Author: Jessica Ortiz

Physiologically Competent white adipose tissue on a chip

Obesity is a global pandemic, particularly severe in the US, where over 70% of adults are overweight including 40% with obesity [CDC: https://www.cdc.gov/nchs/fastats/obesity-overweight.htm]. White adipose tissue (WAT) is a critical organ in obesity and a primary target for treatment discovery, yet a reliable human in vitro model has been lacking due to difficulties in maintaining viability, maturity, and functionality. To address these challenges, we reconstructed a WAT within microphysiological system (WAT-MPS, or fat-chip) using human preadipocytes, mesenchymal stem cells, and induced pluripotent stem cells (hiPSCs). Our WAT-MPS is physiologically comparable to primary adipocytes from human biopsies and maintains over 90% viability for up to six months. With its robust responses to hormones in lipid and glucose metabolism, this fat-chip is an excellent platform for studying obesity and related diseases.

Multi-organ systems: fat-liver

Interorgan crosstalk with WAT is a critical mechanism in triggering systemic metabolic disorders in obesity, but traditional in vitro cultures struggle to model this complexity. The organ-chip platform is particularly suited to mimicking interorgan crosstalk by circulating culture medium between connected chips. Therefore, it is plausible to connect our health and/or obese fat-chips to other organs to study obesity-induced diseases. The liver, being the most relevant metabolic organ due to its direct exposure to factors released from visceral fat through the portal vein, is a key focus. Here, we developed an interconnected fat-liver-chip consisting of isogenic hiPSC-derived white adipocytes, hepatocytes, and macrophages. We further utilized our MPS to identify and characterize pharmacological intervention strategies for hepatic steatosis and systemic insulin resistance. These efforts establish our human isogenic fat-liver MPS as an effective alternative to animal models for studying the etiology and therapy of highly prevalent metabolic diseases. Furthermore, we are working to interconnect more organs in-line, such as fat-liver-islet to form a minimal set of obesity-induced type-2 diabetes milieu, or fat-heart to study obesity-associated cardiac diseases. Our ultimate goal is to emulate a human-on-chip model for studying obesity-associated diseases across multiple organs and tissues.

Engineering and Interrogation of Functional Sexually Dimorphic Liver-Fat in a Microphysiological System

Although both men and women are both susceptible to obesity and these associated diseases (T2D and Fatty Liver), the risk associated varies per sex. Sex differences in the endocrine and immune systems contribute to these observations, recent studies suggest that sex-specific genetic architecture also influences human phenotypes, including reproductive, physiological and disease traits. It is likely that an underlying mechanism is differential gene regulation in males and females, particularly in sex steroid-responsive genes. We are working to establish an isogenic microphysiological model that can recapitulate these effects and demonstrate sex-specific differences.

Obesity related diseases affect billions of people worldwide. 42.5% of adults in the USA are obese and obesity related diseases such as diabetes, cardiovascular disease and certain cancers pose a costly challenge to the American and World healthcare systems. Metabolic disorder treatments that are safe and effective have the potential to transform healthcare and are urgently needed. In recent years targeting brown and beige fat has
garnered much excitement as a potential obesity and diabetes therapeutic. This is because brown and beige fat selectively express the mitochondrial protein UCP1, which induces non-shivering thermogenesis. Treating obesity by converting white fat (WAT) into beige fat (BEAT) has the potential to generate a safe and effective treatment for obesity that is widely adopted by patients.

Matrix Assisted Transplantation of Functional Beige Adipose Tissue

To this end, a tissue engineering approach was first utilized in our lab via the Beige Adipose Tissue Matrix Assisted Cellular Implant (BAT-MACT), wherein subcutaneous implantation of adipose-derived mesenchymal stem cells (ADMSCs) within optimized hydrogels resulted in the establishment of distinct UCP1-expressing implants that successfully attracted host vasculature and persisted for several weeks. Importantly, implant recipients demonstrated elevated core body temperature during cold challenges, enhanced respiration rates, improved glucose homeostasis, and reduced weight gain, demonstrating the therapeutic merit of this highly translatable approach.

Browning-Lipid Nanoparticles (B-LNPs)

In collaboration with the Murthy lab, this project aims to build on the previous BAT- MACT strategy by utilizing a technology that is already FDA approved and more readily applicable, lipid nanoparticle (LNP) based therapeutics. We aim to utilize LNPs carrying mRNA or siRNA cargo that can induce the trans-differentiation of white to beige fat in subcutaneous adipose tissue depots. We have termed this technology as “browning”- LNPs (B-LNPs). Preliminary experiments already show great promise including UCP1 expression upregulation in subcutaneous WAT depots and weight loss in an obese mouse model.

CoQ Transport and Trafficking

Coenzyme Q (CoQ aka ubiquinone) is an essential component of the mitochondrial electron transport chain (ETC) as well as a membrane-incorporated antioxidant. Although it is ubiquitous to all cell types and organelles, little is known about critical mechanisms involving CoQ uptake into cells, trafficking and it’s role outside of the mitochondria. Studies have shown that tissues rich in mitochondria, also have increased CoQ levels and are more sensitive to CoQ deficiencies. Because of this, we are primarily interested in studying CoQ within the context of classical brown adipose tissue (BAT). BAT is a unique type of adipose tissue that is composed of adipocytes with multilocular lipid droplets and a large amount of mitochondria, making it a highly metabolically active organ that is responsible for non-shivering thermogenesis via the expression of uncoupling protein 1 (UCP1). Due to these characteristics, in recent years it has been targeted for both diabetes and obesity therapeutics. We have identified BAT as a major destination for exogenous CoQ, and we identified the specific scavenger receptor CD36 as the main receptor governing CoQ uptake into brown adipocytes. To address the remaining unanswered questions regarding CoQ uptake and trafficking, we have developed novel CoQ-azide conjugates and demonstrate in our preliminary data that these novel compounds can be used to track both the intracellular as well as whole body uptake dynamics of CoQ using fluorescent microscopy and bioluminescent imaging respectively. Our main objective is to understand how CoQ is transported within cells and organisms. Answering important biological questions, ranging from identification of key transporters to time-dependent trafficking will help us identify the factors and mechanisms involved in the uptake of CoQ.

CoQ Deficiency and Thermogenesis

More recent studies in our lab have focused on studying the BAT phenotype under CoQ deficiency conditions. Primary CoQ deficiencies can be caused by hereditary mutations in the biosynthesis pathway while secondary CoQ deficiencies are associated with the pharmacological use of HMG-CoA Reductase inhibitors, statins. Utilizing both cell culture and mouse models of CoQ deficiency we found that CoQ deficiencies in BAT resulted UCP1 downregulation leading to cold intolerance in vivo. Further, CoQ defiency resulted in an upregulation of mitochondrial stress signaling pathways including the integrated stress response (ISR) and mitochondrial unfolded protein response (UPR). The activation of these stress pathways led to an unexpected mitohormetic phenotype in the face of BAT dysfunction whereby BAT to inguinal white adipose tissue (iWAT) interorgan crosstalk via the secreted factor FGF21 led to enhanced energy expenditure and resistance to diet induced obesity in mice. Future studies in the lab aim to understand if the same mechanisms occur during secondary CoQ deficiency via statin treatment. Further, we seek to understand if there are other novel batokines responsible for the mitohormetic phenotype observed in CoQ deficient mice via proximity labeling analysis of secreted proteins.

Investigating the role of intracellular tension in regulating adipocyte thermogenesis

Brown (BAT) and Beige adipose tissue (BeAT) are intriguing weight loss targets due to their high energy expenditure as thermogenic tissues. While most advancements in understanding the molecular mechanism of adipocyte thermogenesis regulation have primarily focused on classical catecholamine downstream signaling cascade, our research has highlighted the critical role of Myh7-mediated intracellular tension in brown adipocyte thermogenesis. We are now particularly interested in exploring the potential for contractile force-mediated thermogenesis regulation in BeAT and the underlying mechanism.

Integrated regulation of adipocyte thermogenesis by intracellular and extracellular tension

Extracellular matrix (ECM) serves as a critical source of mechanical inputs that control cell function through activation of mechanotransducive signaling systems. In collaboration with the Kumar lab at UC Berkeley, we aim to unravel the relation of ECM viscoelastic properties and BeAT thermogenesis capacity using a new multiwell hyaluronic acid (HA) platform that enables modulation of ECM viscoelastic properties in 3D.