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BACKGROUND: Based on our previous researches in mechanism studies and clinical applications of human hair keratin (HHK), a new concept "in vivol in situ tissue engineering" has been proposed. Under the guidance of this theory, a scaffold of HHK-collagan sponge (inner layer) combined with poly (2-hydroxyethyl methacrylate) (PHEMA) (outer layer as a drug delivery carrier) would be developed to investigate its feasibility to be as a dermal dressing. OBJECTIVE: To develop a scaffold composed of HHK-collagan sponge (inner layer) combined with PHEMA film containing polydatin(PD)(outer layer as a drug delivery carrier) and to evaluate the therapeutic efficacy of the HHK-collagen sponge-PHEMA/PD complex on burn wound healing. DESIGN, TIME AND SETTING: A randomized controlled animal experiment was performed at the Department of Histology and Embryology, Southern Medical University between March and December 2005. MATERIALS: Burn was induced in 15 male Sprague-Dawiey (SD) rats, Rat models of burn were evenly randomized to 3 groups: experimental, positive control, and negative control. METHODS: ①HHK-collagen sponge was prepared through combination of a HHK meshwork (1mm × 1 mm in size for each grid) made up of three components (determined according to biochemical procedures of various degrees, i.e., light, medial, and severe) at a ratio of 4:3:3 with primary collagen sponge extracted from bovine tendons in a mould. Sponge film (used as inner layer dressing) was made by vacuum freeze-drying. ② PHEMA was prepared by polymerization. Than PD was added to prepare PHEMNPD film (used as outer layer dressing).③ Degree Ⅱ burn wound models were established in SD rats by scalding, Superficial necrotic tissue was removed from burn wounds at postnatal 3 days and leave the denatured dermis remained. The wounds were either covered with human HHK-collagen- PHEMNPD complex in the experimental group, or with glutaraldehyde-treated porcine skin in the positive control group, and sterile absorbent gauze was used in the negative control group. MAIN OUTCOME MEASURES: ① Complete epithelization was taken as the standards, and at postoperative 7, 14, and 21 days, wound healing was respectively calculated. ② At postoperative 1, 2, 4, 6, and 8 weeks, the whole wound surface and its peripheral tissue were dissected for observing granulation tissue growing under an optical microscope and detecting the collagen fiber and elastic fiber in the newly formed tissue by immunohistochemical staining. RESULTS: ① Gross observation results revealed that in the experimental group, the volume of the diffusate under the ideal moisture was less compared with the positive control group; the healing time was slightly shorter in both the experimental group and the positive control group than in the negative control group (P= 0.000); At postoperative 7, 14, and 21 days, the healing rate was higher in the experimental and positive control groups than in the negative control group (P=0.000), in addition, the experimental group exhibited higher healing rate than the positive control group at postoperative 14 days ( P ＜ 0.05). ②Optical microscope results showed that at postoperative 2 weeks, a small quantity of collagen fibers were found in the wound granulation tissue in all 3 groups, in particular in the experimental group. Immunohistochemical staining results regarding collagen protein and elastin revealed that at postoperative 2 weeks, both the fine strip-like type Ⅰ collagen fibers and a few silk-like elastic fibers were stained yellowish-brown in the dermal matrix in the experimental group, which were weakly positive in the positive control group, while there was no elastin detectable in the negative control group; at postoperative 8 weeks, burn wounds in all the 3 groups werefully recovered. Remodeling of collagen fibers was more obvious in the experimental and positive control groups than in thenegative control group, while the tendency to scar formation with derangement of epithelial cells and collagen fibers in dermis was more prominent in the negative control group than in the remaining two groups.CONCLUSION: HHK-collagen sponge-PHEMA/PD complex may be a new burn dressing via in vivo construction of tissueengineered epidermis, in which PHEMA may be a feasible drug-delivery carrier.
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OBJECTIVE: Inter-oocyte cytoplasmic transfer has been performed recently as a means to improve the outcome of assisted reproductive methods and becomes a hotspot. To overview the recent advances and problems in this area in this article, and put forth a new concept of reverse cloning technique based on ooplasmic transfer. DATA SOURCES: An online search of PUBMED was undertaken to identify correlative English articles published from January 1998 to December 2006 by using the keywords of "ooplasmic transfer, mitochondria heteroplasmy,animal cloning". Meanwhile, relevant Chinese articles published between January 1998 and December 2005 were searched in Wanfang database with the same keywords in Chinese. STUDY SELECTION:The articles were primarily screened, and the citations at the end of each reference were reviewed. Inclusion criteria: The articles related to the research of ooplasmic transfer, cloning or reverse cloning. Exclusion criteria: Repetitive studies or Meta analytical articles. DATA EXTRACTION:Totally 126 relevant articles were collected, and 31 of them were in accordance with the inclusion criteria, while 95 of them were excluded because of repetition or obsolescence. In 31 articles met the criteria, 17 studies were relevant to the ooplasmic transfer, 13 studies related to animal cloning and one study related to reverse cloning. DATA SYNTHESIS: Exciting advances have been achieved in human gamete biology, and ooplasmic transfer have been widely applied. Many studies indicated that mitochondria in oocytoplasm affected obviously the fertilization of the oocytes and early embryo development. Lots of experiments have been done on the interaction of cytoplasm and nucleus as well as the mechanism of genomic reprogramming to further the investigation. Meanwhile, the techniques are progressed. Cloning techniques is excellent in application prospect, while there are some problems at the same time. Therefore the concept of reverse cloning technique was proposed based on ooplasmic transfer. CONCLUSION: The mitochondria in oocytoplasm has remarkable effect on the fertilization of the oocytes and early embryonic development. However, ooplasm transfer can lead to mitochondria DNA heteroplasmy, and more work must be done on potential problems. The reverse cloning technique based on the ooplasmic transfer has a good prospect.