Farnaz Shamsi

Anatomic location of white adipose tissue is a determinant of cardiometabolic risk. To understand differences within/between adipose depots, we generated 65,668 single-nucleus transcriptomes from human subcutaneous or intraabdominal adipose tissue (SAT/IAT). Unsupervised analysis reveals 26 adipose-resident cell clusters including two subpopulations of mature adipocytes, characterized by high vs. low expression of adipocyte maturation genes (ADIPOMAT^hi vs. ADIPOMAT^lo). ADIPOMAT^lo adipocytes demonstrate a low-differentiation, pro-inflammatory, and pro-fibrotic transcriptome. IAT-resident ADIPOMAT^lo were more abundant in higher BMI donors, while SAT-resident ADIPOMAT^lo associated with impaired glycemia. TSHZ3 is identified as a candidate regulator of ADIPOMAT^lo transcriptome. TSHZ3 knockdown in adipogenic progenitors inhibits differentiation, with downregulation of early adipogenic regulators (e.g., CEBPA/B, PPARG) and mature adipocyte genes. Heterozygous deletion of Tshz3 in mice reduces SAT and IAT weight. Here, we show that adipocyte subsets with distinct transcriptomic signature reside in human WAT; altered TSHZ3-mediated transcriptional regulation may contribute to low-maturation subpopulation linked to metabolic disease.

Brown adipose tissue is an evolutionary innovation in placental mammals that regulates body temperature through adaptive thermogenesis. Cold exposure activates brown adipose tissue thermogenesis through coordinated induction of brown adipogenesis, angiogenesis, and sympathetic innervation; however, how these processes are coordinated remains unclear. Here, we show that fragments of Slit guidance ligand 3 (SLIT3) drive crosstalk among adipocyte progenitors, endothelial cells, and sympathetic nerves. Adipocyte progenitors secrete SLIT3, which is cleaved into functionally distinct SLIT3-N and SLIT3-C fragments that independently promote angiogenesis and sympathetic innervation. We identify PLXNA1 as a receptor for SLIT3-C and demonstrate its essential role in sympathetic innervation of brown adipose tissue. Moreover, we identify BMP1 as the first SLIT protease described in vertebrates. Coordinated neurovascular expansion mediated by distinct SLIT3 fragments provides a bifurcated yet integrated mechanism that ensures a synchronized brown adipose tissue response to environmental challenges. Finally, this study reveals a previously unrecognized role for adipocyte progenitors in regulating tissue innervation.

Adipose tissue (AT) is a complex connective tissue with a high relative proportion of adipocytes, which are specialized cells with the ability to store lipids in large droplets. AT is found in multiple discrete depots throughout the body, where it serves as the primary repository for excess calories. In addition, AT has an important role in functions as diverse as insulation, immunity and regulation of metabolic homeostasis. The Human Cell Atlas Adipose Bionetwork was established to support the generation of single-cell atlases of human AT as well as the development of unified approaches and consensus for cell annotation. Here, we provide a first roadmap from this bionetwork, including our suggested cell annotations for humans and mice, with the aim of describing the state of the field and providing guidelines for the production, analysis, interpretation and presentation of AT single-cell data.

Brown adipose tissue (BAT) is an evolutionary innovation that enables placental mammals to regulate body temperature through adaptive thermogenesis. Brown adipocytes are embedded within an intricate network of blood vessels and sympathetic nerves that support their development and thermogenic function. Cold exposure activates BAT thermogenesis through the coordinated induction of brown adipogenesis, angiogenesis, and sympathetic innervation. However, how these distinct processes are coordinated remains unclear. Here, we show that fragments of Slit guidance ligand 3 (Slit3) drive crosstalk among adipocyte progenitors, endothelial cells, and sympathetic nerves. We demonstrate that adipocyte progenitors secrete Slit3, which regulates both angiogenesis and sympathetic innervation in BAT and is essential for BAT thermogenesis in vivo. Proteolytic cleavage of Slit3 generates secreted Slit3-N and Slit3-C fragments, which bind distinct receptors to stimulate angiogenesis and sympathetic innervation, respectively. We identify Plxna1 as a previously unrecognized receptor for Slit3-C and show that it is essential for sympathetic innervation and cold-induced neurite expansion in BAT. Moreover, we introduce bone morphogenetic protein 1 (Bmp1) as the first Slit protease identified in vertebrates. In summary, this work establishes a mechanistic framework for the coordinated regulation of sympathetic innervation and angiogenesis to enhance thermogenic function. The co-regulation of neurovascular expansion by distinct Slit3 fragments offers a bifurcated yet harmonized mechanism to ensure a synchronized BAT response to environmental challenges. Finally, this study provides the first evidence that adipocyte progenitors regulate tissue innervation, revealing a previously unrecognized dimension of cellular interaction within adipose tissue.

Thermogenic adipose tissue, consisting of brown and beige fat, regulates nutrient utilization and energy metabolism. Human brown fat is relatively scarce and decreases with obesity and aging. Hence, inducing thermogenic differentiation of white fat offers an attractive way to enhance whole-body metabolic capacity. Here, we show the role of endothelin 3 (EDN3) and endothelin receptor type B (EDNRB) in promoting the browning of white adipose tissue (WAT). EDNRB overexpression stimulates thermogenic differentiation of human white preadipocytes through cAMP-EPAC1-ERK activation. In mice, cold induces the expression of EDN3 and EDNRB in WAT. Deletion of EDNRB in adipose progenitor cells impairs cold-induced beige adipocyte formation in WAT, leading to excessive weight gain, glucose intolerance, and insulin resistance upon high-fat feeding. Injection of EDN3 into WAT promotes browning and improved whole-body glucose metabolism. The findings shed light on the mechanism of WAT browning and offer potential therapeutics for obesity and metabolic disorders.

Brown adipose tissue (BAT) is responsible for regulating body temperature through adaptive thermogenesis. The ability of thermogenic adipocytes to dissipate chemical energy as heat counteracts weight gain and has gained considerable attention as a strategy against obesity. BAT undergoes major remodeling in a cold environment. This remodeling results from changes in the number and function of brown adipocytes, expanding the network of blood vessels and sympathetic nerves, and changes in the composition and function of immune cells. Such synergistic adaptation requires extensive crosstalk between individual cells in the tissue to coordinate their responses. To understand the mechanisms of intercellular communication in BAT, we apply the CellChat algorithm to single-cell transcriptomic data of mouse BAT. We construct an integrative network of the ligand-receptor interactome in BAT and identify the major signaling inputs and outputs of each cell type. By comparing the ligand-receptor interactions in BAT of mice housed at different environmental temperatures, we show that cold exposure enhances the intercellular interactions among the major cell types in BAT, including adipocytes, adipocyte progenitors, lymphatic and vascular endothelial cells, myelinated and non-myelinated Schwann cells, and immune cells. These interactions are predicted to regulate the remodeling of the extracellular matrix, the inflammatory response, angiogenesis, and neurite growth. Together, our integrative analysis of intercellular communications in BAT and their dynamic regulation in response to housing temperatures provides a new understanding of the mechanisms underlying BAT thermogenesis. The resources presented in this study offer a valuable platform for future investigations of BAT development and thermogenesis.

The development of single cell approaches has facilitated the investigation of cellular heterogeneity and cell type-specific gene expression in complex tissues. Adipose tissue depots contain lipid storing adipocytes as well as a diverse array of cell types that form the adipocyte niche and regulate adipose tissue function. Here, I describe two protocols for the isolation of single cells and nuclei from white and brown adipose tissue. Additionally, I provide a detailed workflow for isolation of cell type- or lineage-specific single nuclei using nuclear tagging and translating ribosome affinity purification (NuTRAP) mouse models. Key words Adipose tissue Adipocyte progenitors Transcriptomics scRNA-seq snRNA-seq

Brown adipose tissue (BAT) is responsible for regulating body temperature through adaptive thermogenesis. Cold exposure remodels BAT by increasing the number and function of brown adipocytes, expanding the network of blood vessels and sympathetic nerves, and changing the makeup and function of immune cells. Such synergistic adaptation requires extensive intercellular crosstalk in adipose niche to coordinate their responses. To understand the mechanisms of intercellular communication, we applied the CellChat algorithm to single-cell transcriptomic data of mouse BAT. We constructed an integrative network of ligand-receptor interactome in BAT and identified the major signaling input and output of each cell type. By comparing the ligand-receptor interactions in BAT of mice housed at different temperatures, we found that cold exposure enhances the interactions among the major cell types in BAT, including adipocytes, adipocyte progenitors, lymphatic and vascular endothelial cells, myelinated and non-myelinated Schwann cells, and immune cells. These interactions regulate the remodeling of the extracellular matrix, inflammatory response, angiogenesis, and neurite growth. Together, our integrative analysis of intercellular communications in BAT and their dynamic regulation establishes a holistic understanding of the mechanisms involved in BAT thermogenesis. The resources presented in this study provide a valuable platform for investigations of BAT development and thermogenesis.

Brown adipose tissue (BAT) is responsible for regulating body temperature through adaptive thermogenesis. The ability of thermogenic adipocytes to dissipate chemical energy as heat counteracts weight gain and has gained considerable attention as a strategy against obesity. BAT undergoes major remodeling in a cold environment. This remodeling results from changes in the number and function of brown adipocytes, expanding the network of blood vessels and sympathetic nerves, and changes in the makeup and function of immune cells. All these processes are essential for enhanced BAT thermogenesis to maintain euthermia in the cold. Such synergistic adaptation requires extensive crosstalk between the individual cells in tissues to coordinate their responses. To understand the mechanisms of intercellular communication in BAT, we applied the CellChat algorithm to single-cell transcriptomic data of mouse BAT. We constructed an integrative network of ligand-receptor interactome in BAT and identified the major signaling input and output of each cell type. By comparing the ligand-receptor interactions in BAT of mice housed at different environmental temperatures, we found that cold exposure enhances the intercellular interactions among the major cell types in BAT, including adipocytes, adipocyte progenitors, lymphatic and vascular endothelial cells, myelinated Schwann cells (MSC), nonmyelinated Schwann cells (NMSC), and immune cells. Furthermore, we identified the ligands and receptors that are regulated at the transcriptional level by temperature. These interactions are predicted to regulate the remodeling of extracellular matrix (ECM), inflammatory response, angiogenesis, and neurite growth. Together, our integrative analysis of intercellular communications in BAT and their dynamic regulation in response to housing temperatures establishes a holistic understanding of the mechanisms involved in BAT thermogenesis. The resources presented in this study provide a valuable platform for future investigations of BAT development and thermogenesis.

Adipogenesis is key to maintaining organism-wide energy balance and healthy metabolic phenotype, making it critical to thoroughly comprehend its molecular regulation in humans. By single-nuclei RNA-sequencing (snRNA-seq) of over 20,000 differentiating white and brown preadipocytes, we constructed a high-resolution temporal transcriptional landscape of human white and brown adipogenesis. White and brown preadipocytes were isolated from a single individual’s neck region, thereby eliminating inter-subject variability across two distinct lineages. These preadipocytes were also immortalized to allow for controlled, in vitro differentiation, allowing sampling of distinct cellular states across the spectrum of adipogenic progression. Pseudotemporal cellular ordering revealed the dynamics of ECM remodeling during early adipogenesis, and lipogenic/thermogenic response during late white/brown adipogenesis. Comparison with adipogenic regulation in murine models revealed several targets for potential adipogenic/thermogenic drivers in humans. Key adipogenic and lipogenic markers revealed in our analysis were applied to analyze publicly available scRNA-seq datasets; these confirmed unique cell maturation features in recently discovered murine preadipocytes, and revealed inhibition of adipogenic expansion in humans with obesity. Overall, our study presents a comprehensive molecular description of both white and brown adipogenesis in humans and provides an important resource for future studies of adipose tissue development and function in both health and metabolic disease state.