Rt2 profiler pcr array mouse adipogenesis

The HFD mice with or without 400 mg/kg/BW/day of LMF-HSFx through oral gavage. After 16 weeks treatment, the mice were anesthetized by inhalant anesthesia with isoflurane before sacrifice. RNA was extracted from adipose tissues by using a Direct-zol RNA Miniprep Plus kit (Cat. No. R2073, Zymo Research, Irvine, CA, USA) and its quantity and quality were estimated by NanoDrop measurement (ThermoFisher, Waltham, MA, USA). The RT PCR processing was performed using a commercially available RT² Profiler™ PCR Array Mouse Adipogenesis (Cat. no. PAMM-049ZA, QIAGEN, Hilden, Germany) for genes of interest. Following, the fold change of target gene shows the ratio of treated group/control group and values > 1.5 or < 0.5 were considered significantly modified.

Total RNA was obtained from frozen tissues, as previously described [36 (link)], using RNeasy Lipid Tissue (Qiagen, Hilden, Germany), according to the manufacturer’s instructions. Gene expression analysis levels were determined using the relative expression method with the threshold crossing point (Ct-value), as described [26 (link),30 (link),37 (link)].
PCR arrays for the RT² Profiler™ PCR Array Mouse Adipogenesis (Qiagen, product no. 330,231 and Cat. No. PAMM-049Z) were performed following the manufacturers’ protocol. Gene expression levels were calculated using the ΔΔCt method after normalization to the housekeeping gene expression and determination of the fold change. Each GeneQuery™ plate contains eight controls, five target housekeeping genes (β-actin, GAPDH, LDHA, NONO, and PPIH), and genes encoding for pre-adipocyte cell markers, proliferation, differentiation and adipogenesis, lipid metabolism, and obesity. Gene expression is presented as the log10 of mean values (n = 3 in each group), as previously described [26 (link),40 (link),41 ] (https://CRAN.R-project.org/package=gplots, accessed on 4 March 2022).