Ursolic Acid Attenuates Diabetic Mesangial Cell Injury through the Up-Regulation of Autophagy via miRNA-21/PTEN/Akt/mTOR Suppression

ObjectiveTo investigate the effect of ursolic acid on autophagy mediated through the miRNA-21-targeted phosphoinositide 3 kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) pathway in rat mesangial cells cultured under high glucose (HG) conditions.MethodsRat glomerular mesangial cells were cultured under normal glucose, HG, HG with the PI3K inhibitor {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002 or HG with ursolic acid conditions. Cell proliferation and hypertrophy were assayed using an MTT assay and the ratio of total protein to cell number, respectively. The miRNA-21 expression was detected using RT-qPCR. The expression of phosphatase and tensin homolog (PTEN)/AKT/mTOR signaling signatures, autophagy-associated protein and collagen I was detected by western blotting and RT-qPCR. Autophagosomes were observed using electron microscopy.ResultsCompared with mesangial cells cultured under normal glucose conditions, the cells exposed to HG showed up-regulated miRNA-21 expression, down-regulated PTEN protein and mRNA expression, up-regulated p85PI3K, pAkt, pmTOR, p62/SQSTMI, and collagen I expression and down-regulated LC3II expression. Ursolic acid and {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002 inhibited HG-induced mesangial cell hypertrophy and proliferation, down-regulated p85PI3K, pAkt, pmTOR, p62/SQSTMI, and collagen I expression and up-regulated LC3II expression. However, {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002 did not affect the expression of miRNA-21 and PTEN. Ursolic acid down-regulated miRNA-21 expression and up-regulated PTEN protein and mRNA expression.ConclusionsUrsolic acid inhibits the glucose-induced up-regulation of mesangial cell miRNA-21 expression, up-regulates PTEN expression, inhibits the activation of PI3K/Akt/mTOR signaling pathway, and enhances autophagy to reduce the accumulation of the extracellular matrix and ameliorate cell hypertrophy and proliferation.     

LncRNAs Expression Signatures of Renal Clear Cell Carcinoma Revealed by Microarray

Background

Long noncoding RNAs (lncRNAs) are an important class of pervasive genes involved in a variety of biological functions. They are aberrantly expressed in many types of cancers. In this study, we described lncRNAs profiles in 6 pairs of human renal clear cell carcinoma (RCCC) and the corresponding adjacent nontumorous tissues (NT) by microarray.

Methodology/Principal Findings

With abundant and varied probes accounting 33,045 LncRNAs in our microarray, the number of lncRNAs that expressed at a certain level could be detected is 17157. From the data we found there were thousands of lncRNAs that differentially expressed (≥2 fold-change) in RCCC tissues compared with NT and 916 lncRNAs differentially expressed in five or more of six RCCC samples. Compared with NT, many lncRNAs were significantly up-regulated or down-regulated in RCCC. Our data showed that down-regulated lncRNAs were more common than up-regulated ones. ENST00000456816, {"type":"entrez-nucleotide","attrs":{"text":"X91348","term_id":"1418768","term_text":"X91348"}}X91348, {"type":"entrez-nucleotide","attrs":{"text":"BC029135","term_id":"23271405","term_text":"BC029135"}}BC029135, {"type":"entrez-nucleotide","attrs":{"text":"NR_024418","term_id":"212286160","term_text":"NR_024418"}}NR_024418 were evaluated by qPCR in sixty-three pairs of RCCC and NT samples. The four lncRNAs were aberrantly expressed in RCCC compared with matched histologically normal renal tissues.

Conclusions/Significance

Our study is the first one to determine genome-wide lncRNAs expression patterns in RCCC by microarray. The results displayed that clusters of lncRNAs were aberrantly expressed in RCCC compared with NT samples, which revealed that lncRNAs differentially expressed in tumor tissues and normal tissues may exert a partial or key role in tumor development. Taken together, this study may provide potential targets for future treatment of RCCC and novel insights into cancer biology.     

Expression Profiles of Long Noncoding RNAs and Messenger RNAs in Mn-Exposed Hippocampal Neurons of Sprague–Dawley Rats Ascertained by Microarray: Implications for Mn-Induced Neurotoxicity

Manganese (Mn) is an essential trace element, while excessive expose may induce neurotoxicity. Recently, lncRNAs have been extensively studied and it has been confirmed that lncRNAs participate in neural functions and aberrantly expressed lncRNAs are involved in neurological diseases. However, the pathological effects of lncRNAs on Mn-induced neurotoxicity remain unclear. In this study, the expression profiles of lncRNAs and messenger RNAs (mRNAs) were identified in Mn-treated hippocampal neurons and control neurons via microarray. Bioinformatic methods and intersection analysis were also employed. Results indicated that 566, 1161, and 1474 lncRNAs meanwhile 1848, 3228, and 4022 mRNAs were aberrantly expressed in low, intermediate, and high Mn-exposed groups compared with the control group, respectively. Go analysis determined that differentially expressed mRNAs were targeted to biological processes, cellular components, and molecular functions. Pathway analysis indicated that these mRNAs were enriched in insulin secretion, cell cycle, and DNA replication. Intersection analysis denominated that 135 lncRNAs and 373 mRNAs were consistently up-regulated while 150 lncRNAs and 560 mRNAs were consistently down-regulated. Meanwhile, lncRNA {"type":"entrez-nucleotide","attrs":{"text":"BC079195","term_id":"50926238"}}BC079195 was significantly up-regulated while lncRNAs uc.229- and {"type":"entrez-nucleotide","attrs":{"text":"BC089928","term_id":"59808777"}}BC089928 were significantly down-regulated in three comparison groups. The relative expression levels of 3 lncRNAs and 4 mRNAs were validated through qRT-PCR. To the best of our knowledge, this study is the first to identify the expression patterns of lncRNAs and mRNAs in hippocampal neurons of Sprague–Dawley rats. The results may provide evidence on underlying mechanisms of Mn-induced neurotoxicity, and aberrantly expressed lncRNAs/mRNAs may be useful in further investigations to detect early symptoms of Mn-induced neuropsychiatric disorders in the central nervous system.     

Differential regulation of microRNAs in response to osmotic,salt and cold stresses in wheat

MicroRNAs (miRNAs) are tiny non-coding regulatory molecules that modulate plant’s gene expression either by cleaving or repressing their mRNA targets. To unravel the plant actions in response to various environmental factors, identification of stress related miRNAs is essential. For understanding the regulatory behaviour of various abiotic stresses and miRNAs in wheat genotype C-306, we examined expression profile of selected conserved miRNAs viz. miR159, miR164, miR168, miR172, miR393, miR397, miR529 and miR1029 tangled in adapting osmotic, salt and cold stresses. The investigation revealed that two miRNAs (miR168, miR397) were down-regulated and miR172 was up-regulated under all the stress conditions. However, miR164 and miR1029 were up-regulated under cold and osmotic stresses in contrast to salt stress. miR529 responded to cold alone and does not change under osmotic and salt stress. miR393 showed up-regulation under osmotic and salt, and down-regulation under cold stress indicating auxin based differential cold response. Variation in expression level of studied miRNAs in presence of target genes delivers a likely elucidation of miRNAs based abiotic stress regulation. In addition, we reported new stress induced miRNAs Ta-miR855 using computational approach. Results revealed first documentation that miR855 is regulated by salinity stress in wheat. These findings indicate that diverse miRNAs were responsive to osmotic, salt and cold stress and could function in wheat response to abiotic stresses.     

Upregulation of the novel lncRNA U731166 is associated with migration,invasion and vemurafenib resistance in melanoma

Our previous work using a melanoma progression model composed of melanocytic cells (melanocytes, primary and metastatic melanoma samples) demonstrated various deregulated genes, including a few known lncRNAs. Further analysis was conducted to discover novel lncRNAs associated with melanoma, and candidates were prioritized for their potential association with invasiveness or other metastasis‐related processes. In this sense, we found the intergenic lncRNA {"type":"entrez-nucleotide","attrs":{"text":"U73166","term_id":"1613889","term_text":"U73166"}}U73166 (ENSG00000230454) and decided to explore its effects in melanoma. For that, we silenced the lncRNA {"type":"entrez-nucleotide","attrs":{"text":"U73166","term_id":"1613889","term_text":"U73166"}}U73166 expression using shRNAs in a melanoma cell line. Next, we experimentally investigated its functions and found that migration and invasion had significantly decreased in knockdown cells, indicating an essential association of lncRNA {"type":"entrez-nucleotide","attrs":{"text":"U73166","term_id":"1613889","term_text":"U73166"}}U73166 for cancer processes. Additionally, using naïve and vemurafenib‐resistant cell lines and data from a patient before and after resistance, we found that vemurafenib‐resistant samples had a higher expression of lncRNA {"type":"entrez-nucleotide","attrs":{"text":"U73166","term_id":"1613889","term_text":"U73166"}}U73166. Also, we retrieved data from the literature that indicates lncRNA {"type":"entrez-nucleotide","attrs":{"text":"U73166","term_id":"1613889","term_text":"U73166"}}U73166 may act as a mediator of RNA processing and cell invasion, probably inducing a more aggressive phenotype. Therefore, our results suggest a relevant role of lncRNA {"type":"entrez-nucleotide","attrs":{"text":"U73166","term_id":"1613889","term_text":"U73166"}}U73166 in metastasis development. We also pointed herein the lncRNA {"type":"entrez-nucleotide","attrs":{"text":"U73166","term_id":"1613889","term_text":"U73166"}}U73166 as a new possible biomarker or target to help overcome clinical vemurafenib resistance.     

Regulation of Gene Expression by PI3K in Mouse Growth Plate Chondrocytes

Background

Endochondral ossification, the process through which long bones are formed, involves chondrocyte proliferation and hypertrophic differentiation in the cartilage growth plate. In a previous publication we showed that pharmacological inhibition of the PI3K signaling pathway results in reduced endochondral bone growth, and in particular, shortening of the hypertrophic zone in a tibia organ culture system. In this current study we aimed to investigate targets of the PI3K signaling pathway in hypertrophic chondrocytes.

Methodology/Principal Findings

Through the intersection of two different microarray analyses methods (classical single gene analysis and GSEA) and two different chondrocyte differentiation systems (primary chondrocytes treated with a pharmacological inhibitor of PI3K and microdissected growth plates), we were able to identify a high number of genes grouped in GSEA functional categories regulated by the PI3K signaling pathway. Genes such as Phlda2 and F13a1 were down-regulated upon PI3K inhibition and showed increased expression in the hypertrophic zone compared to the proliferative/resting zone of the growth plate. In contrast, other genes including Nr4a1 and Adamts5 were up-regulated upon PI3K inhibition and showed reduced expression in the hypertrophic zone. Regulation of these genes by PI3K signaling was confirmed by quantitative RT-PCR. We focused on F13a1 as an interesting target because of its known role in chondrocyte hypertrophy and osteoarthritis. Mouse E15.5 tibiae cultured with {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002 (PI3K inhibitor) for 6 days showed decreased expression of factor XIIIa in the hypertrophic zone compared to control cultures.

Conclusions/Significance

Discovering targets of signaling pathways in hypertrophic chondrocytes could lead to targeted therapy in osteoarthritis and a better understanding of the cartilage environment for tissue engineering.     

Postnatal PPARδ Activation and Myostatin Inhibition Exert Distinct yet Complimentary Effects on the Metabolic Profile of Obese Insulin-Resistant Mice

Background

Interventions for T2DM have in part aimed to mimic exercise. Here, we have compared the independent and combined effects of a PPARδ agonist and endurance training mimetic ({"type":"entrez-nucleotide","attrs":{"text":"GW501516","term_id":"289075981","term_text":"GW501516"}}GW501516) and a myostatin antibody and resistance training mimetic (PF-879) on metabolic and performance outcomes in obese insulin resistant mice.

Methodology/Principal Findings

Male ob/ob mice were treated for 6 weeks with vehicle, {"type":"entrez-nucleotide","attrs":{"text":"GW501516","term_id":"289075981","term_text":"GW501516"}}GW501516, PF-879, or {"type":"entrez-nucleotide","attrs":{"text":"GW501516","term_id":"289075981","term_text":"GW501516"}}GW501516 in combination with PF-879. The effects of the interventions on body composition, glucose homeostasis, glucose tolerance, energy expenditure, exercise capacity and metabolic gene expression were compared at the end of study. {"type":"entrez-nucleotide","attrs":{"text":"GW501516","term_id":"289075981","term_text":"GW501516"}}GW501516 attenuated body weight and fat mass accumulation and increased the expression of genes of oxidative metabolism. In contrast, PF-879 increased body weight by driving muscle growth and altered the expression of genes involved in insulin signaling and glucose metabolism. Despite their differences, both interventions alone improved glucose homeostasis. Moreover, {"type":"entrez-nucleotide","attrs":{"text":"GW501516","term_id":"289075981","term_text":"GW501516"}}GW501516 more effectively improved serum lipids, and PF-879 uniquely increased energy expenditure, exercise capacity and adiponectin levels. When combined the robust effects of {"type":"entrez-nucleotide","attrs":{"text":"GW501516","term_id":"289075981","term_text":"GW501516"}}GW501516 and/or PF-879 on body weight, adiposity, muscle mass, glycemia, serum lipids, energy expenditure and exercise capacity were highly conserved.

Conclusions/Significance

The data, for the first time, demonstrate postnatal inhibition of myostatin not only promotes gains in muscle mass similar to resistance training,but improves metabolic homeostasis. In several instances, these effects were either distinct from or complimentary to those of {"type":"entrez-nucleotide","attrs":{"text":"GW501516","term_id":"289075981","term_text":"GW501516"}}GW501516. The data further suggest that strategies to increase muscle mass, and not necessarily oxidative capacity, may effectively counter insulin resistance and T2DM.     

月季‘绿萼’花器官发育相关microRNA的鉴定及分析

利用高通量测序技术,构建了中国古老月季‘绿萼’(Rosa chinensis ‘Viridiflora’)和‘月月粉’(R.chinensis‘Old Blush’)花蕾期的microRNA(miRNA)文库,并对其进行了测序和序列分析。结果显示,在‘绿萼’文库中,鉴定到已知的miRNA成熟体39个,miRNA前体42个;预测到新的miRNA成熟体56个,前体57个。在‘月月粉’文库中,鉴定到已知RNA成熟体39个,已知miRNA前体40个;预测到新的miRNA成熟体53个,前体57个。与‘月月粉’相比,‘绿萼’中差异表达的miRNA有31个,其中17个上调、14个下调。荧光定量PCR实验结果表明,miR156、miR398和miR535在2种月季的花蕾期表达上调,而miR167、miR172和miR396表达下调。进一步检测miR172和miR156在2种月季不同花器官中的表达差异,发现miR172在‘绿萼’的花瓣、雌、雄蕊中表达显著下调,提示miR172可能通过负调控其靶基因RcAP2的表达,在‘绿萼’花器官发育过程中起重要作用。