Project description:Transcriptional profiling of Homo sapiens inflammatory skin diseases (whole skin biospies): Psoriasis (Pso), vs Atopic Dermatitis (AD) vs Lichen planus (Li), vs Contact Eczema (KE), vs Healthy control (KO) In recent years, different genes and proteins have been highlighted as potential biomarkers for psoriasis, one of the most common inflammatory skin diseases worldwide. However, most of these markers are not psoriasis-specific but also found in other inflammatory disorders. We performed an unsupervised cluster analysis of gene expression profiles in 150 psoriasis patients and other inflammatory skin diseases (atopic dermatitis, lichen planus, contact eczema, and healthy controls). We identified a cluster of IL-17/TNFα-associated genes specifically expressed in psoriasis, among which IL-36γ was the most outstanding marker. In subsequent immunohistological analyses IL-36γ was confirmed to be expressed in psoriasis lesions only. IL-36γ peripheral blood serum levels were found to be closely associated with disease activity, and they decreased after anti-TNFα-treatment. Furthermore, IL-36γ immunohistochemistry was found to be a helpful marker in the histological differential diagnosis between psoriasis and eczema in diagnostically challenging cases. These features highlight IL-36γ as a valuable biomarker in psoriasis patients, both for diagnostic purposes and measurement of disease activity during the clinical course. Furthermore, IL-36γ might also provide a future drug target, due to its potential amplifier role in TNFα- and IL-17 pathways in psoriatic skin inflammation. In recent years, different genes and proteins have been highlighted as potential biomarkers for psoriasis, one of the most common inflammatory skin diseases worldwide. However, most of these markers are not psoriasis-specific but also found in other inflammatory disorders. We performed an unsupervised cluster analysis of gene expression profiles in 150 psoriasis patients and other inflammatory skin diseases (atopic dermatitis, lichen planus, contact eczema, and healthy controls). We identified a cluster of IL-17/TNFα-associated genes specifically expressed in psoriasis, among which IL-36γ was the most outstanding marker. In subsequent immunohistological analyses IL-36γ was confirmed to be expressed in psoriasis lesions only. IL-36γ peripheral blood serum levels were found to be closely associated with disease activity, and they decreased after anti-TNFα-treatment. Furthermore, IL-36γ immunohistochemistry was found to be a helpful marker in the histological differential diagnosis between psoriasis and eczema in diagnostically challenging cases. These features highlight IL-36γ as a valuable biomarker in psoriasis patients, both for diagnostic purposes and measurement of disease activity during the clinical course. Furthermore, IL-36γ might also provide a future drug target, due to its potential amplifier role in TNFα- and IL-17 pathways in psoriatic skin inflammation.
Project description:Exosomal microRNAs have recently been studied as potential diagnostic marker for various malignancies, including hepatocellular carcinoma (HCC). The aim of this study was to investigate serum exosomal microRNA profiles as HCC diagnostic marker. Transmission electron microscopy and western blot were used to identify serum exosomes. Deep sequencing was performed to screen differentially expressed microRNAs between HCC (n=5) and liver cirrhosis (LC, n=5) group. Three upregulated and two downregulated microRNAs were selected for qPCR analysis. The levels of selected microRNAs were normalized to Caenorhabditis elegans miR-39 microRNA mimics. Serum exosomal level of miR-122, miR-148a, and miR-1246 were further analyzed and significantly higher in HCC than LC and normal control (NC) group (P<0.001), but not different from chronic hepatitis group(p>0.05). The receiver operating characteristic curve was used to evaluate diagnostic perfromance of candidate microRNAs. Area under the curve (AUC) of miR-148a was 0.891 [95 % confidence interval (CI), 0.809-0.947] in discriminating HCC from LC, remarkably higher than alpha fetoprotein (AFP) (AUC: 0.712, 95 % CI: 0.607-0.803). Binary logistic regression was adpoted to establish the diagnostic model for discriminating HCC from LC. And the combination of miR-122, miR-148a and AFP increased the AUC to 0.931 (95% CI, 0.857-0.973), which can also be applied for distinguishing early HCC from LC. miR-122 was the best for differentiating HCC from NC (AUC: 0.990, 95% CI, 0.945-1.000). These data suggests that serum exosomal microRNAs signature or their combination with traditional biomarker may be used as a suitable peripheral screening tool for HCC.
Project description:Inflammatory bowel disease (IBD) is a chronic inflammatory disorder of the gastrointestinal tract affecting over 3 million adults in the United States. Despite being widespread, reliable early diagnostic tests are not available. In order to remedy this, we examined exosomal small RNA (smRNA), specifically targeting microRNA (miRNA) and piRNA from the stool samples of IBD model mice, interleukin 10 knockout mice (IL-10 KO), as a potential diagnostic marker. Stool samples were specifically chosen because they are readily available and collection is noninvasive.
Project description:Kynureninase is a member of a large family of catalytically diverse but structurally homologous pyridoxal 5'-phosphate (PLP) dependent enzymes known as the aspartate aminotransferase superfamily or alpha-family. The Homo sapiens and other eukaryotic constitutive kynureninases preferentially catalyze the hydrolytic cleavage of 3-hydroxy-l-kynurenine to produce 3-hydroxyanthranilate and l-alanine, while l-kynurenine is the substrate of many prokaryotic inducible kynureninases. The human enzyme was cloned with an N-terminal hexahistidine tag, expressed, and purified from a bacterial expression system using Ni metal ion affinity chromatography. Kinetic characterization of the recombinant enzyme reveals classic Michaelis-Menten behavior, with a Km of 28.3 +/- 1.9 microM and a specific activity of 1.75 micromol min-1 mg-1 for 3-hydroxy-dl-kynurenine. Crystals of recombinant kynureninase that diffracted to 2.0 A were obtained, and the atomic structure of the PLP-bound holoenzyme was determined by molecular replacement using the Pseudomonas fluorescens kynureninase structure (PDB entry 1qz9) as the phasing model. A structural superposition with the P. fluorescens kynureninase revealed that these two structures resemble the "open" and "closed" conformations of aspartate aminotransferase. The comparison illustrates the dynamic nature of these proteins' small domains and reveals a role for Arg-434 similar to its role in other AAT alpha-family members. Docking of 3-hydroxy-l-kynurenine into the human kynureninase active site suggests that Asn-333 and His-102 are involved in substrate binding and molecular discrimination between inducible and constitutive kynureninase substrates.
Project description:Pancreatic cancer has a dismal prognosis partly due to late diagnosis, where a reliable non-invasive diagnosis is urgently wanted. We here explored, whether serum-derived PaCa exosomes may provide a diagnostic means and microRNA may be advantageous. Exosomes were collected from the serum of 133 patients with pancreatic cancer, 22 patients with non-malignant pancreatic tumors, 25 patients with chronic pancreatitis and 20 healthy donors. Exosomes were isolated by ultracentrifugation and used qRT-PCR for miRNA quantification and miRCURY LNA microRNA Array for miRNA analysis.