碱性成纤维细胞生长因子对增生性瘢痕的作用

目的：探讨碱性成纤维性细胞生长因子（bFGF)对移植至裸鼠皮下的人增生性瘢痕的作用。方法：将烧伤患者手术切除的增生性瘢痕移植到裸鼠背部皮下，动物随机分为5组：对照组，bFGF组，胶原酶组， FGF 胶原酶组和玻璃酸钠组，瘢痕块移植后2周分别用上述不同药物局部注射至瘢痕内共3次，每次间隔3d，后观察疤痕大小的1／2－1／3且软，能使粗大密集的胶原束变松散成团，其结构大部分消失，bFGF 胶原酶组所移植的瘢痕组织块有2／3消失，结论：增生性瘢痕局部应用bFGF可以降低瘢痕胶原，瘢痕中粗大的胶原束结构大部分消失。     

人Ⅰ型前胶原基因反义寡核苷酸对人增生性瘢痕裸鼠移植模型胶原合成的作用

目的人Ⅰ型前胶原基因(ColⅠA1)反义寡核苷酸(antisense oligodeoxyneucleotide,ASODN)对体外培养的人增生性瘢痕成纤维细胞有抑制胶原合成作用,拟进一步探讨ColⅠA1 ASODN对裸鼠移植模型体内的人增生性瘢痕的胶原合成作用。方法 SPF级BALB/c-nunu品系6～8周龄雌性裸鼠60只,体重约20 g;取行瘢痕切除手术患者自愿捐赠的瘢痕组织块去表皮后,移植至裸鼠背部肩胛内侧皮下,每只裸鼠移植1块,制备人增生性瘢痕裸鼠移植模型。将58只成功制备模型的裸鼠根据处理方法不同,随机分成3组,于瘢痕移植2周根据分组行经皮穿刺瘢痕内注射。A组(n=20):5μL浓度为3 mmol/L ColⅠA1 ASODN、3μL脂质体、92μL Opti-MEMⅠ减血清培养基;B组(n=20):3μL脂质体、97μL Opti-MEMⅠ减血清培养基;C组(n=18):100μL Opti-MEMⅠ减血清培养基。实验最初2周每天注射1次,之后隔天1次,至实验取材。注射后2、4、6周,对存活裸鼠进行瘢痕硬度测量后,处死裸鼠取瘢痕组织行胶原染色组织学观察以及透射电镜观察;提取细胞总RNA后行RT-PCR测定ColⅠA1 mRNA相对含量;ELISA法行ColⅠA1蛋白定量分析。结果注射后6周内17只裸鼠死亡;共41只裸鼠40块瘢痕组织符合实验标准,纳入实验;A、B、C组各14、13、14只。注射后2周,3组间瘢痕硬度比较,差异均无统计学意义(P>0.05);4、6周时A组与B、C组比较,差异有统计学意义(P<0.05),B、C组间比较差异无统计学意义(P>0.05)。随时间延长,组织学观察示3组Ⅲ型胶原表达上升,A组最明显,Ⅰ型胶原排列规则。透射电镜观察示A组成纤维细胞退化,胶原纤维排列更趋于一致;B、C组胶原纤维排列也逐渐规则。注射后2、4周3组间ColⅠA1 mRNA和蛋白表达量比较,差异均有统计学意义(P<0.05);6周时A组与B、C组比较,差异有统计学意义(P<0.05),B、C组间比较差异无统计学意义(P>0.05)。结论 在人增生性瘢痕裸鼠移植模型的瘢痕内注射ColⅠA1 ASODN可抑制ColⅠA1 mRNA及Ⅰ型胶原表达,促进瘢痕组织成熟、软化,脂质体可促进该抑制效果。     

Human hypertrophic scar‐like nude mouse model: Characterization of the molecular and cellular biology of the scar process

Hypertrophic scar (HTS) following thermal injury and other forms of trauma is a dermal fibroproliferative disorder that leads to considerable morbidity. Because of the lack of an ideal animal model, research is difficult. We have established an HTS model that involves transplanting human split‐thickness skin graft (STSG) or full‐thickness skin graft (FTSG) onto the backs of nude mice. The animals developed raised, firm, and reddish scars 2 months following transplantation. Histology and micromeasurement indicate raised, thickened engrafted skin with STSG and FTSG. In contrast, thickening was not observed with full‐thickness rat skin grafts used as controls. Masson's trichrome staining demonstrates increased accumulations of collagen fibrils in the dermis in both scars grafted with STSG and FTSG. Staining cells with toludine blue and an antibody for F4/80 showed an increase in the infiltration of mast cells and macrophages. Quantification of fibrocytes reveals increased fibrocytes. Moreover, STSG grafted skin had significantly more macrophages, mast cells, and fibrocytes than FTSG. Real‐time polymerase chain reaction analysis showed significantly elevated mRNA levels for type I collagen, transforming growth factor‐β, connective tissue growth factor and heat shock protein 47 in both types of engrafted skin. These data demonstrate that human skin grafted onto nude mice develops red raised and thickened scars having intrinsic properties that closely resemble HTS formation as seen in humans. Interestingly, STSG developed more scar than FTSG. Furthermore, inflammatory cells and bone marrow‐derived fibrocytes may play a critical role in HTS development in this animal model.     

A new experimental hypertrophic scar model in guinea pigs

Many aspects of the biology and effective therapy of proliferative scars remain undefined, in part due to a lack of an accurate, practical, reproducible, and economical animal model for systematically studying hypertrophic scars. This study was designed to investigate whether hypertrophic scar formation could be induced in guinea pigs by removal of the panniculus carnosus alone, and by a combination of the removal of the panniculus carnosus with application of coal tar afterwards. Whole thickness skin excision or deep partial thickness injury was used to create the lesions on intact skin. Different anatomic locations were tested in different groups. Scars thus developed were examined morphologically by light microscopy and electron microscopy (TEM and SEM) and biochemically by measuring the activity of glucose-6-phosphate dehydrogenase (G6PD) to check whether these scars had morphological and biochemical properties specific to hypertrophic scars. The albino guinea pigs used in this study were divided into three groups. Removal of the panniculus carnosus was performed from the ventral aspect of the torso in animals in groups I and II. On the skin overlying the area of panniculectomy, circular skin excision was performed in group I, and deep partial thickness burn injury was inflicted in group II, to see whether wounds would heal with hypertrophic scars. In group III, dorsal aspect of the torso were used and wounds were produced by circular skin excisions followed by panniculectomy on both sides but coal tar was applied to only one side. Tissue samples were taken from the scars that were hypertrophic in appearance, and from normal scars and normal skin for comparison. Light and electron microscopic examinations and G6PD activity measurements were performed on these samples. While hypertrophic scar development was not seen in group I and group II, scars with morphological and biochemical properties specific to hypertrophic scars developed in one third of animals in group III after healing of the wounds treated with coal tar. In conclusion, it is shown that it is possible to develop experimental hypertrophic scars in guinea pigs with morphological and biochemical properties similar to those of human proliferative scars. Therefore this model is a new, practical, and economical experimental animal model to study proliferative scars, although improvements are needed to increase yield.     

新型增生性瘢痕裸鼠动物模型的建立

目的　建立一种稳定的增生性瘢痕 (HS)动物模型 ,为进一步研究HS的发病机制提供帮助。　方法　在 10 0只裸鼠背部分别作 2 .0cm× 1.5cm全层皮肤缺损创面 ,移植人全厚皮肤 ;皮片存活后用加热的铜棒造成深Ⅱ度烫伤 ,观察创面愈合后的瘢痕增生情况。　结果　皮片完全存活的 86只裸鼠中 ,6 7只有明显、持续的瘢痕增生 ,占 78%。增生瘢痕的外观和组织学特点与人体增生性瘢痕相似。瘢痕增生的平均厚度为 0 .34cm,最厚 0 .6 0cm,增生时间为 6 3～ 2 17d,平均 12 8d。结论　该模型瘢痕增生明显、增生持续时间长 ,可用于观察创面愈合至瘢痕形成的全过程 ,是目前较为稳定的增生性瘢痕动物模型。     

他克莫司抑制兔耳瘢痕增生的作用及其机制研究

目的：他克莫司是临床广泛应用的一种药物,其对增生性瘢痕是否产生作用并无相关报道,为此提出并证实了他克莫司可以抑制兔耳瘢痕增生。方法：建立兔耳增生性瘢痕模型,选10只新西兰大耳白兔双耳腹侧用打孔器制作直径1cm圆形创面,伤后14天创面上皮化后给药,每只兔左耳为空白对照组涂等剂量凡士林软膏,右耳为他克莫司治疗组。分别在伤后14天、21天2、8天3、5天和49天采集标本,行HE染色,观察形态学差异;Real-t i me PCR检测纤维化相关因子TGF-β1、TGF-β2、Col l agen-α1等的表达。结果：HE染色可见他克莫司组成纤维细胞数量及胶原纤维较对照组明显减少,PCR结果可见TGF-β1、TGF-β2及Col l agen-α1表达较对照组在各时间点均减少。结论：实验组较对照组瘢痕明显减轻,证明他克莫司显著抑制兔耳瘢痕增生,可作为临床上治疗及预防瘢痕增生的全新疗法。     

烧伤后增生性瘢痕裸鼠模型中MMP-1和TGF-β1的表达

目的研究MMP-1和TGF-β1在烧伤创面愈合至HS形成过程中的表达及作用.方法将人的正常全厚皮片移植于裸鼠,待其完全成活后造成深Ⅱ度烧伤创面,然后用免疫组织化学染色的方法观察MMP-1和TGF-β1在烧伤创面愈合至瘢痕形成全过程中的表达变化.结果MMP-1于烧伤后第2天出现,第3～15天表达较强,第28天已下降到基础水平.TGF-β1于烧伤后12?h出现,一直持续到烧伤后5个月瘢痕形成.MMP-1和TGF-β1在不同流动性的组织界面明显.结论MMP-1可能主要在创伤愈合的炎症期和增生早期发挥作用,在增生晚期、重塑期作用受到了明显抑制.TGF-β1在伤口愈合过程中广泛和长期的表达模式,与其对细胞功能广泛地调节作用是一致的;与其对MMP-1的抑制作用也是一致的.并且MMP-1和TGF-β1可能在具有不同流动性的组织界面起着特殊的作用.     

人全厚皮片移植裸鼠模型烧伤后 MMP-1、TIMP-1、TGF-β1的表达

目的研究基质金属蛋白酶-1(MMP-1)、金属蛋白酶组织抑制因子-1(TIMP-1)、转移生长因子-1(TGF-β1)在烧伤创面愈合至瘢痕(HS)形成过程中的表达、关系和作用.方法将人的正常全厚皮肤移植于裸鼠,待其完全存活后造成深Ⅱ度烧伤创面.用免疫组织化学染色的方法,观察MMP-1、TIMP-1、TGF-β1在烧伤创面愈合至HS形成全过程中的表达.结果 MMP-1于烧伤后第2天出现,第3～15天表达较强,第28天已下降到基底值.TIMP-1于烧伤后第2天出现,一直持续到烧伤后5个月HS形成,其表达类似于MMP-1的接触界面模式,但无MMP-1规则,且在血管周围有明显的分布.TGF-β1于烧伤后12 h出现,一直持续到烧伤后5个月HS的形成.三种因子在不同结构的接触界面表达明显.结论在创伤愈合的炎症期和增生期,MMP-1和TIMP-1两种力量处于动态平衡;在重塑期,TIMP-1通过抑制MMP-1的表达减少胶原降解,从而诱导HS形成.TGF-β1在创伤愈合过程中广泛和长期的表达模式,与它对细胞功能广泛的调节作用一致,与它对MMP-1的抑制作用也一致.     

建立更加稳定和有效的兔耳瘢痕模型

目的：建立更加稳定和有效的兔耳瘢痕模型。方法：选用16只新西兰大耳白兔,分别在兔耳腹侧中段作1cm×1cm的全层皮肤缺损共192个,以形态学、瘢痕增生指数及羟脯氨酸（HPr）的含量变化对创面愈合后形成的增生块,进行动态组织病理学、细胞增殖活性及胶原纤维合成等检测。结果：兔耳腹侧中段创面可产生类似于人类的增生性瘢痕,其发生率为91．5％,增生块最长持续时间可达120多天。结论：采用本实验的模型复制方法,可以得到更加稳定和有效的兔耳增生性瘢痕。     

建立一种兔耳增生性瘢痕的动物模型

目的　建立由动物自身产生的与人增生性瘢痕类似的增生性动物模型。方法　选用2 4只新西兰大耳白兔 ,分别在兔耳腹侧、背侧面作直径 1.5cm的全层皮肤缺损各 14 4个、72个 ,对创面愈合后形成的增生块进行组织病理学、层黏连蛋白 (LN)mRNA及整合素 β1mRNA检测等检查。结果　兔耳腹侧创面可产生类似人增生性瘢痕样的过度增生 ,其发生率为 71% ,以兔耳腹侧面中部为著 ,增生块最长持续时间超过伤口愈合后 15 0d ,而兔耳背侧面的创面增生块发生率不到3 0 % ,且增生块持续时间短 ,为 76d。切片镜下显示 ,增生块的真皮层存在大量成纤维细胞 ,与人增生性瘢痕结构类似。结论　兔耳腹侧面尤其腹侧面的中部可产生类似人增生性瘢痕样病理改变 ,可望成为研究瘢痕的动物模型。     

一种增生性瘢痕动物模型的建立

目的　建立兔耳瘢痕动物模型 ,观察兔耳腹侧创面在伤后不同时间瘢痕增生的情况。方法　于 32只新西兰白兔的 6 0只兔耳腹面手术切除 2cm× 5cm全层皮肤 ,创面用 1%磺胺嘧啶银冷霜外敷包扎至愈合 ,换药 1次 /周。未作手术的 4只兔耳作对照。 (1)术后连续 12个月观察兔耳创面自然愈合情况。 (2 )用光镜、透射电镜观察兔耳创面瘢痕增生情况。 (3)用计算机图像分析系统测定 1～ 6个月的瘢痕指数。　结果　兔耳创面上皮化后其色泽、厚度和质地均经历从瘢痕形成、成熟到退化的演变过程 ;1～ 2个月的瘢痕指数 2 .2 9± 0 .74较 3～ 4个月 (2 .82± 0 .36 )和 5～ 6个月 (2 .90± 0 .84 )低 (P <0.0 5),其变化与瘢痕增生程度的消长趋势吻合。　结论　兔耳腹面全层皮肤缺损经自然愈合后形成的增生性瘢痕与人体增生性瘢痕相似 ,该模型是研究增生性瘢痕的发生机制及评估其治疗方法的较好的动物模型之一     

增生性瘢痕P物质含量的放射免疫测定

目的测定增生性瘢痕组织中P物质含量是否高于非增生性瘢痕及正常皮肤。方法用放射免疫法分别对增生性瘢痕、非增生性瘢痕及正常皮肤组织中P物质含量做定量分析研究。结果增生性瘢痕组织中P物质含量显著高于非增生性瘢痕及正常皮肤组织(P<0.01)。结论 P物质在增生性瘢痕的发病过程中可能起重要作用。     

增生性瘢痕P物质含量的放射免疫测定

目的测定增生性瘢痕组织中Ｐ物质含量是否高于非增生性瘢痕及正常皮肤。方法用放射免疫法分别对增生性瘢痕、非增生性瘢痕及正常皮肤组织中Ｐ物质含量做定量分析研究。结果增生性瘢痕组织中Ｐ物质含量显著高于非增生性瘢痕及正常皮肤组织（Ｐ＜０．０１）。结论Ｐ物质在增生性瘢痕的发病过程中可能起重要作用。     

胶原酶对裸鼠移植肥厚性瘢痕的治疗作用

目的观察胶原酶对肥厚性瘢痕（ＨＴＳ）胶原降解的影响,探讨它对ＨＴＳ的治疗作用。方法建立ＨＴＳ裸鼠移植模型,局部注射胶原酶于瘢痕组织。治疗后取材行光、电镜观察和分析。结果ＨＴＳ组织胶原纤维被降解,真皮层变薄,瘢痕缩小,质地变软,与对照组差异十分明显。结论胶原酶的局部注射可能是治疗ＨＴＳ的较好方法。     

胶原酶对裸鼠移植肥厚性瘢痕的治疗作用

目的观察胶原酶对肥厚性瘢痕(HTS)胶原降解的影响,探讨它对 HTS 的治疗作用。方法建立 HTS 裸鼠移植模型,局部注射胶原酶于瘢痕组织。治疗后取材行光、电镜观察和分析。结果 HTS 组织胶原纤维被降解,真皮层变薄,瘢痕缩小,质地变软,与对照组差异十分明显。结论胶原酶的局部注射可能是治疗 HTS 的较好方法。     

增生性瘢痕动物实验模型的建立与应用

目的建立由动物自身产生的增生性瘢痕实验模型.方法选用47只日本大耳白兔,在耳腹侧面制作6mm圆形全层皮肤缺损创面、1.5cm×4.5cm长方形创面以及耳背圆形创面共388个,进行组织学等多项观察.结果70%兔耳圆形创面可发生与人增生性瘢痕类似的增生块,增生块持续时间最长为150d;长方形创面增生块发生率为80%以上,持续时间已超过262d.增生块内有特征性的结节或漩涡状结构.创基注入TGF-β1、IFN-y,可以促进或抑制增生块的发生.原位杂交,查明内源性TGF-β1mRNA为强阳性表达.利用此模型证实成纤维细胞凋亡在病理性瘢痕发生、发展过程中起着重要的调控作用.结论兔耳可以产生与人增生性瘢痕类似的病理改变,可以用作瘢痕研究的实验模型.     

High-malignancy orthotopic nude mouse model of human prostate cancer LNCaP.

BACKGROUND: An animal model of human prostate cancer LNCaP demonstrating high rates of spontaneous metastasis from the orthotopic site after tumor implantation would be very valuable for mechanistic and drug discovery studies. We previously developed microsurgical techniques to implant histologically intact tumor tissues orthotopically in nude mice in order to develop high metastatic mouse models of human cancer. METHODS: Intact tissue of the androgen-dependent human prostate cancer cell line, LNCaP, was implanted on the ventral lateral lobes of the prostate gland by surgical orthotopic implantation (SOI) in a series of 20 nude mice. Mice were autopsied, and histopathological examination of primary tumors and relevant organs was done to identify and quantitate micrometastasis. RESULTS: Eighteen of 20 animals transplanted with LNCaP by SOI had tumor growth. Mean primary tumor weight in the prostate was 9.24 g at time of necropsy. Sixty-one percent of the transplanted animals had lymph node metastasis. Forty-four percent had lung metastasis. Mean survival time was 72 days, indicating a high degree of malignancy of the tumor. CONCLUSIONS: The extensive and widespread lung metastasis as well as lymph node metastasis following orthotopic implantation of LNCaP in nude mice and the short survival time provide a high-malignancy nude model of the LNCaP human prostate tumor.     

增生性瘢痕P物质含量的放射免疫测定

目的 测定增生性瘢痕组织中Ｐ物质含量是否高于非增生性瘢痕及正常皮肤。方法 用放射免疫法分别对增生性瘢痕、非增生性瘢痕及正常皮肤组织中Ｐ物质含量做定量分析研究。结果 增生性瘢痕组织中Ｐ物质含量显著高于非增生性瘢痕及正常皮肤组织（Ｐ〈０．０１）。结论 Ｐ物质在增生性瘢痕的发病过程中可能起重要作用。     

增生性瘢痕组织中褪黑素受体的表达

Objective To investigate the expression and its significance of melatonin receptor in human hypertrophic scarring. Methods The expression of melatonin receptor GPR50 was detected with immunohistochemistiy and the melatonin receptors( MT1 、MT2)mRNA were assessed with RT-PCR method in 10 cases of human hypertrophic scar and normal skin. The positive production was sequenced with auto sequencing instrument. Results Positive signals of melatonin receptor could be found in the cell membrane and cytoplasm. The melatonin receptor GPR50 was located in the epithelial basal cells, sweat gland cells and hair follicle in both hypertrophic scar and normal skin. The melatonin receptor GPR50 was extensively expressed in fibroblasts of hypertrophic scar, but not in fibroblasts in normal skin. RT-PCR showed that the expression of melatonin receptor( MT1, MT2 ) mRNA in hypertrophic scar was significantly higher than that in normal skin( P <0. 05). In normal skin and hypertrophic scar group, the expression of MT1 mRNA was higher than MT2 mRNA ( P < 0. 05 ) . In normal skin and hypertrophic scar group, the expression of MT1 mRNA was 0.99081 ±0.26485 and 1.16584 ±0.21829 copy number/μl cDNA, respectively;the expression of MT2 mRNA was 0. 77083 ±0. 15927and 0. 99550 ±0. 14624 copy number/ μl cDNA, respectively. Sequencing results indicated that the positive product coincided with cDNA of human melatonin receptor in GeneBank. Conclusions Positive expression of melatonin receptor can be found in human hypertrophic scar and normal skin, but it is higher in scar. The over expression of melatonin receptor in hypertrophic scar may be related to the development of hypertrophic scar.