两种不同肌筋膜瓣包裹脂肪来源干细胞载体复合物体内成脂效率的比较研究

目的 比较两种不同肌筋膜瓣包裹脂肪来源干细胞(adipose-derived stromal cells,ADSCs)与胶原蛋白支架复合物在体内成脂效率的差异,为临床游离脂肪组织的高效移植提供实验依据. 方法 3～4月龄新西兰大白兔10只,雌雄不限,体重2.0～2.5 kg.取兔颈部皮下脂肪组织分离ADSCs并行原代及传代培养,待第3代细胞生长至瓶底70%～80%面积时,以BrdU(10μg/mL)标记细胞48 h,行免疫荧光染色观察;成脂诱导2周后行油红O染色观察;成骨诱导3周后行茜素红染色观察;成软骨诱导2周后行阿利辛蓝染色观察.另取第3代经BrdU标记的ADSCs成脂诱导2周后,以1×107个/块接种于10 mm×10 mm×5 mm大小的Col Ⅰ中,制备细胞载体复合物.实验分为两组,A组由带血管蒂的右侧背阔肌筋膜瓣包裹复合物,B组由无特定血管蒂的右侧臀大肌筋膜瓣包裹复合物,每只动物均采用自体ADSCs移植及自身对照.术后8周取出移植物,经HE染色和免疫组织化学染色鉴定新生组织,测定两组新生组织的湿重和微血管数,并采用免疫荧光染色鉴定新生组织及微血管内皮细胞的来源. 结果 荧光显微镜下BrdU标记为阳性的ADSCs胞核呈绿色荧光,ADSCs阳性标记率>90%.第3代ADSCs成脂诱导2周,可见细胞内有红色脂滴形成;成骨诱导3周,可见红色钙结节样改变;成软骨诱导2周,可见高度聚集生长的细胞团,阿利辛蓝染色呈蓝色.术后8周取材,A组可见血管增生并长入材料,未见明显纤维包裹;B组有少量血管长入材料.A组新生组织湿重为(0.1495±0.017 3)g,B组为(0.095 3±0.012 7)g,比较差异有统计学意义(P<0.01).HE染色均可见移植物中有新生脂肪组织形成和不同程度微血管长入,支架材料已基本降解吸收.A组微血管数为(31.2±4.5)根,B组为(19.3±2.6)根,比较差异有统计学意义(P<0.01).术后8周新生组织免疫荧光染色示,A、B组新生脂肪细胞的胞核及部分微血管内皮细胞的胞核呈绿色荧光. 结论 带血管蒂的背阔肌筋膜瓣包裹ADSCs与胶原蛋白支架复合物在体内的成脂效率较无特定血管蒂的臀大肌筋膜瓣高.     

组织工程化人工骨血管化研究

目的 比较三种促进组织工程化人工骨体内血管化方法的效果及研究血管化与成骨的相关关系。方法 将HA/β-TCP与PDLLA形成复合支架材料，再复合Ⅰ型胶原、rhBMP-2，与成骨细胞联合培养，仿生制备成组织工程化人工骨，采用包裹带血管蒂筋膜，复合血管内皮细胞或两者联合的促血管化手段。将人工骨修复兔桡骨干骨膜-骨完全缺损，手术后4、8、12周，观察移植物组织学，体视学方法观察血管化，成骨作用及其两者的关系。结果 3种方法均有促血管作用。其优劣依次为；材料包裹带血管蒂筋膜及复合血管内皮细胞大于材料单纯复合血管内皮细胞，后者又大于材料单纯包裹带血管蒂筋膜，术后4周为快速血管化阶段。4-8周间血管化有平缓发展，12周完全血管化，血管化与新骨生成数量成正相关。结论 促血管化手术对组织工程化人工骨移植修复骨缺损的效果起重要促进作用。     

前臂逆行骨间背侧动脉皮瓣手术技巧探讨

目的探讨提高逆行骨间背侧动脉皮瓣修复、皮肤软组织缺损成功率的手术技巧。方法应用前臂逆行骨间背侧动脉皮瓣修复手腕部及前臂下1/3皮肤软组织缺损34例。皮瓣蒂部包含骨间背侧血管、肌间隔、2-3 cm宽深筋膜和皮下组织,部分蒂部尚包括筋膜蒂上的皮条。结果32例皮瓣完全成活,1例皮瓣远端表皮坏死,1例皮瓣远端1.5 cm部分坏死。无一例桡神经深支及其肌支损伤。结论在血管蒂周围带2-3 cm宽的筋膜蒂能使骨间背侧动脉皮瓣更安全,成活更可靠。     

GFP转基因鼠肌卫星细胞系的构建及基因介导成骨分化的实验研究

目的 评价肌卫星细胞(Muscle satellite cells,MSCs)体内外成骨分化的能力,探讨肌卫星细胞作为骨组织工程新的种子细胞的可能性.方法 取新生绿色荧光蛋白转基因小鼠颌面部肌肉,采用酶消化法分离出MSCs,体外原代及传代培养,经腺病毒介导的人骨形成蛋白2(Ad-BMP2)基因转染后,行成骨细胞标志物的活性检测及细胞化学染色,并进行细胞体外矿化及体内成骨的检测.结果 转染后细胞碱性磷酸酶(ALP)组织化学染色呈阳性;骨钙素(OC)免疫细胞化学染色呈阳性且OC活性增强(p＜0.05);21天后见钙结节形成;转染后的细胞与材料复合物植入裸鼠后肢肌肉内,4周后见骨组织形成.结论 体外培养的肌卫星细胞体内外可以成骨分化,可以成为骨组织工程的新的种子细胞.     

带蒂肌瓣包裹异种无机骨与自体红骨髓复合物的实验研究

目的 制备带血运并具有新骨组织形成的,来源于异种无机骨与自体红骨髓复合物的肌骨瓣,方法 6月龄新西兰大白兔12只,实验组,每只兔左前肢尺桡骨旁肌肉内植入异种无机骨与自体红骨髓复合物,对照组,右侧相同部位植入单纯异种无机骨,术后2、8、12周进行X线片,大体标本和组织学观察。结果 两组X线片显示植入物呈松质骨样密度,不随植入时间改变,大体观察见植入后2周植入物与肌肉完全融合,包裹有植入物的带蒂肌瓣的血管清晰可见,将此包裹有植入物的带蒂肌瓣游离30分钟后,植入物仍然与周围组织色泽相同,无缺血坏死改变,组织学观察植入后2、8、12周显示植入部位肌肉组织与植入物相连接,但实验组植入后2周,血管进入复合物中,无新骨形成,8周,异种无机骨边缘有较少的新骨形成征象;12周,新骨组织形成明显增多,成骨细胞位于骨的边缘,周围为未分化间充质细胞,血管和骨基质丰富,骨细胞包绕在骨陷窝中,偶尔可见骨髓腔,对照组直至12周,异种无机骨孔隙间仅有带血管的疏松结缔组织无新骨形成,结论 异种无机骨与自体红骨髓复合物植入肌肉内12周可预制成具有血运的、有新骨形成的肌骨瓣。     

显微外科技术预构组织工程骨血管化的比较

目的 比较3种带血管蒂组织瓣预构组织工程骨血管化的效果,探讨其影响血管化的方式.方法 设计以膝最下血管为蒂的股骨骨膜瓣包裹、贯穿,腹壁浅血管为蒂的筋膜瓣包裹体外培养的磷酸三钙(β-TCP)+骨髓间充质干细胞(MSCs)复合物等3种组织工程预构骨动物模型,以β-TCP+MSCs复合物和单纯β-TCP皮下埋放为对照,通过墨染透明标本、墨染苏木素-伊红(HE)切片、第8因子相关抗原免疫组织化学染色、铸型标本扫描电镜观察等指标,分析带血管蒂组织瓣对组织工程预构骨血管化的影响.结果 新生血管自组织瓣与材料结合的界面向材料内部生长,组织瓣处理各组的血管化指标均显著优于非组织瓣处理组(P＜0.05),前者中血管化的优劣递次为A、B、C组(P＜0.05);材料内部的孔隙中亦可观察到散在的血管,复合了MSCs的D组血管化优于单纯材料的E组(P＜0.01).结论 影响带血管蒂组织瓣组织工程预构骨血管化的因素有:(1)与材料复合的组织瓣血供及其与材料接触面积的大小;(2)提供给材料内部促进血管发生的细胞及细胞因子的种类及密度(包括内源性和外源性的);(3)材料本身的特性,如组织相容性、是否有贯通的微孔等.     

健侧下肢组合血管皮瓣桥携游离(骨)皮瓣转移修复对侧下肢严重创伤

目的 :探讨下肢严重创伤需行游离组织移植而患侧下肢无血管可供吻接时的修复方法。方法 :健侧小腿胫后血管蒂与带蒂保留真皮下血管网皮瓣组成皮瓣桥 ,携带皮瓣或骨皮瓣转移修复患肢。结果 :6周后经血管造影皮瓣与周围组织血运建立。断蒂后皮瓣与骨皮瓣一期成活 ,后期复查移植骨成活并且增粗。结论 :下肢严重创伤无血管可供吻接时健侧胫后血管皮瓣桥可作为一种理想的供血途径 ,这为下肢严重创伤提供一种新的修复方法。     

健侧下肢组合血管皮瓣桥携游离（骨）皮瓣转移修复对侧下？…

目的：探讨下肢严重创作需行游离组织移植而患侧下肢无血管可供吻接时的修复方法。方法：健侧小腿胫后血管蒂与带蒂保留真皮下血管网皮且成皮瓣桥,携带皮瓣或骨皮瓣转移修复患肢。结果：６周后经血管造影皮瓣与周围组织血运建立。断蒂后皮瓣与骨皮瓣一期成活,后期复查移植骨成活并且增粗。结论：下肢严重创伤无血管可供吻接时健侧胫后血管皮瓣桥可作为一种理想的供血途径,这为下肢严重创伤提供一种新的修复方法。     

显微外科与血管化组织工程组织的构建

血运重建是组织再生的基础，要获得组织工程化组织必须保证植入体内的种子细胞尤其是材料内部的种子细胞能够获得及时充分的营养。因此血供的重建就成为组织工程由基础向临床应用的关键性环节。血液供应的重建方法主要有：利用血管内皮细胞生长因子促进血管生长，采用血管内皮细胞与成骨细胞联合移植的方法，或应用显微外科技术。利用显微外科技术，可通过带蒂筋膜瓣包裹、带血供肌瓣包裹或血管束植入的方法，在构建组织工程化组织的同时建立其血液供应。组织工程化组织在临床应用时，如果修复邻近的缺损，只需在原位进行血管化；但在缺损局部软组织条件差、常常无法找到可供利用的筋膜瓣、肌瓣或血管束的情况下，可异位构建血管化的组织工程组织，二期采用显微外科方法通过带血管的组织移植修复缺损。     

骨髓基质细胞体内成骨分化研究

目的探讨将自体骨髓基质干细胞（BMSCs）移植到股骨头缺损坏死部位后能否成活、增殖,并向成骨分化,促进骨修复。方法取狗犬髂骨骨髓,体外培养扩增BMSCs,经5-溴脱氧尿嘧啶核苷（BrdU）标记,移植到股骨头骨缺损模型。术后5周取材。行大体组织观察,组织化学染色,免疫组化检测骨钙素、骨桥素。结果BMSCs能增加骨缺损区内的成骨量、提高骨基质矿化的程度和骨成熟度,提高成骨细胞分化特异性标记物骨钙素、骨桥素的表达。新骨形成越活跃的地方,BrdU阳性细胞数目越多,很多成骨细胞BrdU阳性,新骨形成区的很多血管壁也分布着BrdU阳性细胞。结论移植的骨髓细胞在股骨头内能成活,并能停留在骨坏死、缺损部位生长增殖。移植的BMSCs能分化成为多种细胞,包括成骨细胞、血管壁细胞等。BMSCs不但能直接通过分化成成骨细胞参与成骨,也能通过参与血管形成而促进成骨。     

预构骨肌皮瓣修复下颌骨和皮肤复合缺损的实验研究

目的 探讨BMP-2与胶原复合材料在骨骼肌中异位成骨预构骨肌皮瓣,修复自体下颌骨和皮肤复合缺损的可行性. 方法 4～6周龄新西兰白兔24只,体重2.0～2.5 kg,雌雄不拘,随机分成3组,即实验组、对照组和空白组(n=8).将BMP-2与胶原复合,植入兔背阔肌,预构异位新生骨组织,2、4、6周后行X线片、ALP、Von Kossa、CD31免疫组织化学血管标记及组织学观察.6周后,实验组动物于同侧下颌骨体部制备2 cm × 3 cm皮肤缺损,并形成直径8 mm洞穿性骨缺损;以同侧胸背动脉体表投影为皮瓣纵轴,设计含预制新骨组织的背阔肌肌皮瓣,带蒂移位修复下颌骨及皮肤复合缺损.对照组和空白组实验动物下颌骨体部均造成直径8 mm的洞穿性骨缺损,对照组将复合组织瓣中的骨性部分游离移植修复缺损;空白组缺损不予修复.术后6周,各组取材行四环素荧光染色、X线摄片、组织学观察以及新骨计量等,观察骨和皮肤复合缺损修复效果. 结果 BMP-2与胶原复合材料在兔背阔肌中4～6周成骨,胶原材料于植入后3～5周降解,成骨过程以软骨成骨为主,新骨形态为编织骨,可见明显的微血管分布.修复自体下颌骨缺损6周后,实验组皮肤及骨缺损均愈合良好,对照组缺损修复区基本骨化,空白组尚残留大块骨缺损实验组、对照组及空白组新骨计量分别为(1.594 ±0.674)、(0.801 ±0.036)和(0.079±0.010)mm2,组间两两比较,差异均有统计学意义(P＜0.05). 结论 预构的骨肌皮瓣带血管蒂移位修复兔自体下颌骨和皮肤复合缺损具有可行性和明显优势,有望成为一种血管化骨移植的供区预构形式.     

One-stage reconstruction of facial paralysis associated with skin/soft tissue defects using latissimus dorsi compound flap.

Neurovascular free muscle transfer is now the mainstay for smile reconstruction in the treatment of established facial paralysis. Since facial paralysis due to ablative surgery or some specific disease sometimes accompanies defects of the facial skin and soft tissue, simultaneous reconstruction of defective tissues with facial reanimation is required. The present paper reports results for 16 patients who underwent reconstruction by simultaneous soft tissue flap transfer with latissimus dorsi muscle for smile reconstruction of the paralysed face. Soft tissue flaps comprised skin paddle overlying the latissimus dorsi muscle (n=6), serratus anterior musculocutaneous flap (n=5), serratus anterior muscle flap (n=2), and latissimus dorsi perforator-based flap with a small muscle cuff (n=3). The latissimus dorsi muscle can be elevated as a compound flap of various types, and thus offers the best option as a donor muscle for facial reanimation when soft tissue defects require simultaneous reconstruction.     

The creation of new rotation arc to the rat latissimus dorsi musculo-cutaneous flap with delay procedures

Background

Latissimus dorsi musculocutaneous flap is one of the most frequently performed reconstructive techniques in surgery. Latissimus dorsi muscle has two arcs of rotation. It is classified as type V muscle. This muscle can be elevated on the thoracodorsal artery to cover large defects in the anterior chest and also, the muscle can be elevated on the segmental vessels to cover midline defects posteriorly. The aim of this study was to create a new arc of rotation on a vertical axis for the muscle and investigate effectiveness of vascular and chemical delays on the latissimus dorsi muscle flap with an inferior pedicle in an experimental rat model. We hypothesized that the latissimus dorsi muscle would be based on inferior pedicle by delay procedures.

Methods

We tested two different types of delay: vascular and combination of vascular and chemical. We also tried to determine how many days of "delay" can elicit beneficial effects of vascular and combination delays in an inferior pedicled latissimus dorsi musculocutaneous flap. To accomplish this, 48 male Sprague-Dawley rats were randomly subjected to vascular or combination delay (vascular and chemical). In addition, one ear of each rat was assigned into a delay procedure and the other ear was used as a control. Results were evaluated macroscopically, and micro-angiography and histological examinations were also performed. As a result, there was a significant difference in viable flap areas between vascular delay alone and control groups (p < 0.05).

Results

The higher rate of flap viability was obtained in seven-day vascular delay alone. However, there was no significant difference in the viability between seven-day vascular delay and five-day vascular delay (p < 0.05), so the earliest time when the flap viability could be obtained was at five days. The rate of flap viability was significantly higher in the vascular delay combined with chemical delay than the control group (p < 0.05).

Conclusion

The combination of vascular and chemical delays increased the rate of viability. Nevertheless, there was no significant difference between vascular delay alone and combination of vascular and chemical delays. Chemical delay did not significantly decrease the delay period. Better histological and microangiographical results were achieved in delay groups compared to control groups. We concluded that the arch of the latissimus dorsi musculocutaneous flap can be changed and the flap can be used for various purposes with the delay procedures.     

Experimental study on artificial esophagus created by a latissimus dorsi muscle flap with grafted jejunal mucosa

An experimental study on artificial esophagus was performed in 38 mongrel dogs. Artificial esophagus was created by using a latissimus dorsi muscle, woven dacron vascular prosthesis and jejunal mucosa. A prepared latissimus dorsi muscle flap was cylindrically rolled, and jejunal mucosa with a woven dacron vascular prosthesis as a stent was inserted into this muscle canal. Although, all of these mucosal grafts fell down to necrosis with severe bacterial infections. The main cause of this failure was poor blood supply to the grafted mucosa via the muscle flap. For this reason, artificial granulation tissue with newly developed abundant vessels was made on the muscle surface by means of an insertion of the woven dacron vascular prosthesis into the muscle canal. Two weeks later, inner surface of the muscle canal had good granulations with a rich blood flow. And then the free jejunal mucosa of 10cm in length was grafted into this canal. Consequently, all grafted mucosa were well survived and had normal mucosal structures in the whole transplanted length. Based on these results, a latissimus dorsi muscle flap with the mucosal graft is possible to use as an artificial esophagus.     

Intrathoracic application of the reverse latissimus dorsi muscle flap.

The use of the reverse latissimus dorsi muscle flap based on its paraspinous perforators for posterior trunk wound coverage has been described previously. However, few studies have reported its intrathoracic application. In this study the authors present their experience in treating 3 patients with various intrathoracic defects using the reverse latissimus dorsi muscle flap. There were 1 male and 2 female patients who ranged in age from 4 to 74 years (mean, 49 years). The etiology included an infected aortic graft, a bronchopleural fistula, and a recurrent congenital diaphragmatic hernia. Follow-up ranged from 2 to 24 months. Successful outcomes were achieved in all 3 patients, and there was no recurrence or wound complication identified. Their results demonstrate the versatility and reliability of the reverse latissimus dorsi muscle flap in treating low posterior intrathoracic defects.     

机器人和腔镜背阔肌乳房重建关键技术

机器人和腔镜背阔肌乳房重建是目前乳房重建的重要手术方式,包括背部供区“无痕化”背阔肌乳房重建、“皮岛式”背阔肌乳房重建。其特点为：腋窝无切口,仅做3个trocar孔,在微创视野下通过皮下隧道完成背阔肌皮瓣切取或转移,随后3个trocar孔可用于术后引流,不增加额外切口;背部供区不遗留瘢痕;微创视野下获取背阔肌肌瓣整个过程不需要拉钩。机器人和腔镜辅助背阔肌乳房重建优势为切口隐蔽、创伤小,美容效果好,是一种安全、可行的手术。     

背阔肌肌瓣转移修复Poland综合征胸部畸形

目的 探讨背阔肌肌瓣转移矫治Poland综合征胸部畸形的方法与注意事项。方法 10例患者采用侧胸垂直切口及腋下小切口切取转移背阔肌肌瓣矫正胸壁畸形,注意腋前皱襞的重建。结果 10例背阔肌肌瓣全部成活,随访1～2年,效果良好。结论 背阔肌肌瓣具有稳定、可靠的血供保证,能提供足够的组织量,是修复Poland综合征胸部畸形的理想方法。     

The anatomy of the thoracodorsal artery within the latissimus dorsi muscle

This paper reports the findings of an anatomical study of the primary and secondary muscular branches of the thoracodorsal artery within the latissimus dorsi muscle. Twenty latissimus dorsi muscles were studied, harvested from 13 fresh cadavers and the findings support the hypothesis that it is possible to thin safely a latissimus dorsi muscle flap at the time of flap transfer.     

背阔肌肌皮瓣在前胸部肿瘤切除后皮肤缺损中的应用

目的探讨应用背阔肌肌皮瓣修复前胸部肿瘤切除后皮肤缺损的临床疗效。方法对28例前胸部皮肤肿瘤患者根据其性质切除肿瘤后,按皮肤肌肉缺损面积,设计背阔肌肌皮瓣转移至缺损区修复。结果28例患者相应的局部皮瓣转移修复至缺损区,术后皮瓣均全部成活,供受区切口均Ⅰ期愈合,随访3～24个月,效果满意。结论背阔肌肌皮瓣是临床上修复胸部肿瘤切除后皮肤缺损的实用而有效的方法。     

机器人和腔镜背阔肌乳房重建关键技术

机器人和腔镜背阔肌乳房重建是目前乳房重建的重要手术方式，包括背部供区“无痕化”背阔肌乳房重建、“皮岛式”背阔肌乳房重建。其特点为：腋窝无切口，仅做3个trocar孔，在微创视野下通过皮下隧道完成背阔肌皮瓣切取或转移，随后3个trocar孔可用于术后引流，不增加额外切口；背部供区不遗留瘢痕；微创视野下获取背阔肌肌瓣整个过程不需要拉钩。机器人和腔镜辅助背阔肌乳房重建优势为切口隐蔽、创伤小，美容效果好，是一种安全、可行的手术。