| Throughout the treatment for burn patients, wound healing is an important part of the treatment process of burns. And it also directly related to the patient’s survival rate and the quality of life after wound healing. The problem of how limited donor site skin can be used to cover large surface wounds is still a challenge for burn surgeons. After microskin grafting was clinically applied in 1986, it has successfully cured a large number of patients with severe burns and gradually become one of the main wound healing techniques for patients with extensive burns. However, there are still many problems in microskin grafting, such as the lower rate of wound healing, repeated ulceration after wound healing and the wound scarring. After comparing several approaches of wound healing, our research group proposed an approach of wound healing with mixed transplantation of autologous and allogeneic microskin grafting. And we proved it is effective to improve early coverage of the wound with a limited skin source on wound healing in a rat model of acute skin defect. Meanwhile, the composite grafting of particulate acellular dermal matrix(PADM) and thin layer of skin, which had been independently developed by our group, was performed on the wounds of full thickness skin defect in rats. The results showed that it could effectively improve the quality of wound healing after skin graft, providing experimental foundation for improvement in the quality of wound healing after mixed microskin grafting with dermal matrix.So we expect the mixed grafting of autogeneic and allogeneic microskin had an obvious effect of promoting the wound healing which could cover the wound at the early stage of wound healing. The PADM and the dermal tissue in allogeneic microskin could significantly improve the wound healing and promote the collagen maturation on the wounds, thereby inducing the formation of the basement membrane, as well as improving the skin quality of wound healing after the microskin graft.Objective:1. To summarize the clinical experience by analyzing the clinical data of autologous microskin grafting which was performed in patients with extensive deep burns, and to provide objective parameters for the development of clinical guidline of autologous microskin grafting technique.2. Through clinical prospective study on wound healing of deep burns with mixed transplantation of autologous and allogeneic microskin grafts, to assess the healing condition of wounds in the early stage and the quality of healed wounds in short and long term by observing the wound healing rate, scarring and histological changes, seeking a new wound repair approach for the patients with extensive deep burns.3. To add the PADM into the mixed microskin grafting on the full thickness skin defect model in rats, the rate and the quality of wound healing were observed. It will provide the experimental data for improving mixed microskin grafting in its clinical practice.Materials and methods:1. The autologous microskin grafting was performed in 214 burn patients with extensive deep wounds during the period of November 2002~August 2013 at our burn centre. Their clinical data were collected, and were sorted out into 139 cases with a complete data of wound healing, 115 cases with short term data after healing, and 73 cases with the long term quality of healed wounds. The effects of various factors in therapeutic process on the postoperative mortality, the healing condition of wounds in the early stage and the quality of healed wounds in short and long term were analyzed, through self control.2. The participants were strictly selected by rigid inclusion criteria and exclusion criteria. After enrollment, the corresponding deep wounds on bilateral limbs(or site) were selected and designed as self control according to the same patient’s wounds and were randomly divided into experimental and control groups or sites.(1)The limb was repaired by autogeneic microskin grafting;(2)The other was by mixed graft. Both groups have the same ratio of recipient site to allogeneic donor site surface area,autologous and allogeneic microskin have samer in the group of mixed groups. Through postoperative observation on the general condition, wound healing and hypertrophic scar were assessed after operation every week, its effect of promoting wound healing was studied by methods such as tissue sections. After patient consent, we would taking a small tissue samples for histological examination in reoperation. The structure of collagen bundles and the condition of epidermis were observed by HE staining and Masson staining. Laminin and type Ⅳ collagen in basement membrane was observed by immunohistochemistry examination.3. PADM combined with an optimal mixed proportion of auto and allo microskin on the full thickness skin defect model in rats:Dorsal full thickness skin samples from male SD rats were used for preparing PADM. Male Wistare rats served as alloskin donors. Forty eight female SD rats with full thickness skin defect were enrolled in the study: GroupⅠ(n=12, with autogenous microskin graft at area expansion rate of 10:1); GroupⅡ(n=12, with the PADM and autogenous microskin graft at area expansion rate of 10:1, respectively); Group Ⅲ(n=12, with mixed of autogenous and allogenous microskin graft at area expansion rate of 10:1, respectively); GroupⅣ(n=12,with the PADM into the mixed of autogenous and allogenous microskin graft at area expansion rate of 10:1,respectively). The wound healing rate and wound contraction rate were observed at the 2, 3 and 4 weeks postgraft in 24 rats(every group n= 6). The other rats were biopsied for HE staining, Masson staining, laminin and typeⅣ collagen immunohistochemistry examination on the 2, 3 and 4 weeks postgraft. The distribution and their ratio of typeⅠ/Ⅲ collagen were also examined by sirsus red staining on the 8 weeks postgraft.Result:1. Retrospective analysis in burn patients with autogeneic microskin grafting showed that:(1) the prognosis of patients with extensive deep burns was related to the burned surface area and inhalation injury;(2) the mortality in patients with escharectomy within 1 week and beyond 1 week after the injury was 16.1% and 32.1%(P<0.05), respectively;(3) according to analysis on the wound healing rates of 377 operative sites, the healing rate reached the higher value of(77.23 ±19.91)% when the area expansion ratio was 5~10:1, having an obvious advantage over that with an expansion ratio less than 5:1(P<0.05); at the same healing rate, the healing was as fast as(1.84 ±0.76)%/d when the expansion ratio was 5~10:1, significantly faster than that of(1.53 ± 0.51)%/d when the expansion rate was greater than 10:1(P<0.05);(4) In terms of the short term healing quality of 307 sites wounds, repeated skin ulceration was a common phenomenon after microskin grafting;(5)The long term quality of healed wounds in 189 sites confirmed the obvious correlation between scar hyperplasia and wound healing rates(χ2=11.69, P<0.05).2. The prospective clinical study on the mixed graft of autogeneic and allogeneic microskin in 6 cases of patients with severe burns showed that:(1) The wound healing rates of mixed graft group at 5 and 6 weeks postoperation were(53.57 ±30.78)% and(66.14 ± 30.15)%, respectively, significantly higher than those of the only autogeneic microskin graft group as(36.14± 23.96)% and(46.57±25.8)%(P<0.05).(2) The speed of healing in the mixed graft group was(1.50±0.65)% /d, significantly higher than that of the autogeneic microskin group as(1.06±0.50)% /d(P<0.05).(3) Four samples from two patients at the 7 weeks postgraft and two samples from one patient at the 18 months postgraft were observed by HE staining, Masson staining and immunohistochemical staining. HE staining showed that the epidermis in the healed wounds was thickening apparently and well differentiated with more skin rete pegs in the mixed group at 7 weeks postgraft than those in the autologous skin grafting group. However, there is no significant difference between the two groups at 18 months postgraft, the epidermis and epidermal dermal connection were well developed. Masson staining revealed that the collagen bundles were slender in both groups at the 7 weeks postgraft. The collagen bundles at 18 months postgraft became thick than before, their arrangement became regular and compact. The light positive immunohistochemical staining pattern for typeⅣ collagen was seen in epidermis dermal conjunction in both the two groups at the 7 weeks postgraft. The typeⅣ collagen staining in mixed group, however, was slight strong and more continuous and contact than that in autogeneic microskin graft group. The strong positive staining pattern for typeⅣ collagen was seen in epidermis dermal conjunction in both the two groups at 18 months postgraft. Laminin staining had the similar results to the typeⅣ collagen staining in both groups.3. Animal experiments showed that:(1) The mixed transplantation of autogeneic and allogeneic microskin could cover considerable part of the wounds in early stage. The healing rates in groupⅢ and Ⅳ at 2 weeks after surgery were(43.70±21.25)% and(59.39±19.46)%, respectively, significantly higher than those in the groupⅠand group Ⅱ as(20.59±6.63)% and(23.37±5.90)%(P<0.05). The healing rates of groupⅢ and Ⅳ at the 3 and 4 weeks after operation were also significantly higher than that of other two groups.(2) As for the wound contraction, all groups showed remarkable contraction at the 2, 3 and 4 weeks postgraft,.but the groupⅡand Ⅳ with adding PADM had slightly lower wound contraction rate than in other groups. However,only the contraction rate between groupⅣ(74.57±1.99)% and groupⅠ(79.49±5.70)% at 4 weeks postgraft was statistically significant.(3) In routine HE staining, different degrees of epidermis hyperplasia were found in all groups at 2 weeks postgraft with relative thinner epidermis in groupⅠ and more obvious hyperplasia in groupⅢ and Ⅳ with rete ridges formation. Masson staining revealed that allogeneic dermis or/and PADM existed in wound dermis of groupⅡand Ⅳ at 2 weeks postgraft, the collagen bundles were relatively loose and thick. While the collagen bundles in groupⅠ were thin and compact. At 3 weeks postgraft, different degrees of collagen hyperplasia appeared in all groups. At 4 weeks postgraft, collagen arrangement in the mixed group was loose and approached maturity, among which groupⅣ enjoyed the best result, collagen bundles were thin and arranged densely. Laminin staining was expressed in all groups 2 weeks after the transplantation. And groupⅣ had a strong, complete and continuous expression at the basement membrane, wile weaker continuous expression can be seen in groupⅡ and Ⅲ, only intermittent positive expression was found in groupⅠ. At 3 weeks post graft, relatively strong continuous laminin expression can be seen in group Ⅱ, Ⅲ and Ⅳ while the expression was weaker in groupⅠ. At 4 weeks postgraft, the area of basement membrane in each group has continuously positive expression. The ratios of type Ⅰ to type Ⅲ collagen content in these four groups were(2.23±0.27)%,(4.57±0.24)%,(3.95±0.63)% and(5.08±0.47)%, respectively, and that in the normal skin was(6.08±0.50)%. All the experimental groups were obviously different from normal skin(P<0.05). Compared with groupⅠ, the other three groups were different(P<0.05). There was also an significant difference between the groupⅢ and groupⅣ.Conclusions:1. The patients underwent aggressive excisions can significantly reduce the mortality; appropriate area extension ratio(5: 1 to 10: 1) produce a higher healing rate and speed; improving the wound healing rate of the primary phase would effectively promote the patients’ long term healing quality.2. The autogeneic and allogeneic microskin grafting had an obvious effect of promoting the wound healing in patients with deep burns, which could cover the wound at the early stage of wound healing. The allogeneic microskin can facilitate the reconstruction of basement membrane, helping the establishment of the epidermal dermal junction and increasing the quality of wound healing after the microskin grafting.3. The mixed grafting of autogeneic and allogeneic microskin had an obvious effect of promoting the wound healing in the acute skin defect wound of rats. The PADM and the dermal tissue in allogeneic microskin could significantly improve the wound healing and promoting the collagen maturation on the wounds, thereby inducing the formation of the basement membrane, as well as improving the skin quality of wound healing after the microskin graft. |
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