Impact of Labor on Peripheral Blood Maternal T-Cell Subsets and on Regulatory T and B Cells

2016 ◽  
Vol 24 (2) ◽  
pp. 276-284 ◽  
Author(s):  
Jorge Lima ◽  
Catarina Martins ◽  
Glória Nunes ◽  
Maria-José Sousa ◽  
Jorge C. Branco ◽  
...  
Keyword(s):  
T Cell ◽  
B Cells ◽  
Peripheral Blood ◽  
T Cell Subsets ◽  
T And B Cells

scholarly journals Simultaneous analysis of antigen-specific B and T cells after SARS-CoV-2 infection and vaccination

2021 ◽  
Author(s):  
Krista L Newell ◽  
Mitchell J Waldran ◽  
Stephen J Thomas ◽  
Timothy P Endy ◽  
Adam Tully Waickman
Keyword(s):  
Flow Cytometry ◽  
T Cell ◽  
B Cells ◽  
Simultaneous Detection ◽  
T Cell Subsets ◽  
Cell Populations ◽  
T And B Cells ◽  
Simultaneous Identification ◽  
Immune Profiling ◽  
Insight Into

Conventional methods for quantifying and phenotyping antigen-specific lymphocytes can rapidly deplete irreplaceable specimens. This is due to the fact that antigen-specific T and B cells have historically been analyzed in independent assays each requiring millions of cells. A technique that facilitates the simultaneous detection of antigen-specific T and B cells would allow for more thorough immune profiling with significantly reduced sample requirements. To this end, we developed the B And T cell Tandem Lymphocyte Evaluation (BATTLE) assay, which allows for the simultaneous identification of SARS-CoV-2 Spike reactive T and B cells using an optimized Activation Induced Marker (AIM) T cell assay and dual-color B cell antigen probes. Using this assay, we demonstrate that antigen-specific B and T cell subsets can be identified simultaneously using conventional flow cytometry platforms and provide insight into the differential effects of mRNA vaccination on B and T cell populations following natural SARS-CoV-2 infection.

Improved T Cell Recovery After Transplantation of CD3/CD19 depleted Haploidentical Stem Cell Grafts in Pediatric Patients.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4652-4652
Author(s):  
Heiko-Manuel Teltschik ◽  
Tobias Feuchtinger ◽  
Matthias Pfeiffer ◽  
Michael Schumm ◽  
Ingo Mueller ◽  
...  
Keyword(s):  
T Cell ◽  
B Cells ◽  
Conflicts Of Interest ◽  
Negative Impact ◽  
Conditioning Regimen ◽  
T Cell Subsets ◽  
Control Group ◽  
T And B Cells ◽  
Haploidentical Transplantation ◽  
Significant Difference

Abstract Abstract 4652 Profound depletion of T and B cells is a fundamental prerequisite for haploidentical transplantation and allows to minimize GvHD despite HLA incompatibility. However, posttransplant recovery of donor derived T cells is delayed after various graft manipulation procedures and may result in severe infections. Methods to improve this recovery are of great importance. Here we present immune reconstitution data in patients who received CD3/CD19 depleted stem cells in combination with melphalan based or standard conditioning regimens. 32 patients with ALL (n=14), AML/MDS (n=17), CML (n=1) were included. T and B cells were directly depleted using antiCD3/antiCD19 coated magnetic microbeads and the CliniMACSTM device. The patients received either TBI or Bu i.v. and OKT3 (n=9) or a reduced intensity conditioning (“RIC”: Mel 140mg/m2, Flud 160mg/m2, TT 10mg/kg, OKT3, n= 23). Absolute numbers of lymphocyte subsets per microliter on day 90 were compared within these both groups and with a historical control group (patients with leukemias who received CD34 selected grafts and TBI or Bu based standard conditioning regimen in combination with ATG, n=28). CD3+4+, CD3+8+ and total numbers of CD3+ of patients after CD3/CD19 depletion were significantly higher in the RIC-group than in the TBI/Bu-group (mean numbers: 85.83 vs. 38.84; 133.46 vs. 19.69; 270.27 vs. 63.99; p<0.05, unpaired t-test). Comparison with the whole CD34 historical group showed a faster recovery of CD3+4+ in patients with CD3/19 depletion and RIC (104.26 vs. 54.22; p= 0,034) but no significant difference in CD3+8+ and CD3+. Furthermore, subgroups of the CD34 historical population were compared: patients with CD3/19 depletion and RIC had a significantly faster recovery of CD3+4+, CD3+8+ and CD3+ than CD34 patients with TBI (104.26vs. 25.48; 133.46vs. 43.17; 270.27 vs. 65.86; p<0.05) but had no advantage over CD34 patients with non-TBI conditioning. Conclusions: the type of graft manipulation appeared to have an influence on the speed of CD4+ recovery (CD3/19 depletion > CD34 selection). Moreover, the use of TBI had a clear negative impact on all T cell subsets: patients with TBI had a slower recovery than patients with non-TBI conditioning, independent from graft manipulation procedures and probably due to thymic damage. Thus, the use of RIC-protocols in combination with CD3/CD19 depletion may help to speed up the immune recovery after haploidentical transplantation. Further studies are warranted to evaluate the risk of relapse. Disclosures: No relevant conflicts of interest to declare.

Massive, Clinical Grade Expansion Of Polyclonal T Cells Using Blinatumomab For Adoptive Autologous Cellular Therapy Of CLL Patients

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3272-3272 ◽  
Author(s):  
Josée Golay ◽  
Anna D’amico ◽  
Gianmaria Borleri ◽  
Maria Chiara Finazzi ◽  
Giulia Quaresmini ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Peripheral Blood ◽  
T Cell Subsets ◽  
Th2 Cells ◽  
Adoptive Therapy ◽  
Specific Patient

Abstract Background The combined use of chemotherapy and monoclonal antibodies has proved highly effective for the treatment of CLL but often results in severe life threatening immunosuppression. The development of adoptive therapy with autologous T cells could be clinically relevant to overcome these problems. Methods We have devised a novel, simple and efficient method for ex vivo expansion of normal autologous T cells from the peripheral blood of CLL patients for adoptive therapy, using blinatumomab (CD3xCD19) and rhIL-2 in serum-free medium. The complete phenotype of in vitro expanded T cells was analyzed by flow cytometry and their cytotoxic activity by calcein release assays. Results We performed 18 expansions of T cells, starting from a very small volume of peripheral blood from untreated CLL patients (mean 10.3 ml, range 2-30 ml) that contained a mean of 9.2x106 T cells (range 0.4 to 51x106)(Fig.1). This method allowed reproducible expansion in about 21 days of a mean 410x106 CD3+ T cells (range 71 to 2184x106). The mean fold expansion of T cells in about 3 weeks of in vitro culture was 224 (range 4.4-1326). The only significant contaminant in final Blinatumomab Expanded T cell cultures (BET) were NK cells (mean 18.5%). Indeed addition of blinatumomab and rhIL-2 to the cultures led to a rapid decrease in CLL B cells, which took place from days 7 to 14 onwards and resulted in their complete depletion within 3 weeks (mean 0.2% CLL B cells at days 18-25). Only in one case, a significant percentage of CLL B cells could be observed at the end of culture, but this was due to the particularly high percentage neoplastic cells in the starting population in this patient (98%), resulting in relatively late depletion of these cells, which took place between days 14 and 21, and therefore remained detectable in this case at day 24 (3.8% CLL B cells at day 24). Despite the very low percentage of starting T cells in this specific patient (1.2%), 152x106 T cells could be obtained, equivalent to a 42 fold expansion. In the 18 expansions performed, the resulting BET cells contained both CD4+ and CD8+ cells in varying proportions (median 46.2% and 44.4% respectively). Only in two cases the final product was composed predominantly of CD4+ cells (95%). Expanded T cells were polyclonal, as shown by TCR Vβ expression which was within the normal range by flow cytometry. Indeed CMV specific clones, detected by CMV peptide (pp65495-503)-loaded HLA-A*0201 tetramer, were expanded using this method and detected in equivalent proportion before and after expansion. Final T cells were composed predominantly of the effector and central memory subsets. Th1 were slightly prevalent over Th2 cells (means 20% and 10%, respectively), whereas Th17 and Treg were less than 1%. Since CLL derived T cells have been shown previously to have enhanced expression of the synapse regulators CD272 and CD279 compared to normal T cells, leading to impaired immunological synapse formation, we have analyzed these markers in both starting and BET cells from 4 patients. We observed that CD272 and CD279 diminished in BET compared to the starting CLL T populations (from 73% to 19% and 61% to 18%, respectively). These data suggest that stimulation and expansion with blinatumomab and rhIL-2 has normalized expression of these regulators on CLL T cells. Indeed BET were highly cytotoxic against CD19+ targets cell lines or primary CLL cells, with 70-90% lysis at a 3:1 effector target ratio in presence of blinatumomab. Finally BET were compared to Xcellerated cells expanded using anti-CD3/CD28 Dynabeads and rhIL-2. The expansion protocols using either blinatumomab or anti-CD3/CD28 Dynabeads showed equivalent efficiency and comparable cell composition at the end of culture. Further comparison of the T cell subsets present in BET or CD3/CD28 cultures is in progress. Conclusions These data altogether suggest that the use of blinatumomab and rhIL-2 provides a reproducible, simple and GMP-compliant protocol, allowing expansion of large numbers of autologous polyclonal T cells depleted of CLL cells, from relatively small volumes of peripheral blood from CLL patients. This approach is an attractive option for adoptive therapy in these patients after immunosuppressive treatments. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Modifications in flow cytometric estimation of t cell subsets and b cells in peripheral blood to reduce the cost of investigation

2002 ◽  
Vol 17 (1) ◽  
pp. 69-74 ◽  
Author(s):  
Sanju Jalla ◽  
Sunil Sazawal ◽  
Saikat Deb ◽  
Robert E. Black ◽  
Maharaj K. Bhan
Keyword(s):  
T Cell ◽  
B Cells ◽  
Peripheral Blood ◽  
T Cell Subsets ◽  
Flow Cytometric ◽  
The Cost

scholarly journals Comparison of DNA repair and radiosensitivity of different blood cell populations

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Daniel Heylmann ◽  
Viviane Ponath ◽  
Thomas Kindler ◽  
Bernd Kaina
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Blood Cell ◽  
Peripheral Blood ◽  
Cell Populations ◽  
T And B Cells ◽  
Myeloid Lineage ◽  
Γh2ax Foci

AbstractDespite the frequent use of ionising radiation (IR) in therapy and diagnostics and the unavoidable exposure to external radiation sources, our knowledge regarding the radiosensitivity of human blood cell populations is limited and published data, obtained under different experimental conditions, are heterogeneous. To compare the radiosensitivity of different hematopoietic cell populations, we set out to determine the responses of cells obtained from peripheral blood of healthy volunteers under identical conditions (resting, non-stimulated cells). First, we measured the radiation response of T cells (Treg, Th, CTL), B cells, NK cells, CD34+ progenitor cells and monocytes obtained from peripheral blood and monocyte-derived macrophages (Mph) and immature dendritic cells (iDC) ex vivo and show that T and B cells are highly sensitive, starting to undergo apoptosis following IR with a dose as low as 0.125 Gy. Importantly, there was no clear threshold dose and cell death/apoptosis increased up to a saturation level with a dose of 2 Gy. The sensitivity decreased in the order of T cells > NK and B cells > monocytes > macrophages and iDC. The data confirm a previous report that Mph and iDC are radiation-resistant compared to their progenitor monocytes. Although non-stimulated T and B cells were highly radiation-sensitive compared to monocytes and macrophages, they were competent in the repair of DNA double-strand breaks, as shown by a decline in γH2AX foci in the post-exposure period. CD34+ cells obtained from peripheral blood also showed γH2AX decline post-exposure, indicating they are repair competent. Granulocytes (CD15+) did not display any γH2AX staining following IR. Although peripheral blood lymphocytes, the main fraction are T cells, were significantly more radiation-sensitive than monocytes, they displayed the expression of the repair proteins XRCC1, ligase III and PARP-1, which were nearly non-expressed in monocytes. To assess whether monocytes are depleted in vivo following IR, we measured the amount of T cells and monocytes in cancer patients who received total-body radiation (TBR, 6 × 2 Gy). We observed that the number of T cells in the peripheral blood significantly declined already after the first day of TBR and remained at a low level, which was accompanied by an increase in the number of γH2AX foci in the surviving CD3+ T cell fraction. In contrast, the number of monocytes did not decline extensively, reflecting their radiation resistance compared to T cells. Monocytes also showed an accumulation of γH2AX foci in vivo, but the levels were significantly lower than in T cells. CD56+ NK cells displayed a response similar to T cells. The data support the notion that unstimulated T cell subfractions are nearly equally radiation sensitive. There are, however, remarkable differences in the radiation sensitivity between the lymphoid and the myeloid lineage, with lymphoid cells being significantly more sensitive than cells of the myeloid lineage. In the myeloid lineage, macrophages and iDCs were the most radio-resistant cell types.

HLA-E restricted CD8+ T cell subsets are phenotypically altered in multiple sclerosis patients

2013 ◽  
Vol 20 (7) ◽  
pp. 790-801 ◽  
Author(s):  
Kim Pannemans ◽  
Bieke Broux ◽  
An Goris ◽  
Bénédicte Dubois ◽  
Tom Broekmans ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
T Cells ◽  
T Cell ◽  
B Cells ◽  
T Cell Subsets ◽  
Regulatory Role ◽  
T And B Cells ◽  
Autoreactive T Cell ◽  
Multiple Sclerosis Patients

Background: The importance of Qa-1 restricted CD8+ T cells in regulating autoreactive T cell responses has been demonstrated in animal models for autoimmune disorders, including multiple sclerosis (MS). Objective: We hypothesize that their human variant, HLA-E restricted CD8+ T cells, fulfills a similar regulatory role in man and that these cells are of importance in MS. Methods: A large cohort of MS patients and healthy controls was genotyped for the two known HLA-E polymorphisms. Flow cytometry was used to determine HLA-E expression kinetics and to phenotype HLA-E restricted CD8+ T cells. Immunohistochemistry was performed to investigate HLA-E expression in the central nervous system (CNS) of MS patients. Results: HLA-E is upregulated on immune cells upon in vitro activation and this upregulation is polymorphism-dependent for T and B cells. T and B cells in lesions of MS patients show enhanced HLA-E expression. Furthermore, NKG2C+CD8+ T cells of MS patients have a significantly lower Foxp3 expression, while NKG2A+CD8+ T cells of MS patients produce higher levels of pro-inflammatory cytokines compared to those of healthy individuals. Conclusion: Our study indicates that the HLA-E system is altered in MS and could play a regulatory role in disease.

scholarly journals Detecting specific memory t and b cells in volunteers annually revaccinated with live anthrax vaccine

2019 ◽  
Vol 9 (3-4) ◽  
pp. 495-503
Author(s):  
V. V. Firstova ◽  
A. S. Kartseva ◽  
M. V. Silkina ◽  
M. A. Marin ◽  
Ia. O. Muntian ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Peripheral Blood ◽  
Early Stage ◽  
T Cell Subsets ◽  
Cytotoxic T Cell ◽  
Igg Antibodies ◽  
Specific Memory ◽  
Hla Dr

Currently, live anthrax vaccine has been used for vaccine prophylaxis in Russia and neighbor countries for seve ral decades, but precise mechanism of post-vaccination protection mechanism remains unclear. Here, we provide data on examining serum antibody level against protective antigen (PA) and lethal factor (LF) in repeatedly vaccinated volun teers at early stage (5–8 days) and 1 month after the performing pre-scheduled annual revaccination. Amount of peripheral blood antigen-specific memory T cells after previous vaccinations was analyzed. It was showed that frequency of CD3+CD45RO+CD62L– memory effector T cells was increased in the majority of volunteers on day 5-8 day after performing pre-scheduled annual revaccination that peaked at day 7 by elevating it by 2-fold compared with the control group. Percentage of anthrax-specific central memory T cells did not increase at early stage after vaccination, whereas amount of activated CD3+CD45RO+CD62L+HLA-DR+ subset within this memory T cell population was increased. Likewise, percentage of activated CD3+CD45RO+CD62L–HLA-DR+ effector memory T cell subset was also increased. Moreover, serum anti-PA IgG were detected on day 5–8 day after pre-scheduled annual revaccination in half of volunteers, whereas anti-LF IgG were found only in a single volunteer. Rapidly elevated amount of serum anthrax-specific IgG antibodies evidences about sustained memory B cell response in peripheral blood samples in volunteers after pre-scheduled annual revaccination. However, percentage of CD19+CD27+ memory B cells was not significantly elevated at early stage after revaccination that tended to increase. Both helper and cytotoxic T cell subsets were activated on day 5–8 after revaccination revealed by upregulated expression of CD69 and/or CD25 markers, with the latter predominantly found on helper T cells, thereby accounting for their high proliferative activity, whereas the former — on cytotoxic T cell subsets. Detection of anti-PA IgG antibodies correlates with protection against anthrax, which was confirmed in animal models. Unfortunately, the level of serum anti-PA IgG antibodies rapidly declines after vaccination. Ability of memory B cells to rapidly trigger production of anthrax-specific antibodies in response to revaccination suggests that anti-anthrax immunity may be evaluated by measuring frequency of peripheral blood anthrax-specific memory B and T cells.

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