CD20 B Cell Depleting Therapy Is Associated with up-Regulation of CD8+CD25highFoxp3+ T Regulatory Cells in a Murine Model of Immune Thrombocytopenia (ITP)
Bibliographic record
Abstract
Abstract Primary immune thrombocytopenia (ITP) is an autoimmune disease characterized by increased platelet destruction and/or impaired megakaryocyte production, mediated by autoreactive B cells and T cells. B cell depletion therapy by rituximab, a monoclonal human anti-CD20 antibody, has been shown effective in both anti-platelet antibody positive (B cell mediated) and negative (T cell mediated) ITP patients. Those patients responsive to rituximab therapy showed normalized CD4+ and CD8+ T cell responses (Stasi et al. Blood. 2007), however, the mechanism of T cell regulation by B cell depletion is not clear. One possibility is through normalization of CD4+ T helper cells or up-regulation of CD4+ regulatory T cells (Tregs) (Stasi et al. Blood. 2008). Another possibility is by suppression of activated conventional CD8+ T cells or the up-regulation of CD8+ Tregs. We examined the changes of both CD4+ and CD8+ T cells and Tregs (CD25highFoxp3+) after B cell depletion in vivo in our ITP mouse model. Briefly, BALB/c GPIIIa (CD61) KO mice were either given PBS (ND) or mouse monoclonal anti-CD20 antibody (B-dep, Biogen) at day -1 and day 13 (250ug/mouse, ip). Residual CD19+ B cells in peripheral blood were less than 0.1% within 24hours in the latter group. All mice were immunized by transfusions of wildtype (WT) platelets at day 0, 7, 14, and 21 (1×108/mouse, iv). At day 28, we examined the percentages of T cell subsets in the spleens of the immunized mice. B cell-depleted immune CD61 KO mice showed significantly higher percentages of both CD3+CD8+ T cells and CD8+CD25highFoxp3+ T cells (Table 1). There was no significant difference in the CD3+CD4+ and CD4+CD25highFoxp3+ T cell populations. Both ND and B-dep immune CD61 KO splenocytes showed increased cytotoxicity activity against CD61+ PU5-1.8 target cells in vitro compared with naïve CD61 KO splenocytes, indicating the activation of CD8+ T cells. To test their in vivo effect on ITP development, splenocytes were engrafted from immune mice into irradiated and AsialoGM-1 treated severe combined immunodeficient (SCID) mice at a dose of 2.5×104/mouse and the mice were monitored for weekly platelet counts. ND and in vitro B cell depleted splenocytes from immune KO mice induced persistent ITP during 3 weeks observation whereas splenocytes from B-dep immune mice did not. To further confirm the role of B cell depletion on CD8+ T cell responses, CD8+ T cells from either ND or B-dep immune CD61 KO splenocytes were purified and transferred into SCID mice at 3×104/mouse. CD4+ T cells from ND immune CD61 KO splenocytes were added at 3×104/mouse to all the SCID mice to support the CD8+ T cell survival in vivo. SCID mice received CD8+ T cells from B-dep group showed higher platelet count at Day 14. Overall, our results indicate a protective role of CD8+CD25highFoxp3+ T cells against the development of cell mediated ITP that is enhanced by B cell depleting therapy in vivo. Table 1. CD61 KO MouseSpleens CD3+CD8+(%) CD8+CD25highFoxp3+ (%) Naïve Control 9.12±0.37 0.12±0.08 Immune, ND 6.78±2.37 0.0925±0.03 Immune, B-dep 14.15±5.1 0.2367±0.11 P value (ND vs B-dep) 0.0007 0.0064 Disclosures No relevant conflicts of interest to declare.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.001 | 0.000 |
| Meta-epidemiology (broad) | 0.001 | 0.001 |
| Bibliometrics | 0.001 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.001 | 0.002 |
| Insufficient payload (model declined to judge) | 0.002 | 0.001 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".