Supervisors info:
Ανδρεάδου Ιωάννα , Αναπληρώτρια καθηγήτρια Φαρμακολογίας, Τμήμα Φαρμακευτικής, ΕΚΠΑ
Summary:
Purpose: The role of glycogen synthase kinase 3 beta(GSK3β) in cardioprotection and the link between GSK3β and the mitochondrial Permeability Transition Pore (mPTP) is debated. The purposes of this study were to I. Investigate the pharmacological inhibition and phosphorylation of GSK3β in cardioprotection II. Examine the direct effect of GSK-3β inhibitors on mPTP opening III. Define localization of the kinase on normoxic and ischemic heart
Materials and methods: At first, we examined the established GSK3 inhibitor, BIO, and a series of novel selective analogues (MLS2776-MLS2779) using a rabbit model of 30’ ischemia (I) /3h reperfusion (R). The compounds were administered at the 20th min of I and infarct size (IS) was determined. GSK3β inhibitory phosphorylation (S9) was examined at the 10th min of R. Next, we elucidated the role of GSK3β using the best candidates. C57BL/6 mice underwent 30’ I /2hR and randomly received vehicle, MLS2776 or MLS2778 at the 20th min of I. IS was determined whereas myocardial tissue was obtained at the 10thminute of R to investigate GSK3β inhibition. Additionally, CsA(10mg/kg), MLS2776+CsA and MLS2778+CsA were administered for determination of IS. In order to address the GSK3β-mitochondria interaction, mice were either sham operated or subjected to 30’I/10’R and myocardial mitochondria were isolated, treated with the compounds and Calcium Retention Capacity (CRC) was estimated. GSK3β localization in the cytosolic and mitochondrial fractions was determined. Results: Rabbits treated with MLS series were protected compared to those treated with vehicle (10.6 ± 3.0%, 26.9 ± 2.9%,10.1 ± 1.5%,28.5 ± 4.0% vs 51.9 ± 2.5%, p<0.001). MLS2776 and MLS2778 reduced IS in mice (15.32±1.39% and 16.18± 1.91% vs 45.12±2.14%, p<0.05) and possessed an additional effect when co-administered with CsA indicating a different mechanism of action than CsA.(10.23±0.47%, 11.17±0.76% vs 25.03±1.03% for CsA p<0.05). GSK3β inhibition was confirmed by a decrease in p(Tyr216)-GSK3β and p(Ser33/37/Thr41)-βcatenin(p<0.05), but not by a p(S9)-GSK3β increase. CRC was not altered under normoxic and I/R conditions and GSK3β was primarily located in the cytosol. Conclusions: Pharmacological inhibition of GSK3β attenuates IS beyond mPTP inhibition. However, a direct interaction of GSK3β and mPTP cannot be established and GSK3β-mPTP axis should be further investigated.
