Enhancing the toolbox to study IL-17A in cattle and sheep

Wattegedera, Sean; Corripio-Miyar, Yolanda; Pang, Yvonne; Frew, David; McNeilly, Tom N.; Palarea‐Albaladejo, Javier; McInnes, Colin J.; Hope, Jayne; Glass, Elizabeth; Entrican, Gary

doi:10.1186/s13567-017-0426-5

Cited by 19 publications

(17 citation statements)

References 55 publications

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“…Interleukin (IL)-4 and IL-10 were measured in an indirect ELISA using antibody pairs (mAb clones CC313 and CC314 for IL-4; CC318 and CC320 for IL-10, Bio-Rad Antibody Laboratories, Oxford UK) and quantified using recombinant bovine (rbov) IL-4 of known concentration and recombinant ovine IL-10 of known biological activity [12,17]. IL-17A was assayed and quantified using the bovine IL-17A ELISA kit (Kingfisher Biotech, Minneapolis, MN, USA) with rbov IL-17A standards, according to the manufacturer's instructions; the kit cross-reactivity for ovine IL-17A has previously been reported [18].…”

Section: Cytokine Elisasmentioning

confidence: 99%

Defining immune correlates during latent and active chlamydial infection in sheep

Wattegedera

Livingstone

Maley

et al. 2020

Vet Res

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Ovine enzootic abortion (OEA) caused by the obligate intracellular bacterial pathogen Chlamydia abortus (C. abortus), is an endemic disease in most sheep-rearing countries worldwide. Following infection, C. abortus establishes a complex host-pathogen interaction with a latent phase in non-pregnant sheep followed by an active disease phase in the placenta during pregnancy leading to OEA. Improved knowledge of the host-pathogen interactions at these different phases of disease will accelerate the development of new diagnostic tests and vaccines to control OEA. Current evidence indicates that cellular immunity is essential for controlling C. abortus infection. We have previously described a model of mucosal (intranasal) infection of non-pregnant sheep with C. abortus that replicates the latent and active phases of OEA. We have investigated antigen-specific recall responses of peripheral blood mononuclear cells (PBMC) in sheep infected with C. abortus via the intranasal route to determine how these change during the latent and active phases of disease. By analysing cytokines associated with the major CD4 +ve T helper (T h) cell subsets (Interferon-gamma (IFN-γ)/T h 1; Interleukin (IL)-4/T h 2; IL-17A/T h 17; IL-10/T regulatory), we show that there is selective activation of PBMC producing IFN-γ and/or IL-10 during the latent phase following infection. These cytokines are also elevated during the active disease phase and while they are produced by sheep that are protected from OEA, they are also produced by sheep that abort, highlighting the difficulties in finding specific cellular immunological correlates of protection for complex intracellular pathogens.

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Section: Cytokine Elisasmentioning

confidence: 99%

Defining immune correlates during latent and active chlamydial infection in sheep

Wattegedera

Livingstone

Maley

et al. 2020

Vet Res

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“…IL-17a is the “signature cytokine” secreted by the Th-17 CD4+ve T cell subset. Activation of Th-17-type responses is important not only for host immune control of extracellular bacterial and fungal infections but also associated with chronic inflammation and autoimmunity [ 9 ]. The IL-1RA protein is a naturally occurring antagonist of pro-inflammatory cytokines.…”

Section: Introductionmentioning

confidence: 99%

The role of serum inflammatory cytokines and berberine in the insulin signaling pathway among women with polycystic ovary syndrome

Kuang¹,

Duan²,

Li³

et al. 2020

PLoS ONE

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Objective To study the role of selected serum inflammatory cytokines and berberine in the insulin signaling pathway among women with polycystic ovary syndrome (PCOS). Methods Selected serum inflammatory cytokines were analyzed in the particle cells, which were interfered by berberine, from 78 infertile women who were to be treated with In Vitro Fertilization (IVF) /Intracytoplasmic Sperm Injection-Embryo Transfer (icsi-et). Among them, 49 patients had PCOS infertility, and 29 were non-PCOS patients whose infertility resulted from fallopian tube and male factors. The elisa method was used to detect the changes in the expression levels of inflammatory factors in the cells. The correlations between the serum inflammatory cytokine expression levels and the corresponding clinical hormones were analyzed. The changes in the expression (mRNA and protein) levels of the serum inflammatory cytokines were studied by real-time quantitative PCR and protein printing. Fluorescence microscope and flow cytometry were used to detect the glucose uptake capacity of ovarian granulosa cells in PCOS patients under the action of insulin after berberine. Results In the PCOS group, IL-17a (P = 0.001), IL-1Ra (P<0.0001), and IL-6 (P = 0.035) were significantly higher than those in the non-PCOS group. In the non-PCOS group, AMH level was negatively correlated with inflammatory cytokines IL-17a (r = -0.819;P = 0.004), IL-1a (r = -0.716;P = 0.0.02), IL-1b (r = -0.678;P = 0.031), IL-2 (r = -0.765;P = 0.01), and IL-8 (r = -0.705;P = 0.023). However, in the PCOS group, AMH levels were not significantly correlated with the levels of the examined inflammatory cytokines. Berberine significantly reduced the expression level of mTOR mRNA (P = 0.001), and increased the expression level of IRS-1 mRNA (P = 0.009) in the PCOS granule cells. Conclusion In this study, we find that the elevated levels of serum inflammatory factors IL-17a, IL-1Ra, and IL-6 cause women to be in a subclinical inflammatory state for a long time. Abnormal changes in inflammatory factors alter their original negative correlations with AMH levels, thereby weakening the metabolism of glycolipids, promoting insulin resistance, destroying the normal ovulation and fertilization system of women, leading to polycystic ovary syndrome characterized by menstrual thinning and abnormal ovulation. Berberine can improve the sensitivity of insulin by regulating the signal pathway of insulin receptor substrate-1 (IRS-1) and mammalian target of rapamycin (mTOR) in PCOS patients and achieve a therapeutic effect of treating PCOS.

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“…There are limited commercial monoclonal antibodies available for the research of goat IL-17A so far. Although some researchers have tried to find the cross-reactivities of commercial human or mice monoclonal antibodies available for ruminants [35, 36], or used the commercial bovine IL-17A as an antigen to prepare mAbs that react with goat and sheep [37], there are only a few monoclonal antibodies available for caprine IL-17A. In this study, the prokaryotically expressed soluble recombinant fusion cIL-17A was used as an antigen to prepare monoclonal antibodies against goat IL-17A, and the monoclonal antibody H8 was successfully used in western blot, immunohistochemistry and immunofluorescence.…”

Section: Discussionmentioning

confidence: 99%

Preparation of a novel monoclonal antibody against caprine interleukin-17A and its applications in immunofluorescence and immunohistochemistry assays

et al. 2019

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Background Interleukin-17 (IL-17), the characteristic cytokine secreted by T helper 17 lymphocytes (Th17 cells), plays a pivotal role in host defense and many inflammatory or autoimmune diseases. The aim of this study was to obtain purified protein caprine IL-17A (cIL-17A) as an antigen for preparing an IL-17A-specific monoclonal antibody (mAb). Results The coding sequence (CDS) region of cIL-17A was cloned from the peripheral blood mononuclear cells (PBMCs) of dairy goats and then inserted into the expression vector PET 32a and transformed into competent TransB (DE3) cells. Recombinant fusion protein obtained under optimized conditions was used to immunize BALB/c mice for preparing monoclonal antibodies. Finally, the supernatants of two hybridoma cell lines showing positive reaction with the recombinant fusion protein and negative reaction with fusion tags of PET 32a were collected for western blot, immunofluorescence (IF) and immunohistochemistry (IHC) analysis. Our results showed that the maximum amount of soluble protein could be obtained directly in the supernatant when the recombinant expression cells were induced by isopropyl-β-d-thiogalactoside (IPTG) at a concentration of 0.3 mmol/L at 16 °C for 42 h. Western blot analysis showed that the mAb H8 could recognize the eukaryotically expressed cIL-17A in the supernatant of transfected HEK293T cells. Immunofluorescence and immunohistochemistry assays showed that mAb H8 could strongly recognize both the eukaryotically expressed and natural cIL-17A. Conclusions The monoclonal antibody mAb H8 prepared in this study may be a potential tool for the detection of cIL-17A and beneficial for investigating the pathogenesis of various IL-17-associated diseases. Electronic supplementary material The online version of this article (10.1186/s12896-019-0543-5) contains supplementary material, which is available to authorized users.

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Enhancing the toolbox to study IL-17A in cattle and sheep

Cited by 19 publications

References 55 publications

Defining immune correlates during latent and active chlamydial infection in sheep

Defining immune correlates during latent and active chlamydial infection in sheep

The role of serum inflammatory cytokines and berberine in the insulin signaling pathway among women with polycystic ovary syndrome

Preparation of a novel monoclonal antibody against caprine interleukin-17A and its applications in immunofluorescence and immunohistochemistry assays

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