Single-cell Analysis of Subcutaneous Fat Reveals Pro-fibrotic Cells that Correlate with Visceral Adiposity in HIV.

CONTEXT: Cardiometabolic diseases are common in persons with HIV (PWH) on antiretroviral therapy (ART), which has been attributed to preferential lipid storage in visceral adipose tissue (VAT) compared with subcutaneous adipose tissue (SAT). However, the relationship of SAT-specific cellular and molecular programs with VAT volume is poorly understood in PWH. OBJECTIVE: We characterized SAT cell-type specific composition and transcriptional programs that are associated with greater VAT volume in PWH on contemporary ART. METHODS: We enrolled PWH on long-term ART with a spectrum of metabolic health. Ninety-two participants underwent SAT biopsy for bulk RNA sequencing and 43 had single-cell RNA sequencing. Computed tomography quantified VAT volume and insulin resistance was calculated using HOMA2-IR. RESULTS: VAT volume was associated with HOMA2-IR (p < 0.001). Higher proportions of SAT intermediate macrophages (IMs), myofibroblasts, and MYOC + fibroblasts were associated with greater VAT volume using partial Spearman's correlation adjusting for age, sex, and body mass index (ρ=0.34-0.49, p < 0.05 for all). Whole SAT transcriptomics showed PWH with greater VAT volume have increased expression of extracellular matrix (ECM)- and inflammation-associated genes, and reduced expression of lipolysis- and fatty acid metabolism-associated genes. CONCLUSIONS: In PWH, greater VAT volume is associated with higher proportion of SAT IMs and fibroblasts, and a SAT ECM and inflammatory transcriptome, which is similar to findings in HIV-negative persons with obesity. These data identify SAT cell-type specific changes associated with VAT volume in PWH that could underlie the high rates of cardiometabolic diseases in PWH, though additional longitudinal studies are needed to define directionality and mechanisms.

A prognostic molecular signature of hepatic steatosis is spatially heterogeneous and dynamic in human liver.

Metabolic dysfunction-associated steatotic liver disease (MASLD) prevalence is increasing in parallel with an obesity pandemic, calling for novel strategies for prevention and treatment. We defined a circulating proteome of human MASLD across ≈7000 proteins in ≈5000 individuals from diverse, at-risk populations across the metabolic health spectrum, demonstrating reproducible diagnostic performance and specifying both known and novel metabolic pathways relevant to MASLD (central carbon and amino acid metabolism, hepatocyte regeneration, inflammation, fibrosis, insulin sensitivity). A parsimonious proteomic signature of MASLD was associated with a protection from MASLD and its related multi-system metabolic consequences in >26000 free-living individuals, with an additive effect to polygenic risk. The MASLD proteome was encoded by genes that demonstrated transcriptional enrichment in liver, with spatial transcriptional activity in areas of steatosis in human liver biopsy and dynamicity for select targets in human liver across stages of steatosis. We replicated several top relations from proteomics and spatial tissue transcriptomics in a humanized "liver-on-a-chip" model of MASLD, highlighting the power of a full translational approach to discovery in MASLD. Collectively, these results underscore utility of blood-based proteomics as a dynamic "liquid biopsy" of human liver relevant to clinical biomarker and mechanistic applications.

Relationship of Subcutaneous Adipose Tissue Inflammation-Related Gene Expression With Ectopic Lipid Deposition in Persons With HIV.

OBJECTIVE: Fat redistribution from subcutaneous adipose tissue (SAT) to the abdominal viscera, pericardium, liver, and skeletal muscle contributes to the rising burden of cardiometabolic disease among persons with HIV (PWH). Previous studies found SAT inflammation in PWH impairs lipid storage and persists despite plasma viral suppression on antiretroviral therapy (ART). In this study, we identified SAT immune-related genes associated with ectopic fat deposition in PWH on long-term ART. DESIGN AND METHODS: A total of 92 PWH with well-controlled viremia underwent computed tomography imaging and abdominal SAT biopsy for gene expression analysis. SAT gene expression was measured using a NanoString panel of 255 immune-related genes. Associations between gene expression and computed tomography measurements of the volume and attenuation (radiodensity) of metabolically relevant ectopic fat depots were assessed using multivariable linear regression and network analysis. RESULTS: Greater SAT volume was associated with higher visceral and pericardial adipose tissue volume, but lower skeletal muscle attenuation. Lower SAT attenuation, a measure of lipid content, was associated with lower visceral adipose tissue attenuation. Hierarchical clustering identified a subset of macrophage-related genes in SAT, including CCL2, CCL22, CCL13, CCR1, CD86, CD163, IL-6, IL-10, MRC1, and TREM2, which were associated with an increased lipid deposition in multiple ectopic depots. CONCLUSION: Altered expression of macrophage-related genes in SAT is associated with differences in ectopic fat depot morphometrics among PWH on long-term ART, including in the pericardial and visceral compartments. These findings provide basis for future studies to assess host, virus, and treatment factors shaping the SAT immune environment and its effects on morphometric changes and metabolic comorbidities in PWH.

The soluble epoxide hydrolase inhibitor GSK2256294 decreases the proportion of adipose pro-inflammatory T cells.

Adipose tissue contains a complex immune environment and is a central contributor to heightened systemic inflammation in obese persons. Epoxyeicosatrienoic acids (EETs) are lipid signaling molecules that decrease inflammation in obese animals, but their effect on inflammation in humans is unknown. The enzyme soluble epoxide hydrolase (sEH) hydrolyzes EETs to less active diols, and we hypothesized that pharmacologic sEH inhibition would decrease adipose inflammation in obese individuals. We treated obese prediabetic adults with the sEH inhibitor GSK2256294 versus placebo in a crossover design, collected subcutaneous abdominal adipose tissue via lipoaspiration and characterized the tissue T cell profile. Treatment with GSK2256294 decreased the percentage of pro-inflammatory T cells producing interferon-gamma (IFNγ), but not interleukin (IL)-17A, and decreased the amount of secreted tumor necrosis factor-alpha (TNFα). Understanding the contribution of the EET/sEH pathway to inflammation in obesity could lead to new strategies to modulate adipose and systemic inflammation.

Anticytomegalovirus CD4+ T Cells Are Associated With Subclinical Atherosclerosis in Persons With HIV.

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Nuclear transport modulation reduces hypercholesterolemia, atherosclerosis, and fatty liver.

BACKGROUND: Elevated cholesterol and triglycerides in blood lead to atherosclerosis and fatty liver, contributing to rising cardiovascular and hepatobiliary morbidity and mortality worldwide. METHODS AND RESULTS: A cell-penetrating nuclear transport modifier (NTM) reduced hyperlipidemia, atherosclerosis, and fatty liver in low-density lipoprotein receptor-deficient mice fed a Western diet. NTM treatment led to lower cholesterol and triglyceride levels in blood compared with control animals (36% and 53%, respectively; P<0.005) and liver (41% and 34%, respectively; P<0.05) after 8 weeks. Atherosclerosis was reduced by 63% (P<0.0005), and liver function improved compared with saline-treated controls. In addition, fasting blood glucose levels were reduced from 209 to 138 mg/dL (P<0.005), and body weight gain was ameliorated (P<0.005) in NTM-treated mice, although food intake remained the same as that in control animals. The NTM used in this study, cSN50.1 peptide, is known to modulate nuclear transport of stress-responsive transcription factors such as nuclear factor kappa B, the master regulator of inflammation. This NTM has now been demonstrated to also modulate nuclear transport of sterol regulatory element-binding protein (SREBP) transcription factors, the master regulators of cholesterol, triglyceride, and fatty acid synthesis. NTM-modulated translocation of SREBPs to the nucleus was associated with attenuated transactivation of their cognate genes that contribute to hyperlipidemia. CONCLUSIONS: Two-pronged control of inflammation and dyslipidemia by modulating nuclear transport of their critical regulators offers a new approach to comprehensive amelioration of hyperlipidemia, atherosclerosis, fatty liver, and their potential complications.

Activation of invariant natural killer T cells by lipid excess promotes tissue inflammation, insulin resistance, and hepatic steatosis in obese mice.

Obesity triggers a low-grade systemic inflammation, which plays an important role in the development of obesity-associated metabolic diseases. In searching for links between lipid accumulation and chronic inflammation, we examined invariant natural killer T (iNKT) cells, a subset of T lymphocytes that react with lipids and regulate inflammatory responses. We show that iNKT cells respond to dietary lipid excess and become activated before or at the time of tissue recruitment of inflammatory leukocytes, and that these cells progressively increase proinflammatory cytokine production in obese mice. Such iNKT cells skew other leukocytes toward proinflammatory cytokine production and induce an imbalanced proinflammatory cytokine environment in multiple tissues. Further, iNKT cell deficiency ameliorates tissue inflammation and provides protection against obesity-induced insulin resistance and hepatic steatosis. Conversely, chronic iNKT cell stimulation using a canonical iNKT cell agonist exacerbates tissue inflammation and obesity-associated metabolic disease. These findings place iNKT cells into the complex network linking lipid excess to inflammation in obesity and suggest new therapeutic avenues for obesity-associated metabolic disorders.

Assessing tumor growth and distribution in a model of prostate cancer metastasis using bioluminescence imaging.

Bioluminescence imaging (BLI) has greatly facilitated the development of animal models of cancer, allowing sensitive detection of luciferase-expressing cancer cells in living mice. Previous efforts characterizing such models have involved small numbers of animals, limiting understanding of their performance features. We employed BLI to serially image the growth and distribution of a prostate cancer cell line, 22Rv1, after intracardiac injection into scid mice (n = 85). This approach models hematogenous dissemination of cancer cells and allows inquiry of the process of metastatic colonization at various organ sites, although accurately injecting cancer cells into the left ventricle remains challenging. Therefore, to predict injection success we measured the ratio of the thoracic bioluminescence signal to the whole body bioluminescence signal (T/WB ratio) immediately following intracardiac injection. A T/WB ratio less than 0.50 predicted the development of tumors outside of the thoracic cavity while a T/WB greater than 0.50 predicted the development of tumors entirely within the thoracic cavity, suggestive of a failed injection. Progressive tumor growth was quantified using BLI. Tumors colonized multiple organ sites including bone, liver, and adrenal glands resembling the spectrum of metastases in autopsy studies of patients with prostate cancer. Tumors growing in bone exhibited mixed osteolytic and osteoblastic features, eliciting a spiculated periosteal response. With the ability to more accurately predict injection success, we can now monitor efficacy of intracardiac injections facilitating the performance of this model.