创面愈合过程中创缘表皮干细胞的异位

目的观察创缘表皮干细胞在全层皮肤创面愈合过程中的分布特征,初步探讨其在创面愈合过程中的作用.方法将已行BrdU活体标记的 20只Wistar大鼠背部制备4个为2.54 cm2的全层皮肤创面,分别于伤后3、7、14和21 d(n=5)行组织学检查,动态观察创面愈合情况,并行β1整合素、角蛋白19(K19)与BrdU免疫组织化学法检测表皮干细胞在创面愈合过程中的分布情况.结果创面愈合率为83.75%(61/80).所有创面肉芽组织于各时相点均未见β1整合素、K19阳性细胞出现,但于创缘表皮的棘层或颗粒层均出现散在的β1整合素、K19和BrdU阳性细胞.且越接近创面阳性细胞越密集,组织学上与基底层的阳性细胞无直接联系;其数量随创面的缩小逐渐增加,直至创面愈合.创面上皮化后,阳性细胞逐渐减少,并随愈合创面表皮脚的出现而消失.而感染创面的阳性细胞数量明显少于未感染创面.结论表皮干细胞能主动参与创面的修复,创缘的表皮干细胞异位的主要功能可能是促进创面再上皮化.     

表皮干细胞在糖尿病创面愈合过程中的动态变化

目的 观察糖尿病（diabetes mellitus，DM）大鼠创伤后不同时期表皮厚度、表皮干细胞（epidermalste mcells，ESCs）数量变化和分布特征及创面愈合率等动态变化，探讨ESCs与DM皮肤创面难愈之间的关系。方法 48只Wistar大鼠，雄性，体重180～200g。随机分为DM组和正常对照组，各24只。DM组大鼠制备DM大鼠慢性创面愈合模型。两组大鼠同时用特制打孔器在大鼠背部脊柱两侧约1．5cm制作直径1．8cm、面积2．54cm。的全层皮肤缺损（共计96个创面），分别在致伤后第3、7、14和21天拍摄创面，计算创面愈合率；取创缘及肉芽组织，行HE染色及角蛋白19（keratin19，K19）和β1整合素抗体免疫组织化学染色，显微图像分析系统测量表皮厚度、阳性部分面积及灰度值。结果 致伤后第3、7、14和21天，正常对照组大鼠创面愈合率分别为24．48％±3．37％、50．46％±1．26％、92．82％±2．12％和99．41％±0．66％，而DM组分别为2．43％±1．02％、40．59％±1．65％、80．77％±3．57％和85．40％±0．94％，两组同时期比较差异有统计学意义（P〈0．01）。致伤后第3、7、14和21天，正常对照组表皮厚度分别为26．43±3．21、33．29±3．52、31．53±3．35和26．01±3．19p．m，DM组表皮厚度分别为23．58±2．33、31．02±3．38、33．72±5．49和21．80±4．02p．m，致伤后第3、21天，二者比较差异有统计学意义（P〈0．01）。致伤后第3、7、14和21天，正常对照组K19染色的平均阳性单位（positiveunit，PU）值分别为91．68％、93．14％、72．27％和70．31％，而DM组分别为40．29％、40．79％、29．94％和10．37％；正常对照组β1整合素染色的平均PU值分别为49．60％、91．16％、77．13％和57．17％，DM组分别为38．94％、24．16％、61．36％和38．83％，与正常对照组同时期比较，DM组K19和β1整合素染色的平均PU值都降低，且差异有统计学意义（P〈0．05）。结论 ESCs数量少和活性低可能是DM创面难愈的重要机制之一。     

SDF-1对创缘表皮干细胞定向趋化及促创面愈合效应的研究

目的：观察干细胞趋化因子SDF-1在创面愈合过程中调控表皮干细胞定向迁移促进创面愈合的过程。方法：制作大鼠背部全层皮肤缺损模型,随机分为三个实验组：①SDF-1组;②AMD3100（CXCR4受体的拮抗剂）组;③空白对照组。分别于致伤后1天、3天、5天、7天和12天照相计算伤后不同时间创面占初始创面的百分比并取材。运用HE染色和免疫组化技术观察创缘表皮干细胞定向迁移分化的特点,寻找其规律。结果：与其他两个处理组相比,SDF-1处理组愈合时间缩短,其创缘皮表皮基底层细胞分裂增殖旺盛,创缘表皮干细胞数目明显多于其他两组。结论：在创面愈合过程中,SDF-1可以调控表皮干细胞向创缘迁移,加速创面上皮化。     

不同来源的表皮干细胞促进皮肤创面愈合的研究

目的 观察表皮干细胞(ESCs)及去分化来源的表皮干细胞(DESCs)对皮肤创面愈合的促进作用及其异同.方法 由新鲜人包皮标本以贴壁法分离ESCs,随机分为3组,第1组作为原代ESCs:第2组传代培养至第6代时可见细胞成熟呈表皮细胞(ECs)形态,由原来的大克隆生长的小圆细胞形态变为较为肥大的不规则型细胞,无法形成克隆样增殖;其细胞表型也由β1整合素、CK19强阳性、CK10阴性变为β1整合素、CK19阴性,而CK10强阳性,予以100μg/L的碱性成纤维细胞生长因子(bFGF)培养48 h进行去分化诱导,48 h后ECs周边开始出现小圆细胞并呈克隆样生长,β1整合素、CK19、CK10等细胞表型也趋于与ESCs一致,即DESCs;第3组传代培养至第6代获得ECs.3组细胞皆以D-Hanks液重悬调其终质量浓度为2×106个/ml.将12只裸鼠每只背部左右各制备1个直径6 mm创面,共24个创面.将全部创面随机等分为4组,每组6个,分别以ESCs、DESCs、Ecs及生理盐水(NS)各0.2 ml进行创面注射.于术后0、3、7 d测量创面面积进行统计学分析,并于术后7 d行创面取材苏木素-伊红(HE)染色.结果 ESCs组与DESCs组在术后3 d时创面面积分别为(11.758±2.544)、(11.515±1.351)mm2,7 d时创面面积分别为(1.795±1.063)、(2.043±1.138)mm2,修复速度要明显快于ECs组与NS组[3 d时创面面积分别为(17.857±1.722)、(16.192±2.256)mm2,7 d时创面面积分别为(5.367±1.219)、(5.070±1.357)mm2,P＜0.01];而ESCs与DESCs组之间创面修复速度比较差异无统计学意义(P＞0.05);ECs组与NS组之间比较差异无统计学意义(P＞0.05).HE染色提示ESCs组与DESCs组的创面愈合质量优于ECs组与NS组,可见有皮肤附属腺的再生且纤维瘢痕组织较少.结论 ESCs与DESCs皆有促进创面愈合的作用,相互之间无明显差异.

Abstract：

Objective To investigate the enhancing effect of epithelial stem cells (ESCs) and dedifferentiation derived epithelial stem cells (DESCs) in the healing of epidermal wound. Methods ESCs were isolated from the fresh circumcised foreskins by using adherence method and randomly divided into three cohorts: ESCs cohort, DESCs cohort and epithelial cells (ECs) cohort. The final cell density was adjusted to 2 × 106/ml with D-Hanks in each group. The model of BALB/C mice full-thickness skin loss wounds was established. Two circular 6 mm-diameter skin loss wounds were cut on every BALB/C mouse back. Twenty-four wounds of 12 BALB/C mice were divided equally and randomly into 4 groups:DESCs, ESCs, ECs and NS. The cell suspensions of DESCs, ESCs and ECs were injected respectively into the wound edges with the density of 2 × 106/ml. And the volme was 0. 2 ml per treatment wound. The same volume of normal saline was injected in NS group. On the postoperative day 0, 3, 7, all of the wound areas were measured and analyzed statistically. On the postoperative 7 day, the sections were made and observed by using hematoxylin and eosin (HE) staining. Results The wound areas in DESCs group and ESCs group were ( 11. 758 ± 2. 544) and ( 11.515 ± 1.351 ) mm2 on the 3rd day postoperatively, and ( 1. 795 ±1. 063) and (2. 043 ± 1. 138) mm2 on the 7th day postoperatively, respectively. The wound areas in ECs group and NS group were ( 17. 857 ± 1. 722) and ( 16. 192 ±2. 256) mm2 on the 3rd day postoperatively,and ( 1. 795 ± 1. 063) and (2. 043 ± 1. 138) mm2 on the 7th day postoperatively, respectively. There was statistically significant difference in wound area between ESCs or DESCs group and ECs, NS groups on the 3rd, 7th day postoperatively ( P ＜ 0. 01 ), but there was no statistically significant difference between DESCs and ESCs groups ( P ＞ 0. 05 ). HE staining showed the repairing quality of treatment wounds was superior to that in the control wounds, and regenerating glands and less fibrous connective tissues were seen in ESCs and EDSCs groups. Conclusion Both ESCs and DESCs could promote the wound repair.     

人脂肪干细胞与表皮生长因子促进皮肤创面愈合的研究

目的:观察人脂肪干细胞(hASCs)与人表皮生长因子(hEGF)对皮肤缺损创面修复的影响。方法:选择郑州大学第二附属医院2020年1月6例患者抽脂减肥的脂肪组织作为hASCs的来源,以酶消化法从人脂肪组织中提取hASCs,将其培养至第3代。利用倒置显微镜对细胞形态进行观察;使用流式细胞仪鉴定细胞表型及分化能力检测。随机...     

Rac1蛋白调控表皮干细胞迁移促进创面愈合的研究

目的 观察Rac1蛋白对表皮干细胞(ESC)迁移运动的调控作用,为完善创面愈合的基础理论以及指导临床应用提供参考.方法 将慢病毒空载体FUGW单独和分别与Rac1蛋白抑制型突变体Rac1T17N、Rac1蛋白活化型突变体Rac1Q61L融合后转染入ESC,按照随机数字表法分为3部分进行如下实验.(1)将ESC分别接种于Ⅰ型胶原溶液(20 μg/mL)、Ⅳ胶原溶液(20 μg/mL)或纤连蛋白溶液(10 μg/mL)包被的24孔细胞培养板,采用CytoTox 96 比色试剂盒检测ESC对不同基质的黏附率.(2)选取上述黏附于Ⅳ型胶原的1000个ESC,四甲基异硫氰酸罗丹明标记的鬼笔环肽染色,激光扫描共聚焦显微镜下观察黏附细胞的形态延展并比较面积大小.(3)选用Transwell小室,上室加入ESC、下室中加入含基质细胞衍生因子1(SDF-1)的限定性角质形成细胞无血清培养液(以不加SDF-1的培养液为对照),倒置相差显微镜下观察ESC的趋化能力,结果以细胞迁移变化率表示.(4)将ESC接种于6孔细胞培养板孵育12 h,加入含4 μg/mL丝裂霉素C的培养液继续孵育2 h,于单层贴壁细胞划痕,6、12 h后统计剩余划痕宽度百分率.对数据进行t检验.结果 与转染FUGW的ESC比较,转染Rac1Q61L的ESC对Ⅰ型胶原的黏附率明显增加(t＝5.302,P<0.05),转染Rac1T17N的ESC对Ⅰ型胶原(t＝13.741,P<0.05)、Ⅳ型胶原(t＝15.676,P<0.05)及纤连蛋白(t＝8.256,P<0.05)的黏附率均明显下降.激光扫描共聚焦显微镜下观察,与转染FUGW的ESC比较,转染Rac1Q61L的ESC面积明显增大,细胞边缘有层板状伪足伸出;转染Rac1T17N的ESC面积显著缩小.在趋化因子SDF-1作用下,与转染FUGW的ESC比较,转染Rac1Q61L的ESC迁移变化率升高43.4%,转染Rac1T17N的ESC迁移变化率下降78.0%;无SDF-1作用时,与转染FUGW的ESC比较,转染Rac1T17N的ESC迁移变化率下降55.2%,转染Rac1Q61L的ESC迁移变化率未见明显变化(升高1.7%).划痕后6、12 h,转染Rac1Q61L的ESC剩余划痕宽度百分率分别为(39±9)%、(6±5)%,低于转染FUGW的ESC[(43±5)%,t=1.027,P>0.05;(18±7)%,t=4.389,P<0.05];划痕后6、12 h,转染Rac1T17N的ESC剩余划痕宽度百分率分别为(81±9)%、(71±11)%,明显高于转染FUGW的ESC(t值分别为11.386、11.726,P值均小于0.05).结论 Rac1蛋白可通过影响ESC的黏附、延展以及趋化能力调控细胞迁移,并可能因此参与ESC促进创面愈合的进程.

Abstract：

Objective To investigate modulatory role of Rac1 protein in epidermal stem cell (ESC) migration during wound healing,in order to provide a reference for enriching basic theory of wound healing and guiding clinical application. Methods Constitutively active mutant of Rac1 protein (Rac1Q61L) or dominant negative isoform of Rac1 protein (Rac1T17N) was transfected into ESC using a retroviral vector FUGW,and retroviral vector FUGW transfected into ESC in singles was used as blank control. The cells were divided into 3 parts according to the random number table and treated as follows. First,equal numbers of cells were inoculated into 24-well plates coated with collagen Ⅰ (20 μg/mL),collagen Ⅳ (20 μg/mL) or fibronectin (10 μg/mL). Cells adhered to above matrices were quantitated using CytoTox 96 colorimetric kit. Second,1000 cells adhered to collagen Ⅳ,after being stained with tetramethyl rhodamine isothiocyanate-phalloidin,were collected for observation of cell morphology and comparison of spreading area under confocal laser scanning microscope. Third,ESC with density of 2×105 cells per well were placed in upper compartment of Transwell chamber,DK-SFM culture medium alone or that containing stromal cell derived factor 1 (SDF-1) was added into lower compartment of Transwell chamber. Migration of ESC was observed using inverted phase contrast microscope,and the result was denoted as migration rate. Lastly,ESC with density of 7.5×105 cells per well was inoculated into 6-well plates for 12 hours,and treated with 4 μg/mL mitomycin C for 2 hours. The remaining scratch width of monolayer was respectively measured 6 hours or 12 hours after scratching to calculate the percentage of remaining scratch width. Data were processed with t test.Results Compared with that of blank control,the number of Rac1Q61L-transfected cells adhered to collagen Ⅰ was significantly increased (t＝5.302,P<0.05),while the number of Rac1T17N-transfected cells adhered to collagen Ⅰ,Ⅳ,and fibronectin were all obviously decreased (with t value respectively 13.741,15.676,8.256,P values all below 0.05). Confocal laser scanning microscope showed that spreading area of Rac1Q61L-transfected ESC (with laminate pseudopodia on edge) and Rac1T17N-transfected ESC was respectively larger and smaller as compared with that of blank control. With SDF-1 effect,the migration rate of Rac1T17N-transfected ESC was decreased by 78.0% and Rac1Q61L-transfected ESC was increased by 43.4% as compared with that of blank control. Without SDF-1 effect,the migration rate of Rac1T17N-transfected ESC was decreased by 55.2%,while the migration rate of Rac1Q61L-transfected ESC was close to that of blank control. Six or 12 hours after scratching,the percentage of remaining scratch width in Rac1Q61L-transfected ESC was lower as compared with that in blank control[(39±9)% vs. (43±5)%,(6±5)% vs. (18±7)%,with t value respectively 1.027,4.389,with P value respectively above and below 0.05],while that in Rac1T17N-transfected ESC[(81±9)%,(71±11)%,respectively]was obviously higher as compared with that in blank control (with t value respectively 11.386,11.726,P values all below 0.05). Conclusions Rac1 protein may control the migration of ESC by regulating its adhesion,spreading,and chemotaxis,and it plays an active role in wound healing accelerated by ESC.     

创面愈合过程中表皮生长因子及其受体变化的临床研究

创面愈合是一个极其复杂的病理过程,涉及了机体许多病理变化,如出血凝血、修复重建等。许多研究表明：生长因子可以促进创面愈合,加快组织的修复［1］。笔者曾研究过大鼠供皮区创面愈合过程中内源性表皮生长因子（epidermal growth factor, EGF）、表皮生长因子受体（epidermal growth factor receptor,EGFr）有明显的表达,其变化与创面修复有关［2,3］。本实验通过研究人供皮区创面愈合过程中EGF、EGFr变化,探讨其与组织修复的关系。资料与方法1.标本留取：选取烧伤或整形患者中厚断层供皮区创面共22例,男15例,女7例,年龄18…     

脂肪干细胞促进人表皮角质形成细胞创面模型愈合的研究

目的 探讨脂肪干细胞(ADSCs)促进人表皮角质形成细胞(HEKa)创面模型愈合的可能性.方法 体外培养大鼠ADSCs(rADSCs)(n=10).实验组为rADSCs与HEKa直接共培养组(n=10),对照组为在transwell培养板建立的rADSCs与HEKa间接共培养(间接组,n=8)和单纯HEKa培养组(单纯组,n=8).刮擦生长融合成片的HEKa细胞制备创面.培养24、48、72 h后,计数迁移到创面的HEKa细胞数量并计算创面愈合率,检测HEKa细胞吸收脱氧胸腺嘧啶核苷的放射活性以判定HEKa细胞的增殖活性.结果 实验组、间接组、单纯组在伤后24 h分别有(9.2±0.2)、(5.0±0.3)、(4.2±0.3)个细胞/高倍视野;48 h分别有(58.5±0.4)、(26.5±0.3)、(20.7±0.5)个细胞/高倍视野;72 h分别有(125.8±0.4)、(43.0±0.5)、(35.6±0.5)个细胞/高倍视野越过创缘进入创面,实验组均明显优于两对照组(P<0.05).三组创面愈合率在伤后72 h分别为61.0% ±3.0% 、35.0% ±2.5% 、32.0% ±2.1% ,实验组均明显优于两对照组(P<0.05).脱氧胸腺嘧啶核苷掺入培养基法表明三组HEKa细胞的放射性强度为(1440±210)、(1050±280)、(1130±390)cpm/105细胞,实验组与两对照组之间差异有统计学意义(P<0.05).结论 rADSCs通过直接接触促进HEKa细胞的分裂增殖和迁移.     

毛囊干细胞与创面愈合研究进展

目前临床上广泛借助各种技术,包括自异体皮,培养的皮肤细胞和胶原基质制作组织工程皮肤等,用以治疗溃疡、创伤以及大面积烧伤.创面愈合除表皮覆盖外,另一问题就是皮肤附件结构的重建.     

表皮生长因子促进皮肤创面愈合的研究

为了观察外源性表皮生长因子对创面愈合的影响，在３０例大鼠皮肤创面愈合模型中，局部外用表皮生长因子（ＥＧＦ），以软膏基质作为对照，分别于１，２周测量创面面积，测定创面组织中的ＤＮＡ、蛋白质及羟脯氨酸含量，记录创面完全愈合时间。结果表明，实验组与对照组创面完全愈合的时间分别为（１４．６±１．２）天和（１８．５±２．０６）天（Ｐ＜０．０１），ＥＧＦ能显著增加组织中的ＤＮＡ、蛋白质及羟脯氨酸含量（Ｐ＜０．０１）。结果提示，ＥＧＦ能显著加速创面的修复，缩短愈合时间。     

离体冻伤模型创面愈合过程中基质细胞衍生因子1促表皮干细胞迁移的研究

Objective To study the promotion effect of stromal cell-derived factor 1 (SDF-1) on the migration of epidermal stem cells (ESC) in the healing process of frostbite-wound model ex vivo. Methods A three-dimensional model of full-thickness frostbite of skin was constructed (with slot-like wound) out of skin equivalent. The expression of SDF-1 in wound stroma was observed with immunohisto-chemistry staining on post injury days (PID) 3 and 7. The model frostbite wounds were divided into control group (treated with PBS 50 μL per wound) , SDF-1 group (treated with 100 ng/mL SDF-1, 50 μL per wound) , and AMD3100 group [treated with 100 ng/mL AMD3100 (50 μL per wound) for 30 minutes, and then SDF-1 50 μL was added per wound]. The redistribution of ESC around wound was observed. Results The expression of SDF-1 in wound stroma increased gradually on PID 3 and 7. Compared with those in control and AMD3100 groups, there were more ESC and epithelial cell layers, and more integrin β1-positive cells appeared at the basal layer of wound in SDF-1 group, and some of the positive cells migrated upward to epidermis. Conclusions SDF-1 contributes to wound repair through promoting ESC to migrate toward and gather around wound edge. This may be one of the mechanisms of ESC participating in wound repair.     

离体冻伤模型创面愈合过程中基质细胞衍生因子1促表皮干细胞迁移的研究

Objective To study the promotion effect of stromal cell-derived factor 1 (SDF-1) on the migration of epidermal stem cells (ESC) in the healing process of frostbite-wound model ex vivo. Methods A three-dimensional model of full-thickness frostbite of skin was constructed (with slot-like wound) out of skin equivalent. The expression of SDF-1 in wound stroma was observed with immunohisto-chemistry staining on post injury days (PID) 3 and 7. The model frostbite wounds were divided into control group (treated with PBS 50 μL per wound) , SDF-1 group (treated with 100 ng/mL SDF-1, 50 μL per wound) , and AMD3100 group [treated with 100 ng/mL AMD3100 (50 μL per wound) for 30 minutes, and then SDF-1 50 μL was added per wound]. The redistribution of ESC around wound was observed. Results The expression of SDF-1 in wound stroma increased gradually on PID 3 and 7. Compared with those in control and AMD3100 groups, there were more ESC and epithelial cell layers, and more integrin β1-positive cells appeared at the basal layer of wound in SDF-1 group, and some of the positive cells migrated upward to epidermis. Conclusions SDF-1 contributes to wound repair through promoting ESC to migrate toward and gather around wound edge. This may be one of the mechanisms of ESC participating in wound repair.     

离体冻伤模型创面愈合过程中基质细胞衍生因子1促表皮干细胞迁移的研究

目的 了解离体皮肤冻伤模型创面愈合过程中,基质细胞衍生因子1(SDF-1)对创缘表皮干细胞(ESC)的运动趋化作用.方法 体外构建三维皮肤等价物全层冻伤模型(创面呈孔形),免疫组织化学染色观察冻伤后3、7 d创缘间质内SDF-1的表达情况.另将冻伤模型分为对照组(创面区添加PBS 50 μL/孔)、SDF-1组(创面区添加100 ng/mL SDF-1,50μL/孔)和AMD3100组[创面区用100 ng/mL AMD3100(50μL/孔)处理30 min后,添加SDF-1 50μL/孔],观察各组创缘中ESC的分布变化.结果 免疫组织化学染色显示,冻伤后3、7 d创缘间质内SDF-1的表达逐渐增加.与对照组及AMD3100组相比,SDF-1组创缘基底层整合素β1阳性细胞数量增多,且部分阳性细胞向上层表皮迁移聚集,创缘ESC数量更多,表皮细胞层数增加.结论 SDF-1促进ESC向创缘迁移聚集参与创面修复,这可能是ESC参与创面修复过程的机制之一.     

局部应用表皮细胞生长因子促进创面愈合

表皮细胞生长因子(EGF)是由53个氨基酸组成的多肽类物质。它能刺激多种细胞mRNA、mDNA和蛋白质的合成,除了能刺激体外皮肤表皮细胞的分裂以外,它还能促使二度烧伤和皮肤供皮区表皮的再生和真皮的增殖,增加外科切口的张力强度。调节正常表皮再生的分子水平机理尚不清楚,有可能这类肽类生长因子对自体分泌或旁分泌起了重要作用。作者用双盲法作前瞻性随机临床试验以评价EGF对供皮区愈合的效果。糖尿病、癌症、妊娠、胶原血管疾病、免疫抑制紊乱、化学或电烧伤、热力烧伤体表面积大于     

干细胞疗法在创面愈合中的研究进展

创面愈合是一项十分复杂的生物学过程,其具体机制远未阐明.目前已知,它与许多因素都有关联,例如糖尿病、肾功能衰竭、营养不良、吸烟、辐射、感染和免疫功能下降等.这些因素都可以造成创面愈合不良,从而形成慢性溃疡.此外,愈合过程中也会出现一定程度的瘢痕,引起功能障碍.创面愈合过程中引发的愈合过度,还可导致增生性瘢痕的形成,造成局部瘙痒、发热和疼痛等一系列不适,这些都是我们在临床中经常要面对的挑战.近20年来,随着干细胞技术的发展,对创面修复的研究也有了很大的进展.胚胎干细胞（embryonic stem cell,ESCs）、诱导多能干细胞（induced pluripotent stem cell,iPS细胞）及成体干细胞（adult stem cell,ASCs）等均具有强大自我分化能力干细胞的发现和应用,使得干细胞技术成为了治疗创面愈合的一种新策略.本文就以上几种干细胞技术在创面愈合中应用的优缺点做一综述.     

P物质与表皮干细胞联用对糖尿病大鼠创面愈合与神经再生的影响

目的 观察感觉神经肽P物质与表皮干细胞(ESC)联合应用对糖尿病大鼠创面愈合与神经再生的作用. 方法 分离培养SD大鼠ESC(经鉴定),接种于羊膜滋养层上构建羊膜-ESC备用.选择48只糖尿病模型大鼠,每只背部制作4个全层皮肤缺损创面.按随机抽签法将此192个创面分为ESC+P物质组、ESC组、P物质组、对照组,每组48个创面.ESC+P物质组和ESC组创面均移植羊膜-ESC,P物质组、对照组创面移植羊膜.移植后,ESC+P物质组、P物质组在创周及创面中央注射1×10-7 mol/L的P物质250μL,ESC组、对照组在创周及创面中央注射PBS 250μL作对照,各组每日注射2次,连用4d.于大鼠伤后4、7、10、14、17、23 d,观察并计算创面愈合率(每时相点8个创面),HE染色观察创面组织结构改变.伤后4、7、10 d,行Masson染色观察创面组织总胶原分布,免疫组织化学染色观察Ⅰ、Ⅲ型胶原沉积量.伤后14、23 d,用免疫组织化学染色法观察创面组织中蛋白基因产物9.5(PGP 9.5)及P物质阳性神经纤维分布情况.对数据行单因素方差分析和t检验. 结果(1)ESC+P物质组伤后14 d创面愈合率达100.0％,明显早于ESC组、P物质组、对照组完全愈合时间(伤后17、17、23 d).HE染色显示ESC+P物质组创面愈合质量明显优于其余3组.(2)伤后10 d,ESC+P物质组与P物质组创面组织中胶原着色深、面积广;其余2组胶原染色较浅、面积较小.随着伤后时间推移,各组创面Ⅰ型胶原沉积量逐渐升高,Ⅲ型胶原沉积量逐渐下降.伤后4、7、10 d,ESC+P物质组Ⅰ型胶原沉积量明显高于ESC组(t值分别为32.72、118.21、26.71,P值均小于0.01)和对照组(t值分别为44.37、22 76、30.32,P值均小于0.01);ESC+P物质组与P物质组水平相对接近.伤后4、7、10d,ESC+P物质组创面Ⅲ型胶原沉积量明显高于ESC组(t值分别为32.27、28 68、14.51,P值均小于0.01)和对照组(t值分别为35 68、22.52、22 24,P值均小于0.01).(3)ESC +P物质组与P物质组创面组织中有大量PGP 9.5和P物质阳性神经纤维再生,创面深层部分神经纤维末梢向表皮延伸.ESC组、对照组仅见创面深层有少量PGP 9.5和P物质阳性神经纤维,且未向表皮延伸.伤后14、23 d,ESC+P物质组创面PGP 9.5阳性神经纤维面积占(3.86±0.25)％、(7 03±0.28)％,明显高于ESC组[(1.48±0.30)％、(3.01±0 43)％,t值分别为23 95、30 27,P值均小于0.01]和对照组[(1 46±0 23)％、(2.84±0.29)％,t值分别为27.35、40.32,P值均小于0.01].伤后14、23 d,ESC+P物质组创面P物质阳性神经纤维面积占(2.01±0 14)％、(1.19±0 11)％,明显高于ESC组[(0.85±0 17)％、(1.34±0 21)％,t值分别为20.50、2.60,P＜0.05或P＜0.01]和对照组[(0.74 ±0.15)％、( 1.30 ±0.17)％,t值分别为23 98、2.41,P＜0.05或P＜0.01]. 结论 感觉神经肽P物质和ESC联合应用,可以有效促进糖尿病大鼠创面愈合与神经再生.     

烫伤大鼠不同深度创面组织中表皮干细胞分布的初步研究

目的初步观察烫伤大鼠不同深度创面组织中表皮干细胞的分布情况。方法将32只SD大鼠分别造成Ⅰ、浅Ⅱ、深Ⅱ和Ⅲ度烫伤，伤后24h取创面组织标本，制作切片。采用细菌蛋白一生物素标记法(LSAB)进行免疫组织化学染色，以α2、β1整合素及角蛋白10(K10)作为一抗，观察不同创面组织中表皮干细胞的表达和分布情况。结果Ⅰ度烧伤创面组织中，K10阳性细胞分布于表皮的棘细胞层、颗粒层、透明层，α2、β1整合素阳性细胞分布于表皮基底层，数量多。浅Ⅱ度创面中，α2、β1整合素阳性细胞位于残留的基底层和皮肤附属器(主要是毛囊)中，数量较少。深Ⅱ度创面中，α2、β1整合素阳性细胞仅存在于真皮深层健存的皮肤附属器中，数量很少。Ⅲ度创面中，罕见α2、β1整合素阳性细胞。结论表皮干细胞在烧伤创面中的分布与烧伤深度有关，残存的表皮干细胞可能是创伤修复过程中再上皮化的细胞来源。     

脂肪干细胞对创面愈合作用的研究进展

创面愈合是由多种细胞及细胞因子参与的复杂过程,难愈或不愈创面是整形外科的一大挑战.随着人们对脂肪干细胞基础研究及临床应用研究的深入,利用脂肪干细胞来提高创面愈合已被证明是一种非常有前景的治疗策略.在此就脂肪干细胞促进创面愈合的机制、治疗策略的改进、细胞移植途径及临床应用研究等方面的进展进行综述.     

表皮与真皮之间的相互作用对创面愈合的影响

创面愈合是一个复杂有序的过程,包括炎性细胞、修复细胞的聚集,受损组织的清除,细胞外基质的产生和重塑,最后完成再上皮化或形成瘢痕。它涉及细胞的运动、黏附、通讯、增殖和分化等,也包含细胞内、外物质的代谢和基因的启动、调控等一系列生化和分子生物学反应。这一过程包括炎症反应、细胞增殖、创面成熟和重建3个阶段。     

脂肪间充质干细胞在创面愈合中的应用

皮肤的创面愈合是一个相对复杂的生物学过程,此过程中需密切协调级联反应来恢复和修复损伤部位,包括细胞迁移和增殖、血管生成、细胞外基质沉积和重塑.在愈合过程中,ADSCs起着不可或缺的作用,而经过特殊处理后的ADSCs可大大加快创面愈合过程,特别是对类似于癌性创面、失神经创面、内分泌疾病等引发的难愈性溃疡创面等特殊创面的治...