烧伤后早期增生性瘢痕相关细胞骨架基因表达研究

目的　筛选烧伤后早期增生性瘢痕细胞骨架相关基因 ,并探讨其在瘢痕挛缩产生中的作用。　方法　以包含 4 0 96种人类基因的基因芯片研究烧伤后早期增生性瘢痕组织中相关细胞骨架基因表达 ;并利用原位杂交技术观察其中一条基因在烧伤后早期增生性瘢痕组织中的定位分布表达。　结果　在 3例临床标本中 ,共找到瘢痕相关细胞骨架基因 13条 ,均表达上调。在烧伤后早期增生性瘢痕组织中 ,表达人成纤维细胞肌原蛋白基因的细胞明显增多。　结论　烧伤后早期增生性瘢痕组织中就发生了多种细胞骨架相关基因表达上调 ,这可能是烧伤后增生性瘢痕挛缩的重要原因。     

不同胎龄胎儿皮肤和增生性瘢痕中细胞外信号调节激酶5及其mapkk基因表达的变化

目的探讨细胞外信号调节激酶5(extracellular-signal regulated protein kinase 5,erk5)及其上游信号分子mapkk(mek5)在不同胎龄的胎儿皮肤和不同形成时期的增生性瘢痕中的基因表达变化规律及其可能的生物学意义.方法用病理学技术检测不同发育时期的皮肤和增生性瘢痕的结构特征后,提取18例不同胎龄(13～32周)的胎儿皮肤、6例少儿皮肤、16例不同发生时期的增生性瘢痕(4个月-11年)和8例正常皮肤组织的总RNA后,分离mRNA,用RT-PCR方法检测这2种基因在不同组织中的表达变化特征.结果erk5和mek5基因在不同发育阶段的皮肤和增生性瘢痕中都有表达.在早期妊娠胎儿的皮肤中,这2种基因表达较强,随着胎龄的增加,基因表达水平逐渐降低,在少儿皮肤组织中,这两种基因的转录本含量明显减少(P<0.05).在正常皮肤和不同形成时期的增生性瘢痕中,mek5基因表达水平相近,相互间差异不显著,而erk5基因在正常皮肤中的表达水平较低,在增殖期和成熟期增生性瘢痕中表达水平明显增强(P<0.01).结论erk5与mek5基因在不同发育时期的皮肤组织内都有表达,显示erk5介导的信号通路可能对皮肤的发生、结构功能的维持以及伤后修复十分重要.在早期妊娠胎儿皮肤中erk5和mek5基因的高表达可能是胎儿皮肤组织细胞快速增殖,创面无瘢痕愈合的机制之一,而增生性瘢痕发生和形成也可能与erks基因表达增强有关.     

增生性瘢痕形成和成熟过程中三种成纤维细胞生长因子及其受体基因表达的变化

目的探讨成纤维细胞生长因子-7(FGF-7)、酸性成纤维细胞生长因子(aFGF)、碱性成纤维细胞生长因子(bFGF)及其受体(FGFR1和FGFR2)在不同形成时期的增生性瘢痕中的基因表达变化规律及其可能的生物学意义.方法用病理学技术检测增生性瘢痕和正常皮肤的结构特征后,提取16例不同发生时期的增生性瘢痕(4个月～11年)和8例正常皮肤组织的总RNA后,分离mRNA,用RT-PCR方法检测这5种基因在不同组织中的表达.结果在正常皮肤组织中,FGF-7,bFGF,FGFR1和FGFR2基因表达较低,而在增殖期的增生性瘢痕组织中,这4种基因转录本的灰密度值分别为正常皮肤的2.1、2.1、3.6和2.8倍,基因表达显著增强(P<.05);在成熟期的增生性瘢痕中,FGF-7,bFGFR1和bFGFR2基因的表达量都低于增殖期的增生性瘢痕组织,而bFGF仍保持高水平的基因表达.在正常皮肤和增殖期的增生性瘢痕中,aFGF基因呈低水平表达,而在成熟期的增生性瘢痕中aFGF基因表达明显增强.结论FGF-7、bFGF及其受体基因表达升高可能是增生性瘢痕形成的机制之一,而FGF-7、FGFR1和FGFR2基因表达降低,aFGF表达增强可能与增生性瘢痕达到相对稳定的动态平衡状态有关.     

几种深Ⅱ度烧伤创面处理方法的回顾及改善创面微循环的初步实验研究

目的回顾性分析几种深Ⅱ度烧伤创面的修复方法,探讨改善创面微循环对创面愈合的意义. 方法 (1)对于笔者单位烧伤患者的深Ⅱ度创面,应用削痂疗法治疗614例、磨痂疗法治疗32例、清创后异体皮覆盖86例、外用磺胺嘧啶银后创面暴露1 836例、外用中药京万红烫伤膏包扎治疗408例.统计、分析各种疗法的治疗效果.(2)制作大鼠深Ⅱ度烫伤模型.伤后5 min内分别由其尾静脉注入等渗盐水(对照组,10只)、巴曲酶(治疗组,10只),创面均外用磺胺嘧啶银.测定两组大鼠伤前及伤后0.5-72.0 h的创面皮肤血流灌注单位,计算其伤后14、18 d的创面愈合率、收缩率及创面愈合时间.用组织学方法观察两组大鼠创面愈合后的皮肤毛囊数. 结果 (1)削痂疗法术后2-3周创面愈合,其中烧伤总面积50%～79%TBSA的患者治愈率94.8%,总面积80%～98%TBSA者治愈率93.4%.磨痂疗法磨痂+异体皮覆盖术后(13.8±2.1)d创面愈合,无瘢痕形成.清创后异体皮覆盖其中82例患者术后(18.0±2.3)d创面愈合.外用磺胺嘧啶银后暴露其中1 658例患者用药后(26.0±3.2)d痂下愈合.外用京万红烫伤膏后包扎患者多有细菌感染,其中下肢创面愈合时间为(26.0±2.8)d.(2)治疗组大鼠伤后2.0-72.0 h创面局部血流灌注单位均明显高于对照组(P<0.01).伤后14、18 d,治疗组创面愈合率明显高于对照组(P<0.01),但两组创面收缩率接近(P>0.05).治疗组创面愈合时间短于对照组(P<0.01).伤后30 d,对照组大鼠真皮层中残存少量毛囊,数量明显少于治疗组(P<0.01). 结论深Ⅱ度烧伤后早期采用削痂、磨痂或清创后覆盖异体皮的方法处理创面,可减轻感染、缩短疗程、提高治愈率和愈合质量.使用巴曲酶可改善深Ⅱ度烧伤创面微循环,加快愈合速度.     

微小RNA21和TGF-β_1在正常皮肤和增生性瘢痕组织中的表达研究

目的:探讨微小RNA-21(miRNA-21)基因与转化生长因子-β_1(transforming growth factor-β,TGF-β_1)基因在正常皮肤组织和增生性瘢痕组织中的表达,并分析其相关性。方法:选取本院烧伤整形科收治的16例烧伤整形患者的增生性瘢痕组织及其临近的正常皮肤组织,提取组织中的总RNA,采用实时荧光定量PCR(realtime fluorescence quantitative PCR,RT-q PCR)技术分别检测增生性瘢痕组织和正常组织中miRNA-21与TGF-β_1基因表达量,对检测结果进行分析比较,并行相关性分析。结果:增生性瘢痕组织中miRNA-21基因表达明显上调,差异具有统计学意义(P0.05),其表达水平是正常皮肤组织的2.18倍;增生性瘢痕中TGF-β_1的表达量多于正常皮肤组织,差异具有统计学意义(P0.05),其表达水平是正常皮肤组织的2.31倍;增生性瘢痕组织中miRNA21基因表达量与TGF-β_1基因表达量呈正相关(r=0.836,P0.01)。结论:增生性瘢痕组织中miRNA-21和TGF-β_1基因表达异常,可能与瘢痕组织形成相关。     

增生瘢痕皮肤中转化生长因子基因表达

目的探讨转化生长因子(TGF)-β1和转录因子Smad 2,Smad 3三种基因影响增生性瘢痕形成和胎儿皮肤伤口无瘢痕愈合的调节机制.方法 32份被检测标本中包括增生性瘢痕8份,其对应的正常皮肤组织8份,胎儿和成人皮肤组织各8份.用逆转录-多聚酶链反应方法(RT-PCR)检测这3种基因在不同的组织细胞内的表达变化规律.结果 TGF-β1、Smad 2 和Smad 3三种基因在增生性瘢痕、胎儿和出生后机体皮肤组织细胞内都有表达.在所检测8对增生性瘢痕和其对应正常皮肤细胞中,这3种不同基因在增生性瘢痕细胞中的表达量高于正常皮肤细胞的组数分别为TGF-β1基因有5对,Smad 2基因有8对,而Smad 3基因有5对.在胎儿皮肤细胞内,TGF-β1的mRNA含量明显低于成人皮肤细胞(t=2.204,P<0.05),差异有显著意义;Smad 3基因表达也呈相似的变化规律,mRNA含量显著低于成人皮肤细胞(t=4.269,P<0.01),差异有非常显著意义;而Smad 2基因的mRNA含量却明显高于成人皮肤细胞(t=6.685,P<0.01),差异亦有非常显著意义.结论 TGF-β1和它的下游信号分子Smad 2,Smad 3可能与增生性瘢痕形成密切相关.在增生性瘢痕细胞内,这3种基因高表达可诱导胞外基质大量沉积,加速成纤维细胞增殖,促进组织纤维化.TGF-β1和Smad 3基因在胎儿皮肤组织细胞内低表达可能是皮肤伤口无瘢痕愈合的机制之一.     

瘢痕中肌动蛋白及肌球蛋白的实验研究

目的　探讨增生性瘢痕、瘢痕疙瘩及正常皮肤成纤维细胞中肌动蛋白和肌球蛋白Ⅱ的不同表达情况及其与瘢痕挛缩的关系。方法　取增生性瘢痕15例,瘢痕疙瘩10例,正常皮肤15例,用免疫组织化学方法检测其中肌动蛋白和肌球蛋白Ⅱ的表达情况,同时做细胞培养,用流式细胞术检测成纤维细胞中肌动蛋白和肌球蛋白Ⅱ的表达情况。结果　增生性瘢痕肌球蛋白Ⅱ的免疫组织化学染色呈阳性,而瘢痕疙瘩及正常皮肤肌球蛋白Ⅱ的表达均为阴性。增生性瘢痕肌球蛋白Ⅱ的表达较其它两种组织有非常显著性差异(P<0.01),而瘢痕疙瘩和正常皮肤无显著性差异(P>0.05)。流式细胞术检测增生性瘢痕、瘢痕疙瘩及正常皮肤三种组织中肌球蛋白Ⅱ的阳性率分别为(95.11±2.78)%、(16.86±7.11)%及(5.31±1.79)%,增生性瘢痕肌球蛋白Ⅱ表达的阳性率较其它两种组织有非常显著性差异(P<0.01),而瘢痕疙瘩和正常皮肤无显著性差异(P>0.05)。三种组织中肌动蛋白的免疫组织化学染色均为阳性,无显著性差异(P>0.05)。流式细胞术检测三种组织中肌动蛋白的阳性率分别为(77.77±15.43)%、(88.89±10.29)%及(82.92±13.48)%,其表达的阳性率无显著性差异(P>0.05)。结论　瘢痕挛缩与肌球蛋白Ⅱ密切相关,而肌动蛋白在细胞中除作为细胞的收缩蛋白之一外,同时与细胞的活动及运动有关,是细胞生命活动中必不可少的蛋白之一。     

喷雾按摩在烧伤瘢痕康复中的应用

目的：探讨烧伤后增生性瘢痕的康复处理效果。方法：采用中草药离子喷雾、面膜及按摩方法对烧伤后增生性瘢痕进行康复处理,结果：48例深Ⅱ度烧伤后不同时期与形状的增生性瘢痕,采用本法处理,效果满意,总有效率达100％。结论：深Ⅱ度烧伤创面愈合后开始康复处理越早效果越好,本法对片块状瘢痕效果理想。     

烧伤后增生性瘢痕组织中基质金属蛋白酶2mRNA表达的临床研究

目的观察烧伤后增生性瘢痕组织中基质金属蛋白酶2(MMP2)的动态表达情况。方法术中切取23例深Ⅱ度烧伤患者创面自然愈合后形成的早、中、晚期增生性瘢痕组织23块。采用原位杂交法检测MMP2mRNA在各期瘢痕组织中的表达和分布,并计算MMP2的阳性细胞百分比。结果MMP2主要由瘢痕组织中的成纤维细胞表达。早期瘢痕组织中MMP2mRNA阳性表达少,其阳性细胞百分比为(13.9±3.6)%;中期瘢痕组织中MMP2mRNA阳性表达增多,其阳性细胞占(68.7±5.2)%;晚期瘢痕组织中MMP2mRNA表达极低,其阳性细胞仅占(2.0±0.9)%。结论MMP2与瘢痕组织重塑过程密切相关。     

异种(猪)脱细胞真皮基质一次性包扎治疗深Ⅱ度烧伤

目的探讨应用异种(猪)脱细胞真皮基质一次性包扎治疗深Ⅱ度烧伤创面的临床应用效果。方法1997年1月—2004年1月,应用异种(猪)脱细胞真皮基质一次包扎治疗50%~95%总体表面积(TBSA)、深Ⅱ度烧伤的患者67例[异种(猪)脱细胞真皮基质治疗组];同期保痂治疗的50%~95%TBSA、深Ⅱ度的患者10例(保痂治疗组),观察创面愈合时间和愈合质量及其并发症发生情况。治愈患者经过3个月~2年的随诊,观察瘢痕增生情况。结果异种(猪)脱细胞真皮基质治疗组深Ⅱ度创面中途基本不需换药,创面愈合时间缩短,平均(12.2±2.6)d,而保痂治疗组愈合时间为(27.4±3.5)d,差异具有统计学意义(P<0.05);同时,异种(猪)脱细胞真皮基质治疗组瘢痕增生情况较保痂治疗组明显减轻或者无瘢痕增生。结论一次性覆盖异种(猪)脱细胞真皮基质可有效地治疗深Ⅱ度烧伤创面,能加快创面愈合,减轻瘢痕增生,从而降低烧伤感染和炎症反应综合征的发生。     

A porcine deep dermal partial thickness burn model with hypertrophic scarring

We developed a reproducible model of deep dermal partial thickness burn injury in juvenile Large White pigs. The contact burn is created using water at 92 degrees C for 15s in a bottle with the bottom replaced with plastic wrap. The depth of injury was determined by a histopathologist who examined tissue sections 2 and 6 days after injury in a blinded manner. Upon creation, the circular wound area developed white eschar and a hyperaemic zone around the wound border. Animals were kept for 6 weeks or 99 days to examine the wound healing process. The wounds took between 3 and 5 weeks for complete re-epithelialisation. Most wounds developed contracted, purple, hypertrophic scars. On measurement, the thickness of the burned skin was approximately 1.8 times that of the control skin at week 6 and approximately 2.2 times thicker than control skin at 99 days after injury. We have developed various methods to assess healing wounds, including digital photographic analysis, depth of organising granulation tissue, immunohistochemistry, electron microscopy and tensiometry. Immunohistochemistry and electron microscopy showed that our porcine hypertrophic scar appears similar to human hypertrophic scarring. The development of this model allows us to test and compare different treatments on burn wounds.     

Control of hypertrophic scar from inception by using xenogenic (porcine) acellular dermal matrix (ADM) to cover deep second degree burn

OBJECTIVE: We have spent 7 years to investigate the method of applying porcine acellular dermal matrix (ADM) on deep partial thickness burn wound until the wound heals without dressing change. Known as "Feng's pig skin method" by our hospital, the method appears to encourage rapid re-epithilization with minimum scarring. METHOD: The deep partial thickness burn wound was rinsed cleanly under anesthesia when the patient admitted. ADM was applied on the wound after the detached epidermis was thoroughly removed, wrapped and fixed by sterile gauze and bandages. The dressing was removed within two weeks and the wound completely healed. The outcome of the treatment was analyzed by using the modified Vancouver Burn Scar Assessment Scale. RESULT: All the wounds healed with one dressing within 2 weeks, and the time of wound re-epithelialization shortened to 7-12 days. Scar hyperplasia did not occur, or it was greatly ameliorated compared with traditional treatment after a followed-up period of 3 months to 2 years. The Scar Index was significant lower than that of the traditional exposure method. CONCLUSION: Using ADM to cover deep second degree burn can preserve maximally residual dermal tissue and epithelium, help accelerate the regeneration of epithelial and stem cells, thus shorten the healing time, remodel the skin structure, and consequently has the effect of controlling hypertrophic scar at inception.     

Imaging mass spectrometry for accessing molecular changes during burn wound healing

The spatiotemporal analysis of the proteomic profile during human wound healing is a critical investigative step that can establish the complex interplay of molecular events that comprise the local response to burn injury. Partial‐thickness wound samples with adjacent “normal” skin were collected from twenty‐one patients with burn wounds and examined across a time spectrum ranging from the acute injury period at 3, 6, 11 days to the later hypertrophic scar period at 7 and 15 months. The techniques used for histology‐directed tissue analyses highlighted inflammatory protein markers at the early time points after injury with diminished expression as burn wounds progressed into the proliferative phase. The datasets show the usefulness of MALDI MS and imaging mass spectrometry as discovery approaches to identify and map the cutaneous molecular sequence that is activated in response to the unique systemic inflammatory response following burn trauma. This information has the potential to define the unique factors that predispose human burn victims to disfiguring hypertrophic scar formation.     

Thermal injury model in the rabbit ear with quantifiable burn progression and hypertrophic scar

Hypertrophic scar is a major clinical outcome of deep‐partial thickness to full thickness thermal burn injury. Appropriate animal models are a limitation to burn research due to the lack of, or access to, animal models which address the endpoint of hypertrophic scar. Lower species, such as rodents, heal mainly by contracture, which limits the duration of study. Higher species, such as pigs, heal more similarly to humans, but are associated with high cost, long duration for scar development, challenges in quantifying scar hypertrophy, and poor manageability. Here, we present a quantifiable deep‐partial thickness burn model in the rabbit ear. Burns were created using a dry‐heated brass rod for 10 and 20 seconds at 90 °C. At the time of eschar excision on day 3, excisional wounds were made on the contralateral ear for comparison. Burn wound progression, in which the wound size expands over time is a major distinction between excisional and thermal injuries, was quantified at 1 hour and 3 days after the injuries using calibrated photographs and histology and the size of the wounds was found to be unchanged from the initial wound size at 1 hour, but 10% in the 20 seconds burn wounds at 3 days. A quantifiable hypertrophic scar, measured by histology as the scar elevation index, was present in both 20 seconds burn wounds and excisional wounds at day 35. ImageJ measurements revealed that the 20 seconds burn wound scars were 22% larger than the excisional wound scars and the 20 seconds burn scar area measurements from histology were 26% greater than in the excisional wound scar. The ability to measure both burn progression and scar hypertrophy over a 35‐day time frame suits this model to screening early intervention burn wound therapeutics or scar treatments in a burn‐specific scar model.     

巴曲酶对深Ⅱ度烫伤创面微循环血流变化及愈合的影响

目的　探讨巴曲酶对大鼠烫伤早期创面淤滞带微循环血流变化及愈合过程的影响。　方法　Wistar雄性大鼠 2 0只 ,随机分为烫伤对照组及巴曲酶治疗组 ,在各组动物背部均造成 4cm× 4cm大小深Ⅱ度烫伤。分别于伤前、伤后 0 .5、2、4、6、12、2 4、72h测定创面局部皮肤血流量 ;于伤后立即、伤后 14、18d测定烫伤面积及残留创面面积 ;于伤后 30d将大鼠处死取材行组织学观察及毛囊计数。　结果　大鼠伤后 2 72h皮肤血流量进行性减少 ;与对照组相比 ,给予巴曲酶后皮肤血流量得到明显改善 ,创面愈合速度明显加快 ,愈合后皮肤附件明显增多。　结论　巴曲酶能逆转烧伤后创面淤滞带循环 ,加快创面愈合速度和提高皮肤愈合质量     

荷负电气溶胶治疗Ⅱ度烧伤创面的临床效果及病理学观察

目的观察荷负电气溶胶(下称气溶胶)治疗Ⅱ度烧伤创面的效果。方法选择单纯浅Ⅱ、深Ⅱ度烧伤患者,随机分为:(1)气溶胶组:浅Ⅱ度180例、深Ⅱ度100例,伤后6h~2d开始用气溶胶治疗创面,l~2次/d,1.5h/次。(2)对照组:浅Ⅱ、深Ⅱ度患者各30例,常规治疗。(3)自身对照组:浅Ⅱ、深Ⅱ度患者各10例,同上用气溶胶治疗,但同一患者部分创面覆盖无菌金属片屏蔽气溶胶(屏蔽组),部分创面不屏蔽(非屏蔽组)。观察气溶胶治疗过程中患者创面的大体变化,治疗前后进行创面细菌培养,并监测其肝、肾功能及血生化指标有无改变。记录各组患者创面愈合时间。另制作深Ⅱ度烫伤大鼠模型,同前分为气溶胶组和对照组并治疗。取两组大鼠治疗前及治疗后1、2、3周的创面组织标本,作病理学观察。结果气溶胶治疗后患者创面渗出少,治疗前后均无细菌生长。总体来讲,气溶胶治疗前后患者肝、肾功能及血生化指标无明显改变。气溶胶组患者浅Ⅱ度创面伤后(6.3±1.6)d愈合,深Ⅱ度创面(15.1±3.1)d愈合,明显短于对照组相同深度创面[(11.3±1.4)、(21.2±1.4)d,P<0.01]。自身对照组中,相同烧伤深度的非屏蔽组与屏蔽组比较,创面愈合时间也明显缩短(P<0.01)。病理学检查显示,气溶胶组大鼠治疗后第3周皮肤结构已基本恢复正常,而对照组此时恢复较差。结论气溶胶能有效促进Ⅱ度烧伤创面的愈合且使用安全。     

烧伤后24小时内削痂对深Ⅱ度创面局部炎症反应的影响

目的探讨烧伤后24h内行削痂手术对深Ⅱ度创面局部炎症反应以及组织损害的影响。方法选择12例烧伤患者,伤后24h内对深Ⅱ度创面行削痂手术,将患者同一创面分为3个标本采集区,即手术前、手术后和未手术区。手术前标本取自削痂术前创面,手术后和未手术区创面标本于伤后5—7d获取。采用组织培养和比色法测定创面组织释放白细胞介素(IL)8、髓过氧化物酶(MPO)和丙二醛(MDA)的水平,通过HE和Masson染色对创面组织坏死程度进行形态学观察。结果削痂手术后,患者创面组织局部释放IL8、MPO、MDA的水平分别为(6.83±1.85)μg/L、(4.07±0.87)U/g、(8.94±5.66)μmol/g,与未手术创面比较明显降低(P<0.01)。形态学观察显示,手术前创面有凝固性坏死灶;未手术创面炎症反应明显,坏死组织范围扩大;削痂手术后创面局部的炎症反应得到改善,未见坏死组织范围扩大。结论伤后24h内行削痂手术,可以改善深Ⅱ度烧伤创面局部的炎症反应,防止创面进行性加深,有利于创面及早愈合。