title: Blockade of the OX40/OX40L pathway and induction of PD-L1 synergistically protects mouse islet allografts from rejection.
abstract: BACKGROUND
  OX40/OX40 ligand (OX40/OX40L) and programmed death-1/programmed death ligand-1 (PD-1/PD-L1) costimulatory signals play important roles in T cell-induced immune responses. The aim of this study was to investigate the roles of OX40/OX40L and PD-1/PD-L1 costimulatory pathways in mouse islet allograft rejection.
  
  
  METHODS
  Lentiviral vectors containing OX40L siRNA sequences and an adenovirus vector containing the PD-L1 gene were constructed. The streptozotocin-induced model of diabetes was established in C57BL/6 (H-2(b)) mice. Diabetic C57BL/6 mice were randomly allocated into five groups: group 1, untreated control; group 2, Ad-EGFP treatment; group 3, Ad-PD-L1 treatment; group 4, OX40L-RNAi-LV treatment; group 5, OX40L-RNAi-LV combined with Ad-PD-L1 treatment. Lentiviral vector and the adenovirus vector were injected, singly or combined, into the caudal vein one day before islet transplantation. The islets of DBA/2 (H-2(d)) mice were transplanted into the renal subcapsular space of the diabetic recipients. Recipient blood glucose and the survival time of the allografts were monitored. Antigen-specific mixed lymphocyte reaction was also evaluated.
  
  
  RESULTS
  The recombinant lentiviral RNA interference vector OX40L-RNAi-LV reduced OX40L protein expression by 70%. The recombinant adenovirus vector Ad-PD-L1 increased PD-L1 protein expression in vivo in C57BL/6 recipient mice. Combined OX40L-RNAi-LV/Ad-PD-L1 treatment induced a synergistic protective effect in pancreatic islet allografts. Allograft survival time in the combined treatment group was (92.27±9.65) days, not only longer than that of the control ((6.51±0.27) days) and Ad-EGFP groups ((7.09±0.13) days) (P < 0.01), but also significantly longer than that of Ad-PD-L1 and OX40L-RNAi-LV single treatment groups ((40.64±3.95) days and (55.14±5.48) days respectively, P < 0.01). The blood glucose concentration of recipient mice in the combined treatment group was also stable and kept within the normal range. Flow cytometry analysis showed that combined OX40L-RNAi-LV/Ad-PD-L1 treatment significantly decreased proliferation in an antigen-specific mixed lymphocyte reaction. After donor DBA/2 lymphocyte stimulation, 89.71% of lymphocytes from recipient combination treatment C57BL/6 mice were not split and proliferated. In contrast, after stimulation with third party Lewis rat lymphocytes, only 45.84% lymphocytes of C57BL/6 mice were not split and proliferated.
  
  
  CONCLUSIONS
  This study demonstrates the successful construction of the recombinant lentivirus vector OX40L-RNAi-LV and adenovirus vector Ad-PD-L1 for the blockade of OX40/OX40L and activation of PD-1/PD-L1 costimulatory pathways simultaneously in pancreatic islet allografts in diabetic mice. Combination therapy with these two vectors resulted in inhibition of T cell activation, synergistically prolonging the survival time of pancreatic islet allografts.
authors.affiliation: Department of Hepatobiliary Surgery, Peking University People's Hospital, Beijing 100044, China.
authors.forename: Tao
authors.initials: T
authors.lastname: Li
authors.affiliation: Department of Surgical Intensive Care Unit, Peking University People's Hospital, Beijing 100044, China.
authors.forename: Rui
authors.initials: R
authors.lastname: Ma
authors.affiliation: Department of Hepatobiliary Surgery, Peking University People's Hospital, Beijing 100044, China. Email: gandanwk@vip.sina.com.
authors.forename: Jiye
authors.initials: J
authors.lastname: Zhu
authors.affiliation: Department of Hepatobiliary Surgery, Peking University People's Hospital, Beijing 100044, China.
authors.forename: Fushun
authors.initials: F
authors.lastname: Wang
authors.affiliation: Department of Hepatobiliary Surgery, Peking University People's Hospital, Beijing 100044, China.
authors.forename: Lei
authors.initials: L
authors.lastname: Huang
authors.affiliation: Department of Hepatobiliary Surgery, Peking University People's Hospital, Beijing 100044, China.
authors.forename: Xisheng
authors.initials: X
authors.lastname: Leng
mesh_terms: D000818:Animals; D060890:B7-H1 Antigen / Q000235:genetics / Q000378:metabolism*; D006084:Graft Rejection / Q000235:genetics / Q000517:prevention & control*; D016381:Islets of Langerhans Transplantation / Q000276:immunology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D053278:OX40 Ligand / Q000235:genetics / Q000378:metabolism*; D053262:Receptors, OX40 / Q000235:genetics / Q000378:metabolism*; D014184:Transplantation, Homologous
chemical_list: D060890:B7-H1 Antigen; C498919:Cd274 protein, mouse; D053278:OX40 Ligand; D053262:Receptors, OX40; C505717:Tnfrsf4 protein, mouse