scholarly journals Hypertrophied human adipocyte spheroids as in vitro model of weight gain and adipose tissue dysfunction

2021 ◽  
Author(s):  
Anna Ioannidou ◽  
Shemim Alatar ◽  
Matilda Ahlander ◽  
Amanda Hornell ◽  
Rachel M Fisher ◽  
...  
Keyword(s):  
Weight Gain ◽  
Lipid Droplets ◽  
Molecular Mechanisms ◽  
Metabolic Dysfunction ◽  
Obesity Prevalence ◽  
Human Adipocytes ◽  
Human Adipocyte ◽  
Vitro Model

The rise in obesity prevalence has created an urgent need for new and improved methods to study human adipocytes and the pathogenic effects of weight gain in vitro. Despite numerous studies showing the advantages of culturing adipocyte progenitors as 3D structures, the majority continue using traditional 2D cultures which result in small, multilocular adipocytes with poor representability. We hypothesized that providing differentiating pre-adipocytes with a vascular growth niche would mimic in vivo adipogenesis and improve the differentiation process. Here we present a simple, easily applicable culture protocol that allows for the differentiation and culturing of human adipocytes with a more unilocular morphology and larger lipid droplets than previous protocols. We moreover offer a protocol for inducing adipocyte enlargement in vitro, resulting in larger lipid droplets and the development of several key features of adipocyte dysfunction, including altered adipokine secretions and impaired lipolysis. Taken together, our hypertrophied human adipocyte spheroids offer an improved culture system for studying the cellular and molecular mechanisms causing metabolic dysfunction and inflammation during weight gain.

Identity of the renin cell is mediated by cAMP and chromatin remodeling: an in vitro model for studying cell recruitment and plasticity

2008 ◽  
Vol 294 (2) ◽  
pp. H699-H707 ◽  
Author(s):  
Ellen Steward Pentz ◽  
Maria Luisa S. Sequeira Lopez ◽  
Magali Cordaillat ◽  
R. Ariel Gomez
Keyword(s):  
Chromatin Remodeling ◽  
In Vitro Model ◽  
Molecular Mechanisms ◽  
Fluorescent Protein ◽  
Yellow Fluorescent Protein ◽  
Histone H4 Acetylation ◽  
Renin Gene ◽  
Vitro Model

The renin-angiotensin system (RAS) regulates blood pressure and fluid-electrolyte homeostasis. A key step in the RAS cascade is the regulation of renin synthesis and release by the kidney. We and others have shown that a major mechanism to control renin availability is the regulation of the number of cells capable of making renin. The kidney possesses a pool of cells, mainly in its vasculature but also in the glomeruli, capable of switching from smooth muscle to endocrine renin-producing cells when homeostasis is threatened. The molecular mechanisms governing the ability of these cells to turn the renin phenotype on and off have been very difficult to study in vivo. We, therefore, developed an in vitro model in which cells of the renin lineage are labeled with cyan fluorescent protein and cells actively making renin mRNA are labeled with yellow fluorescent protein. The model allowed us to determine that it is possible to culture cells of the renin lineage for numerous passages and that the memory to express the renin gene is maintained in culture and can be reenacted by cAMP and chromatin remodeling (histone H4 acetylation) at the cAMP-responsive element in the renin gene.

scholarly journals SOX2 and BRN2 as Key Transcription Factors in Neural Rosette Formation In Vitro

2021 ◽  
Vol 1 (1) ◽  
Author(s):  
Zuzana Hudáčová ◽  
Keyword(s):  
Transcription Factors ◽  
Neural Development ◽  
Molecular Mechanisms ◽  
Rosette Formation ◽  
In Vivo Studies ◽  
Confounding Variables ◽  
Vitro Model ◽  
Underlying Processes

Although neurogenesis has been well studied, its molecular mechanisms remain largely unknown due to the challenges posed by the complexity of the underlying processes. Whilst in vivo studies can be used to study neurogenesis, the inability to control confounding variables complicate findings. Therefore, the purpose of this study was to identify the markers of in vitro neural rosette formation and describe the formation of neural rosettes from pluripotent stem cells using immunofluorescence analysis. The protocol of stem cell cultivation and induction of neural rosette formation was tested. Following, two transcription factors, BRN2 and SOX2, were fluorescently labelled and cells were imaged over a period of eight days. It was identified that SOX2 and BRN2 are expressed during in vitro neural rosette formation. These results are concurrent with in vivo neurogenesis, which suggests that neural rosettes could be a suitable in vitro model for researching neural development. Given that mistakes can arise during neurogenesis, such as neural tube defects, developing robust models to understand the formation of the nervous system is important. Moving forward, a detailed molecular understanding of neural rosette formation has the potential to be used for targeting specific transcription factors to treat or prevent problematic neurogenesis.

scholarly journals Cytotoxicity of atropine to human corneal endothelial cells by inducing mitochondrion-dependent apoptosis

2016 ◽  
Vol 241 (13) ◽  
pp. 1457-1465 ◽  
Author(s):  
Qian Wen ◽  
Ting-Jun Fan ◽  
Cheng-Lei Tian
Keyword(s):  
Corneal Endothelium ◽  
Molecular Mechanisms ◽  
Anticholinergic Drug ◽  
Death Receptor ◽  
Time Dependent ◽  
Dependent Manner ◽  
Plasma Membrane Permeability ◽  
Vitro Model

Atropine, a widely used topical anticholinergic drug, might have adverse effects on human corneas in vivo. However, its cytotoxic effect on human corneal endothelium (HCE) and its possible mechanisms are unclear. Here, we investigated the cytotoxicity of atropine and its underlying cellular and molecular mechanisms using an in vitro model of HCE cells and verified the cytotoxicity using cat corneal endothelium (CCE) in vivo. Our results showed that atropine at concentrations above 0.3125 g/L could induce abnormal morphology and viability decline in a dose- and time-dependent manner in vitro. The cytotoxicity of atropine was proven by the induced density decrease and abnormality of morphology and ultrastructure of CCE cells in vivo. Meanwhile, atropine could also induce dose- and time-dependent elevation of plasma membrane permeability, G1 phase arrest, phosphatidylserine externalization, DNA fragmentation, and apoptotic body formation of HCE cells. Moreover, 2.5 g/L atropine could also induce caspase-2/-3/-9 activation, mitochondrial transmembrane potential disruption, downregulation of anti-apoptotic Bcl-2 and Bcl-xL, upregulation of pro-apoptotic Bax and Bad, and upregulation of cytoplasmic cytochrome c and apoptosis-inducing factor. In conclusion, atropine above 1/128 of its clinical therapeutic dosage has a dose- and time-dependent cytotoxicity to HCE cells in vitro which is confirmed by CCE cells in vivo, and its cytotoxicity is achieved by inducing HCE cell apoptosis via a death receptor-mediated mitochondrion-dependent signaling pathway. Our findings provide new insights into the cytotoxicity and apoptosis-inducing effect of atropine which should be used with great caution in eye clinic.

scholarly journals REPTOR/CREBRF encode key regulators of muscle energy metabolism

2021 ◽  
Author(s):  
Pedro Saavedra ◽  
Phillip A Dumesic ◽  
Yanhui Hu ◽  
Patrick Jouandin ◽  
Richard Binari ◽  
...  
Keyword(s):  
Energy Metabolism ◽  
Molecular Mechanisms ◽  
In Vivo Studies ◽  
Metabolic Flexibility ◽  
Metabolic Dysfunction ◽  
Glucose Content ◽  
Muscle Energy ◽  
Muscle Energy Metabolism

Metabolic flexibility of muscle tissue describes the capacity to use glucose or lipids as energy substrates and its disruption is associated with metabolic dysfunction. Cancer-induced cachexia is a metabolic syndrome linked with muscle wasting, changes in muscle energy metabolism and lower life expectancy in cancer patients. The molecular mechanisms driving metabolic changes in muscle, however, are poorly characterized. Here, using a Drosophila model of systemic metabolic dysfunction triggered by yorkie-induced gut tumors, we identify the transcription factor REPTOR as a key regulator of energy metabolism in muscle. We show that REPTOR is upregulated in muscles of adult flies with gut yorkie-tumors, where it is necessary to modulate glucose metabolism. REPTOR expression in muscles is induced by ImpL2, a tumor-derived insulin binding protein that reduces systemic insulin signaling, or by nutritional restriction. Further, in vitro and in vivo studies indicate that high activity of REPTOR is sufficient to increase glucose content, transcriptionally repress phosphofructokinase and increase mitochondrial respiration. Consistent with the fly studies, higher levels of CREBRF, the mammalian ortholog of REPTOR, reduce glycolysis in mouse myotubes while promoting an oxidative phenotype. Altogether, our results implicate REPTOR/CREBRF as key regulators of muscle metabolism and metabolic flexibility that share a conserved function as repressors of glycolysis and promoters of oxidative phosphorylation.

scholarly journals Hypertrophied human adipocyte spheroids as in vitro model of weight gain and adipose tissue dysfunction

2021 ◽  
Author(s):  
Anna Ioannidou ◽  
Shemim Alatar ◽  
Ruby Schipper ◽  
Fabiana Baganha ◽  
Matilda Åhlander ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Weight Gain ◽  
In Vitro Model ◽  
Human Adipocyte ◽  
Adipose Tissue Dysfunction ◽  
Vitro Model

Sepsis induces the transcription of the glucocorticoid receptor in skeletal muscle cells

2003 ◽  
Vol 105 (3) ◽  
pp. 383-391 ◽  
Author(s):  
Xiaoyan SUN ◽  
Joshua M. V. MAMMEN ◽  
Xintian TIAN
Keyword(s):  
Skeletal Muscle ◽  
Molecular Mechanisms ◽  
L6 Myotubes ◽  
Breakdown Rates ◽  
Dependent Protein ◽  
Vitro Model ◽  
Energy Dependent ◽  
L6 Myocytes

Evidence from a recent study indicates that glucocorticoids (GCs) mediate skeletal muscle proteolysis during sepsis via the GC receptor (GR) pathway. Attempts to identify the mechanisms regulating GR gene expression in skeletal muscle during sepsis have been hampered by the lack of an appropriate in vitro model system that can mimic in vivo septic conditions. In the present study, we report that GR gene transcription in L6 myocytes in vitro is up-regulated by treatment with sera from septic rats in a manner similar to that measured in septic rats in vivo. Sera from septic rats were collected from animals in which sepsis was induced by caecal ligation and puncture and from control rats that were sham-operated. Finally, by treating L6 myotubes with the GR antagonist RU 38486, thereby preventing sepsis-induced GR transcription, we confirmed that the possible septic effect on the GR was due to increased GCs. L6 myocytes treated with sera from septic rats might therefore be useful as an experimental model for identifying the molecular mechanisms by which the GR regulates muscle cachexia during sepsis. Furthermore, RU 38486 inhibited the sepsis-induced increase in total and myofibrillar energy-dependent protein breakdown rates in incubated extensor digitorum longus muscles from septic and sham-operated rats, as measured by release of tyrosine and 3-methylhistidine respectively. Our results demonstrate for the first time that sepsis induces GR transcription in skeletal muscle, and supports the hypothesis that the GC-induced proteolysis under sepsis is partially a consequence of GR activation.

scholarly journals Bisphenol A effects on gene expression in adipocytes from children: association with metabolic disorders

2015 ◽  
Vol 54 (3) ◽  
pp. 289-303 ◽  
Author(s):  
Ciro Menale ◽  
Maria Teresa Piccolo ◽  
Grazia Cirillo ◽  
Raffaele A Calogero ◽  
Alfonso Papparella ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Bisphenol A ◽  
Metabolic Disorders ◽  
Molecular Mechanisms ◽  
Endocrine Function ◽  
Human Adipocytes ◽  
Fat Cell ◽  
Endocrine Disrupting Chemical

Bisphenol A (BPA) is a xenobiotic endocrine-disrupting chemical.In vitroandin vivostudies have indicated that BPA alters endocrine-metabolic pathways in adipose tissue, which increases the risk of metabolic disorders and obesity. BPA can affect adipose tissue and increase fat cell numbers or sizes by regulating the expression of the genes that are directly involved in metabolic homeostasis and obesity. Several studies performed in animal models have accounted for an obesogen role of BPA, but its effects on human adipocytes – especially in children – have been poorly investigated. The aim of this study is to understand the molecular mechanisms by which environmentally relevant doses of BPA can interfere with the canonical endocrine function that regulates metabolism in mature human adipocytes from prepubertal, non-obese children. BPA can act as an estrogen agonist or antagonist depending on the physiological context. To identify the molecular signatures associated with metabolism, transcriptional modifications of mature adipocytes from prepubertal children exposed to estrogen were evaluated by means of microarray analysis. The analysis of deregulated genes associated with metabolic disorders allowed us to identify a small group of genes that are expressed in an opposite manner from that of adipocytes treated with BPA. In particular, we found that BPA increases the expression of pro-inflammatory cytokines and the expression ofFABP4andCD36, two genes involved in lipid metabolism. In addition, BPA decreases the expression ofPCSK1, a gene involved in insulin production. These results indicate that exposure to BPA may be an important risk factor for developing metabolic disorders that are involved in childhood metabolism dysregulation.

Flavonoids as P-gp Inhibitors: A Systematic Review of SARs

2019 ◽  
Vol 26 (25) ◽  
pp. 4799-4831 ◽  
Author(s):  
Jiahua Cui ◽  
Xiaoyang Liu ◽  
Larry M.C. Chow
Keyword(s):  
Molecular Mechanisms ◽  
Metastatic Cancer ◽  
Drug Candidates ◽  
Chemical Structures ◽  
Transporter Family ◽  
Promising Area ◽  
New Generation ◽  
Nontoxic Inhibitors

P-glycoprotein, also known as ABCB1 in the ABC transporter family, confers the simultaneous resistance of metastatic cancer cells towards various anticancer drugs with different targets and diverse chemical structures. The exploration of safe and specific inhibitors of this pump has always been the pursuit of scientists for the past four decades. Naturally occurring flavonoids as benzopyrone derivatives were recognized as a class of nontoxic inhibitors of P-gp. The recent advent of synthetic flavonoid dimer FD18, as a potent P-gp modulator in reversing multidrug resistance both in vitro and in vivo, specifically targeted the pseudodimeric structure of the drug transporter and represented a new generation of inhibitors with high transporter binding affinity and low toxicity. This review concerned the recent updates on the structure-activity relationships of flavonoids as P-gp inhibitors, the molecular mechanisms of their action and their ability to overcome P-gp-mediated MDR in preclinical studies. It had crucial implications on the discovery of new drug candidates that modulated the efflux of ABC transporters and also provided some clues for the future development in this promising area.

Targeting PPARalpha in Alzheimer's Disease

2018 ◽  
Vol 15 (4) ◽  
pp. 345-354 ◽  
Author(s):  
Barbara D'Orio ◽  
Anna Fracassi ◽  
Maria Paola Cerù ◽  
Sandra Moreno
Keyword(s):  
Oxidative Stress ◽  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Molecular Mechanisms ◽  
Redox Homeostasis ◽  
Antioxidant Response ◽  
Peroxisome Proliferator ◽  
Prevailing Paradigm

Background: The molecular mechanisms underlying Alzheimer's disease (AD) are yet to be fully elucidated. The so-called “amyloid cascade hypothesis” has long been the prevailing paradigm for causation of disease, and is today being revisited in relation to other pathogenic pathways, such as oxidative stress, neuroinflammation and energy dysmetabolism. The peroxisome proliferator-activated receptors (PPARs) are expressed in the central nervous system (CNS) and regulate many physiological processes, such as energy metabolism, neurotransmission, redox homeostasis, autophagy and cell cycle. Among the three isotypes (α, β/δ, γ), PPARγ role is the most extensively studied, while information on α and β/δ are still scanty. However, recent in vitro and in vivo evidence point to PPARα as a promising therapeutic target in AD. Conclusion: This review provides an update on this topic, focussing on the effects of natural or synthetic agonists in modulating pathogenetic mechanisms at AD onset and during its progression. Ligandactivated PPARα inihibits amyloidogenic pathway, Tau hyperphosphorylation and neuroinflammation. Concomitantly, the receptor elicits an enzymatic antioxidant response to oxidative stress, ameliorates glucose and lipid dysmetabolism, and stimulates autophagy.

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