Details

Autor(en) / Beteiligte

• Kříž, J

Titel

USE OF LABORATORY MICE IN DIABETOLOGY

Ist Teil von

• Physiological research, 2023-09, Vol.72 (5), p.P11-P11

Ort / Verlag

Praha: Institute of Physiology

Erscheinungsjahr

2023

Quelle

EZB Electronic Journals Library

Beschreibungen/Notizen

• The intensity of IBMIR (Instant Blood-Mediated Inflammatory Reaction) depends on the amount of tissue factor (TF) molecules on the islet cells. At the same time, TF is an important growth factor stimulating islet graft revascularization. To reduce the intensity of IBMIR and to improve the early function of islet graft, we tested the possibility of the short-term inhibition of TF synthesis in islet cells using RNA interference. Male Brown Norway rats (250-270 g) served both as pancreatic islet (PI) donors and recipients. PI were isolated using collagenase digestion according to standard protocol. After overnight cultivation Pls were transfected with anti-TF siRNA (s130189, ThermoFisher Scientific, USA) using lipofection (Lipofectamine RNAiMAX, 50 nM; n=6) or microporation (Neon, 2 pulses, 950 V, 20 ms, 200 nM; n=6). After 24 h, treated PIs were transplanted to portal vein of streptozotocin diabetic animals in marginal dose (2 PI/g)- Both methods led to a comparable decrease in TF mRNA expression - microporation of siRNA reduced the amount of TF mRNA by 76/55 %, lipofection by 75/70% after 24/48 h, respectively. Both methods led to a significant decrease in TF protein expression as proven by western blot and a significant reduction of liver ischemia after PI transplantation as proven by MRI. PIs normalized glycemia of all recipients transplanted with lipofected PIs but none with microporated PIs. In conclusion, antiTF-siRNA transfected by microporation efficiently reduced the amount of TF for 24 and 48 h but did not improve the function of transplanted PIs. AntiTF-siRNA transfected by lipofection efficiently reduced amount of TF for 24 h and significantly improved the function of transplanted PIs. Microporation likely caused the Off-target effect in PI graft.