皮瓣延迟术对大鼠三血管体穿支皮瓣成活的影响及其机制

Effects of surgical delay procedure on the survival of perforator flap with three angiosomes in rat and its mechanism

目的 观察皮瓣延迟术对大鼠三血管体穿支皮瓣成活的影响并探讨其可能机制，方法 本实验采用大鼠背部右侧以旋髂深动脉为蒂的三血管体穿支皮瓣模型,将3个血管体之间的2个交界区域由皮瓣蒂部开始以远依次命名为闭塞区域1、闭塞区域2.按随机数字表法将110只SD大鼠分为常规皮瓣组40只、单纯延迟组30只、延迟皮瓣组40只，（1）于常规皮瓣组中,按随机数字表法选取30只大鼠直接行皮瓣手术,术后0（即刻）、1、2、3、7d分别处死6只,取2个闭塞区域全层皮肤组织行HE染色检测血管密度及外径.剩余10只大鼠颈外静脉置管后行皮瓣手术,从术后0d（5只）或术后1d（5只）开始每隔2d注射1.5 mL荧光素钠（100 g/L）观察皮瓣血运变化,每只注射4次;术后7d,计算10只大鼠皮瓣成活率,采用改良氧化铅-明胶灌注法行血管造影观察皮瓣内血管形态与分布，（2）单纯延迟组大鼠仅通过局部皮肤小切口手术结扎右侧胸背动脉,术后0（即刻）、1、2、3、7 d分别处死6只大鼠同常规皮瓣组部位取皮肤标本测量血管密度及外径.（3）延迟皮瓣组大鼠按单纯延迟组术式行延迟手术,术后7d按与常规皮瓣组相同的方法选取、分配及处理本组所有大鼠并检测相应指标.对数据行成组样本t检验、析因设计方差分析、SNK检验， 结果 （1）3组间比较,术后7d闭塞区域1、闭塞区域2血管密度差异均明显（F值分别为2.69、2.76,P值均小于0.05），延迟皮瓣组大鼠术后7d闭塞区域1、闭塞区域2血管密度分别为（29±7）、（31±8）个/mm^2,显著高于常规皮瓣组的（23±5）、（23±3）个/mm2（q值分别为5.67、6.01,P值均小于0.05）,且显著高于组内术后0d水平（q值分别为6.42、7.14,P值均小于0.05），术后3、7d,常规皮瓣组、延迟皮瓣组大鼠闭塞区域1血管外径显著高于单纯延迟组（q值为8.15～11.13,P值均小于0.05）;延迟皮瓣组术后0、1、2、3、7 d[（65±8）、（63±13）、（69 ±9）、（67±8）、（64±13） μm]及单纯延迟组术后3、7d闭塞区域2Ⅱ血管外径显著高于常规皮瓣组相应时相点血管外径水平[术后0、1、2、3、7d分别为（46±10）、（40 ±9）、（43±13）、（46±12）、（47±11）μm],q值为7.29～10.79,P值均小于0.05.常规皮瓣组、单纯延迟组大鼠术后3、7d以及延迟皮瓣组大鼠术后0、1、2d组内闭塞区域1与闭塞区域2间血管外径比较,差别明显（q值为5.32～9.56,P值均小于0.05），术后3、7d,常规皮瓣组及延迟皮瓣组大鼠闭塞区域1、单纯延迟组大鼠闭塞区域2血管外径显著高于组内术后0d（q值为6.12～8.13,P值均小于0.05），（2）术后7d内,常规皮瓣组、延迟皮瓣组大鼠皮瓣内来自蒂部的血液都顺利通过闭塞区域1覆盖血流动力学供区，术后0d,2组大鼠皮瓣内的血液越过闭塞区域1后均发生持续约3 min的血流阻塞现象,常规皮瓣组出现在闭塞区域2附近,延迟皮瓣组出现在闭塞区域2远端约1 cm处，（3）术后7d,延迟皮瓣组大鼠皮瓣成活率为（95±12）％,显著高于常规皮瓣组的（80±9）％（t=2.91,P ＜0.01）,皮瓣部分坏死仅出现在潜在供区，（4）术后7d,与未行手术的左侧皮肤血管相比,常规皮瓣组大鼠皮瓣内闭塞区域1的血管扩张明显,相邻血管树间界限模糊,而闭塞区域2的血管变化轻微;延迟皮瓣组大鼠皮瓣内2个闭塞区域的血管均明显扩张， 结论 本实验的延迟方法可促进大鼠三血管体穿支皮瓣潜在供区的成活,其主要是通过皮瓣术前扩张闭塞区域2的闭塞血管实现的。

Objective To observe the effects of surgical delay procedure on the survival of perforator flap with three angiosomes in rat,and to explore its possible mechanism.Methods The flap model was a perforator flap with three angiosomes which located on the right dorsal side of a rat based on thc right deep circumflex iliac vessel.The two connection areas between the three angiosomes were successively named choke zone （CZ） 1 and CZ 2 beginning from the pedicle to the remote area.A total of 110 SD rats were divided into routine flap group （RF,n =40）,delay only group （DO,n =30）,and delay flap group （DF,n =40） according to the random number table.（1） In group RF,30 rats were selected according to the random number table,and flap surgery was performed directly.Six rats were sacrificed on post operation day （POD） 0,1,2,3,7 respectively to collect the full-thickness skin samples at both CZs for HE staining to measure the vascular density and diameter.The rest 10 rats underwent flap surgery immediately after a catheter was successfully implanted into their external jugular vein.A volume of 1.5 mL sodium fluorescein solution （100 g/L） was injected to the 10 rats on POD 0 （5 rats） or POD 1 （5 rats） each time with a 2-day interval to learn the change in flap circulation.Each rat was injected for 4 times.The flap survival rate of the 10 rats was calculated on POD 7,and the configuration and distribution of the vessels in the flap were observed through angiography with the improved perfusion method of lead oxide-gelatin.（2） In group DO,the right thoracodorsal perforators of all the rats were surgically ligated through a small skin incision,and 6 rats were sacrificed on POD 0,1,2,3,7 respectively.The skin samples of each rat at the same area as in group RF were harvested to measure the vascular density and diameter.（3） In group DF,rats were treated with ligation surgery as in group DO,and then they were assigned and treated as in group RF on POD 7 with corresponding indexes detected later.Data were processed with group t test,analysis of variance with factorial design,and SNK test.Results （1） Significant differences of vascular density at both CZ 1 and CZ 2 were found on POD 7 among the three groups （with F values respectively 2.69 and 2.76,P values below 0.05）.The vascular density values of CZ 1 and CZ 2 of rats in group DF were （29 ±7） and （31±8） per mm2 on POD 7,which were significantly higher than those of group RF [（23 ± 5） and （23 ± 3） per mm2,with q values respectively 5.67 and 6.01,P values below 0.05] and those within group DF on POD 0 （withqvalues respectively 6.42 and 7.14,Pvaluesbelow0.05）.On POD3 and7,the vascular diameter values of CZ 1 of rats in groups RF and DF were significantly higher than those of group DO （with q values from 8.15 to 11.13,P values below 0.05）.The vascular diameter values of CZ 2 of rats in group DF on POD 0,1,2,3,7 [（65±8）,（63±13）,（69±9）,（67 ±8）,（64±13） μm] and in group DO on POD 3 and 7 were significantly higher than those in group RF [respectively （46 ±10）,（40 ± 9）,（43±13）,（46±12）,（47±11） μmonPOD0,1,2,3,7] at corresponding time point （withqvalues from 7.29 to 10.79,P values below 0.05）.The difference in vascular diameter between CZ 1 and CZ 2 was statistically significant in groups RF and DO on POD 3 and 7,and in group DF on POD 0,1,and 2 （with q values from 5.32 to 9.56,P values below 0.05）.Compared with that on POD 0 within each group,the vascular diameter of CZ 1 in groups RF and DF and that of CZ 2 in group DO increased significantly on POD 3 or7 （with q values from 6.12 to 8.13,P values below 0.05）.（2） In groups DF and RF,blood from the pedicle ran through CZ 1 and covered the dynamic territory successfully within POD 7.On POD 0,the blood within all flaps was blocked for about 3 min after going through CZ 1 at 1 cm distal from CZ 2 in group DF and around CZ 2 in group RF.（3） Flap survival rate of rats in group DF was （95 ±12） %,which was statistically higher than that of group RF [（80 ±9） %,t =2.91,P 〈0.01].All the partial flap necrosis occurred in potential territory.（4） Compared with the vessels in the left dorsal side without surgery,the vessels of CZ 1 in group RF were dilated obviously,and the boundary between vascular trees became indistinct,but the vessels in CZ 2 changed slightly; the vessels in both CZs in group DF were dilated dramatically.Conclusions The delay method could enhance the survival of potential territory in perforator flap with three angiosomes,and it acted mainly by dilating the choke vessels in CZ 2 before flap surgery.