Antiproinflammatory effects of iodixanol (OptiPrep)-based density gradient purification on human islet preparations.
Islet isolation and purification using a continuous density gradient may reduce the volume of tissue necessary for implantation into patients, therefore minimizing the risks associated with intraportal infusion in islet transplantation. On the other hand, the purification procedure might result in a decreased number of islets recovered due to various stresses such as exposure to cytokine/chemokine. While a Ficoll-based density gradient has been widely used in purification for clinical trials, purification with iodixanol (OptiPrep) has been recently reported in islet transplant series with successful clinical outcomes. The aim of the current study was to compare the effects of the purification method using OptiPrep-based and Ficoll-based density gradients. Human islet isolations were performed using a modified automated method. After the digestion phase, pre-purification digests were divided into two groups and purified using a semiautomated cell processor with either a continuous Ficoll- or OptiPrep-based density gradient. The quantity, purity, viability, and cellular composition of islet preparations from each group were assessed. Cytokine/chemokine and tissue factor production from islet preparations after 48-h culture were also measured. Although islet purity, post-purification IEQ, islet recovery rate, FDA/PI, and fractional β-cell viability were comparable, β-cell mass after 48-h culture significantly improved in the OptiPrep group when compared to the Ficoll group. The production of cytokine/chemokine including IL-1β, TNF-α, IFN-γ, IL-6, IL-8, MIP-1β, MCP-1, and RANTES but not tissue factor from the OptiPrep group was significantly lower during 48-h culture after isolation. Each preparation contained the similar number of ductal cells and macrophages. Endotoxin level in both gradient medium was also comparable. The purification method using OptiPrep gradient media significantly reduced cytokine/chemokine production but not tissue factor from human islet preparations and improved β-cell survival during pretransplant culture. Our results suggest that the purification method using OptiPrep gradient media may be of assistance in increasing successful islet transplantation.
Mita
A
A
Cell Transplant Center, Diabetes Research Institute, University of Miami Leonard M. Miller School of Medicine, Miami, FL, USA.
Ricordi
C
C
Messinger
S
S
Miki
A
A
Misawa
R
R
Barker
S
S
Molano
R D
RD
Haertter
R
R
Khan
A
A
Miyagawa
S
S
Pileggi
A
A
Inverardi
L
L
Alejandro
R
R
Hering
B J
BJ
Ichii
H
H
D000293:Adolescent; D000328:Adult; D000818:Animals; D000893:Anti-Inflammatory Agents / Q000494:pharmacology*; D002469:Cell Separation / Q000295:instrumentation / Q000379:methods*; D002470:Cell Survival; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient / Q000295:instrumentation / Q000379:methods*; D018925:Chemokines / Q000378:metabolism; D003287:Contrast Media; D016207:Cytokines / Q000378:metabolism; D005260:Female; D006801:Humans; D050417:Insulin-Secreting Cells / Q000166:cytology / Q000187:drug effects* / Q000276:immunology / Q000502:physiology; D007515:Islets of Langerhans / Q000166:cytology*; D016381:Islets of Langerhans Transplantation / Q000379:methods; D008297:Male; D051379:Mice; D008819:Mice, Nude; D008875:Middle Aged; D013925:Thromboplastin / Q000378:metabolism; D014283:Triiodobenzoic Acids / Q000494:pharmacology*
D000893:Anti-Inflammatory Agents; D018925:Chemokines; D003287:Contrast Media; D016207:Cytokines; D014283:Triiodobenzoic Acids; D013925:Thromboplastin; C044834:iodixanol