heart and skin transplantation induced antidonor T cell responses of similar magnitude, specificity, and cytokine profile. The heart-graft-primed T cells transiently infiltrated the graft and ultimately induced the development of chronic transplant vasculopathy. Increasing the frequency of donor-reactive T cells by presensitization or by using TCR (CD8+ antimale)-transgenic recipients did not mediate acute rejection but accelerated the pace and severity of the vasculopathy. Surprisingly, decreasing the tissue mass of the donor heart by 50% resulted in acute rejection of these smaller grafts without increasing the frequency of antidonor effector T cells in the recipients. In complementary studies, placement of one or two male skin grafts on a single recipient did not affect the frequency or cytokine profile of the induced antimale T cell repertoire. Nonetheless, the recipients of single grafts acutely rejected the transplanted skin while the recipients of two skin grafts did not. These results provide new insight into the pathogenesis of transplant vasculopathy and provide an explanation for the difference in outcome between murine skin and heart transplants by highlighting the novel concept that the efficiency of transplant-reactive T cell immunity is heavily influenced by the tissue burden it encounters at the effector stage.     

Clinical application of autologous cultured epithelia for the treatment of burn wounds and burn scars

This report presents our experience with autologous cultured human epithelia grafting on burn wounds, burn scars, and skin-graft donor sites in seven patients. Dispersed epidermal cells were cultured with 3T3 cells treated with mitomycin C. After 2 to 3 weeks, cultured epithelia (total 350 to 2250 cm2) were grafted to the wound. The results showed that cultured epithelia grafts did not take so completely compared to the meshed skin grafts used for the coverage of burn wounds. However, cultured grafts placed on aseptic wounds adhered well and showed good appearance. In the histologic findings, normal differentiation of epidermal cells was found. Cultured grafts were bordered from subepidermal granulative tissue with basement membrane. A rete ridge and the adnexal structures were absent in the specimens that adhered to the burn wounds. However, in the specimens that took on abraded wounds, a gently sloping rete ridge and elastic fibers were seen. The histologic findings showed structures resembling normal skin.     

Blood group substances, T6 antigen and heterochromatin pattern as species markers in the nude mouse/human skin model

Antibodies to blood group antigens and to human T6 antigen and the DNA-specific fluorochrome D287/170 were applied to human skin transplanted to nude mice and to the surrounding murine skin. It was found that the two epithelia maintained their characteristic, different blood group B antigen distributions and heterochromatin patterns. Furthermore, it was shown that human Langerhans' cells persisted in the grafted epidermis. During an observation period of until 48 weeks after transplantation only one of 32 grafts showed signs of invasion by murine epidermis.     

Violagabriellae variant of Staphylococcus epidermidis on normal human skin

Twenty-seven strains of a reddish violet pigment-producing variety of Staphylococcus epidermidis have been recovered from normal human skin. They closely resemble the previously unique Castellani strain, named Micrococcus violagabriellae, which was placed by K. J. Steel in S. epidermidis. These strains are classified as S. epidermidis by using the Baird-Parker schema; however, besides the pigment produced, they also differ from S. epidermidis in proteolytic activity and effect on litmus milk. The variety seems to be part of the normal flora of the human axilla.     

Effect of an abundant human skin melanosomal 66 kDa protein (MP 66) on m urine tyrosinase: Its physiological implications on melanogenesis

A single polypeptide protein (MP 66) of molecular weight 66 kDa purified to homogeneity from melanosomes of normal human skin epidermal melanocytes, was partially characterized. The isoelectric point of MP 66 is in the range of 7.3 to 7.6. This protein, which was shown to inhibit partially purified human skin tyrosinase activity at pH 6.8, also inhibits murine tyrosinase at pH 6.8. However, at pH 5.0, it stimulates murine tyrosinase activity. The physiological implications of these results are discussed.     

The cell type mediating hyperacute rejection of allogeneic mouse skin grafts.

Allogeneic murine skin grafts can be rejected hyperacutely, as white or “red” grafts, by a cell-mediated mechanism. The hyperacute rejection is transferable by sensitized, nylon adherent, θ-bearing lymph node cells draining skin graft sites.     

腋臭患者腋窝皮肤需氧微生物群的研究

对１６例女性腋患者腋窝皮肤菌群从微生态学的角度进行定性、定量分析。并将结果同２４例正常女性腋窝皮肤菌群做了比较。结果表明,美白喉杆菌及表皮葡萄球力为腋臭患者腋窝皮肤的主要菌生机制及用生态制剂等调整菌群,改善微环境,,达到防治目的有所帮助     

Tissue engineering of cultured skin substitutes

Skin replacement has been a challenging task for surgeons ever since the introduction of skin grafts by Reverdin in 1871. Recently, skin grafting has evolved from the initial autograft and allograft preparations to biosynthetic and tissue-engineered living skin replacements. This has been fostered by the dramatically improved survival rates of major burns where the availability of autologous normal skin for grafting has become one of the limiting factors. The ideal properties of a temporary and a permanent skin substitute have been well defined. Tissue-engineered skin replacements: cultured autologous keratinocyte grafts, cultured allogeneic keratinocyte grafts, autologous/allogeneic composites, acellular biological matrices, and cellular matrices including such biological substances as fibrin sealant and various types of collagen, hyaluronic acid etc. have opened new horizons to deal with such massive skin loss. In extensive burns it has been shown that skin substitution with cultured grafts can be a life-saving measure where few alternatives exist. Future research will aim to create skin substitutes with cultured epidermis that under appropriate circumstances may provide a wound cover that could be just as durable and esthetically acceptable as conventional split-thickness skin grafts. Genetic manipulation may in addition enhance the performance of such cultured skin substitutes. If cell science, molecular biology, genetic engineering, material science and clinical expertise join their efforts to develop optimized cell culture techniques and synthetic or biological matrices then further technical advances might well lead to the production of almost skin like new tissue-engineered human skin products resembling natural human skin.     

Correction of junctional epidermolysis bullosa by transplantation of genetically modified epidermal stem cells

The continuous renewal of human epidermis is sustained by stem cells contained in the epidermal basal layer and in hair follicles. Cultured keratinocyte stem cells, known as holoclones, generate sheets of epithelium used to restore severe skin, mucosal and corneal defects. Mutations in genes encoding the basement membrane component laminin 5 (LAM5) cause junctional epidermolysis bullosa (JEB), a devastating and often fatal skin adhesion disorder. Epidermal stem cells from an adult patient affected by LAM5-beta3-deficient JEB were transduced with a retroviral vector expressing LAMB3 cDNA (encoding LAM5-beta3), and used to prepare genetically corrected cultured epidermal grafts. Nine grafts were transplanted onto surgically prepared regions of the patient's legs. Engraftment was complete after 8 d. Synthesis and proper assembly of normal levels of functional LAM5 were observed, together with the development of a firmly adherent epidermis that remained stable for the duration of the follow-up (1 year) in the absence of blisters, infections, inflammation or immune response. Retroviral integration site analysis indicated that the regenerated epidermis is maintained by a defined repertoire of transduced stem cells. These data show that ex vivo gene therapy of JEB is feasible and leads to full functional correction of the disease.     

Serine proteinase activation of latent human skin collagenase

Latent and active collagenase were demonstrated following direct extraction from normal skin homogenates with 0.1M calcium chloride at 60 degrees C. 83% of the collagenase activity was in latent form and could be maximally activated with trypsin. Partial activation of the latent enzyme could also be demonstrated by incubation of the skin extract without added trypsin. This endogenous activation was inhibited by the addition of soya bean trypsin inhibitor, trasylol, di-isopropylphosphofluoridate and phenylmethanesulphonylfluoride, none of which inhibited collagenase directly. This suggests that the skin extracts contain a collagenase activating enzyme with the inhibition profile of a serine proteinase. A chymotryptic proteinase with a similar inhibition profile was extracted from normal human skin and partially purified. This enzyme activated fibroblast procollagenase derived from tissue culture of normal skin. The procollagenase was also partially activated by plasmin and chymotrypsin. This is the first demonstration of a collagenase activating enzyme in human skin and raises the possibility that collagenase activation by this mechanism may be responsible for collagen degradation in some disease processes.     

Effects of fibroblasts of different origin on long term maintenance of xenotransplanted human epidermal kerationocytes in immunodeficient mice

We examined effects of fibroblasts of different origin on long-term maintenance of xenotransplanted human epidermal keratinocytes. A suspension of cultured epidermal cells, originating from adult human trunk skin, was injected into double mutant immunodeficient (BALB/c nu/scid) mice subcutaneously, with or without cultured fibroblastic cells of different origin. At one week after transplantation, the epidermal cells generated epidermoid cysts consisting of human epidermis-like tissue. When the epidermal cells were injected alone or together with fibroblastic cells derived from human bone marrow, muscle fascia, or murine dermis, organized epidermoid cysts regressed within 6 weeks. In contrast, when the epidermal cells were injected together with human dermal fibroblasts, generated epidermoid cysts were maintained in vivo for more than 24 weeks. Histological examination showed that the reorganized epidermis, after injection of both epidermal keratinocytes and dermal fibroblasts, retained normal structures of the original epidermis during 6 to 24 weeks after transplantation. The results indicate that human dermal fibroblasts facilitate the long-term maintenance of the reorganized epidermis after xenotransplantation of cultured human epidermal keratinocytes by supporting self renewal of the human epidermal tissue in vivo.     

Vascular endothelial growth factor and Kaposi's sarcoma cells in human skin grafts.

Human cancer cells often produce tumors in animal models that incompletely reproduce the histology of the parental tumor. Kaposi's sarcoma (KS) cells, in particular, have not produced durable angiogenic lesions in animal models that resemble those of KS in humans. We investigated the contribution of transformed KS cells, vascular endothelial growth factor (VEGF), and human skin tissue on tumor development in a human skin graft/mouse model. High levels of serum VEGF (322 pg/ml) were seen in HIV-1-infected persons with KS compared with HIV-1-infected persons without KS (115 pg/ml). Human KS lesions expressed VEGF in the spindle cells. Transformed KS cells expressed the mitogenically active 121-amino acid and 165-amino acid isoforms of VEGF. Tumors induced by KS cells implanted in the SCID mice grew preferentially in human skin grafts rather than in ungrafted murine skin. Tumors induced in the presence of human skin grafts developed numerous lumens expressing alpha(v)beta(3) integrin. KS cells inoculated with neutralizing anti-VEGF antibody did not form tumors. This study supports an important role for VEGF in tumor development and shows how a human tissue can preferentially promote tumor growth.     

Protective effect of vitamin E on ischaemia-reperfusion injury in ovarian grafts

Ovarian cortical tissue cryopreservation with subsequent autografting is a potential strategy for the preservation of fertility in patients undergoing systemic chemotherapy and pelvic radiotherapy. Non-vascular implants are first subjected to a period of ischaemia before revascularization and are, therefore, vulnerable to ischaemia-reperfusion injury from reactive oxygen species. Ischaemia-reperfusion injury was investigated during the first week after surgery in murine ovarian grafts and human ovarian xenografts in mice with severe combined immune deficiency (SCID) by measuring total lipid peroxides and malondialdehyde concentrations with a colorometric assay. The effects of administering an antioxidant, vitamin E, on these concentrations were also tested. Products of lipid peroxidation were higher in non-supplemented murine autografts compared with control ovaries (P < 0.05), and were significantly reduced on day 3 by vitamin E administration (P < 0.05). Similarly, in human xenografts, there was a significant reduction in lipid peroxidation with vitamin E administration. These results correspond to a significantly greater total follicle survival in the murine grafts of the supplemented group (45 versus 72%; P < 0.05). They suggest that antioxidant treatment improves the survival of follicles in ovarian grafts by reducing ischaemia-reperfusion injury.     

Survival of Skin Graft between Transgenic Cloned Dogs and Non-Transgenic Cloned Dogs

Whereas it has been assumed that genetically modified tissues or cells derived from somatic cell nuclear transfer (SCNT) should be accepted by a host of the same species, their immune compatibility has not been extensively explored. To identify acceptance of SCNT-derived cells or tissues, skin grafts were performed between cloned dogs that were identical except for their mitochondrial DNA (mtDNA) haplotypes and foreign gene. We showed here that differences in mtDNA haplotypes and genetic modification did not elicit immune responses in these dogs: 1) skin tissues from genetically-modified cloned dogs were successfully transplanted into genetically-modified cloned dogs with different mtDNA haplotype under three successive grafts over 63 days; and 2) non-transgenic cloned tissues were accepted into transgenic cloned syngeneic recipients with different mtDNA haplotypes and vice versa under two successive grafts over 63 days. In addition, expression of the inserted gene was maintained, being functional without eliciting graft rejection. In conclusion, these results show that transplanting genetically-modified tissues into normal, syngeneic or genetically-modified recipient dogs with different mtDNA haplotypes do not elicit skin graft rejection or affect expression of the inserted gene. Therefore, therapeutically valuable tissue derived from SCNT with genetic modification might be used safely in clinical applications for patients with diseased tissues.     

淤积性皮炎与皮肤菌群关系的研究

目的：探讨淤积性皮炎与皮肤菌群之间的关系。方法：对21例小腿淤积性皮炎患者皮损，邻近皮肤及鼻孔的皮肤菌群进行检测。并将结果同20例正常人小腿皮肤及鼻孔菌群做了比较，其数据进行统计学处理。结果：金黄色葡萄球菌为淤积性皮炎患者皮肤的主要菌种，其分离率及密度均高于正常对照组。结论：金黄色葡萄球菌可能是淤积性皮炎发生的一个重要因素。     

正常菌群对常见致病菌拮抗作用的研究

生物拮抗作用是微生物种群关系研究的一个侧面,实验中我们使用从皮肤上分离的常住菌——痤疮丙酸杆菌(A14-1)和表皮葡萄球菌(F65)对常见致病菌如金黄色葡萄球菌(C189)、绿脓杆菌(C8514)和埃希氏大肠杆菌(C1356)的体外拮抗试验表明,它们具有明显的拮抗作用,拮抗作用一般从48小时开始,72小时明显。而皮肤常住菌间无拮抗作用,相反它们还有协同作用,这对于皮肤的自净以及维持皮肤的微生态平衡具有重要的作用。此外,实验中还进行了乳杆菌和双歧杆菌等正常菌群分离株与上述常见致病菌的体外拮抗试验,结果表明也     

慢性湿疹患者皮损、鼻粘膜需氧微生物群的定位、定量研究

本文首次应用定位、定量方法对23例小腿部慢性湿疹患者皮损、邻近皮肤及鼻孔的菌群,从微生态学的角度进行探讨,并将结果同24例正常人小腿皮肤及鼻孔的菌群做了比较。其结果表明:金黄色葡萄球菌为皮损及邻近皮肤的主要菌种,并且其分离率及密度有按正常皮肤、邻近皮肤、皮损依次增高现象;类白喉杆菌的数量在皮损部位明显低于正常皮肤与邻近皮肤;表皮葡萄球菌分离率皮损及邻近皮肤均明显降低。本组结果同正常对照组相比其皮肤微生物群菌谱虽无不同,但其数量的变异是显著的,这对探讨湿疹的病因及用生态制剂等调整皮肤菌群使其恢复微生态平衡达到防治目的有所裨益。