Disease Markers

We have previously identified an allele of the human SP‐A2 gene that occurs with greater frequency in an RDS population [12]. Because of the importance of SP‐A in normal lung function and its newly emerging role in innate host defense and regu‐lation of inflammatory processes, we wish to better characterize genotypes of both SP‐A1 and SP‐A2 genes. It has been determined that SP‐D shares similar roles in immune response. Therefore, in this report we 1) describe a novel, non radioactive PCR based‐cRFLP method for genotyping both SP‐A and SP‐D; 2) describe two previously unpublished biallelic polymorphisms within the SP‐D gene; 3) present the partial sequence of one new SP‐A1 allele (6A¹⁴) and describe other new SP‐A1 and SP‐A2 alleles; and 4) describe additional methodologies for SP‐A genotype assessment. The ability to more accurately and efficiently genotype samples from individuals with various pulmonary diseases will facilitate population and family based association studies. Genetic poly‐morphisms may be identified that partially explain individual disease susceptibility and/or treatment effectiveness.

Background.Recent reports indicate that matrix metalloproteinase-7 (MMP-7) and CC-chemokine ligand 18 (CCL18) are potential disease markers of idiopathic pulmonary fibrosis (IPF). The objective of this study was to perform direct comparisons of these two biomarkers with three well-investigated serum markers of IPF, Krebs von den Lungen-6 (KL-6), surfactant protein-A (SP-A), and SP-D.Methods.The serum levels of MMP-7, CCL18, KL-6, SP-A, and SP-D were evaluated in 65 patients with IPF, 31 patients with bacterial pneumonia, and 101 healthy controls. The prognostic performance of these five biomarkers was evaluated in patients with IPF.Results.The serum levels of MMP-7, KL-6, and SP-D in patients with IPF were significantly elevated compared to those in patients with bacterial pneumonia and in the healthy controls. Multivariate survival analysis showed that serum MMP-7 and KL-6 levels were independent predictors in IPF patients. Moreover, elevated levels of both KL-6 and MMP-7 were associated with poorer survival rates in IPF patients, and the combination of both markers provided the best risk discrimination using the C statistic.Conclusions.The present results indicated that MMP-7 and KL-6 were promising prognostic markers of IPF, and the combination of the two markers might improve survival prediction in patients with IPF.

Background:Micro-RNAs (miRNAs) regulate one-third of all protein-coding genes and are fundamental in the pathophysiology of a wide range of diseases. We studied the expression of several miRNA species (miR-200 family, miR-205 and miR-192) in the urinary sediment of patients with IgA nephropathy (IgAN).

Methods:We studied 43 patients with biopsy-proven IgAN. Urinary expression of miRNAs was determined and compared to that from 13 healthy controls.

Results:The levels of urinary miR-200a, miR-200b and miR-429, but not miR-200c, miR-141, miR-205, or miR-192, were down-regulated in patients with IgAN. Proteinuria significantly correlated with urinary expression of miR-200a (r= −0.483,P< 0.001), miR-200b (r= −0.448,P= 0.001) and miR-429 (r= −0.466,P= 0.001). Baseline renal function significantly correlated with urinary expression of miR-200b (r= 0.512,P< 0.001) and miR-429 (r= 0.425, P = 0.005). Urinary gene expression of ZEB2 inversely correlated with miR-200b (r= −0.321,P= 0.017); and vimentin expression inversely correlated with that of miR-200a (r= −0.360,P= 0.007), miR-200b (r= −0.416,P= 0.002) and miR-429 (r= −0.375,P= 0.005). After 33.4 ± 12.6 months, the rate of renal function decline significantly correlated with urinary expression of miR-200b (r= 0.316,P= 0.034).

Conclusions:Urinary expression of miR-200a, miR-200b and miR-429 were down-regulated in patients with IgAN, and the degree of reduction correlated with disease severity and rate of progression. The results suggested that these miRNA species might play important roles in the pathophysiology of IgAN. Further studies are needed to clarify the role of urinary miRNA repression as a non-invasive marker of IgAN.

Schizophrenia is one of the most common neuropsychiatric disorders affecting nearly 1% of the human population. Current diagnosis of schizophrenia is based on complex clinical symptoms. The use of easily detectable peripheral molecular markers could substantially help the diagnosis of psychiatric disorders. Recent studies showed that peripheral blood lymphocytes (PBL) express subtypes of D1 and D2 subclasses of dopamine receptors. Recently, dopamine receptor D₃ (DRD3) was found to be over‐expressed in schizophrenic PBL and proposed to be a diagnostic and follow‐up marker for schizophrenia. In this study we screened PBL of 13 drug‐naive/drug‐free schizophrenic patients to identify additional markers of schizophrenia. One of the benefits of our study is the use of blood samples of non‐medicated, drug‐naive patients. This excludes the possibility that changes detected in gene expression levels might be attributed to the medication rather than to the disorder itself. Among others, genes for dopamine receptor D₂ (DRD2) and the inwardly rectifying potassium channel (Kir2.3) were found to be over‐expressed in microarray analysis. Increased mRNA levels were confirmed by quantitative real‐time PCR (QRT‐PCR) using the SybrGreen method and dual labeled TaqMan probes. The use of both molecular markers allows a more rapid and precise prediction of schizophrenia and might help find the optimal medication for schizophrenic patients.

Regulation of gene expression is essential for normal physiological functions; thus deregulation of gene expression is common in disease conditions. One level of regulation of gene expression is performed by noncoding RNAs, among which microRNAs (miRNA) are the best studied. Abnormal expression of these molecular players can lead to pathogenic processes such as heart disease, immune system abnormalities, and carcinogenesis, to name but a few. Of a length of 18–25 nucleotides miRNAs are involved in binding partial complementary sequences within the 3′-UTR (3′-untranslated region) of the target mRNAs. Depending on the type of neoplastic transformation, miRNAs can act both as oncogenes (oncomirs) or as tumor suppressors. Because of the great importance of miRNAs, most researches focus on either their role as biomarkers or their potential as therapeutic targets. Herein, we present the review of microRNA biology, function, and tumorigenic potential with emphasis on their role in lung cancer.

Molecular mechanisms responsible for acute relapse of multiple sclerosis (MS) remain currently unknown. The aim of this study is to identify the relapse‐specific biomarker genes in T lymphocytes of relapsing‐remitting MS (RRMS). Total RNA of CD3⁺ T cells isolated from six RRMS patients taken at the peak of acute relapse and at the point of complete remission was processed for DNA microarray analysis. We identified a set of 43 differentially expressed genes (DEG) between acute relapse and complete remission. By using 43 DEG as a discriminator, hierarchical clustering separated the cluster of relapse from that of remission. The molecular network of 43 DEG investigated by KeyMolnet, a bioinformatics tool for analyzing molecular interaction on the curated knowledge database, showed the most significant relationship with aberrant regulation of gene expression by the nuclear factor‐kappa B (NF‐κB) in T cells during MS relapse. These results support the logical hypothesis that NF‐κB plays a central role in triggering molecular events in T cells responsible for induction of acute relapse of MS, and suggest that aberrant gene regulation by NF‐κB on T‐cell transcriptome might serve as a molecular biomarker for monitoring the clinical disease activity of MS.

Background and Aim. Cholangiocarcinoma (CCA) is the most frequent biliary malignancy, which poses high mortality rate due to lack of early detection. Hence, most CCA cases are present at the advanced to late stages with local or distant metastasis at the time of diagnosis. Currently available tumor markers including CA19-9 and CEA are inefficient and of limited usage due to low sensitivity and specificity. Here, we attempt to identify serum tumor markers for CCA that can effectively distinguish CCA from benign biliary tract diseases (BBTDs).Methods. Serum samples from 19 CCA patients and 17 BBTDs were separated by SDS-PAGE followed with LC-MS/MS and were subjected to statistical analysis and cross-validation to identify proteins whose abundance was significantly elevated or suppressed in CCA samples compared to BBTDs.Results. In addition to identifying several proteins previously known to be differentially expressed in CCA and BBTDs, we also discovered a number of molecules that were previously not associated with CCA. These included FAM19A5, MAGED4B, KIAA0321, RBAK, and UPF3B.Conclusions. Novel serum biomarkers to distinguish CCA from BBTDs were identified using a proteomic approach. Further validation of these proteins has the potential to provide a biomarker for differentiating CCA from BBTDs.

Interleukin 23 receptor (IL23R) gene has been reported as a genetic factor strongly associated with inflammatory bowel disease, psoriasis, and ankylosing spondylitis. We investigated the association betweenIL23Rgene single nucleotide polymorphisms (SNPs) and susceptibility to sarcoidosis, including the clinical manifestation of uveitis.

Ninety-one sarcoidosis subjects (58 with and 33 without uveitis) and 104 healthy controls were genotyped for eleven IL23R SNPs. DNA was amplified using specific PCR primers and genotyped by denaturing HPLC and/or direct DNA sequencing. Case-control frequency comparisons were analyzed using Chi square test.

ThreeIL23RSNPs, rs7517847 (intron 6), rs11465804 (intron 8), and rs11209026 (exon 9, c.1142G>A, p.Arg381Gln) were associated with sarcoidosis in our population (p< 0.05): rs7517847 showed increased frequencies in sarcoidosis compared to controls, but rs11465804 and rs11209026 were decreased. Two of these SNPs were associated with the uveitis subgroup compared to controls: rs11465804 (0.9% vs. 7.2%, OR = 0.11, P = 0.013) and rs11209026 (1.8% vs. 7.3%, OR = 0.23, P = 0.038).

This finding indicates the association ofIL23Rpolymorphism with sarcoidosis, especially with sarcoid uveitis.IL23Rmay be a common susceptibility gene shared by several autoimmune disorders including inflammatory bowel disease, psoriasis, and ankylosing spondylitis and sarcoid uveitis.

Aims. Cancer cells use the indoleamine 2,3-dioxygenase 1 (IDO1) pathway to suppress the host’s immune response in order to facilitate survival, growth, invasion, and metastasis of malignant cells. Higher IDO1 expression was shown to be involved in colorectal cancer (CRC) progression and to be correlated with impaired clinical outcome. However, the potential correlation between the expression of IDO1 in a CRC population with a low mutation rate of theAPCgene remains unknown.Material and Methods. Tissues and blood samples were collected from 192 CRC patients. The expressions of IDO1, tryptophan 2,3-dioxygenase (TDO2), and beta-catenin proteins were analyzed by immunohistochemistry. Microsatellite instability (MSI) was determined by PCR amplification of microsatellite loci.Results. The results showed that high IDO1 or TDO2 protein expression was associated with characteristics of more aggressive phenotypes of CRC. For the first time, they also revealed a positive correlation between the abnormal expression of beta-catenin and IDO1 or TDO2 proteins in a CRC population with a low mutation rate ofAPC.Conclusion. We concluded that an IDO1-regulated molecular pathway led to abnormal expression of beta-catenin in the nucleus/cytoplasm of CRC patients with low mutation rate ofAPC, making IDO1 an interesting target for immunotherapy in CRC.

Background. Thyroid-associated orbitopathy (TAO) is the most common autoimmune disease of the orbit. It occurs more often in patients presenting with hyperthyroidism, characteristic of Graves’ disease, but may be associated with hypothyroidism or euthyroidism. The diagnosis of TAO is based on clinical orbital features, radiological criteria, and the potential association with thyroid disease. To date, there is no specific marker of the orbital disease, making the early diagnosis difficult, especially if the orbital involvement precedes the thyroid dysfunction.Summary. The goal of this review is to present the disease and combine the available data in the literature concerning investigation of TAO biomarkers.Conclusions. Despite the progress done in the understanding of TAO disease, some important pieces are still missing. Typically, for the future, major efforts have to be done in the discovery of new biomarkers, validation of the suspected candidates on multicenter cohorts with standardized methodologies, and establishment of their clinical performances on the specific clinical application fields in order to improve not only the management of the TAO patients but also the therapeutic options and follow-up.