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Methods in Molecular Biology (Clifton,... 2019The 3T3-L1 murine preadipocyte cell line is a commonly used tool for analysis of the subcellular pathways involved in preadipocytic cell differentiation (a process also...
The 3T3-L1 murine preadipocyte cell line is a commonly used tool for analysis of the subcellular pathways involved in preadipocytic cell differentiation (a process also commonly known as adipogenesis). The major characteristic of adipogenesis is the intracellular accumulation of membrane-bound lipid droplets. Here, we describe methods used for the culture and transformation of these preadipocytes into mature adipocytes and quantification of intracellular lipid accumulation using the lipid specific dye, Oil Red O.
Topics: 3T3-L1 Cells; Adipocytes; Adipogenesis; Animals; Cell Culture Techniques; Cell Differentiation; Lipid Metabolism; Lipids; Mice
PubMed: 30535703
DOI: 10.1007/978-1-4939-8994-2_25 -
The Journal of Experimental Biology Mar 2018White adipose tissue (AT) is the main lipid storage depot in vertebrates. Initially considered to be a simple lipid store, AT has recently been recognized as playing a... (Review)
Review
White adipose tissue (AT) is the main lipid storage depot in vertebrates. Initially considered to be a simple lipid store, AT has recently been recognized as playing a role as an endocrine organ that is implicated in processes such as energy homeostasis and as a rich source of stem cells. Interest in adipogenesis has increased not only because of the prevalence of obesity, metabolic syndrome and type 2 diabetes in humans, but also in aquaculture because of the excessive fat deposition experienced in some cultured fish species, which may compromise both their welfare and their final product quality. Adipocyte development is well conserved among vertebrates, and this conservation has facilitated the rapid characterization of several adipogenesis models in fish. This Review presents the main findings of adipogenesis research based in primary cultures of the preadipocytes of farmed fish species. Zebrafish has emerged as an excellent model for studying the early stages of adipocyte fish development Nevertheless, larger fish species are more suitable for the isolation of preadipocytes from visceral AT and for studies in which preadipocytes are differentiated to form mature adipocytes. Differentiated adipocytes contain lipid droplets and express adipocyte marker genes such as those encoding the peroxisome proliferator activated receptor γ (), CCAAT-enhancer-binding protein α (), lipoprotein lipase (), fatty acid synthase (), fatty acid binding protein 11 (), fatty acid transporter protein1 (), adiponectin and leptin. Differentiated adipocytes also have elevated glycerol 3-phosphate (G3P) dehydrogenase (GPDH) activity. To better understand fish adipocyte development and regulation, different adipokines, fatty acids, growth factors and PPAR agonists have been studied, providing relevant insights into which factors affect these processes and counterbalance AT dysregulation.
Topics: Adipocytes; Adipogenesis; Animals; Aquaculture; Cell Differentiation; Fishes
PubMed: 29514876
DOI: 10.1242/jeb.161588 -
Life Sciences Feb 2023In recent decades, obesity has extensively emerged to the level of pandemics. It's significantly associated with serious co-morbidities that could decrease life quality... (Review)
Review
In recent decades, obesity has extensively emerged to the level of pandemics. It's significantly associated with serious co-morbidities that could decrease life quality and even life expectancy. Obesity has several determinants, such as age, sex, endocrine, and genetic factors. The miRNAs have emerged as genetic factors affecting obesity. The miRNAs are small noncoding nucleic acids that can modify gene expression and hence, control biological processes. The miRNAs can greatly affect many biological processes in obesity, such as adipogenesis, lipid metabolism, and homeostasis. As a result, the entry of miRNAs in obesity therapeutic approaches has been strongly advised as miRNAs mimics, inhibitors, and stimulators. Hence, this review aims to point out a summarized and updated overview of miRNAs and their roles in obesity and its included processes, such as adipogenesis and lipid metabolism. Besides, we also review recent applications of miRNAs as a treatment approach for obesity.
Topics: Humans; Adipogenesis; Lipid Metabolism; MicroRNAs; Obesity
PubMed: 36639051
DOI: 10.1016/j.lfs.2023.121382 -
Annual Review of Physiology Feb 2021Adipose tissue depots in distinct anatomical locations mediate key aspects of metabolism, including energy storage, nutrient release, and thermogenesis. Although... (Review)
Review
Adipose tissue depots in distinct anatomical locations mediate key aspects of metabolism, including energy storage, nutrient release, and thermogenesis. Although adipocytes make up more than 90% of adipose tissue volume, they represent less than 50% of its cellular content. Here, I review recent advances in genetic lineage tracing and transcriptomics that reveal the identities of the heterogeneous cell populations constituting mouse and human adipose tissues. In addition to mature adipocytes and their progenitors, these include endothelial and various immune cell types that together orchestrate adipose tissue development and functions. One salient finding is the identification of progenitor subtypes that can modulate adipogenic capacity through paracrine mechanisms. Another is the description of fate trajectories of monocyte/macrophages, which can respond maladaptively to nutritional and thermogenic stimuli, leading to metabolic disease. These studies have generated an extraordinary source of publicly available data that can be leveraged to explore commonalities and differences among experimental models, providing new insights into adipose tissues and their role in metabolic disease.
Topics: Adipocytes; Adipogenesis; Adipose Tissue; Animals; Humans; Thermogenesis
PubMed: 33566675
DOI: 10.1146/annurev-physiol-031620-095446 -
Molecular and Cellular Biochemistry Jul 2021Lipid metabolism, which encompasses synthesis and degradation of lipids, is critical for a wide range of cellular functions, including structural and morphological... (Review)
Review
Lipid metabolism, which encompasses synthesis and degradation of lipids, is critical for a wide range of cellular functions, including structural and morphological properties of organelles, energy storage, signalling, and the stability and function of membrane proteins. Adipose tissue is a dynamic tissue type that performs a lot of significant physiological functions, including secretion, and is involved in maintaining homeostasis and in regulatory roles of other tissues based on paracrine or endocrine. More recently, several classes of non-coding RNAs (ncRNAs), such as long non-coding RNA (lncRNA), microRNA (miRNA) and circular RNA (circRNA), have been discovered in adipocytes, and they act as critical regulators of gene expression in adipogenesis and regulate adipogenesis through multiple pathways. In the present paper, we discussed several classes of non-coding RNAs and summarized the latest research on the regulatory role of ncRNAs in bovine adipogenesis. We gave examples for known modes of action to look forward to providing reference information future scientific research in cattle breeding.
Topics: Adipogenesis; Adipose Tissue, White; Animals; Cattle; MicroRNAs; RNA, Long Noncoding
PubMed: 33730298
DOI: 10.1007/s11010-021-04132-2 -
Nature Communications Jan 2023Fibro-adipogenic progenitors (FAPs) play a crucial role in skeletal muscle regeneration, as they generate a favorable niche that allows satellite cells to perform...
Fibro-adipogenic progenitors (FAPs) play a crucial role in skeletal muscle regeneration, as they generate a favorable niche that allows satellite cells to perform efficient muscle regeneration. After muscle injury, FAP content increases rapidly within the injured muscle, the origin of which has been attributed to their proliferation within the muscle itself. However, recent single-cell RNAseq approaches have revealed phenotype and functional heterogeneity in FAPs, raising the question of how this differentiation of regenerative subtypes occurs. Here we report that FAP-like cells residing in subcutaneous adipose tissue (ScAT), the adipose stromal cells (ASCs), are rapidly released from ScAT in response to muscle injury. Additionally, we find that released ASCs infiltrate the damaged muscle, via a platelet-dependent mechanism and thus contribute to the FAP heterogeneity. Moreover, we show that either blocking ASCs infiltration or removing ASCs tissue source impair muscle regeneration. Collectively, our data reveal that ScAT is an unsuspected physiological reservoir of regenerative cells that support skeletal muscle regeneration, underlining a beneficial relationship between muscle and fat.
Topics: Humans; Muscle, Skeletal; Adipose Tissue; Cell Differentiation; Adipogenesis; Muscular Diseases
PubMed: 36604419
DOI: 10.1038/s41467-022-35524-7 -
Molecular Metabolism Aug 2022Regulation of proteasomal activity is an essential component of cellular proteostasis and function. This is evident in patients with mutations in proteasome subunits and...
OBJECTIVE
Regulation of proteasomal activity is an essential component of cellular proteostasis and function. This is evident in patients with mutations in proteasome subunits and associated regulators, who suffer from proteasome-associated autoinflammatory syndromes (PRAAS). These patients display lipodystrophy and fevers, which may be partly related to adipocyte malfunction and abnormal thermogenesis in adipose tissue. However, the cell-intrinsic pathways that could underlie these symptoms are unclear. Here, we investigate the impact of two proteasome subunits implicated in PRAAS, Psmb4 and Psmb8, on differentiation, function and proteostasis of brown adipocytes.
METHODS
In immortalized mouse brown pre-adipocytes, levels of Psmb4, Psmb8, and downstream effectors genes were downregulated through reverse transfection with siRNA. Adipocytes were differentiated and analyzed with various assays of adipogenesis, lipogenesis, lipolysis, inflammation, and respiration.
RESULTS
Loss of Psmb4, but not Psmb8, disrupted proteostasis and adipogenesis. Proteasome function was reduced upon Psmb4 loss, but partly recovered by the activation of Nuclear factor, erythroid-2, like-1 (Nfe2l1). In addition, cells displayed higher levels of surrogate inflammation and stress markers, including Activating transcription factor-3 (Atf3). Simultaneous silencing of Psmb4 and Atf3 lowered inflammation and restored adipogenesis.
CONCLUSIONS
Our study shows that Psmb4 is required for adipocyte development and function in cultured adipocytes. These results imply that in humans with PSMB4 mutations, PRAAS-associated lipodystrophy is partly caused by disturbed adipogenesis. While we uncover a role for Nfe2l1 in the maintenance of proteostasis under these conditions, Atf3 is a key effector of inflammation and blocking adipogenesis. In conclusion, our work highlights how proteasome dysfunction is sensed and mitigated by the integrated stress response in adipocytes with potential relevance for PRAAS patients and beyond.
Topics: Adipocytes, Brown; Adipogenesis; Animals; Inflammation; Lipodystrophy; Mice; Proteasome Endopeptidase Complex
PubMed: 35636710
DOI: 10.1016/j.molmet.2022.101518 -
Current Opinion in Organ Transplantation Oct 2019It has been increasingly common to use adipose tissue for regenerative and reconstructive purposes. Applications of autologous fat transfer and different stem cell... (Review)
Review
PURPOSE OF REVIEW
It has been increasingly common to use adipose tissue for regenerative and reconstructive purposes. Applications of autologous fat transfer and different stem cell therapies have significant limitations and adipose tissue engineering may have the potential to be an important strategy in the reconstruction of large tissue defects. A better understanding of adipogenesis will help to develop strategies to make adipose tissue more effective for repairing volumetric defects.
RECENT FINDINGS
We provide an overview of the current applications of adipose tissue transfer and cellular therapy methods for soft tissue reconstruction, cellular physiology, and factors influencing adipogenesis, and adipose tissue engineering. Furthermore, we discuss mechanical properties and vascularization strategies of engineered adipose tissue, and its potential applications in the clinical settings.
SUMMARY
Autologous fat tissue transfer is the standard of care technique for the majority of surgeons; however, high resorption rates, poor perfusion within a large volume fat graft and widely inconsistent graft survival are the main limitations. Adipose tissue engineering is a promising field to reach the first goal of producing adipose tissue which has more predictable survival and higher graft retention rates. Advancements of scaffold and vascularization strategies will contribute to metabolically and functionally more relevant adipose tissue engineering.
Topics: Adipogenesis; Adipose Tissue; Cell- and Tissue-Based Therapy; Humans; Therapy, Soft Tissue; Tissue Engineering; Tissue Scaffolds; Transplantation, Autologous
PubMed: 31397728
DOI: 10.1097/MOT.0000000000000694 -
American Journal of Physiology. Cell... Sep 2023CROP-Seq combines gene silencing using CRISPR interference with single-cell RNA sequencing. Here, we applied CROP-Seq to study adipogenesis and adipocyte biology. Human...
CROP-Seq combines gene silencing using CRISPR interference with single-cell RNA sequencing. Here, we applied CROP-Seq to study adipogenesis and adipocyte biology. Human preadipocyte SGBS cell line expressing KRAB-dCas9 was transduced with a sgRNA library. Following selection, individual cells were captured using microfluidics at different timepoints during adipogenesis. Bioinformatic analysis of transcriptomic data was used to determine the knockdown effects, the dysregulated pathways, and to predict cellular phenotypes. Single-cell transcriptomes recapitulated adipogenesis states. For all targets, over 400 differentially expressed genes were identified at least at one timepoint. As a validation of our approach, the knockdown of and (which encode key proadipogenic transcription factors) resulted in the inhibition of adipogenesis. Gene set enrichment analysis generated hypotheses regarding the molecular function of novel genes. knockdown led to downregulation of transcriptional response to proinflammatory cytokine TNF-α in preadipocytes and to decreased CXCL-16 and IL-6 secretion. knockdown resulted in increased expression of adipogenesis markers. In summary, this powerful, hypothesis-free tool can identify novel regulators of adipogenesis, preadipocyte, and adipocyte function associated with metabolic disease. Genomics efforts led to the identification of many genomic loci that are associated with metabolic traits, many of which are tied to adipose tissue function. However, determination of the causal genes, and their mechanism of action in metabolism, is a time-consuming process. Here, we use an approach to determine the transcriptional outcome of candidate gene knockdown for multiple genes at the same time in a human cell model of adipogenesis.
Topics: Humans; RNA, Guide, CRISPR-Cas Systems; Adipogenesis; Adipocytes; Cell Line; Metabolic Diseases; Cell Differentiation
PubMed: 37486064
DOI: 10.1152/ajpcell.00148.2023 -
Molecular Metabolism Aug 2023Adipogenesis is key to maintaining organism-wide energy balance and healthy metabolic phenotype, making it critical to thoroughly comprehend its molecular regulation in...
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 Identified several novel transcription factors as potential targets for adipogenic/thermogenic drivers in humans. Among these novel candidates, we explored the role of TRPS1 in adipocyte differentiation and showed that its knockdown impairs white adipogenesis in vitro. 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.
Topics: Humans; Animals; Mice; Adipogenesis; RNA-Seq; Adipose Tissue, Brown; Adipose Tissue, White; Cell Differentiation; Repressor Proteins
PubMed: 37286033
DOI: 10.1016/j.molmet.2023.101746