The Hemodynamic Characteristics Of The Extended Flap And The Preliminary Investigation Of Mechanism Underlying Choke Vessel Dilation

ObjectiveThe flap is the method to repair the defected tissues in clinical practice.The most commonly used flaps are the perforator flap,the perforator-based fasicocutaneous flap and the random flap.This study adopted the laser doppler flowmetry for measurement of blood perfusion on zone ??? and ? of the above-mentioned three kinds of flap at the multiple time points.One of the three models was chosen to test the expression,location and activity of matrix metalloproteinase-2(MMP-2)and matrix metalloproteinase-9(MMP-9)in the choke area through which the mechanism of flap survival was investigated,providing the basis for clinical flap transplantation to repair the defect.Methods1.The caudal border of the flap with a dimension of 3×10cm was located about 1cm posterior to the point where the iliolumbar perforator was located.Perforator flap: the flap was completely detached from the surrounding skin,depending only on the iliolumbar vessels for perfusion and drainage;the perforator-based fasciocutaneous flap: the caudal side of the flap is not detached from the surrounding skin,allowing double pathways for supply and drainage for the flap from both the iliolumbar vessels and the remaining skin pedicle;the random flap: the iliolumbar vessels were severed,leaving the flap supplied and drained only through the skin pedicle.Laser doppler flowmeter was adopted for measurement of blood perfusion on zone???and ? of the flaps on the three groups at the multiple time points and compared with each other.2.Using the dorsal skinfold chamber technique and the numeric control devices to observe the continuous and focal morphological change of the vessels and capture a panoramic view of the vascular evolution of the whole flap;using the transmission electron microscopy to observe micrographic images of vessels in the choke zone and the degeneration of axons after flap harvest;using immunohistochemical to observe the location of MMP-2 and MMP-9;using protein immunoblot(Western blot)to detect the expression of MMP-2 and MMP-9 at the multiple time points;using gelation zymography to test the activity of MMP-2 and MMP-9 at the multiple time points.Results1.The necrotic rates of the perforator flap,the perforator-based fasciocutaneous flap and the random flap had statistical difference,but there was no statistical difference between the perforator flap and the perforator-based fasciocutaneous flap.Immediately after surgery,the perfusion value of zone? and? in the perforator flap group was larger than that in the perforator-based fasciocutaneous flap group with statistical significance.Perfusion value of the three zones in the random flap group was lower than their counterparts of the perforator flap group and the perforator-based fasciocutaneous flap.7 d after surgery,the vascular network between the iliolumbar perforator and the sacrococcygeal perforator underwent tremendous enlargement in diameter.2.The micrographic images of vessels in the choke zone showed: the arteries had three layers,and the veins had one to two layers of smooth muscle cells on day 0.On day 1,no particular ultrastructural changes of the arteries and veins could be observed.On day 3,abundance of rough endoplasmic reticulum in the arterial wall could be observed,indicating a proliferative phenotype.And widespread edema of mitochondria and disappearance of cristae could be observed in the venous wall.Splitting of lamellae,inward folding and disintegration of the myelinated sheath around the axons could be observed,and cellular organelles within the axons were also breaking down.On day 7,6 layers of smooth muscle cells existed in an arterial wall but were arranged in a seemingly chaotic order.A striking phenomenon was the adherence of a plethora of lymphocytes along the vascular adventitia.3.The immunohistochemical results showed: MMP-2 immunopositive cells increased on day 1,3,5 and 7.Immunoreactivity of MMP-2 was localized predominantly in the endothelium,diffuse staining also appeared in the tunica media and tunica adventitia of vessels.Almost no MMP-9 immunopositive cells could be observed on skin tissues of day 0.On the following days after flap harvest,the immunopositive cells were mainly located in the lumen inclined to adhere to the endothelium of vessels.As for PGP 9.5,strong immunostaining corresponding to existence of axons within nerve fascicles could be observed on day 0 and 1.Weak staining of axons began from day 3 to day 7.4.Western blots showed increased expression of MMP-2 and MMP-9 with statistical significance in the choke areas after flap harvest.Post-hoc analysis showed that the expression of MMP-9 on day 1 was significantly higher than on day 0;Gelation zymography showed increased activity of MMP-9 with statistical significance while no increased activity of MMP-2 was noted after flap harvest.Post-hoc analysis showed that the activity of MMP-9 on day 1 was significantly higher than that on the other four time points,which was corresponding to results of western blots.ConclusionsThere is no significant difference on the survival capacity between the perforator flap and the perforator-based fasciocutaneous flap.The survival of the random flap depends on the dilation of the vascular network between the pedicle and the nearest potential perforator.Two methods were adopted for observation of the morphological evolution of vessels within the flap.The dorsal skinfold chamber technique cominbed with the numeric control device can provide dynamic and high-quality images of the vascular evolution within the flap.The morphology observed in this study indicated that the venous blood returned to the pedicle through the bypass route in the first three days after surgery and then through the denervated veins later on.MMP-9 might play a more important role than MMP-2 in the vascular evolution of an extended flap.