MicroRNAs are small and non-coding RNA molecules with the master role in regulation of gene expression at post-transcriptional/translational levels. Many methods have been developed for microRNA loss-of-function study, such as antisense inhibitors and sponges; however, the robustness, specificity, and stability of these traditional strategies are not highly satisfied. CRISPR/cas9 system is emerging as a novel genome editing tool in biology/medicine research, but its indication in microRNA research has not been studied exclusively. In this study, we clone CRISPR/cas9 constructs with single-guide RNAs specifically targeting biogenesis processing sites of selected microRNAs; and we find that CRISPR/cas9 can robustly and specifically reduce the expression of these microRNAs up to 96%. CRISPR/cas9 also shows an exclusive benefit in control of crossing off-target effect on microRNAs in the same family or with highly conserved sequences. More significantly, for the first time, we demonstrate the long term stability of microRNA knockdown phenotype by CRISPR/cas9 in both in vitro and in vivo models.
Genetic searches for tumor suppressors have recently linked small nucleolar RNA misregulations with tumorigenesis. In addition to their classically defined functions, several small nucleolar RNAs are now known to be processed into short microRNA-like fragments called small nucleolar RNA-derived RNAs. To determine if any small nucleolar RNA-derived RNAs contribute to breast malignancy, we recently performed a RNA-seq-based comparison of the small nucleolar RNA-derived RNAs of two breast cancer cell lines (MCF-7 and MDA-MB-231) and identified small nucleolar RNA-derived RNAs derived from 13 small nucleolar RNAs overexpressed in MDA-MB-231s. Importantly, we find that inhibiting the most differentially expressed of these small nucleolar RNA-derived RNAs (sdRNA-93) in MDA-MB-231 cells results primarily in a loss of invasiveness, whereas increased sdRNA-93 expression in either cell line conversely results in strikingly enhanced invasion. Excitingly, we recently determined sdRNA-93 expressions in small RNA-seq data corresponding to 116 patient tumors and normal breast controls, and while we find little sdRNA-93 expression in any of the controls and only sporadic expression in most subtypes, we find robust expression of sdRNA-93 in 92.8% of Luminal B Her2+tumors. Of note, our analyses also indicate that at least one of sdRNA-93’s endogenous roles is to regulate the expression of Pipox, a sarcosine metabolism-related protein whose expression significantly correlates with distinct molecular subtypes of breast cancer. We find sdRNA-93 can regulate the Pipox 3′UTR via standard reporter assays and that manipulating endogenous sdRNA-93 levels inversely correlates with altered Pipox expression. In summary, our results strongly indicate that sdRNA-93 expression actively contributes to the malignant phenotype of breast cancer through participating in microRNA-like regulation.
Objectives: Whether uric acid levels were associated with the progression of chronic kidney disease (CKD) remained controversial. This meta-analysis was aimed to assess the effect of lowering serum uric acid therapy on the progression of CKD to clarify the role of uric acid in the progression of CKD indirectly.Methods: Pubmed, Embase, the Cochrane library, CBM were searched for randomized controlled trials (RCTs) that assessed the efficiency of lowering serum uric acid therapy on the progression of CKD without language restriction. Summary estimates of weighted mean differences (WMDs) and relative risk (RR) were obtained by using random-effect or fixed-effect models. Sensitivity analyses were performed to identify the source of heterogeneity.Results: A total of 12 randomized controlled trials with 832 CKD participants were included in the analysis. Pooled estimate for eGFR was in favor of lowering serum uric acid therapy with a mean difference (MD) of 3.88 ml/min/1.73 m2, 95% CI 1.26–6.49 ml/min/1.73 m2, p = .004 and this was consistent with results for serum creatinine. The risk of worsening of kidney function or ESRD or death was significantly decreased in the treatment group compared to the control group (RR 0.39, 95% CI 0.28–0.52, p< .01).Conclusions: Uric acid-lowering therapy may be effective in retarding the progression of CKD. Further randomized controlled trials should be performed to confirm the effect of lowering serum uric acid therapy on the progression of CKD.
LncRNAs represent a large class of noncoding RNA molecules that have important functions and play key roles in a variety of human diseases. There is an urgent need to develop bioinformatics tools as to gain insight into lncRNAs. This study developed a sequence-based bioinformatics method, LncDisease, to predict the lncRNA-disease associations based on the crosstalk between lncRNAs and miRNAs. Using LncDisease, we predicted the lncRNAs associated with breast cancer and hypertension. The breast-cancer-associated lncRNAs were studied in two breast tumor cell lines, MCF-7 and MDA-MB-231. The qRT-PCR results showed that 11 (91.7%) of the 12 predicted lncRNAs could be validated in both breast cancer cell lines. The hypertension-associated lncRNAs were further evaluated in human vascular smooth muscle cells (VSMCs) stimulated with angiotensin II (Ang II). The qRT-PCR results showed that 3 (75.0%) of the 4 predicted lncRNAs could be validated in Ang II-treated human VSMCs. In addition, we predicted 6 diseases associated with the lncRNA GAS5 and validated 4 (66.7%) of them by literature mining. These results greatly support the specificity and efficacy of LncDisease in the study of lncRNAs in human diseases. The LncDisease software is freely available on the Software Page: http://www.cuilab.cn/.
Triptolide is a major active component of Tripterygium wilfordii Hook F, which exerts marked immunosuppressive, anti-inflammatory and podocyte-protective effects. In this study, the ability of triptolide to inhibit inflammation and attenuate podocyte injury was examined in a rat model of diabetic nephropathy (DN). Type II diabetic rats with DN were treated with triptolide at a dose of 100 μg.kg−1.day−1. Following 8 weeks of triptolide treatment, the urine albumin level, kidney weight/body weight and the number of cells positive for ED-1 (a marker for rat mononuclear macrophages) in the kidney were assessed. The effects of triptolide on podocyte injury and chronic inflammation were analyzed using quantitative polymerase chain reaction (qPCR), western blotting and immunohistochemistry. Following triptolide treatment, the albuminuria in the type II diabetic rats was significantly reduced. Furthermore, the glomerular hypertrophy and foot process effacement were improved, and there was a recovery of the slit diaphragm associated with nephrin and podocin expression. The inflammation in the kidneys was also attenuated. Furthermore, triptolide significantly reduced the expression of transforming growth factor-β1 and osteopontin, and the infiltration of ED-1-positive cells into the kidney. The results demonstrated that triptolide markedly attenuated albuminuria and podocyte injury in the rat model of DN, which may have been correlated with the inhibition of inflammation and macrophage infiltration in the kidneys.
The immune system plays an essential and central role in tumor cell differentiation, proliferation, angiogenesis, apoptosis, invasion, and metastasis. Over the past decade, cancer therapy has rapidly evolved from traditional approaches, such as surgery, chemotherapy, and radiotherapy, to revolutionary new treatment options with immunotherapy. This new era of cancer treatment options has now been clinically tested and applied to many forms of human malignancies, often with quite dramatic results. As we develop more effective combinations of cancer treatment, several agents have been recently investigated, putatively identified as anticancer agents, or immunostimulatory molecules. One such agent is metformin, originally developed as a fairly standard first-line therapy for patients with type-2 diabetes mellitus (T2DM). Given the underlying mechanisms of action, researchers began to examine the alternative functions and possible utility of metformin, finding that the cancer risk in patients with T2DM was reduced. It appears that metformin, at least in part, has an antitumor effect through activation of the 5' adenosine monophosphate-activated protein kinase (AMPK) signaling pathway. Moreover, numerous studies have demonstrated that metformin interferes with key immunopathological mechanisms that are involved in the pathological processes or associated with malignant progression. Such insights may shed light on further analyzing whether metformin enhances the effectiveness of the immunotherapy and overcomes the immunotherapy resistance in the patients. Herein, we provide a comprehensive review of the literature examining the impact of metformin upon the host immune system and cancer immunity.
BACKGROUND: Polymorphism of acyl coenzyme A: diacylglycerol acyltransferase 1 (DGAT1) affects the fat content of milk and meat in cattle. To investigate the effects of DGAT1 on sheep meat quality traits, the cDNA of DGAT1 was cloned. The association between single nucleotide polymorphisms of the DGAT1 gene and sheep meat quality traits was assessed.
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