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Equine allogeneic bone marrow-derived mesenchymal stromal cells elicit antibody responses in vivo.

Pezzanite LM, Fortier LA, Antczak DF, Cassano JM, Brosnahan MM, Miller D, Schnabel LV - Stem Cell Res Ther (2015)

Bottom Line: Donor MSCs, which were previously isolated and immunophenotyped, were thawed and culture expanded to achieve between 30x10(6) and 50x10(6) cells for intradermal injection into the recipient's neck.Anti-ELA-A2 antibody responses were varied both in terms of strength and timing.Allogeneic MSCs are capable of eliciting antibody responses in vivo that can be strong and also cross-reactive with MHC types other than that of the donor.

View Article: PubMed Central - PubMed

Affiliation: Department of Clinical Sciences, College of Veterinary Medicine, Cornell University, Ithaca, NY, 14853, USA. lmp238@cornell.edu.

ABSTRACT

Introduction: This study tested the hypothesis that Major Histocompatibility Complex (MHC) incompatible equine mesenchymal stromal cells (MSCs) would induce cytotoxic antibodies to donor MHC antigens in recipient horses after intradermal injection. No studies to date have explored recipient antibody responses to allogeneic donor MSC transplantation in the horse. This information is critical because the horse is a valuable species for assessing the safety and efficacy of MSC treatment prior to human clinical application.

Methods: Six MHC heterozygote horses were identified as non-ELA-A2 haplotype by microsatellite typing and used as allogeneic MHC-mismatched MSC recipients. MHC homozygote horses of known ELA-A2 haplotype were used as MSC and peripheral blood leukocyte (PBL) donors. One MHC homozygote horse of the ELA-A2 haplotype was the recipient of ELA-A2 donor MSCs as an MHC-matched control. Donor MSCs, which were previously isolated and immunophenotyped, were thawed and culture expanded to achieve between 30x10(6) and 50x10(6) cells for intradermal injection into the recipient's neck. Recipient serum was collected and tested for the presence of anti-donor antibodies prior to MSC injection and every 7 days after MSC injection for the duration of the 8-week study using the standard two-stage lymphocyte microcytotoxicity dye-exclusion test. In addition to anti-ELA-A2 antibodies, recipient serum was examined for the presence of cross-reactive antibodies including anti-ELA-A3 and anti-RBC antibodies.

Results: All MHC-mismatched recipient horses produced anti-ELA-A2 antibodies following injection of ELA-A2 MSCs and developed a wheal at the injection site that persisted for the duration of the experiment. Anti-ELA-A2 antibody responses were varied both in terms of strength and timing. Four recipient horses had high-titered anti-ELA-A2 antibody responses resulting in greater than 80% donor PBL death in the microcytotoxicity assays and one of these horses also developed antibodies that cross-reacted when tested on lymphocyte targets from a horse with an unrelated MHC type.

Conclusions: Allogeneic MSCs are capable of eliciting antibody responses in vivo that can be strong and also cross-reactive with MHC types other than that of the donor. Such responses could limit the effectiveness of repeated allogeneic MSC use in a single horse, and could also result in untoward inflammatory responses in recipients.

No MeSH data available.


Related in: MedlinePlus

Cytotoxic antibody responses to allogeneic (major histocompatibility complex class I and class II mismatched) donor ELA-A2 mesenchymal stromal/stem cells by recipient horses A to F. The cytotoxic antibody titer was determined by the highest dilution (N) of each recipient serum sample that resulted in killing of at least 80% of donor peripheral blood leukocytes (PBLs). Ab, antibodies detected by the presence of donor PBL death, but with donor PBL death below the cutoff value of 80%; No Ab, no antibodies detected (no donor PBL death). As shown, four recipient horses (Horses B, C, E, and F) had strong anti-ELA-A2 antibody responses which persisted for the duration of the experiment, while two recipient horses (Horses A and D) had weak responses. ∨Both weak responders A and D received a second injection of donor mesenchymal stromal/stem cells at 5 weeks (35 days). ELA-A2, equine leukocyte antigen haplotype.
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Fig2: Cytotoxic antibody responses to allogeneic (major histocompatibility complex class I and class II mismatched) donor ELA-A2 mesenchymal stromal/stem cells by recipient horses A to F. The cytotoxic antibody titer was determined by the highest dilution (N) of each recipient serum sample that resulted in killing of at least 80% of donor peripheral blood leukocytes (PBLs). Ab, antibodies detected by the presence of donor PBL death, but with donor PBL death below the cutoff value of 80%; No Ab, no antibodies detected (no donor PBL death). As shown, four recipient horses (Horses B, C, E, and F) had strong anti-ELA-A2 antibody responses which persisted for the duration of the experiment, while two recipient horses (Horses A and D) had weak responses. ∨Both weak responders A and D received a second injection of donor mesenchymal stromal/stem cells at 5 weeks (35 days). ELA-A2, equine leukocyte antigen haplotype.

Mentions: Data reported as the percentage death of ELA-A2 donor leukocytes. For each time point, the value in italics is the cytotoxic antibody titer (the highest dilution of each antisera that resulted in killing of at least 80% of the donor leukocytes as reported in Figure 2). ELA-A2, equine leukocyte antigen haplotype.


Equine allogeneic bone marrow-derived mesenchymal stromal cells elicit antibody responses in vivo.

Pezzanite LM, Fortier LA, Antczak DF, Cassano JM, Brosnahan MM, Miller D, Schnabel LV - Stem Cell Res Ther (2015)

Cytotoxic antibody responses to allogeneic (major histocompatibility complex class I and class II mismatched) donor ELA-A2 mesenchymal stromal/stem cells by recipient horses A to F. The cytotoxic antibody titer was determined by the highest dilution (N) of each recipient serum sample that resulted in killing of at least 80% of donor peripheral blood leukocytes (PBLs). Ab, antibodies detected by the presence of donor PBL death, but with donor PBL death below the cutoff value of 80%; No Ab, no antibodies detected (no donor PBL death). As shown, four recipient horses (Horses B, C, E, and F) had strong anti-ELA-A2 antibody responses which persisted for the duration of the experiment, while two recipient horses (Horses A and D) had weak responses. ∨Both weak responders A and D received a second injection of donor mesenchymal stromal/stem cells at 5 weeks (35 days). ELA-A2, equine leukocyte antigen haplotype.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4414005&req=5

Fig2: Cytotoxic antibody responses to allogeneic (major histocompatibility complex class I and class II mismatched) donor ELA-A2 mesenchymal stromal/stem cells by recipient horses A to F. The cytotoxic antibody titer was determined by the highest dilution (N) of each recipient serum sample that resulted in killing of at least 80% of donor peripheral blood leukocytes (PBLs). Ab, antibodies detected by the presence of donor PBL death, but with donor PBL death below the cutoff value of 80%; No Ab, no antibodies detected (no donor PBL death). As shown, four recipient horses (Horses B, C, E, and F) had strong anti-ELA-A2 antibody responses which persisted for the duration of the experiment, while two recipient horses (Horses A and D) had weak responses. ∨Both weak responders A and D received a second injection of donor mesenchymal stromal/stem cells at 5 weeks (35 days). ELA-A2, equine leukocyte antigen haplotype.
Mentions: Data reported as the percentage death of ELA-A2 donor leukocytes. For each time point, the value in italics is the cytotoxic antibody titer (the highest dilution of each antisera that resulted in killing of at least 80% of the donor leukocytes as reported in Figure 2). ELA-A2, equine leukocyte antigen haplotype.

Bottom Line: Donor MSCs, which were previously isolated and immunophenotyped, were thawed and culture expanded to achieve between 30x10(6) and 50x10(6) cells for intradermal injection into the recipient's neck.Anti-ELA-A2 antibody responses were varied both in terms of strength and timing.Allogeneic MSCs are capable of eliciting antibody responses in vivo that can be strong and also cross-reactive with MHC types other than that of the donor.

View Article: PubMed Central - PubMed

Affiliation: Department of Clinical Sciences, College of Veterinary Medicine, Cornell University, Ithaca, NY, 14853, USA. lmp238@cornell.edu.

ABSTRACT

Introduction: This study tested the hypothesis that Major Histocompatibility Complex (MHC) incompatible equine mesenchymal stromal cells (MSCs) would induce cytotoxic antibodies to donor MHC antigens in recipient horses after intradermal injection. No studies to date have explored recipient antibody responses to allogeneic donor MSC transplantation in the horse. This information is critical because the horse is a valuable species for assessing the safety and efficacy of MSC treatment prior to human clinical application.

Methods: Six MHC heterozygote horses were identified as non-ELA-A2 haplotype by microsatellite typing and used as allogeneic MHC-mismatched MSC recipients. MHC homozygote horses of known ELA-A2 haplotype were used as MSC and peripheral blood leukocyte (PBL) donors. One MHC homozygote horse of the ELA-A2 haplotype was the recipient of ELA-A2 donor MSCs as an MHC-matched control. Donor MSCs, which were previously isolated and immunophenotyped, were thawed and culture expanded to achieve between 30x10(6) and 50x10(6) cells for intradermal injection into the recipient's neck. Recipient serum was collected and tested for the presence of anti-donor antibodies prior to MSC injection and every 7 days after MSC injection for the duration of the 8-week study using the standard two-stage lymphocyte microcytotoxicity dye-exclusion test. In addition to anti-ELA-A2 antibodies, recipient serum was examined for the presence of cross-reactive antibodies including anti-ELA-A3 and anti-RBC antibodies.

Results: All MHC-mismatched recipient horses produced anti-ELA-A2 antibodies following injection of ELA-A2 MSCs and developed a wheal at the injection site that persisted for the duration of the experiment. Anti-ELA-A2 antibody responses were varied both in terms of strength and timing. Four recipient horses had high-titered anti-ELA-A2 antibody responses resulting in greater than 80% donor PBL death in the microcytotoxicity assays and one of these horses also developed antibodies that cross-reacted when tested on lymphocyte targets from a horse with an unrelated MHC type.

Conclusions: Allogeneic MSCs are capable of eliciting antibody responses in vivo that can be strong and also cross-reactive with MHC types other than that of the donor. Such responses could limit the effectiveness of repeated allogeneic MSC use in a single horse, and could also result in untoward inflammatory responses in recipients.

No MeSH data available.


Related in: MedlinePlus