Abstract:Obesity has become one of the leading pathophysiologic disorders in recent years. Adipose tissue is the main tissue related to obesity and is known to play a role in various physiological complications, including type 2 diabetes. To better understand how the fat tissue develops, we used an in vitro live cell imaging system to quantify the adipogenesis by means of nondestructive digital imaging to monitor the accumulation of intracellular lipid droplets (LDs), a hallmark of adipogenesis, from the macro- to the … Show more
“…The results indicated an increase in the Hippo-signaling pathway as represented by the nuclear mechano-sensors YAP1 and Lamin A/C with an increase in the niche rigidity (Figure 4). These findings support the idea that static stretched conditions lead to accelerated adipogenesis (Hara et al, 2011;Lustig et al, 2018;Mor-Yossef Moldovan et al, 2019;Shoham & Gefen, 2012).…”
Section: Discussionsupporting
confidence: 87%
“…week. Differentiation was induced when the cultures reached a confluence of approximately 90% according to the protocol described in an earlier publication (Mor-Yossef Moldovan et al, 2019).…”
Adipose tissue plays a leading role in obesity, which, in turn, can lead to Type 2 diabetes. Adipocytes (AD) respond to the biomechanical stimulation experienced in fat tissue under static stretch during prolonged sitting or lying. To investigate the effect of such chronic stimulation on adipocyte cell metabolism, we used an in vitro system to mimic the static stretch conditions. Under in vitro culture stretching, cells were analyzed at the single‐cell level and we measured an increase in the projected area of the AD and higher content of lipid droplets. A decrease in the projected area of these cells’ nucleus is associated with peroxisome proliferator‐activated receptor‐gamma expression and heterochromatin. This is the first study to reveal proteins that were altered under static stretch following a mass spectrometry analysis and main pathways that affect cell fate and metabolism. Bioinformatics analysis of the proteins indicated an increase in mitochondrial activity and associated pathways under static stretch stimulation. Quantification of the mitochondrial activity by 3‐[4,5‐dimethylthiazol‐2‐yl]‐2,5‐diphenyl tetrazolium bromide (MTT) assay and the ATPase related proteins specifically measured ATP5B indicated an increase in adipogenesis which points to a higher rate of cell metabolism under static stretch. In summary, our results elaborate on the metabolism of AD exposed to biomechanical stimulation, that is, associated with altered cellular protein profile and thereby influenced cell fate. The static stretch stimulation accelerated adipocyte differentiation through increased mitochondrial activity. Hence, in this study, we introduce a new perspective in understanding the molecular regulation of mechano‐transduction in adipogenesis.
“…The results indicated an increase in the Hippo-signaling pathway as represented by the nuclear mechano-sensors YAP1 and Lamin A/C with an increase in the niche rigidity (Figure 4). These findings support the idea that static stretched conditions lead to accelerated adipogenesis (Hara et al, 2011;Lustig et al, 2018;Mor-Yossef Moldovan et al, 2019;Shoham & Gefen, 2012).…”
Section: Discussionsupporting
confidence: 87%
“…week. Differentiation was induced when the cultures reached a confluence of approximately 90% according to the protocol described in an earlier publication (Mor-Yossef Moldovan et al, 2019).…”
Adipose tissue plays a leading role in obesity, which, in turn, can lead to Type 2 diabetes. Adipocytes (AD) respond to the biomechanical stimulation experienced in fat tissue under static stretch during prolonged sitting or lying. To investigate the effect of such chronic stimulation on adipocyte cell metabolism, we used an in vitro system to mimic the static stretch conditions. Under in vitro culture stretching, cells were analyzed at the single‐cell level and we measured an increase in the projected area of the AD and higher content of lipid droplets. A decrease in the projected area of these cells’ nucleus is associated with peroxisome proliferator‐activated receptor‐gamma expression and heterochromatin. This is the first study to reveal proteins that were altered under static stretch following a mass spectrometry analysis and main pathways that affect cell fate and metabolism. Bioinformatics analysis of the proteins indicated an increase in mitochondrial activity and associated pathways under static stretch stimulation. Quantification of the mitochondrial activity by 3‐[4,5‐dimethylthiazol‐2‐yl]‐2,5‐diphenyl tetrazolium bromide (MTT) assay and the ATPase related proteins specifically measured ATP5B indicated an increase in adipogenesis which points to a higher rate of cell metabolism under static stretch. In summary, our results elaborate on the metabolism of AD exposed to biomechanical stimulation, that is, associated with altered cellular protein profile and thereby influenced cell fate. The static stretch stimulation accelerated adipocyte differentiation through increased mitochondrial activity. Hence, in this study, we introduce a new perspective in understanding the molecular regulation of mechano‐transduction in adipogenesis.
“…In this study, we also described a new concept of adding NPs into the production process of the biocomposites in order to serve as a delivery system, for example by conjugating NPs and growth factors for biological and chemical signals in the scaffold. We demonstrated endocytosis uptake of FITC-NPs by the cells and following our earlier study, we reported that NPs could induce gene activation, proliferation, and differentiation in human mesenchymal cells [29]. We followed mesenchymal cells uptake of the FITC labeled NPs and evaluated their effect on cell viability and morphology in a 3D biocomposite and in 2D in-vitro systems over time.…”
Mesenchymal stem cells serve as the body's reservoir for healing and tissue regeneration. In cases of severe tissue trauma where there is also a need for tissue organization, a scaffold may be of use to support the cells in the damaged tissue. Such a scaffold should be composed of a material that can biomimic the mechanical and biological properties of the target tissues in order to support autologous cell-adhesion, their proliferation, and differentiation. In this study, we developed and assayed a new biocomposite made of unique collagen fibers and alginate hydrogel that was assessed for the ability to support mesenchymal cell-proliferation and differentiation. Analysis over 11 weeks in vitro demonstrated that the scaffold was biocompatible and supports the cells viability and differentiation to produce tissue-like structures or become adipocyte under differentiation medium. When the biocomposite was enriched with nano particles (NPs), mesenchymal cells grew well after uptake of fluorescein isothiocyanate (FITC) labeled NPs, maintained their viability, migrated through the biocomposite, reached, and adhered to the tissue culture dish. These promising findings revealed that the scaffold supports the growth and differentiation of mesenchymal cells that demonstrate their full physiological function with no sign of material toxicity. The cells' functionality performance indicates and suggests that the scaffold is suitable to be developed as a new medical device that has the potential to support regeneration and the production of functional tissue.Biomolecules 2020, 10, 458 2 of 11 new tissue to improves regeneration and healing processes [1,[5][6][7][8][9][10][11]. The use of naturally made scaffolds is the best approach, but they may lack the physical/mechanical properties needed for the new tissue formed.We have developed a composite biomaterial scaffold made from collagen fibers embedded in alginate hydrogel [12][13][14][15][16][17]. The collagen used as a biopolymer for producing implantable medical devices is usually obtained through chemical extraction and reconstitution processes, resulting in products with poor mechanical strength [18][19][20]. However, the biocomposite we describe here focuses on the unique coral-derived collagen fibers that were recently identified for their molecular composition and structure [13,14] and mechanical properties [12,[15][16][17]. These attributes make our biocomposite uniquely suitable for use in a new generation of scaffolds that can be tailored to meet the mechanical properties of various target tissues. The biological and mechanical properties of these collagen fibers facilitate the desired physiological activity, together with the promotion of cell proliferation and differentiation. The collagen fibers are embedded in an alginate hydrogel, a polysaccharide extracted from algae that has already been applied in a wide spectrum of uses such as food, pharmaceutics, and medical device industries [21][22][23][24]. The addition of the collagen fibers reinforces the alginate in ...
“…As the experiment progressed and the LDs accumulated further, the mean cell projected area increased (Figs. 3f and 3g) , 2018;Lustig et al, 2018aLustig et al, , 2018b. The lipid contents in a specific FOV increased during the differentiation process (Fig.…”
Section: Monitoring Adipocyte Differentiation At the Single-cell Levelmentioning
3T3-L1 cells serve as model systems for studying adipogenesis and research of adipose tissue-related diseases, e.g. obesity and diabetes. Here, we present two novel and complementary nondestructive methods for adipogenesis analysis of living cells which facilitate continuous monitoring of the same culture over extended periods of time, and are applied in parallel at the macro-and micro-scales. At the macro-scale, we developed visual differences mapping (VDM), a novel method which allows to determine level of adipogenesis (LOA)-a numerical index which quantitatively describes the extent of differentiation in the whole culture, and percentage area populated by adipocytes (PAPBA) across a whole culture, based on the apparent morphological differences between preadipocytes and adipocytes. At the micro-scale, we developed an improved version of our previously published image-processing algorithm, which now provides data regarding single-cell morphology and lipid contents. Both methods were applied here synergistically for measuring differentiation levels in cultures over multiple weeks. VDM revealed that the mean LOA value reached 1.11 ± 0.06 and the mean PAPBA value reached >60%. Micro-scale analysis revealed that during differentiation, the cells transformed from a fibroblast-like shape to a circular shape with a build-up of lipid droplets. We predict a vast potential for implementation of these methods in adipose-related pharmacological research, such as in metabolic-syndrome studies.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.