[Biological dressing with human hair keratin-collagen sponge-poly 2-hydroxyethyl methacrylate composite promotes burn wound healing in SD rats].

OBJECTIVE: To develop a composite material containing human hair keratin (HHK), collagen sponge (inner layer) and poly 2-hydroxyethyl methacrylate (PHEMA) film that allows sustained release of polydatin and test its effect as a biological dressing in promoting burn wound healing in SD rats. METHODS: Three HHK materials with fast, moderate, and low degradation rates were mixed at the ratio of 4:3:3 to prepare a reticular structure, which was processed into a composite material with bovine tendon-derived collagen sponge, and further complexed with HEMA film containing PD prepared by polymerization. Degree II burn wound was induced in SD rats by scalding and within postburn day 2-5, the wounds were cleansed and covered with the composite material or with glutaraldehyde-treated porcine skin (positive control). At week 1, 2, 4, 6 and 8 following wound dressing, 6 full-thickness skin samples were harvested from the wounds for histological observation and immunohistochemical detection of collagen and elastic fibers, and the wound healing time and healing rate were recorded. RESULTS: The prepared collagen sponge film was transparent and porous (50-300 microm in diameter) and allowed sustained PD release into normal saline within 48 h. Compared with the porcine skin, the composite material reduced exudation and maintained ideal moisture of the wound, and significantly shortened the wound healing time (P=0.000). On day 7, 14, and 21 following dressing, the composite material and porcine skin significantly increased the wound healing rate as compared with the negative control group (P=0.000), and on day 14, the composite achieved significantly greater healing rate than the porcine skin (P<0.05). CONCLUSION: HHK-collagen sponge-PHEMA/PD composite as a dressing material promotes burn wound healing in rats by allowing in vivo construction of tissue engineered epidermis. PHEMA is feasible for sustained drug delivery in this composite.

Toxicity evaluation of chicken calamus keratin conduit as a tissue-engineering scaffold biomaterial.

OBJECTIVE: To evaluate the toxicity of chicken calamus keratin (CCK) conduit as a tissue-engineered scaffold material. METHODS: The chemical composition of the leaching solution of CCK was determined by means of ultraviolet spectrometry, and the toxic effects of the solution was evaluated by skin sensitization test in rats, intracutaneous stimulation test in rabbits, acute systemic toxicity test in mice, and cytotoxicity test in L929 cells. RESULTS: The leaching solution of CCK consisted mainly of middle-molecular-weight peptides with a small quantity of macromolecular proteins. Skin sensitization test in rats showed that application of the CCK leaching solution caused no obvious skin reddening, regional edema, or skin necrosis. Intracutaneous injection of the leaching solution in rabbits did not induce obvious skin stimulation manifested by intradermal erythema or edema. In acute systemic toxic test, administration of the leaching solution in mice caused no death, organ dysfunction, cyanosis, tremor, severe peritoneal irritation, ptosis, or dyspnoea. In vitro cytotoxicity test indicated that the cell toxicity of the CCK leaching solution was approximately at 0 level. CONCLUSION: CCK contained in the treated chicken calamus easily undergoes hydrolysis to release mainly some peptides which do not induce obvious toxic effects, suggesting the safe potential applications of CCK conduit as a tissue-engineering biomaterial.

Collagen gel coating or cyclosporine A for improving histocompatibility of chicken calamus keratin.

OBJECTIVE: To improve the histocompatibility of chicken calamus keratin (CCK) graft by collagen-gel coating or using of cyclosporine A (CsA). METHODS: Thirty SD rats were equally randomized into 5 groups, and in 4 of them, CCK implantation into the bilateral erector spinae was performed on different treatment protocols. In group A, the rats received daily intraperitoneal injection of CsA (5 mg/kg) for two consecutive weeks after CCK implantation; in group B, CCK was soaked in CsA (2.5 mg/ml) solution at 4 degrees Celsius; for 48 h before grafting; in group C, CCK coated with collagen gel was grafted; and in group D, only CCK was implanted. Rats in the fifth group received only cutaneous incision as well as muscular dissection to serve as the blank control. CCK degradation and its effect on the surrounding tissues were observed at 2, 4 and 8 weeks after grafting. Immunohistochemistry was performed to identify T lymphocyte infiltration in the host tissues. RESULTS: All the rats survived the operation. Numerous macrophages, especially multinucleated giant cells occurred on the peripheral of the CCK grafts, and small degraded CCK pieces were observed in their cytoplasm. Only a few inflammatory cells were seen in the host tissues. At 2, 4 and 8 weeks after CCK implantation, only a few CD3-positive cells were found in all the groups, and in group A and B, the density of T lymphocytes was significantly lower than that in group D, and there was no significant difference between group A and the blank control group. CONCLUSIONS: CsA significantly improves the histocompatibility of CCK material, and short-term systemic CsA administration achieves the best results. Macrophages, especially multinucleated giant cells participate in CCK degradation in vivo.

[From meridians and acupoints to self-supervision and control system: a hypothesis of the 10th functional system based on anatomical studies of digitized virtual human].

Chinese acupuncture and moxibustion has been widely accepted as a useful therapeutics all over the world, but its mechanism has not been fully defined. For this purpose, a reticular framework of whole-body fascia and connective tissues has been established by means of digitized virtual human technique. The virtual acupoints represented three-dimensionally were compared with the sites for stimulation in practice of traditional Chinese medicine (TCM) acupuncture therapy. The results showed that the fascial network constituted by the connective tissues may be the anatomical basis for acupuncture therapy. We found that the acupoints were mainly located where thick connective tissues were present. In this fascial network, sensitive nerve endings, active cells and lymphatic vessels abounded in the sites with thick connective tissue, and needling at these sites induced definite biological effects. In light of biological phylogeny and embryo development, we believe that the connective tissue network may constitute a new functional system in the human body, the Self-supervision and control system (Fasciology), which provides a theoretical base for acupuncture therapy.

[Preliminary study of chicken calamus conduit as a scaffold material for tissue engineering].

OBJECTIVE: To observe the unique structural features of chicken calamus keratin (CCK) conduit as a candidate scaffold material for tissue engineering and its in vivo degradation and histocompatibility after its implantation into living tissues. METHODS: Chicken calami were taken from healthy chickens and treated through sequential, controllable physical and biochemical procedures for preparation of three types of CCK conduits, namely CCK-I (mildly treated), CCK-II (moderately treated) and CCK-III (intensely treated). Light microscopy (LM) and scanning electron microscopy (SEM) were performed for morphological observation. Each of these three types of CCK pieces (experimental group) and the untreated ones (control group) was implanted into the dorsal muscular tissue on both sides of SD rats, respectively. Routine tissue sectioning and HE stain were performed to identify the morphological changes under light microscope. Each of the CCK threads (experimental group) and the untreated chicken calamus threads (control group) was also grafted within the sciatic nerve bundles of SD rats, respectively. RESULTS: The wall of the chicken calamus was composed of 4 compact parts from inside to outside on cross sections, namely the innermost basophilic homogenous coarse line, 3-5 layers of acidophilic corneum, 60-100 layers of circular keratin tracts containing massive pigment granules, and 10-20 outmost layers of keratin tracts with only a few pigment granules. The three-dimensional surface features of chicken calamus identified by SEM, as compared with untreated chicken calamus, was characterized by loose arrangement containing horizontal and vertical keratins with obvious pores of different sizes and depths on its surface. At 8 weeks after implantation into the muscular tissue in experimental groups, the CCK grafts were degraded into thin filaments or/and dispersed pieces and fine granules with the appearance of blood vessels, which facilitated the absorption of the degradation products; at 12 weeks, the grafts were markedly degraded into tiny fragments. In the control group, in contrast, the grafts remains intact throughout the experiment. After implantation of the material into the nerve bundles, similar cell infiltration and tissue responses to the grafts were observed as compared to those occur in intramuscular grafting. The degradation products did not seem to cause nerve tissue degeneration or necrosis. CONCLUSIONS: Fresh chicken calamus is a natural tube composed of multi-layered compact keratin tracts with pigment granules and small amount of matrix, and is non-absorbable in vivo, and therefore does not favor the purpose for use directly as a candidate biological scaffold. After proper treatment, the chicken calamus becomes loosely arranged porous material, and can be degraded and absorbed in vivo without resulting in tissue degradation or necrosis, suggesting its potential for applications in tissue engineering.

Ooplasmic transfer: problems and prospects.

Cytoplasmic transfer between human oocytes, which represents a complete cytoplasmic exchange, has been performed recently as a means to improve the outcome of assisted reproduction and becomes a hotspot of researches. Many studies have indicated that mitochondria in the oocytoplasm obviously affect fertilization of the oocytes and early embryo development. However, ooplasmic transfer can lead to mitochondrial DNA heteroplasmy and the prospect of mitochondrial heteroplasmy and its potential problems necessitate further studies. The authors reviews the ooplasmic transfer, the relation between ooplasm and fertilization and embryo development, and the mitochondrial heteroplasmy. The authors also propose a new theory of "reverse cloning technique".

Preparation and bioactivity of human hair keratin-collagen sponge, a new type of dermal analogue.

OBJECTIVE: To develop a three-dimensional porous film of human hair keratin (HHK)-collagen sponge complex for use as a dermal substitute. METHODS: The three components F, B, and Z derived from healthy human hair were weaved into a meshwork and integrated with purified soluble type I collagen extracted from bovine tendons to prepare a highly porous film with vacuum freeze-drying followed by secondary cross-linking with glutaraldehyde. The film was grafted beneath the dorsal skin in 21 SD rats (experimental group), with simple collagen sponge serving as the negative control. The rats receiving surgical operation but without graft served as the blank control. The graft and its surrounding tissue were harvested on days 3, 7 and at weeks 2, 4, 6, 8, 12 after implantation for evaluation of tissue compatibility, vascularization and degradation. RESULTS: The prepared collagen sponge film was semitransparent and porous. Three to 7 days after grafting, inflammatory reaction was relieved gradually, and several fibroblasts and blood vessels were found adherent to the grafts in the experimental groups. At week 4, the wounds healed in the experimental groups, and the fibroblasts were actively secreting collagen and the film degraded obviously with the appearance of elastic fibers. At weeks 6 and 8, new collagen fibers thickened and assumed regular arrangement, and the collagen sponge films disappeared completely. In the control groups, the changes were less obvious and total HHK degradation occurred till week 12. CONCLUSION: The degradable and absorbable HHK-collagen sponge film has relatively satisfactory tissue compatibility and can accelerate wound healing by stimulating cell proliferation and vascularization, showing the potential as an optimal dermal substitute.