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Background and Aims: We aimed to investigate the underlying mechanism of action of risankizumab, a monoclonal antibody targeting the IL-23 p19 subunit, previously reported to induce clinical and endoscopic remission in a randomised phase II study in patients with active Crohn's disease., Methods: Ileum and colon biopsies obtained at screening and Week 12 from a subgroup of patients [n = 106] in the risankizumab phase II study were analysed by transcriptome-wide RNA-Seq profiling. Univariate associations were assessed using linear modelling., Results: By Week 12, risankizumab significantly decreased [p < 0.005] the expression of 1880 and 765 genes in the colon [false-discovery rate = 0.02] and ileum [false-discovery rate = 0.05], respectively. These genes were associated with the IL-23/IL-17 axis, Th1 pathway, innate immunity, and tissue turnover. Colonic transcriptomic profiles following risankizumab treatment reflected the transcriptomic changes observed in patients achieving endoscopic response and remission at Week 12 and were significantly different from placebo [p < 0.005]. The colonic transcriptomic profile, significantly modulated by risankizumab at Week 12, was indicative of suppression of pathways associated with epithelial biology. Furthermore, pathways associated with Crohn's disease modulated by risankizumab treatment included second messenger-mediated signalling, immune response, lymphocyte and leucocyte activation, lymphocyte differentiation and cell-cell adhesion., Conclusions: Endoscopic remission and response observed with risankizumab in patients with active Crohn's disease was associated with significant transcriptomic changes in the colon, compared with placebo. Differentiated expression of genes associated with the IL-23/IL-17 axis was observed in the colon and ileum 12 weeks after risankizumab treatment.

Dipeptidyl peptidase 4 inhibitors and angiotensin II receptor blockers attenuate chronic kidney disease progression in experimental diabetic and non-diabetic nephropathy in a blood pressure and glucose independent manner, but the exact molecular mechanisms remain unclear. MicroRNAs (miRNAs) are short, non-coding RNA species that are important post-transcriptional regulators of gene expression and play an important role in the pathogenesis of nephropathy. miRNAs are present in urine in a remarkably stable form, packaged in extracellular vesicles. Here, we investigated linagliptin and telmisartan induced effects on renal and urinary exosomal miRNA expression in 5/6 nephrectomized rats. In the present study, renal miRNA profiling was conducted using the Nanostring nCounter technology and mRNA profiling using RNA sequencing from the following groups of rats: sham operated plus placebo; 5/6 nephrectomy plus placebo; 5/6 nephrectomy plus telmisartan; and 5/6 nephrectomy plus linagliptin. TaqMan Array miRNA Cards were used to evaluate which of the deregulated miRNAs in the kidney are present in urinary exosomes. In kidneys from 5/6 nephrectomized rats, the expression of 13 miRNAs was significantly increased (>1.5-fold, P < 0.05), whereas the expression of 7 miRNAs was significantly decreased (>1.5-fold, P < 0.05). Most of the deregulated miRNA species are implicated in endothelial-to-mesenchymal transition and inflammatory processes. Both telmisartan and linagliptin suppressed the induction of pro-fibrotic miRNAs, such as miR-199a-3p, and restored levels of anti-fibrotic miR-29c. In conclusion, the linagliptin and telmisartan-induced restorative effects on miR-29c expression were reflected in urinary exosomes, suggesting that miRNA profiling of urinary exosomes might be used as a biomarker for CKD progression and monitoring of treatment effects. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) are short non-coding RNA species which are important post-transcriptional regulators of gene expression and play an important role in the pathogenesis of diabetic nephropathy. miRNAs are present in urine in a remarkably stable form packaged in extracellular vesicles, predominantly exosomes. In the present study, urinary exosomal miRNA profiling was conducted in urinary exosomes obtained from 8 healthy controls (C), 8 patients with type II diabetes (T2D) and 8 patients with type II diabetic nephropathy (DN) using Agilent´s miRNA microarrays. In total, the expression of 16 miRNA species was deregulated (>2-fold) in DN patients compared to healthy donors and T2D patients: the expression of 14 miRNAs (miR-320c, miR-6068, miR-1234-5p, miR-6133, miR-4270, miR-4739, miR-371b-5p, miR-638, miR-572, miR-1227-5p, miR-6126, miR-1915-5p, miR-4778-5p and miR-2861) was up-regulated whereas the expression of 2 miRNAs (miR-30d-5p and miR-30e-5p) was down-regulated. Most of the deregulated miRNAs are involved in progression of renal diseases. Deregulation of urinary exosomal miRNAs occurred in micro-albuminuric DN patients but not in normo-albuminuric DN patients. We used qRT-PCR based analysis of the most strongly up-regulated miRNAs in urinary exosomes from DN patients, miRNAs miR-320c and miR-6068. The correlation of miRNA expression and micro-albuminuria levels could be replicated in a confirmation cohort. In conclusion, urinary exosomal miRNA content is altered in type II diabetic patients with DN. Deregulated miR-320c, which might have an impact on the TGF-β-signaling pathway via targeting thrombospondin 1 (TSP-1) shows promise as a novel candidate marker for disease progression in type II DN that should be evaluated in future studies. [ABSTRACT FROM AUTHOR]

Background: Normalization of microarrays is a standard practice to account for and minimize effects which are not due to the controlled factors in an experiment. There is an overwhelming number of different methods that can be applied, none of which is ideally suited for all experimental designs. Thus, it is important to identify a normalization method appropriate for the experimental setup under consideration that is neither too negligent nor too stringent. Major aim is to derive optimal results from the underlying experiment. Comparisons of different normalization methods have already been conducted, none of which, to our knowledge, comparing more than a handful of methods. Results: In the present study, 25 different ways of pre-processing Illumina Sentrix BeadChip array data are compared. Among others, methods provided by the BeadStudio software are taken into account. Looking at different statistical measures, we point out the ideal versus the actual observations. Additionally, we compare qRT-PCR measurements of transcripts from different ranges of expression intensities to the respective normalized values of the microarray data. Taking together all different kinds of measures, the ideal method for our dataset is identified. Conclusions: Pre-processing of microarray gene expression experiments has been shown to influence further downstream analysis to a great extent and thus has to be carefully chosen based on the design of the experiment. This study provides a recommendation for deciding which normalization method is best suited for a particular experimental setup. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) are short, non-coding RNA species that are important post-transcriptional regulators of gene expression and play an important role in the pathogenesis of non-alcoholic fatty liver disease. Here, we investigated the phosphodiesterase 5 (PDE5) inhibitor induced effects on hepatic and plasma exosomal miRNA expression in CCl4-treated rats. In the present study, hepatic miRNA profiling was conducted using the Nanostring nCounter technology and mRNA profiling using RNA sequencing from PDE5 treated rats in the model of CCl4-induced liver fibrosis. To evaluate if the PDE5 inhibitor affected differentially expressed miRNAs in the liver can be detected in plasma exosomes, qRT-PCR specific assays were used. In livers from CCl4-treated rats, the expression of 22 miRNAs was significantly increased (>1.5-fold, adj. p < 0.05), whereas the expression of 16 miRNAs was significantly decreased (>1.5-fold, adj. p < 0.05). The majority of the deregulated miRNA species are implicated in fibrotic and inflammatory processes. The PDE5 inhibitor suppressed the induction of pro-fibrotic miRNAs, such as miR-99b miR-100 and miR-199a-5p, and restored levels of anti-fibrotic miR-122 and miR-192 in the liver. In plasma exosomes, we observed elevated levels of miR-99b, miR-100 and miR-142-3p after treatment with the PDE5-inhibitor compared to CCl4/Vehicle-treated. Our study demonstrated for the first time that during the development of hepatic fibrosis in the preclinical model of CCl4-induced liver fibrosis, defined aspects of miRNA regulated liver pathogenesis are influenced by PDE5 treatment. In conclusion, miRNA profiling of plasma exosomes might be used as a biomarker for NASH progression and monitoring of treatment effects. [ABSTRACT FROM AUTHOR]

Psoriasis is a chronic inflammatory disease characterized by demarcated, raised, and scaling skin lesions. It often serves as a model for immune-mediated disorders. Gene expression profiling of affected skin has allowed insights into psoriasis pathogenesis. However, the mechanisms leading to specific mRNA expression alterations in psoriasis are barely understood. To perform integrated microRNA-mRNA expression studies of non-lesional, peri-lesional, and lesional skin from psoriasis patients. Cutaneous microRNA and mRNA expression profiles of 14 patients using Nanostring nCounter-technology and RNA sequencing as well as in vitro keratinocyte stimulation and qPCR studies. Only 3.5 % of microRNAs manifested a robust gradual expression trend from non-lesional to paired lesional skin, with 61 % being upregulated and 39 % being downregulated. Relevance of these microRNA regulations was supported by their inverse association with 57 % of the mRNA species found to be regulated during psoriatic lesion development. Many of the involved mRNAs were downregulated and functionally related to keratinocyte metabolism, barrier function, and neuronal signaling, and were already regulated in peri-lesional skin. An integrated correlation analysis revealed a robust interaction for 134 microRNAs/mRNAs pairs. In vitro keratinocyte studies of selected microRNAs/mRNAs revealed regulations of all analyzed microRNAs in a psoriasis-like manner by IL-17A/TNF-α (e.g. hsa-miR-23a-3p), IFN-γ (e.g. hsa-miR-106a-5p/miR-17-5p), or IL-24 (e.g. hsa-miR-203a-3p). Moreover, most of their predicted target mRNAs (e.g. ID4, EPHB2) were respectively altered by the same cytokines. Our study suggests that, during development of psoriatic lesions, defined aspects of psoriasis pathogenesis are regulated by the action of microRNAs. [ABSTRACT FROM AUTHOR]