Session Type: ACR Poster Session B
Session Time: 9:00AM-11:00AM
Background/Purpose: Lymphopenia has been associated with the development of autoimmune diseases. Particularly, in patients with Systemic Lupus Erythematosus (SLE) is a common clinical feature. However, the molecular mechanisms related to lymphopenia in SLE have not been fully addressed. Besides, the E3 ligase Cbl-b has been shown to regulate T cell responsiveness and its deficiency has been documented in SLE. The aim of this study was to assess CD4+ T cell number and proliferation regulated by cell cycle inhibitors ubiquitination by the E3 ligase Cbl-b in SLE patients.
Methods: We included 20 patients with SLE (8 in remission and 12 with active untreated disease) according to the classification criteria of the American College of Rheumatology and 20 age and gender-matched healthy controls. Absolute peripheral lymphocyte count was obtained for each subject. PBMCs were isolated by density gradient and effector (CD4+CD25–) T cells were purified by magnetic selection and stimulated with anti-CD3+anti-CD28 beads. The expression of total ubiquitin, Cbl-b, p27kip1 and p21cip1 was analyzed by Western blotting. Interaction between Cbl-b, p27kip and p21cip1 was addressed by immunoprecipitation (IP). Proliferative responses were assessed by CFSE. Differences were assessed by t Student test and Spearman correlation coefficient was also used. p<0.05 was considered as statistically significant.
Results: We found decreased Cbl-b expression in effector T cells from SLE patients in comparison to healthy controls (1.6±0.2 vs 3.8±1.3, p=0.003), which was associated with lower proliferation upon TCR stimulation vs healthy controls (p=0.022). Decreased proliferation and absolute number of effector T cells correlated with Cbl-b expression (r=0.553, p=0.020; r=0.680, p=0.003, respectively). Moreover, this phenomenon was associated with diminished ubiquitination of the cell cycle inhibitor p27kip1 in effector T cells from SLE patients when compared to healthy controls. We did not find differences in ubiquitination and expression of another cell cycle inhibitor, p21cip1 in comparison to healthy controls. Interestingly, we also found by IP assays, that p27kip1 and p21cip1interact with Cbl-b. We found no significant differences regarding to disease activity.
Conclusion: Our data suggest that the deficiency of the E3 ligase Cbl-b is associated with the presence of lymphopenia in SLE patients. This phenomenon is related to decreased ubiquitination of the cell cycle inhibitor p27kip1 in effector T cells from SLE patients, which triggers its sustained expression as well as decreased proliferation of lupus lymphocytes. Furthermore, this effect is specific to p27kip1, since even though, p21cip1 also interacts with Cbl-b, its ubiquitination and expression was not related to this E3 ligase in SLE patients. To our knowledge, this is the first evidence that suggest deficient ubiquitination as a molecular mechanism of lymphopenia in SLE patients.
To cite this abstract in AMA style:Gómez-Martín D, Romo-Tena J, Merayo-Chalico J, Alcocer-Varela J. CD4+ T Cell Lymphopenia Is Associated with Deficient Ubiquitination of the Cell Cycle Inhibibitor p27kip1 By the E3 Ligase Casitas B-Lineage Lymphoma-b in Patients with Systemic Lupus Erythematosus [abstract]. Arthritis Rheumatol. 2016; 68 (suppl 10). https://acrabstracts.org/abstract/cd4-t-cell-lymphopenia-is-associated-with-deficient-ubiquitination-of-the-cell-cycle-inhibibitor-p27kip1-by-the-e3-ligase-casitas-b-lineage-lymphoma-b-in-patients-with-systemic-lupus-erythematosus/. Accessed October 28, 2020.
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/cd4-t-cell-lymphopenia-is-associated-with-deficient-ubiquitination-of-the-cell-cycle-inhibibitor-p27kip1-by-the-e3-ligase-casitas-b-lineage-lymphoma-b-in-patients-with-systemic-lupus-erythematosus/