Serum Of Patients Research Articles

Exosome-transported circ_0061407 and circ_0008103 play a tumour-repressive role and show diagnostic value in non-small-cell lung cancer

BackgroundCircular RNAs (circRNAs), one of the major contents of exosomes, have been shown to participate in the occurrence and progression of cancers. The role and the diagnostic potential of exosome-transported circRNAs in non-small-cell lung cancer (NSCLC) remain largely unknown.MethodsThe NSCLC-associated exosomal circ_0061407 and circ_0008103 were screened by circRNA microarray. The role of circ_0061407 and circ_0008103 in NSCLC was examined in vitro and in vivo. The encapsulation of the two circRNAs into exosomes and the transport to recipient cells were observed by confocal microscopy. The effects of exosome-transported circ_0061407 and circ_0008103 on recipient cells were investigated using a co-culture device. Bioinformatics analyses were performed to predict the mechanisms by which circ_0061407 and circ_0008103 affected NSCLC. The quantitative polymerase chain reaction was used to quantify the exosome-containing circ_0061407 and circ_0008103 in the serum samples of healthy, pneumonia, benign lung tumours, and NSCLC. The diagnostic efficacy was evaluated using receiver operating characteristic curves.ResultsThe levels of circ_0061407 and circ_0008103 within exosomes were down-regulated in the serum of patients with NSCLC. The up-regulation of circ_0061407 and circ_0008103 inhibited the proliferation, migration/invasion, cloning formation of NSCLC cells in vitro and inhibited lung tumour growth in vivo. Circ_0061407 and circ_0008103 were observed to be packaged in exosomes and transported to recipient cells, where they inhibited the proliferation, migration/invasion, and cloning formation abilities of the recipient cells. Moreover, circ_0061407 and circ_0008103 might be involved in the progression of NSCLC by interacting with microRNAs and proteins. Additionally, lower serum exosomal circ_0061407 and circ_0008103 levels were associated with advanced pathological staging and distant metastasis.ConclusionsThis study identified two novel exosome-transported circRNAs (circ_0061407 and circ_0008103) associated with NSCLC. These findings may provide additional insights into the development of NSCLC and potential diagnostic biomarkers for NSCLC.

Elevated serum vasohibin-1 levels in atopic dermatitis: Implications for disease chronicity.

Atopic dermatitis (AD) is often characterized by chronic skin changes of dermal fibrosis, typically regulated by inflammatory and angiogenic factors. However, the significance of angiogenesis inhibitory factors in the development of AD is poorly understood. The present study investigated the potential role of an angiogenesis inhibitory factor, vasohibin-1 (VASH1), in AD by evaluating serum and skin VASH1 levels and their correlation with clinical features. The results showed that VASH1 expression levels in both the serum and skin of patients with AD were significantly elevated compared to healthy controls. Immunohistochemical staining of AD skin showed increased VASH1 expression in dermal vascular endothelial cells. Notably, there was a significant correlation between serum VASH1 levels and disease duration as well as VASH1 and vascular endothelial growth factor A expression levels in the skin tissue of patients with AD. These results may suggest a pathogenesis of increased angiogenesis and associated elevated inhibitory processes accompanying inflammation in the chronic phase of AD.

Milk Fat Globule Proteins Are Relevant Bovine Milk Allergens in Patients with α‐Gal Syndrome

Alpha-gal syndrome (AGS) is a mammalian meat allergy associated with tick bites and specific IgE to the oligosaccharide galactose-α-1,3-galactose (α-gal). Recent studies have shown that 10-20% of AGS patients also react to the dairy proteins. Considering the already described role of the meat lipid fraction in AGS manifestations, the aim of this work has been to investigate whether the milk fat globule proteins (MFGPs) could be involved in AGS. The MFGPs are extracted and their recognition by the IgE of AGS patients is proved through immunoblotting experiments. The identification of the immunoreactive proteins by LC-HRMS analysis allows to demonstrate for the first time that butyrophillin, lactadherin, and xanthine oxidase (XO) are α-gal glycosylated. The role of xanthine oxidase seems to be prevalent since it is highly recognized byboth the anti-α-gal antibody and AGS patient sera. The results obtained in this study provide novel insights in the characterization of α-Gal carrying glycoproteins in bovine milk, supporting the possibility that milk, especially in its whole form, may give reactions in AGS patients. Although additional factors are probably associated with the clinical manifestations, the avoidance of milk and milk products should be considered in individuals with AGS showing symptoms related to milk consumption.

Measurement of Serum Vitamins B2 and B6 in Patients with Hashimoto’s Thyroiditis by LC–MS/MS Method

AbstractVitamin B2 and vitamin B6 are cofactors and coenzymes in many biochemical reactions, and their sufficient serum concentrations may be crucial for the proper functioning of the thyroid gland. The aim of the study was to develop and validate a novel high-performance liquid chromatography–tandem mass spectrometry (LC–MS/MS) method for the determination of vitamin B2 and vitamin B6 in the serum of patients with Hashimoto’s thyroiditis (HT). Detection of the analytes was performed on a triple-quadrupole MS with multiple reaction monitoring using electrospray ionization. The mobile phase consisted of water and methanol, both containing 0.1% (v/v) formic acid. The analytes were isolated from serum by the one-step sample preparation of the trichloroacetic acid precipitation. Validation parameters, including linearity, accuracy, precision, limits of detection and quantification, and stability, were determined and fulfilled the criteria for analyzing compounds in biological fluids. The method was applied for the determination of the above vitamin concentrations in serum samples of 74 patients with HT and 33 healthy controls. The concentration of vitamin B2 was significantly lower in patients with HT compared to the control group (7.20 ± 7.86 ng/mL vs. 12.03 ± 9.62 ng/mL) and the concentration of vitamin B6 was also lower, but without a statistically significant difference (30.09 ± 19.16 ng/mL vs. 37.29 ± 23.02 ng/mL). We concluded that the vitamins concentration measurement may be helpful in the diagnosis of vitamin deficiencies and the supplementation strategy.

Evaluation of Melatonin and Antioxidant Levels in the Serum of Breast Cancer Patients

Metabolites and antioxidants can be altered in patients with various diseases, particularly in those with cancer. This study aims to measure melatonin and total antioxidant levels in the serum of breast cancer patients and compare them with a healthy control group. Both groups had serum samples collected at 2:00 a.m. and 9:00 a.m. Melatonin levels were determined using High-performance liquid chromatography (HPLC), while total antioxidant levels were assessed by Enzyme-linked immunosorbent assay (ELISA). Levels of melatonin and total antioxidants differed between the groups. In the control group, the levels of melatonin and total antioxidants were considerably greater compared to the diagnostic group, with a p-value of 0.001. Melatonin and total antioxidant levels were consistently greater at 2:00 than at 9:00. The maximum melatonin levels recorded in the diagnostic and control groups were 39.3 pg/mL and 65.9 pg/mL, respectively. Melatonin levels varied based on the age of the participants, with higher amounts observed in younger participants compared to older people in both groups. The BMI affected the levels of melatonin, with a greater BMI leading to elevated melatonin concentrations. The diagnostic group had a total antioxidant level of 39.3 pg/mL, while the control group had a level of 65.9 pg/mL. The elevated levels of melatonin and total antioxidants in the control group indicate a healthy status. The research demonstrated a significant correlation between melatonin and antioxidant levels in breast cancer patients, with reduced melatonin levels and total antioxidant levels in the diagnosed group. Elevated BMI was linked to reduced melatonin levels.

Single-sEV profiling identifies the TACSTD2 + sEV subpopulation as a factor of tumor susceptibility in the elderly

BackgroundAging is a very complex physiological phenomenon, and sEVs are involved in the regulation of this mechanism. Serum samples from healthy individuals under 30 and over 60 years of age were collected to analyze differences in sEVs proteomics.ResultsBased on PBA analysis, we found that sEVs from the serum of elderly individuals highly express TACSTD2 and identified a subpopulation marked by TACSTD2. Using ELISA, we verified the upregulation of TACSTD2 in serum from elderly human and aged mouse. In addition, we discovered that TACSTD2 was significantly increased in samples from tumor patients and had better diagnostic value than CEA. Specifically, 9 of the 13 tumor groups exhibited elevated TACSTD2, particularly for cervical cancer, colon cancer, esophageal carcinoma, liver cancer and thyroid carcinoma. Moreover, we found that serum sEVs from the elderly (especially those with high TACSTD2 levels) promoted tumor cell (SW480, HuCCT1 and HeLa) proliferation and migration.ConclusionTACSTD2 was upregulated in the serum of elderly individuals and patients with tumors, and could serve as a dual biomarker for aging and tumors.

Exosomal YB-1 facilitates ovarian restoration by MALAT1/miR-211-5p/FOXO3 axis

Premature ovarian failure (POF) affects many adult women less than 40 years of age and leads to infertility. Mesenchymal stem cells-derived small extracellular vesicles (MSCs-sEVs) are attractive candidates for ovarian function restoration and folliculogenesis for POF due to their safety and efficacy, however, the key mediator in MSCs-sEVs that modulates this response and underlying mechanisms remains elusive. Herein, we reported that YB-1 protein was markedly downregulated in vitro and in vivo models of POF induced with H2O2 and CTX respectively, accompanied by granulosa cells (GCs) senescence phenotype. Notably, BMSCs-sEVs transplantation upregulated YB-1, attenuated oxidative damage-induced cellular senescence in GCs, and significantly improved the ovarian function of POF rats, but that was reversed by YB-1 depletion. Moreover, YB-1 showed an obvious decline in serum and GCs in POF patients. Mechanistically, YB-1 as an RNA-binding protein (RBP) physically interacted with a long non-coding RNA, MALAT1, and increased its stability, further, MALAT1 acted as a competing endogenous RNA (ceRNA) to elevate FOXO3 levels by sequestering miR-211-5p to prevent its degradation, leading to repair of ovarian function. In summary, we demonstrated that BMSCs-sEVs improve ovarian function by releasing YB-1, which mediates MALAT1/miR-211-5p/FOXO3 axis regulation, providing a possible therapeutic target for patients with POF.

The sclerotic component of metabolic syndrome: Fibroblast activities may be the central common denominator driving organ function loss and death.

Fibrosis is a common feature of more than 50 different diseases and the cause of more than 35% of deaths worldwide, of which liver, kidney, skin, heart and, recently, lungs are receiving the most attention. Tissue changes, resulting in loss of organ function, are both a cause and consequence of disease and outcome. Fibrosis is caused by an excess deposition of extracellular matrix proteins, which over time results in impaired organ function and organ failure, and the pathways leading to increased fibroblast activation are many. This narrative review investigated the common denominator of fibrosis, fibroblasts, and the activation of fibroblasts, in response to excess energy consumption in liver, kidney, heart, skin and lung fibrosis. Fibroblasts are the main drivers of organ function loss in lung, liver, skin, heart and kidney disease. Fibroblast activation in response to excess energy consumption results in the overproduction of a range of collagens, of which types I, III and VI seem to be the essential drivers of disease progression. Fibroblast activation may be quantified in serum, enabling profiling and selection of patients. Activation of fibroblasts results in the overproduction of collagens, which deteriorates organ function. Patient profiling of fibroblast activities in serum, quantified as collagen production, may identify an organ death trajectory, better enabling identification of the right treatment for use in different metabolic interventions. As metabolically activated patients have highly elevated risk of kidney, liver and heart failure, it is essential to identify which organ to treat first and monitor organ status to correct treatment regimes. In direct alignment with this, it is essential to identify the right patients with the right organ deterioration trajectory for enrolment in clinical studies.

Dominant Circulating Cell-free Mycobacterial Proteins in in-use Machining Fluid and their Antigenicity Potential.

Occupational exposure to industrial Metalworking Fluid (MWF) colonized by Mycobacterium immunogenum (MI) has been associated with immune lung disease hypersensitivity pneumonitis (HP) in machinists. This warrants regular fluid monitoring for early detection of mycobacterial proteins, especially those with antigenic potential. To detect and identify dominant MI proteins and antigens directly from the field-drawn in-use MWF using an integrated immunoproteomic and immunoinformatic approach. An MI-positive MWF selected by DNA-based screening of several field-drawn MWF samples were cultured to isolate the colonizing strain and profiled for dominant circulating cell- free (ccf) MI proteins, including antigens using an integrated immunoproteomic (1D- and 2Dgel fractionation of seroreactivity proteins combined with shotgun proteomic analysis using LC-MS/ MS) and immunoinformatic strategy. A new MI strain (MJY-27) was identified. The gel fractionated MI protein bands (1Dgel) or spots (2D-gel) seroreactive with anti-MI sera probes (Rabbit and Patient sera) yielded 86 MI proteins, 29 of which showed peptide abundance. T-cell epitope analysis revealed high (90-100%) binding frequency for HLA-I& II alleles for 13 of the 29 proteins. Their antigenicity analysis revealed the presence of 6 to 37 antigenic determinants. Interestingly, one of the identified candidates corresponded to an experimentally validated strong B- and T-cell antigen (AgD) from our laboratory culture-based studies. This first report on dominant proteins, including putative antigens of M. immunogenum prevalent in field in-use MWF, is a significant step towards the overall goal of developing fluid monitoring for exposure and disease risk assessment for HP development in machining environments.

Serum exosomes miR-206 and miR-549a-3p as potential biomarkers of traumatic brain injury

Traumatic brain injury (TBI) is one of the leading causes of death and disability worldwide. However, effective diagnostic, therapeutic and prognostic biomarkers are still lacking. Our research group previously revealed through high-throughput sequencing that the serum exosomes miR-133a-3p, miR-206, and miR-549a-3p differ significantly in severe TBI (sTBI), mild or moderate TBI (mTBI), and control groups. However, convincing experimental evidence is lacking. To solve this problem, we used qPCR in this study to further verify the expression levels of serum exosomes miR-133a-3p, miR-206 and miR-549a-3p in TBI patients. The results showed that the serum exosomes miR-206 and miR-549a-3p showed good predictive value as biomarkers of TBI. In addition, in order to further verify whether serum exosomes miR-206 and miR-549a-3p can be used as potential biomarkers in patients with TBI and to understand the mechanism of their possible effects, we further determined the contents of SOD, BDNF, VEGF, VEGI, NSE and S100β in the serum of TBI patients. The results showed that, serum exosomes miR-206 and miR-549a-3p showed good correlation with BDNF, NSE and S100β. In conclusion, serum exosomes miR-206 and miR-549a-3p have the potential to serve as potential biomarkers in patients with TBI.

Abstract PO3-19-04: The CROWN STUDY: A multicenter prospective cohort study of cardiovascular outcomes in premenopausal women with hormone receptor positive breast cancer treated with ovarian suppression and an aromatase inhibitor

Abstract Background: Treatment for premenopausal women with high or intermediate risk hormone receptor (HR)+ breast cancer (BC) includes the concurrent use of ovarian function suppression (OFS) and an aromatase inhibitor (AI) to induce near complete estrogen deprivation (NCED). The long-term cardiovascular (CV) sequela for women treated with NCED is unknown. Premature menopause in non-cancer populations is associated with CV disease. The CROWN study (CaRdiac Outcomes With Near complete estrogen deprivation) will use sophisticated imaging assessments of cardiac dysfunction to understand the evolution of CV injury, as well as biomarker and demographic correlates, with the goal of developing tools to assess and mitigate CV risk. Methods: This is a NIH funded prospective cohort study conducted at 3 regional NCI-supported Cancer Centers (Atrium Health Wake Forest Baptist, Virginia Commonwealth and Duke) that will include premenopausal women, age ≤ 55, with Stage I-III BC following completion of planned chemotherapy, surgery and radiation with an ECOG 0-1. HR+ BC patients will receive an AI and OFS. Women with HR- BC are included as comparators. CV imaging and biomarkers will be obtained at baseline, 1 year and 2 years (Table 1). These assessments will include serial cardiac magnetic resonance (CMR) and coronary computed tomography angiography (CCTA) imaging as well as laboratory measurements, including exploratory biomarkers. The primary objective is to determine the 24-month difference in stress myocardial blood flow as measured by adenosine CMR imaging in both groups. We will correlate CMR imaging with CCTA to provide complementary detail of coronary plaque changes. The study will also assess the relevance of pre-existing risk factors on study outcomes, including an emphasis on racial disparities, and dynamic change in modifiable and treatment related risk factors. We plan to enroll 90 women, 67 in the NCED group and 23 in the HR-group, allowing for a 10% drop out rate. There are two primary types of statistical analyses. The first includes testing hypotheses between group (NCED vs HR-) and within group (longitudinal changes within the NCED group). The second analyses, involve developing predictive equations utilizing a stepwise linear regression approach to determine if patient demographics, clinical parameters and serum biomarkers are associated with cardiovascular changes over time. A total of 17 patients are enrolled as of 07/2023. NCT05309655 Study Procedures XX- XX Citation Format: Susan Dent, Emily Douglas, Sarah Hatcher, Katherine Ansley, Carolyn Park, Nathaniel O'Connell, Mary Hackney, Ralph D'Agostino, Rani Bansal, Renee Avecilla, Monica Brown, Kacey Clayton-Stiglbauer, Igor Klem, Jennifer Jordan, Alexandra Thomas. The CROWN STUDY: A multicenter prospective cohort study of cardiovascular outcomes in premenopausal women with hormone receptor positive breast cancer treated with ovarian suppression and an aromatase inhibitor [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO3-19-04.

Abstract PO1-13-04: Distinct immune signatures discriminate between patients defined as exceptional survivors of metastatic breast cancer compared to those with rapidly progressing treatment resistant breast cancers

Abstract Background: "Atypical responders" can encompass three sub- categories of patients: "exceptional responders" (those with an unusually favourable treatment response), "rapid progressors" (unusually poor or no response), and "exceptional/long-term survivors" (outlived initial prognosis). Here, we aim to investigate the drivers of immune surveillance mechanisms (local, lymphoid and peripheral) and the nature of immunological tumour recognition in exceptional survivors of stage 4 metastatic disease following standard-of-care chemo- and targeted therapies. Methods: We conducted a complementary multi-platform immune profiling study to define differences in the phenotypic immune landscape including mass-spectrometry-based proteomics on peripheral blood mononuclear cells (PBMCs), multiplex cytokine profiling of patient serum, and high-dimensional flow cytometry on whole blood samples. By staining 500μl of whole blood using 2 panels, 353 immune-cell-related parameters were obtained for comparisons among the following groups: "exceptional survivors," "exceptional responders," "rapid progressors," "non-exceptional metastatic patients," "early breast cancer patients," and "healthy volunteers". Patients were matched for age, breast cancer receptor status, and sites of metastases where possible. Results: Preliminary analysis of serum biomarkers and proteome of the PBMCs of the exceptional survivors showed elevated levels of NK-cell-related proteins such as KCTD10 and RBBP7. Flow cytometry analysis revealed the presence of increased activated CD56dim NK cells, CD56bright NK cells, and central memory (CM) (CD45RA-CD27+) CD8 T cells in both the exceptional survivors and the rapid progressors. However, expression of CD25 appears to be the main mode of activation in rapid progressors compared with increased NKG2D expression in exceptional survivors. Additionally, activation of the unconventional gd T cells was evident in both groups with terminally differentiated effector memory cells re-expressing CD45RA (TEMRA) Vd1 cell predominating in the exceptional survivors. In the rapid progressors, the increased CD25+ TEMRA Vd2 cells coupled with elevated serum IL-17A and IL-1b suggest their ability to generate IL-17. Th2 cytokines, IL-5 and IL-13 were also enriched in rapid progressors and concomitant increase in CD25+ Th2 cells reveals a strong Th2-driven immune signature in these patients. Finally increased CD86+ atypical double negative (DN) B cells, mostly comprised of the DN2 subset were also significantly enriched in the rapid progressors compared to the exceptional survivors where DN1 B cells were prevalent. Conclusion: The multi-platform approach to investigating immune responses present in atypical responders has identified several distinct immunophenotypes in which the extreme outliers differ in their potential immune surveillance mechanisms. Functional validation of these findings through activation and cytotoxicity assays is currently underway and future work aims to understand the spatial biology of these circulating immune cells within the context of their primary tumours, metastases, and lymph nodes. Citation Format: Helen Kakkassery, Thanussuyah Alaguthurai, Zuza Kozik, Paul Buckley, Ruifang Lu, Esme Carpenter, Rosalind Graham, Gheed Alhity, Farhana Hossain, Sadiyah Mukhtar, Syed Haider, Jyoti Choudhary, Sheeba Irshad. Distinct immune signatures discriminate between patients defined as exceptional survivors of metastatic breast cancer compared to those with rapidly progressing treatment resistant breast cancers [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-13-04.

A novel indirect ELISA for serodiagnosis of mucormycosis using antigens from Rhizopus arrhizus.

Due to a delay in diagnosis by conventional techniques and high mortality, the development of a standardised and rapid non-culture-based technique is an unmet need in pulmonary, gastrointestinal, and disseminated forms of mucormycosis. Though limited studies have been conducted for molecular diagnosis, there are no established serologic tests for this highly fatal infection. To develop and evaluate an indirect in-house enzyme-linked immunosorbent assay (ELISA) utilising antigens of Rhizopus arrhizus for detecting anti-Rhizopus antibodies (IgG and IgM) in sera of patients with mucormycosis. We extracted both secretory and mycelial Rhizopus antigens using standardised protocols. Bradford assay was used for protein quantification. We then standardised an indirect ELISA using R. arrhizus mycelial and secretory antigens (10.0 μg/mL in bicarbonate buffer pH 9.2) for detecting anti-Rhizopus IgG and IgM antibodies in patient sera. We included patients with mucormycosis, other fungal infections, and healthy controls. Antibody index value (E-value) was calculated for each patient sample. Asparagine broth culture filtrate utilising 85% ammonium sulphate salt fractionation and mycelial homogenate grown in yeast extract peptone dextrose (YPD) broth precipitated with trichloroacetic acid (TCA) yielded a large amount of good-quality protein for the assay. We included 55 patients with mucormycosis (rhino-orbito-cerebral mucormycosis [ROCM, n = 39], pulmonary [n = 15], gastrointestinal [n = 1]), 24 with other fungal infections (probable aspergillosis [n = 14], candidiasis [n = 10]), and healthy controls (n = 16). The sensitivity of the antibody test for diagnosing mucormycosis ranged from 83.6-92.7% for IgG and 72.7-87.3% for IgM, with a specificity of 91.7-92.5% for IgG and 80-82.5% for IgM. The sera from patients with other fungal infections and healthy individuals did not show significant cross-reactivity. The detection of anti-Rhizopus IgG antibody performed significantly better in comparison to IgM-based ELISA for diagnosing both ROCM (sensitivity of 84.6% vs. 69.2%) and pulmonary cases (86.6% vs. 80.0%). More extensive studies are required to confirm our findings.

Trim14-IκBα signaling regulates chronic inflammatory pain in rats and osteoarthritis patients

Chronic inflammatory pain is the highest priority for people with osteoarthritis when seeking medical attention. Despite the availability of NSAIDs and glucocorticoids, central sensitization and peripheral sensitization make pain increasingly difficult to control. Previous studies have identified the ubiquitination system as an important role in the chronic inflammatory pain. Our study displayed that the E3 ubiquitin ligase tripartite motif-containing 14 (Trim14) was abnormally elevated in the serum of patients with osteoarthritis and pain, and the degree of pain was positively correlated with the degree of Trim14 elevation. Furthermore, CFA-induced inflammatory pain rat model showed that Trim14 was significantly increased in the L3-5 spinal dorsal horn (SDH) and dorsal root ganglion (DRG), and in turn the inhibitor of nuclear factor Kappa-B isoform α (IκBα) was decreased after Trim14 elevation. After intrathecal injection of Trim14 siRNA to inhibit Trim14 expression, IκBα expression was reversed and increased, and the pain behaviors and anxiety behaviors of rats were significantly relieved. Overall, these findings suggested that Trim14 may contribute to chronic inflammatory pain by degrading IκBα, and that Trim14 may become a novel therapeutic target for chronic inflammatory pain.

Serum netrin-1 levels are high in Rheumatoid arthritis associated interstitial lung disease

BackgroundRecent data show that netrin-1 has a role in development of pulmonary fibrosis. This study was aimed to investigate serum netrin-1 level and its relation to interstitial lung disease(ILD) in patients with rheumatoid arthritis (RA). Method42 RA patients with RA-ILD, 58 RA patients without RA-ILD (RA non-ILD group), and 61 healthy volunteers were included in this study. The modified DAS28-ESR score was used to calculate disease activity in RA patients. Using the quantitative immunoassay method, Serum netrin-1 levels were measured with an ELISA kit (Catalog number: E-EL-H2328; lab science, lot number: GZWTKZ5SWK, Texas, USA). ResultsThe median value of netrin-1 was found to be significantly higher in the RA-ILD group (82.9 [59.9–124]) compared to both the RA non-ILD group(52.9 [49.5–73.1])(B = −0.006, OR = 0.994, CI 95 %=0.989–0.999, P = 0.018) and the control group(53.5 [49.5–87.5]) (B: −0.005, OR: 0.994, CI 95 %: 0.990–0.999, p: 0.022). A cut-off value of 61.78 for netrin-1 was found to have a sensitivity of 73.8 % and a specificity of 69 % for the diagnosis of RA-ILD (AUC [95 %Cl] = 0.771 [0.679–0.862], p < 0.0001).It was found that high serum netrin-1 level was strongly associated with the RA-usual interstitial pneumonia(UIP) pattern and poorly related to the RA-nonspecific interstitial pneumonia(NSIP) pattern compared to the RA non-ILD group. ConclusionsNetrin-1 is elevated in the serum of patients with RA-ILD, especially in the UIP pattern. Netrin-1 may be a potential candidate for predicting the development of RA-ILD that should be investigated in the pathophysiological and therapeutic fields.

Repetitive transcranial magnetic stimulation for post-stroke non-fluent aphasia: a systematic review and meta-analysis of randomized controlled trials

ObjectiveTo systematically evaluate the efficacy and safety of repetitive transcranial magnetic stimulation (rTMS) on language function in patients with non-fluent aphasia post-stroke.MethodsWe selected randomized clinical trials (RCT) that involved stroke patients with non-fluent aphasia, whose intervention was rTMS vs. no therapy or other therapy. Two researchers autonomously reviewed the literature based on the specified criteria for inclusion and exclusion and completed the process of data extraction, data verification, and quality evaluation. Meta-analysis was performed using RevMan 5.41 and Stata MP 172, while the assessment of risk of bias was carried out utilizing the Risk of Bias version 2 tool (RoB2)3.ResultsThe meta-analysis involved 47 RCTs, encompassing 2,190 patients overall. The indexes indicated that rTMS has the potential to decrease the severity of non-fluent aphasia in stroke patients, including improvement of the capability of repetition, naming, and spontaneous language. The determination of BDNF in the serum of patients was also increased. In addition, rTMS reduced the likelihood of depression in stroke patients.ConclusionTo summarize the relevant studies, rTMS has significant effects on improving the language abilities of stroke patients suffering from non-fluent aphasia, including the abilities of repetition, naming, and spontaneous language.

Tetraspanin immunoassay for the detection of extracellular vesicles and renal cell carcinoma

AbstractHalf of patients with renal cell carcinoma (RCC) develop metastases. New and noninvasive biomarkers are needed for the diagnosis of RCC. The study aims to develop an EV‐based assay for the detection of RCC using a highly sensitive nanoparticle‐aided time‐resolved fluorescence immunoassay (NP‐TRFIA). To confirm the presence of tetraspanins on EVs, size exclusion chromatography is used to separate EV‐ and PE‐fractions from RCC4, 786‐O, and HEK293 cell lines. EV‐ and PE‐fractions are quantified using NP‐TRFIA assays established for CD9, CD63, CD81, and CD151. Tetraspanins are measured from RCC CCM and serum samples of RCC (n = 14), benign (n = 17), and healthy (n = 9) individuals. Among the tetraspanins, CD63 exhibits 3‐5‐fold higher expression on RCC4 and 786‐O CCM compared to HEK293. A sandwich CD63‐CD63 assay demonstrates significant discrimination of RCC patients from benign (p = 0.0003), and healthy (p = 0.005) individuals, respectively. Similarly, the CD81‐CD81 assay also enables significant separation of RCC patients compared to benign (p = 0.014), and healthy (p = 0.003) controls, respectively. This suggests that RCC cell lines and serum of RCC patients show higher amounts of CD63‐ and CD81‐EVs compared to controls. Detection of these EVs using NP‐TRFIA approach may play a vital role in the detection of RCC.

Expression of 10 circulating cytokines/chemokines in HBV-related liver disease

BackgroundCytokines/chemokines play essential roles in the occurrence and progression of hepatitis B virus (HBV) infection. This study aimed to observe the expression patterns of 10 related cytokines/chemokines in the serum of healthy individuals, self-limited patients and HBV-infected patients at different stages of disease (chronic hepatitis B (CHB), liver cirrhosis (LC), hepatocellular dysplastic nodules (DNs) and hepatocellular carcinoma (HCC)) and to analyze the relationships of these cytokines/chemokines with disease progression.MethodsThe levels of six cytokines (FGF-2, IFN-α2, IL-4, IL-6, IL-10 and VEGF-A) and four chemokines (GRO-α, IL-8, IP-10 and MCP-1) were quantified using Luminex multiplex technology.ResultsThere were no significant differences in the expression of the 10 cytokines/chemokines between healthy individuals and self-limited patients. The levels of IL-4, IL-6, and IL-8 increased significantly in the CHB and LC groups. IL-10 was highly expressed in the HCC group. The level of IP-10 was significantly greater in all liver disease groups (CHB, LC, DN and HCC) than in the HI and SL-HBV groups, while the level of GRO was significantly lower in all liver disease groups than in the HI and SL-HBV groups. The levels of the 10 cytokines/chemokines were not significantly different between the preoperative group and the two-day postoperative group. Significant increases in the levels of IL-4, VEGF-A and IL-8 and significant decreases in those of IL-10 and GRO-α were observed 3 months after surgery. Correlation analysis revealed that most of the cytokines/chemokines with significant correlation differences were positively correlated before and after HCC surgery.ConclusionOur results highlight the fluctuating status of specific cytokines in HBV infection-related disease progression. It is speculated that these cytokines may be used as serum markers to monitor dynamic changes during the progression of HBV-related liver disease and to predict patient prognosis.

LncRNA Myocardial Infarction-Associated Transcript (MIAT)/miR-505-5p Axis Regulates Proliferation and Migration of Vascular Smooth Muscle Cells of Hypertension Mice

To clarify the role of long non-coding RNA (lncRNA) MIAT in regulating proliferative and migratory abilities in VSMCs extracted from hypertension mice via downregulating microRNA-505-5p (miR-505-5p). Serum levels of MIAT and miR-505-5p in enrolled 20 hypertension patients and 20 healthy volunteers were detected. VSMCs were extracted from hypertension mice and healthy mice. Regulatory effects of MIAT and miR-505-5p on proliferative and migratory abilities in VSMCs were examined. At last, the interaction between MIAT and miR-505-5p was explored by dual-luciferase reporter assay and rescue experiments. Serum level of MIAT was higher in hypertension patients than those of healthy subjects, while miR-505-5p was downregulated. MIAT level was negatively correlated to miR-505-5p level in serum of hypertension patients. Knockdown of MIAT suppressed proliferative and migratory abilities in VSMCs extracted from hypertension mice. In addition, knockdown of MIAT upregulated E-cadherin and downregulated Vimentin and Snail-1. MiR-505-5p was verified to be the target binding MIAT. Knockdown of miR-505-5p reversed regulatory effects of MIAT on VSMCs phenotypes. LncRNA MIAT stimulates VSMCs in hypertension mice to proliferate and migrate through downregulating miR-505-5p, which may be a promising target for diagnosis and treatment of hypertension.

Characterization of antigenic proteins of the Taenia solium postoncospheral form.

Neurocysticercosis is the leading cause for acquired epilepsy worldwide, and it is caused by the larval stage of the parasite Taenia solium. Several proteins of this stage have been characterized and studied to understand the parasite-host interaction, however, the proteins from the early cysticercus stages (the postoncospheral form) have not yet been characterized. The study of the postoncospheral form proteins is important to understand the host-parasite relationship in the early stages of infection. The aim of this work was to identify postoncospheral form antigenic proteins using sera from neurocysticercosis patients. T. solium activated oncospheres were cultured in HCT-8 cells to obtain the postoncospheral form. Soluble total and excretory/secretory proteins were obtained from the postoncospheral form and were incubated with both pool sera and individual serum of neurocysticercosis positive human patients. Immunoblotting showed target antigenic proteins with apparent molecular weights of 23kDa and 46-48kDa. The 46-48kDa antigen bands present in soluble total and excretory/secretory postoncospheral form proteins were analyzed by LC-MS/MS; proteins identified were: nuclear elongation factor 1 alpha, enolase, unnamed protein product/antigen diagnostic GP50, calcium binding protein calreticulin precursor and annexin. The postoncospheral form expresses proteins related to interaction with the host, some of these proteins are predicted to be exosomal proteins. In conclusion, postoncospheral proteins are consistent targets of the humoral immune response in human and may serve as targets for diagnosis and vaccines.