We have reported that high-hydrostatic-pressure (HHP) technology is safe and useful for producing various kinds of decellularized tissue. However, the preparation of decellularized or inactivated skin using HHP has not been reported. The objective of this study was thus to prepare inactivated skin from human skin using HHP, and to explore the appropriate conditions of pressurization to inactivate skin that can be used for skin reconstruction. Human skin samples of 8 mm in diameter were packed in bags filled with normal saline solution (NSS) or distilled water (DW), and then pressurized at 0, 100, 150, 200 and 1000 MPa for 10 minutes. The viability of skin after HHP was evaluated using WST-8 assay. Outgrowth cells from pressurized skin and the viability of pressurized skin after cultivation for 14 days were also evaluated. The pressurized skin was subjected to histological evaluation using hematoxylin and eosin staining, scanning electron microscopy (SEM), immunohistochemical staining of type IV collagen for the basement membrane of epidermis and capillaries, and immunohistochemical staining of von Willebrand factor (vWF) for capillaries. Then, human cultured epidermis (CE) was applied on the pressurized skin and implanted into the subcutis of nude mice; specimens were subsequently obtained 14 days after implantation. Skin samples pressurized at more than 200 MPa were inactivated in both NSS and DW. The basement membrane and capillaries remained intact in all groups according to histological and immunohistological evaluations, and collagen fibers showed no apparent damage by SEM. CE took on skin pressurized at 150 and 200 MPa after implantation, whereas it did not take on skin pressurized at 1000 MPa. These results indicate that human skin could be inactivated after pressurization at more than 200 MPa, but skin pressurized at 1000 MPa had some damage to the dermis that prevented the taking of CE. Therefore, pressurization at 200 MPa is optimal for preparing inactivated skin that can be used for skin reconstruction.

Platelet-rich plasma (PRP) contains numerous growth factors to promote wound healing and angiogenesis. The objective of this study was to explore the efficacy of biodegradable gelatin hydrogel impregnated with PRP releasate (PRPr) in the wound healing process compared with the single application of PRPr prepared from mouse PRP centrifuged by a double-spin method. Gelatin hydrogel disks with an isoelectric point of 5.0 were used in this study. A total of 180 mice (n = 45/group) were randomly assigned to the following 4 experimental groups: control group, biodegradable gelatin hydrogel group, PRPr group and gelatin hydrogel impregnated with PRPr (PRPrG) group. Wound area and epithelialization were compared on days 1, 5, 7, 14 and 21 post-wounding. After complete epithelialization, wound contraction was also evaluated. Neovascularization using immunohistochemical staining of von Willebrand factor was analyzed on day 14. The wound area of PRPrG on days 5, 7 and 14 was smaller than that in the other groups (p < 0.01). The epithelialization lengths of PRPrG on days 7 and 14 were significantly longer than the others (p < 0.01). The capillary formation of PRPrG was also superior to those in all other groups on day 14. On day 21, all wounds were completely epithelialized and PRPrG prevented wound contraction the most. It is concluded that the sustained-release system of gelatin impregnated with PRPr can stimulate angiogenesis and accelerate wound healing compared with the single application of PRP.

Giant congenital melanocytic nevi are intractable lesions associated with a risk of melanoma. High hydrostatic pressure (HHP) technology is a safe physical method for producing decellularized tissues without chemicals. We have reported that HHP can inactivate cells present in various tissues without damaging the native extracellular matrix (ECM). The objectives of this study were to inactivate human nevus tissue using HHP and to explore the possibility of reconstructing skin using inactivated nevus in combination with cultured epidermis (CE). Human nevus specimens 8 mm in diameter were pressurized by HHP at 100, 200, 500, and 1000 MPa for 10 min. The viability of specimens just after HHP, outgrowth of cells, and viability after cultivation were evaluated to confirm the inactivation by HHP. Histological evaluation using hematoxylin-eosin staining and immunohistochemical staining for type IV collagen was performed to detect damage to the ECM of the nevus. The pressurized nevus was implanted into the subcutis of nude mice for 6 months to evaluate the retention of human cells. Then, human CE was applied on the pressurized nevus and implanted into the subcutis of nude mice. The viability of pressurized nevus was not detected just after HHP and after cultivation, and outgrowth of fibroblasts was not observed in the 200, 500, and 1000 MPa groups. Human cells were not observed after 6 months of implantation in these groups. No apparent damage to the ECM was detected in all groups; however, CE took on nevus in the 200 and 500 MPa groups, but not in the 1000 MPa group. These results indicate that human nevus tissue was inactivated by HHP at more than 200 MPa; however, HHP at 1000 MPa might cause damage that prevents the take of CE. In conclusion, all cells in nevus specimens were inactivated after HHP at more than 200 MPa and this inactivated nevus could be used as autologous dermis for covering full-thickness skin defects after nevus removal. HHP between 200 and 500 MPa will be optimal to reconstruct skin in combination with cultured epidermal autograft without damage to the ECM.

One-hundred-ninety elderly (mean age 75.8 years), disabled patients at risk of being institutionalized due to stroke, acute medical problems, dementia, and other illnesses completed a rehabilitation program at the Geriatric Rehabilitation Unit (GRU). A retrospective study to measure possible impacts of rehabilitation on functional status and placement at discharge from the GRU and on living situation 1 year postdischarge showed an improvement of functional status after rehabilitation in nearly all patients. By discharge, the number of patients with partial or total activities of daily living (ADL) independence increased from 87 to 173, ambulatory patients increased from 42 to 127, continent patients from 89 to 141 and mentally clear patients from 40 to 91. Higher ratings in these parameters at discharge were associated with better placement and significantly lower mortality 1 year postdischarge. Patients placed in noninstitutional settings had a lower (21%) mortality rate when compared with patients placed in nursing homes (mortality 45%). The GRU program is clearly associated with improved outcome of care.