TRPV1 Activation Counters Vascular Dysfunction by Increasing PPARs, SiRT-1, PGC1α, and UCP-1 Expression in the Thoracic Aorta Kaylan Schilling2, Laurel Markert1, Joy Watkins2, Kelvin Kinyatta2, with SteveN McAllister2, Padmamalini Baskaran1, and Baskaran Thyagarajan1. Molecular signaling laboratory, School of Pharmacy, University of Wyoming1 Division of Health Science and Public Safety, Central Wyoming College2 Good morning everyone. Our presentation reviews research that TRPV1 activation counters vascular dysfunction by increasing PPARS, SiRT-1, PGC1α, and UCP-1 Expression in the Thoracic Aorta. My name is Kaylan Schilling. I am a junior in the molecular biology program. This research is supported by the SPREM grant and through the Wyoming INBRE undergrad fellowship.

Background Obesity occurs due to imbalance between energy intake and expenditure Leads to increased visceral fat accumulation Foreshadows metabolic diseases and causes complications such as: Impaired glucose metabolism Non alcoholic fatty liver disease Cardiovascular diseases Vascular dysfunction associated obesity causes: Hypertension Heart disease Myocardial Infarction Obesity is an increasing problem in our country, where approximately 30% of the nation is considered obese.

Hypothesis Activation of TRPV1 enhances expression in the thoracic aorta vasculature suppresses the development of hypertension and vascular damage Enhances the expression of metabolically important: Peroxisome proliferator activated receptor alpha (PPAR ) Sirtuin-1 (SiRT-1; central cellular metabolic sensor) PPAR coactivator 1 (PGC-1) Uncoupling protein 1 (UCP-1). Pppar gamma deacytalization insilun sensitization

TRPV1 Pathway Recent research suggests that activating transient receptor potential vanilloid subfamily 1 (TRPV1) pathway with capsaicin is a good strategy to counter obesity and metabolic complications. Capsaicin sensitizes transient receptor potential vanilloid subfamily 1 (TRPV1), allows for increased ca2+ influx into the cell. Phospholipase C (PLC) hydrolysis of PIP2 (Phosphatidylinositol 4,5-bisphosphate)- secondary messenger, triggers cascade Cleaves into diacylglycerol (DAG)- Ca2+ binds, activates PKC stimulate ca influx ca. Inositol trisphosphate (IP3) attaches to IP3 receptor in ER and triggers release of Ca2+ stores. calcium/calmodulin-dependent kinase II (CaMKII) can phosphorlate SiRT1 and TRPV1 Sirtuin-1 (SiRT-1) NAD+ dependant deacatylates Peroxisome proliferator activated receptor(PPARα) and PPARγ coactivator 1α (PGC1α ) which both are coactivators and are transcription factors for Uncoupling Protien 1 (UCP1)

Methods Wild type and TRPV 1 -/- mice were fed diets consisting of: Normal chow diet High fat diet (60% calories from fat) High fat diet with capsaicin (0.01% w/w; 1.33 mg/kg) Diets began at 6 weeks of age and continued for 32 weeks At 38 weeks of age: Non-invasive tail cuff blood pressure Echocardiogram Thoracic aorta harvested Western blot analysis Echocardiogram observing cardiac function

TRPV1 Expressed in Mammalian Thoracic Aorta IgG from pre immune serum rabbit shows non specific band UNT –untransfected cells used as negative control, TRPV1 Overexpression positive control

Blood pressure of Mice receiving HFD ± Capsaicin Capsaicin can increase heart rate because it increases metabolism

Echocardiograph data of C57BL6 mice fed with HFD ± 0.01% CAP Non significance

Echocardiograph data of C57BL6 mice fed with HFD ± 0.01% CAP

Western Blot Analysis of Thoracic Aorta

Western Blot Analysis of Thoracic Aorta

Western Blot Analysis of Thoracic Aorta PKCα – Known to increase hypertension by vasoconstriction. PKCε – target for treating hypertension.

Summary Our diet induced obese mice were hypertensive and also had poor left ventricular function Capsaicin increased PPARα, PGC1α, SIRT1 and UCP1 – thereby preventing lipid accumulation and increasing insulin sensitivity and energy homeostasis. Capsaicin decreased PKCα- a vasoconstrictor, but increased PKCε which is known to inhibit hypertension. Thus, capsaicin improved vascular functions by preserving TRPV1 expression and activity in thoracic aorta.

Acknowledgments This project was supported in part by grants from the National Center for Research Resources (P20RR016474) and the National Institute of General Medical Sciences (P20GM103432) from the National Institutes of Health. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. We thank the University of Wyoming INBRE Network for their support in this research.