Low levels of soluble DPP4 among Saudis may have constituted a risk factor for MERS endemicity.

Most of the cases of Middle East respiratory syndrome coronavirus (MERS-CoV) were reported in Saudi Arabia. Dipeptidyl peptidase-4 (DPP4) was identified as the receptor for the virus. The level of soluble DPP4 (sDPP4) was found to be reduced in MERS-CoV infected patients while high levels of sDPP4 were suggested to be protective against MERS-CoV in animal models. We investigated whether the Saudi population has lower levels of sDPP4 which makes them more susceptible to MERS-CoV infection and, therefore, could explain the larger number of cases from the country. Blood samples were collected from 219 Saudi blood donors and 200 blood donors from other ethnic groups. The plasma level of sDPP4 was measured by ELISA and the following SNPs in the DPP4 gene; rs35128070, rs1861978, rs79700168, and rs17574, were genotyped by TaqMan SNP genotyping assay. The average level of plasma sDDP4 was significantly lower in Saudis than other Arabs and non-Arabs (P value 0.0003 and 0.012, respectively). The genotypes AG of rs35128070 and GT of rs1861978 were significantly associated with lower sDPP4 among Saudis (P value 0.002 for each). While both genotypes AA and AG of rs79700168 and rs17574 were associated with significantly lower average sDPP4 level in Saudis compared to other ethnic groups (P value 0.031 and 0.032, and 0.027 and 0.014, respectively). Herein, we report that the Saudi population has lower levels of plasma sDPP4 than other ethnic groups, which is associated with genetic variants in the DPP4 gene. This may have contributed to increase the susceptibility of the Saudi population to MERS-CoV infection and could be a factor in the long-lasting persistence of the virus in the country.

Leukocyte populations in peripheral blood of dromedary camels with clinical endometritis.

Endometritis represents the main cause of reproductive failure in dromedary camels. In dromedary camels, associations between endometritis-causing pathogen-species, disease severity, and systemic changes in the immune system have not been evaluated. In the current study, there was use of flow cytometry and immunofluorescence of membrane proteins for the evaluation of leukocyte subsets and the cellular phenotype in blood of camels with clinical endometritis and evaluations of associations with disease severity and endometritis-causing pathogens. Animals with endometritis had markedly larger numbers of total leukocytes and neutrophils. Although total lymphocyte and monocyte counts did not differ between camels with and without clinical endometritis, there were lesser numbers of total and effector CD4-positive T cells in camels with endometritis. Among monocytes, number of camel inflammatory monocytes (Mo-II) was markedly greater, whereas Mo-III numbers were less in the blood of camels with clinical endometritis. Number of inflammatory monocytes was also indicative of endometritis severity grade. Among camels with clinical endometritis, E. coli- and S. aureus-infected animals had similar endometritis grades and comparable phenotype and composition patterns of leukocytes. Neutrophils and monocytes of camels with clinical endometritis had fewer cell adhesion molecules (i.e., CD11a and CD18). Collectively, the results from the current study allowed for identification of associations between endometritis severity grade and larger numbers of inflammatory monocytes. The results also indicate there is no association between endometritis pathogen-species and changes in phenotype or composition of blood leukocytes.

Neutrophil degranulation differentially modulates phenotype and function of bovine monocyte subsets.

Monocytes and neutrophils are important players in the innate immune response and cooperate during infection and inflammation. In our study we analyzed the effects of neutrophil degranulation products (polymorphonuclear granulocytes degranulation products, PMN-DGP) on the activation, the adhesion and the migration of three bovine monocyte subsets, as well as their effects on monocyte-macrophage differentiation. Cross-linking of surface CD18 molecules on bovine PMN resulted in the release of primary, secondary and tertiary granules as well as of secretory vesicles. PMN-DGP induced a significant Ca2+-influx in classical (classical monocytes, cM) and intermediate monocytes (intermediate monocytes, intM) but not in non-classical monocytes (non-classical monocytes, ncM). A selective and up-regulated expression induced by PMN-DGP was only seen for CD11a and CD31 on intM. PMN-DGP induced a selective migration of intM in vitro. The presence of PMN-DGP during the differentiation of cM or intM into macrophages resulted in increased expression of membrane CD163 and reduced expression of MHC-II molecules. PMN-DGP-derived macrophages produced more IL-12 and IL-10 and showed enhanced phagocytosis and ROS production capacities. In conclusion, PMN-DGP selectively attract bovine intM and skew the functional maturation of cM and intM.

The chemokine CCL5 induces selective migration of bovine classical monocytes and drives their differentiation into LPS-hyporesponsive macrophages in vitro.

Human and mouse studies indicate distinct roles of selected chemokines for monocyte subset attraction. We therefore analyzed the still unknown sensitivity and response of bovine monocyte subsets toward two monocyte-attracting chemokines (CCL2, CCL5). Only CCL5 induced a significant Ca(2+)influx and migration response in bovine monocytes, with classical and intermediate monocytes being significantly stimulated and attracted compared to nonclassical monocytes. The presence of CCL5 during in vitro macrophage differentiation did not alter their capacity to phagocytize or to generate reactive oxygen species upon stimulation with E. coli. However, macrophages differentiated in the presence of CCL5 displayed an altered phenotype with significantly less expressed CD14 and MHC class II molecules, whereas CD16 was upregulated. Moreover, CCL5-differentiated macrophages displayed a reduced upregulation of CXCL8, ARG1, IL6 and IL10 mRNA. Taken together, CCL5 but not CCL2 mainly attract bovine classical monocytes and promote their differentiation into LPS-hypo-responsive macrophages.

Recombinant bovine S100A8 and A9 enhance IL-1ß secretion of interferon-gamma primed monocytes.

Calgranulin A (S100A8) and B (S100A9) are found at high levels in inflamed tissue and have been associated with acute and chronic inflammatory disorders. Calgranulins are discussed as damage-associated molecular patterns (DAMPs). To analyze the role of calgranulins for inflammatory responses, bovine S100A8 and S100A9 were cloned, successfully expressed and FPLC-purified. Both molecules did not induce NF-κB activation in boTLR4-transfected HEK293 cells and stimulation of bovine monocytes with both proteins did not result in interleukin 1ß (IL-1ß) secretion or an upregulated mRNA expression of selected genes (IL1B, TNF, CXCL8, IL10, IL12). However, Interferon γ (IFN-γ) primed bovine monocytes released significantly higher amounts of IL-1ß after stimulation with S100A8, S100A9, and co-stimulation with adenosine triphosphate (ATP). In IL-4/IL-13-primed monocytes, the IL-1ß release was completely abrogated. The results imply that TLR4/MyD88/NF-κB-independent S100A8/A9-mediated activation of the inflammasome in cattle is favored in a Th1 environment and that S100A8 and S100A9 act as a DAMP in cattle.

Inflammasome activation in bovine monocytes by extracellular ATP does not require the purinergic receptor P2X7.

Extracellular adenosine triphosphate (ATP) is a second signal for the assembly of the NLR family, pyrin domain-containing 3 (NLRP3) inflammasome, which form a framework to activate caspase 1, leading to the processing and secretion of the pro-inflammatory cytokine interleukin-1ß (IL-1ß). The aim of the present study was to investigate the role of the ATP-gated ion channel subtype P2X7 receptor in the inflammasome activation of bovine monocytes. ATP-induced inflammasome assembly in bovine monocytes was shown by caspase-1 activation and the release of IL-1ß by LPS/ATP-stimulated bovine cells. The IL-1ß release depended on potassium efflux but was independent of reactive oxygen generation of bovine monocytes. Unlike in the human system, a P2X7 receptor antagonist did not block the ATP-induced release of IL-1ß of LPS-primed bovine cells. P2X7 mediated pore formation was observed in subsets of bovine T lymphocytes (CD4+>CD8+) but not in monocytes. In addition, ATP and 2-MeSATP but not the high affinity P2X7 agonist BzATP induced calcium influx in bovine monocytes. The data indicate that ROS generation plays no role in the ATP-induced activation of inflammasome in bovine monocytes and that P2X7-mediated pore formation is not necessary for the release of Interleukin-1ß.