Macrophage effector responses of horses are influenced by expression of CD154

Sponseller, Brett A.; Clark, Sandra K.; Gilbertie, Jessica M.; Wong, D.; Hepworth, Kate; Wiechert, Sarah; Chandramani, Prashanth; Sponseller, Beatrice T.; Alcott, Cody J.; Bellaire, Bryan H.; Petersen, Andrew C.; Jones, Douglas E.

doi:10.1016/j.vetimm.2016.08.001

Cited by 2 publications

(5 citation statements)

References 26 publications

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“…According to the consistent detection of bands of 38 and 18 kDa matching the predicted MW of membrane-bound and cleaved (soluble) equine CD154, respectively ( 20 , 22 ), the anti-human CD154 mAb 5C8 was considered specifically cross-reactive with equine CD154 and used in the following flow cytometry experiments.…”

Section: Resultsmentioning

confidence: 99%

“…In mice and man, CD154 has been established as a marker of T helper cell (Th) activation by antigens ( 8 , 16 , 17 ). CD154 is a member of the type II transmembrane protein of the TNF superfamily ( 17 – 19 ) with a predicted MW of 28.7 kDa in horses [GenBank accession: AQM73588.1 ( 20 )]. In humans and mice, the predicted MW of CD154 is 29 kDa, but the actual MW of CD154 is usually 33 or 39 kDa due to post-translational modifications of the protein ( 17 , 19 , 21 ).…”

Section: Introductionmentioning

confidence: 99%

“…Recombinant equine CD154 was used as an experimental vaccine adjuvant of a West Nile Virus subunit vaccine for horses in vivo and improved the induction of neutralizing antibodies compared to the subunit vaccine without CD154 ( 24 ). Furthermore, CHO cell expressed recombinant ( r ) equine CD154 stimulated equine monocyte-derived macrophages to produce reactive oxygen species in vitro ( 20 ). In this study, an anti-human CD154 antibody was used for the confirmation of r equine CD154 protein expression ( 20 ).…”

Section: Introductionmentioning

confidence: 99%

“…Furthermore, CHO cell expressed recombinant ( r ) equine CD154 stimulated equine monocyte-derived macrophages to produce reactive oxygen species in vitro ( 20 ). In this study, an anti-human CD154 antibody was used for the confirmation of r equine CD154 protein expression ( 20 ). In humans and pigs CD154 has been used as an indicator of antigen-specific T cell activation and for the enrichment of reactive T cells ( 8 , 15 , 16 , 25 , 26 ), detected with another anti-human CD154 monoclonal antibody (mAb), clone 5C8 ( 27 ).…”

Section: Introductionmentioning

confidence: 99%

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CD154 Expression Indicates T Cell Activation Following Tetanus Toxoid Vaccination of Horses

et al. 2022

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Despite the relevance of adaptive immunity against equine pathogens antigen-specific T cell responses of horses are not well characterized and the lack of insight into T cell responses hampers the understanding of the pathogeneses of important diseases. In this study we used tetanus toxoid (TT) as a well-defined antigen to characterize antigen-reactive T cells. Six healthy adult horses received a routine booster against tetanus with an immune stimulating complex (ISCOM)-based vaccine and were followed for 28 days. TT-specific serum antibodies were quantified by ELISA and increased in all horses by day 7 after vaccination. CD154 is an established indicator of antigen-reactive T helper cells in other species, but has not been characterized in horses. CD154 detection in equine PBMC by an anti-human CD154 antibody (clone 5C8) was confirmed by Western blots and then applied for flow cytometry. As a common indicator of equine T cell activation, cytokine induction was studied in parallel. T cells were analyzed by multicolor flow cytometry of PBMC after re-stimulation with TT in vitro. Reactive T helper (Th) cells were characterized by increased frequencies of CD4+CD154+ lymphocytes in in vitro TT-re-stimulated PBMC on day 14 after vaccination of the horses compared to pre-vaccination. The majority of all CD154+ cells after TT re-stimulation were CD4+ Th cells, but CD154 was also induced on CD4- cells albeit in lower frequencies. CD154+CD4+ Th cells were enriched in cytokine-expressing cells compared to CD154-CD4+ Th cells. Similar to the CD4+CD154+ frequencies, CD4+IL-4+, CD4+IFN-γ+ and CD4+TNF-α+ were increased after vaccination, but IL-4+ increased later than IFN-γ+ and CD4+TNF-α+, which already exceeded pre-vaccination frequencies on day 7. CD4+CD154+ frequencies correlated positively with those of CD4+IL-4+ (Th2) on day 14, and negatively with CD4+IFN-γ+ induction on day 7, but did not correlate with CD4+TNF-α+ frequencies or TT-specific antibody concentrations. CD154 appears to be a useful marker of antigen-reactive equine Th cells in combination with cytokine expression. The T cell analyses established here with TT can be applied to other antigens relevant for infections or allergies of horses and in horse models for translational research.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

CD154 Expression Indicates T Cell Activation Following Tetanus Toxoid Vaccination of Horses

et al. 2022

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“…However, Kergosien and Rice (1998) found that a single low dose (but not higher doses) of tributyltin oxide resulted in enhanced macrophage secretory function and the respiratory burst in mice six days after intraperitoneal injection. Because the burst in a macrophage can take up to 24 h to detect (Sponseller et al 2016), the zebrafish RBA may not be sensitive enough to detect a macrophage respiratory burst after only 2.5 h of PMA treatment (instead primarily detecting neutrophil respiratory bursts). This interpretation is supported by the flow cytometry data in this study wherein it was observed that there were roughly 10-fold more neutrophils at 96 hpf as compared to macrophages (see Figure 5).…”

Section: Compounds With Developmental But No Observable Immunotoxicitymentioning

confidence: 99%

In vivoassessment of respiratory burst inhibition by xenobiotic exposure using larval zebrafish

Phelps

Fletcher

Rodríguez-Nunez

et al. 2020

Journal of Immunotoxicology

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Currently, assessment of the potential immunotoxicity of a given agent involves a tiered approach for hazard identification and mechanistic studies, including observational studies, evaluation of immune function, and measurement of susceptibility to infectious and neoplastic diseases. These studies generally use costly low-throughput mammalian models. Zebrafish, however, offer an excellent alternative due to their rapid development, ease of maintenance, and homology to mammalian immune system function and development. Larval zebrafish also are a convenient model to study the innate immune system with no interference from the adaptive immune system. In this study, a respiratory burst assay (RBA) was utilized to measure reactive oxygen species (ROS) production after developmental xenobiotic exposure. Embryos were exposed to non-teratogenic doses of chemicals and at 96 h post-fertilization, the ability to produce ROS was measured. Using the RBA, 12 compounds with varying immune-suppressive properties were screened. Seven compounds neither suppressed nor enhanced the respiratory burst; five reproducibly suppressed global ROS production, but with varying potencies: benzo[a]pyrene, 17b-estradiol, lead acetate, methoxychlor, and phenanthrene. These five compounds have all previously been reported as immunosuppressive in mammalian innate immunity assays. To evaluate whether the suppression of ROS by these compounds was a result of decreased immune cell numbers, flow cytometry with transgenic zebrafish larvae was used to count the numbers of neutrophils and macrophages after chemical exposure. With this assay, benzo[a]pyrene was found to be the only chemical that induced a change in the number of immune cells by increasing macrophage but not neutrophil numbers. Taken together, this work demonstrates the utility of zebrafish larvae as a vertebrate model for identifying compounds that impact innate immune function at non-teratogenic levels and validates measuring ROS production and phagocyte numbers as metrics for monitoring how xenobiotic exposure alters the innate immune system.

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Macrophage effector responses of horses are influenced by expression of CD154

Cited by 2 publications

References 26 publications

CD154 Expression Indicates T Cell Activation Following Tetanus Toxoid Vaccination of Horses

CD154 Expression Indicates T Cell Activation Following Tetanus Toxoid Vaccination of Horses

In vivoassessment of respiratory burst inhibition by xenobiotic exposure using larval zebrafish

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