TXNIP as a Mediator of Palmitic Acid‐Induced Programmed Cell‐Death in Cardiomyocytes
Authors: D'Souza Karen M. , Oh Charles , Zhang Weiyang , Reaven Peter D. 
Published: June 2021

INTRODUCTION

ThioredoxinInteracting Protein (TXNIP), an endogenous inhibitor of the thioredoxinantioxidant system, is known to be elevated in diabetes, a condition characterized by abnormalities in both glucose and lipid metabolism. Underhyperglycemic conditions TXNIP is regulated in part by glucose responsive factors MondoA and ChREBP (carbohydrate response element binding protein) but littleis known about its role in mediating cardiac lipotoxicity. Plasma saturatedfatty acid (SFA) levels are elevated in type 2 diabetes and may lead tocardiomyocyte death and subsequently diabetic cardiomyopathy (DCM), a leading cause of morbidity and mortality in diabetic patients. The current aim is toelucidate the role of TXNIP in SFA‐driven programmed cell death (eitherapoptosis or pyroptosis) in vitro inAC16 cardiomyocytes and to identify potential mechanisms involved.

METHODSAND RESULTS

AC16 cardiomyocytes were treated with 150μMand 300μM palmitic acid(PA) for a period of 16 hours. TXNIP expression was determined by westernblotting of whole cell lysates in comparison to vehicle (control). Cytosolicand nuclear fractions were prepared using a suitable kit and analyzed bywestern blotting to determine activation of transcription factors MondoA andChREBP in response to PA. Apoptosis was measured by flow cytometry usingAnnexin V/PI staining while pyroptosis was determined by assessing inflammatorymarkers using RT‐PCR. To confirm the regulation of TXNIP by MondoA/ChREBP andthe involvement of TXNIP in PA‐driven cell death, TXNIP was inhibited bypre‐treating (24 hours) cells with 10μMSBI‐477, a chemical inhibitor of MondoA/ChREBP.

PA showed adose‐dependent increase in TXNIP protein expression (1.5 and 2.3 ‐fold increaseat 150μM and 300μM PA respectively; ANOVA p<0.01 between groups; n=3) in comparison to the control. PA did not affect MondoA andChREBP protein expression but activated these transcription factors as demonstrated by an approximately 2‐fold increase in their nuclear expression. PA induceddose‐dependent apoptotic cell death in AC16 cardiomyocytes (1.3 and 4.4 ‐fold increase at 150μM and 300μM PA respectively; ANOVA p<0.001 between groups;n=4) which was inhibited by pre‐treatment with inhibitor SBI‐477 (81% declinerelative to 300μM PA alone; t‐test p<0.05; n=3). 300μM PA increased IL‐6 and IL‐8 gene expression by 56 and 15 ‐fold respectively but showed nosignificant change in the pyroptosis marker IL‐18.

CONCLUSION

Ourdata reflects a central role for TXNIP in SFA‐driven apoptosis. TXNIP could represent a key link between lipotoxicity and cardiomyocyte cell death, providing an attractive therapeutictarget to prevent SFA‐mediated cell death and in doing so stall the development of DCM. Further studies will be needed to establish the key players involved in SFA‐driven cardiomyocyte cell death and determine the programmed cell death pathways that predominate.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.