ACSL4 Directs Intramuscular Adipogenesis and Fatty Acid Composition in Pigs

The intramuscular fat is a major quality trait of meat, affecting sensory attributes such as flavor and texture. Several previous GWAS studies identified Acyl-CoA Synthetase Long Chain Family Member 4 (ACSL4) gene as the candidate gene to regulate intramuscular fat content in different pig populations, but the underlying molecular function of ACSL4 in adipogenesis within pig skeletal muscle is not fully investigated. In this study, we isolated porcine endogenous intramuscular adipocyte progenitors and performed ACSL4 loss- and gain-of-function experiments during adipogenic differentiation. Our data showed that ACSL4 is a positive regulator of adipogenesis in intramuscular fat cells isolated from pigs. More interestingly, the enhanced expression of ACSL4 in pig intramuscular adipocytes could increase the cellular content of monounsaturated and polyunsaturated fatty acids, such as gamma-L eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA). The above results not only confirmed the function of ACSL4 in pig intramuscular adipogenesis and meat quality attributes, but also provided new clues for the improvement of the nutritional value of pork for human health.

Bone Marrow Mesenchymal Stromal Cells: Identification, Classification, and Differentiation

Bone marrow mesenchymal stromal cells (BMSCs), identified as pericytes comprising the hematopoietic niche, are a group of heterogeneous cells composed of multipotent stem cells, including osteochondral and adipocyte progenitors. Nevertheless, the identification and classification are still controversial, which limits their application. In recent years, by lineage tracing and single-cell sequencing, several new subgroups of BMSCs and their roles in normal physiological and pathological conditions have been clarified. Key regulators and mechanisms controlling the fate of BMSCs are being revealed. Cross-talk among subgroups of bone marrow mesenchymal cells has been demonstrated. In this review, we focus on recent advances in the identification and classification of BMSCs, which provides important implications for clinical applications.

Cold-responsive adipocyte progenitors couple adrenergic signaling to immune cell activation to promote beige adipocyte accrual

The full array of cold-responsive cell types within white adipose tissue that drive thermogenic beige adipocyte biogenesis remains undefined. We demonstrate that acute cold challenge elicits striking transcriptomic changes specifically within DPP4+ PDGFRβ+ adipocyte precursor cells, including a β-adrenergic receptor CREB-mediated induction in the expression of the prothermogenic cytokine, Il33. Doxycycline-inducible deletion of Il33 in PDGFRβ+ cells at the onset of cold exposure attenuates ILC2 accumulation and beige adipocyte accrual. These studies highlight the multifaceted roles for adipocyte progenitors and the ability of select mesenchymal subpopulations to relay neuronal signals to tissue-resident immune cells in order to regulate tissue plasticity.

Constitutive loss of DNMT3A causes morbid obesity through misregulation of adipogenesis

DNA Methyltransferase 3A (DNMT3A) is an important facilitator of differentiation of both embryonic and hematopoietic stem cells. Heterozygous germline mutations in DNMT3A lead to Tatton-Brown-Rahman Syndrome (TBRS), characterized by obesity and excessive height. While DNMT3A is known to impact feeding behavior via the hypothalamus, here we investigated a role in adipocyte progenitors utilizing heterozygous knockout mice that recapitulate cardinal TBRS phenotypes. These mice become morbidly obese due to adipocyte enlargement and tissue expansion. Adipose tissue in these mice exhibited defects in preadipocyte maturation and precocious activation of inflammatory gene networks, including interleukin-6 signaling. Adipocyte progenitor cell lines lacking DNMT3A exhibited aberrant differentiation. Furthermore, mice in which Dnmt3a was specifically ablated in adipocyte progenitors showed enlarged fat depots and increased progenitor numbers, partly recapitulating the TBRS obesity phenotypes. Loss of DNMT3A led to constitutive DNA hypomethylation, such that the DNA methylation landscape of young adipocyte progenitors resemble that of older wild-type mice. Together, our results demonstrate that DNMT3A coordinates both the central and local control of energy storage required to maintain normal weight and prevent inflammatory obesity.

Abstract P186: Chemical And Mechanical Activation Of The Mechanosensor Piezo1 Alters Adipogenesis In Pvat Preadipocytes

During hypertension, vascular remodeling allows the blood vessel to withstand high blood pressure (BP). This process is well characterized in the media and intima layers of the vessel. In the perivascular adipose tissue (PVAT) there is evidence for fibrosis development during hypertension; but PVAT remodeling is poorly understood. In stem cells (i.e., adipocyte progenitors) from non-PVAT depots, mechanical forces affect adipogenesis' commitment and lipogenic stages. The mechanism involves PIEZO1, a mechanosensor that boosts the differentiation of preadipocytes towards osteogenic and fibroblastic lineages. However, PVAT's particular anatomical location continuously exposes it to forces generated by blood flow that could affect adipogenesis during normotensive and hypertensive states. Our objective was to evaluate PIEZO1's role in the adipogenic potential of preadipocytes. We hypothesize that activation of PIEZO1 reduces Adipogenesis in PVAT preadipocytes. Aortic (APVAT) was collected from male SD rats at 10 weeks of age (n=15) to harvest preadipocytes by Liberase™ digestion. Nonselective cationic channel PIEZO1 activity was evaluated with Ca 2+ indicator Fluo-4AM. Piezo1 was reduced with siRNA. Preadipocytes were differentiated for 4 d in adipogenic media containing PIEZO1 agonist Yoda1 (CON=0; YODA1=10μM). Mechanical strain (MS) was applied with FlexCell System at 12%, half-sine at 1 Hz for 4 d (MS+; MS-). Adipogenesis was evaluated by quantification of adipogenic gene network expression using PCR; lipid accumulation using lipophilic stains (Bodipy, siRNA experiments) or Oil Red O (FlexCell experiments). Adipogenesis efficiency is reported as Adipocyte/Total cells as measured in the IncuCyte Live-Cell ® system. Yoda1 reduced adipogenesis by 33% compared with CON and as expected, increased cytoplasmic Ca 2+ . In si Piezo1 cells, the anti-adipogenic effect of Yoda1 was reversed. MS+ reduced adipogenesis efficiency (0.15±0.06) compared with MS- (0.22±0.1). These data demonstrate that Piezo1 activation in PVAT may be an adaptive or pathogenic mechanism by which adipocyte populations are reduced, thus minimizing their secretion of vasoactive adipokines, and enhancing the deleterious impact of hypertension on PVAT function.

Breast Cancer Endocrine Therapy Promotes Weight Gain With Distinct Adipose Tissue Effects in Lean and Obese Female Mice

Breast cancer survivors treated with tamoxifen and aromatase inhibitors report weight gain and have an elevated risk of type 2 diabetes, especially if they have obesity. These patient experiences are inconsistent with, preclinical studies using high doses of tamoxifen which reported acute weight loss. We investigated the impact of breast cancer endocrine therapies in a preclinical model of obesity and in a small group of breast adipose tissue samples from women taking tamoxifen to understand the clinical findings. Mature female mice were housed at thermoneutrality and fed either a low-fat/low-sucrose (LFLS) or a high-fat/high-sucrose (HFHS) diet. Consistent with the high expression of Esr1 observed in mesenchymal stem cells from adipose tissue, endocrine therapy was associated with adipose accumulation and more preadipocytes compared with estrogen-treated control mice but resulted in fewer adipocyte progenitors only in the context of HFHS. Analysis of subcutaneous adipose stromal cells revealed diet- and treatment-dependent effects of endocrine therapies on various cell types and genes, illustrating the complexity of adipose tissue estrogen receptor signaling. Breast cancer therapies supported adipocyte hypertrophy and associated with hepatic steatosis, hyperinsulinemia, and glucose intolerance, particularly in obese females. Current tamoxifen use associated with larger breast adipocyte diameter only in women with obesity. Our translational studies suggest that endocrine therapies may disrupt adipocyte progenitors and support adipocyte hypertrophy, potentially leading to ectopic lipid deposition that may be linked to a greater type 2 diabetes risk. Monitoring glucose tolerance and potential interventions that target insulin action should be considered for some women receiving life-saving endocrine therapies for breast cancer.

Adipocytes and Stromal Cells Regulate Brown Adipogenesis Through Secretory Factors During the Postnatal White-to-Brown Conversion of Adipose Tissue in Syrian Hamsters

Brown adipose tissue (BAT) is a specialized tissue that regulates non-shivering thermogenesis. In Syrian hamsters, interscapular adipose tissue is composed primarily of white adipocytes at birth, which is converted to BAT through the proliferation and differentiation of brown adipocyte progenitors and the simultaneous disappearance of white adipocytes. In this study, we investigated the regulatory mechanism of brown adipogenesis during postnatal BAT formation in hamsters. Interscapular adipose tissue of a 10-day-old hamster, which primarily consists of brown adipocyte progenitors and white adipocytes, was digested with collagenase and fractioned into stromal–vascular (SV) cells and white adipocytes. SV cells spontaneously differentiated into brown adipocytes that contained multilocular lipid droplets and expressed uncoupling protein 1 (Ucp1), a marker of brown adipocytes, without treatment of adipogenic cocktail such as dexamethasone and insulin. The spontaneous differentiation of SV cells was suppressed by co-culture with adipocytes or by the addition of white adipocyte-conditioned medium. Conversely, the addition of SV cell-conditioned medium increased the expression of Ucp1. These results indicate that adipocytes secrete factors that suppress brown adipogenesis, whereas SV cells secrete factors that promote brown adipogenesis. Transcriptome analysis was conducted; however, no candidate suppressing factors secreted from adipocytes were identified. In contrast, 19 genes that encode secretory factors, including bone morphogenetic protein (BMP) family members, BMP3B, BMP5, and BMP7, were highly expressed in SV cells compared with adipocytes. Furthermore, the SMAD and MAPK signaling pathways, which represent the major BMP signaling pathways, were activated in SV cells, suggesting that BMPs secreted from SV cells induce brown adipogenesis in an autocrine manner through the SMAD/MAPK signaling pathways. Treatment of 5-day-old hamsters with type I BMP receptor inhibitor, LDN-193189, for 5 days reduced p38 MAPK phosphorylation and drastically suppressed BAT formation of interscapular adipose tissue. In conclusion, adipocytes and stromal cells regulate brown adipogenesis through secretory factors during the postnatal white-to-brown conversion of adipose tissue in Syrian hamsters.

Breast Cancer Endocrine Therapy Exhausts Adipocyte Progenitors Promoting Weight Gain and Glucose Intolerance

Breast cancer survivors treated with anti-estrogen therapies report weight gain and have an elevated risk of type 2 diabetes. Here, we show that current tamoxifen use did not influence body mass index but associated with larger breast adipocyte diameter only in women with obesity, suggesting adipose tissue may be targeted by breast cancer therapies. To understand the mechanisms behind these clinical findings, we investigated the impact of estrogen deprivation and tamoxifen in a relevant pre-clinical murine model of obesity. Specifically, mature female mice were housed at thermoneutrality and fed either a low-fat/low-sucrose (LFLS) or a high-fat/high-sucrose (HFHS) diet. Consistent with the high expression of Esr1 observed in single-cell RNA sequencing of mesenchymal stem cells from mouse adipose tissue, endocrine therapies associated with adipose accumulation and preadipocyte expansion, but resulted in adipocyte progenitor depletion only in the context of HFHS. Consequently, 7-week endocrine therapy supported adipocyte hypertrophy and was associated with hepatic steatosis, hyperinsulinemia, insulin resistance, and glucose intolerance, particularly in HFHS fed females. We administered HFHS fed females either metformin or pioglitazone, glucose lowering drugs used to treat diabetes, or treadmill interval exercise during endocrine therapy with the goal of improving whole body metabolism. All interventions prevented the effects of tamoxifen but not estrogen deprivation on adipocyte size and insulin resistance in HFHS-fed mice. This translational study suggests that endocrine therapies may act via ER-alpha to directly disrupt adipocyte progenitors and support adipocyte hypertrophy, leading to ectopic lipid deposition that may promote hyperinsulinemia, insulin resistance and type 2 diabetes. Interventions that target insulin action should be considered for some women receiving life-saving endocrine therapies for breast cancer.

AGMO Inhibitor Reduces 3T3-L1 Adipogenesis

Alkylglycerol monooxygenase (AGMO) is a tetrahydrobiopterin (BH4)-dependent enzyme with major expression in the liver and white adipose tissue that cleaves alkyl ether glycerolipids. The present study describes the disclosure and biological characterization of a candidate compound (Cp6), which inhibits AGMO with an IC50 of 30–100 µM and 5–20-fold preference of AGMO relative to other BH4-dependent enzymes, i.e., phenylalanine-hydroxylase and nitric oxide synthase. The viability and metabolic activity of mouse 3T3-L1 fibroblasts, HepG2 human hepatocytes and mouse RAW264.7 macrophages were not affected up to 10-fold of the IC50. However, Cp6 reversibly inhibited the differentiation of 3T3-L1 cells towards adipocytes, in which AGMO expression was upregulated upon differentiation. Cp6 reduced the accumulation of lipid droplets in adipocytes upon differentiation and in HepG2 cells exposed to free fatty acids. Cp6 also inhibited IL-4-driven differentiation of RAW264.7 macrophages towards M2-like macrophages, which serve as adipocyte progenitors in adipose tissue. Collectively, the data suggest that pharmacologic AGMO inhibition may affect lipid storage.