Rapamycin induces of protective autophagy in vascular endothelial cells exposed to oxygen-glucose deprivation.
The protective potential of rapamycin has been reported in a few experimental models of brain ischemia, both in vivo and in vitro. Although the precise cellular processes underlying the neuroprotective effects of rapamycin in experimental models of stroke remain unknown, the current experimental data suggest that the mechanism of action of the drug may result from the mTOR-mediated autophagy induction. However, it is unclear whether the activation of autophagy acts as a pro-death or pro-survival factor in vascular endothelial cells in ischemic brain damage. It seems to be very important, since stroke affects not only neurons and astrocytes but also microvessels. In the present study, we used human umbilical vein endothelial cells (HUVEC) subjected to ischemia-simulating conditions (combined oxygen and glucose deprivation, OGD) for 6h to determine potential effect of rapamycin-induced autophagy on HUVEC damage. The drug at concentrations of 100 and 1000nM increased the expression of Beclin 1 and LC3-II together with a significant increase in the p62 degradation in ischemic HUVEC. Treatment with rapamycin in OGD significantly increased the cell viability, indicating that the drug exerts cytoprotective effect. The inhibition of Beclin 1 by siRNAs significantly attenuated the expression of autophagy-related proteins and reduced HUVEC viability following OGD and rapamycin treatment. Our findings demonstrated that toxicity of simulated ischemia conditions were enhanced in HUVEC when autophagy was blocked, and that rapamycin effectively prevented OGD-evoked damage by induction of protective autophagy via inhibition of the mTOR pathway.
Department of General and Vascular Surgery, Medical University of Silesia, Ziołowa 45/47, PL 40-635 Katowice, Poland.
Tomasz
T
Urbanek
Department of General and Vascular Surgery, Medical University of Silesia, Ziołowa 45/47, PL 40-635 Katowice, Poland.
Wacław
W
Kuczmik
Department of Experimental Neuroendocrinology, Institute of Pharmacology, Polish Academy of Sciences, Smętna 12, PL 31-343 Kraków, Poland.
Agnieszka
A
Basta-Kaim
Department of Pharmacology, Medical University of Silesia, Medyków 18, PL 40-752 Katowice, Poland. Electronic address: bgabryel@interia.pl.
Bożena
B
Gabryel
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D001343:Autophagy / Q000187:drug effects* / Q000502:physiology; D000071186:Beclin-1; D015687:Cell Hypoxia; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D005786:Gene Expression Regulation / Q000187:drug effects; D005947:Glucose / Q000172:deficiency*; D061307:Human Umbilical Vein Endothelial Cells / Q000187:drug effects* / Q000502:physiology; D006801:Humans; D007511:Ischemia / Q000188:drug therapy / Q000503:physiopathology; D008565:Membrane Proteins / Q000378:metabolism; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D018696:Neuroprotective Agents / Q000494:pharmacology*; D010100:Oxygen / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D000071456:Sequestosome-1 Protein; D015398:Signal Transduction / Q000187:drug effects; D020123:Sirolimus / Q000494:pharmacology*; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; C491997:BECN1 protein, human; D000071186:Beclin-1; C477012:MAP1LC3A protein, human; D008565:Membrane Proteins; D008869:Microtubule-Associated Proteins; D018696:Neuroprotective Agents; D034741:RNA, Small Interfering; C099718:SQSTM1 protein, human; D000071456:Sequestosome-1 Protein; C546842:MTOR protein, human; D058570:TOR Serine-Threonine Kinases; D005947:Glucose; D010100:Oxygen; D020123:Sirolimus
Autophagy; Endothelial cells; Ischemia; Rapamycin; mTOR