Modifications in routine protocol of RNA isolation can improve quality of RNA purified from adipocytes

Sinitsky, M. Yu.; Матвеева, В. Г.; Asanov, Maxim А.; Понасенко, А. В.

doi:10.1016/j.ab.2017.12.020

Cited by 11 publications

(8 citation statements)

References 10 publications

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“…Before RNA extraction, all work surfaces and laboratory equipment were treated with RNase Zap ™ RNase Decontamination Solution (Invitrogen, Carlsbad, CA, USA). Total RNA purification from isolated adipocytes was performed using the commercial RNeasy® Plus Universal Mini Kit (Qiagen, Hilden, Germany) according to the manufacturer's protocol with some modifications described previously (17). Briefly, not more than 5 × 10 5 cells were washed with phosphate-buffered saline (PBS) and homogenized with 750 μL of QIAzol™ Lysis Reagent (Qiagen).…”

Section: Methodsmentioning

confidence: 99%

Adipocytes Directly Affect Coronary Artery Disease Pathogenesis via Induction of Adipokine and Cytokine Imbalances

et al. 2019

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This study aimed to investigate the adipokine and cytokine profiles of adipocytes from epicardial and subcutaneous adipose tissues in interconnection with the visceral adipose tissue area and the biochemical and clinical characteristics of patients with coronary artery disease. We assessed 84 patients with coronary artery disease (65 men, 19 women) and divided them into two groups based on the presence of visceral obesity. We sampled epicardial and subcutaneous adipose tissues from the patients with visceral obesity. We then cultured the adipocytes and evaluated their adipokine profiles and pro-inflammatory activity. Results show that the mRNA expression of adiponectin in cultures of epicardial adipocytes from patients with and without visceral obesity was lower than that in subcutaneous adipocytes. Moreover, adiponectin mRNA expression in cultures of subcutaneous and epicardial adipocytes from patients with visceral obesity was lower than that in patients without obesity. For leptin, the reverse pattern was observed, with expression higher in cultures of epicardial adipocytes than in subcutaneous adipocytes and higher in epicardial adipocytes from patients with visceral obesity than in those from subjects without visceral obesity. In addition, in epicardial adipocytes, increased expression of proinflammatory cytokine genes (IL6, TNF) was observed compared with that in subcutaneous adipocytes. In contrast, expression of IL10 was higher in cultures of subcutaneous adipocytes than in epicardial adipocytes. The epicardial adipose tissue area was associated with the presence of higher levels of leptin and TNF-α within adipocytes and serum, increased lipid and carbohydrate metabolism. Coronary artery disease, in the context of the status of epicardial adipocytes, can be characterized as “metabolic inflammation,” suggesting the direct involvement of adipocytes in pathogenesis through the development of adipokine imbalances and activation of proinflammatory processes.

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Section: Methodsmentioning

confidence: 99%

Adipocytes Directly Affect Coronary Artery Disease Pathogenesis via Induction of Adipokine and Cytokine Imbalances

et al. 2019

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“…Adipocyte RNA was isolated by following the modified steps from the routine RNeasy Plus Universal Mini Kit protocol (Qiagen, Hilden, Germany) [ 24 ]. RNA was reverse transcribed by random primers and quantitative reverse transcription PCR was performed with SYBR Green master mix reagent (Applied Biosystems, Foster City, CA, USA).…”

Section: Methodsmentioning

confidence: 99%

Disruption of adipocyte YAP improves glucose homeostasis in mice and decreases adipose tissue fibrosis

Han

Aslam

Misra

et al. 2022

Molecular Metabolism

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“…Total RNA was isolated from adipocytes using the commercial RNeasy ® Plus Uni-versal Mini Kit (Qiagen, Hilden, Germany), according to the manufacturer’s instructions with slight modifications, as described previously ( 13 ). The quantity and quality of puri-fied RNA were assessed using a NanoDrop 2000 Spectrophotometer (Thermo Fisher Scientific) by measuring the light absorbance at 280 nm, 260 nm, and 230 nm and calcu-lating the 260/280 (A260/280) and 260/230 (A260/230) ratios.…”

Section: Methodsmentioning

confidence: 99%

Characteristics of adipocytokine expression by local fat depots of the heart: Relationship with the main risk factors for cardio-vascular diseases

Груздева

Dyleva²,

Белик³

et al. 2022

Front. Endocrinol.

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In our study we investigated the relationships between adipocytokines in adipose tissue (AT) and cardiovascular disease (CVD) risk factors; (2) Methods: fat tissue biopsies were obtained from 134 patients with stable CAD undergoing coronary artery bypass grafting and 120 patients undergoing aortic or mitral valve replacement. Adipocytes were isolated from subcutaneous (SAT), epicardial (EAT), and perivascular AT (PVAT) samples, and cultured for 24 h, after which gene expression of adipocytokines in the culture medium was determined; (3) Results: men showed reduced ADIPOQ expression in EAT and PVAT, LEP expression in PVAT, and LEPR expression in SAT and PVAT compared to women. Men also exhibited higher SAT and lower PVAT IL6 than women. Meanwhile, dyslipidemia associated with decreased ADIPOQ expression in EAT and PVAT, LEPR in EAT, and IL6 in PVAT. Arterial hypertension (AH) associated with low EAT and PVAT ADIPOQ, and high EAT LEP, SAT, as well as PVAT LEPR, and IL6 in SAT and EAT. ADIPOQ expression decreased with increased AH duration over 20 years against an increased LEP background in ATs. Smoking increased ADIPOQ expression in all ATs and increased LEP in SAT and EAT, however, decreased LEPR in PVAT. Patients 51–59 years old exhibited the highest EAT and PVAT LEP, IL-6, and LEPR expression compared to other age groups; (4) Conclusions: decreased EAT ADIPOQ expression against an increased pro-inflammatory IL6 background may increase atherogenesis and contribute to CAD progression in combination with risk factors including male sex, dyslipidemia, and AH.

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Modifications in routine protocol of RNA isolation can improve quality of RNA purified from adipocytes

Cited by 11 publications

References 10 publications

Adipocytes Directly Affect Coronary Artery Disease Pathogenesis via Induction of Adipokine and Cytokine Imbalances

Adipocytes Directly Affect Coronary Artery Disease Pathogenesis via Induction of Adipokine and Cytokine Imbalances

Disruption of adipocyte YAP improves glucose homeostasis in mice and decreases adipose tissue fibrosis

Characteristics of adipocytokine expression by local fat depots of the heart: Relationship with the main risk factors for cardio-vascular diseases

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