scholarly journals Dysregulated Alternative Splicing Pattern of PKCδ during Differentiation of Human Preadipocytes Represents Distinct Differences between Lean and Obese Adipocytes

ISRN Obesity ◽  
2013 ◽  
Vol 2013 ◽  
pp. 1-9 ◽  
Author(s):  
Gay Carter ◽  
André Apostolatos ◽  
Rekha Patel ◽  
Abhishek Mathur ◽  
Denise Cooper ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Expression Pattern ◽  
Molecular Mechanisms ◽  
Developmental Process ◽  
Splice Variants ◽  
Apoptosis Pathway ◽  
Antiapoptotic Proteins ◽  
Alternatively Spliced ◽  
Human Preadipocytes

Obesity and its comorbidities affect millions of people. Here, we demonstrate that human preadipocytes are susceptible to programmed cell death (apoptosis) while mature adipocytes are resistant to apoptosis. The molecular mechanisms underlying the phenotype of apoptosis-resistant adipocytes are lesser known. To study the role of apoptosis and define molecular differences in the developmental process of adipogenesis, human preadipocytes were differentiated in vitro to mature adipocytes. Many genes in the apoptosis pathway are alternatively spliced. Our data demonstrates that during differentiation PKCδ, Bclx, and Caspase9 switch to their prosurvival splice variants along with an increase in Bcl2 expression when the cells terminally differentiate into mature adipocytes. Next we determined the expression pattern of these genes in obesity. Our data indicated high expression of PKCδVIII in adipose tissue of obese patient in different depots. We demonstrate a shift in the in vitro expression of these splice variants in differentiating preadipocytes derived from obese patients along with a decrease in adipogenesis markers. Hence, the programmed splicing of antiapoptotic proteins is a pivotal switch in differentiation that commits adipocytes to a prosurvival pathway. The expression pattern of these genes is dysregulated in obesity and may contribute to adipose tissue dysfunction.

scholarly journals Signal Transducer and Activator of Transcription 3 (STAT3) Suppresses STAT1/Interferon Signaling Pathway and Inflammation in Senescent Preadipocytes

Antioxidants ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 334
Author(s):  
Aisha Y. Madani ◽  
Yasser Majeed ◽  
Houari B. Abdesselem ◽  
Maha V. Agha ◽  
Muneera Vakayil ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Molecular Mechanisms ◽  
Adenosine Monophosphate ◽  
Human Adipose Tissue ◽  
Premature Aging ◽  
Guanosine Monophosphate ◽  
Negative Regulator ◽  
Signal Transducer ◽  
Interferon Signaling

Obesity promotes premature aging and dysfunction of white adipose tissue (WAT) through the accumulation of cellular senescence. The senescent cells burden in WAT has been linked to inflammation, insulin-resistance (IR), and type 2 diabetes (T2D). There is limited knowledge about molecular mechanisms that sustain inflammation in obese states. Here, we describe a robust and physiologically relevant in vitro system to trigger senescence in mouse 3T3-L1 preadipocytes. By employing transcriptomics analyses, we discovered up-regulation of key pro-inflammatory molecules and activation of interferon/signal transducer and activator of transcription (STAT)1/3 signaling in senescent preadipocytes, and expression of downstream targets was induced in epididymal WAT of obese mice, and obese human adipose tissue. To test the relevance of STAT1/3 signaling to preadipocyte senescence, we used Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR associated protein 9 (CRISPR/Cas9) technology to delete STAT1/3 and discovered that STAT1 promoted growth arrest and cooperated with cyclic Guanosine Monophosphate-Adenosine Monophosphate (GMP-AMP) synthase-stimulator of interferon genes (cGAS-STING) to drive the expression of interferon β (IFNβ), C-X-C motif chemokine ligand 10 (CXCL10), and interferon signaling-related genes. In contrast, we discovered that STAT3 was a negative regulator of STAT1/cGAS-STING signaling—it suppressed senescence and inflammation. These data provide insights into how STAT1/STAT3 signaling coordinates senescence and inflammation through functional interactions with the cGAS/STING pathway.

Age-associated features of the expression level of apoptosis markers in cardiomyocytes of patients with dilated cardiomyopathy

2022 ◽  
pp. 885-890
Author(s):  
К.П. Кравченко ◽  
К. Л. Козлов ◽  
А.О. Дробинцева ◽  
Д.С. Медведев ◽  
В.О. Полякова
Keyword(s):  
Cell Death ◽  
Dilated Cardiomyopathy ◽  
Molecular Mechanisms ◽  
Molecular Level ◽  
Confocal Laser ◽  
Control Group ◽  
Cellular Mechanisms ◽  
Apoptosis Pathway ◽  
Bax Protein

Для понимания патогенеза дилатационной кардиомиопатии (ДКМП) необходимо установить молекулярно-клеточные механизмы старения миокарда, в том числе связанные с программируемой клеточной гибелью, молекулярные механизмы которого практически не изучены. Цель работы - изучение маркеров апоптоза в кардиомиоцитах у пациентов с ДКМП in vitro. В работе использовали метод первичных диссоциированных клеточных культур и метод иммунофлюоресцентной конфокальной лазерной микроскопии. Для моделирования клеточного старения использовали клетки 3-го и 14-го пассажей, соответствующие «молодым» и «старым» культурам. На молекулярном уровне старение клеток кардиомиоцитов сопровождалось повышением экспрессии р16 в 2 раза по сравнению с «молодыми культурами» как в контрольной, так и в группе с ДКМП. Также установлено, что экспрессия р16 в культурах, взятых от пациентов с патологией, была в 2 раза выше, чем в аналогичных культурах от здоровых пациентов. Экспрессия р21 была повышена в группе с ДКМП по сравнению с контрольной группой, однако при старении культуры экспрессия p21 не изменялась, оставаясь на высоком уровне. Наиболее значимые различия были получены при сравнении экспрессии Bax в культуре клеток кардиомиоцитов из группы с ДКМП в «молодой» культуре с нормой - в 3,2 раза. Старение клеток миокарда на молекулярном уровне проявлялось в повышении экспрессии белка Baх, именно он является запускающим механизмом митохондриального пути апоптоза. Возможно, этот путь клеточной гибели является превалирующем при ДКМП. To understand the pathogenesis of dilated cardiomyopathy (DCMP), it is necessary to establish the molecular-cellular mechanisms of myocardial aging, including those associated with programmed cell death, the molecular mechanisms of which have not been practically studied. The aim of this work is to study markers of apoptosis in cardiomyocytes of patients with DCMP in vitro. We used the method of primary dissociated cell cultures and the method of immunofluorescence confocal laser microscopy. Cells of the 3 and 14 passages, corresponding to «young» and «old» cultures, were used to simulate cellular senescence. Results. At the molecular level, aging of cardiomyocyte cells was accompanied by a twofold increase in the expression of p16 compared to «young cultures» both in the control group and in the group with DCMP. It was also found that the expression of p16 in cultures taken from patients with pathology was 2 times higher than in similar cultures from healthy patients. The expression of p21 was increased in the group with DCMP compared to the control; however, with aging of the culture, the expression of p21 did not change, remaining at a significant level. The most significant differences were obtained when comparing the expression of Bax in the cell culture of cardiomyocytes from the group with DCMP in a «young» culture compared with the norm, 3,2 times. Aging of myocardial cells at the molecular level was manifested in an increase in the expression of the Bax protein, which is the triggering mechanism of the mitochondrial apoptosis pathway. It is possible that this pathway of cell death is prevalent in DCMP.

scholarly journals Hypoxia Alters the Expression of Dipeptidyl Peptidase 4 and Induces Developmental Remodeling of Human Preadipocytes

2016 ◽  
Vol 2016 ◽  
pp. 1-9 ◽  
Author(s):  
Helena H. Chowdhury ◽  
Jelena Velebit ◽  
Nataša Radić ◽  
Vito Frančič ◽  
Marko Kreft ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Transmembrane Protein ◽  
Human Adipose Tissue ◽  
Dipeptidyl Peptidase ◽  
Dipeptidyl Peptidase 4 ◽  
Hypoxic Conditions ◽  
Human Preadipocytes ◽  
Developmental Remodeling

Dipeptidyl peptidase 4 (DPP4), a transmembrane protein, has been identified in human adipose tissue and is considered to be associated with obesity-related type 2 diabetes. Since adipose tissue is relatively hypoxic in obese participants, we investigated the expression of DPP4 in human preadipocytes (hPA) and adipocytes in hypoxia, during differentiation and upon insulin stimulation. The results show that DPP4 is abundantly expressed in hPA but very sparsely in adipocytes. During differentiationin vitro, the expression of DPP4 in hPA is reduced on the addition of differentiation medium, indicating that this protein can be hPA marker. Long term hypoxia altered the expression of DPP4 in hPA. Inin vitrohypoxic conditions the protease activity of shed DPP4 is reduced; however, in the presence of insulin, the increase in DPP4 expression is potentiated by hypoxia.

scholarly journals Perspectives for Future Use of Extracellular Vesicles from Umbilical Cord- and Adipose Tissue-Derived Mesenchymal Stem/Stromal Cells in Regenerative Therapies—Synthetic Review

2020 ◽  
Vol 21 (3) ◽  
pp. 799 ◽  
Author(s):  
Joanna Lelek ◽  
Ewa K. Zuba-Surma
Keyword(s):  
Adipose Tissue ◽  
Umbilical Cord ◽  
Extracellular Vesicles ◽  
Stromal Cells ◽  
Tissue Repair ◽  
Molecular Mechanisms ◽  
Skin Diseases ◽  
Differentiation Potential

Mesenchymal stem/ stromal cells (MSCs) represent progenitor cells of various origin with multiple differentiation potential, representing the most studied population of stem cells in both in vivo pre-clinical and clinical studies. MSCs may be found in many tissue sources including extensively studied adipose tissue (ADSCs) and umbilical cord Wharton’s jelly (UC-MSCs). Most of sanative effects of MSCs are due to their paracrine activity, which includes also release of extracellular vesicles (EVs). EVs are small, round cellular derivatives carrying lipids, proteins, and nucleic acids including various classes of RNAs. Due to several advantages of EVs when compare to their parental cells, MSC-derived EVs are currently drawing attention of several laboratories as potential new tools in tissue repair. This review focuses on pro-regenerative properties of EVs derived from ADSCs and UC-MSCs. We provide a synthetic summary of research conducted in vitro and in vivo by employing animal models and within initial clinical trials focusing on neurological, cardiovascular, liver, kidney, and skin diseases. The summarized studies provide encouraging evidence about MSC-EVs pro-regenerative capacity in various models of diseases, mediated by several mechanisms. Although, direct molecular mechanisms of MSC-EV action are still under investigation, the current growing data strongly indicates their potential future usefulness for tissue repair.

scholarly journals Structural analysis of the DNA-binding domain of alternatively spliced steroid receptors

2002 ◽  
Vol 173 (3) ◽  
pp. 429-436 ◽  
Author(s):  
L Wickert ◽  
J Selbig
Keyword(s):  
Amino Acid ◽  
Dna Binding ◽  
Splice Variants ◽  
Steroid Receptor ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Amino Acid Residues ◽  
Receptor Function ◽  
Alternatively Spliced

We have generated 24 DNA-binding domain structure models of alternatively spliced or mutated steroid receptor variants by homology-based modeling. Members of the steroid receptor family dispose of a DNA-binding domain which is built from two zinc fingers with a preserved sequence homology of about 90%. Data from crystallographic analysis of the glucocorticoid receptor DNA-binding domain are therefore appropriate to serve as a template structure. We inserted or deleted amino acid residues in order to study the structural details of the glucocorticoid, mineralocorticoid, and androgen receptor splice variants. The receptor variants are created by QUANTA- and MODELLER-based modeling. Subsequently, the resulting energy-minimized models were compared with each other and with the wild-type receptor respectively. A prediction for the receptor function based mainly on the preservation or destruction of secondary structures has been carried out. The simulations showed that amino acid insertions of one, four or nine additional residues of existing steroid receptor splice variants were tolerated without destruction of the secondary structure. In contrast, a deletion of four amino acids at the splice site junction leads to modifications in the secondary structure of the DNA-recognition helix which apparently disturb the receptor-DNA interaction. Furthermore, an insertion of 23 amino acid residues between the zinc finger of the androgen receptor leads to a large loop with an additional alpha-helical structure which seems to disconnect a specific contact from its hormone response element. Thereafter, the prediction of receptor function based on the molecular models was compared with the available experimental results from the in vitro function tests. We obtained a close correspondence between the molecular modeling-based predictions and the conclusions of receptor function drawn from in vitro studies.

scholarly journals The Suv420h histone methyltransferases regulate PPAR-γ and energy expenditure in response to environmental stimuli

2019 ◽  
Vol 5 (4) ◽  
pp. eaav1472 ◽  
Author(s):  
Simona Pedrotti ◽  
Roberta Caccia ◽  
Maria Victoria Neguembor ◽  
Jose Manuel Garcia-Manteiga ◽  
Giulia Ferri ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Molecular Mechanisms ◽  
Target Genes ◽  
Ex Vivo ◽  
Peroxisome Proliferator ◽  
Histone Methyltransferases ◽  
Environmental Stimuli ◽  
Ppar Γ ◽  
Brown Adipose

Obesity and its associated metabolic abnormalities have become a global emergency with considerable morbidity and mortality. Epidemiologic and animal model data suggest an epigenetic contribution to obesity. Nevertheless, the cellular and molecular mechanisms through which epigenetics contributes to the development of obesity remain to be elucidated. Suv420h1 and Suv420h2 are histone methyltransferases responsible for chromatin compaction and gene repression. Through in vivo, ex vivo, and in vitro studies, we found that Suv420h1 and Suv420h2 respond to environmental stimuli and regulate metabolism by down-regulating peroxisome proliferator–activated receptor gamma (PPAR-γ), a master transcriptional regulator of lipid storage and glucose metabolism. Accordingly, mice lacking Suv420h proteins activate PPAR-γ target genes in brown adipose tissue to increase mitochondria respiration, improve glucose tolerance, and reduce adipose tissue to fight obesity. We conclude that Suv420h proteins are key epigenetic regulators of PPAR-γ and the pathways controlling metabolism and weight balance in response to environmental stimuli.

From mice to men – mouse models in obesity research: What can we learn?

2013 ◽  
Vol 110 (10) ◽  
pp. 634-640 ◽  
Author(s):  
Ilse Scroyen ◽  
Bianca Hemmeryckx ◽  
H. Roger Lijnen
Keyword(s):  
Adipose Tissue ◽  
Mouse Models ◽  
Developmental Process ◽  
Proteolytic System ◽  
Extracellular Matrix Remodelling ◽  
Adipose Tissue Development ◽  
Experimental Mouse

summaryObesity has become a world-wide epidemic and is associated with diseases such as diabetes, dyslipidaemia, cardiovascular disease and certain types of cancers. Understanding the adipose tissue developmental process, involving adipogenesis, angiogenesis and extracellular matrix remodelling, is therefore crucial to reveal the pathobiology of obesity. Experimental mouse models are extensively used to gain new insights into these processes and to evaluate the role of new key players, in particular proteolytic system components, in adipose tissue development and obesity. In this paper, we will review available in vitro and in vivo murine models of obesity and discuss their value in understanding the mechanisms contributing to obesity.

scholarly journals HAND2 is a novel obesity-linked adipogenic transcription factor regulated by glucocorticoid signalling

Diabetologia ◽  
2021 ◽  
Author(s):  
Maude Giroud ◽  
Foivos-Filippos Tsokanos ◽  
Giorgio Caratti ◽  
Stefan Kotschi ◽  
Sajjad Khani ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Adipose Tissue ◽  
Mouse Model ◽  
Molecular Mechanisms ◽  
Adipocyte Differentiation ◽  
Gene Clusters ◽  
Data Availability ◽  
Primary Mouse

Abstract Aims/hypothesis Adipocytes are critical cornerstones of energy metabolism. While obesity-induced adipocyte dysfunction is associated with insulin resistance and systemic metabolic disturbances, adipogenesis, the formation of new adipocytes and healthy adipose tissue expansion are associated with metabolic benefits. Understanding the molecular mechanisms governing adipogenesis is of great clinical potential to efficiently restore metabolic health in obesity. Here we investigate the role of heart and neural crest derivatives-expressed 2 (HAND2) in adipogenesis. Methods Human white adipose tissue (WAT) was collected from two cross-sectional studies of 318 and 96 individuals. In vitro, for mechanistic experiments we used primary adipocytes from humans and mice as well as human multipotent adipose-derived stem (hMADS) cells. Gene silencing was performed using siRNA or genetic inactivation in primary adipocytes from loxP and or tamoxifen-inducible Cre-ERT2 mouse models with Cre-encoding mRNA or tamoxifen, respectively. Adipogenesis and adipocyte metabolism were measured by Oil Red O staining, quantitative PCR (qPCR), microarray, glucose uptake assay, western blot and lipolysis assay. A combinatorial RNA sequencing (RNAseq) and ChIP qPCR approach was used to identify target genes regulated by HAND2. In vivo, we created a conditional adipocyte Hand2 deletion mouse model using Cre under control of the Adipoq promoter (Hand2AdipoqCre) and performed a large panel of metabolic tests. Results We found that HAND2 is an obesity-linked white adipocyte transcription factor regulated by glucocorticoids that was necessary but insufficient for adipocyte differentiation in vitro. In a large cohort of humans, WAT HAND2 expression was correlated to BMI. The HAND2 gene was enriched in white adipocytes compared with brown, induced early in differentiation and responded to dexamethasone (DEX), a typical glucocorticoid receptor (GR, encoded by NR3C1) agonist. Silencing of NR3C1 in hMADS cells or deletion of GR in a transgenic conditional mouse model results in diminished HAND2 expression, establishing that adipocyte HAND2 is regulated by glucocorticoids via GR in vitro and in vivo. Furthermore, we identified gene clusters indirectly regulated by the GR–HAND2 pathway. Interestingly, silencing of HAND2 impaired adipocyte differentiation in hMADS and primary mouse adipocytes. However, a conditional adipocyte Hand2 deletion mouse model using Cre under control of the Adipoq promoter did not mirror these effects on adipose tissue differentiation, indicating that HAND2 was required at stages prior to Adipoq expression. Conclusions/interpretation In summary, our study identifies HAND2 as a novel obesity-linked adipocyte transcription factor, highlighting new mechanisms of GR-dependent adipogenesis in humans and mice. Data availability Array data have been submitted to the GEO database at NCBI (GSE148699). Graphical abstract

Differences in the translation efficiency and mRNA stability mediated by 5′-UTR splice variants of human SP-A1 and SP-A2 genes

2005 ◽  
Vol 289 (3) ◽  
pp. L497-L508 ◽  
Author(s):  
Guirong Wang ◽  
Xiaoxuan Guo ◽  
Joanna Floros
Keyword(s):  
Gene Expression ◽  
Mrna Stability ◽  
Mrna Decay ◽  
Molecular Mechanisms ◽  
Luciferase Activity ◽  
Splice Variants ◽  
Translational Efficiency ◽  
Translation Efficiency ◽  
Control Vector

Surfactant protein A (SP-A) plays an important role in host defense, modulation of inflammatory processes, and surfactant-related functions of the lung. The human SP-A (hSP-A) locus consists of two functional genes, SP-A1 and SP-A2. Several hSP-A 5′-untranslated region (UTR) splice variants for each gene have been characterized and shown to be translated in vitro and in vivo. In this report, we investigated the role of hSP-A 5′-UTR splice variants on SP-A production and molecular mechanisms involved. We used in vitro transient expression of hSP-A 5′-UTR constructs containing luciferase as the reporter gene and quantitative real-time PCR to study hSP-A 5′-UTR-mediated gene expression. We found that 1) the four (A′D′, ABD, AB′D′, and A′CD′) 5′-UTR splice variants under study enhanced gene expression, by increasing luciferase activity from 2.5- to 19.5-fold and luciferase mRNA from 4.3- to 8.8-fold compared with the control vector that lacked hSP-A 5′-UTR; 2) all four 5′-UTR splice variants studied regulated mRNA stability. The ABD variant exhibited the lowest rate of mRNA decay compared with the other three constructs (A′D′, AB′D′, and A′CD′). These three constructs also exhibited significantly lower rate of mRNA decay compared with the control vector; 3) based on the indexes of translational efficiency (luciferase activity/mRNA), ABD and AB′D′ exhibited higher translational efficiency compared with the control vector, whereas the translational efficiency of each A′D′ and A′CD′ was lower than that of the control vector. These findings indicate that the hSP-A 5′-UTR splice variants play an important role in both SP-A translation and mRNA stability.

scholarly journals ANTXR1 Facilitates Tumor Growth in Glioma via Deactivating MAPK Signaling Pathway

2020 ◽  
Author(s):  
Aijun Liang ◽  
Chaoyang Zhou ◽  
Jianzhong Zhang ◽  
Jingxing Leng ◽  
Bin Xi ◽  
...  
Keyword(s):  
Cell Growth ◽  
Glioma Cell ◽  
Molecular Mechanisms ◽  
Mapk Signaling ◽  
Tumor Promoter ◽  
Normal Brain ◽  
Apoptosis Pathway ◽  
Xenograft Models

Abstract Background: Gliomas are commonly known as primary brain tumors and associated with frequent recurrence and an unsatisfactory prognosis in despite of extensive research in the underlying molecular mechanisms. In this study, we aimed to examine the role of ANTXR1 in glioma tumorigenesis and explore its downstream regulatory mechanism. Methods: We detected overexpression of ANTXR1 in glioma cell lines (SHG-44 and U251) in comparison with those in normal brain tissues.Result: Glioma cell growth and migratory ability were dramatically impaired as a result of silencing ANTXR1 by shANTXR1 lentivirus. ANTXR1 blockade also accelerated cell apoptosis since ANTXR1 held back apoptosis via targeting G2 phrase during cell mitosis. In vivo xenograft models verified in vitro findings above that the solid tumors stripped from mice were much lighter and smaller after depletion of ANTXR1 than controls. We also mechanically probed the downstream pathways and disclosed that overexpression of ANTXR1 abrogated the levels of MAKP9 and apoptisis-related protein HTRA2, but augmenting CCND1 and CDK6 levels in glioma cells. Our findings allow us to demonstrate that ANTXR1 acts as a tumor promoter in glioma induction through attenuating MAPK9-mediated gene transcription and HTRA2-induced apoptisis but intensifying CCND1-mediated proliferation. Conclusion: Together, we declare that ANTXR1 plays an indispensable role in glioma tumorigenesis via deactivating MAPK signaling and apoptosis pathway but activating PI3K/AKT-mediated cell growth. Our study provide a valuable clue to targeting ANTXR1 as a molecular target and a promising anticancer agent in glioma clinical therapeutics.

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