Promotion Of Neurological Recovery In Rat Spinal Cord Injury By CD80 Monoclonal Antibody Loaded On PLLA Scaffold Through Alleviating Inflammation

Objective: Using a new type of microsol electrospun technology with immune modulators and biological materials to create a local microenvironment,which can not only control the secondary inflammation of SCI,but also bridge the damage nerve,together to promote the SCI repair.Methods: The poly(L-lactide)(PLLA)electrospun fibrous scaffolds were prepared by electrospinning.Agonist mouse anti-human CD80 antibody(CD80mAb)is grafted to the the PLLA electrospun fibrous scaffolds through microsol technology.(1)The physical and chemical properties of PLLA,PLLA-IgG and PLLA –CD80mAb were explored by transmission electron microscope(TEM)and scanning electron microscopy(SEM),respectively.The tensile stress of the electrospun fibrous scaffolds were tested by mechanical tensile test.(2)The released profile of CD80mAb on PLLA scaffolds.(3)Cytotoxicity of BMSCs is evaluated with CCK-8 kit.(4)T9 right spinal cord hemisection injury model in rats were established: 34 health six week old male SD rats were randomly divided into four groups: 1.Sham operation group(n = 7);2.Control group(n = 10);3.PLLA-IgG transplantation group(n =7);4.PLLA-CD80mAb transplantation group(n = 10).The rat mortality was recorded within 8 weeks after surgery;The Basso-Beattie-Bresnahan(BBB)score was performed once a week after SCI for 8 weeks.(5)At 8th week post injury we did histological evaluation.For immonohistochemistry,we incubated the spinal cord sections with primary antibody: anti-GFAP、anti-i NOS、anti-IL-1β 、anti-NF-200、anti-IL-6.Results:(1)TEM showed successful core-sheath structure of the scafffold,the diameters of PLLA,PLLA-IgG,PLLA –CD80mAb fiber were similar.Mechanical tensile test showed that the mechanical properties of fiber scaffolds were not changed before and after grafting.(2)The fluorescent signal is observed in PLLA electrospun fibrous scaffolds conjugated with FITC-labeled antibodies,indicating the successful grafting of IgG or CD80mAb onto the scaffolds.The release curve indicated that PLLA –CD80mAb could release CD80mAb with 100 h.(3)CCK-8 proved that CD80mAb released from PLLA –CD80mAb had no cytotoxicity.(4)Totally 4 rats were eliminated because of incomplete right hemisection.(2 from control group,1 from PLLA-IgG group,1from PLLA –CD80mAb group),and 30 rats finally were included.The mortality rate of PLLA –CD80mAb group was lower than that of PLLA-IgG group and Control group,the recovery of right hind limb function was faster than other groups.At 8 week,the BBB score of PLLA –CD80mAb group was significantly higher than that of PLLA group and control group.(5)Immonohistochemistry results showed that numbers of NF-200 staining positive cells in PLLA –CD80mAb group were more than those in control group and PLLA group,while GFAP,IL-1β,IL-6.INOS expression increased in group at 7 weeks.Conclusion: The electrospun fiber loaded CD80mAb was realized by the microsol technology,ensuring sustained release of the fiber scaffold and the biological activity of the antibody and the properties of the fibrous scaffold itself were retained.The local inflammatory environment after spinal cord injury was regulated by inhibiting the M1 macrophages,together with physical bridge role of the scaffold promotes the recovery of spinal cord injury.