骨髓间充质干细胞对肾移植受者T淋巴细胞分化和miRNA-155表达的影响

目的探讨骨髓间充质干细胞（MSCs）对肾移植受者T淋巴细胞分化和miRNA-155表达的影响。 方法选取2013年1月至2017年12月于福州总医院接受MSCs诱导+同种异体肾移植术的受者20例（MSCs组）,对照组为同期配对的异体肾移植受者20例。两组患者术后免疫抑制方案均为霉酚酸酯+他克莫司+强的松。两组患者分别于移植术前、术后第15天抽取静脉血,流式细胞仪检测外周血CD4⁺ CD25⁺ FoxP3⁺ Treg细胞/?CD4⁺细胞亚群比值;ELISA检测白细胞介素2（IL-2）、肿瘤坏死因子α（TNF-α）和白细胞介素10（IL-10）浓度;免疫磁珠分选外周血T淋巴细胞后,Rea1-time PCR法检测外周血T淋巴细胞中miRNA-?155的表达。两组间均数比较采用独立t检验,治疗前后均数比较采用配对t检验。 结果移植前两组肾移植受者外周血CD4⁺ CD25⁺ FoxP3⁺ Treg细胞/?CD4⁺细胞比值、IL-2、IL-?10、TNF-α、T淋巴细胞miRNA-155表达水平差异均无统计学意义（P均> 0.05）;术后第15天,与对照组相比,MSCs组外周血CD4⁺ CD25⁺ FoxP3⁺ Treg细胞/?CD4⁺细胞亚群比值（30.44﹪?± 4.23﹪? vs 26.06﹪?±4.77﹪,t = 2.365,P = 0.042）、IL-10水平（20.35?ng/?L?±?5.10?ng/?L? vs 16.63?ng/?L±6.26?ng/?L,t = 2.062,P?=?0.046）上升,而IL-2（27.47ng/?L±4.30 ng/?L vs 31.40?ng/?L±5.33 ng/L,t = 2.252,P = 0.015）、TNF-α（41.52?ng/?L±8.32?ng/L vs 46.67?ng/?L±6.71?ng/L,t = 2.157,P = 0.037）和T淋巴细胞miRNA-155表达水平（1.61±0.31 vs 1.89±0.15,t = 3.688,P?= 0.001）则降低。 结论MSCs能够升高肾移植受者外周血CD4⁺ CD25⁺ FoxP3⁺ Treg细胞/?CD4⁺细胞比值和IL-10水平,降低IL-2、TNF-α和T淋巴细胞miRNA-155表达水平,与MSCs的免疫耐受诱导有关。     

窒息新生儿血清miR-21、miR-210及miR-199a的表达及临床意义

目的:探讨窒息新生儿血清mi RNA-21、mi RNA-210及mi RNA-199a的表达及临床意义。方法:选取2015年9月1日到2017年7月31日我院新生儿科收治的窒息新生儿40例作为观察组,另选取同期在我院出生的健康新生儿40例作为对照组,根据阿氏(Apgar)评分将观察组的新生儿分为重度窒息组(12例)和轻度窒息组(28例)。检测所有新生儿血清中mi RNA-21、mi RNA-210及mi RNA-199a的表达水平,并比较观察组与对照组、重度窒息组与轻度窒息组血清中mi RNA-21、mi RNA-210及mi RNA-199a水平,并分析其相关性。结果:观察组的血清mi RNA-21、mi RNA-210水平高于对照组,mi RNA-199a水平低于对照组,差异有统计学意义(P0.05)。重度窒息组的血清mi RNA-21、mi RNA-210水平均高于轻度窒息组,mi RNA-199a水平低于轻度窒息组,差异有统计学意义(P0.05)。观察组血清mi RNA-21与mi RNA-210呈正相关(P0.05),血清mi RNA-21、mi RNA-210与mi RNA-199a水平呈负相关(P0.05)。结论:窒息新生儿血清中mi RNA-21、mi RNA-210呈现高表达,mi RNA-199a呈现低表达,且其表达水平与窒息严重程度相关。     

miRNA-148a and miRNA-30c expressions as potential biomarkers in breast cancer patients

BackgroundBreast cancer is an extensively identified malignant tumor and is a prime cause of cancer mortalities in females. It has been shown that alteration of miRNAs expression (up or down regulation) can affect the initiation and progression of many malignancies. We aimed to evaluate the role of circulating miRNA-148a and miRNA-30c in female patients with breast cancer and estimate their usage as potential biomarkers in the diagnosis, prognosis and survival of breast cancer.MethodsThis study included 75 breast cancer female patients.They were compared with 55 apparently healthy female subjects. miRNAs expression analysis was assessed via real-time PCR.ResultsTo discriminate breast cancer patients from controls, miR-30c showed the best performance at a cut off value of ≤20.6 (AUC = 0.998, 97.33% sensitivity, 96.36% specificity, p < 0.001), followed by miR-148a (AUC = 0.995, 94.67% sensitivity, 90.91% specificity, p < 0.001 at a cut off value of ≤0.1), CA 15-3 (AUC = 0.930, 88.0% sensitivity, 81.82% specificity, p < 0.001 at a cut off value of >21.3), and finally CEA (AUC = 0.751, 70.67% sensitivity, 63.64% specificity, p < 0.001 at a cut off value of >2.5).ConclusionmiRNA-148a and miRNA-30c expressions were down regulated in female patients with breast cancer and might be considered as potential blood biomarkers. Both also might have rule in disease treatment and selection of therapeutic targets. Future studies are needed to improve their role in predicting response to treatment and prognosis.     

LncRNA NKILA inhibits the proliferation and promotes the apoptosis of CSCC cells by downregulating miRNA-21

Long noncoding RNA (lncRNA) NKILA has been well studied in several types of human tumors as a tumor suppressor, while its involvement in cervical squamous cell carcinoma (CSCC) remains unclear. In our studies, we found that serum NKILA was at lower levels and serum microRNA-21 (miRNA-21) was at higher levels in patients with early stage CSCC than in the healthy female. Altered expression of NKILA and miRNA-21 can effectively separate patients with CSCC at an early stage from healthy controls. Serum levels of NKILA were significantly and negatively correlated with miRNA-21 in patients with CSCC but not in normal controls. Overexpression of NKILA mediated the inhibited expression of miRNA-21 in CSCC cells, but mimic transfection of miRNA-21 did not significantly change the expression level of NKILA. Overexpression of NKILA repressed the proliferation and promoted the apoptosis of CSCC cells, while miRNA-21 showed opposite functions. In addition, miRNA-21 mimic transfection reduced the effects of NKILA on CSCC cells. Collectively, lncRNA NKILA could repress the proliferation and promote the apoptosis of CSCC cells by downregulating miRNA-21.     

Serum levels of interleukin-13 and interferon-gamma from adult patients with asthma in Mysore

Serum protein analysis for noninvasive quantification of airway inflammation in asthma is a promising research tool in the field of lung diseases. Cytokines are believed to have major role in inflammatory process of the airways of the lung. There is an imbalance between T-helper (Th)-2 cells, which secrete interleukin (IL)-4 and interleukin (IL)-13, and Th1 cells, which secrete interferon (IFN)-gamma in asthma. To test the hypothesis that serum IL-13 and IL-4 levels may be elevated whereas IFN-gamma would be decreased in this cohort of patients, a property that could make them possible candidate biomarkers in determining asthma occurrence and severity, we measured concentrations of IL-4, IL-13 and IFN-gamma in serum samples of 88 subjects (44 normal, 12 with mild asthma, 16 with moderate asthma, and 16 with severe asthma). Serum Levels of IL-4, IL-13, and IFN-gamma were determined by an enzyme-linked immune-sorbent assay (ELISA). Median serum level of IFN-gamma in asthmatic patients was 8.0pg/ml, while it was 11.4pg/ml in healthy controls. However, the difference was not significant. Among the different age groups in whom IFN-gamma was assessed, the highest median value in both cases and controls was observed in the age group of 31-40years. The median serum level of IL-13 was 40.0pg/ml in asthmatic patients and 58.25pg/ml in healthy controls. The difference was not significant. On subgroup analysis, no significant difference of IFN-gamma and IL-13 between asthma of different severities was observed. The study also revealed nonsignificant difference of serum cytokines with the duration of asthma, number of allergens, and severity of sensitization. Normal serum levels of IFN-gamma and IL-13 in asthmatic patients suggest their neutral role in the inflammatory process; however, more studies are required to establish the effect of these cytokines in adulthood asthma in different ethnic populations.     

哮喘患儿血白细胞介素-10、13检测的临床意义

目的探讨哮喘患儿外周血白细胞介素-10(IL-10)、13的变化及其在哮喘发病机制中的作用。方法用ELISA双抗体夹心法测定20例哮喘患儿及18例正常儿童血浆IL-10、13的含量。结果哮喘组血浆IL-13水平明显高于正常对照组,IL-10水平明显低于正常对照组(p均<0.05)。结论IL-10、13等细胞因子参与儿童哮喘发病的病理生理过程,可为判断病情提供较好的实验室参数。     

Clinical significance of the expression of miRNA-21, miRNA-31 and miRNA-let7 in patients with lung cancer

ObjectiveTo explore the expression differences of miRNA-21, miRNA-31 and miRNA-let7 between lung cancer patient and healthy people, thereby providing reference for early diagnosis of lung cancer.MethodReal-time PCR was employed to determine the expression difference between lung cancer patients (50 cases) and healthy people (24 cases). The clinical data of lung cancer patients were analyzed to explore the correlation between clinicopathological characteristics and expression level of miRNA-21, miRNA-31, miRNA-let7.ResultsThe relative expression levels of miRNA-21 and miRNA-31 in lung cancer group were obviously higher than those in healthy control group, and the relative expression level of miRNA-let7 in lung cancer group was slightly higher than that in healthy control group. Lung cancer patients with lymph node metastasis had higher expression level than those without lymph node metastasis. The ROC curve showed that the three miRNAs had clinical diagnosis efficiency for lung cancer, and the combined detection of the three miRNAs were more efficient in diagnosing lung cancer. Survival curve analysis suggested that the median survival times of patients in the miRNA-21 and miRNA-31 high expression groups were shorter than those in the low expression groups, and the median survival time of patients in miRNA-let7 high expression group was longer than that in the low expression group.ConclusionPlasma miRNA-21, miRNA-31 and miRNA-let7 may be diagnostic marker for lung cancer.     

The diagnostic and prognostic role of MiRNA 15b and MiRNA 378a in neonatal sepsis

Background and objectivesSepsis is one of the major factors for both term and preterm babies with morbidity and mortality. On the basis of recent clinical trials, altered circulating micro-RNAs (miRNAs) may serve as possible biomarkers in sepsis for diagnosis and prognosis. The aim of this research is to assess the diagnostic and prognostic biomarkers of miRNA 15b and miRNA 378a for neonatal sepsis.Subjects & methodsThis study was carried out 25 neonates with sepsis admitted to neonatal ICU of Menoufia University Hospital and 25 healthy controls from February 2019 to May 2020. The relative quantification (RQ) of miRNA-15b and miRNA-378a expression was assessed using real time PCR technique. Results: Our results demonstrated that patients with sepsis had significantly higher level of MiRNA-15b than the healthy volunteers. On the other hand, patients with sepsis had significantly lower level of MiRNA-378a than the healthy volunteers. The ROC curve showed that the serum MiRNA-15b was a significant discriminator of sepsis with a combined sensitivity and specificity of 76% and 88% with cutoff point of 3.24. In addition, serum MiRNA-378a was a significant discriminator of sepsis with a combined sensitivity and specificity of 60% and 88% with cutoff point of 0.361. The miRNA-15b expression significantly correlated positive with respiratory rate (r =0.415,p =0.039), WBCs (r = 0.408, p =0.043), and CRP (r =0.407, p=0.043). Likewise, miRNA-378a expression significantly correlated negative with respiratory rate (r =-0.415p =0.024), WBCs (r =- 0.442, p =0.027), and CRP (r =- 0.459, p=0.021).ConclusionBoth MiRNA 15b and 378a are promising biomarker for neonatal sepsis.     

229例低复制慢性乙型肝炎患者血清相关指标的分析

目的探讨血清HBV—DNA低复制的CHB患者相关指标的变化及意义。方法收集HBV—DNA低复制的CHB患者血液标本229例,ALT、乙肝标志物、mIL-2R、IL-10、miRNA-122等指标均采用全自动分析仪测定,对结果进行比较和线性相关分析。结果低复制CHB患者中e抗原阳性患者占38．00％,e抗原阴性患者占51．53％;229例患者ALT升高者占总数的66．81％,患者血浆miRNA-122的表达量的变化与ALT浓度呈正相关（r=0．841）。e抗原阳性与e抗原阴性组IL-10、mIL-2R浓度与对照组比较差异均有统计学意义（P〈0．05）。结论HBV—DNA低复制CHB患者ALT、mlL-2R、IL-10、miRNA-122等指标的变化证明此类患者部分病例存在肝细胞实质性的损害,临床应高度重视,尤其要与非活动性乙型肝炎病原携带状态相鉴别,不可一概而论。     

Overexpression of microRNA-155 increases IL-21 mediated STAT3 signaling and IL-21 production in systemic lupus erythematosus

IntroductionInterleukin (IL)-21 is a key cytokine in autoimmune diseases such as systemic lupus erythematosus (SLE) by its regulation of autoantibody production and inflammatory responses. The objective of this study is to investigate the signaling capacity of IL-21 in T and B cells and assess its possible regulation by microRNA (miR)-155 and its target gene suppressor of cytokine signaling 1 (SOCS1) in SLE.MethodsThe signaling capacity of IL-21 was quantified by stimulating peripheral blood mononuclear cells (PBMCs) with IL-21 and measuring phosphorylation of STAT3 (pSTAT3) in CD4+ T cells, B cells, and natural killer cells. Induction of miR-155 by IL-21 was investigated by stimulating purified CD4+ T cells with IL-21 and measuring miR-155 expression levels. The functional role of miR-155 was assessed by overexpressing miR-155 in PBMCs from SLE patients and healthy controls (HCs) and measuring its effects on STAT3 and IL-21 production in CD4+ and CD8+ T cells.ResultsInduction of pSTAT3 in CD4+ T cells in response to IL-21 was significantly decreased in SLE patients compared to HCs (p < 0.0001). Further, expression levels of miR-155 were significantly decreased and SOCS1 correspondingly increased in CD4+ T cells from SLE patients. Finally, overexpression of miR-155 in CD4+ T cells increased STAT3 phosphorylation in response to IL-21 treatment (p < 0.01) and differentially increased IL-21 production in SLE patients compared to HCs (p < 0.01).ConclusionWe demonstrate that SLE patients have reduced IL-21 signaling capacity, decreased miR-155 levels, and increased SOCS1 levels compared to HCs. The reduced IL-21 signaling in SLE could be rescued by overexpression of miR-155, suggesting an important role for miR-155 in the reduced IL-21 signaling observed in SLE.

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-015-0660-z) contains supplementary material, which is available to authorized users.     

miRNA-155 and miRNA-181a as prognostic biomarkers for pediatric acute lymphoblastic leukemia

Mortality rates of acute lymphoblastic leukemia (ALL) have improved over the past 20 years; however, a significant portion of deaths stems from the lack of prognostic biomarkers, which can direct therapy and overcome drug resistance. microRNA-155a (miRNA-155a) and miRNA-181a are two single-stranded miRNAs involved in the pathogenesis of many types of leukemia and lymphoma and is linked to drug resistance. We investigated their expression levels in 55 patients, 45 diagnosed with ALL and 10 as a control group. We found that miRNA-155a and miRNA-181a were significantly upregulated in the ALL group with both being linked to high levels of minimal residual disease and poor prognosis. miRNA-155a cutoff value was significant in discriminating between high- and low-risk ALL patients as well as between ALL patients and healthy controls, miRNA-181a cutoff value, however, was not significant. Both markers levels were significantly downregulated after therapy. We conclude that miR-155 is correlated with poor prognosis in ALL, whereas we couldn’t link miRNA-181a to the prognosis in ALL. Moreover, the marked decrease in their expression after therapy could reflect their impact on disease outcome.     

Serum microRNA biomarkers for detection of non-small cell lung cancer

Non small cell lung cancer (NSCLC) is the leading cause of cancer-related mortality world-wide and the majority of cases are diagnosed at late stages of disease. There is currently no cost-effective screening test for NSCLC, and the development of such a test is a public health imperative. Recent studies have suggested that chest computed tomography screening of patients at high risk of lung cancer can increase survival from disease, however, the cost effectiveness of such screening has not been established. In this Phase I/II biomarker study we examined the feasibility of using serum miRNA as biomarkers of NSCLC using RT-qPCR to examine the expression of 180 miRNAs in sera from 30 treatment naive NSCLC patients and 20 healthy controls. Receiver operating characteristic curves (ROC) and area under the curve were used to identify differentially expressed miRNA pairs that could distinguish NSCLC from healthy controls. Selected miRNA candidates were further validated in sera from an additional 55 NSCLC patients and 75 healthy controls. Examination of miRNA expression levels in serum from a multi-institutional cohort of 50 subjects (30 NSCLC patients and 20 healthy controls) identified differentially expressed miRNAs. A combination of two differentially expressed miRNAs miR-15b and miR-27b, was able to discriminate NSCLC from healthy controls with sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of 100% in the training set. Upon further testing on additional 130 subjects (55 NSCLC and 75 healthy controls), this miRNA pair predicted NSCLC with a specificity of 84% (95% CI 0.73-0.91), sensitivity of 100% (95% CI; 0.93-1.0), NPV of 100%, and PPV of 82%. These data provide evidence that serum miRNAs have the potential to be sensitive, cost-effective biomarkers for the early detection of NSCLC. Further testing in a Phase III biomarker study in is necessary for validation of these results.     

Long noncoding RNA HAND2-AS1 inhibits cancer cell proliferation,migration, and invasion in esophagus squamous cell carcinoma by regulating microRNA-21

Long noncoding RNA (lncRNA) HAND2-AS1 is a well-characterized tumor suppressor in several types of malignancies, while its role in esophagus squamous cell carcinoma (ESCC) is unknown. In this study, we found that lncRNA HAND2-AS1 was downregulated, while microRNA-21 ( miRNA-21) was upregulated in tumor tissues than in adjacent healthy tissues of ESCC patients. Expression levels of lncRNA HAND2-AS1 and miRNA-21 were significantly and inversely correlated in tumor tissues but not in healthy tissues. Plasma levels of lncRNA HAND2-AS1 were lower in ESCC patients than in healthy controls, and downregulation of plasma lncRNA HAND2-AS1 distinguished early stage ESCC patients from healthy controls. lncRNA HAND2-AS1 overexpression resulted in downregulation of miRNA-21 in cells of ESCC cell lines and inhibited cell proliferation, migration, and invasion. miRNA-21 overexpression failed to affect lncRNA HAND2-AS1 expression but significantly attenuated the inhibitory effect of lncRNA HAND2-AS1 overexpression on cancer cell proliferation, migration, and invasion. Therefore, lncRNA HAND2-AS1 may inhibit cancer cell proliferation, migration, and invasion in ESCC by regulating miRNA-21.     

Increased interleukin-4 and decreased interferon-γ levels in serum of children with asthma

Background and purposeImmune and inflammatory responses, mediated by cytokines, play important roles in the pathophysiology of asthma. These responses are associated with over expression of T helper (Th)-2 cytokine, particularly interleukin (IL)-4 and IL-5, and decreased expression of Th-1 cytokine, IL-2 and IFN-γ. We hypothesized that there would be an imbalance in the levels of circulating IL-4 and IFN-γ in the asthmatic subjects.MethodWe investigated serum levels of IL-4 and IFN-γ among eighty children (18 steroid-naïve, 30 steroid-treated children with asthma and 32 healthy controls) using commercially available ELISA kits.ResultsSerum level of IL-4 was significantly higher in steroid-naïve group of asthmatic children compared to the healthy control subjects and was lower in steroid-treated group though the level was statistically not significant. In contrast, serum levels of IFN-γ were significantly lower in both steroid-naïve and steroid-treated groups of asthmatic children compared to healthy control subjects.ConclusionThe results of our study suggest that serum level of IL-4 may be elevated in concert with decreased level of IFN-γ in asthma. Determination of serum levels of IL-4 and IFN-γ may be a useful tool for understanding the disease processes in asthma.     

In vitro knock-out of miR-155 suppresses leukemic and HCV virus loads in pediatric HCV-4–associated acute lymphoid leukemia: A promising target therapy

Hepatitis C virus (HCV) infection is a major public health problem, having a high prevalence in Egypt. Leukemia and lymphoma have been associated with HCV infection. MicroRNA-155 (miR-155) has been reported to play a regulatory role in cancer, inflammation, and immune response to infection. The expression level of miR-155 in HCV viremic patients is controversial; although high miR-155 levels were demonstrated in HCV genotypes 1,2, and 3, low levels of miR-155 were detected in Egyptian patients with HCV genotype 4. Several studies have investigated the correlation between the levels of miRNA-155 and the replication of HCV, others have evaluated miRNA-155 as a prognostic biomarker in different types of cancer. No studies have investigated the impact of miRNA-155 knockdown on HCV pediatric patients associated with childhood acute lymphoblastic leukemia (ALL). We knocked-out the miR_155a in cultured polymorphonuclear cells (PBMCs) obtained from 60 children with ALL; 30 were associated with HCV-4 infection and 30 were HCV negative. The miR_155a, HCV viral load, and cell proliferation werre assessed in treated and untreated cells using TaqMan assay quantitative polymerase chain reaction. We found that miRNA-155 was significantly upregulated by seven folds in the HCV-4 associated ALL group; while being linked to high HCV viral load and leukemic burden, miR_155a knock-out can improve the disease outcome. We conclude that miR-155 is a critical miRNA that is considered a therapeutic target in pediatric HCV leukemic patients.     

Novel serological biomarker panel using protein microarray can distinguish active TB from latent TB infection

BackgroundRapid laboratory technologies which can effectively distinguish active tuberculosis (ATB) from controls and latent tuberculosis infection (LTBI) are lacked.The objective of this study is to explore MTB biomarkers in serum that can distinguish ATB from LTBI.MethodsWe constructed a tuberculosis protein microarray containing 64 MTB associated antigens. We then used this microarray to screen 180 serum samples, from patients with ATB and LTBI, and healthy volunteer controls. Both SAM (Significance analysis of microarrays) and ROC curve analysis were used to identify the differentially recognized biomarkers between groups. Extra 300 serum samples from patients with ATB and LTBI, and healthy volunteer controls were employed to validate the identified biomarkers using ELISA-based method.ResultsAccording to the results, the best biomarker combinations of 4 proteins (Rv1860, RV3881c, Rv2031c and Rv3803c) were selected. The biomarker panel containing these 4 proteins has reached a sensitivity of 93.3% and specificity of 97.7% for distinguishing ATB from LTBI, and a sensitivity of 86% and specificity of 97.6% for distinguishing ATB from HC.ConclusionThe biomarker combination in this study has high sensitivity and specificity in distinguishing ATB from LTBI, suggesting it is worthy for further validation in more clinical samples.