Transplantation of rat islets transduced with human heme oxygenase-1 gene using adenovirus vector.
OBJECTIVE
To investigate whether human heme oxygenase-1 (HO-1) gene has protective action on islets cultured in vitro, and to explore whether transduction of HO-1 gene to donor islets could enhance engrafted islets survival and suppress local lymphocytic infiltration in islet grafts.


METHODS
Newly isolated rat islets were isolated from the Sprague-Dawley rats and were divided into 3 groups in vitro study as follows: enhanced green fluorescent protein (EGFP) group, islets transduced with adenovirus vectors containing EGFP gene using multiplicities of infection (MOI) = 2, 5, 10, and 20 to determine the transduction efficacy; HO-1 group, islets transduced with adenovirus vectors containing human HO-1 gene using MOI = 20; and control group, mock transduced islets. Flow cytometry was used to detect apoptotic cells after induction by recombinant human tumor necrosis factor-alpha (rTNF-alpha) and cycloheximide (CHX) for 48 hours. Diabetic recipients were randomly divided into the following groups: HO-1 group (n = 9), receiving islets transduced with recombinant adenovirus-HO-1; control group (n = 9), receiving mock transduced islets; and phosphate buffer solution (PBS) group (n = 6), receiving only 0.8 mL PBS. About 1200 rat islet equivalents were transplanted into each recipient rendered by streptozotocin using the portal vein as transplant site. Allograft survival, apoptosis, and the state of lymphocytic infiltration were analyzed.


RESULTS
After treatment with rTNF-alpha and CHX, the apoptotic ratio of islet cells was 4.22% +/- 2.38% in the HO-1 group (MOI = 20), significantly lower than 23.81% +/- 8.51% in the control group (P < 0.05), and 28.76% +/- 14.76% in the EGFP group (MOI = 20; P < 0.05). Maintenance of normoglycemia was prolonged in the HO-1 group, indicated by results that islet survival time was 10.56 +/- 4.33 days significantly longer than that of untreated islets which was 5.33 +/- 4.18 days (P < 0.05). The lymphocytic infiltration degree in engrafted islets treated with HO-1 gene was lower than that in the control group.


CONCLUSIONS
HO-1 gene overexpression in rat islets by adenovirus transduction can protect cultured islets against rTNF-alpha and CHX-mediated cytotoxicity. HO-1 gene has cytoprotective effects on engrafted islets, which could prolong engrafted islets survival in allogenic transplantation model, and diminish the degree of lymphocytic infiltration in islet grafts. These findings suggest a potential therapeutic application for HO-1 gene in improving islet survival/function in human islet transplantation.
Li
Yongxiang
Y
Department of Urology and Center of Renal Transplantation, Shanghai Organ Transplant and Research Center, Shanghai First People's Hospital, China. yongxiangli@yahoo.com.cn
Li
Ge
G
Dong
Weiping
W
Chen
Jing
J
Lu
Daru
D
Tan
Jianming
J
D000256:Adenoviridae; D000465:Algorithms; D000818:Animals; D017209:Apoptosis; D002478:Cells, Cultured; D003520:Cyclophosphamide / Q000633:toxicity; D003921:Diabetes Mellitus, Experimental / Q000601:surgery*; D004195:Disease Models, Animal; D005434:Flow Cytometry; D005822:Genetic Vectors; D051547:Heme Oxygenase-1 / Q000235:genetics* / Q000378:metabolism; D007515:Islets of Langerhans / Q000166:cytology; D016381:Islets of Langerhans Transplantation / Q000473:pathology / Q000502:physiology*; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley; D014161:Transduction, Genetic; D014409:Tumor Necrosis Factor-alpha / Q000633:toxicity
D014409:Tumor Necrosis Factor-alpha; D003520:Cyclophosphamide; D051547:Heme Oxygenase-1