Cell Activation In Patients Research Articles

Transmembrane stem factor nanodiscs enhanced revascularization in a hind limb ischemia model in diabetic, hyperlipidemic rabbits

Therapies to revascularize ischemic tissue have long been a goal for the treatment of vascular disease and other disorders. Therapies using stem cell factor (SCF), also known as a c-Kit ligand, had great promise for treating ischemia for myocardial infarct and stroke, however clinical development for SCF was stopped due to toxic side effects including mast cell activation in patients. We recently developed a novel therapy using a transmembrane form of SCF (tmSCF) delivered in lipid nanodiscs. In previous studies, we demonstrated tmSCF nanodiscs were able to induce revascularization of ischemia limbs in mice and did not activate mast cells. To advance this therapeutic towards clinical application, we tested this therapy in an advanced model of hindlimb ischemia in rabbits with hyperlipidemia and diabetes. This model has therapeutic resistance to angiogenic therapies and maintains long term deficits in recovery from ischemic injury. We treated rabbits with local treatment with tmSCF nanodiscs or control solution delivered locally from an alginate gel delivered into the ischemic limb of the rabbits. After eight weeks, we found significantly higher vascularity in the tmSCF nanodisc-treated group in comparison to alginate treated control as quantified through angiography. Histological analysis also showed a significantly higher number of small and large blood vessels in the ischemic muscles of the tmSCF nanodisc treated group. Importantly, we did not observe inflammation or mast cell activation in the rabbits. Overall, this study supports the therapeutic potential of tmSCF nanodiscs for treating peripheral ischemia.

Damage-associated cellular markers in the clinical and pathogenic profile of vaccine-induced immune thrombotic thrombocytopenia

BackgroundAdenoviral vector-based COVID-19 vaccine-induced immune thrombotic thrombocytopenia (VITT) is rare but carries significant risks of mortality and long-term morbidity. The underlying pathophysiology of severe disease is still not fully understood. The objectives were to explore the pathophysiological profile and examine for clinically informative biomarkers in patients with severe VITT. MethodsTwenty-two hospitalized patients with VITT, 9 pre- and 21 post-ChAdOx1 vaccine controls, were recruited across England, United Kingdom. Admission blood samples were analyzed for cytokine profiles, cell death markers (lactate dehydrogenase and circulating histones), neutrophil extracellular traps, and coagulation parameters. Tissue specimens from deceased patients were analyzed. ResultsThere were strong immune responses characterized by significant elevations in proinflammatory cytokines and T helper 1 and 2 cell activation in patients with VITT. Markers of systemic endothelial activation and coagulation activation in both circulation and organ sections were also significantly elevated. About 70% (n = 15/22) of patients met the International Society for Thrombosis and Haemostasis criteria for disseminated intravascular coagulation despite negligible changes in the prothrombin time. The increased neutrophil extracellular trap formation, in conjunction with marked lymphopenia, elevated lactate dehydrogenase, and circulating histone levels, indicates systemic immune cell injury or death. Both lymphopenia and circulating histone levels independently predicted 28-day mortality in patients with VITT. ConclusionThe coupling of systemic cell damage and death with strong immune-inflammatory and coagulant responses are pathophysiologically dominant and clinically relevant in severe VITT.

Effect of miR-125b on T Cell Activation in Aplastic Anemia by Targetting B7-H4

To investigate the effect of miR-125b on T cell activation in patients with aplastic anemia (AA) and its molecular mechanism. A total of 30 AA patients were enrolled in department of hematology, Binzhou Medical University Hospital from January 2018 to October 2021, as well as 15 healthy individuals as healthy control (HC) group. Peripheral blood mononuclear cells (PBMCs) were isolated, in which the levels of miR-125b and B7-H4 mRNA were detected by RT-qPCR. Immunomagnetic beads were used to separate naive T cells and non-naive T cells from AA patients and healthy people to detect the levels of miR-125b and B7-H4 mRNA. Lentivirus LV-NC inhibitor and LV-miR-125b inhibitor were transfected into cells, and T cell activation was detected by flow cytometry. The dual-luciferase reporter gene assay was used to detect the targetting relationship between miR-125b and B7-H4. RT-qPCR and Western blot were used to detect the levels of miR-125b, CD40L, ICOS, IL-10 mRNA and B7-H4 protein. Compared with HC group, the expression of miR-125b was up-regulated but B7-H4 mRNA was down-regulated in PBMCs of AA patients (P <0.05), and the proportions of CD4+CD69+ T cells and CD8+CD69+ T cells in PBMCs of AA patients were higher (P <0.05). The expression of miR-125b was significantly up-regulated but B7-H4 mRNA was down-regulated in both naive T cells and non-naive T cells of AA patients (P <0.05), and non-naive T cells was more significant than naive T cells (P <0.05). Compared with NC inhibitor group, the expression of miR-125b was significantly decreased, the expression level of CD69 on CD4+ and CD8+ T cells in PBMCs was also significantly decreased, while the luciferase activity was significantly increased after co-transfection of miR-125b inhibitor and B7-H4-3'UTR-WT in the miR-125b inhibitor group (P <0.05). Compared with NC inhibitor group, the mRNA and protein levels of B7-H4 were significantly increased in the miR-125b inhibitor group (P <0.05). Compared with miR-125b inhibitor+shRNA group, the expression levels of CD69 on CD4+ and CD8+ T cells were significantly increased, and the levels of CD40L, ICOS and IL-10 mRNA were also significantly increased in the miR-125b inhibitor+sh-B7-H4 group (P <0.05). MiR-125b may promote T cell activation by targetting B7-H4 in AA patients.

Experimentally Induced Peripheral Venous Congestion Exacerbates Inflammation, Oxidative Stress, and Neurohormonal and Endothelial Cell Activation in Patients With Systolic Heart Failure

Venous congestion (VC) is a hallmark of symptomatic heart failure (HF) requiring hospitalization; however, its role in the pathogenesis of HF progression remains unclear. We investigated whether peripheral VC exacerbates inflammation, oxidative stress and neurohormonal and endothelial cell (EC) activation in patients with HF with reduced ejection fraction (HFrEF). Two matched groups of patients with HFrEF and with no peripheral VC vs without recent HF hospitalization were studied. We modeled peripheral VC by inflating a cuff around the dominant arm, targeting ∼ 30 mmHg increase in venous pressure (venous stress test [VST]). Blood and ECs were sampled before and after 90 minutes of VST. We studied 44 patients (age 53 ± 12 years, 32% female). Circulating endothelin-1, tumor necrosis factor-α, interleukin-6, isoprostane, angiotensin II (ang-2), angiopoietin-2, vascular cell adhesion molecule-1, and CD146 significantly increased after the VST. Enhanced endothelin-1 and angiopoietin-2 responses to the VST were present in patients with vs without recent hospitalization and were prospectively associated with incident HF-related events; 6698 messenger ribonucleic acid (mRNA probe sets were differentially expressed in ECs after VST. Experimental VC exacerbates inflammation, oxidative stress, neurohormonal and EC activation and promotes unfavorable transcriptome remodeling in ECs of patients with HFrEF. A distinct biological sensitivity to VC appears to be associated with high risk for HF progression.

Retracted: Analysis of the Relationship between Scleritis and T Cell Activation in Patients with Hepatocellular Carcinoma Treated with PD-1 Carrelizumab.

[This retracts the article DOI: 10.1155/2022/4853481.].

No evidence of oligoclonal bands, intrathecal immunoglobulin synthesis and B cell recruitment in acute ischemic stroke.

Within the past 10 years, immune mechanisms associated with acute ischemic stroke (AIS) have been brought into focus, but data on B cell activation and intrathecal Ig production is still scarce. In this study, we determined the prevalence of an elevated IgG index, positive oligoclonal bands (OCBs) and chemokine C-X-C motif ligand 13 (CXCL13) levels in the cerebrospinal fluid (CSF) as markers of intrathecal IgG synthesis and B cell activation in patients with AIS. In a retrospective study we analyzed the cerebrospinal fluid (CSF) from 212 patients with AIS from December 2013 to May 2018 assessing intrathecal Ig synthesis, OCBs and CXCL13 concentrations. Overall, 5.7% (12/212) of AIS patients showed an intrathecal IgG synthesis, 0.5% (1/212) with isolated elevated IgG index, 5.2% (7/136) isolated positive OCBs and 2.9% (4/136) both elevated IgG index and positive OCBs. CXCL13 levels were elevated in 3.6% (3/83) of the patients. Approximately one third of these patients had simultaneously chronic inflammatory CNS disease (multiple sclerosis, neuromyelitis optica spectrum disorder, neurosarcoidosis). There was no significant association between CSF findings and stroke characteristics including vascular territory, localization, volume, etiology, acute treatment, or blood-brain barrier dysfunction. Intrathecal IgG synthesis was more common in patients with prior stroke. Longitudinal CSF analysis did not reveal any newly-occurring, but instead mostly persistent or even disappearing intrathecal IgG synthesis after AIS. We found no evidence of a relevant B cell recruitment and intrathecal IgG synthesis in patients with AIS. In fact, the occurrence of intrathecal IgG synthesis was associated with concurrent chronic inflammatory CNS disease or previous stroke. Consequently, in patients with first-ever AIS and intrathecal IgG synthesis, physicians should search for concomitant inflammatory CNS disease.

Corneal nerve loss and increased Langerhans cells are associated with disease severity in patients with rheumatoid arthritis.

Rheumatoid arthritis (RA) is a multisystem autoimmune disorder characterized by articular and extra-articular manifestations. Neuropathy is a poorly studied manifestation of RA. The aim of this study was to utilize the rapid non-invasive ophthalmic imaging technique of corneal confocal microscopy to identify whether there is evidence of small nerve fibre injury and immune cell activation in patients with RA. Fifty consecutive patients with RA and 35 healthy control participants were enrolled in this single-centre, cross-sectional study conducted at a university hospital. Disease activity was assessed with the 28-Joint Disease Activity Score and erythrocyte sedimentation rate (DAS28-ESR). Central corneal sensitivity was measured with a Cochet-Bonnet contact corneal esthesiometer. A laser scanning in vivo corneal confocal microscope was used to quantify corneal nerve fibre density (CNFD), nerve branch density (CNBD), nerve fibre length (CNFL), and Langerhans cell (LC) density. Corneal sensitivity (P = 0.01), CNFD (P = 0.02), CNBD (P < 0.001), and CNFL (P < 0.001) were lower, and mature (P = 0.001) and immature LC densities (P = 0.011) were higher in patients with RA compared to control subjects. CNFD (P = 0.016) and CNFL (P = 0.028) were significantly lower in patients with moderate to high (DAS28-ESR > 3.2) compared to mild (DAS28-ESR ≤ 3.2) disease activity. Furthermore, the DAS28-ESR score correlated with CNFD (r = -0.425; P = 0.002), CNBD (ρ = -0.362; P = 0.010), CNFL (r = -0.464; P = 0.001), total LC density (ρ = 0.362; P = 0.010) and immature LC density (ρ = 0.343; P = 0.015). This study demonstrates reduced corneal sensitivity, corneal nerve fibre loss and increased LCs which were associated with the severity of disease activity in patients with RA.

PGE2 expression by HPV6/11-induced respiratory papillomas blocks NK cell activation in patients with recurrent respiratory papillomatosis.

Recurrent respiratory papillomatosis (RRP), a rare chronic disease caused primarily by human papillomavirus types 6 and 11, consists of repeated growth of premalignant papillomas in the airway. RRP is characterized by multiple abnormalities in innate and adaptive immunity. Natural killer (NK) cells play important roles in immune surveillance and arepart of the innate immune responses that help prevent tumor growth. We identified that papillomas lack classical class I MHC and retain nonclassical class I MHC expression. Moreover, in this study, we have identified and characterized the mechanism that blocks NK cell targeting of papilloma cells. Here, we show for the first time that the PGE2 secreted by papilloma cells directly inhibits NK cells activation/degranulation principally through the PGE2 receptor EP2, and to a lesser extent through EP4 signaling. Thus, papilloma cells have a potent mechanism to block NK cell function that likely supports papilloma cell growth.

Coordinated Loss and Acquisition of NK Cell Surface Markers Accompanied by Generalized Cytokine Dysregulation in COVID-19.

Coronavirus disease 2019 (COVID-19), caused by the SARS-CoV-2 virus, is accompanied by a dysregulated immune response. In particular, NK cells, involved in the antiviral response, are affected by the infection. This study aimed to investigate circulating NK cells with a focus on their activation, depletion, changes in the surface expression of key receptors, and functional activity during COVID-19, among intensive care unit (ICU) patients, moderately ill patients, and convalescents (CCP). Our data confirmed that NK cell activation in patients with COVID-19 is accompanied by changes in circulating cytokines. The progression of COVID-19 was associated with a coordinated decrease in the proportion of NKG2D+ and CD16+ NK cells, and an increase in PD-1, which indicated their exhaustion. A higher content of NKG2D+ NK cells distinguished surviving patients from non-survivors in the ICU group. NK cell exhaustion in ICU patients was additionally confirmed by a strong negative correlation of PD-1 and natural cytotoxicity levels. In moderately ill patients and convalescents, correlations were found between the levels of CD57, NKG2C, and NKp30, which may indicate the formation of adaptive NK cells. A reduced NKp30 level was observed in patients with a lethal outcome. Altogether, the phenotypic changes in circulating NK cells of COVID-19 patients suggest that the intense activation of NK cells during SARS-CoV-2 infection, most likely induced by cytokines, is accompanied by NK cell exhaustion, the extent of which may be critical for the disease outcome.

Complement and endothelial cell activation in COVID-19 patients compared to controls with suspected SARS-CoV-2 infection: A prospective cohort study.

BackgroundThromboinflammation may influence disease outcome in COVID-19. We aimed to evaluate complement and endothelial cell activation in patients with confirmed COVID-19 compared to controls with clinically suspected but excluded SARS-CoV-2 infection.MethodsIn a prospective, observational, single-center study, patients presenting with clinically suspected COVID-19 were recruited in the emergency department. Blood samples on presentation were obtained for analysis of C5a, sC5b-9, E-selectin, Galectin-3, ICAM-1 and VCAM-1.Results153 cases and 166 controls (suffering mainly from non-SARS-CoV-2 respiratory viral infections, non-infectious inflammatory conditions and bacterial pneumonia) were included. Hospital admission occurred in 62% and 45% of cases and controls, respectively. C5a and VCAM-1 concentrations were significantly elevated and E-selectin concentrations decreased in COVID-19 out- and inpatients compared to the respective controls. However, relative differences in outpatients vs. inpatients in most biomarkers were comparable between cases and controls. Elevated concentrations of C5a, Galectin-3, ICAM-1 and VCAM-1 on presentation were associated with the composite outcome of ICU- admission or 30-day mortality in COVID-19 and controls, yet more pronounced in COVID-19. C5a and sC5b-9 concentrations were significantly higher in COVID-19 males vs. females, which was not observed in the control group.ConclusionsOur data indicate an activation of the complement cascade and endothelium in COVID-19 beyond a nonspecific inflammatory trigger as observed in controls (i.e., “over”-activation).

Aqueous Humor Biomarkers of Retinal Glial Cell Activation in Patients With or Without Age-Related Cataracts and With Different Stages of Diabetic Retinopathy.

PurposeTo clarify the expression of biomarkers of retinal glial cell activation in the aqueous humor (AH) of patients with and without age-related cataracts (ARCs) at different stages of diabetic retinopathy (DR).MethodsPatients were stratified by the presence of ARCs and then grouped by the presence of diabetes mellitus (DM), nonproliferative DR (NPDR), proliferative DR (PDR), and controls. Water channel aquaporin 1 (AQP1), water channel aquaporin 4 (AQP4), inwardly rectifying potassium channel 4.1 (Kir4.1), and glial fibrillary acidic protein (GFAP) were assayed in AH samples by ELISAs.ResultsWe enrolled 82 patients. The AQP1 concentration was higher in AH from cataract control patients than in control patients without cataracts (P < 0.05). The APQ1 concentration was also higher in patients with DM, NPDR, and PDR than in controls (P < 0.05). The concentrations of AQP4 and GFAP were significantly increased in patients with NPDR and PDR (P < 0.05) but not in patients with DM. Kir4.1 concentration was significantly decreased in patients with NPDR and PDR (P < 0.05), but the decrease in patients with DM did not reach significance. There were no differences in AQP4, Kir4.1, and GFAP between patients with and without ARCs.ConclusionsIncreased AQP1 in AH may be a biomarker for ARCs in patients without diabetes and a biomarker for retinal glial cell activation in patients with diabetes without cataracts. AQP4, Kir4.1, and GFAP levels in AH suggested that retinal glial cell activation was affected by the progression of DR.

T peripheral helper cells in autoimmune diseases.

Pathologic T cell-B cell interactions underlie many autoimmune diseases. The T cells that help B cells in autoimmune diseases vary in phenotype and include T cells that lack typical features of T follicular helper cells, such as expression of CXCR5 and BCL6. A population of PD-1hi CXCR5- T peripheral helper (Tph) cells has now been recognized in multiple autoantibody-associated diseases. Tph cells display a distinctive set of features, merging the ability to provide B cell help with the capacity to migrate to inflamed peripheral tissues. Here, we review the scope of immune-related conditions in which Tph cells have been implicated and provide a perspective on their potential contributions to pathologic B cell activation in autoimmune diseases. We discuss Tph cells as a promising therapeutic strategy in autoimmunity and consider the utility of tracking Tph cells in blood as a biomarker to quantify aberrant T cell-B cell activation in patients with autoimmune diseases.

Analysis of the Relationship between Scleritis and T Cell Activation in Patients with Hepatocellular Carcinoma Treated with PD-1 Carrelizumab.

In order to explore the function of inhibiting the immune effect, the relationship between programmed death receptor 1 (PD-1) carrelizumab in the treatment of hepatocellular carcinoma-induced scleritis and T cell activation is investigated. A total of 120 patients with primary liver cancer treated in the department of oncology of our hospital from July 2020 to January 2022 are selected and treated with carrelizumab. According to the occurrence of PD-1 carrelizumab treatment, the patients are divided into the scleritis group and nonscleritis group. The levels of T cells, PD-1, PD-L1 proteins, and serum inflammatory factors at different time points are compared. The experimental results show that the occurrence of scleritis after liver cancer treatment with PD-1 carrelizumab is closely associated with Treg cells, the percentage of Th17 cells, the expression of PD-1, PD-L1 proteins, and inflammatory factors. It is clearly evident that PD-1 carrelizumab can increase the risk of scleritis by affecting T cell activation.

Subclinical Corneal Nerve Fiber Damage and Immune Cell Activation in Systemic Lupus Erythematosus: A Corneal Confocal Microscopy Study.

PurposeThe purpose of this study was to evaluate the utility of corneal confocal microscopy (CCM) in identifying small nerve fiber damage and immune cell activation in patients with systemic lupus erythematosus (SLE).MethodsThis cross-sectional comparative study included 39 consecutive patients with SLE and 30 healthy control participants. Central corneal sensitivity was assessed using a Cochet-Bonnet contact corneal esthesiometer and a laser scanning CCM (Heidelberg, Germany) was used to quantify corneal nerve fiber density (CNFD), nerve branch density (CNBD), nerve fiber length (CNFL), and Langerhans cell (LC) density.ResultsAge was comparable among patients with SLE (33.7 ± 12.7) and controls (35.0 ± 13.7 years, P = 0.670) and the median duration of disease was 3.0 years (2.0–10.0 years). CNBD (P = 0.003) and CNFL (P = 0.019) were lower and mature LC density (P = 0.002) was higher, but corneal sensitivity (P = 0.178) and CNFD (P = 0.198) were comparable in patients with SLE compared with controls. The SELENA-SLEDAI score correlated with CNFD (ρ = −0.319, P = 0.048) and CNFL (ρ = −0.373, P = 0.019), and the total and immature LC densities correlated with CNBD (ρ = −0.319. P = 0.048, and ρ = −0.328, P = 0.041, respectively). Immature LC density was higher (P = 0.025), but corneal sensitivity and nerve fiber parameters were comparable between patients with (33%) and without neuropsychiatric symptoms and SLE.ConclusionsCorneal confocal microscopy identifies distal corneal nerve fiber loss and increased immune cell density in patients with SLE and corneal nerve loss was associated with disease activity.Translational RelevanceCorneal confocal microscopy may enable the detection of subclinical corneal nerve loss and immune cell activation in SLE.

The Thalidomide Analogue (Actimid) Induces T-Cell Activation at Day 28 in Patients with Relapsed Refractory Myeloma but Does Not Correlate with Improved Outcome.

Introduction:Thalidomide analogues such as CC-4047 (Actimid, Celgene) have shown efficacy in treatment of relapsed myeloma but the exact mode of action remains unclear (Streetly M, et al ASH 2003). Preliminary work (Schey S, JCO 2004) suggests that thalidomide analogues can activate T-cells. We have demonstrated that measurement of serum free light chains at day 7 of treatment with Actimid can indicate likelihood of response (Patten P, et al, ASH 2003). Interleukin-2 (IL-2) secretion is increased in patients with myeloma on Actimid although this is not associated with clinical response (Ahsan G, et al Haematologica. Suppl.1 2005). We have now looked at expression of activation markers on T cells following 28 days of CC-4047.Aim:The aim of this study was to see if CC-4047 induces activation of T-cells and whether this correlated with clinical response as assessed by use of the serum free light chain (SFL) assay.Patients and Method:17 Patients (median age 59, range 34–75 years) participated in a phase I/II dose escalation study with CC-4047. All patients received treatment with oral CC-4047, on alternate day schedules. Serum was collected for IL-2 and serum free light chain. EDTA samples were taken for FACS analysis, which was performed on separated lymphocytes. 4 colour flow cytometry was performed using CD3/CD4/CD8/CD56 and CD69 monoclonal antibodies (Beckman Coulter, UK). Analysis for IL-2 and serum free light chains were performed as previously described (Ahsan et al Haematologica 2005 suppl.1)Results:Using conventional assessment 9/17 patients had evidence of response (CR, VGPR, PR) and 8 had stable disease (<25% reduction in paraprotein), In 9 patients there was complete serum free light chain data pre and at day 7. There was significant increased expression of CD69 (p=0.01) on the T-cells. A linear regression analysis was performed between changes in CD69 levels and changes in SFL ratio. There was no significant correlation found (R=0.1) table 1.Table 1PatientDose (mg)CD69 (Absolute count)Serum Free Light ChainBest responseAlt day10/lPrePostPrePost1128.426963SD223.65.516595SD3216.7--PR424961835.5CR552.4915065VGPR657.67.42527VGPR7510.652.315.41SD8515.210.63632SD9524.294830VGPR10531.19.75928SD11107.44.6--MR12101.77.9--VGPR13105.65.3--MRConclusions:This study showed that CD69 expression is increased on T cells in myeloma patients treated with CC-4047. This was detectable at day 28 of treatment. There did not appear to be a correlation between CD69 expression at day 28 and change in serum free light chain ratio. In line with other studies we have identified T cell activation in patients treated with the thalidomide analogue CC-4047. This does not appear to correlate with likelihood of response and it may be that response is due to activation of NK cells or that sustained activation of T-cells after 28 days is important for response. Further work is on-going to address these questions.

P60.05 Influence of Different Concentrations of Nivolumab on T Cell Activation in Patients with Non Small Cell Lung Cancer: in Vitro Study

P60.05 Influence of Different Concentrations of Nivolumab on T Cell Activation in Patients with Non Small Cell Lung Cancer: in Vitro Study

MiR-150-5p regulate T cell activation in severe aplastic anemia by targeting Bach2

MiR-150-5p is an immune-related miRNA and elevated in the plasma of patients with aplastic anemia (AA), but its role in T cell activation in patients with severe aplastic anemia (SAA) is unclear. This study aims to explore the role of miR-150-5p in T cell activation of SAA. RT-PCR and Western blot were used to detect the expression of mRNA and protein. The cell proportion was detected by flow cytometry. The lentiviruses expressing miR-150-5p inhibitor and Bach2 shRNA were respectively infected to produce stable miR-150-5p or Bach2 knockout cells. Brdu incorporation method was used to detect T cell proliferation. SAA mouse model was induced with cyclophosphamide and busulfan, and intravenous injection of LV inhibitor NC and LV-miR-150-5p inhibitor. The miR-150-5p expression is up-regulated in SAA, which is negatively correlated with Bach2. Inhibition of miR-150-5p reduces the activation of T cells. MiR-150-5p directly targeted 3'UTR of Bach2. Moreover, the expression of miR-150-5p and the activation of T cells were decreased in SAA mouse model. MiR-150-5p promotes T cell activation in SAA by targeting Bach2. Targeting miR-150-5p may be a new molecular therapy for SAA patients.

Programmed cell death 1-expressing CD56-negative natural killer (NK) cell expansion is a hallmark of chronic NK cell activation during dasatinib treatment.

Dasatinib treatment markedly increases the number of large granular lymphocytes including natural killer (NK) cells in a proportion of Ph+ leukemia patients, which associates with a better prognosis. In‐depth immune profiling of NK cells can predict therapeutic response in these patients. In the present study, we showed that CD56‐negative (CD56neg) NK cells increased exclusively in cytomegalovirus‐seropositive (CMV+) patients treated with dasatinib. The increase longitudinally paralleled with progressive differentiation of CD56dim NK cells during dasatinib therapy driven by CMV reactivation as shown by principal component analysis on 19 NK cell markers. The CD56neg NK cells showed downregulation of NK‐activating receptors, upregulation of PD‐1, and lower cytotoxicity and cytokine production, indicating that these cells are anergic and dysfunctional as seen in chronic infections with HIV‐1 or hepatitis C virus. Moreover, cytolytic activity of CD56dim and CD56neg NK cells against leukemia cells was partially restored by nivolumab in proportion to the frequency of PD‐1+ NK cells. The proportion of patients who achieved deep molecular responses at 2 years was significantly higher in dasatinib‐treated patients with ≥3% CD56neg NK cells than in those with fewer CD56neg NK cells (54.5% vs 15.8%, P = .0419). These findings suggest that CD56neg NK cells may be an exhausted population induced by chronic activation through CMV reactivation during dasatinib therapy. Expansion of CD56neg NK cells is a hallmark of chronic NK cell activation in patients treated with dasatinib and may predict a better clinical outcome. Furthermore, PD‐1 blockade may enhance anti‐leukemia responses of such NK cells.

Mesenchymal stem cells negatively regulate CD4+ T cell activation in patients with primary Sj&ouml;gren syndrome through the miRNA‑125b and miRNA‑155 TCR pathway.

Treatment with mesenchymal stem cells (MSCs) has been revealed to suppress CD4+ T cells and autoimmunity in both mouse models and patients with primary Sjögren syndrome (pSS); however, the underlying mechanism remains unclear. MicroRNAs (miRNAs or miRs) mediate CD4+ T cell activation, but the mechanism is not understood, particularly for CD4+ T cells treated with MSCs. Characterization of miRNAs may reveal pSS pathogenesis, guide MSC treatment and provide more personalized management options. The present study aimed to perform an miRNome analysis of quiescent and T cell receptor (TCR)-activated CD4+ T cells treated with MSCs via miRNA profiles and bioinformatics. Following 72 h of co-culture, MSCs inhibited TCR-induced CD4+ T cell activation and decreased IFN-γ levels. The numbers of aberrant miRNAs in pSS naïve (vs. healthy naïve), pSS activation (vs. pSS naïve), MSC treatment and pre-IFN-γ MSC treatment (vs. pSS activation) groups were 42, 55, 27 and 32, respectively. Gene enrichment analysis revealed that 259 pathways were associated with CD4+ T cell stimulation, and 240 pathways were associated with MSC treatment. Increased miRNA-7150 and miRNA-5096 and decreased miRNA-125b-5p and miRNA-22-3p levels in activated CD4+ T cells from patients with pSS were reversed by MSC treatment. Notably, the proliferation of CD4+ T cells and CD4+ IFN-γ+ cells, expression levels of miRNA-125b-5p and miRNA-155 in CD4+ T cells and supernatant IFN-γ secretion were associated with disease activity. miRNA may play a vital role in MSC treatment for activated CD4+ T cells. The results indicated that the expression levels of miRNA-125b-5p and miRNA-155 in TCR-activated CD4+ T cells from patients with pSS may provide insight regarding autoimmune diseases and offer a novel target for prospective treatment. Therefore, these results may be crucial in providing MSC treatment for pSS.

Biomarkers of Hypercoagulability and Endothelial Cell Activation in Patients with Severe COVID-19 Admitted at the Intensive Care Unit. the Roadmap-COVID-19 Prospective Observational Study

Introduction: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes coronavirus disease 2019 (COVID-19) characterized by acute pneumonia which may progress to respiratory failure and life-threatening complications, including acute respiratory distress syndrome (ARDS) and multisystem organ failure with fatal outcome. COVID-19 should be regarded as a systemic disease involving multiple systems. COVID-19 is associated with excessive inflammation, platelet activation, endothelial dysfunction, blood coagulation activation and fibrin formation. The development of a screening algorithm for the evaluation of endothelial and blood coagulation activation is an urgent need for the optimisation of the management of the patients with severe COVID-19. To this aim biomarkers of endothelial cell activation and hypercoagulability have central placeAim: The prospective observational monocentric cohort study ROADMAP-COVID-19 study aimed to evaluate the biomarkers of hypercoagulability and endothelial cell activation which they could be used in the development of a screening algorithm in patients with COVID-19Methods: The ROADMAP-COVID-19 study enrolled 90 consecutive patients hospitalized at Tenon University Hospital (AP-HP, Paris) between March 18th and April 30th, 2020 with confirmed SARS-CoV-2 infection admitted to the ICU with clinically deteriorated severe COVID-19 (ICU- group). Patients were assessed upon the first day to the ICU admission. The control group consisted 30 healthy individuals (C-group). All patients and controls were evaluated for, clotting times (PT and aPTT), fibrinogen, D-Dimers, antithrombin (AT), protein C (PC) and protein S (Ps) activity, clotting factors V (FV), VIII (FVIII) and XII (FXII), high molecular weight kininogen (HK), von Willebrand factor (FvW), thrombin generation assay (TG) using TF 20 pM-PPP-Reagent® (Thrombogram-Thrombinoscope from Stago), fibrin monomers (FM), free TFPI and TF activity, thrombomodulin activity (TM), TF expressing microparticles (the ratio TF/TFPI was also calculated) and Heparanase. The IL-6 levels were also measured. All patients hospitalized in conventional medical department or in ICU routinely received thromboprophylaxis with body weight adapted enoxaparin. The ratio of the mean values of each studied biomarker in the ICU and the C-group was calculated. Biomarkers were classified in four clusters (Decrease, Stable, Slight Increase, Moderate Increase and High Increase).Results :The ICU-group consisted of 102 patients; 87% of these patients were admitted to the ICU directly from the emergency department. Males were 76 out of 102 patients. Age ranged from 30 to 93 years in the W-group. The changes of the biomarkers in patients with COVID-19 admitted at the ICU as compared to healthy individuals with are shown in Table 1. The ICU patients with COVID-19 were characterized by marked increase of IL-6 followed by increase of heparanase and D-dimers. The PPL-ct showed the most important decrease. Patients showed a significant decrease in FXII levels which was associated with a moderate increase of kininogen levels. The levels of the natural coagulation inhibitors as well as the clotting times (PT and aPTT) did not significantly modify in patients as compared to the controls. The rate of the propagation phase of thrombin generation (MRI) and the Peak of thrombin were significantly decrease and the lag-time was prolonged reflecting the antithrombotic effect of treatment with LMWH. Heparanse, free TFPI, and TM showed moderate or high increase. The decreased Ratio of TF/TFPI indicated that the free TFPI release dominated over the release of soluble TF activity.Conclusion: Patients with severe COVID-19 requiring ICU admission showed inhaled inflammatory reaction, hyperocoagulability and endothelial cell activation whereas platelet activation appeared to be secondary. Among the studied biomarkers the PPL-ct D-dimers and heparanase followed by TFPI, FvW, FM and TM levels appear to be mandatory for hypercoagulability of cellular origin. Interestingly patients with severe COVID-19 showed consumption of FXII indicating intrinsic clotting pathway activation and high levels of HK which also indicates endothelial cell activation. The clinical relevance of these biomarkers for early detection of hypercoagulability of cellular origin in patients with COVID-19 is being evaluated in prospective study.DisclosuresNo relevant conflicts of interest to declare.