Association between treatment with colchicine and improved survival in a single-centre cohort of adult hospitalised patients with COVID-19 pneumonia and acute respiratory distress syndrome.

OBJECTIVES: The outbreak of COVID-19 posed the issue of urgently identifying treatment strategies. Colchicine was considered for this purpose based on well-recognised anti-inflammatory effects and potential antiviral properties. In the present study, colchicine was proposed to patients with COVID-19, and its effects compared with 'standard-of-care' (SoC). METHODS: In the public hospital of Esine, northern Italy, 140 consecutive inpatients, with virologically and radiographically confirmed COVID-19 admitted in the period 5-19 March 2020, were treated with 'SoC' (hydroxychloroquine and/or intravenous dexamethasone; and/or lopinavir/ritonavir). They were compared with 122 consecutive inpatients, admitted between 19 March and 5 April 2020, treated with colchicine (1 mg/day) and SoC (antiviral drugs were stopped before colchicine, due to potential interaction). RESULTS: Patients treated with colchicine had a better survival rate as compared with SoC at 21 days of follow-up (84.2% (SE=3.3%) vs 63.6% (SE=4.1%), p=0.001). Cox proportional hazards regression survival analysis showed that a lower risk of death was independently associated with colchicine treatment (HR=0.151 (95% CI 0.062 to 0.368), p<0.0001), whereas older age, worse PaO2/FiO2, and higher serum levels of ferritin at entry were associated with a higher risk. CONCLUSION: This proof-of-concept study may support the rationale of use of colchicine for the treatment of COVID-19. Efficacy and safety must be determined in controlled clinical trials.

Relationship between different subpopulations of circulating CD4+ T lymphocytes and microvascular or systemic oxidative stress in humans.

BACKGROUND AND OBJECTIVE: Different components of the immune system, including innate and adaptive immunity (T effector lymphocytes and T regulatory lymphocytes - TREGs) may be involved in the development of hypertension, vascular injury and inflammation. However, no data are presently available in humans about possible relationships between T-lymphocyte subtypes and microvascular oxidative stress. Our objective was to investigate possible relationships between T-lymphocyte subtypes and systemic and microvascular oxidative stress in a population of normotensive subjects and hypertensive patients. PATIENTS AND METHODS: In the present study we enrolled 24 normotensive subjects and 12 hypertensive patients undergoing an elective surgical intervention. No sign of local or systemic inflammation was present. All patients underwent a biopsy of subcutaneous fat during surgery. A peripheral blood sample was obtained before surgery for assessment of T lymphocyte subpopulations by flow cytometry and circulating indices of oxidative stress. RESULTS: A significant direct correlation was observed between Th1 lymphocytes and reactive oxygen species (ROS) production (mainly in microvessels). Additionally, significant inverse correlations were observed between ROS and total TREGs, or TREGs subtypes. Significant correlations were detected between circulating indices of oxidative stress/inflammation and indices of microvascular morphology/Th1 and Th17 lymphocytes. In addition, a significant inverse correlation was detected between TREGs in subcutaneous small vessels and C reactive protein. CONCLUSIONS: Our data suggest that TREG lymphocytes may be protective against microvascular damage, probably because of their anti-oxidant properties, while Th1-Th17 lymphocytes seem to exert an opposite effect, confirming an involvement of adaptive immune system in microvascular damage.

Serum levels of granzyme B decrease in patients with rheumatoid arthritis responding to abatacept.

OBJECTIVES: A possible role of granzyme B (GZMB) in the pathogenesis of joint erosions in rheumatoid arthritis (RA) has been suggested. Since CD28neg T-cells may be an important source of GZMB, and we have previously shown that co-stimulation blockade by abatacept can prevent the generation of the CD28neg T-cell populations, we evaluated the effect of abatacept therapy on GZMB serum levels in patients with RA. METHODS: The serum levels of GZMB were evaluated by an indirect solid-phase enzyme immunoassay before the start of treatment with abatacept (T0) in 53 patients with RA and after 6 months of therapy (T6) in 25 patients. RESULTS: At T0, GZMB serum levels were correlated with disease activity measured by DAS28-CRP (p=0.0022) and percentages of circulating CD4+CD28neg and CD8+CD28neg T-cells (p=0.007; p=0.031). The levels of GZMB in 18 patients with a moderate or good EULAR clinical response to ABA significantly decreased from T0 to T6 (p=0.023), whereas no variation was observed in 7 non responders. The variation of GZMB levels was directly correlated with that of DAS28-PCR (p=0.040), but not with those of circulating CD28-neg T-cell subsets. CONCLUSIONS: Costimulation blockade by ABA can decrease the serum levels of GZMB in RA patients responding to the treatment, suggesting that this might be one of the mechanism by which abatacept can prevent radiographic erosions. However, the lack of correlation of such decrease with the numbers of circulating CD28-neg T cells suggests that these cells probably are not the main source of serum GZMB.

Reduced T-cell repertoire restrictions in abatacept-treated rheumatoid arthritis patients.

BACKGROUND: CD28(neg) T cells, which display functional characteristic of oligoclonally expanded cytotoxic memory T lymphocytes, are believed to be pathologically relevant in rheumatoid arthritis manifestation. The CD28 co-stimulation blockade by abatacept can prevent the generation of CD28(neg) T-cell populations in these patients. METHODS: Samples were obtained before and after 12 months of abatacept therapy. T-cell phenotype and T-cell receptor diversity were evaluated by flow cytometry and complementarity-determining region-3 spectratyping, respectively, while telomerase reverse-transcriptase gene level was measured by real-time PCR. RESULTS: Abatacept induces a decrease of the percentage and number of CD4(+)CD28(neg) T cells and a reduction of T-cell repertoire restrictions; these features are directly correlated. Thymic output and telomerase activity are not modified by the therapy. CONCLUSIONS: Abatacept-induced decrease of peripheral T-cell repertoire restrictions can due to a reduced generation of senescent, chronically stimulated CD4(+)CD28(neg) T cells.

Anti-Ro/SSA-p200 antibodies in the prediction of congenital heart block. An Italian multicentre cross-sectional study on behalf of the 'Forum Interdisciplinare per la Ricerca nelle Malattie Autoimmuni (FIRMA) Group'.

OBJECTIVES: To verify the association between the presence of specific anti-52 Ro/SSA-p200 antibodies and congenital heart block (CHB). METHODS: 207 pregnant Italian women carrying anti-Ro/SSA Ab were retrospectively evaluated. Anti-p200 Ab were investigated in the mothers' sera by ELISA (Euro-Diagnostica,Wieslab SS-A p200). RESULTS: CHB occurred in 42 children (34 complete CHB), whereas 165 were not affected. All CHB cases were previously identified with an ELISA screening for anti-Ro/SSA 60 kD Ab. Anti-p200 Ab were more frequently positive (81.0% vs. 59.1%, p=0.013) and at a higher titer in CHB mothers (Absorbance ratio: 2.030 (0.208-4.052) vs. 0.925 (0.200-3.816); p=0.017). This association was maintained even when the 42 mothers of children with CHB were compared with a control group matched for age and diagnosis (80.9% vs. 50.0%; p=0.006). The presence of anti-p200 Ab provided an odds ratio (OR) for CHB of 2.98 (CI: 1.30-6.83), which was higher than that of other variables, such as maternal disease and other antibody specificities. CHB risk significantly decreased in the absence of this fine specificity (OR:0.34, CI: 0.15-0.77). However, while the negative predictive value related to anti-Ro/SSA 60 kD Ab ELISA was 100%, almost 20% of mothers negative for anti-p200 Ab delivered babies with CHB. CONCLUSIONS: Anti-p200 antibodies seem to be associated with CHB with a higher probability than anti-Ro/SSA Ab, and therefore may be an additional test to identify mothers at higher risk to deliver affected children. An ELISA screening for anti-Ro/SSA 60 kD Ab is nevertheless mandatory given the probability of developing CHB also in the absence of anti-p200 Ab.

Treatment with COLchicine in hospitalized patients affected by COVID-19: The COLVID-19 trial.

OBJECTIVE: To evaluate whether the addition of colchicine to standard of care (SOC) results in better outcomes in hospitalized patients with COVID-19. DESIGN: This interventional, multicenter, randomized, phase 2 study, evaluated colchicine 1.5 mg/day added to SOC in hospitalized COVID-19 patients (COLVID-19 trial) and 227 patients were recruited. The primary outcome was the rate of critical disease in 30 days defined as need of mechanical ventilation, intensive care unit (ICU), or death. RESULTS: 152 non-anti-SARS-CoV-2-vaccinated patients (colchicine vs controls: 77vs75, mean age 69.1±13.1 vs 67.9±15 years, 39% vs 33.3% females, respectively) were analyzed. There was no difference in co-primary end-points between patients treated with colchicine compared to controls (mechanical ventilation 5.2% vs 4%, ICU 1.3% vs 5.3%, death 9.1% vs 6.7%, overall 11 (14.3%) vs 10 (13.3%) patients, P=ns, respectively). Mean time to discharge was similar (colchicine vs controls 14.1±10.4 vs 14.7±8.1 days). Older age (>60 years, P=0.025), P/F<275 mmHg (P=0.005), AST>40 U/L (P<0.001), pre-existent heart (P=0.02), lung (P=0.003), upper-gastrointestinal (P=0.014), lower-gastrointestinal diseases (P=0.009) and cancer (P=0.008) were predictive of achieving the primary outcome. Diarrhoea (9.1% vs 0%, p=0.0031) and increased levels of AST at 6 days (76.9±91.8 vs 33.5±20.7 U/l, P=0.016) were more frequent in the colchicine group. CONCLUSION: Colchicine did not reduce the rate and the time to the critical stage. Colchicine was relatively safe although adverse hepatic effects require caution. We confirm that older (>60 years) patients with comorbidities are characterized by worse outcome.

Thymic and bone marrow output in patients with common variable immunodeficiency.

OBJECTIVE: The study aims to obtain more information about the immune deficit of common variable immunodeficiency (CVID) patients. MATERIALS AND METHODS: A new real-time PCR assay was used to quantify T and B lymphocyte mobilization from the production and maturation sites through the detection of T cell receptor excision circles (TRECs) and kappa-deleting recombination circles (KRECs) and to allow the estimation of the average number of B cell divisions. T and B lymphocyte subsets were analyzed by flow cytometry. RESULTS: The number of TREC(+) lymphocytes, which depends on age and gender, was significantly reduced in CVID patients. Similarly, KREC concentration was lower than in controls. Classification of patients according to the percentage of memory switched B cells showed that patients belonging to MB2 group and therefore with conserved B cell maturation have the lowest new B cell output but increased average peripheral divisions, leading to the highest B cell number. CONCLUSIONS: TREC and KREC quantification can be helpful for a more complete and informative understanding of a heterogeneous disease such as CVID.

Detection of anti-IFI16 antibodies by ELISA: clinical and serological associations in systemic sclerosis.

OBJECTIVES: To validate the clinical significance of anti-IFI16 autoantibodies in SSc and assess their associations with serological markers of SSc. METHODS: A semi-quantitative ELISA was used to detect anti-IFI16 autoantibodies in the sera of 344 SSc patients from seven Italian hospitals and 144 healthy controls. SSc-associated autoantibodies [anti-RNA polymerase III (anti-RNAP III) antibodies, anti-centromere, anti-topo I] and IF patterns were evaluated using commercial assays. Statistical analyses were performed to test clinical and serological associations. RESULTS: The results of this study confirm a significant prevalence (29%) of anti-IFI16 antibodies in the SSc population (n = 344). Anti-IFI16 antibodies were also detected in 30% of the SSc patients who tested negative for both ACAs and anti-topo I (anti-Scl70) antibodies. In this subgroup of patients, anti-IFI16 antibodies were significantly associated with the limited cutaneous form of SSc with a sensitivity of 40% and a specificity of 81%. Moreover, analysis of the distribution of anti-RNAP III antibodies vs anti-IFI16 in the same SSc population showed that they were mutually exclusive. IIF revealed no association between anti-IFI16 and fluoroscopic patterns, due to a lack of IFI16 autoantigen in HEp-2 cells. Anti-IFI16 antibody levels were also significantly associated with heart involvement. CONCLUSIONS: Anti-IFI16 autoantibodies are frequently detected in SSc, displaying clinical and laboratory associations, and being particularly useful for diagnosis and disease classification in patients who are negative for other SSc serological markers.

The number of circulating recent thymic emigrants is severely reduced 1 year after a single dose of alemtuzumab in renal transplant recipients.

To better understand the kinetics of the delayed reconstitution of peripheral CD4+ T-cells after depletion with a single administration of alemtuzumab (AL) for renal transplantation, we evaluated in these patients the percentage and absolute number of recent thymic emigrants (RTEs) CD4+ T cells, together with naive and memory subsets, defined by the analysis of CD31, CD45RA and CCR7 expression, and compared with patients treated with a nondepleting protocol based on basiliximab, and with healthy controls. In AL-treated patients, the number of circulating CD4+ T cells was greatly reduced 1 year after the infusion (P < 0.01), but the proportions of central memory, effector memory and terminally differentiated effector memory subsets among CD4+ cells were significantly increased. On the contrary, the proportion and the absolute number of naïve CD4+ T cells, although progressively increasing with time, were severely reduced. In particular, the absolute number of RTEs had only very slight increase with time (P = 0.049) and was dramatically low 1 year after the therapy (P < 0.01 vs. healthy controls; P < 0.05 vs. basiliximab-treated transplant recipients). These data suggest that a prolonged defective thymic output after AL therapy in renal transplant recipients is one of the main causes of the persistent CD4+ T-cell lymphopenia observed in these patients.

Anti-inflammatory therapies for pericardial diseases in the COVID-19 pandemic: safety and potentiality.

: The COVID-19 pandemic is challenging our cardiovascular care of patients with heart diseases. In the setting of pericardial diseases, there are two possible different scenarios to consider: the patient being treated for pericarditis who subsequently becomes infected with SARS-CoV-2, and the patient with COVID-19 who develops pericarditis or pericardial effusion. In both conditions, clinicians may be doubtful regarding the safety of nonsteroidal anti-inflammatory drugs (NSAIDs), corticosteroids, colchicine, and biological agents, such as anti-IL1 agents (e.g. anakinra), that are the mainstay of therapy for pericarditis.For NSAIDs, there is no clear scientific evidence linking ibuprofen and other NSAIDs to worsening of COVID-19; however, it seems prudent to continue them, if necessary to control pericarditis, and on the other hand, to prefer paracetamol for fever and systemic symptoms related to COVID-19. Treatments with corticosteroids, colchicine, and anakinra appear well tolerated in the context of COVID-19 infection and are currently actively evaluated as potential therapeutic options for COVID infection at different stages of the disease. On this basis, currently most treatments for pericarditis do not appear contraindicated also in the presence of possible COVID-19 infection and should not be discontinued, and some (corticosteroids, colchicine, and anakinra) can be considered to treat both conditions.

Incidence and prevalence of systemic sclerosis in Valcamonica, Italy, during an 18-year period.

Objectives: To investigate the epidemiology of systemic sclerosis in Valcamonica, an Italian Alpine valley, during an 18-year-long period. Methods: Patients with systemic sclerosis living in Valcamonica between 1999 and 2016 were identified by capture/recapture method using: (1) clinical databases of the only secondary Rheumatology Unit present in the valley and of the tertiary referral center for this area; (2) administrative data, extracting records with the International Classification of Diseases, 10th Revision, code for systemic sclerosis. Patients were included in the analysis when either the 1980 American Rheumatism Association classification criteria for systemic sclerosis or the 2013 American College of Rheumatology/European League Against Rheumatism criteria were satisfied. To study temporal changes, mean yearly incidence during three different 6-year interval was calculated. Prevalence rates were estimated at four different time points. Results: General population with age over 14 years living in Valcamonica varied during the evaluated period between 85,168 and 91,245 inhabitants. A total of 65 patients with systemic sclerosis were identified (female 84.6%, limited cutaneous systemic sclerosis: 84.6%; anticentromere: 64.6%). Systemic sclerosis incidence and prevalence increased during the study period (p = 0.029 and p < 0.0001, respectively). The increase of incidence was accounted for by cases satisfying only the 2013 criteria, with limited cutaneous systemic sclerosis, and with anticentromere, whereas the incidence of systemic sclerosis cases classified according to the 1980 criteria did not significantly increase. The prevalence at 31 December 2016 was 58.6 (95% confidence interval, 44.8-76.6) per 100,000 persons aged >14 years. Survival at 10 years after systemic sclerosis diagnosis was 83.0% (standard error, 5.6). Conclusion: Systemic sclerosis incidence and prevalence increased over time in this area, due to the increased recruitment of patients with milder forms of the disease.

Microvascular Density and Circulating Endothelial Progenitor Cells Before and After Treatment with Incretin Mimetics in Diabetic Patients.

INTRODUCTION: Glucagon-like peptide 1-receptor agonists (incretin mimetics) and dipeptidyl peptidase-4 inhibitors (incretin enhancers) have been recently introduced in the treatment of diabetes mellitus. In particular, incretin mimetics seems to have ancillary antioxidant/antinflammatory properties that might be involved in endothelial protection. AIM: To investigate the effect of incretin mimetic therapy (liraglutide, exenatide) given to 11 patients with type 2 diabetes mellitus, on circulating endothelial progenitor cells (EPCs) (bone marrow-derived cells possibly participating in neovascularization and endothelial protection and repair) and capillary density. METHODS: Four diabetic patients were treated with exenatide (5 µg twice daily for 4 weeks and then 10 µg twice daily for 3 weeks) and 7 with liraglutide (0.6 mg per day for 1 week and then 1.2 mg per day for 3 weeks). Peripheral venous blood samples were obtained before treatment (basal) and after 4 week in patients treated with liraglutide, and after 4 and 7 weeks in patients treated with exenatide, since drug titration is usually longer. EPCs were evaluated by flow cytometry as CD34+/KDR+ cells. Capillary density was evaluated by videomicroscopy, before and after venous congestion, in the dorsum of the 4th finger. RESULTS: Patients treated with liraglutide (6 males 1 female, age 54 ± 12 years) showed a decrease in body mass index and blood pressure during treatment, while patients treated with exenatide (3 males 1 female, age 57 ± 6 years) did not show any relevant change. EPCs were significantly increased after treatment with exenatide, but not after treatment with liraglutide. Capillary density was slightly increased only after 4 weeks of treatment with exenatide, however the increase was no longer present at the final evaluation. CONCLUSIONS: Treatment with exenatide, but not with liraglutide, was able to increase the number of circulating EPCs, possibly through an antioxidative/antiinflammatory effect.

Relationship Between Different Subpopulations of Circulating CD4+ T-lymphocytes and Microvascular Structural Alterations in Humans.

BACKGROUND: Different components of the immune system, including innate and adaptive immunity (T-effector lymphocytes and T-regulatory lymphocytes-TREGs) may be involved in the development of hypertension. In addition, it was demonstrated in animal models that TREGs may prevent angiotensin II-induced hypertension and vascular injury/inflammation. However, no data are presently available in humans about possible relationships between T-lymphocyte subtypes and microvascular structural alterations. METHODS: For this purpose, in the present study, we enrolled 24 normotensive subjects and 12 hypertensive patients undergoing an elective surgical intervention. No sign of local or systemic inflammation was present. All patients underwent a biopsy of subcutaneous fat during surgery. Subcutaneous small resistance arteries were dissected and mounted on a wire myograph and the media to lumen ratio (M/L) was calculated. In addition, retinal arteriolar structure was evaluated noninvasively by scanning laser Doppler flowmetry. Capillary density in the nailfold, dorsum of the finger, and forearm were evaluated by videomicroscopy. A peripheral blood sample was obtained before surgery for assessment of T-lymphocyte subpopulations by flow cytometry. RESULTS: Significant negative correlations were observed between indices of microvascular structure (M/L of subcutaneous small arteries and wall to lumen ratio of retinal arterioles) and circulating TREG lymphocytes. A direct correlation was observed between M/L of subcutaneous small arteries and circulating Th17 lymphocytes. In addition, total capillary density was correlated with a TREG effector memory subpopulation. CONCLUSION: Our data suggest that some lymphocyte subpopulations may be related to microvascular remodeling, confirming previous animal data, and opening therapeutic possibilities.

Predictive factors of abatacept therapy discontinuation in patients with rheumatoid arthritis.

The aim of this paper was to look for predictors of abatacept (ABA) therapy discontinuation in patients with rheumatoid arthritis (RA). Seventy-one RA patients treated with ABA were followed up. Demographical, clinical, and laboratory parameters of the patients, including peripheral blood T and B cell populations, different rheumatoid factor and anti-cyclic citrullinated peptide autoantibodies isotypes, and serum free light chains were evaluated. Comparing patients who discontinued ABA with those still in therapy we observed: a higher proportion of smokers (51.9 vs 25.6 %; p = 0.03); a non significant lower proportion of anti-cyclic citrullinated peptide positivity (76 vs 89.5 %; p = 0.13); a lower proportion of terminally differentiated effector memory cells (TDEM) among total CD8+ T lymphocytes at baseline (22.0 % (7.8-39.2) vs 38.7 % (20.7-55.9); p = 0.002). Logistic multivariate analysis showed that only the proportion of CD8+TDEM T cells was an independent predictive factor of therapy discontinuation (OR (95 % IC) = 6.2 (1.2 to 30.8); p = 0.026). Receiver-operating characteristic analysis showed a significant performance of this biomarker for prediction of therapy discontinuation (using a cut-off of 30.6 %: AUC: 0.760 ± 0.07; p = 0.002). Patients with a low proportion of CD8+TDEM at baseline had a higher probability of discontinuing the treatment during time (log-rank test: p < 0.01). T cell characterization for identification of TDEM CD8+ T cells might be a useful test to predict discontinuation of ABA therapy.