Observation Of Effect Of Long-term Combination Of Low-dose FK506 And Lithium Chloride On Nerve Regeneration After Sciatic Nerve End-to-side Anastomosis In Rats

Objective:1. To investigate the FK506 and lithium chloride promote the effect of regeneration after sciatic nerve injury in rats and the mechanism;2. we investigated the feasibility and superiority of FK506 and lithium chloride of repair of rat sciatic nerve injury;3. To investigate the different dose of FK506 and lithium chloride on the effect of regeneration and functional recovery after sciatic nerve injury, to lay a good foundation for the clinical treatment of peripheral nerve injury repair.Method:49 adult male Sprague Dawley rats which received end-to-side neurorrhaphy were randomly divided into 7 groups: Group A, 7 rats received intraperitoneal injection with saline water 1mg/kg/d;Group B, 7 rats received intraperitoneal injection with FK506 1mg/kg/d;Group C,7 rats received intraperitoneal injection with FK506 0.5mg/kg/d;Group D, 7 rats received intraperitoneal injection with Li Cl 85mg/kg/d;Group E, 7 rats received intraperitoneal injection with Li Cl 40mg/kg/d;Group F,7 rats received intraperitoneal injection with Li Cl 40mg/kg/d and FK 506 0.5mg/kg /d;Group G, 7 rats received intraperitoneal injection with Li Cl 85mg/kg/d and FK506 1mg/kg/d.After operation,all of the rat received intraperitoneal injection everyday, respectively,both for 12 wk. Histological and morphological examinations were performed 12 wk after the operation.In the 2nd, 4th, 6th, 8th, and 12 th wk after operation,function recovery analysis was performed.Results:1 General observationsAll rats postoperative incision healing,no infection and death.A few see a foot ulcer in rats,all healing within two weeks.Early postoperative all rats hind limbs movement clumsy,knee flexion dysfunction.After12 weeks observation,the experimental group rats of knee flexion dysfunction improved,acupuncture is responsive.While the control group to improve is poorer,acupuncture response is relatively slow.Obviously,after the application of FK506(group B、C、F、G) animal weight is significantly lower than other groups(A、D、E)(P>0.05).The application FK506 also have significant differences in drug group(C>F>G>B,P<0.05).There is no statistical difference between group D and group E(P>0.05).(Fig.5)2 Sciatic nerve function recovery(SFI)SFI of the analysis of the rat sciatic nerve index measurement and comparison on postoperative 2,4,6,8,10,12 weeks:in groups,the normal gait footprints(left) shows for various toe hind PAWS at full stretch,and side(right) is shown as a limb transplantation after claw "foot drop and flexion contracture,each toe adduction.No difference was observed in the 2nd and 4th week(P>0.05)between three groups. In the 12 wk,the analyzed variables did not show any differences among the six operated groups(P>0.05),which were statistically superior to group A(P<0.05).(Table.1)3 Muscle mass preservation12 weeks after operation,all rats are killed and get the extensor digitorum longus muscles,weighting out the muscle wet weight with the analysis of the balance and observe the calf muscle atrophy.According to the following formula: The experimental side muscle wet weight as a percentage of the normal side= The experimental side muscle weight/ the contralateral muscle weight.Significant difference between control group and experimental group(P>0.05),but there is no difference between B group and G group(experimental group)(P>0.05),however they are better than other groups(P<0.05).(Table.1)4 Immunohistochemical staining12 weeks after operation,Experimental group shows a lot of regenerating myelinated nerve fibers and fibers mixed unmyelinated,Schwann cells are morphologically normal,surrounding the regeneration of axons form myelinated fibers,most of the myelin sheath structure are well developed,and most of the myelin sheath structure is well developed,in concentric lamellar-like structure,minority for immature myelin is not completely closed.The control group,the regenerated nerve fibers is sparse,poor myelination.A positive area of S100 and NF in Immunochemistry Stain.All the analyzed variables did not show any differences among the six groups With drug treatment(P>0.05),which were statistically superior to group A(P<0.05).(Table.3;Fig.3;Fig.4)5 In vivo electrophysiological analysisTo quantify the functional recovery of regenerated nerves,electrophysiological analysis was performed at 6 and 12 weeks post operation to determine the response in the hind limb gastrocnemius muscle to electrical stimulation at the proximal end of the grafts.Statistically significantly different(P <0.05)from the contralateral sural nerves,at each time point.In the 12 th wk,each groups exhibited significantly batter than other groups in the 6th(P <0.05).(Table.4)Conclusion:1.FK506 and lithium chloride for the regeneration of peripheral nerve injury have significant role in promoting.2.Low-dose combination of FK506 and chlorination can quickly reduce immune rejection after peripheral nerve injury,to rapidly improve the regeneration microenvironment after peripheral nerve injury,and to improve the quality and regeneration rate of nerve,made with the same effect of high dose which is separate application and it has no fatal adverse reactions,and has potential significance for clinical immunosuppressant application.