Epigenetic Application of ATAC-Seq Based on Tn5 Transposase Purification Technology.

Background: Assays of transposase accessible chromatin sequencing (ATAC-seq) is an efficient assay to investigate chromatin accessibility, which depends on the activity of a robust Tn5 transposase to fragment the genome while cutting in the sequencing adapters. Methods: We propose reliable approaches for purifying hyperactive Tn5 transposase by chitin magnetic bead sorting. Double-stranded DNA of J76 cells and 293T cells were digested and subjected to tagmentation as test samples with Tn5 transposase, and libraries were established and sequenced. Sequencing data was then analyzed for peak calling, GO enrichment, and motif analysis. Results: We report a set of rapid, efficient, and low-cost methods for ATAC-seq library construction and data analysis, through large-scale and rapid sequencing. These methods can provide a reference for the study of epigenetic regulation of gene expression.

Re-evaluation of the prognostic significance of oropharyngeal dysphagia in idiopathic inflammatory myopathies.

OBJECTIVE: To investigate the prognostic significance of videofluorographic swallowing study (VFSS)-confirmed oropharyngeal dysphagia in idiopathic inflammatory myopathies (IIMs). METHOD: We reviewed the medical records of patients who were diagnosed with IIM between 2009 and 2020 at Seoul St Mary's Hospital. All oropharyngeal dysphagia cases were limited to VFSS-confirmed dysphagia found during the initial diagnostic work-up for IIM. We described the findings on VFSS and the course of the dysphagic symptoms. Logistic regression and survival analyses were performed to evaluate the risk of pneumonia and mortality, respectively. RESULTS: We found 88 patients with IIM who met the criteria. Among them, 17 patients (19%) had oropharyngeal dysphagia. Except for two cases lost to follow-up and one deceased case, all of the patients with dysphagia (14 of 14) had swallowing function restored within 6 months. The risk of pneumonia within 3 months from the diagnosis of IIM was significant [odds ratio = 4.49, 95% confidence interval (CI) 1.07-18.88]. The median follow-up duration was 34 and 27 months for the groups without and with dysphagia, respectively. The survival analysis failed to demonstrate that the presence of oropharyngeal dysphagia increased the risk of death (hazard ratio = 0.77, 95% CI: 0.085-7.00). CONCLUSIONS: Oropharyngeal dysphagia found at the initial diagnosis of IIM improved within 3-6 months in nearly all cases. Furthermore, IIM patients who had oropharyngeal dysphagia at the initial diagnosis of IIM were not likely to have shorter survival, even if the risk of pneumonia was increased in the short term.

Association between antiphospholipid antibodies and arterial thrombosis in patients with rheumatoid arthritis.

OBJECTIVES: Antiphospholipid antibodies (aPL) are present in a proportion of patients with rheumatoid arthritis but their clinical significance remains unclear. We investigated the association between aPL and thrombotic events in rheumatoid arthritis patients. METHODS: In this cross-sectional study, aPL profiles were evaluated in 376 rheumatoid arthritis patients in accordance with the standard guidelines. Clinical and radiographic data were retrospectively collected. RESULTS: aPL were identified in 39 patients (10.4%). Lupus anticoagulant was the most common subtype (n = 25, 6.6%); anti-cardiolipin antibodies and anti-ß2 glycoprotein I antibodies were detected in six and 12 patients (1.6% and 3.2%), respectively. Compared to the aPL-negative group, aPL-positive patients included more male patients (41.0% vs. 15.4%, P < 0.001) and more smokers (41.0% vs. 16.0%, P = 0.001). There was no difference between the two groups in age, disease duration and body mass index, or the frequency of diabetes, hypertension or dyslipidaemia. Of note, arterial thromboses were more common in the aPL-positive than the aPL-negative group (12.8% vs. 2.1%, P = 0.004), whereas the frequency of venous thrombosis did not differ between the two groups (0.0% vs. 0.9%, P = 1.000). On multivariate regression analysis, aPL, age, hypertension, dyslipidaemia and baseline C-reactive protein level were independently associated with arterial thrombotic events (all P values < 0.05). CONCLUSION: aPL was found in a subset of rheumatoid arthritis patients, who were more often smokers, and aPL was independently associated with development of arterial thrombosis. This result suggests that aPL may contribute to an increased risk of arterial thrombosis in rheumatoid arthritis patients.

Prediction of subclinical atherosclerosis by serum osteoprotegerin in premenopausal women with systemic lupus erythematous: correlation of osteoprotegerin with monocyte chemotactic protein-1.

OBJECTIVE: Patients with systemic lupus erythematosus (SLE) have increased risk for cardiovascular disease. Previous studies disclosed the association of serum osteoprotegerin (OPG) with the presence of symptomatic atherosclerosis in the general population and several disease conditions. We thus investigated the association between serum OPG levels and subclinical atherosclerosis in premenopausal SLE patients. METHODS: Serum OPG levels and carotid artery intima-media thickness (IMT) were measured in 181 premenopausal SLE patients and age-matched 85 control subjects. Traditional cardiovascular risk factors and SLE-related factors were analyzed. RESULTS: Patients with SLE had significantly increased serum OPG levels (1086 versus 517 pg/ml, p < 0.001) and carotid IMT (0.63 versus 0.45 mm, p < 0.001) compared with control subjects. Carotid IMT significantly increased across the quartiles of OPG. Logistic regression analysis revealed that compared to the lowest OPG quartile, the odds ratio (OR, 95% confidence interval) for increased carotid IMT in quartile 2, 3, and 4 was 1.126 (1.013-1.801), 1.562 (1.268-2.799), and 4.460 (1.126-7.128), respectively, after multiple adjustments (p for trend across quartiles < 0.001). These associations remained significant after further adjustment for inflammatory parameters. Interestingly, serum monocyte chemotactic protein-1 (MCP-1) levels were positively correlated with serum OPG levels (γ = 0.332, p < 0.001). Parallel analysis showed that serum MCP-1 was also an independent predictor of carotid IMT incrassation, but this association was lost when serum OPG was included in the model. CONCLUSION: Serum OPG levels were increased and correlated with serum MCP-1 levels in premenopausal SLE patients. Increased serum OPG was independently associated with subclinical atherosclerosis in these patients.

Ab initio investigation of the abstraction reactions by H and D from tetramethylsilane and its deuterated substitutions.

Thermal rate constants for chemical reactions using the corrections of zero curvature tunneling (ZCT) and of small curvature tunneling (SCT) methods are reported. The general procedure is implemented and used with high-quality ab initio computations and semiclassical reaction probabilities along the minimum energy path (MEP). The approach is based on a vibrational adiabatic reaction path and is applied to the H + Si(CH3)4 → H2 + Si(CH3)3CH2 reaction and its isotopically substituted variants. All of the degrees of freedom are optimized, and harmonic vibrational frequencies and zero-point energies are calculated at the MP2(full) level with the cc-pVTZ basis set. Single-point energies are calculated at a higher level of theory with the same basis set, namely, CCSD(T,full). The influence of the basis set superposition error (BSSE) on the energetics is tested. The method is further exploited to predict primary and secondary kinetic isotope effects (KIEs and SKIEs, respectively). Rate constants computed with the ZCT and SCT methods over a wide temperature range (180-2000 K) show important quantum tunneling effects at low temperatures when compared to rates obtained from the purely classical transition-state theory (TST) and from the canonical variational transition state theory (CVT). For the H + Si(CH3)4 reaction, they are given by the following expressions: k(TST/ZCT) = 9.47 × 10(-19) × T(2.65) exp(-2455.7/T) and k(CVT/SCT) = 7.81 × 10(-19) × T(2.61) exp[(2704.2/T) (in cm(3) molecule(-1) s(-1)). These calculated rates are in very good agreement with those from available experiments.

Analysis and prediction of 5-year survival in patients with cutaneous melanoma: a model-based period analysis.

Background: The survival and prognosis of patients are significantly threatened by cutaneous melanoma (CM), which is a highly aggressive disease. It is therefore crucial to determine the most recent survival rate of CM. This study used population-based cancer registry data to examine the 5-year relative survival rate of CM in the US. Methods: Period analysis was used to assess the relative survival rate and trends of patients with CM in the Surveillance, Epidemiology, and End Results (SEER) database during 2004-2018. And based on the data stratified by age, gender, race and subtype in the SEER database, a generalized linear model was 12established to predict the 5-year relative survival rate of CM patients from 2019 to 2023. Results: The 5-year relative survival increased to various degrees for both total CM and CM subtypes during the observation period. The improvement was greatest for amelanotic melanoma, increasing from 69.0% to 81.5%. The 5-year overall relative survival rates of CM were 92.9%, 93.5%, and 95.6% for 2004-2008, 2009-2013, and 2014-2018, respectively. Females had a marginally higher survival rate than males for almost all subtypes, older people had lower survival rates than younger people, white patients had higher survival rates than nonwhite ones, and urban locations had higher rates of survival from CM than rural locations did. The survival rate of CM was significantly lower for distant metastasis. Conclusion: The survival rate of patients with CM gradually improved overall during 2004-2018. With the predicted survival rate of 96.7% for 2019-2023, this trend will still be present. Assessing the changes experienced by patients with CM over the previous 15 years can help in predicting the future course of CM. It also provides a scientific foundation that associated departments can use to develop efficient tumor prevention and control strategies.

A prognostic nomogram for the cancer-specific survival of white patients with invasive melanoma at BANS sites based on the Surveillance, Epidemiology, and End Results database.

Objective: The purpose of this study was to develop a comprehensive nomogram for the cancer-specific survival (CSS) of white patients with invasive melanoma at back, posterior arm, posterior neck, and posterior scalp (BANS) sites and to determine the validity of the nomogram by comparing it with the conventional American Joint Committee on Cancer (AJCC) staging system. Methods: This study analyzed the patients with invasive melanoma in the Surveillance, Epidemiology, and End Results (SEER) database. R software was used to randomly divide the patients into training and validation cohorts at a ratio of 7:3. Multivariable Cox regression was used to identify predictive variables. The new survival nomogram was compared with the AJCC prognosis model using the concordance index (C-index), area under the receiver operating characteristic (ROC) curve (AUC), net reclassification index (NRI), integrated discrimination index (IDI), calibration plotting, and decision-curve analysis (DCA). Results: A novel nomogram was established to determine the 3-, 5-, and 8-year CSS probabilities of patients with invasive melanoma. According to the nomogram, the Age at Diagnosis had the greatest influence on CSS in invasive melanoma, followed by Bone Metastasis, AJCC, Stage, Liver Metastasis, Histologic Subtype, Brain Metastasis, Ulceration, and Primary Site. The nomogram had a higher C-index than the AJCC staging system in both the training (0.850 versus 0.799) and validation (0.829 versus 0.783) cohorts. Calibration plotting demonstrated that the model had good calibration ability. The nomogram outperformed the AJCC staging system in terms of AUC, NRI, IDI, and DCA. Conclusion: This was the first study to develop and evaluate a comprehensive nomogram for the CSS of white patients with invasive melanoma at BANS sites using the SEER database. The novel nomogram can assist clinical staff in predicting the 3-, 5-, and 8-year CSS probabilities of patients with invasive melanoma more accurately than can the AJCC staging system.

Clinical, molecular- and cytogenetic analysis of a case of severe radio-sensitivity.

In radiotherapy the normal tissue reaction is often a limiting factor for radiation treatment. Still there is no screening method, which predicts normal tissue reaction on radiotherapy, especially in comparison to tumor tissue, and therefore allows tailoring of the radiation dose to each patient. Here, we present a case of severe radiation-related side effects. We applied classical cytogenetic techniques (Giemsa-banding and staining of centromeric regions), the comet assay as well as multicolor fluorescence in situ hybridization on peripheral blood lymphocytes of this patient in order to determine the radio-sensitivity on the DNA level and to correlate these findings with the clinical outcome. Our investigations revealed abnormalities on chromosome 9, deficiencies in the DNA-repair capacity after radiation exposure and a high number of radiation induced chromosomal aberrations. A detected high amount of residual damage two or three hours after radiation exposure and repair as well as the high number of chromosomal aberrations (ChAs) suggests a correlation between repair capacity and radiation induced ChAs. We concluded that the detected abnormalities might serve as a genetic basis for the radio-sensitive phenotype of this patient. Taken together this report strengthens the idea that intensive DNA genomic analysis of individual patients can serve as the basis for more favourable treatment of cancer patients.

Diffuse alveolar hemorrhage in systemic lupus erythematosus: risk factors and clinical outcome: results from affiliated hospitals of Catholic University of Korea.

This study was undertaken to investigate clinical characteristics of diffuse alveolar hemorrhage (DAH) in patients with systemic lupus erythematosus (SLE) and to determine risk factors and clinical outcomes of DAH in SLE patients. Among the 1521 patients with SLE admitted between January 1993 and June 2009 to affiliated hospitals of Catholic University of Korea, 21 SLE were admitted for DAH. The inclusion criteria for DAH was defined as new infiltrates on chest radiographs, an acute hemoglobin drop of at least 1.5 g/dl in the absence of an obvious source of bleeding, and one or more of the following signs: hemoptysis, hypoxemia, bronchoscopic or biopsy evidence of DAH. Included as disease controls were 83 SLE patients, matched for age and sex, who were admitted for other manifestations. Data based on medical records were analyzed retrospectively. There were no significantly differing demographic characteristics between SLE patients with DAH and those with other manifestations. Multivariate analysis demonstrated coexisting neuropsychiatric lupus (p = 0.002) and high SLE disease activity index scores (SLEDAI > 10) as independent risk factors in the development of DAH (p = 0.029). Among the 21 SLE patients with DAH, 13 died during the admission period (in-hospital mortality rate: 61.9%). Mortality was associated with infection and requirements of mechanical ventilation. Collectively, SLE patients who have neuropsychiatric manifestations or are in the active stage of the disease have an increased risk for developing DAH. Due to the high mortality of SLE patients with DAH, early recognition of risk factors and appropriate intervention is essential.

Risk factors and outcomes of Escherichia coli bacteremia caused by strains that produce CTX-M or non-CTX-M extended-spectrum-beta-lactamases.

To determine whether there are differences in risk factors and outcomes among patients with E. coli bacteremia caused by strains that produce CTX-M or non-CTX-M extended-spectrum beta-lactamases. From 1 July 2005 to 30 June 2007, patients with positive blood culture of extended-spectrum ß-lactamases (ESBL)-producing E. coli were reviewed. Sixty patients with ESBL-producing E. coli bacteremia were identified. These included 41 (68.3%) isolates with CTX-M ß-lactamases. CTX-M-14 accounted for 31 (75.6%) and CTX-M-3 for 9 (22.0%) of the 41 CTX-M isolates. Patients with CTX-M strains were less likely, by univariate analysis, to have significant risk factors for infection including age ≥ 65 years, chronic renal insufficiency, ICU stay at bacteremia onset, central venous catheter use and mechanical ventilation. Multivariate analysis revealed that chronic renal failure and ICU stay were independent predictors. Antibiograms were similar for CTX-M and non-CTX-M producers except that CTX-M strains were significantly more susceptible to cefmetazole (92.7 vs 36.8%, p < 0.0001). The overall mortality and length of hospitalization were not significantly different between the two groups. E. coli with CTX-M ß-lactamases was more likely than non-CTX-M strains to invade non-compromised patients. There were no differences in clinical outcomes between the two groups.

Theory of vibronic interactions in D2 and H2: a comparison between multichannel-quantum-defect and coupled-equation approaches.

New experimental energy levels for the 2pπC(1)Π(u)(-) state of D(2) are reported extending up to the dissociation limit and including rotational quantum numbers up to N = 10. These data are extracted from recent high resolution optical emission spectra, and they are used for a detailed comparison of two theoretical approaches, both of which are fully ab initio and are based on the same state-of-the-art clamped-nuclei potential energy curves. These are the coupled differential equations (CE) and the multichannel quantum defect theory (MQDT) approaches, each of which accounts for adiabatic corrections and non-adiabatic couplings. Both theoretical approaches reproduce the experimental levels to within a fraction of a wavenumber unit (cm(-1)) for the lower vibrational quantum numbers, with the MQDT surpassing the CE method. As the dissociation limit is approached, the residuals observed-calculated increase up to several cm(-1) and the MQDT method is up to a factor of two less accurate than the CE method. The same analysis is carried out with existing data for the H(2) isotopomer and yields similar results. An analogous comparison is also made for the 3pπD(1)Π(u)(-) and 4pπD('1)Π(u)(-) states for both isotopomers, where the MQDT is found to be superior to the CE approach.

Seroprevalence of SARS-CoV-2 antibodies, associated epidemiological factors and antibody kinetics among healthcare workers in Connecticut.

BACKGROUND: Healthcare workers (HCWs) are at the front line of the ongoing coronavirus 2019 (COVID-19) pandemic. Comprehensive evaluation of the seroprevalence of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) among HCWs in a large healthcare system could help to identify the impact of epidemiological factors and the presence of symptoms on the immune response to the infection over time. AIM: To determine the seroprevalence of SARS-CoV-2-specific antibodies among HCWs, identify associated epidemiological factors and study antibody kinetics. METHODS: A longitudinal evaluation of the seroprevalence and epidemiology of SARS-CoV-2-specific antibodies was undertaken in approximately 30,000 HCWs in the largest healthcare system in Connecticut, USA. FINDINGS: At baseline, the prevalence of SARS-CoV-2 antibody among 6863 HCWs was 6.3% [95% confidence interval (CI) 5.7-6.9%], and was highest among patient care support (16.7%), medical assistants (9.1%) and nurses (8.2%), and lower for physicians (3.8%) and advanced practice providers (4.5%). Seroprevalence was significantly higher among African Americans [odds ratio (OR) 3.26 compared with Caucasians, 95% CI 1.77-5.99], in participants with at least one symptom of COVID-19 (OR 3.00, 95% CI 1.92-4.68), and in those reporting prior quarantine (OR 3.83, 95% CI 2.57-5.70). No symptoms were reported in 24% of seropositive participants. Among the 47% of participants who returned for a follow-up serological test, the seroreversion rate was 39.5% and the seroconversion rate was 2.2%. The incidence of re-infection in the seropositive group was zero. CONCLUSION: Although there is a decline in the immunoglobulin G antibody signal over time, 60.5% of seropositive HCWs had maintained their seroconversion status after a median of 5.5 months.

Effect of oestrogen on T cell apoptosis in patients with systemic lupus erythematosus.

Defective control of T cell apoptosis is considered to be one of the pathogenetic mechanisms in systemic lupus erythematosus (SLE). Oestrogen has been known to predispose women to SLE and also to exacerbate activity of SLE; however, the role of oestrogen in the apoptosis of SLE T cells has not yet been documented. In this study, we investigated the direct effect of oestrogen on the activation-induced cell death of T cells in SLE patients. The results demonstrated that oestradiol decreased the apoptosis of SLE T cells stimulated with phorbol 12-myristate 13-acetate (PMA) plus ionomycin in a dose-dependent manner. In addition, oestradiol down-regulated the expression of Fas ligand (FasL) in activated SLE T cells at the both protein and mRNA levels. In contrast, testosterone increased FasL expression dose-dependently in SLE T cells stimulated with PMA plus ionomycin. The inhibitory effect of oestradiol on FasL expression was mediated through binding to its receptor, as co-treatment of tamoxifen, an oestrogen receptor inhibitor, completely nullified the oestradiol-induced decrease in FasL mRNA expression. Moreover, pre-treatment of FasL-transfected L5178Y cells with either oestradiol or anti-FasL antibody inhibited significantly the apoptosis of Fas-sensitive Hela cells when two types of cells were co-cultured. These data suggest that oestrogen inhibits activation-induced apoptosis of SLE T cells by down-regulating the expression of FasL. Oestrogen inhibition of T cell apoptosis may allow for the persistence of autoreactive T cells, thereby exhibiting the detrimental action of oestrogen on SLE activity.

Synchrotron vacuum ultraviolet radiation studies of the D (1)Π(u) state of H(2).

The 3pπD (1)Π(u) state of the H(2) molecule was reinvestigated with different techniques at two synchrotron installations. The Fourier transform spectrometer in the vacuum ultraviolet wavelength range of the DESIRS beamline at the SOLEIL synchrotron was used for recording absorption spectra of the D (1)Π(u) state at high resolution and high absolute accuracy, limited only by the Doppler contribution at 100 K. From these measurements, line positions were extracted, in particular, for the narrow resonances involving (1)Π(u) (-) states, with an accuracy estimated at 0.06 cm(-1). The new data also closely match multichannel quantum defect calculations performed for the Π(-) components observed via the narrow Q-lines. The Λ-doubling in the D (1)Π(u) state was determined up to v=17. The 10 m normal incidence scanning monochromator at the beamline U125/2 of the BESSY II synchrotron, combined with a home-built target chamber and equipped with a variety of detectors, was used to unravel information on ionization, dissociation, and intramolecular fluorescence decay for the D (1)Π(u) vibrational series. The combined results yield accurate information on the characteristic Beutler-Fano profiles associated with the strongly predissociated Π(u) (+) parity components of the D (1)Π(u) levels. Values for the parameters describing the predissociation width as well as the Fano-q line shape parameters for the J=1 and J=2 rotational states were determined for the sequence of vibrational quantum numbers up to v=17.

Incorrect diagnoses in patients with neutralizing anti-interferon-gamma-autoantibodies.

OBJECTIVES: Early diagnosis of adult-onset immunodeficiency associated with neutralizing anti-interferon-gamma autoantibodies (anti-IFNγ Abs) remains difficult given the lack of a distinctive phenotype and a routine test. This study aimed to investigate the determinants of incorrect tentative diagnoses and useful clues for early disease recognition. METHODS: This study enrolled adult patients who had unexplained opportunistic infections diagnosed at six hospitals and identified those having neutralizing anti-IFNγ Abs (cases). Demographics, medical history, initial presentations and laboratory data, causative pathogens, tentative diagnoses, and treatment were analysed and compared among individuals having neutralizing anti-IFNγ Abs (cases) and those without (controls). RESULTS: Among the 154 patients enrolled, neutralizing anti-IFN-γ Abs were detected in 50 (71%) of 70 patients with disseminated non-tuberculous mycobacterial infection (dNTM) but not in 84 patients without dNTM. The median time from disease onset to the recognition of dNTM associated with neutralizing anti-IFNγ Abs was 1.6 years (range, 0.25-19 years). Incorrect tentative diagnoses resulted in the administration of anti-tuberculosis regimens (60%, 30/50), immunosuppressants (48%, 24/50), and systemic chemotherapy (2%, 10/50) to the 50 cases. Multivariate analysis revealed that case patients were more likely than controls to present with multiple bone lesions (adjusted odds ratio (OR), 27.16; 95% confidence interval (CI), 1.21-609.59) and leukocytosis (adjusted OR, 1.48; 95% CI, 1.12-1.95); however, the controls had a higher rate of mycobacterial bloodstream infection (adjusted OR, 0.05; 95% CI 0.00-0.66). CONCLUSIONS: The high rate of incorrect tentative diagnoses led to frequent inappropriate management in patients with neutralizing anti-IFNγ Abs, and highlighted the need for increased awareness among clinicians.

False-positive Aspergillus galactomannan immunoassays associated with intravenous human immunoglobulin administration.

OBJECTIVES: Evidence of false-positive galactomannan enzyme immunoassay (GM-EIA) results associated with intravenous immunoglobulin (IVIG) administration is scarce. Here, we aimed to determine the false-positive rate of GM-EIA after IVIG administration and to identify the related factors. METHODS: Standard GM-EIA was performed using diluted and pure human IVIG samples with and without heat treatment. We also included adult patients who had at least one GM-EIA result within 1 week of IVIG administration for analysis. Those who had prior invasive aspergillosis within 1 year before IVIG therapy were excluded. The clinical characteristics and galactomannan index (GMI) kinetics between patients with false-positive and true-positive GMI were compared. RESULTS: All diluted and pure IVIG samples tested positive for GM. Heat treatment resulted in the considerable elevation of GMI. Of 48 patients with positive GM-EIA results within 1 week of IVIG administration, 22 (45.8%) were considered to have false-positive antigenaemia (false-positive group, FPG). After the completion of IVIG administration, a decline in GMI was observed in all FPG patients but in only 18 out of 26 patients (69.2%) with true-positive results (true-positive group, TPG). By 7, 14, and 18 days of IVIG administration, GMI reverted to negative values in 7/15 (46.7%), 18/20 (90%) and 22/22 (100%) FPG patients, respectively, and 6/24 (25%), 14/24 (58.3%), and 16/26 (61.5%) of TPG patients, respectively. The TPG was more likely to have two or more consecutively positive GMIs after IVIG administration than the FPG (adjusted odds ratio, 9.01; 95% confidence interval, 1.99-40.9). CONCLUSIONS: IVIG treatment may produce false-positive GM-EIA results. A positive GMI among patients receiving human IVIG should be interpreted with caution.

Up-regulated macrophage migration inhibitory factor protects apoptosis of dermal fibroblasts in patients with systemic sclerosis.

Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine that has been demonstrated to regulate the apoptosis of several cell types. Dysregulated apoptosis of fibroblasts has been implicated in a variety of fibrotic diseases, including systemic sclerosis (SSc). In this study, we investigated the role of MIF in the apoptosis of dermal fibroblasts. The concentrations of MIF were measured in sera and in culture supernatants of peripheral blood mononuclear cells (PBMCs) and dermal fibroblasts by enzyme-linked immunosorbent assay. The degree of apoptosis was determined by colorimetric assay, and signalling pathways were examined by Western blot. The results showed that serum levels of MIF were significantly higher in patients with SSc (n = 47) than in healthy controls (n = 56). Stimulation of PBMCs by anti-CD3 and anti-CD28 increased the production of MIF by fourfold over the constitutive levels. SSc dermal fibroblasts produced higher amounts of MIF than normal dermal fibroblasts. When treated with sodium nitroprusside (SNP), SSc dermal fibroblasts showed a lower degree of apoptosis compared with normal dermal fibroblasts. Exogenous MIF (1-100 ng/ml) inhibited SNP-induced apoptosis of dermal fibroblasts dose-dependently. Both extracellular regulated kinase (ERK) inhibitor (PD98059) and protein kinase B (Akt) inhibitor (LY294002) almost completely blocked the inhibitory effect of MIF on apoptosis. Furthermore, MIF increased the expression of Bcl-2, phospho-ERK and phospho-Akt activity in dermal fibroblasts. Taken together, our data suggest that MIF released by activated T cells and dermal fibroblasts decreases the apoptosis of dermal fibroblasts through activation of ERK, Akt and Bcl-2 signalling pathways, which might be associated with excessive fibrosis in SSc.

Mesenchymal stem cells overexpressing interleukin-10 attenuate collagen-induced arthritis in mice.

Mesenchymal stem cells (MSCs) have the inherent ability to migrate to multiple organs and to exert immunosuppressive activity. The aim of this study was to investigate the anti-arthritogenic effects of interleukin (IL)-10-transduced MSCs (IL-10-MSC) on the development of inflammatory arthritis. DBA/1 mice were immunized with type II collagen (CII) to induce inflammatory arthritis and then injected weekly three times with IL-10-MSCs 21 days after primary immunization. Control mice received vehicle or MSCs alone. Serum anti-CII antibody and T cell response to CII were determined. The results showed that cultured IL-10-MSCs were able to secrete high amounts of IL-10 in vitro. Injection of IL-10-MSCs decreased the severity of arthritis significantly. However, there was no difference in arthritis severity between mice treated with MSC and vehicle alone. Anti-CII antibody titres in the sera and T cell proliferative response to CII in lymph node cells were decreased significantly in mice treated with IL-10-MSCs compared with vehicle-treated mice. Serum IL-6 level was also decreased by the administration of IL-10-MSCs. In contrast, spleen cells of IL-10-MSC-treated mice produced higher amounts of IL-4 than those of control mice. Interestingly, although not as potent as IL-10-MSCs, injection of naive MSCs alone decreased serum levels of IL-6 and anti-CII antibody, while increasing IL-4 production from cultured splenic cells. Taken together, systemic administration of genetically modified MSCs overexpressing IL-10 inhibits experimental arthritis not only by suppressing autoimmune response to CII but also by regulating cytokine production, and thus would be a new strategy for treating rheumatoid arthritis.

Use of QuantiFERON-TB Gold In-tube assay in screening for neutralizing anti-interferon-γ autoantibodies in patients with disseminated nontuberculous mycobacterial infection.

OBJECTIVE: Anti-interferon- γ (IFN-γ) autoantibodies (anti-IFN-γ Abs) have been increasingly recognized as an important cause of disseminated nontuberculous mycobacterial (DNTM) infection, and identification of this immunodeficiency impacts clinical management. However, the protean disease manifestations and inaccessibility to diagnostic tests in clinical settings hamper its early diagnosis. Here, we sought to determine whether QuantiFERON-TB Gold In-tube (QFT-GIT), a commercialized IFN-γ release assay, could be used to screen for neutralizing anti-IFN-γ Abs among previously healthy adults with DNTM infection. METHODS: Non-HIV patients with DNTM infection were prospectively enrolled for the QFT-GIT assays. We measured their plasma concentration of anti-IFN-γ Abs and their neutralizing capacity through enzyme-linked immunosorbent assay and flow cytometry. We then analysed the correlation between QFT-GIT results and the presence of neutralizing anti-IFN-γ Abs among patients with and without previously recognized immunosuppression, respectively. RESULTS: Irrespective of the autoantibody concentration or disease activity, all patients with neutralizing anti-IFN-γ Abs (100%, 30/30) had indeterminate QFT-GIT results because of extremely low or undetectable IFN-γ levels in the mitogen tubes. None of the four DNTM patients who were previously healthy and tested negative of anti-IFN-γ Abs had an indeterminate QFT-GIT result, and their IFN-γ levels in the mitogen tube were significantly higher than those of the patients with anti-IFN-γ Abs (8.28 IU/mL vs. 0.05 IU/mL, p 0.001). CONCLUSION: An indeterminate QFT-GIT result because of undetectable or extremely low IFN-γ level in the mitogen tube suggests the presence of neutralizing anti-IFN-γ Abs in a previously healthy patient with DNTM infection.