Abstract:| INTRODUC TI ONMicroRNAs (miRNAs) are highly conserved, small (21-25 nucleotides in length), noncoding RNAs that are present in biofluids such as serum, plasma, and urine. 1 Through the posttranscriptional regulation of mRNA expression, miRNAs control diverse physiological activities. 2 Recently, it has been reported that biofluid-derived miRNAs can serve as novel noninvasive biomarkers for the early detection of cancers, and degenerative and metabolic diseases, as well as therapeutic targets for such conditi… Show more
“…RNAseq and dPCR generate relative and absolute quantification of genes through different methodologies, which could explain the mismatch. One potential source of discrepancy is the library preparation process required by RNAseq, but not dPCR 20 …”
Section: Discussionmentioning
confidence: 99%
“…Others normalize quantitative reverse‐transcription PCR (qPCR) to plasma input volume 19 . RNA deep‐sequencing (RNAseq) allows relative and absolute quantification based on normalization across millions of all mapped reads 20 . Digital droplet PCR (dPCR) provides absolute quantification without a normalization gene by measuring tens of thousands of PCR reactions in parallel and assaying against a standard curve for 6‐carboxyfluorescein (FAM) fluorescence 17 .…”
Section: Introductionmentioning
confidence: 99%
“…19 RNA deep-sequencing (RNAseq) allows relative and absolute quantification based on normalization across millions of all mapped reads. 20 Digital droplet PCR (dPCR) provides absolute quantification without a normalization gene by measuring tens of thousands of PCR reactions in parallel and assaying against a standard curve for 6-carboxyfluorescein (FAM) fluorescence. 17 qPCR and dPCR for miRNAs in lung cancer have high correlation between the assays, with dPCR having lower coefficient of variation and greater reliability.…”
Background: Differentiating benign from canine malignant mammary tumors requires invasive surgical biopsy. Circulating microRNAs (miRNA) may represent promising minimally invasive cancer biomarkers in people and animals.Objectives: To evaluate the serum mRNA profile between dogs with and without mammary carcinoma, and to determine if any of these markers have prognostic significance.Animals: Ten healthy client-owned female dogs (5 intact, 5 spayed) and 10 dogs with histologically confirmed mammary carcinoma were included; 9 were client-owned, whereas 1 was a research colony dog.Methods: Retrospective study. Serum miRNA was evaluated by RNA deep-sequencing (RNAseq) and digital droplet PCR (dPCR).Expression of candidate biomarkers miR-18a, miR-19b, miR-29b, miR-34c, miR-122, miR-125a, and miR-181a was compared with clinical characteristics, including grade, metastasis, and survival.Results: 452 unique serum miRNAs were detected by RNAseq. Sixty-five individual miRNAs were differentially expressed (>±1.5-fold) and statistically significant between groups. Serum miR-19b (P = .003) and miR-125a (P < .001) were significantly higher in the mammary carcinoma group by dPCR. Both had high accuracy based on receiver operator characteristic area under the curve (0.930 for miR-125a; 0.880 for miR-19b). Circulating miR-18a by RNAseq was significantly higher in mammary carcinoma dogs with histologic evidence of lymphatic invasion (P = 0.03). There was no significant association with any miRNA and survival or inflammatory status. Conclusions and Clinical Importance: Circulating miRNAs are differentially expressed in dogs with mammary carcinoma. Serum miR-19b and miR-18a represent candidate biomarkers for diagnosis and prognosis, respectively. Abbreviations: AUC, area under the curve; CMT, canine mammary tumor(s); dPCR, digital droplet PCR; FAM, 6-carboxyfluorescein; miRs/miRNA, microRNA; qPCR, quantitative PCR; RNAseq, RNA deep-sequencing; ROC, receiver operator characteristic.
“…RNAseq and dPCR generate relative and absolute quantification of genes through different methodologies, which could explain the mismatch. One potential source of discrepancy is the library preparation process required by RNAseq, but not dPCR 20 …”
Section: Discussionmentioning
confidence: 99%
“…Others normalize quantitative reverse‐transcription PCR (qPCR) to plasma input volume 19 . RNA deep‐sequencing (RNAseq) allows relative and absolute quantification based on normalization across millions of all mapped reads 20 . Digital droplet PCR (dPCR) provides absolute quantification without a normalization gene by measuring tens of thousands of PCR reactions in parallel and assaying against a standard curve for 6‐carboxyfluorescein (FAM) fluorescence 17 .…”
Section: Introductionmentioning
confidence: 99%
“…19 RNA deep-sequencing (RNAseq) allows relative and absolute quantification based on normalization across millions of all mapped reads. 20 Digital droplet PCR (dPCR) provides absolute quantification without a normalization gene by measuring tens of thousands of PCR reactions in parallel and assaying against a standard curve for 6-carboxyfluorescein (FAM) fluorescence. 17 qPCR and dPCR for miRNAs in lung cancer have high correlation between the assays, with dPCR having lower coefficient of variation and greater reliability.…”
Background: Differentiating benign from canine malignant mammary tumors requires invasive surgical biopsy. Circulating microRNAs (miRNA) may represent promising minimally invasive cancer biomarkers in people and animals.Objectives: To evaluate the serum mRNA profile between dogs with and without mammary carcinoma, and to determine if any of these markers have prognostic significance.Animals: Ten healthy client-owned female dogs (5 intact, 5 spayed) and 10 dogs with histologically confirmed mammary carcinoma were included; 9 were client-owned, whereas 1 was a research colony dog.Methods: Retrospective study. Serum miRNA was evaluated by RNA deep-sequencing (RNAseq) and digital droplet PCR (dPCR).Expression of candidate biomarkers miR-18a, miR-19b, miR-29b, miR-34c, miR-122, miR-125a, and miR-181a was compared with clinical characteristics, including grade, metastasis, and survival.Results: 452 unique serum miRNAs were detected by RNAseq. Sixty-five individual miRNAs were differentially expressed (>±1.5-fold) and statistically significant between groups. Serum miR-19b (P = .003) and miR-125a (P < .001) were significantly higher in the mammary carcinoma group by dPCR. Both had high accuracy based on receiver operator characteristic area under the curve (0.930 for miR-125a; 0.880 for miR-19b). Circulating miR-18a by RNAseq was significantly higher in mammary carcinoma dogs with histologic evidence of lymphatic invasion (P = 0.03). There was no significant association with any miRNA and survival or inflammatory status. Conclusions and Clinical Importance: Circulating miRNAs are differentially expressed in dogs with mammary carcinoma. Serum miR-19b and miR-18a represent candidate biomarkers for diagnosis and prognosis, respectively. Abbreviations: AUC, area under the curve; CMT, canine mammary tumor(s); dPCR, digital droplet PCR; FAM, 6-carboxyfluorescein; miRs/miRNA, microRNA; qPCR, quantitative PCR; RNAseq, RNA deep-sequencing; ROC, receiver operator characteristic.
“…Overall, research on miR-147 and its role in the development of CKD is lacking. To our best knowledge, this study is the first to document upregulation of miR-147 in CKD, which reemphasizes the importance of an unbiased approach using sequencing technology for miRNA discovery 37 .…”
Dogs with X-linked hereditary nephropathy (XLHN) are an animal model for Alport syndrome in humans and progressive chronic kidney disease (CKD). Using mRNA sequencing (mRNA-seq), we have characterized the gene expression profile affecting the progression of XLHN; however, the microRNA (miRNA, miR) expression remains unknown. With small RNA-seq and quantitative RT-PCR (qRT-PCR), we used 3 small RNA-seq analysis tools (QIAGEN OmicSoft Studio, miRDeep2, and CPSS 2.0) to profile differentially expressed renal miRNAs, top-ranked miRNA target genes, and enriched biological processes and pathways in CKD progression. Twenty-three kidney biopsies were collected from 5 dogs with XLHN and 4 age-matched, unaffected littermates at 3 clinical time points (T1: onset of proteinuria, T2: onset of azotemia, and T3: advanced azotemia). We identified up to 23 differentially expressed miRNAs at each clinical time point. Five miRNAs (miR-21, miR-146b, miR-802, miR-142, miR-147) were consistently upregulated in affected dogs. We identified miR-186 and miR-26b as effective reference miRNAs for qRT-PCR. This study applied small RNA-seq to identify differentially expressed miRNAs that might regulate critical pathways contributing to CKD progression in dogs with XLHN.
“…Several studies reported the analysis and characterization of exosomes derived from canine-species MSCs and presented differential expression of cytokines and growth factors in adipose tissue- and bone-marrow-derived MSC exosomes [ 55 ]. Research studies using canine reported the comparison of miRNA expression levels in exosomes according to the sources of biofluids, such as serum and urine [ 56 ]. Stem cell-derived exosomes not only exert immune defenses, but also play a crucial antitumor role [ 57 , 58 , 59 ].…”
Adipose tissue-derived mesenchymal stem cells (AD-MSCs) release extracellular vesicles such as exosomes, apoptotic bodies, and microparticles. In particular, exosomes are formed inside cells via multivesicular bodies (MVBs), thus their protein, DNA, and RNA content are similar to those of the parent cells. Exosome research is rapidly expanding, with an increase in the number of related publications observed in recent years; therefore, the function and application of MSC-derived exosomes could emerge as cell-free therapeutics. Exosomes have been isolated from feline AD-MSCs and feline fibroblast cell culture media using ultracentrifugation. Feline exosomes have been characterized by FACS, nanoparticle tracking analysis, and transmission electron microscopy imaging. Moreover, cytokine levels were detected by sandwich enzyme-linked immunosorbent assay in exosomes and LPS-induced THP-1 macrophages. The size of the isolated exosomes was that of a typical exosome, i.e., approximately 150 nm, and they expressed tetraspanins CD9 and CD81. The anti-inflammatory factor IL-10 was increased in feline AD-MSC-derived exosomes. However, pro-inflammatory factors such as IL-1β, IL-8, IL-2, RANTES, and IFN-gamma were significantly decreased in feline AD-MSC-derived exosomes. This was the first demonstration that feline AD-MSC-derived exosomes enhance the inflammatory suppressive effects and have potential for the treatment of immune diseases or as an inflammation-inhibition therapy.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.