激光佐治慢性皮肤溃疡疗效观察及诱导HSP70表达研究

目的:探讨激光治疗慢性皮肤溃疡的疗效及诱导创面组织热休克蛋白70(heat shock prote in 70,HSP70)表达情况。方法:将64例84处慢性皮肤溃疡创面随机分为传统治疗组和激光治疗组,激光治疗组在传统治疗基础上加用激光治疗,所有创面在治疗三周进行疗效判定;同期随机切取5例创面组织,并设正常皮肤为对照,行组织切片HSP70免疫组化染色,计数HSP70阳性细胞数并检测阳性细胞灰度值进行统计学分析;设计引物行RT-PCR检测HSP70mRNA表达情况。结果:两组慢性皮肤溃疡患者经相应治疗均较治疗前明显改善,但激光治疗组创面的治愈率和总有效率明显高于传统治疗组(P<0.05或P<0.01);免疫组化染色显示,正常皮肤和慢性溃疡组织中未见明显HSP70阳性表达细胞,激光治疗后三周后可见较多量的HSP70表达阳性细胞,其阳性细胞计数明显高于其它两组(P<0.01),细胞信号灰度表达明显低于其它两组(P<0.05),而传统治疗组与正常对照组无显著差异;RT-PCR结果发现激光治疗组组织中扩增到一条特异性泳带,大小约268 bp,正常皮肤和传统治疗组组织中未扩增到明显HSP70mRNA基因片段。结论:激光佐治慢性皮肤溃疡具有良好临床疗效,可能与激活创面细胞的热休克内源性保护机制而发挥抗感染和促进愈合作用有关。     

联合应用NGF 和胰岛素对糖尿病大鼠烫伤创面表皮干细胞的影响

目的:通过局部联合应用神经生长因子和胰岛素对糖尿病大鼠深II度烫伤创面表皮干细胞标记物β1整合素和角蛋白19(K19)表达的影响,探讨神经生长因子和胰岛素联合应用于糖尿病烫伤创面治疗后对创面愈合的影响。方法:雄性wistar大鼠腹腔注射链脲佐菌素(STZ)建立糖尿病模型60只,1月后在大鼠背部造成深II度烫伤。将大鼠随机分为糖尿病对照组(B)、胰岛素治疗组(C)、神经生长因子治疗组(D)、神经生长因子联合胰岛素治疗组(E),每组15只。另取15只正常雄性wistar大鼠作为正常对照组(A)。观察伤后3、7、11、15、21 d各组创面愈合情况,检测创面β1整合素和角蛋白19(K19)的表达并计算创面愈合率。结果:E组创面愈合率自第7天起较A、B、C、D组增加,为[(25.33±2.32)%,(P<0.05)];A、C、D组创面愈合率较B组增加分别为[(22.51±1.78)%,(16.68±1.95)%,(18.29±1.70)%,(P<0.05)]。E组整合素β1和角蛋白19(K19)表达自伤后第7至21天各时相点显著增加,(P<0.05)。结论:糖尿病大鼠深II度烫伤创面局部联合应用神经生长因子和胰岛素可促进表皮干细胞的增殖与分化从而加速创面的愈合。     

外源性VEGF促进大鼠Ⅱ度烫伤创面中晚期愈合

目的探究Ⅱ度烫伤时外源性血管内皮生长因子(vascular endothelial growth factor,VEGF)对皮肤愈合及其对表皮干细胞(epidermal stem cells,ESCs)迁移、分化的影响方法健康Wistar大鼠随机分为VEGF组、空白对照组、阿西替尼(Axitinib,VEGF抑制剂)组。采用水浴烫伤法制备Ⅱ度烫伤模型,分别以0.2μg/ml VEGF、PBS溶液和10μg/ml阿西替尼处理各组创面,各组均治疗7d,从烫伤至创面愈合分别在第2d、8d、14d及21d测量创面愈合情况,并取创面组织作组织学检测,运用免疫组化技术检测ECSs的分布及数量。结果①创面愈合率:烫伤后21d VEGF组>对照组>阿西替尼组;②愈合速度:烫伤后1-7d空白对照组>阿西替尼组>VEGF组,其后VEGF组愈合速度逐渐加快,第14d开始愈合速度表现为VEGF组>空白对照组>阿西替尼组;③组织学变化:烫伤后8-21d,VEGF组表皮细胞增殖明显,表皮修复和毛囊再生迅速,均早于空白对照组及阿西替尼组。④ECSs阳性细胞率变化:烫伤后第8-14dVEGF组ECSs阳性细胞率明显高于空白对照组和阿西替尼。结论Ⅱ度烫伤时,外源性VEGF在愈合中晚期加快愈合速度使愈合时间明显缩短,并且促进毛囊汗腺的再生使修复后的创面在外观、功能与正常皮肤相近,有助于提高全层皮肤创面的愈合质量。     

负压封闭引流对兔颅骨外露创面愈合效果的实验研究

目的:探讨负压封闭引流技术(VSD)对兔颅骨外露缺损创面愈合的治疗效果。方法:选取成年新西兰大白兔76只,平均分为四组并建立兔颅骨外露实验模型。其中,A组(19只):于兔颅骨上方制作直径为2.0cm的圆形创面,保留骨膜,采用-120mmHg负压引流和常规换药治疗;B组(19只):实验动物处理同A组,仅采用常规换药治疗;C组(19只):在兔颅骨上制作直径2.0cm的圆形创面,剔除骨膜,治疗方法同A组;D组(19只):实验动物处理同C组,治疗方法同B组。每组各抽取10只,观察创面愈合率和创面愈合时间;其余9只分别在第7天、10天、20天、30天进行取材检测,分析疗效机制。结果:A组创面愈合时间为19.40±1.65天,B组为24.00±2.31天;C组为25.40±4.43天,D组为30.00±5.50天。运用VSD治疗和常规治疗创面愈合时间比较有统计学意义(P0.05)。结论:VSD治疗兔骨外露缺损创面能有效缩短创面愈合时间,促进血管再生,胶原蛋白合成。     

创面生物活性玻璃修复材料促进家猪创面愈合的实验研究

目的:观察创面生物活性玻璃修复材料对家猪皮肤创面的促愈合作用。方法:选择14头家猪,随机分成7组,每组2头,在每头猪的脊柱两旁制造3个4×4cm的全层皮肤缺损的创面模型,每头猪6个创面又分成实验组和空白对照组,于试验后每天观察创面愈合情况,第1、3、7、14、21、28、35天图像分析计算创面愈合率,并同时取创面组织行组织学染色,观察各组材料对家猪皮肤全层缺损创面愈合的影响。结果:在涂材料的实验组和空白对照组创面愈合时间分别是23.19±1.27d、29.52±1.54d两组组间比较,具有统计学意义(P<0.05);实验组的创面愈合率在各时间段均高于空白对照组,差异有统计学意义(P<0.05);组织学观察实验组的上皮化程度、表皮生长、成纤维细胞、毛细血管数量均好于空白对照组。结论:创面生物活性玻璃修复材料对家猪皮肤创面愈合具有促进作用,可作为一种新型的促愈合覆盖材料进一步研究。     

创面生物活性玻璃修复材料促进家猪创面愈合的实验研究

目的：观察创面生物活性玻璃修复材料对家猪皮肤创面的促愈合作用。方法：选择14头家猪,随机分成7组,每组2头,在每头猪的脊柱两旁制造3个4×4cm的全层皮肤缺损的创面模型,每头猪6个创面又分成实验组和空白对照组,于试验后每天观察创面愈合情况,第1、3、7、14、21、28、35天图像分析计算创面愈合率,并同时取创面组织行组织学染色,观察各组材料对家猪皮肤全层缺损创面愈合的影响。结果：在涂材料的实验组和空白对照组创面愈合时间分别是23.19±1.27d、29.52±1.54d两组组间比较,具有统计学意义（P〈0.05）;实验组的创面愈合率在各时间段均高于空白对照组,差异有统计学意义（P〈0.05）;组织学观察实验组的上皮化程度、表皮生长、成纤维细胞、毛细血管数量均好于空白对照组。结论：创面生物活性玻璃修复材料对家猪皮肤创面愈合具有促进作用,可作为一种新型的促愈合覆盖材料进一步研究。     

在烫伤条件下VEGF对大鼠皮肤eNOS基因表达的影响

目的:探讨大鼠烫伤愈合过程中VEGF对皮肤组织eNOS基因表达的影响。方法:采用了大鼠深II度烫伤模型,分别以VEGF和阿西替尼(VEGF抑制剂)干预,观察大鼠烫伤后愈合过程的组织学变化,PCR检测创面eNOS基因的表达。结果:肉眼观察VEGF组大鼠创面最早愈合,对照组大鼠愈合速度其次,阿西替尼组创面愈合最晚;HE染色结果显示伤后第2、8、21天VEGF组炎性细胞浸润程度、新生毛细血管数量均高于对照组和阿西替尼组,PCR结果显示烫伤后第2、8天VEGF组的eNOS基因的表达明显上调,且同时间点与对照组比较有统计学意义(P0.05)。结论:VEGF可诱导烫伤创面eNOS基因的表达,增加血管生成重要因子NO的生成,有利于创面愈合。     

雷公藤对哮喘大鼠气道重构及磷脂酰肌醇3激酶表达的影响

目的:探讨雷公藤内酯醇对哮喘气道重构及磷脂酰肌醇3激酶(PI3K)表达的影响。方法:将40只SD大鼠随机分为5组(n=8):A组(正常对照组);B组(哮喘4周组);C组(哮喘6周组);D组(给药4周组);E组(给药6周组)。测定气道反应性并观察气道壁嗜酸性粒细胞浸润;图像分析软件测定支气管壁厚度、支气管平滑肌厚度及支气管平滑肌细胞核数量;免疫组织化学染色、逆转录聚合酶链式反应(RT-PCR)检测PI3K蛋白及mRNA表达。结果:①B组、C组PI3Kp85α的蛋白及mRNA表达水平显著高于A组(P均<0.01),E组上述指标较B组、C组、D组均显著降低(P<0.01、P<0.01、P<0.05);②B组及C组支气管壁厚度、支气管壁平滑肌厚度、支气管壁平滑肌细胞核数量均较A组明显增加(P均<0.01),而E组上述指标较B组、C组、D组均显著降低(P均<0.01);③B组、C组的气道反应性均高于A组(P均<0.01),E组较B组、C组、D组均显著降低(P<0.01、P<0.01、P<0.05)。结论:气道平滑肌增生是气道重构的一个显著特征,PI3K可能在此起促进作用。雷公藤内酯醇可能通过下调PI3K的表达而减轻哮喘气道高反应性及抑制气道平滑肌增生,对哮喘气道重构有一定治疗作用。     

miR-152-3p调控Notch1/DLL4通路对家兔深II度烧伤创面血管生成的影响

摘要 目的：探究微小核糖核酸（miR）-152-3p调控果蝇Notch同源物1（Notch1）/Delta样配体4（DLL4）通路对家兔深II度烧伤创面血管生成的影响。方法：将50只新西兰家兔随机分为对照组、模型组、miR-152-3p拮抗剂（antagomir）组、miR-152-3p antagomir阴性对照+空载组、miR-152-3p antagomir+Notch1敲低组,每组10只,除对照组外其余各组家兔构建深II度烧伤模型,分组给药处理后,实时荧光定量聚合酶链式反应（qRT-PCR）检测各组家兔创面组织miR-152-3p与Notch1、DLL4 mRNA表达;检测各组家兔创面愈合率及微循环血流灌注值（MPD）;免疫组织化学染色检测各组家兔创面微血管密度（MVD）;酶联免疫吸附反应（ELISA）检测各组家兔血清血管内皮细胞生长因子（VEGF）及促血管生成素1（Ang1）水平;免疫印迹检测各组家兔创面组织VEGF、Ang1与Notch1/DLL4通路蛋白表达;双荧光素酶报告基因实验检测兔脐静脉内皮细胞中miR-152-3p对Notch1及DLL4的靶向调节。结果：与对照组相比,模型组家兔创面组织miR-152-3p与Notch1、DLL4 mRNA表达升高（P<0.05）,创面MPD及MVD、血清VEGF及Ang1水平、创面组织VEGF与Ang1蛋白表达降低（P<0.05）。与模型组相比,miR-152-3p antagomir组家兔创面组织miR-152-3p mRNA表达降低（P<0.05）,创面愈合率、创面MPD及MVD、血清VEGF及Ang1水平、创面组织Notch1、DLL4 mRNA及蛋白表达、创面组织VEGF与Ang1蛋白表达升高（P<0.05）;miR-152-3p antagomir阴性对照+空载组家兔各指标无明显差异（P＞0.05）;与miR-152-3p antagomir组相比,miR-152-3p antagomir+Notch1敲低组家兔创面组织miR-152-3p mRNA表达无明显差异（P＞0.05）,创面愈合率、创面MPD及MVD、血清VEGF及Ang1水平、创面组织Notch1、DLL4 mRNA及蛋白表达、创面组织VEGF与Ang1蛋白表达降低（P<0.05）。miR-152-3p可靶向下调兔脐静脉内皮细胞中Notch1及DLL4的表达。结论：敲低miR-152-3p可通过上调Notch1/DLL4通路而增强家兔深II度烧伤创面血管生成,进而促进其创面愈合。     

皮肤创伤愈合过程中短暂扩增细胞增生及皮肤干细胞迁移

目的研究皮肤疤痕组织形成过程中皮肤干细胞分布、增殖分化迁移特征,初步探讨这些特征与皮肤创伤修复的关系。方法利用眼科显微外科剪对2日龄昆明小鼠背部皮肤进行人工统一造创,定期获取皮肤创面样品,常规病理染色观察创面愈合形态;应用免疫荧光染色法,以细胞转录因子Sox2和角蛋白14抗体分别检测皮肤干细胞的分布及干细胞增殖分化时所形成的短暂扩增细胞,并结合细胞增殖EdU荧光染色初步分析皮肤干细胞的迁移方向。结果创面愈合过程中表皮层中表达Sox2的阳性细胞逐渐连贯,并且发现真皮乳头层中Sox2和角蛋白14同时大量表达,可见致密细胞网和向下凸起的新生毛囊样结构形成。同时,创面愈合初期细胞迁移主要由创面底部开始,向上迁移并填充创面。结论创面愈合过程中,创面底部皮肤干细胞首先开始大量分裂增殖,并向创面迁移,创面上部皮肤干细胞分裂增殖迟于创面底部;迁移的皮肤干细胞以不对称分裂的形式增殖形成大量短暂扩增细胞,并在增厚的疤痕乳头层部位形成毛囊样结构填充皮肤疤痕。     

脱细胞小肠黏膜下层与脱细胞心包修复大鼠腹壁缺损的对比研究

目的：观察小肠黏膜下层（small intestinal submucosa,SIS）和脱细胞心包（pericardium,PC）修复大鼠腹壁缺损的效果,比较两种生物材料相容性。方法：SD大鼠40只,体重200～250g,手术造成3 cm×2 cm全层腹壁缺损,随机分为二组（n=20）,分别采用相同面积的小肠黏膜下层（small intestinal submucosa,SIS）和脱细胞真皮基质（acellular dermal matr,ADM）补片进行修补。术后1、2、4和8周分批取出腹壁修复材料,行动物一般情况观察、腹腔内粘连情况评价、力学强度测定及组织学观察。结果：术后动物都成活,两种材料术后8周均无疝瘘发生,缺损得到完整修复。术后各期SIS组的腹腔粘连评分明显低于PC组。术后4、8周,SIS组力学强度强于PC组,有统计学意义;组织学观察两组未见明显免疫排斥反应,SIS组的组织再生和重塑、血管化优于PC组;术后炎症反应两组无明显差异。结论：SIS和PC均能修复大鼠腹壁全层缺损,SIS在生物相容性方面优于PC。     

Small intestinal submucosa in abdominal wall repair after TRAM flap harvesting in a rat model

The strength of porcine small intestinal submucosa in abdominal wall repair after transverse rectus abdominis myocutaneous flap harvesting was examined in a rat model. Changes in the levels of selected molecular markers of inflammation after small intestinal submucosa implantation were also studied. Eighty-three rats were divided into three groups. In experimental group I, an abdominal wall defect created by removal of the rectus abdominis muscle was repaired with placement of a 1.5 x 5-cm2 patch of small intestinal submucosa. In experimental group II, the muscle defect was repaired with a combination of small intestinal submucosa patch placement and fascial closure. In the control group, the defect was repaired with direct fascial closure. At postoperative times of 3 days, 2 weeks, 1 month, and 2 months, the muscle tissues adjacent to the abdominal wall repair site were subjected to biopsies for assessment of inflammation markers. Full-thickness sections of the abdominal wall from the repair site in each animal were removed for tensile strength testing and histological examinations. The results demonstrated that interleukin-6 and interferon-gamma levels were increased in the two experimental, small intestinal submucosa-treated groups at 3 days and 2 weeks postoperatively. The results of mechanical testing demonstrated that the average tensile strength of the repaired abdominal wall in the repair model with combined small intestinal submucosa placement and fascial repair was significantly greater than the values for repairs with fascial closure or small intestinal submucosa placement alone. The use of small intestinal submucosa placement in combination with fascial repair can significantly improve the strength of the repaired abdominal wall after transverse rectus abdominis myocutaneous flap harvesting.     

Evaluation of Dermal Substitute in a Novel Co-Transplantation Model with Autologous Epidermal Sheet

The development of more and more new dermal substitutes requires a reliable and effective animal model to evaluate their safety and efficacy. In this study we constructed a novel animal model using co-transplantation of autologous epidermal sheets with dermal substitutes to repair full-thickness skin defects. Autologous epidermal sheets were obtained by digesting the basement membrane (BM) and dermal components from rat split-thickness skins in Dispase II solution (1.2 u/ml) at 4°C for 8, 10 and 12 h. H&E, immunohistochemical and live/dead staining showed that the epidermal sheet preserved an intact epidermis without any BM or dermal components, and a high percentage of viable cells (92.10±4.19%) and P63 positive cells (67.43±4.21%) under an optimized condition. Porcine acellular dermal matrixes were co-transplanted with the autologous epidermal sheets to repair full-thickness skin defects in Sprague-Dawley rats. The epidermal sheets survived and completely re-covered the wounds within 3 weeks. Histological staining showed that the newly formed stratified epidermis attached directly onto the dermal matrix. Inflammatory cell infiltration and vascularization of the dermal matrix were not significantly different from those in the subcutaneous implantation model. Collagen IV and laminin distributed continuously at the epidermis and dermal matrix junction 4 weeks after transplantation. Transmission electron microscopy further confirmed the presence of continuous lamina densa and hemidesmosome structures. This novel animal model can be used not only to observe the biocompatibility of dermal substitutes, but also to evaluate their effects on new epidermis and BM formation. Therefore, it is a simple and reliable model for evaluating the safety and efficacy of dermal substitutes.     

Vascularized acellular dermal matrix island flaps for the repair of abdominal muscle defects

The potential widespread use of tissue-engineered matrices in soft-tissue reconstruction has been limited by the difficulty in fabricating and confirming a functional microcirculation. Acellular dermal matrix placed in a soft-tissue pocket acts as a scaffold to be incorporated by the host's fibrovascular tissue. A new method for noninvasive real-time observation of functional microvascular networks using orthogonal polarization spectral (OPS) imaging has recently been reported. Arterioles, venules, and capillaries can be directly visualized, and the movement of individual blood cells through them can be observed. The present study was performed to investigate the use of prefabricated acellular dermal matrix with an arteriovenous unit for the repair of abdominal muscle defects. OPS imaging was used to determine the presence of a functional microcirculation in the neovascularized matrix. In Sprague-Dawley rats, vascularized matrix was prefabricated by placing the superficial epigastric artery and vein on a 2-cm x 2-cm implant-type acellular dermal matrix in the thigh. Three weeks after implantation, the matrix-arteriovenous unit was elevated as an axial-type flap and a 2-cm x 2-cm full-thickness block of abdominal muscle immediately superior to the inguinal ligament was resected. Additional procedures were performed according to group: no repair (group 1, n = 20); repair with nonvascularized acellular dermal matrix (group 2, n = 20); repair with devascularized acellular dermal matrix (group 3, = 20); and repair with vascularized acellular dermal matrix (group 4, n = 20). OPS imaging (field of view, 1 mm in diameter; scan depth range, 0.2 mm) was performed on both sides of each flap on a total of 10 random distal regions before and after pedicle transection in group 3 and with the pedicle preserved in group 4. Hernia rate and duration of survival were compared for 21 days. OPS imaging showed directional blood cell movement through the capillary network in all areas scanned in group 4. No microvascular perfusion was observed after pedicle transection in group 3. Hernia rates of 100, 80, 90, and 0 percent were seen in groups 1, 2, 3, and 4, respectively. Median survival times of 9, 11.5, 9, and 21 postoperative days were noted in groups 1, 2, 3, and 4, respectively. Histopathologic analysis with factor VIII revealed full-thickness infiltration of the matrix by endothelial cells, signifying newly formed blood vessels. Repair of abdominal muscle defects using vascularized acellular dermal matrix resulted in no hernia and survival of all animals for the duration of study. However, repairs using avascular or devascularized matrix resulted in significant rates of hernia and decreased survival. Acellular dermal matrix can be prefabricated into vascularized tissue using an arteriovenous unit and used successfully to repair abdominal muscle defects. OPS imaging allowed for high-contrast direct visualization of microcirculation in previously acellular tissue following prefabrication with an arteriovenous unit.     

Composite thin film and electrospun biomaterials for urologic tissue reconstruction

A replacement material for autologous grafts for urinary tract reconstruction would dramatically reduce the complications of surgery for these procedures. However, acellular materials have not proven to work sufficiently well, and cell‐seeded materials are technically challenging and time consuming to generate. An important function of the urinary tract is to prevent urine leakage into the surrounding tissue—a function usually performed by the urothelium. We hypothesize that by providing an impermeable barrier in the acellular graft material, urine leakage would be minimized, as the urothelium forms in vivo. However, since urothelial cells require access to nutrients from the supporting vasculature, the impermeable barrier must degrade over time. Here we present the development of a novel biomaterial composed of the common degradable polymers, poly(ε‐caprolactone) and poly(L ‐lactic acid) and generated by electrospinning directly onto spin‐coated thin films. The composite scaffolds with thin films on the luminal surface were compared to their electrospun counterparts and commercially available small intestinal submucosa by surface analysis using scanning electron microscopy and by analysis of permeability to small molecules. In addition, the materials were examined for their ability to support urothelial cell adhesion, proliferation, and multilayered urothelium formation. We provide evidence that these unique composite scaffolds provide significant benefit over commonly used acellular materials in vitro and suggest that they be further examined in vivo. Biotechnol. Bioeng. 2011; 108:207–215. © 2010 Wiley Periodicals, Inc.     

Histochemistry and morphology of porcine mast cells

Summary Mast cells have been described extensively in rodents and humans but not in pigs, and the objective of this study was to characterize porcine mast cells by histochemistry and electron microscopy. Carnoy's fluid proved to be a good fixative but fixation with neutral buffered formalin blocked staining of most mast cells. Alcian Blue stained more mast cells than did Toluidine Blue (pH 0.5), although Alcian Blue also stained goblet cells. In pigs, unlike rodents, the Alcian Blue method did not distinguish between mast cells in the intestinal mucosa and those in the connective tissue of the intestinal submucosa, tongue and skin. Mast cells were significantly larger in adult pigs than in piglets; in adult pigs and piglets, mast cells in the intestinal mucosa were significantly larger than those in submucosal connective tissue, and they were more varied in shape in piglets and adults. Granules in mast cells in the intestinal mucosa stained less intensely than those in mast cells in connective tissue of tongue, skin and intestinal submucosa. Mast cells in the connective tissue of the tongue, skin and intestinal submucosa fluoresced strongly when stained with berberine sulphate or with a mixture of berberine sulphate and Acridine Orange, but mast cells in the intestinal mucosa did not. All mast cells reacted positively in an enzyme-histochemical method previously used to detect human tryptase but not in a method previously used to detect human chymase. Mast cells in the medulla of thymus stained similarly to mast cells in the intestinal mucosa. Ultrastructural differences between mast cells were not detected.     

Conservative treatment of cutis aplasia

Three cases of cutis aplasia are presented. The defects involved include full-thickness defects of scalp and cranium as well as full-thickness skin defects of the abdomen and thigh. All patients were treated conservatively with the use of Silvadene cream dressings. Healing was obtained in all patients.     

Histological and mechanical evaluation of antifreeze peptide (Afp1m) cryopreserved skin grafts post transplantation in a rat model

The objective of this study was to evaluate the use of Afp1m as a cryopreservative agent for skin by examining the transplanted skin histological architecture and mechanical properties following subzero cryopreservation. Thirty four (34) rats with an average weight of 208 ± 31 g (mean ± SD), were used. Twenty four (n = 24) rats were equally divided into four groups: (i) immediate non-cryopreserved skin autografts (onto same site), (ii) immediate non-cryopreserved skin autografts (onto different sites), (iii) skin autografts cryopreserved with glycerol for 72 h and (iv) skin autografts cryopreserved with Afp1m for 72 h at −4 °C. Rounded shaped full-thickness 1.5–2.5 cm in diameter skin was excised from backs of rats for the autograft transplantation. Non-cryopreserved or cryopreserved auto skin graft were positioned onto the wound defects and stitched. Non-transplanted cryopreserved and non-cryopreserved skin strips from other ten rats (n = 10) were allowed for comparative biomechanical test. All skin grafts were subjected to histological and mechanical examinations at the end of day 21. Histological results revealed that tissue architecture especially the epidermal integrity and dermal-epidermal junction of the Afp1m cryopreserved skin grafts exhibited better histological appearance, good preservation of tissue architecture and structural integrity than glycerolized skin. However, there was no significant difference among these groups in other histological criteria. There were no significant differences among the 4 groups in skin graft mechanical properties namely maximum load. In conclusion, Afp1m were found to be able to preserve the microstructure as well as the viability and function of the skin destined for skin transplantation when was kept at −4 °C for 72 h.     

The acquirement of growth ability for third-stage larvae of Strongyloides ratti during the head-passage in the rat

The role of larval passage through the head in the course of the migration of Strongyloides ratti in rats was investigated. Third-stage larvae (L3) recovered from various portions of donor rats were re-injected into the skin, cranial cavity and small intestine of recipient rats to check their ability for further growth. Cultured L3 (L3c) and the L3 recovered from the skin of donor rats (L3s) did not survive in the small intestine after intestinal inoculation. However, intestinal inoculation of L3 recovered from the head of donor rats (L3h) revealed growth to the adult stage. Cultured L3 injected into the cranial cavity of rats also became adult worms in the small intestine. L3 incubated in the cranial cavity for more than 24 h could grow in the small intestine of the recipient rats. These experiments suggest that S. ratti L3 acquire their ability to mature in the small intestine during their migration through the head of rats.