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Longitudinal analysis of T and B cell phenotype and function in renal transplant recipients with or without rituximab induction therapy.

Kamburova EG, Koenen HJ, van den Hoogen MW, Baas MC, Joosten I, Hilbrands LB - PLoS ONE (2014)

Bottom Line: In addition, the ratio between the percentage of central memory CD4+ and CD4+ regulatory T cells was significantly higher up to 24 months post-transplant compared to pre-transplant levels.Addition of a single dose of rituximab resulted in a long-lasting B-cell depletion.Rituximab treatment had no effect on the T-cell phenotype and function post-transplant.

View Article: PubMed Central - PubMed

Affiliation: Department of Laboratory Medicine - Medical Immunology, Radboud University Medical Centre, Nijmegen, The Netherlands.

ABSTRACT

Background: Prevention of rejection after renal transplantation requires treatment with immunosuppressive drugs. Data on their in vivo effects on T- and B-cell phenotype and function are limited.

Methods: In a randomized double-blind placebo-controlled study to prevent renal allograft rejection, patients were treated with tacrolimus, mycophenolate mofetil (MMF), steroids, and a single dose of rituximab or placebo during transplant surgery. In a subset of patients, we analyzed the number and phenotype of peripheral T and B cells by multiparameter flow cytometry before transplantation, and at 3, 6, 12, and 24 months after transplantation.

Results: In patients treated with tacrolimus/MMF/steroids the proportion of central memory CD4+ and CD8+ T cells was higher at 3 months post-transplant compared to pre-transplant levels. In addition, the ratio between the percentage of central memory CD4+ and CD4+ regulatory T cells was significantly higher up to 24 months post-transplant compared to pre-transplant levels. Interestingly, treatment with tacrolimus/MMF/steroids resulted in a shift toward a more memory-like B-cell phenotype post-transplant. Addition of a single dose of rituximab resulted in a long-lasting B-cell depletion. At 12 months post-transplant, the small fraction of repopulated B cells consisted of a high percentage of transitional B cells. Rituximab treatment had no effect on the T-cell phenotype and function post-transplant.

Conclusions: Renal transplant recipients treated with tacrolimus/MMF/steroids show an altered memory T and B-cell compartment post-transplant. Additional B-cell depletion by rituximab leads to a relative increase of transitional and memory-like B cells, without affecting T-cell phenotype and function.

Trial registration: ClinicalTrials.gov NCT00565331.

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Related in: MedlinePlus

Longitudinal analysis of circulating B cells in renal transplant recipients after treatment with tacrolimus, MMF and steroids, and a single dose of rituximab (RTX) during transplant surgery.(A) Representative dot plots of CD19+ B cells within the CD45+ lymphocyte population for a RTX-treated and a triple immunosuppression (IS)-treated patient before transplantation (pre-Tx) and at 3, 12, and 24 months after transplantation. (B) Shown are the absolute numbers of CD19+ B cells for RTX-(gray, n = 12) and triple IS-treated (white, n = 14) patients before transplantation (t = 0) and up to 24 months after transplantation (n = 10 and n = 9 at t = 24 m, respectively). (C) Pie charts depicting the distribution the different B cells subsets over time using the Bm1-Bm5 classification as depicted in Figure 3B: Bm1 (IgD+CD38−), Bm2 (IgD+CD38+), Bm2’ (IgD+CD38++), Bm3+4 (IgD−CD38++), Early Bm5 (IgD−CD38+) and Late Bm5 (IgD−CD38−) cells within the CD19+ B-cell population. Data are represented as means of 14 triple IS+RTX-treated and 12 IS-treated patients before transplantation (pre-Tx) and at 3, 12, and 24 months after transplantation (n = 10 and n = 9 at t = 24 m, respectively). (D) Shown are the percentages of CD80+, CD95+ and BAFF-R+ cells within the CD19+ B-cell population for RTX- (gray, n = 12) and triple IS-treated (white, n = 14) patients before transplantation (t = 0) and up to 24 months after transplantation. Results are shown as box plots displaying the median, 25th and 75th percentiles as the box, and the 5th and 95th percentiles as whiskers. Significant differences are indicated by asterisks: **P<0.01.
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pone-0112658-g004: Longitudinal analysis of circulating B cells in renal transplant recipients after treatment with tacrolimus, MMF and steroids, and a single dose of rituximab (RTX) during transplant surgery.(A) Representative dot plots of CD19+ B cells within the CD45+ lymphocyte population for a RTX-treated and a triple immunosuppression (IS)-treated patient before transplantation (pre-Tx) and at 3, 12, and 24 months after transplantation. (B) Shown are the absolute numbers of CD19+ B cells for RTX-(gray, n = 12) and triple IS-treated (white, n = 14) patients before transplantation (t = 0) and up to 24 months after transplantation (n = 10 and n = 9 at t = 24 m, respectively). (C) Pie charts depicting the distribution the different B cells subsets over time using the Bm1-Bm5 classification as depicted in Figure 3B: Bm1 (IgD+CD38−), Bm2 (IgD+CD38+), Bm2’ (IgD+CD38++), Bm3+4 (IgD−CD38++), Early Bm5 (IgD−CD38+) and Late Bm5 (IgD−CD38−) cells within the CD19+ B-cell population. Data are represented as means of 14 triple IS+RTX-treated and 12 IS-treated patients before transplantation (pre-Tx) and at 3, 12, and 24 months after transplantation (n = 10 and n = 9 at t = 24 m, respectively). (D) Shown are the percentages of CD80+, CD95+ and BAFF-R+ cells within the CD19+ B-cell population for RTX- (gray, n = 12) and triple IS-treated (white, n = 14) patients before transplantation (t = 0) and up to 24 months after transplantation. Results are shown as box plots displaying the median, 25th and 75th percentiles as the box, and the 5th and 95th percentiles as whiskers. Significant differences are indicated by asterisks: **P<0.01.

Mentions: The participation of renal transplant recipients in a randomized double-blind placebo-controlled study evaluating the efficacy and safety of RTX when added to triple drug immunosuppression gave us the opportunity to study the effects of additional B-cell depletion on the phenotype and function of T and B cells. RTX treatment resulted in a nearly complete depletion of B cells from the peripheral lymphocyte population up to 12 months after transplantation. Remarkably, the B-cell depletion after a single dose of RTX was long lasting. The absolute numbers of B cells remained quite low at 24 months after RTX treatment with a median of 9.7 B cells/µl (range 3.0–294.8) compared to a median of 116.4 B cells/µl (range 49.7–379.7) in patients not treated with RTX (Figure 4A–B). Interestingly, at 12 months the percentage of Bm2’ (IgD++CD38++) and Bm3+4 (IgD−CD38++) cells was significantly higher in RTX-treated patients, while the percentages of Bm1 (IgD+CD38−) and Bm2 (IgD+CD38+) cells were lower in RTX-treated patients. (Figure 4C). Accordingly, the percentages of CD24++CD38++ transitional and IgD−CD27+ switched memory B cells were higher in RTX-treated patients, while the percentages of IgD+CD27− naïve B cells were lower in RTX-treated patients (Figure S3A). Fitting with the relative increase of Bm3+4 (IgD−CD38++) cells, there was an increase in the percentage of CD80+ and CD95+ B cells at 12 and/or 24 months after transplantation (Figure 4D). Remarkably, the percentage of BAFF-R+ B cells was lower in RTX-treated patients at 3 and 12 months after transplantation. In addition, the BAFF-R expression (MFI) on B cells of RTX-treated patients was lower up to 24 months after transplantation (Figure 4D).


Longitudinal analysis of T and B cell phenotype and function in renal transplant recipients with or without rituximab induction therapy.

Kamburova EG, Koenen HJ, van den Hoogen MW, Baas MC, Joosten I, Hilbrands LB - PLoS ONE (2014)

Longitudinal analysis of circulating B cells in renal transplant recipients after treatment with tacrolimus, MMF and steroids, and a single dose of rituximab (RTX) during transplant surgery.(A) Representative dot plots of CD19+ B cells within the CD45+ lymphocyte population for a RTX-treated and a triple immunosuppression (IS)-treated patient before transplantation (pre-Tx) and at 3, 12, and 24 months after transplantation. (B) Shown are the absolute numbers of CD19+ B cells for RTX-(gray, n = 12) and triple IS-treated (white, n = 14) patients before transplantation (t = 0) and up to 24 months after transplantation (n = 10 and n = 9 at t = 24 m, respectively). (C) Pie charts depicting the distribution the different B cells subsets over time using the Bm1-Bm5 classification as depicted in Figure 3B: Bm1 (IgD+CD38−), Bm2 (IgD+CD38+), Bm2’ (IgD+CD38++), Bm3+4 (IgD−CD38++), Early Bm5 (IgD−CD38+) and Late Bm5 (IgD−CD38−) cells within the CD19+ B-cell population. Data are represented as means of 14 triple IS+RTX-treated and 12 IS-treated patients before transplantation (pre-Tx) and at 3, 12, and 24 months after transplantation (n = 10 and n = 9 at t = 24 m, respectively). (D) Shown are the percentages of CD80+, CD95+ and BAFF-R+ cells within the CD19+ B-cell population for RTX- (gray, n = 12) and triple IS-treated (white, n = 14) patients before transplantation (t = 0) and up to 24 months after transplantation. Results are shown as box plots displaying the median, 25th and 75th percentiles as the box, and the 5th and 95th percentiles as whiskers. Significant differences are indicated by asterisks: **P<0.01.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4231065&req=5

pone-0112658-g004: Longitudinal analysis of circulating B cells in renal transplant recipients after treatment with tacrolimus, MMF and steroids, and a single dose of rituximab (RTX) during transplant surgery.(A) Representative dot plots of CD19+ B cells within the CD45+ lymphocyte population for a RTX-treated and a triple immunosuppression (IS)-treated patient before transplantation (pre-Tx) and at 3, 12, and 24 months after transplantation. (B) Shown are the absolute numbers of CD19+ B cells for RTX-(gray, n = 12) and triple IS-treated (white, n = 14) patients before transplantation (t = 0) and up to 24 months after transplantation (n = 10 and n = 9 at t = 24 m, respectively). (C) Pie charts depicting the distribution the different B cells subsets over time using the Bm1-Bm5 classification as depicted in Figure 3B: Bm1 (IgD+CD38−), Bm2 (IgD+CD38+), Bm2’ (IgD+CD38++), Bm3+4 (IgD−CD38++), Early Bm5 (IgD−CD38+) and Late Bm5 (IgD−CD38−) cells within the CD19+ B-cell population. Data are represented as means of 14 triple IS+RTX-treated and 12 IS-treated patients before transplantation (pre-Tx) and at 3, 12, and 24 months after transplantation (n = 10 and n = 9 at t = 24 m, respectively). (D) Shown are the percentages of CD80+, CD95+ and BAFF-R+ cells within the CD19+ B-cell population for RTX- (gray, n = 12) and triple IS-treated (white, n = 14) patients before transplantation (t = 0) and up to 24 months after transplantation. Results are shown as box plots displaying the median, 25th and 75th percentiles as the box, and the 5th and 95th percentiles as whiskers. Significant differences are indicated by asterisks: **P<0.01.
Mentions: The participation of renal transplant recipients in a randomized double-blind placebo-controlled study evaluating the efficacy and safety of RTX when added to triple drug immunosuppression gave us the opportunity to study the effects of additional B-cell depletion on the phenotype and function of T and B cells. RTX treatment resulted in a nearly complete depletion of B cells from the peripheral lymphocyte population up to 12 months after transplantation. Remarkably, the B-cell depletion after a single dose of RTX was long lasting. The absolute numbers of B cells remained quite low at 24 months after RTX treatment with a median of 9.7 B cells/µl (range 3.0–294.8) compared to a median of 116.4 B cells/µl (range 49.7–379.7) in patients not treated with RTX (Figure 4A–B). Interestingly, at 12 months the percentage of Bm2’ (IgD++CD38++) and Bm3+4 (IgD−CD38++) cells was significantly higher in RTX-treated patients, while the percentages of Bm1 (IgD+CD38−) and Bm2 (IgD+CD38+) cells were lower in RTX-treated patients. (Figure 4C). Accordingly, the percentages of CD24++CD38++ transitional and IgD−CD27+ switched memory B cells were higher in RTX-treated patients, while the percentages of IgD+CD27− naïve B cells were lower in RTX-treated patients (Figure S3A). Fitting with the relative increase of Bm3+4 (IgD−CD38++) cells, there was an increase in the percentage of CD80+ and CD95+ B cells at 12 and/or 24 months after transplantation (Figure 4D). Remarkably, the percentage of BAFF-R+ B cells was lower in RTX-treated patients at 3 and 12 months after transplantation. In addition, the BAFF-R expression (MFI) on B cells of RTX-treated patients was lower up to 24 months after transplantation (Figure 4D).

Bottom Line: In addition, the ratio between the percentage of central memory CD4+ and CD4+ regulatory T cells was significantly higher up to 24 months post-transplant compared to pre-transplant levels.Addition of a single dose of rituximab resulted in a long-lasting B-cell depletion.Rituximab treatment had no effect on the T-cell phenotype and function post-transplant.

View Article: PubMed Central - PubMed

Affiliation: Department of Laboratory Medicine - Medical Immunology, Radboud University Medical Centre, Nijmegen, The Netherlands.

ABSTRACT

Background: Prevention of rejection after renal transplantation requires treatment with immunosuppressive drugs. Data on their in vivo effects on T- and B-cell phenotype and function are limited.

Methods: In a randomized double-blind placebo-controlled study to prevent renal allograft rejection, patients were treated with tacrolimus, mycophenolate mofetil (MMF), steroids, and a single dose of rituximab or placebo during transplant surgery. In a subset of patients, we analyzed the number and phenotype of peripheral T and B cells by multiparameter flow cytometry before transplantation, and at 3, 6, 12, and 24 months after transplantation.

Results: In patients treated with tacrolimus/MMF/steroids the proportion of central memory CD4+ and CD8+ T cells was higher at 3 months post-transplant compared to pre-transplant levels. In addition, the ratio between the percentage of central memory CD4+ and CD4+ regulatory T cells was significantly higher up to 24 months post-transplant compared to pre-transplant levels. Interestingly, treatment with tacrolimus/MMF/steroids resulted in a shift toward a more memory-like B-cell phenotype post-transplant. Addition of a single dose of rituximab resulted in a long-lasting B-cell depletion. At 12 months post-transplant, the small fraction of repopulated B cells consisted of a high percentage of transitional B cells. Rituximab treatment had no effect on the T-cell phenotype and function post-transplant.

Conclusions: Renal transplant recipients treated with tacrolimus/MMF/steroids show an altered memory T and B-cell compartment post-transplant. Additional B-cell depletion by rituximab leads to a relative increase of transitional and memory-like B cells, without affecting T-cell phenotype and function.

Trial registration: ClinicalTrials.gov NCT00565331.

Show MeSH
Related in: MedlinePlus