Date: Tuesday, June 14, 2016
Session Time: 6:00pm-7:00pm
Presentation Time: 6:00pm-7:00pm
Location: Halls C&D
Background: Instant Blood Mediated Inflammatory Reaction (IBMIR) has been shown to severely affect islet engraftment. We previously reported that Withaferin A (WA) can protect islets from IBMIR induced damage. MicroRNA 375 has been recently identified as a reliable marker for beta cell damage.
Objective: To corroborate the finding that WA could protect islets from damage caused by IBMIR using the novel beta cell damage marker miRNA 375.
Material and Methods: Human islets were isolated and blood were drawn from cadaveric donors. Human islets were mixed with autologous blood in a heparinized tube and placed at 37C in a shaker incubator. Islets were pretreated with 1[micro]g/mL WA for 1hr prior to mixing with blood. Plasma samples were collected at various time points and stored at -80C until analysis. Total microRNA was isolated from plasma using exiqon biofluid miRNA isolation kit. MicroRNA were reverse transcribed and real time PCR was performed using exiqon universal cDNA synthesis kit and real time quantitative PCR kit respectively as per manufacturer's protocol. LNA based primers for miRNA 375, miRNA 423-3p, and Unisp6 were used. Statistical significance was determined by two-way ANOVA. (Significance are denoted as *P<0.05, **P<0.01, ***P<0.001, ****P<0.00001)
Results: In this study we observed that miRNA 375 was significantly increased in the serum immediately after mixing islets in autologous blood and the level of miR375 increased with time. WA treated islets showed significantly reduced miR375 level at 1hr, 3hr and 6hr time points compared to controls. Relative expression of miRNA 375 in serum of control vs Serum of WA group at 1hr (407.26 vs. 53.99; P=0.0042), 3hr (470.9 vs. 204.77; P=0.037) and at 6hr (452.07 vs. 196.2; P=0.0462). These data correlated with the c-peptide and pro-insulin data for the same samples .
Conclusion: This data supports that withaferin A has the potential to be used as antiinflammatory drug during islet transplantation to overcome IBMIR.
CITATION INFORMATION: Kanak M, Takita M, Yoshimatsu G, Lawrence M, Levy M, Naziruddin B. NFκB Inhibitor Withaferin A Blocks IBMIR In Vitro: miRNA 375 Analysis. Am J Transplant. 2016;16 (suppl 3).
To cite this abstract in AMA style:Kanak M, Takita M, Yoshimatsu G, Lawrence M, Levy M, Naziruddin B. NFκB Inhibitor Withaferin A Blocks IBMIR In Vitro: miRNA 375 Analysis. [abstract]. Am J Transplant. 2016; 16 (suppl 3). https://atcmeetingabstracts.com/abstract/nfb-inhibitor-withaferin-a-blocks-ibmir-in-vitro-mirna-375-analysis/. Accessed May 17, 2021.
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