Immune Response to SARS-CoV-2 Vaccine in 2 Men

<b><i>Introduction:</i></b> In the trials of corona virus vaccines, detailed analyses of subsets of lymphocytes were not carried out. We present perhaps the most comprehensive immunological analysis of 29 subsets of B and T cells in 2 healthy subjects receiving 2 doses of the Pfizer SARS-CoV-2 (COVID-19) vaccine. <b><i>Methods:</i></b> Analyses were performed prior to vaccination, 3 weeks following the 1st dose, and 4 weeks following the 2nd dose. Total, naïve (T_N), and different memory and effector subsets (T_CM, T_EM, and T_EMRA) of CD4+ and CD8+ T cells; SARS-CoV-2 spike protein-specific tetramer+, and cytotoxic CD8+ T; subsets of T follicular cells (T_FH, T_FH1, T_FH2, T_FH1/T_FH17, and T_FH17); B-cell subsets (mature B cells, naive B cells, transitional B cells, marginal zone B cells, class-switched memory B cells, germinal center B cells, and CD21^low B cells), and plasmablasts; and regulatory lymphocytes (CD4+ Treg, CD8+ Treg, Breg, and T_FR cells) were evaluated with specific monoclonal antibodies by flow cytometry. <b><i>Results:</i></b> A lack of COVID-19 IgG antibodies after the 1st dose in one of 2 subjects was associated with increased regulatory lymphocytes and decreased plasmablasts. Seroconversion after the 2nd dose in this subject was associated with decreased T_FR cells and increased plasmablasts. In both subjects, CD4 T_EM and CD8 T_CM were markedly increased following the 2nd dose. T_FH1 and regulatory lymphocytes were increased (except Breg) following the 1st dose. A striking increase in SARS-CoV-2-specific CD8+ T cells was observed following the 2nd dose. <b><i>Conclusion:</i></b> Our data support the need for 2nd dose of vaccine to induce strong SARS-CoV-2 CD8 T-cell specific response and generation of memory subsets of CD4+ and CD8+ T cells. Regulatory lymphocytes appear to play a role in the magnitude of response.

Follicular Regulatory T Cells in Systemic Lupus Erythematosus

Follicular regulatory T (Tfr) cells are the regulatory T cell subset mainly localized in the germinal center (GC), acting as modulators of GC responses. They can disrupt Tfh cell- and B cell-linked recognition, induce Tfh apoptosis, and suppress B cell function. Evidences show that dysregulated Tfr cells are associated with the disease activity index and serum autoantibody levels, influencing the development of systemic lupus erythematosus (SLE). This review focuses on the interaction among Tfr, Tfh, and B cells, summarizes the characterization and function of Tfr cells, concludes the imbalance of CD4+T subsets in SLE, and presents potential therapies for SLE. In general, we discuss the roles of Tfr cells in the progress of SLE and provide potential treatments.

AB0017 IMMUNE CHARACTERISTICS OF PERIPHERAL BLOOD IN SECONDARY SJOGREN’S SYNDROME PATIENTS WITH RHEUMATOID ARTHRITIS

Background:Secondary Sjogren’s Syndrome (sSS) is diagnosed when symptoms of SS coexist with other systemic connective tissue disease, often secondary to rheumatoid arthritis(RA).The occurrence of SS secondary with RA will worsen the course of disease and increase the high incidence and mortality of RA. At present, the immune characteristics of peripheral blood of sSS with RA are not clear.Objectives:To observe the difference of immune Immune characteristics in peripheral blood between sSS secondary to RA, primary Sjogren’s syndrome(pSS) and RA patients.Methods:20 sSS with RA patients, 20 pSS paients and 20 RA pateints hospitalized in ShanXi medical university the second Hospital were enrolled. The percentage and absolute numbers of lymphocyte phenotypes and CD4+ T subsets in peripheral blood were examined by flow cytometry.Results:As for the percentage and absolute number of total T, B, NK, CD4+T,CD8+ T and the ratio of CD4 + T to CD8+ T cells, there was no significant difference between the sSS with RA, RA, and SS group. There was also no statistical difference in the percentage of CD4+T subsets(Th1,Th2,Th17 and Treg) between the three groups. But the ratio of Th17 and Treg in sSS with RA group was increased than pSS group.About comparison of absolute number of CD4+T subsets, there was no statistical difference among the three groups except that the Th1 cells in RA group was significantly higher than SS group.Conclusion:Elevated Th17/Treg may be an immunological feature that differentiates sSS with RA patients from pSS patients. In addition, in general, peripheral blood of patients with RA and SS have similar immune characteristics.References:[1]Wei W,Ahmad S S, Chi S. From Molecular Mechanism to the Etiology of Sjogren Syndrome.Curr Pharm Des. 2018;24(35):4177-4185.[2]Hajiabbasi A, Masooleh I S,Alizadeh Y, Secondary Sjogren’s Syndrome in 83 Patients With Rheumatoid Arthritis.Acta Med Iran.2016;54(7):448-53.Figure 1.The comparsion about the lymphocyte phenotypes and CD4+ T subsets in peripheral blood of sSS with RA(n=20), pSS(n=20) and RA patients(n=20).(*p<0.05,**p<0.001,*p<0.0001)Disclosure of Interests:None declared

POS0396 THE LEVEL OF PERIPHERAL REGULATORY T CELLS IS ASSOCIATED WITH THE CHANGES OF INTESTINAL MICROBIOTA IN PATIENTS WITH RHEUMATOID ARTHRITIS

Background:Rheumatoid arthritis (RA) is a systemic autoimmunity inflammation disease characterized with chronic aggressive arthritis and the presence of abnormal antibodies. Several observations showed that the breakdown of immune tolerance caused by many complex interactions was involved in the development of RA[1]. However, the pathogenesis of RA remained unclear. It has been confirmed that the imbalance of Th17 and Treg cells play a crucial role in destroying immune tolerance [2]. Besides, researches showed that intestinal microbiota can influence host immunity by acting on the immune cells to play pro-inflammatory or anti-inflammatory effect, and in turn immune system can also regulate the microbiota[3, 4]. Thus, a frontier point of view in the field of rheumatism, immune microecology, was proposed, which is a novel concept for the breakdown of immune tolerance. Studies have confirmed that there was an imbalance of intestinal microbiota in patients with RA [4]. But the relationship between the CD4+T subsets cells and intestinal microbiota in RA is unknown.Objectives:We detected and compared the absolute number of CD4+T cells subsets in the peripheral blood and the proportion or abundance of intestinal microbiota in patients with RA and healthy adults, and then analyzed the relationship between them to explore the role of CD4+T cells subsets and intestinal microbiota in the pathogenesis of RA.Methods:We collected the sample of stool and blood from 15 patients with RA hospitalized at the Second Hospital of Shanxi Medical University and 8 age and gender-matched healthy controls(HC). The absolute number of CD4+T cells subsets including Th1, Th2, Th17 and Treg cells were detected by flow cytometry. The 16S rRNA in the stool specimens were sequenced by the Roche/45 high-throughput sequencing platform. We analyzed whether there was correlarion between CD4+T subsets cells and intestinal microbiota.Results:Patients with RA had a higher level of Christensenellaceae and a lower level of Pseudomonadaceae as compared with those of HCs at the family level (p<0.05). And at the genus level, the patients with RA had higher levels of Ruminococcus torques, Christensenellaceae R-7, Ruminiclostridium 9 and Ruminococcus 1 compared with those of HCs (p<0.05) (Figure 1).And the Ruminococcus torques at the genus level was negative correlated with the absolute number of Treg cells (p<0.001) (Figure 2).Conclusion:The results here suggested that there were different proportion or abundance of intestinal microbiota between the patients with RA andHCs. And the changes of intestinal microbiota such as Ruminococcus torques were associated with Treg cells, further indicating that the imbalance of intestinal microbiota in RA can destory the immune tolerance. The above results uncovered that the intestinal microbiota had immunomodulatory function, which may be the upstream mechanism participated in the pathogenesis of RA.References:[1]Weyand CM, Goronzy JJ. The immunology of rheumatoid arthritis. Nat Immunol 2021, 22(1): 10-18.[2]Weyand CM, Goronzy JJ. Immunometabolism in the development of rheumatoid arthritis. Immunol Rev 2020, 294(1): 177-187.[3]Brown EM, Kenny DJ, Xavier RJ. Gut Microbiota Regulation of T Cells During Inflammation and Autoimmunity. Annu Rev Immunol 2019, 37: 599-624.[4]du Teil Espina M, Gabarrini G, Harmsen HJM, Westra J, van Winkelhoff AJ, van Dijl JM. Talk to your gut: the oral-gut microbiome axis and its immunomodulatory role in the etiology of rheumatoid arthritis. FEMS Microbiol Rev 2019, 43(1).Figure 1.At the family level (a-b) and the genus level(c-f), the relative abundance of intestinal microbiota in patients with RA and HCs were different. Data were expressed as median (Q1, Q3) and analyzed by Wilcoxon test. (*** P < 0.001, **P < 0.01 and *P < 0.05).Figure 2.A heatmap shows the correlation between the intestinal microbiota and CD4+T cells in patients with RA, and Ruminococcus torques at the genus level was negative related with Treg cells. (Colors indicate the Spearman rank correlation, *** P < 0.001).Disclosure of Interests:None declared

Increased Mitochondrial Mass Related To Heightened Pyroptosis of CD4+T Cells In HIV-1 Infection

Background: In HIV-1 infection, over 90% CD4+T cells die of caspase-1 mediated pyroptosis. What governs the increased susceptibility of CD4+T cells to pyroptosis is poorly understood. Method: Blood samples were obtained from 31 ART-naive HIV-infected patients, 29 ART-exposed HIV-infected patients, and 21 healthy control donors. Plasma levels of IL-18 and IL-1β, activated caspase-1, mitochondrial mass (MM) and mitochondrial fusion/fisson genes of CD4+T subsets were measured. Results: Significantly higher IL-18 level of plasma and MM level of CD4+T cells were found in HIV-infected patients than that in healthy controls, and the MMhigh phenotype manifested more sensitivity to caspase-1 mediated pyroptosis. Moreover, the increased MM was more pronounced in the early differentiated and inactivated CD4+T cells. However, higher MM was not intrinsically linked to T cell differentiation disorder or excessive activation of the CD4+T cells. Mechanistically, the increased mitochondrial mass was significantly correlated with an elevated expression level of the mitochondrial fusion gene-mitofusin1.Conclusion: MM increase associates with heightened sensitivity of CD4+T cells to pyroptosis even in early differentiated and unactivated CD4+T cells in patients with HIV-1 infection, regardless of whether the patients are on HAART or not. These new revelations uncovered a previously unappreciated challenge to immune reconstitution with antiretroviral therapy.

Traditional Thai Massage Promoted Immunity in the Elderly via Attenuation of Senescent CD4+ T Cell Subsets: A Randomized Crossover Study

The beneficial physiological effects of traditional Thai massage (TTM) have been previously documented. However, its effect on immune status, particularly in the elderly, has not been explored. This study aimed to investigate the effects of multiple rounds of TTM on senescent CD4+ T cell subsets in the elderly. The study recruited 12 volunteers (61–75 years), with senescent CD4+ T cell subsets, who received six weekly 1-h TTM sessions or rest, using a randomized controlled crossover study with a 30-day washout period. Flow cytometry analysis of surface markers and intracellular cytokine staining was performed. TTM could attenuate the senescent CD4+ T cell subsets, especially in CD4+28null NKG2D+ T cells (n = 12; p < 0.001). The participants were allocated into two groups (low < 2.75% or high ≥ 2.75%) depending on the number of CD4+28null NKG2D+ T cells. After receiving TTM over 6 sessions, the cell population of the high group had significantly decreased (p < 0.001), but the low group had no significant changes. In conclusion, multiple rounds of TTM may promote immunity through the attenuation of aberrant CD4+ T subsets. TTM may be provided as a complementary therapy to improve the immune system in elderly populations.

Low-dose IL-2 therapy limits the reduction in absolute numbers of circulating regulatory T cells in rheumatoid arthritis

Background: Circulating regulatory T cells (Tregs) are responsible for mediating immune tolerance and maintaining immunological homeostasis. Decreases in Tregs may be involved in the onset of rheumatoid arthritis (RA). Low-dose interleukin-2 (IL-2) has been considered for the treatment of inflammatory diseases mediated by T cells. This study focused on the status of circulating CD4+T subsets and the clinical feasibility of IL-2 therapies in patients with RA. Methods: The subjects included 888 patients with RA and 100 healthy controls (HCs); 233 RA patients received IL-2 treatment with 0.5 million international units (MIU)/day from days 1 through 5. The demographic features, disease activity, and levels of CD4+T cells measured by modified flow cytometry were collected in all RA patients before and after treatment. Results: RA patients had lower absolute Treg counts (but not Th17) compared with HCs, which was associated with disease activity; previously treated RA patients had the fewest circulating Tregs ( p < 0.05). Patients treated with low-dose IL-2 had a three-fold increase in absolute anti-inflammatory Treg counts, as well as a two-fold increase in the other CD4+T subsets. Moreover, post-treatment levels of markers of disease activity in RA patients treated with IL-2 were significantly lower than the baseline values ( p < 0.001), with no apparent side effects. Conclusion: Decreased absolute counts of circulating CD4+T lymphocyte subsets were observed in patients with RA. Circulating Tregs, which mediate immune tolerance, may be involved in the pathogenesis and progression of RA; however, this was ameliorated by low-dose IL-2, without obvious side effects. Plain language summary Low-dose IL-2 treatment for rheumatoid arthritis • Circulating Tregs may be involved in the pathogenesis and progression of RA. • The absolute count of Tregs was significantly correlated with disease activity measures. • Low-dose IL-2 was able to effectively expade Tregs and help for RA patients’ symptoms remission without evaluated side effects.

Low-Dose IL-2 Therapy Limits the Reduction in Absolute Numbers of Circulating Regulatory T Cells in Rheumatoid Arthritis

Background Circulating regulatory T cells (Tregs) are responsible for mediating immune tolerance and maintaining immunological homeostasis. Decreases in Tregs may be involved in the onset of rheumatoid arthritis (RA). Low-dose interleukin-2 (IL-2) has been considered for the treatment of inflammatory diseases mediated by T cells. This study focused on the status of circulating CD4+T subsets and the clinical feasibility of IL-2 therapies in patients with RA.Methods The subjects included 888 patients with RA and 100 healthy controls (HCs); 233 RA patients received IL-2 treatment of at 0.5 million international units (MIU)/day from days 1 through 5. The demographic features, disease activity, and levels of CD4+T cells measured by modified flow cytometry were collected in all RA patients before and after treatment.Results RA patients had lower absolute Treg counts (but not Th17) compared with HCs, which was associated with disease activity; previously treated RA patients had the fewest circulating Tregs (P < 0.05). Patients treated with low-dose IL-2 had a three-fold increase in absolute anti-inflammatory Treg counts, as well as a two-fold increase in the other CD4+T subsets. Moreover, post-treatment levels of markers of disease activity in RA patients treated with IL-2 were significantly lower than the baseline values (P < 0.001), with no apparent side effects.Conclusions Decreased absolute counts of circulating CD4+T lymphocyte subsets were observed in patients with RA. Circulating Tregs, which mediate immune tolerance, may be involved in the pathogenesis and progression of RA; however, this was ameliorated by low-dose IL-2, without obvious side effects.

Dissecting Common and Unique Effects of Anti-α4β7 and Anti-Tumor Necrosis Factor Treatment in Ulcerative Colitis

Background and Aims Vedolizumab is an anti-α4β7 antibody approved for the treatment of ulcerative colitis [UC]. Although it is assumed that vedolizumab blocks intestinal homing of lymphocytes, its effects on different intestinal cell populations are not fully stablished. In order to establish the unique mechanisms of action of vedolizumab in UC patients, we compared its effects to those induced by anti-tumour necrosis factor [TNF]. Methods Patients with active UC [endoscopic Mayo score &gt;1] starting vedolizumab [n = 33] or anti-TNF [n = 45] and controls [n = 22] were included. Colon biopsies [at weeks 0, 14 and 46] and blood samples [at weeks 0, 2, 6, 14, 30 and 46] were used for cell phenotyping, transcriptional analysis [qPCR], and to measure receptor occupancy. Results Vedolizumab, in contrast to anti-TNF, significantly reduced the proportion of α4β7+ cells within intestinal T subsets while preserving the percentage of α4β7+ plasma cells. The marked decrease in α4β7 did not change the percentage of colonic αEβ7+ cells [at 46 weeks]. Both vedolizumab and anti-TNF significantly downregulated inflammation-related genes in the colon of responders [Mayo score &lt; 2]. Moreover, both treatments significantly decreased the percentage of intestinal, but not blood, total lymphocytes [T and plasma cells], as well as the proportion of α4β1+ cells within intestinal T lymphocytes. Conclusions Our data show that while vedolizumab and anti-TNF block two unrelated targets, they induce remarkably similar effects. On the other hand, vedolizumab’s unique mechanism of action relies on blocking intestinal trafficking of α4β7 T cells, despite effectively binding to B and plasma cells that express α4β7.

Intra-tumoral injection of caerin 1.1 and 1.9 peptides in vaccinated TC-1 tumour bearing mice with PD-1 blockade modulates macrophage heterogeneity and the activation of CD8+ T cells in the tumour microenvironment

Development of a vaccine formula that alters the tumour-infiltrating lymphocytes to be more immune active against a tumour is key to the improvement of clinical responses to immunotherapy. Here, we demonstrate that, in conjunction with E7 antigen specific immunotherapy, and IL-10 and PD-1 blockade, intra-tumoral administration of caerin 1.1 and 1.9 peptides further improves the tumour microenvironment (TME) when compared with injection of a control peptide. We used single cell transcriptomics and mass spectrometry-based proteomics to quantify changes in cellular activity across different cell types within the TME. We show that the injection of caerin 1.1/1.9 increases immune activating macrophages and NK cells, while reducing immunosuppressive macrophages with M2 phenotype, and increased numbers of activated CD8+ T cells with higher populations of memory and effector-memory CD8+ T subsets. Proteomic profiling demonstrated activation of Stat1 modulated apoptosis and production of nitric oxide. Further, computational integration of the proteome with the single cell transcriptome was consistent with deactivation of immune suppressive B cell function following caerin 1.1 and 1.9 treatment.