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The impact of obesity upon bone metabolism is controversial since both beneficial or harmful effects have been reported. Bone remodeling is modulated by the central nervous system through cytokines, hormones and neuromodulators. The present study aimed to evaluate the effects evoked by bilateral retroperitoneal white adipose tissue (rWAT) denervation (Dnx) upon bone mineral metabolism and remodeling in an experimental model of obesity in rats. Male Wistar rats were fed during 18 weeks with high-fat diet (HFD) or standard diet (SD) as controls, and rWAT Dnx or Sham surgery was performed at the 14th week. Biochemical and hormonal parameters, bone histomorphometry, rWAT and hypothalamus protein and gene expression were analyzed. The HFD group presented decreased bone formation parameters, increased serum and bone leptin and FGF23, increased serum and hypothalamic neuropeptide Y (NPY) and decreased serum 1,25-dihydroxyvitamin D 3 and PTH. After rWAT Dnx, bone markers and histomorphometry showed restoration of bone formation, and serum and hypothalamic NPY decreased, without alteration in leptin levels. The present study shows that the denervation of rWAT improved bone formation in obese rats mediated by a preferential reduction in neurohormonal actions of NPY, emphasizing the relevance of the adipose tissue-brain-bone axis in the control of bone metabolism in obesity.

Reverse transcription-quantitative polymerase chain reaction (RT-PCR) is the gold standard technique for gene expression analysis, but the choice of quantitative reference genes (housekeeping genes, HKG) remains challenging. Identify the best HKG is essential for estimating the expression level of target genes. Therefore, the aim of this study was to determine the best HKG for an in vitro model with mouse mesangial cells (MMCs) stimulated with 5 ng/mL of TGF-β. Five candidates HKG were selected: Actb, Hprt, Gapdh, 18S and Ppia. After quantitative expression, the best combination of these genes was analyzed in silico using six software programs. To validate the results, the best genes were used to normalize the expression levels of fibronectin, vimentin and α-SMA. In silico analysis revealed that Ppia, Gapdh and 18S were the most stable genes between the groups. GenEX software and Spearman's correlation determined Ppia and Gapdh as the best HKG pair, and validation of the HKG by normalizing fibronectin, vimentin and α-SMA were consistent with results from the literature. Our results established the combination of Ppia and Gapdh as the best HKG pair for gene expression analysis by RT-PCR in this in vitro model using MMCs treated with TGF-β., (© 2022. The Author(s).)

Sepsis contributes to the high prevalence of acute kidney injury (AKI), which mainly occurs in hospitalized patients. The delay in AKI detection is a risk factor for death and chronicity; thus, early diagnosis is essential for initiating proper treatment strategies. Although serum creatinine is used as biomarker, it is increased in plasma serum creatinine only at late stages of AKI. MicroRNAs (miRNAs), a class of noncoding RNAs responsible for gene regulation, can be found in biological fluids within vesicles such as exosomes and may be promising tools for the early detection of AKI. We aimed to identify potential blood miRNAs that can be used as early biomarkers of sepsis-induced AKI in rats. Adult male Wistar rats received a single dose of lipopolysaccharide (LPS). The earliest significant increase in serum creatinine was detected 4 h after LPS administration. To evaluate whether miRNAs could act as early biomarkers, blood samples were collected before and 2 h after LPS infusion. Serum NGAL levels were used as a comparative marker. Serum miRNAs were derived from exosomes, and their expression were evaluated by the PCR array. miR-181a-5p and miR-23b-3p showed higher expression in LPS-treated rats than in the control animals ( p < 0.05). Bioinformatics analysis showed that both miRNAs target molecules associated with transcription factors that regulate genes related to proinflammatory cytokines. Considering that LPS activates transcription factors that lead to the production of proinflammatory cytokines, possible premature changes in the serum levels of miR-181a-5p and miR-23b-3p may be used to identify sepsis-induced AKI earlier., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Da-Silva, Anauate, Guirao, Novaes, Maquigussa and Boim.)

Gastric cancer (GC) is one of the leading types of fatal cancer worldwide. Epigenetic manipulation of cancer cells is a useful tool to better understand gene expression regulatory mechanisms and contributes to the discovery of novel biomarkers. Our research group recently reported a list of 83 genes that are potentially modulated by DNA methylation in GC cell lines. Herein, we further explored the regulation of one of these genes, LRRC37A2 , in clinical samples. LRRC37A2 expression was evaluated by RT-qPCR, and DNA methylation was studied using next-generation bisulphite sequencing in 36 GC and paired adjacent nonneoplastic tissue samples. We showed that both reduced LRRC37A2 mRNA levels and increased LRRC37A2 exon methylation were associated with undifferentiated and poorly differentiated tumours. Moreover, LRRC37A2 gene expression and methylation levels were inversely correlated at the +45 exon CpG site. We suggest that DNA hypermethylation may contribute to reducing LRRC37A2 expression in undifferentiated and poorly differentiated GC. Therefore, our results show how some genes may be useful to stratify patients who are more likely to benefit from epigenetic therapy. Abbreviations: AR: androgen receptor; 5-AZAdC: 5-aza-2'-deoxycytidine; B2M : beta-2-microglobulin; GAPDH : glyceraldehyde-3-phosphate dehydrogenase; GC: gastric cancer; GLM: general linear model; LRRC37A2 : leucine-rich repeat containing 37 member A2; SD: standard deviation; TFII-I: general transcription factor II-I; TSS: transcription start site; XBP1: X-box binding protein 1.

Autosomal Dominant Polycystic Kidney Disease (ADPKD) is the most common inherited renal disorder, characterized by renal cyst development leading to end-stage renal disease. Although the appropriate choice of suitable reference is critical for quantitative RNA analysis, no comparison of frequently used "housekeeping" genes is available. Here, we determined the validity of 7 candidate housekeeping genes (Actb, Actg1, B2m, Gapdh, Hprt, Pgam1 and Ppia) in kidney tissues from mouse models orthologous to ADPKD, including a cystic mice (CY) 10-12 weeks old (Pkd1 flox/flox :Nestin cre /Pkd1 flox/- :Nestin cre , n = 10) and non-cystic (NC) controls (Pkd1 flox / flox /Pkd1 flox / - , n = 10), Pkd1-haploinsufficient (HT) mice (Pkd1 +/- , n = 6) and wild-type (WT) controls (Pkd1 +/+ , n = 6) and a severely cystic (SC) mice 15 days old (Pkd1 V/V , n = 7) and their controls (CO, n = 5). Gene expression data were analyzed using six distinct statistical softwares. The estimation of the ideal number of genes suggested the use of Ppia alone as sufficient, although not ideal, to analyze groups altogether. Actb, Hprt and Ppia expression profiles were correlated in all samples. Ppia was identified as the most stable housekeeping gene, while Gapdh was the least stable for all kidney samples. Stat3 expression level was consistent with upregulation in SC compared to CO when normalized by Ppia expression. In conclusion, present findings identified Ppia as the best housekeeping gene for CY + NC and SC + CO groups, while Hprt was the best for the HT + WT group., (© 2021. The Author(s).)