CD4 T lymphocyte autophagy is upregulated in the salivary glands of primary Sjögren’s syndrome patients and correlates with focus score and disease activity

Background: Primary Sjögren’s syndrome (pSS) is a common chronic autoimmune disease characterized by lymphocytic infiltration of exocrine glands and peripheral lymphocyte perturbation. In the current study, we aimed to investigate the possible pathogenic implication of autophagy in T lymphocytes in patients with pSS. Methods: Thirty consecutive pSS patients were recruited together with 20 patients affected by sicca syndrome a nd/or chronic sialoadenitis and 30 healthy controls. Disease activity and damage were evaluated according to SS disease activity index, EULAR SS disease activity index, and SS disease damage index. T lymphocytes were analyzed for the expression of autophagy-specific markers by biochemical, molecular, and histological assays in peripheral blood and labial gland biopsies. Serum interleukin (IL)-23 and IL-21 levels were quantified by enzyme-linked immunosorbent assay. Results: Our study provides evidence for the first time that autophagy is upregulated in CD4+ T lymphocyte salivary glands from pSS patients. Furthermore, a statistically significant correlation was detected between lymphocyte autophagy levels, disease activity, and damage indexes. We also found a positive correlation between autophagy enhancement and the increased salivary gland expression of IL-21 and IL-23, providing a further link between innate and adaptive immune responses in pSS. Conclusions: These findings suggest that CD4+ T lymphocyte autophagy could play a key role in pSS pathogenesis. Additionally, our data highlight the potential exploitation of T cell autophagy as a biomarker of disease activity and provide new ground to verify the therapeutic implications of autophagy as an innovative drug target in pSS

Standardisation of labial salivary gland histopathology in clinical trials in primary Sjögren's syndrome

Labial salivary gland (LSG) biopsy is used in the classification of primary Sjögren's syndrome (PSS) and in patient stratification in clinical trials. It may also function as a biomarker. The acquisition of tissue and histological interpretation is variable and needs to be standardised for use in clinical trials. A modified European League Against Rheumatism consensus guideline development strategy was used. The steering committee of the ad hoc working group identified key outstanding points of variability in LSG acquisition and analysis. A 2-day workshop was held to develop consensus where possible and identify points where further discussion/data was needed. These points were reviewed by a subgroup of experts on PSS histopathology and then circulated via an online survey to 50 stakeholder experts consisting of rheumatologists, histopathologists and oral medicine specialists, to assess level of agreement (0–10 scale) and comments. Criteria for agreement were a mean score ≥6/10 and 75% of respondents scoring ≥6/10. Thirty-nine (78%) experts responded and 16 points met criteria for agreement. These points are focused on tissue requirements, identification of the characteristic focal lymphocytic sialadenitis, calculation of the focus score, identification of germinal centres, assessment of the area of leucocyte infiltration, reporting standards and use of prestudy samples for clinical trials. We provide standardised consensus guidance for the use of labial salivary gland histopathology in the classification of PSS and in clinical trials and identify areas where further research is required to achieve evidence-based consensus

Diabetes-Related Autoantibodies in Children With Acute Lymphoblastic Leukemia

[No abstract available

Cytokines and inflammatory mediators: 25. Certolizumab Pegol has a Different Profile from the other Anti-TNFS, Including Golimumab, in a Variety of in Vitro Assays

Background: Activities of the anti-TNFs, certolizumab pegol (CZP), etanercept (ETA), infliximab (IFX) and adalimumab (ADA), have been compared in a range of in vitro assays. CZP is the only licensed PEGylated Fab' anti-TNF; ETA is a fusion protein with an IgG1 Fc, and IFX and ADA are both antibodies with an IgG1 Fc. Golimumab (GLM) is a monoclonal IgG1 TNF inhibitor recently approved for a number of indications; it is thus of interest to assess the in vitro activity of GLM. In vitro assays previously used were neutralisation of TNF in the L929 bioassay, inhibition of LPS-driven cytokine production by monocytes, induction of apoptosis in activated lymphocytes and monocytes, and induction of neutrophil necrosis. Methods: Neutralisation of human TNF was assessed in the L929 bioassay using a range of concentrations of the anti-TNFs and a fixed concentration of TNF (100 pg/mL). Activity of the anti-TNFs at inhibiting LPS-driven IL-1β secretion by monocytes was assessed by incubating peripheral blood monocytes with various concentrations of the anti-TNF for 1 hour (hr) and then washing the cells. LPS was added for 4 hrs, the supernatants collected and the IL-1β level measured by ELISA. To assess induction of apoptosis, peripheral blood lymphocytes were activated for 2 days with 2 μg/mL CD3/CD28 and monocytes with 300 U/mL IL-4 and GMCSF for 3 days. The effect of the anti-TNFs on apoptosis was assessed by Annexin V staining using flow cytometry 24 hrs later. The effect of the anti-TNFs on neutrophil necrosis was determined by measuring myeloperoxidase release after 12 hrs. An isotype-matched control was used in all assays except the L929 bioassay. Results: IC90 neutralisation activity of the anti-TNFs in the L929 bioassay was 0.3 ng/mL for ETA, 4 ng/mL for GLM, 15 ng/mL for ADA, and 20 ng/mL for IFX, compared with 2.5 ng/mL for CZP. CZP was the most potent inhibitor of LPS-driven IL-1β secretion (IC50 ∼0.1 ng/mL), followed by GLM (20 ng/mL) and IFX (50 ng/mL). GLM, ADA, IFX and ETA induced apoptosis of monocytes and lymphocytes to a similar degree reaching a level of 23% and ∼40% at 100 μg/mL, respectively. CZP caused no increase in apoptosis above the levels seen with the isotype-matched control. In the neutrophil necrosis assay, ADA,IFX and GLM caused ∼70% necrosis at 100 μg/mL, and ETA 48%. CZP did not increase the level of necrosis above the level of the control. Conclusions: Bioactivity of the IgG1 molecules GLM, IFX and ADA in neutralising human TNF was inferior to that of CZP and ETA. CZP, the only PEGylated anti-TNF, had a different profile to the other anti-TNFs as it was the most potent at inhibiting LPS-driven IL-1β production by monocytes, did not induce apoptosis of activated monocytes and lymphocytes, and did not cause neutrophil necrosis. The clinical relevance of these in vitro effects is unknown. Nevertheless, these assays show interesting in vitro differences between the anti-TNFs. Disclosure statement: G.F. and A.N. are employees of UC

Characterization and outcomes of 414 patients with primary SS who developed haematological malignancies

Objective: To characterize 414 patients with primary SS who developed haematological malignancies and to analyse how the main SS- and lymphoma-related features can modify the presentation patterns and outcomes. Methods: By January 2021, the Big Data Sjögren Project Consortium database included 11 966 patients fulfilling the 2002/2016 classification criteria. Haematological malignancies diagnosed according to the World Health Organization (WHO) classification were retrospectively identified. Results: There were 414 patients (355 women, mean age 57 years) with haematological malignancies (in 43, malignancy preceded at least one year the SS diagnosis). A total of 376 (91%) patients had mature B-cell malignancy, nearly half had extranodal marginal zone lymphoma (MZL) of mucosa-associated lymphoid tissue (MALT lymphoma) (n = 197), followed by diffuse large B-cell lymphoma (DLBCL) (n = 67), nodal MZL lymphoma (n = 29), chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) (n = 19) and follicular lymphoma (FL) (n = 17). Rates of complete response, relapses and death were 80%, 34% and 13%, respectively, with a 5-year survival rate of 86.5% after a mean follow-up of 8 years. There were significant differences in age at diagnosis (younger in MALT, older in CLL/SLL), predominant clinical presentation (glandular enlargement in MALT lymphoma, peripheral lymphadenopathy in nodal MZL and FL, constitutional symptoms in DLBCL, incidental diagnosis in CLL/SLL), therapeutic response (higher in MALT lymphoma, lower in DLBCL) and survival (better in MALT, nodal MZL and FL, worse in DLBCL). Conclusion: In the largest reported study of haematological malignancies complicating primary SS, we confirm the overwhelming predominance of B-cell lymphomas, especially MALT, with the salivary glands being the primary site of involvement. This highly-specific histopathological scenario is linked with the overall good prognosis with a 5-year survival rate of nearly 90%

Left Ventricular Function in Insulin-Dependent and in Non-Insulin-Dependent Diabetic Patients: Radionuclide Assessment.

The aim of this study was to compare, by gated radionuclide angiography, systolic and diastolic ventricular function in insulin-dependent (IDDM) and non-insulin-dependent (NIDDM) diabetic patients without overt cardiovascular disease. The study population consisted of 20 IDDM patients (15 male, 5 female; 40.7 \ub1 10.3 years), 14 NIDDM patients (9 male, 5 female; 47.0 \ub1 7.5 years) and 12 healthy subjects (7 male, 5 female; 41.5 \ub1 6.3 years) as a control (C) group. The duration of diabetes (DD) and glycosylated hemoglobin (HbA1c) levels were significantly higher in the IDDM patients. The ventricular ejection fraction and peak ejection rate (PER) were assessed by gated radionuclide left ventriculography and were similar in three groups, while the peak filling rate (PFR) was lower in the NIDDM patients compared to the IDDM patients (p < 0.05) and controlled healthy subjects (p < 0.01, IDDM = 3.39 \ub1 1.14; NIDDM = 2.65 \ub1 0.83; C = 3.55 \ub1 0.73), the time to PFR was significantly more prolonged in the NIDDM group than in the IDDM (p < 0.05) and C groups (p < 0.05, NIDDM = 162 \ub1 26; IDDM = 140 \ub1 28; C = 142 \ub1 23). The PFR/PER ratio was near the normal value (-1) in the IDDM patients and controlled subjects, while in the NIDDM patients it was reduced (= 0.84 \ub1 0.18). Seven IDDM and 4 NIDDM patients had borderline signs of cardiovascular autonomic neuropathy, unrelated to DD, HbAic and scintigraphic parameters. Left ventricular systolic performance was substantially normal and similar in both the IDDM and NIDDM patients. Ventricular diastolic filling was impaired in the NIDDM patients, as shown by the decrease in PFR and in particular in the PFR/PER ratio. Our radionuclide data suggest that the NIDDM patients had a prevalent abnormality of ventricular diastolic performance, with respect to the IDDM patients, although the latter patients had higher DD and HbA1cvalues