Feasibility of Indirect immunofluorescence (IIF) alone as a screening method for antinuclear antibody in connective diseases in India's sub-Himalayan region

Background: For the management of connective tissue disorders (CTDs), antinuclear antibody (ANA) testing is essential, both from diagnostic and prognostic points of view. Usually, patterns obtained by ANA-IIF testing correlates to specific autoantibodies as obtained from the test for ENA (by LIA/ELISA, etc.). But to apply these data from western studies, we may need validation in the local population like our subjects in sub-Himalayan (Garhwal region) area where CTDs are common. Also, suppose ANA-IFA pattern's correlation is reliably known in our population, it can minimize the cost of managing CTDs by limiting ENA testing, which is 10 times costlier than ANA-IIF. Hence, this study was undertaken to know the specific autoantibody targets (ENA by LIA) against ANA-IIF patterns in our local population. Materials and Methods: In this retrospective cross-sectional work, serum samples of CTDs were tested for ANA by IIF (Euroimmune AG) and ENA by LIA (Euroline ANA-3G) continuously for 36 months. The manufacturer's kit insert was followed, and results were analyzed applying appropriate statistical methods. Results: Major ANA-IIF patterns were found to be associated with specific autoantibodies, for example, Nuclear homogenous with dsDNA, nucleosomes, histones; speckled pattern with nRNP/Sm, Sm, SSA/Ro-52, SSB; nucleolar pattern with Scl-70, Pm-Scl 100 and centromere pattern with CENP-B. Anticytoplasmic (ACA) are found to be linked with some ANA negative (by IIF) samples, emphasizing the need for careful observation for ACA especially where ANA is not found. Conclusions: In most subjects, specific ENA targets correlated well with ANA-IIF patterns, implying effective cost minimization in CTD management. Similar future prospective studies (with clinical data) can provide a database and reference for our population.

Avances de la bioinformática en Costa Rica: vista retrospectiva y perspectivas / Bioinformatics advances in Costa Rica: retrospective view and perspectives

Resumen Introducción: La disciplina científica de la bioinformática tiene el potencial de generar aplicaciones innovadoras para las sociedades humanas. Costa Rica, pequeña en tamaño y población en comparación con otros países de América Latina, ha ido adoptando la disciplina de manera progresiva. El reconocer los avances permite determinar hacia dónde puede dirigirse el país en este campo, así como su contribución a la región latinoamericana. Objetivo: En este manuscrito se reporta evidencia de la evolución de la bioinformática en Costa Rica, para identificar debilidades y fortalezas que permitan definir acciones a futuro. Métodos: Se realizaron búsquedas en bases de datos de publicaciones científicas y repositorios de secuencias, así como información de actividades de capacitación, redes, infraestructura, páginas web y fuentes de financiamiento. Resultados: Se observan avances importantes desde el 2010, incluyendo un aumento en oportunidades de entrenamiento y número de publicaciones, aportes significativos a las bases de datos de secuencias y conexiones por medio de redes. Sin embargo, ciertas áreas, como la masa crítica y la financiación requieren más desarrollo. La comunidad científica y sus patrocinadores deben promover la investigación basada en bioinformática, invertir en la formación de estudiantes de posgrado, aumentar la formación de profesionales, crear oportunidades laborales para carreras en bioinformática y promover colaboraciones internacionales a través de redes. Conclusiones: Se sugiere que para experimentar los beneficios de las aplicaciones de la bioinformática se deben fortalecer tres aspectos clave: la comunidad científica, la infraestructura de investigación y las oportunidades de financiamiento. El impacto de tal inversión sería el desarrollo de proyectos ambiciosos pero factibles y colaboraciones extendidas dentro de la región latinoamericana. Esto permitiría realizar contribuciones significativas para abordar los desafíos globales y la aplicación de nuevos enfoques de investigación, innovación y transferencia de conocimiento para el desarrollo de la economía, dentro de un marco de ética de la investigación.

Abstract Introduction: The scientific discipline of bioinformatics has the potential to generate innovative applications for human societies. Costa Rica, small in size and population compared to other Latin American countries, has been progressively adopting the discipline. Recognizing progress makes it possible to determine where the country can go in this field, as well as its contribution to the Latin American region. Objective: This manuscript reports evidence of the evolution of bioinformatics in Costa Rica, to identify weaknesses and strengths allowing future actions plans. Methods: We searched databases of scientific publications and sequence repositories, as well as information on training activities, networks, infrastructure, web pages and funding sources. Results: Important advances have been observed since 2010, such as increases in training opportunities and the number of publications, significant contributions to the sequence databases and connections through networks. However, areas such as critical mass and financing require further development. The scientific community and its sponsors should promote bioinformatics-based research, invest in graduate student training, increase professional training, create career opportunities in bioinformatics, and promote international collaborations through networks. Conclusions: It is suggested that in order to experience the benefits of bioinformatics applications, three key aspects must be strengthened: the scientific community, the research infrastructure, and funding opportunities. The impact of such investment would be the development of ambitious but feasible projects and extended collaborations within the Latin American region and abroad. This would allow significant contributions to address global challenges and the implementation of new approaches to research, innovation and knowledge transfer for the development of the economy, within an ethics of research framework.

Detection of Anti-Extractable Nuclear Antigens in Patients with Systemic Rheumatic Disease via Fluorescence Enzyme Immunoassay and Its Clinical Utility

High expression of ENA-78 in lymphatic epithelial cells associated with cholangiocarcinoma promotes lymphangiogenesis / 医学研究生学报

ObjectiveLymphatic epithelial cells (LECs) are important links involved in lymphatic metastasis in the microenvironment of cholangiocarcinoma. This study aims to detect the modulation of inflammatory factors and chemokines secreted by LECs after stimulation of cholangiocarcinoma cells, and observe the effects of highly expressed factors on lymphangiogenesis.MethodsThe culture medium of cholangiocarcinoma (RBE, HCCC9810), LECs stimulated by cholangiocarcinoma cell culture medium (CCM), and normal LECs were prepared. Inflammatory factors and chemokines in the culture medium were detected using protein chip. The experiments are divided into the following groups, including a blank control group, CCM group, CCM coupled with Anti-ENA-78 group, Anti-ENA-78 group, ENA-78 group, ENA-78 coupled with SB2252002, and SB225002 group. The relationship between the content of factor and time was investigated using ELISA, while the relation between target factors and lymphangiogenesis obtained by cell proliferation and tubule formation assay.ResultsWe found ENA-78, IP-10, GCP-2, MCP-2, MCP-3, MIP-3a, HCC-1, and Lymphotactin expression increased in LECs supernatant after CCM stimulation. However, I-TAC, MIP-1d, IL-10, MIG, PDGF-BB, and CXCL16 factors showed down-regulation. The secretion of ENA-78 in CCM was relatively low. By ELISA, we found that the ENA-78 protein in RBE-LECs and HCCC9810-LECs gradually increased over time, and reached the plateau phase at the point of 48h. The lymphatic tube forming ability of LECs cultured in CCM was significantly increased compared with that of the control group, and this ability could be partially weakened by ENA-78 neutralizing antibodies. In the exogenous ENA-78 protein group, the lymphatic tube formation ability was as well significantly increased compared with that in the control group, and this ability could be effectively blocked by the IL-8B inhibitor.ConclusionThe increased secretion ENA-78 of lymphatic epithelial cells induced by cholangiocarcinoma may play a role in promoting lymphangiogenesis through the IL-8B receptor.

High expression of ENA-78 in lymphatic epithelial cells associated with cholangiocarcinoma promotes lymphangiogenesis / 医学研究生学报

ObjectiveLymphatic epithelial cells (LECs) are important links involved in lymphatic metastasis in the microenvironment of cholangiocarcinoma. This study aims to detect the modulation of inflammatory factors and chemokines secreted by LECs after stimulation of cholangiocarcinoma cells, and observe the effects of highly expressed factors on lymphangiogenesis.MethodsThe culture medium of cholangiocarcinoma (RBE, HCCC9810), LECs stimulated by cholangiocarcinoma cell culture medium (CCM), and normal LECs were prepared. Inflammatory factors and chemokines in the culture medium were detected using protein chip. The experiments are divided into the following groups, including a blank control group, CCM group, CCM coupled with Anti-ENA-78 group, Anti-ENA-78 group, ENA-78 group, ENA-78 coupled with SB2252002, and SB225002 group. The relationship between the content of factor and time was investigated using ELISA, while the relation between target factors and lymphangiogenesis obtained by cell proliferation and tubule formation assay.ResultsWe found ENA-78, IP-10, GCP-2, MCP-2, MCP-3, MIP-3a, HCC-1, and Lymphotactin expression increased in LECs supernatant after CCM stimulation. However, I-TAC, MIP-1d, IL-10, MIG, PDGF-BB, and CXCL16 factors showed down-regulation. The secretion of ENA-78 in CCM was relatively low. By ELISA, we found that the ENA-78 protein in RBE-LECs and HCCC9810-LECs gradually increased over time, and reached the plateau phase at the point of 48h. The lymphatic tube forming ability of LECs cultured in CCM was significantly increased compared with that of the control group, and this ability could be partially weakened by ENA-78 neutralizing antibodies. In the exogenous ENA-78 protein group, the lymphatic tube formation ability was as well significantly increased compared with that in the control group, and this ability could be effectively blocked by the IL-8B inhibitor.ConclusionThe increased secretion ENA-78 of lymphatic epithelial cells induced by cholangiocarcinoma may play a role in promoting lymphangiogenesis through the IL-8B receptor.

A study on the correlation between serum ENA -78 expression and pelvic inflammatory adhesion and pain grading among patients with endometriosis / 预防医学

Objective To observe the changes of ENA -78 expression levels in patients with endometriosis (EM)before and after treatment,and the role of ENA -78 expression in pain and pelvic inflammation.Methods A total of 160 patients with EMwere selected as the research subjects for I phase,II phase,III phase,IV phase patients.In the same period,40 cases of non EMpatients were selected as control group.Serum ENA -78 expression levels were detected among patients with EM before treatment,3 months after treatment and 6 months after treatment,and the control group before treatment by the method of ELISA .The correlation between ENA -78 expression and pelvic inflammatory adhesion and pain grading among patients with EMwas analyzed.Results There was significant difference in the level of serum ENA -78 expression between the two groups at different time points (P <0.01).The expression level of serum ENA -78 of case group decreased with time (P <0.01),while the expression level of serum ENA -78 of EM patients with different pain grading was different (P <0.05).The expression level of ENA -78 in severe group was higher than that in mild group and moderate group (P <0.05).The degree of pelvic adhesion in EMpatients was positively correlated with the level of ENA -78 expression (rs =0.675,P =0.000).The expression of ENA -78 in serum of EMpatients with different degrees of pelvic inflammation was different (P <0.05 ),and the expression of V -grade group ENA -78 was higher than other <br> groups (P <0.05).The level of ENA -78 expression was positively correlated with the degree of pain in patients with EM (rs =0.601,P =0.000).Conclusion Serum ENA -78 expression levels among EM patients were high,and positively correlated with pelvic inflammatory adhesion and pain grading.Serum ENA -78 decreased after GnRh -a treatment.

Expressions and significance of CXC chemotactic factors about GROα, ENA-78 and NAP-2 at rat asthma / 中华微生物学和免疫学杂志

Objective To observe the expressions of grouth-related oncogen (GRO)α, epithelial neutrophil activating protein-78 (ENA-78) and neutrophil-activating peptide-2 (NAP-2) of rat asthma. And to investigate the role of neutrophil in the pathogenesis of asthma exacerbation. Methods In this experi-ment, the rat model of asthma were randomly divided into two groups on average, including asthma group and control group. Levels of ENA-78 at blood neutrophil were detected by flow cytometry method. The ex-pressions of GROα protein at bronchial wall and NAP-2 protein at blood neutrophil were detected by immuno-histochemieal method. Results Levels of GROα, ENA-78 and NAP-2 proteins in asthma group [0.138 ±0.009(A value), 97.65±13.99(MFI), 0.198±0.016(A value), respectively]were significantly higher than those in control group[0.077±0.010(A value), 50.79±8.66(MFI), 0.079±0.015(A value), re-spectively], all P < 0.01. Conclusion Levels of GROα, ENA-78 and NAP-2 were increased at rat asth-ma. They may be participate in inflammation of asthma exacerbation. Neutrophil may promote inflammatory cells influxing into airway wall via increasing synthesis of CXC chemotactic factors.

Detection of Anti-ENA and anti-dsDNA Antibodies Using Line Immunoassay in Systemic Autoimmune Diseases / 대한진단검사의학회지

BACKGROUND: Detection of antibodies to extractable nuclear antigens (ENAs) and dsDNA is needed for the diagnosis of and predicting prognosis in systemic autoimmune diseases. Recently introduced line immunoassay (LIA) has the advantage of detecting several autoantibodies simultaneously, and we evaluated its usefulness in the diagnosis of autoimmune diseases in comparison with enzyme-linked immunosorbent assay (ELISA). METHODS: Samples were collected from 437 patients referred by rheumatologists. FANA (fluorescent antinuclear antibody) test and LIA for the detection of 13 different autoantibodies, including 6 ENAs and dsDNA were performed. LIA-positive samples for ENA or dsDNA antibodies were further tested with ELISA. Final diagnosis was made by rheumatologists according to the diagnostic criteria. Agreement of results between LIA and ELISA was analyzed in 53 selected patients with systemic autoimmune diseases. RESULTS: The LIA detected antibodies to ENA and dsDNA in 118 and 22 patients, respectively, and ELISA detected 70.3% (83/118) and 45.5% (10/22) of LIA positive samples. Especially, 60.2% (71/118) of patients with positive ENA antibody on LIA was diagnosed as systemic autoimmune diseases. Patients having strong FANA titer and homogenous/speckled pattern showed higher prevalence of autoantibodies, but a small proportion of FANA negative patients also showed positive reactivity (LIA 10.8%, ELISA 5.2%). LIA showed a good agreement with ELISA for the anti-ENA antibodies (> or =80%), and a lower agreement for the anti-dsDNA antibody (67.9%). CONCLUSIONS: LIA detecting several autoantibodies simultaneously might replace ELISA for anti-ENA antibodies, but not for anti-dsDNA antibodies. When LIA is performed considering clinical manifestations and FANA, it could contribute to the diagnosis of systemic autoimmune disease.

Research of enalapril on rat myocardial fibrosis and its mechanism / 中国药理学通报

Aim To investigate the effects of enalapril(Ena) on cardiac fibrosis induced by isoprenaline(Iso) and to explore its mechanism.Methods Effect of Ena and captopril(Cap) on collagen content,HW/BW,LVI,hydroxyprolnie(Hyp) level,and TGF-?1 protein expression was observed with IH and WB in rats induced by Iso subcutaneous injection.Results Different doses of Ena could decrease HW/BW,LVI and Hyp level dramatically,and decrease TGF-?1 protein expression which was closely related to myocardial fibrosis(MF).Conclusion Enalapril can inhibit TGF-?1 protein expression,by which inhibit myocardial fibrosis.

The role of chemokines ENA-78 in endometriosis / 医学研究生学报

Endometriosis is a common gynecologic disorder defined by the presence,growth,and invasion of endometrial tissue outside the uterine cavity.Although many hypotheses have been proposed,the etiology of the disease remains an enigma.Recent studies have suggested that factors within the peritoneal fluid contribute to endometriotic implant invasion,neovascularization,and proliferation.Chemokines,especially ENA-78,similar to IL-8,are postulated to play an essential role in the local inflammatory reactions seen in endometriosis.

GRO-alpha, IL-8 and ENA-78: Expressed by Stimulated Endothelial Cells and Increased PMN Adhesion / 결핵

BACKGROUND: Inflammation, where vascular endothelial cells are activated by cytokines, recruits circulating leukocytes such as neutrophils into the tissues. Mononuclear phagocytes as well as tissue cells activated by these stimuli produce these chemokines. In this study, the effects of IL-1 and LPS on the expression of CXC chemokines such as GRO-alpha, IL-8 and ENA-78 in vascular endothelial cells and the neutrophil adnesion effects of ENA-78 and GRO-alpha was investigated. METHODS: Human umbilical vein endothelial cells were cultured and stimulated with various concentrations of IL-1 and LPS. The concentrations of the GRO-alpha, IL-8 and ENA-78 secreted were measured using enzyme-linked immunosorbent assay. The effects of ENA-78 and GRO-alpha on neutrophil adhesion to the endothelial cells were also investigated. RESULTS: The addition of IL-1 and LPS to the vascular endothelial cells induced GRO-alpha, IL-8 and ENA-78 secretion in a time- and dose-dependent manner. The neutrophil adhesion was also increased by induction of ENA-78 and GRO-alpha to the vascular endothelial cells in a dose-dependent manner. CONCLUSION: CXC chemokines such as GRO-alpha, IL-8 and ENA-78 secreted by the vascular endothelial cells play an important role in the acute inflammatory responses by stimulating neutrophil adhesion to the vascular endothelial cells, raising the possibility that the CXC chemokines are one of the targets in the clinical application of acute inflammation.

Detection of Autoantibodies for Extractable Nuclear Antigens by LG Immunoblot Kit / 대한임상병리학회지

BACKGROUND: Identification of antibodies recognizing extractable nuclear antigens (ENAs) is use-ful in the diagnosis and characterization of a variety of connective tissue diseases. Recently, LG ENA Immunoblot (LGCI, Seoul, Korea) was introduced for detecting various autoantibodies to ENAs simultaneously. Performance of this kit was evaluated in this study. METHODS: Sera from 108 SLE patients and 103 RA patients were tested for the presence of spe-cific autoantibodies to ENAs by LG ENA Immunoblot and DID. Concordance rates in each autoan-tibody were obtained. After discordant results were resolved by EIA (ENA ELISA TEST SYSTEM, Zeus Scientific Inc., NJ, USA) and western blot (ANA Western Blot Immunoassay, IMMCO Diag-nostics Inc., NY, USA), sensitivity and specificity of LG ENA Immunoblot were evaluated. Between-day precision was also tested. RESULTS: Concordance rates in each autoantibody in two methods were as follows: anti-RNP (88.0%, 95/108; 100%, 103/103), anti-Sm (87.0%, 94/108; 97.1%, 100/103), anti-SSA (94.4%, 102/108; 99.0%, 102/103), anti-SSB (97.2%, 105/108; 98.1%, 101/103), anti-Scl70 (99.1%, 107/108; 100%, 103/103) in SLE and RA patients, respectively. Sensitivity and specificity of Immunoblot were 92.0% and 99.6% for anti-RNP, 100% and 99.6% for anti-Sm, 100% and 98.6% for anti-SSA, 90.0% and 98.5% for anti-SSB, and 100% and 100% for anti-Scl70, respectively. Between-day precisions were 100% in all anti-ENA antibodies. CONCLUSIONS: LG ENA Immunoblot showed good concordance rates with the conventional DID method and high sensitivity (>90%) and specificity (>98.5%) in detecting all kinds of anti-ENA autoantibodies. LG Immunoblot has another merit in that it can detect several autoantibodies simul-taneously. It is suggested that LG ENA Immunoblot can replace DID for anti-ENA detection without any problem.

Detection Of Antibodies To Ena In The Systemic Lupus Erythematosus By Immunoblotting: A Cornparision Of Irnmunoblotting And Double Immunodiffusion / 대한류마티스학회지

OBJECTIVE: Systemic Lupus Erythematosus(SLE) is characterized by various autoantibodies to a variety of nuclear antigens. Certain extractable nuclear antigens(ENA) have been served as extremly useful aids in differentiation of clinical subset, diagnostic marker and in the detection of early forms of systemic rheumatic diseases. This study was to employ immunoblot to determine the prevalences of antibodies to ENA in SLE compared with immunodiffusion. METHODS: Sera were obtained from 127 SLE patients. Antibodies to ENA were assessed by double immunodiffusion (DID) and immunoblot method. RESULTS: Using the immunblot method, the prevalences of antibodies to ENA were as follows: The antibody to Sm was 27%, UIRNP 33.6%, Ro 41.8%, La 14%, ribosomal P 14% and Ku 4%. The prevalences of antibodies to ENA by DID were as follows: The antibody to Sm was 15%, RNP 24.5%, Ro 54.3% and La 9.4%. CONCLUSIONS: Compared with immunodiffusion, results using immunoblot showed greater sensitivity in the detection of autoantibodies to ENA in SLE.