Circulating miR-199a and long noncoding-RNA ANRIL as Promising Diagnostic Biomarkers for Inflammatory Bowel Disease.

BACKGROUND & AIMS: Inflammatory bowel disease (IBD), involving both Crohn's disease (CD) and ulcerative colitis (UC), represents a chronic, immune-mediated inflammatory disease due to an uncontrolled, ongoing inflammatory response to intestinal bacteria in those with genetic susceptibility. MicroRNA (miRNA) extrusion from relevant remote organs or tissues is reflected in the expression of miRNAs in serum and plasma. Both UC and CD patients had higher blood levels of expressed miR-199a. Long noncoding RNA (lncRNA) ANRIL is a proinflammatory gene that mediates nuclear factor κB to play a role in inflammatory diseases, such as IBD. The aim of the current study is to investigate the potential role of both miR-199a and ANRIL in diagnosing IBD in adult patients. METHODS: Sixty-seven IBD patients diagnosed clinically, radiologically, endoscopically, and histologically were included in this prospective cohort study. Participants were classified into 3 groups: the UC group (n = 35), the CD group (n = 32), and the control group (n = 30). Demographics, history taking, laboratory characteristics, and treatments were recorded. Tumor necrosis factor α , miR-199a, and ANRIL were measured. RESULTS: The findings suggested that miR-199a and ANRIL might be associated with the occurrence or progression of IBD because both genes were substantially expressed in the peripheral blood of patients with this condition. Receiver-operating characteristic curve analysis indicated that the detection of miR-199a and ANRIL had a predictive sensitivity of 62.9% and 88.6% and a specificity of 70.7% and 96.7% for the occurrence of UC cases, respectively, and a predictive sensitivity of 72.4% and 46.9% and a specificity of 96.7% and 34.7% for the occurrence of CD cases, respectively. CONCLUSIONS: Both miR-199a and ANRIL are abundant in the sera of IBD adult Egyptian patients (UC and CD). Both can represent a noninvasive marker for early disease diagnosis.

This study investigated the relation between tumor necrosis factor α, ANRIL, and miR-199a in inflammatory bowel disease patients to determine the probability of using them as prognostic and diagnostic biomarkers, as well as distinguishing between Crohn's disease and ulcerative colitis patients. Our findings showed that ANRIL and miR-199a can represent noninvasive biomarkers for early disease diagnosis.

Expression profile of LncRNA ANRIL, miR-186, miR-181a, and MTMR-3 in patients with preeclampsia.

Preeclampsia (PE) is a leading cause of maternal and neonatal morbidity and mortality worldwide. Several studies demonstrated the role of lncRNAs and miRNAs in the pathogenesis of preeclampsia; the aim was to detect the expression profiles of serum LncRNA ANRIL, miR-186, miR-181a, and MTMR-3 in patients with preeclampsia. The study included 160 subjects divided into 80 subjects considered as a control group, 80 patients with preeclampsia. We found that there was a significant difference between the preeclampsia and control groups with up-regulation of miR-186 median (IQR) = 4, 29 (1.35-7.73) (P < 0.0001), miR-181a median (IQR) = 2.45 (0.83-6.52) (P = 0.028), and downregulation of lncRNA ANRIL median (IQR) = 0.35(0.28-0.528) (P < 0.0001), MTMR median (IQR) = 0.32(0.155-1.11), (P < 0.0001). ROC curve of lncRNA ANRIL, miR-186, miR-181a, and MTMR-3 in preeclampsia patients showing the roles of these markers in the diagnosis of preeclampsia. In conclusion, serum LncRNA ANRIL, miR-186, miR-181a, and MTMR-3 could be promising biomarkers in the diagnosis of preeclampsia.

Lnc-HULC, miR-122, and sirtulin-1 as potential diagnostic biomarkers for psoriasis and their association with the development of metabolic syndrome during the disease course.

Psoriasis is a persistent inflammatory skin disorder driven by T cells. The disease is characterized by aberrant keratinocytes (KCs) differentiation, epidermal proliferation, and excessive hyperplasia of veins and arteries. The purpose of the study was to identify the levels of circulating lnc-HULC, miR-122, and Sirtuin 1 (SIRT-1) in psoriatic patients, evaluate their possible roles as diagnostic biomarkers, and link their levels with the development of metabolic syndrome during psoriasis progression. This study included 176 participants. The subjects were divided into four groups, with 44 participants in each group. All patients have undergone a complete history taking and clinical examination. Laboratory investigations included Low-density lipoprotein (LDL), High-density lipoprotein (HDL), Triglycerides (TG), Fasting blood sugar (FBS), and cholesterol plasma levels. Serum levels of miR-122 and lnc-HULC were examined by qRT-PCR. Serum levels of SIRT-1 were examined by ELISA. The serum concentrations of lnc-HULC and miR-122 were significantly higher in psoriatic participants compared to controls. Psoriatic patients' serum concentrations of SIRT-1 were much lower than those of healthy individuals. There was a negative association between SIRT-1 concentration and BMI, disease duration, PASI score, LDL, and cholesterol levels. The blood levels of lnc-HULC, miR-122, and SIRT-1 in psoriasis patients provide a promising role as diagnostic biomarkers in patients with and without metabolic syndrome.

Long non-coding RNA NBAT1, TUG1, miRNA-335, and miRNA-21 as potential biomarkers for acute ischemic stroke and their possible correlation to thyroid hormones.

Objective: RNA-based mechanisms of epigenetic modification related to acute ischemic stroke (AIS) have been widely studied recently. The current work aimed to determine the potential roles of four ncRNAs (TUG1 and its target miR-21, NBAT1, and miR-335) as promising diagnostic biomarkers in AIS as well as their involvement in the disease pathogenesis. Methods: The levels of the studied lncRNAs and miRNAs were measured in the serum for two different groups, including patients with AIS (60) and healthy controls (60). All individuals were subjected to a full history investigation and clinical examination. Blood samples were tested for FBS, 2HPP, TAG, HDL, LDL, TSH, T3, and T4 levels. Results: The serum levels of TUG1 were significantly increased in AIS patients compared to control subjects. It is worthwhile to note that serum TUG1 levels were positively correlated with cholesterol, triglycerides, LDL, carotid IMT (Intima-media thickness), and miR-21, while they were negatively correlated with HDL levels. Our study showed that NBAT1 serum expression levels were elevated in AIS patients compared to controls. NBAT1 expression levels were observed to be positively correlated with triglycerides, TUG1, and miR-21. NBAT1 could distinguish between AIS patients and controls with a sensitivity of 100% and specificity of 100% at a cut-off point of 1.45. Regarding miR-335, we found that its expression levels were downregulated in AIS patients compared with healthy controls. It could distinguish between AIS patients and controls with a sensitivity of 73.3% and a specificity of 100% at a cut-off point of 0.796. Conclusion: Our results revealed that serum TUG1, miR-21, NBAT1, and miR-335 could be promising molecular diagnostic markers for AIS as these biomarkers could discriminate between AIS patients and healthy controls.

The potential role of serum expression profile of long non coding RNAs, Cox2 and HOTAIR as novel diagnostic biomarkers in systemic lupus erythematosus.

BACKGROUND: The role of the long non-coding RNAs (lncRNAs) in the pathogenesis of systemic lupus erythematosus (SLE) is mostly unknown, despite increasing evidence that lncRNAs extensively participate in physiological and pathological conditions. AIM: To detect the level of lncRNA-Cox2, HOTAIR, IL-6, and MMP-9 in the serum of SLE patients and to correlate these levels with disease activity and patients' clinical and laboratory data to evaluate the value of these biomarkers for SLE diagnosis and assessment of disease activity. METHODS: Blood samples from 58 SLE patients, and 60 healthy controls (HCs) were used for detection of lncRNAs-Cox2 and HOTAIR expression levels by real-time polymerase chain reaction. Both IL-6 and MMP-9 serum levels were assayed by enzyme-linked immunosorbent assay. Lupus activity was assessed with the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI). RESULTS: The serum expression levels of lncRNA-Cox2 and HOTAIR were significantly up-regulated in SLE patients vs HCs (fold change [median (IQR) was 1.29(0.81-1.71, P<0.0001) and 2.68(0.95-3.67), P = 0.038) for lncRNA-Cox2 and HOTAIR, respectively. Serum levels of both IL-6 and MMP-9 were significantly high in SLE patients compared with HCs (P≤0.001 for each). The up-regulated lncRNA-Cox2 was positively associated with the presence of neurological manifestations in SLE patients (P = 0.007). Furthermore, HOTAIR expression level had significantly positive correlation with IL-6 (r = 0.578, P<0.0001), MMP-9 level (r = 0.762, P<0.0001), nephritis grades (r = 0.296, P = 0.024) and proteinuria (r = 0.287, P = 0.035). LncRNA-Cox2 showed sensitivity and specificity 72.4%, and 100.0% respectively. HOTAIR sensitivity was 60.3%, and specificity was 100.0%. By multiple logistic regression analysis, lncRNA-Cox2 and HOTAIR were found as SLE independent predictors. CONCLUSION: LncRNA-COX2 and HOTAIR can be used as new non-invasive biomarkers for the diagnosis of SLE.

The Influence of rs1859168 Polymorphism on Serum Expression of HOTTIP and Its Target miR-615-3p in Egyptian Patients with Breast Cancer.

BACKGROUND: Polymorphisms of long noncoding RNAs are lately documented as hazardous factors for the development of numerous tumors. Furthermore, the evaluation of noncoding RNAs has emerged as a novel detector of breast cancer patients. We aimed to genotype the HOXA transcript at the distal tip (HOTTIP) rs1859168 and assess its relationship with the levels of the serum HOTTIP and its target miR-615-3p in patients with breast cancer (BC). METHODS: One hundred and fifty-one patients with BC, 139 patients with fibroadenoma (FA), and 143 healthy participants were incorporated into the current study. The genotyping of rs1859168 and the measurements of the HOTTIP and miR-615-3p levels were assessed using quantitative real-time PCR. RESULTS: We revealed a significant association between each of the CC genotypes, C allele, dominant and recessive models, and the increased risk of BC (p = 0.013, p < 0.001, p < 0.001, and p < 0.001, respectively) relative to the healthy controls. Similarly, the CC genotype, C allele, and recessive model were observed to be related to the increased incidence of BC with respect to FA (p < 0.001 for all). A significant upregulation of HOTTIP and a marked decrease of miR-615-3p were verified in patients with BC compared to each of the healthy individuals, patients with FA, and the non-BC group (healthy subjects + FA) (p < 0.001 for all). A significant negative correlation was demonstrated between the expression of HOTTIP and miR-615-3p in the serum of patients with BC. The HOTTIP expression was upregulated, while that of miR-615-3p was downregulated in patients with BC who carried the CC genotype with respect to those who carried the AA or AC genotypes (p < 0.05 for all). CONCLUSIONS: The genetic variants of rs1859168 are linked to an increased susceptibility to BC. Moreover, HOTTIP and miR-615-3p may be used as novel indicators and targets for the treatment of patients with BC.

Association between LINC00657 and miR-106a serum expression levels and susceptibility to colorectal cancer, adenomatous polyposis, and ulcerative colitis in Egyptian population.

Colorectal cancer (CRC) represented the second cause of mortality among cancer patients. Long noncoding RNAs and microRNAs (miRNAs) serve as noninvasive biomarkers for CRC surveillance and introduce new therapeutic approaches. LINC00657 and miR-106a expression levels play a pivotal role in CRC. This study included 190 Egyptian subjects, and the expression levels of LINC00657 and miR-106a in serum were measured by using quantitative real-time polymerase chain reaction. We found that upregulation of LINC00657 and downregulation of miR-106a are significantly associated with the development of CRC. Also, a positive correlation was detected between their serum levels. In addition, serum LINC00657 can distinguish adenomatous polyposis (AP) patients and/or ulcerative colitis (UC) patients from controls. Also the miRNA-106a expression level discriminates AP but not UC from healthy individuals. Our study cited new diagnostic biomarkers for CRC, AP, and UC among Egyptians in addition to be noninvasive screening tools for CRC in both healthy subjects and those having precancerous lesions. © 2019 IUBMB Life, 71(9):1322-1335, 2019.

Expression Profile of Long Noncoding RNAs, lnc-Cox2, and HOTAIR in Rheumatoid Arthritis Patients.

Despite the increased proof that long noncoding RNAs (lncRNAs) can control gene expression and broadly affect the normal physiological and disease conditions, the part of lncRNAs in rheumatoid arthritis (RA) is not well known. This study aimed to assess the serum expression levels of lnc-Cox2 and HOTAIR in RA and to investigate their role as novel noninvasive biomarkers in diagnosis of RA. Also, their relations with the levels of interleukin (IL)-6 and matrix metalloproteinase (MMP)-9 and with other clinicolaboratory data in RA patients were analyzed. LncRNAs-Cox2 and HOTAIR expression levels were detected in serum by real-time quantitative polymerase chain reaction. Both IL-6 and MMP-9 levels in serum were measured by enzyme-linked immunosorbent assay. The mRNA expression of lncRNA-Cox2 and HOTAIR was significantly upregulated in RA patients compared with healthy controls. Serum levels of both IL-6 and MMP-9 were significantly higher in RA patients than in healthy subjects (P < 0.001 each). Receiver operating characteristic (ROC) curve demonstrated that lncRNA-Cox2 and HOTAIR could discriminate RA patients from healthy controls. HOTAIR (not lnc-Cox2) was observed to be an independent predictor for RA using multiple logistic regression analysis. We concluded that lnc-Cox2 and HOTAIR serum expression levels can be used as novel noninvasive biomarkers for the diagnosis of RA.

Diagnostic and prognostic role of serum miR-20b, miR-17-3p, HOTAIR, and MALAT1 in diabetic retinopathy.

Noncoding RNAs are emerging biomarkers for many diseases including diabetic retinopathy (DR). This study aimed to measure the expression levels of serum miR-20b, miR-17-3p, HOTAIR, and MALAT1 in DR patients. A total of 80 patients diagnosed as type 2 diabetes (T2D) and 81 healthy subjects were recruited in this study. T2D patients were divided into three groups: nondiabetic retinopathy (NDR) group (30 patients), nonproliferative diabetic retinopathy (NPDR) group (30 patients), and proliferative diabetic retinopathy (PDR) group (20 patients). Quantitative real-time polymerase chain reaction (PCR) was used to assess the expression of serum miR-20b, miR-17-3p, HOTAIR, and MALAT1. We found a significant decrease in serum miR-20b and a significant increase in serum HOTAIR and MALAT1 in NDR patients compared to healthy subjects. Also, we revealed a significant decrease in serum miR-20b and miR-17-3p and a significant increase in serum HOTAIR and MALAT1 in each of NPDR and PDR groups when compared with healthy subjects. Furthermore, we reported a significant decrease in miR-20b and miR-17-3p and a significant increase in HOTAIR and MALAT1in DR as well as in PDR patients when compared with NDR patients. However, on comparing NPDR with NDR patients, no significant difference was observed regarding the expression levels of miR-20b and miR-17-3p, in contrast, significant elevation of serum HOTAIR and MALAT1 was found in NPDR. Moreover, we observed a significant decrease in serum miR-20b and miR-17-3p and a significant increase in serum HOTAIR and MALAT1 in PDR group relative to NPDR group. Receiver operating characteristic (ROC) curve was used for evaluating the diagnostic value of the examined serum noncoding RNAs as novel biochemical indicators detecting severity of DR. Our analyses suggested that the examined serum noncoding RNAs may discriminate DR (PDR and NPDR) from NDR. Furthermore, these noncoding RNAs (less importantly miR-17) can be used as promising novel biomarkers for prediction DR severity, distinguishing PDR from NPDR patients. We can conclude that serum miR-20b, miR-17-3p, HOTAIR, and MALAT1 may be used as noninvasive biomarkers for screening of DR and early diagnosis of PDR. © 2018 IUBMB Life, 71(3):310-320, 2019.

LncRNAs, MALAT1 and lnc-DC as potential biomarkers for multiple sclerosis diagnosis.

Long non-coding RNAs (lncRNAs) play an important role in gene regulation and show greater tissue specificity and complexity of biological functions. There is on-going research in their contribution in autoimmune diseases like multiple sclerosis (MS). Our study aimed at the evaluation of serum levels of lncRNAs, MALAT1 and lnc-DC in MS patients and the investigation of the association between these lncRNAs and the disease activity. Serum from 45 MS patients and 45 healthy controls was separated. MALAT1 and lnc-DC expression levels were assayed by qRT-PCR. MALAT1 and lnc-DC were significantly increased in MS patients (P=0.004 and P=0.006, respectively) in comparison with controls. There was a significant increase in expression of MALAT1 in secondary progressive MS (SPMS) subgroup compared with controls (P<0.0001); however, significant elevation of lnc-DC was demonstrated in relapsing remitting MS (RRMS) subtype (P=0.003) compared with normal controls. A positive association between the expression levels of MALAT1 and lnc-DC (r = 0.513, P < 0.0001) in MS patients was detected. Moreover, positive correlation was observed between MALAT1and lnc-DC in RRMS (r = 0.569, P = 0.001). Serum levels of MALAT1 and lnc-DC may serve as potential novel molecular biomarkers for MS diagnosis and may provide a new direction for its treatment.

Transforming growth factor-beta 1 in diabetic nephropathy.

In diabetic nephropathy the extent of matrix accumulation in both glomeruli and the interstitium correlates strongly with the degree of renal insufficiency and proteinuria. Factors responsible for the deposition and accumulation of extra cellular matrix material within the kidney are therefore of considerable interest. Such factors include the potent fibrotic cytokine TGF-beta. We measured serum TGF-beta1 in patients with various stages of diabetic nephropathy, and correlated its level with different biochemical parameters. The study was conducted on: Group I: 30 patients with diabetic nephropathy (Subgroup IA: 20 patients with microalbuminuria; Subgroup IB: 10 patients with overt nephropathy), Group II: 19 diabetic patients without nephropathy (positive control), Group III: 20 healthy volunteers (negative control). Serum creatinine, Fasting and postprandial blood glucose, Fasting serum cholesterol, Glycated haemoglobin (HbA1c), Microalbumin estimation in urine, Serum TGF-beta1 estimation were done for all the studied groups. Our results showed a statistically significantly higher serum TGF-beta1 level in patients with diabetic nephropathy versus diabetic patients without nephropathy (mean +/- SD, 47.66 +/- 21.92 and 27.07 +/- 15.46 respectively) (P<0.001). Also in patients with diabetic nephropathy versus healthy controls (mean +/- SD, 47.66 +/- 21.29 and 27.05 +/- 8.95 respectively) (P<0.001). While serum TGF-beta1 concentrations were almost similar in diabetic patients without nephropathy and in healthy controls. Serum TGF-beta1 was statistically significantly higher in patients with overt nephropathy versus patients with microalbuminuria (mean+/-SD, 73.5 +/- 2.41 and 34.9 +/- 12.41) (P<0.001). Serum TGF-beta1 was significantly positively correlated with albumin excretion rate, fasting and postprandial blood glucose levels, serum cholesterol and HbA1c, these correlations were only found in diabetic patients with nephropathy but not in those without nephropathy or the control group. (r=0.86, P<0.001, r=0.444, P<0.05, r=0.375, P<0.05, r=0.532, P <0.01, r=0.696, P<0.001 respectively. HbA1c was found to be predictor of 68% of changes of serum TGF-beta1 (P<0.001) and serum cholesterol was predictor of 73% of changes of serum TGF-beta1 concentration (P<0.01). In conclusion, our results suggest that TGF-beta1 may play a key role in the development and progression of diabetic nephropathy. Accordingly, it may be also directly implicated in the functional deterioration of the kidney functions seen in patients with diabetic nephropathy, therefore beside proper glycemic control, strategies aiming at antagonizing TGF- beta1 for example by the use of specific antibodies or a specific inhibitor of TGF-beta1 may help to prevent the development or attenuate the progression of nephropathy in diabetic patients.