bluuebunny/crossref_metadata_2025 huggingface dataset:
Dermal fibroblasts from different layers of human skin are heterogeneous in expression of collagenase and types I and III procollagen mRNA
Authors: Ali Bahar Mohammad , Bauer Barbara , Tredget Edward E. , Ghahary Aziz
Published: February 2004

We have previously shown that the expression of collagenase mRNA and activity are suppressed in fibroblasts derived from postburn hypertrophic scar. Although it is known that differential synthesis of collagen and collagenase in postburn hypertrophic scar is one of the main reasons for the excessive accumulation of collagen, it is not clear why the expression of collagenase in hypertrophic scar fibroblasts is suppressed relative to that in normal fibroblasts. In this study, we hypothesized that dermal fibroblasts from deeper layers of skin, which normally migrate toward the wound site, have a different capacity to express key extracellular matrix proteins such as collagenase and types I and III procollagen. To test this hypothesis, we established four different pairs of hypertrophic and site‐matched normal cell strains from four different patients with postburn hypertrophic scarring. In another set of experiments, ten different cell strains from two normal human skin samples that were horizontally sectioned into five different layers (layers 1–5 correspond to upper to deeper layers, respectively) were established in culture. Cells at the same passages were harvested, total RNA was extracted, and Northern analysis was conducted to determine the level of collagenase and types I and III procollagen mRNA expression in each cell strain. The results of Northern blot analysis showed two transcripts each for the pro α1(I) collagen chain (5.8 and 4.8 kb) and for the pro α1(III) collagen chain (5.4 and 4.8 kb) in all cell strains examined. The intensity of the pro α1(I) chain of type I procollagen mRNA varied, ranging from 50.8 to 137.1 and 44.4 to 131.5 densitometry units among either normal or hypertrophic scar cells examined, respectively. Similarly, the relative quantity of type III procollagen transcript also varied, ranging from 10.4 to 91.1 and 28.7 to 116.1 among normal and hypertrophic scar cells, respectively. When five different skin layers from each tissue sample were evaluated for functional cell heterogeneity, the results showed a marked variation in expression of mRNA for type I and III procollagen. Although the level of collagenase mRNA also varied among different cell strains examined, the expression of collagenase mRNA was lower in fibroblasts derived from deeper layers of each skin sample. In conclusion, cells from different layers of normal skin samples are heterogeneous in their constitutive expression of some key extracellular matrix components such as collagenase and types I and III procollagen.
Keloid fibroblast responsiveness to epidermal growth factor and activation of downstream intracellular signaling pathways
Authors: Satish Latha , Babu Mary , Tran Kien T. , Hebda Patricia A. , Wells Alan
Published: February 2004

Keloids, which overgrow the boundaries of the original injury, represent aberrations in the fundamental process of wound healing that include over‐abundant cell in‐migration, cell proliferation, and inflammation, as well as increased extracellular matrix synthesis and defective remodeling. To understand the key events that result in the formation of these abnormal scars would open new avenues for better understanding of excessive repair, and might provide new therapeutic options. We examined epidermal growth factor receptor (EGFR)‐induced cell motility in keloid fibroblasts, as this receptor initiates cell migration during normal wound repair. We show that keloid fibroblasts respond to EGF‐induced cell migration but the response is somewhat diminished compared to normal adult fibroblasts (∼30% reduced); the mitogenic response was similarly blunted (∼5% reduced). Keloid fibroblasts express near normal levels of EGFR (82%), but show a much more attenuated activation of EGFR itself and the motility‐associated phospholipase C‐γ. This was reflected in part by rapid loss of EGFR upon exposure to EGF. Interestingly, while extracellular signal‐regulated kinase/mitogen‐activated protein kinase (ERK‐MAPK) activation was relatively robust in keloid fibroblasts, the downstream triggering of the motility‐associated calpain activity was blunted. This was reflected by high cell–substratum adhesiveness in the keloid fibroblasts. Thus, the blunted migratory response to EGF noted in keloid fibroblasts appears due to limited activation of two important biochemical switches for cell motility.
Connective tissue growth factor siRNA modulates mRNA levels for a subset of molecules in normal and TGF‐β1–stimulated porcine skin fibroblasts
Authors: Wang Jian Fei , Olson Merle E. , Ma Lingling , Brigstock David R. , Hart David A.
Published: February 2004

Previous studies in a pig model of skin wound healing showed a coordinate expression of transforming growth factor‐β (TGF‐β) and connective tissue growth factor (CTGF), and exposure of porcine skin fibroblasts in vitro to recombinant human CTGF significantly up‐regulated mRNA levels for a number of molecules. Therefore, based on recent reports that small interfering RNA (siRNA; double‐stranded RNA) can effect silencing of the expression of gene(s), this approach has now been used with CTGF‐specific siRNA to better understand the function of this growth factor in regulating matrix homeostasis and repair. Normal skin fibroblasts from Yorkshire pigs were treated with 0.1–0.8 µM CTGF siRNA, TGF‐β, or TGF‐β plus CTGF siRNA for 12–48 hours. Total RNA was isolated and quantified, and then mRNA levels for specific molecules were analyzed by reverse transcription‐polymerase chain reaction. Protein levels for CTGF and HSP47 were assessed by Western‐blot analysis. CTGF siRNA transfection led to significant decreases in mRNA and protein levels for CTGF in both a dose‐ and time‐dependent manner. mRNA levels for types I and III procollagen, decorin, HSP47, tissue inhibitor of metalloproteinase ‐1, ‐2, ‐3, and basic fibroblast growth factor were also significantly and uniquely decreased following exposure of cells to CTGF siRNA. Addition of TGF‐β to the cells led to increases in CTGF mRNA levels that were blocked by CTGF siRNA. CTGF siRNA exposure also significantly and selectively down‐regulated TGF‐β–mediated increases in mRNA levels for types I and III procollagen. The results indicate that CTGF can regulate extracellular matrix molecule, growth factor, and proteinase inhibitor gene expression, and that some of the TGF‐β effects on skin fibroblasts are via a CTGF‐dependent pathway. (WOUND REP REG 2004;12:–)
Genomic analysis of insulin‐like growth factor‐I gene transfer in thermally injured rats
Authors: Dasu Mohan R. K. , Herndon David N. , Spies Marcus , Perez‐Polo J. Regino
Published: February 2004

Thermal trauma causes tissue damage by membrane destabilization and energy depletion at the cellular level, resulting in tissue necrosis and inflammation leading to delayed cell death. One therapeutic approach is to block the immediate triggering of the inflammatory cascade that results in prolonged hypermetabolic responses and immune dysfunction while promoting the expression of growth factors. In the present study, we determined hepatic gene expression responses to insulin‐like growth factors‐i (IGF‐I) gene transfer to burned rats using high‐density DNA microarray assays. The expression of 123 out of ~8,800 genes assyed (1.4% of total) were significantly altered. Of these, 42 genes were altered irrespective of treatment by burn trauma (p < 0.05). Changes in gene expression were confirmed by measuring mRNA levels using reverse transcription‐polymerase chain reaction and protein levels by Western blot assays. DNA microarray analyses showed two major mechanisms that mediated beneficial outcomes after IGF‐I gene transfer in the burned rat livers. These mechanisms were the stimulation of IGF binding protein potentiation of peripheral IGF‐I and the inhibition of the burn‐augmented pro‐apoptotic and oxidative mitochondrial metabolites stimulated by thermal trauma.
Lansoprazole promotes gastric mucosal cell proliferation and migration by activating p44/p42 mitogen‐activated protein kinase
Authors: Suzuki Hidekazu , Masaoka Tatsuhiro , Minegishi Yuriko , Motosugi Yuichiro , Miura Soichiro , Ishii Hiromasa
Published: February 2004

Cell proliferation and migration are important repair mechanisms in cell defect type mucosal injuries, such as peptic ulcers. To evaluate the level of cell restitution in vitro, we established a normalized assay system for analyzing the area of a tissue defect created in the center of a cultured cell layer. Although proton pump inhibitors are known to be potently effective in the treatment of peptic ulcers by inducing acid suppression, they are also effective in low‐acid conditions, such as in gastric ulcers associated with severe atrophic gastritis of the corpus. The present study was designed to examine the pH‐independent effect of lansoprazole (LPZ) on cell restitution in vitro. The mouse gastric mucosal cell line, GSM06, was cultured to confluence. A 4‐fluoric ethylene‐tipped aluminum stick was then used to produce a cell‐free area in the center of the culture well. After measuring the area of the cell defect using a digital analyzer equipped with an inverted microscope, LPZ was added to each well; the area of the residual cell defect was then measured 6 and 24 hours after LPZ administration. To investigate the involvement of the p44/p42 mitogen‐activated protein kinase (MAPK) and p38 MAPK in this process, PD98059 (a MEK inhibitor) or FR167653 (a p38 MAPK inhibitor) was added to the cell cultures. In a separate experiment, GSM06 cells were cultured to the subconfluent level, each test agent was added, and the cell number in each well was measured using an MTT assay 16 hours after the administration of the agents. Six hours after the addition of LPZ, a slight but significant increase in the cell restitution rate was observed in the LPZ‐treated groups compared with that in the control group. After 24 hours, a further significant increase in the cell restitution rate was observed in the LPZ groups compared with that in the control group. While the addition of PD98059 significantly attenuated the cell restitution rate in the LPZ groups, the addition of FR167653 had no such effect. The total cell number in the subconfluent cell cultures was significantly increased in the LPZ‐treated groups compared with that in the control group. In conclusion, LPZ promotes the healing of injured gastric mucosal cells following injury by enhancing cell proliferation and migration. Furthermore, the mechanism by which cell proliferation and migration is promoted by LPZ may involve the activation of p44/p42 MAPK.
The effect of vacuum‐assisted closure therapy on the pig femoral artery vasomotor responses
Authors: Wackenfors Angelica , Sjögren Johan , Algotsson Lars , Gustafsson Ronny , Ingemansson Richard , Malmsjö Malin
Published: February 2004

Vacuum‐assisted closure (VAC) is frequently used to treat wound infections. The aim of the present study was to evaluate the effect of VAC therapy on blood vessels. Vasodilatation and vasoconstriction were studied in isolated ring segments of the pig femoral artery after continuous VAC therapy of an inguinal wound for 12 hours. Vasoconstriction induced by endothelin‐1 (ET‐1), which is mainly an endothelin type A receptor agonist (Emax = 181 ± 2% of potassium), and the endothelin type B receptor agonist, sarafotoxin 6c (Emax = 30 ± 1%), were significantly increased after VAC therapy (ET‐1; 325 ± 3% and sarafotoxin 6c; 69 ± 1%). The norepinephrine‐, phenylephrine‐, and angiotensin II‐induced vasoconstrictions were not affected by VAC therapy. Acetylcholine induced an endothelium‐dependent dilatation that was enhanced after VAC therapy (Rmax = 38 ± 1% of norepinephrine‐preconstriction after sham and 47 ± 1% after VAC therapy, p < 0.05). The dilatory response was mediated by nitric oxide (Rmax = 39 ± 1%), prostaglandins (5 ± 1%) and endothelium‐derived hyperpolarizing factor (16 ± 1%), which were all significantly increased after VAC therapy. In conclusion, VAC therapy for 12  hours enhances an endothelin type A and type B receptor‐mediated vasoconstriction. This may be compensated for by a more efficacious endothelium‐dependent vasodilatation. No spontaneous bleeding, perforation, dissection, or other macroscopic change could be observed in the arteries exposed to VAC therapy.
Cranial bone defect healing is accelerated by mesenchymal stem cells induced by coadministration of bone morphogenetic protein‐2 and basic fibroblast growth factor
Authors: Akita Sadanori , Fukui Masashi , Nakagawa Hiroshi , Fujii Tohru , Akino Kozo
Published: February 2004

To facilitate bone healing in difficult circumstances, and to replace conventional therapeutic modalities, highly purified bone marrow‐derived human mesenchymal stem cells (hMSCs) were investigated for induction of their osteogenic lineage upon provision of cytokine cues in vitro and in the cranial defect model in vivo. Alkaline phosphatase‐expressing cells were most frequently observed when the hMSCs were treated with 2.5 ng/ml of basic fibroblast growth factor (bFGF) and 50 ng/ml of bone morphogenetic protein (BMP)‐2 for 4 days in culture after a 6‐day incubation in osteogenic medium containing dexamethasone, ascorbic acid‐2‐phosphate, and β‐glycerophosphate. Four‐millimeter full‐thickness cranial defect wounds were made in male nude rats (F344/NJCl‐rnu), whose deficit in the T cell compartment prevented T‐cell–mediated cellular rejection. The animals were treated for 4 weeks with hMSCs and application of 10 µg each of bFGF and BMP‐2 that had been soaked into a gelatin sponge carrier. Significant bone mineral density was observed by dual X‐ray absorptiometry and this treatment also produced histologically mature osteocytes surrounded by both osteoblasts and osteoclasts expressing alkaline phosphatase and osteocalcin. The bone mineral densities and histological structures were matched at 8 weeks post‐transplantation. Therefore, human bone marrow‐derived mesenchymal stem cells are able to differentiate into an osteogenic lineage upon cytokine stimulation and accelerate healing in a nude rat cranial bone healing model.
Bacterial load in relation to vacuum‐assisted closure wound therapy: A prospective randomized trial
Authors: Mouës Chantal M. , Vos Margreet C. , Van Den Bemd Gert‐Jan C.M. , Stijnen Theo , Hovius Steven E.R.
Published: February 2004

Vacuum‐assisted closure has become a new technique in the challenging management of contaminated, acute, and chronic wounds. Although promising clinical results have been described, scientific proof to substantiate the mechanism of action of this therapy is scarce. In the present study, we examined whether the positive effect on wound healing found in vacuum‐assisted closure–treated wounds could be explained by an effect on the bacterial load. Fifty‐four patients who needed open wound management before surgical closure were included in this study. Wounds were randomized to either vacuum‐assisted closure therapy (n= 29) or treatment by conventional moist gauze therapy (n= 25). Healing was characterized by development of a clean granulating wound bed (“ready for surgical therapy”) and reduction of wound surface area. To quantify bacterial load, biopsies were collected. No significant difference was found in time needed to reach “ready for surgical therapy” comparing both therapies. Wound surface area reduction was significantly larger in vacuum‐assisted closure–treated wounds: 3.8 ± 0.5 percent/day (mean ± SEM) compared to conventional‐treated wounds (1.7 ± 0.6 percent/day; p < 0.05). The total quantitative bacterial load was generally stable in both therapies. However, nonfermentative gram negative bacilli showed a significant decrease in vacuum‐assisted closure–treated wounds (p < 0.05), whereas Staphylococcus aureus showed a significant increase in vacuum‐assisted closure–treated wounds (p < 0.05). In conclusion, this study shows a positive effect of vacuum‐assisted closure therapy on wound healing, expressed as a significant reduction of wound surface area. However, this could not be explained by a significant quantitative reduction of the bacterial load.
009  Calcium‐Zinc Alginate and Recombinant Human bFGF; Its efficacy from the histological standpoint
Authors: Otsuka T , Tosa Y , Hosaka Y
Published: February 2004

Aim: To evaluate the efficacy of wound dressing material, Calcium‐Zinc Alginate: Curasorb Zn® as a carrier of the recombinant human bFGF, histological detection was performed.

Methods: Full thickness skin defect on the back of Wistar rat was made for four groups; the control group, bFGF group; topical application of recombinant human bFGF for 5 μg per day, Ca‐Zn Alginate group; topical use of the dressing and the group that both the bFGF and Ca‐Zn Alginate were applied, nine rats for respective group.

Results: Significant granulation was noted both bFGF group and Ca‐Zn Alginate group compared to the control group. Fibrosis was noted in the control and Ca‐Zn Alginate group whereas no fibrosis was noted in the FGF group. Numerous amounts of cells were noted in the FGF group which indicate strong inflammation. Also, marked abscess formation was noted in the bFGF group. The greatest thickness of granulation was obtained in the group that both the bFGF and Ca‐Zn Alginate were applied.

Conclusions: It was considered that the appropriate use of the carrier for the bFGF is important to control infection. The Ca‐Zn Alginate dressing was effective to suppress abscess formation.
011  Effects of Granulocyte Colony‐Stimulating Factor in Rats with Severe Acute Pancreatitis
Authors: HongFang Tuo , Masanobu Nakashima , Nobutsugu Abe , Masanori Sugiyama , Yutaka Atomi
Published: February 2004

Aim: It is reported that Granulocyte colony‐stimulating factor (G‐CSF) increases the number and functions of the neutrophils of blood in animal models. In the present study, we observed the effects of G‐CSF in rats with severe acute pancreatitis.

Methods: Pancreatitis was induced by injection of 0.2 ml of 3% taurocholate acid into biliopancreatic duct of the Wistar rat. Thirty rats were randomized into three groups. 1: Control group (C group). 2: Acute pancreatitis (AP) group. 3: AP + G‐CSF groups. G‐CSF was administrated via jugular veins 1 hour before induction of pancreatitis. Blood and ascites at 1 and 3 hours after induction of pancreatitis were measured for the number of neutrophils, their phagocytosis and bactericidal activity and the concentrations of TNF‐á, IL‐6 and IL‐1â.

Results: The phagocytic neutrophils increased in AP + G‐CSF group (22 ± 4.3 × 105) compared with AP group (10 ± 1.9 × 105)(p < 0.05); the bactericidal neutrophils increased in AP + G‐CSF group (60 ± 14.5 × 105) compared with AP group (20 ± 6.1 × 105)(p < 0.05). G‐CSF did not increase any the concentration of TNF‐á, IL‐6 and IL‐1â in blood and ascites.

Conclusions: G‐CSF increases the number of phagocytic and bactericidal neutrophils of blood and ascites in SAP rat, without increasing the concentration of TNF‐á, IL‐6 and IL‐1â, therefore improves host defense against infection.
013  A New Method of Closing the Sternum with Absorbable Suture and Absorbable Fixed Pins
Authors: Hosomura N , Nagamine H , Yamahata T , Nakayama M , Kikuchi K
Published: February 2004

Aim: In case of using metal wire in closing after the median sternotomy, several disadvantages have been reported, including long lasting subject's sense of incongruity, risks of tear of the wire and cutting to be made by the wire, tenderness and becoming an abstacle for the use of CT or MRI. In our facilities, in order to solve these problems, absorbable suture and absorbable rib fixing pins have been used in closing the sternum.

Methods: Objects of the median sternotomy we’ve had since October, 2001, are six cases. The suture used was 6 PDS codes of 1 mm in diameter and it went through the sternum and ligated it. Three absorbable pins of poly‐L‐lactide were used.

Results: A progress observation period after the operation is from one month to fourteen months.The prognosis have been good and complications not recognized.

Conclusions: The way of closing the sternum with absorbing suture and absorbing rib fixing pins will make it possible to solve problems incurred from the metal wire suturing.
015  Three Cases of Pressure Ulcer cured by Concomitant Use of bFGF and Dressing
Authors: Toshihiro Kitte , Shinya Arinaga , Toshiro Tatsuma , Akira Nakamura
Published: February 2004

Introduction: It is reported on 3 cases of pressure ulcer which were hard to cure surgically by flap plastic operation or else, were completely healed by concomitant use of the basic fibroblast growth factor (bFGF) and the dressing.

Case report: CASE 1:A 55‐year‐old male with a pressure ulcer 3 cm in diameter formed on the buttock. After excising the margin and spraying bFGF, the wound was covered with a dressing made of hydrofiber. It was completely cured in 3 months. CASE 2:A 65‐year‐old male with a pressure ulcer on the sacral region 30 cm the major axis. The pocket was cut open, and bFGF was sprayed inside. Amounts of the dressing material depended on volumes of the exudate. It was cured completely in 6 months. CASE 3:A 85‐year‐old male with a pressure ulcer 10 cm in the major axis on the greater trochanteric region of the right side. The margin was excised to open the pocket, bFGF was sprayed, and hydrogel was used as the dressing. It was cured in 6 months completely.

Discussion: Usefulness of the conservative treatment is reported based on the experience referring to these 3 cases with pressure ulcer, which were completely cured upon concomitant use of bFGF and the dressing material suitable to the conditions of the wound after thorough surgical debridement and washings.
016  Is This Ulcer A True “Chronic” Ulcer?
Authors: Hiroshi Yasuda , Koji Murata , Ryoko Kinoshita , Hideka Nakagawa , Miwa Kobayashi , Yoshiki Tokura
Published: February 2004

Aim: Lethal diseases occasionally present with a “chronic” ulcer. If a physician can not realize this fact, the ulcer might give rise to a misery result. We present three educational cases of chronic ulcers that give caution to physicians.

Case report: The first case was a hand ulcer due to postradiation squamous cell carcinoma, which eventually led to amputation of index and middle finger. The second case was a leg ulcer due to postburn squamous cell carcinoma, resulting in death caused by visceral metastasis. Both cases were treated as simple chronic ulcers at other clinics for a long time. The third case was a lower leg ulcer caused by infection with cryptococcus neoformans. The ulcer was covered with a split thickness skin graft after administration of anti‐fungal drugs.

Conclusions: As seen in these cases, we must always have serious doubts why the ulcer is not easily healed. If the causative factor is not easily found, biopsies from the ulcer would be helpful for the true diagnosis of the ulcerative lesions.
017  An Investigation into Bacteria of Intraperitoneal Washed Fluid on Surgical Site Infection
Authors: Takumi Irie , Hiroyuki Baba , Mituo Endo
Published: February 2004

Aim: Surgical Site Infection (SSI) has been considered to be caused by the sticking bacteria on the surgical site. We have investigated how the sticking bacteria may influence the SSI in clean‐contaminated surgery.

Methods: In clean‐contaminated operations with a Protractor we washed intraperitoneal space of 26 patients, 14 male and 12 female, whose mean age were 67.2 ± 7.6 years old, with 3L of saline for the operations of upper gastrointestinal tract and 5L for under. We collected the washed fluid at pre‐operation and post‐operation, and abdominal fat for the culture of the sticking bacteria in those specimens.

Results: All SSI rate was 3.8%. Each detective rates of bacteria were 38.5%, 50%, and 46.2%, but each SSI rates were all 0%. We used the Protractor which may protect the sticking of bacteria on surgical sites, however many bacteria have been detected on them. There was only one patient who had been resulted in SSI. In this case the patient was suffered from hypovolemic shock in pancreatic‐tail resection, however we could not detect any sticking bacteria on the surgical site.

Conclusions: Sticking of bacteria on surgical sites is not a chief cause of SSIs in clean‐contaminated surgery. We guess there may be another cause as the bacterial‐translocation for the SSIs in clean‐contaminated surgery.
018  A Case Report: Successful Wet Treatment of a Dissociated and Infected Surgical Wound After Radical Vulvectomy
Authors: Setsuko Murata
Published: February 2004

A 72‐year‐old female, who had slight impaired glucose tolerance, was diagnosed the second stage of carcinoma of the vulva. On the second day after radical vulvectomy with inguinal lymphadenectomy, the center of the wound changed color into deep purple. On the 8th postoperative days, the swelling of the wound extended over the mons pubis and a thigh, so the infected wound, which reached the epimysium, was opened for the debridement. Pseudomonas aeruginosa and gram negative bacilli were detected from the wound. Firstly, the wound was cleaned by a physiological salt solution, streptokinase, and sulfadiazine silver. The gynecologist and nurses in charge discussed about the therapy under the consultation of a dermatologist and treated the wound by the wet treatment that a piece of gauze moisten by a physiological salt solution was exchanged 3 or 4 times a day after disinfecting by povidone iodine. On the13th postoperative days, the infection was controlled and on the19th days the skin was grafted from a thigh. After the successful skin grafting the patient was discharged. Carcinoma of the vulva tends to occur in elder women. Wounds are often dissociated because of an interruption in the circulation by the excessive extension of the skin. From the successful experience of wet treatment after a surgical wound infection, we recognized the necessity of close observation on surgical site and the importance of cooperation of the doctors and the nurses in charge.
019  2 Cases of Mediastinitis After Median Sternotomy Surgery Treated with Vacuum‐Assisted Closure
Authors: Hideyuki Yanagi , Hiroto Terashi , Shinya Tahara
Published: February 2004

Introduction: Mediastinitis, which is at times encountered after thoracic surgery with median sternotomy, is often resistant for treatments and in cases leads to death. We applied vacuum‐assisted closure (V.A.C.) for 2 cases of post‐sternotomy mediastinitis, in an original manner using readily available materials and made them healed. The details are reported.

Methods: We applied polyurethane foam (HydroSite®, Smith & Nephew) with silicone tube (BLAKE®, ETHICON) on the debrided wound, covered it with surgical draping film (Ioban® 2, 3M Health Care), and applied continuous negative pressure with a wall suction. The dressing was changed at intervals of 2 to 3 days.

Results:(Case 1) A 53‐years old male with complications of diabetes mellitus and hypertension underwent coronal artery bypass graft surgery for angina pectoris. Sternal infection occurred in postoperative period and the wound was opened. (Case 2) A 78‐years old male underwent total aortic arch replacement surgery for aortic dissection. The wound was opened for a postoperative infection. We applied vacuum‐assisted closure therapy for these 2 cases in the method presented above and observed promoted wound granulation and rapid contraction of wounds, with no manifestations of increasing infection or changes of circular conditions. Each of the cases was healed with vacuum‐assisted closure in 7 Weeks without any surgical procedures.

Conclusions: We viewed vacuum‐assisted closure therapy as another choice of the treatment for mediastinitis.
020  Study on the Wound Cure after Laparoscopic Operation
Authors: Kenji Kakisako , Fumitaka Yoshizumi , Kimio Yamaguchi , Kazutoshi Kaketani
Published: February 2004

Aim: Regarding the recent change in the surgical technique of infraumbilical incision for laparoscope insertion from lateral incision to longitudinal incision to reach inside the umbilicus, we conducted this study to find the merits and demerits of change as well as to clarify the circumstance in which poor wound cure occurs after laparoscopic operation.

Methods: Total 608 patients who underwent laparoscopic operations between February 2000 and July 2002 at this hospital were used as the subjects of this study. Accordingly, the comparison was made between the 343 cases in the early stage (lateral incision) and the 380 cases in the late stage (longitudinal incision).

Results: Postoperative wound trouble occurred in 24 (3.9%) of the 608 cases. Infection occurred in the wound made by open laparoscopy in 5 patients. The trouble in the trocker insertion site itself was observed in 19 cases (3.1%). The infraumbilical wound trouble occurred in 10 cases in the early stage and 5 cases in late stage indicated, indicating lower trouble rate in the latter.
021  “Epithelization Unit”: A New Concept in Skin Wound Healing
Authors: Naoto Yamamoto , Tomoharu Kiyosawa , Katsuyuki Arai
Published: February 2004

Aim: Development of adequate connective tissue as a bed below new epidermis is important for successful epitherialization. However, how the dermal fibrous architecture newly forms and changes chronologically in that process is unknown.The aim of the present study was to determine the regeneration process of neodermis in skin wound healing from the viewpoint of neoformation of its fibrous architecture

Methods: We investigated the repair process of an excised skin wound made on experimental animals with the use of alkali water cell‐maceration scanning electron microscopy, which allowed a three‐dimensional demonstration of individual collagen fibrils in their natural from and location

Results: Both new papillary dermis and new epidermis underwent synchronous development in epithelization, as if forming “a unit”. The scar tissue has the same layered structure as seen in normal skin, namely epidermis including basement membrane, papillary dermis, and reticular dermis. The new epidermis on the new papillary dermis demonstrated a mature appearance and a mechanically stable condition.

Conclusions: The healing process proceeds as forming a unit composed by epidermis including basement and papillary dermis, and is finally completed with total coverage of the wound by this unit. We would like to call this unit, the “epithelization unit”.
022  Preincubation Reduces Epithelialization Period after Grafting in Cell‐Precomfluent Cultured Skin
Authors: Yasumi Saso , Takeshi Kawazoe , Shigehiko Suzuki , Kenji Tomihata
Published: February 2004

Purpose: We have developed a cultured skin with both epidermal and dermal components using two different types of collagen sponges as a scaffold. In addition, we succeeded in grafting cell‐preconfluent cultured skin immediately after seeding the cells. In this study, we examined whether the incubation before grafting can reduce the period of epithelization after grafting.

Method: We produced cultured skin as follow. We seeded fibroblasts on the dermal collagen sponge with a seeding density of 100,000 cells per square centimeter. After incubation for 4 hours, an epidermal collagen sponge was put on the dermal collagen sponge, and keratinocytes were seeded on the epidermal collagen sponge with a seeding density again of 100,000 cells per square centimeter. The cultured skin was incubated for 1, 2, and 3 days (n = 3), and grafted them onto skin defects in SCID mice. Three, 5, 7, and 14 days after grafting, tissue specimens were harvested for histological and immunohistochemical examination.

Result: Three days after grafting, in 3 days incubation group, the cultured skins formed epithelium with a few layers, while in other groups, they did not. Five days after grafting, in 3 days incubation group, the cultured skins formed 6–8 epithelium layers, while in other groups, they did fewer layers. Seven days after grafting, in 3 days incubation group, the cultured skin formed epithelial layers consisiting of cornified layers, and human type IV collagen was stained at the dermal‐epidermal junction. In other groups, it was not stained. Fourteen days after grafting, all samples formed epithelial layers, and human type IV collagen was stained.

Conclusion: In 3 days incubation group, the epithelization was completed earlier than that in the other groups.
023  Development of Tissue Engineering Skin Based on Acellular Dermal Matrix
Authors: Ryo Yamaguchi , Yoshihiro Takami , Jingping Guo , Hideharu Tanaka , Syuji Shimazaki
Published: February 2004

Aim: In tissue engineering of the skin, the selection of a scaffold or a matrix in which a cultured cells grow is quite important. We have developed a tissue engineering skin composed of human keratinocytes and fibroblasts on an acellular allogenic dermal matrix (ADM), derived from cryopreserved human skin.

Methods: ADM was prepared from cryopreserved split‐thickness human skin by treating with Dispase and Triton X‐100. The tissue engineering skin were produced by seeding human keratinocytes and fibroblasts on ADM. Several days after seeding, the tissue engineering skin was exposed to an air‐liquid interface for another 7 days. Then, the histological structure of the skin and the production of growth factors by the skin were investigated.

Results: The produced ADM was found to be completely acellular with remaining structure of the basement membrane components, such as type IV collagen and laminin. The developed tissue engineering skin had stratified keratinocytes on the surface of the ADM migrating fibroblasts in the dermal collagen structure, resembling to the normal skin appearance. It was found that several important growth factors in wound healing process, such as TGF‐α, TGF‐β and VEGF, were produced by the tissue engineering skin.

Conclusions: It was suggested that ADM is suitable for a scaffold in tissue engineering of the skin.
026  Treatment with Autologous Cultured Dermal Substitutes (CDS) for Burn Scar Contracture in Children
Authors: Yasushi Fujimori , Koichi Ueda , Yuka Omiya , Kentaro Kubo , Hiromichi Matsui , Yoshimitsu Kuroyanagi
Published: February 2004

Aim: A treatment of extensive burn contracture in children needs to repeat the autologous skin graft. This study was designed to evaluate the application of autologous CDS to prepare the proper wound bed acceptable for the split‐thickness autologous skin graft.

Methods: Prior to the clinical study, the master cell banking system was established using a small piece of skin derived from each patient. The autologous CDS was prepared by plating the patient's own fibroblasts, cultured from the master cells, on a spongy matrix of hyaluronic acid and atelo‐collagen. The CDS was applied on the skin defects left behind surgical excision to release scar contracture. The split‐thickness autologous skin graft (6∼8/1000 inch) was applied on the wound bed, prepared by using the CDS.

Results: The clinical trials were conducted in 5 cases. When the autologous CDS was applied on the skin defect, exposing subcutaneous fatty tissue, the highly vascularlized wound bed was prepared within about 1 week. Although a split‐thickness skin graft was very thin, the severe contracture was not observed over a period of several months.

Conclusion: The application of autologous CDS is promising for the treatment for extensive burn scar contracture in children.
028  Clinical Trials of Allogenic Cultured Dermal Substitute for the Treatment of Burn Wounds
Authors: Tadaaki Morotomi , Hiroshi Kamiishi , Kentaro Kubo , Hiromichi Matsui , Shizuko Kagawa , Yoshimitsu Kuroyanagi
Published: February 2004

Aim: Recently, various types of cultured skin substitutes have been developed and some of them are used clinically. This study was designed to evaluate the efficacy of allogenic cultured dermal substitute which was applied to burn injuries in clinical trials such as deep dermal burns and dermal burns.

Methods: Allogenic cultured dermal substitute (CDS) was simply applied to the burn wound, over which covering materials were applied to protect CDS.

Results: The application of CDS to deep dermal burns was proved to facilitate healthy granulation tissue formation at early stage and epithelialization from the outer margins. When CDS applied to the debrided wound surface of dermal burns, an excellent wound bed was generated which was suitable for the graft take of an autologous patch.

Conclusion: CDS provides an excellent epithelialization and granulation for burn wounds.
029  Treatment of Fresh DDB Wounds with Allogenic Cultured Fibroblasts, Allogenic Cultured Dermal Substitute
Authors: Kazutaka Soejima , Takamitsu Nakano , Hajime Matsumine , Daiki Nakamori , Motohiro Nozaki , Kentaro Kubo , Hiromichi Matsui , Sizuko Kagawa , Yoshimitsu Kuroyanagi
Published: February 2004

Aim: Multi‐center's clinical study for the application of the cultured dermal substitute (CDS) composed of hyaluronic acid and collagen spongy matrix with allogenic cultured fibroblasts1 is in progress since 2001. In this study, effect of the CDS for the treatment of fresh DDB wounds was tested.

Methods: Six cases of second‐degree burns, diagnosed as DDB by observation of blood stream of the dermal capillary using Compact Micro Vision System (Hi‐Scope®, Hi Rox Co., Japan), were treated with the cultured allogenic fibroblasts, CDS.

Results: The epithelialization of the wounds was obtained on 8.2 ± 2.6 (mean ± SD) days after the application.

Conclusion: The results of our clinical experiences suggest that the allogenic cultured fibroblasts have beneficial effect on the wound healing of the DDB.
030  Clinical Trials With Allogeneic Cultured Dermal Substitutes (CDS)(Regenerating Medical Millennium Project of the Ministry of Health, Labor and Welfare)
Authors: Yasushi Fujimori , Koichi Ueda , Yuka Omiya , Kentaro Kubo , Hiromichi Matsui , Yoshimitsu Kuroyanagi
Published: February 2004

Aim: Clinical researches using allogeneic CDS, developed in the R & D Center for Artificial Skin of Kitasato University, have been carried out in 30 medical centers across Japan. The clinical results in our hospital, especially focusing on the treatment of refractory ulcers and dermal burns, were reported in this study.

Methods: The CDS was prepared by plating cultured fibroblasts on a spongy matrix of hyaluronic acid and atelo‐collagen. The CDS was used in 13 clinical trials, including 6 refractory ulcers, 4 dermal burns, 1 skin defect left behind preparing skin flaps, and 2 skin defects left behind removal of scar. The use of CDS in conjunction with a conventional ointment‐gauze dressing was repeated at an interval of 4 to 7 days over a period of 2 to 6 weeks.

Results: The successful application with CDS was achieved in all cases. Especially, in case of refractory ulcers, failed to heal even by using trafermin (Fiblast spray®) or other ointments, a complete healing was achieved in one case and wound size reduction was observed in other 5 cases within 6 weeks.

Conclusions: Our results suggest that the CDS is effective in the treatment of refractory ulcers and other skin defects. The excellent clinical results seem to be related to the cytokines released from the CDS.
031  Clinical Trials with Allogeneic Cultured Dermal Substitutes for the Treatment of Burns and Skin Ulcers (Regenerating Medical Millennium Project of the Ministry of Health, Labor and Welfare)
Authors: Hiroaki Oka , Kosaku Suenobu , Takahiko Moriguchi , Kentaro Kubo , Hiromichi Matsui , Yoshimitsu Kuroyanagi
Published: February 2004

Aim: Multi‐center clinical trials were performed using allogeneic cultured dermal substitutes (CDS) newly developed in the R & D Center for Artificial Skin of Kitasato University.

Method: This study was designed to evaluate the efficacy of CDS for the treatment of burns in 5 cases and the treatment of skin ulcers in 4 cases. Complications with the skin ulcers were diabetes (1 case), varix (1 case), and collagen diseases (2 cases). The wound surface area was measured with picture analysis software during the wound healing process.

Results: The clinical evaluation was made using a protocol that includes a standard for the development of new wound dressings. According to the comprehensive judgment, the results achieved were evaluated as excellent for 4 burn cases and good for one case. Regarding the skin ulcers, results were excellent in 3 cases and good for one case. No harmful complications were observed during this study.

Conclusions: CDS were useful for burn and ulcer treatment but the wound surface must be checked rigorously for the occurrence of infection during the healing process.
032  Treatment of Chronic Ulcer with Allogeneic Cultured Dermal Substitute (CDS) composed of Collagen Sponge with Fibroblasts
Authors: Hideo Oshima , Hajime Inoue , Masayoshi Tanabe , Yu Takahashi , Yoshimitsu Kuroyanagi , Norio Kumagai
Published: February 2004

We have been cultivating human epidermal cells for therapeutic purpose according to the original methods developed by Rheinwald and Green. Cultured epithelium (CE) was applied to patients with severe skin defects, burn wounds, chronic skin ulcers and cutaneous disorders like hypomelanosis. Autologous CE allows to restore massive skin surface in a short period compared with other conventional treatments. For grafts take, it is important to manage wound beds properly prior to CE grafting. The CDS was applied to prepare wound bed acceptable for CE grafting. The CDS was designed to secrete various types of cytokines, i.e., VEGF and KGF to stimulate wound healing. The successful management of deep wounds like chronic skin ulcer or burn ulcer requires granulation tissue formation and epithelialization to wound closure. This study aimed to evaluate the application of CDS in conjunction with CE for patients with chronic skin ulcer and burn ulcer. In some cases the wounds were cured by using CDS, and followed by CE grafting. All clinical trials achieved excellent or good results, showing no contracture and no hypertrophic scar after wound closure. The CDS was found to be useful to prepare wound beds and to facilitate wound management.
033  Clinical Trials with Allogeneic Cultured Dermal Substitutes (CDS) for Treatment of Skin Ulcers (Regenerating Medical Millennium Project of the Ministry of Health, Labor and Welfare)
Authors: Koichi Hirotomi , Taro Mikami , Kensaku Ohki , Kayo Daigo , Jiro Maegawa , Katsuyuki Torikai , Kentarou Kubo , Hiromichi Matsui , Yoshimitsu Kuroyanagi
Published: February 2004

Aim: Multi‐center's clinical study, using allogeneic CDS are proceeded in 30 hospitals as the Millennium Project of the Ministry of Health, Labor and Welfare. The clinical trial in our hospital was designed to evaluate the efficacy of CDS for the treatment of refractory skin ulcers.

Methods: The application of CDS on the debrided wound was repeated at an interval of 3 to 5 days. The evaluation was performed according to the protocol, especially focused on granulation tissue formation, wound size reduction, and epithelialization.

Results: The clinical trials were conducted in 6 cases, including 5 cases with various complications, i.e., diabetes (2 cases), collagen diseases (1 case), renal failure (1 case), and skin defect after resection of tumor (1 case). According to the comprehensive judgement, 5 cases were evaluated as achieving excellent, and 1 case was evaluated as achieving good.

Conclusion: The successful application of CDS is dependent on the control of wound conditions free from infection. Appropriate debridment was necessary prior to application of CDS.
034  Application of Allogeneic Cultured Dermal Substitute to Skin Defects After Excision of Skin Cancer
Authors: Yokoo K , Aoyama H , Tamada Y , Hashimoto T , Kuroyanagi Y , Kagawa S , Kubo K , Matsui H
Published: February 2004

Aim: This study was aimed to confirm the usefulness of allogeneic cultured dermal substitute (CDS) in the treatment of skin defects after excision of skin cancer.

Methods: Eleven elderly patients (mean age of 76.3 year old) with skin cancer were included in this study. Most of the patients had basic disease such as diabetes or ASO. Allogeneic CDS used in the study were produced at Kitasato University. The CDS were applied to skin defects with exposing bone or tendon after oncological surgery, and changed once or twice a week until the open wound became suitable for autologous skin grafting or healed completely.

Results: Healthy granulation tissue was formed to cover the exposed bone or tendon in all cases. Only in one case, the treatment with allogeneic CDS was abandoned owing to undesirable infection. In eight cases, appropriate wound beds acceptable for autologous skin graft were prepared. In other two cases, the skin defect became smaller and eventually closed without skin graft. In ten cases with or without skin graft, undesirable scar contracture was not observed over prolonged follow‐up.

Conclusions: Elderly patients with skin cancer provide reconstructive surgeons with challenging problems when bone or bare tendon is exposed after oncological surgery. Flap transfer might be complicated especially when the patients suffered from a basic disease such as diabetes or ASO. This study has confirmed that the use of allogeneic CDS is a safe and reliable method to achieve wound healing in those high‐risk patients.
035  Flat Resection Technique by CO2 Laser for Lentigo
Authors: Masashi Fukatsu , Naohiro Ishii , Yoshiaki Kubota , Yuji Nakanishi
Published: February 2004

It has been said that CO2 laser vaporization and resection conventionally performed for elevated lentigo may cause depressed scar formation when the diameter of the lesion exceeds 5 mm. Accordingly, in consideration of cosmetic factors, we have devised a method for the removal of even a larger elevated lentigo by the Flat Resection technique which can be used effectively without causing a depressed scar to form. Upon the primary laser irradiation, the lesion was resected at the horizontal plane of the skin, and the resected tumor was subjected to histopathological analysis. If the lesion remained about one month after the primary irradiation, additional laser treatment was performed at the patient's request. From our experience, satisfactory outcomes were obtained for up to 12‐mm lesions. Following the flat resection, residual lesions were pathologically observed in 52.5%, and macroscopically observed in 45%. Additional laser treatment was performed for 35%. It was considered that this technique is useful for sites such as the base of ala nasi and the vicinity of the eyebrow, of which simple reefing requires skills. However, since the possibility of malignant degeneration of lentigo could not be ruled out in a longer‐term follow‐up, particular care is needed to select a case suitable for this technique.
036  A New Method for Skin Abrasion Therapy using the Pulsed Carbon Dioxide Laser with Scanner
Authors: Yuki Iijima , Tatsunori Tanaka , Kei Yoneda , Eiji Miyamoto
Published: February 2004

We have treated superficial skin lesions using pulsed carbon dioxide laser with scanner (NIDEK COL‐1040). We can choose the shape, size and power of irradiation programmed in the scanner. It makes the treatment more effective and uniform to combine the scanner mode with the pulse mode. However, for a lesion of complicated shape, it is difficult to avoid skin damage caused by multiple irradiations to a small area and/or irradiation to the normal skin. The result may be unsatisfactory with pigmentation or unsightly scar in such case. As the water‐solvability is a characteristic of the carbon dioxide laser beam, we used water film around the lesion to avoid unexpected irradiation to the normal skin. This method provided us better results.