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2.
Vaccines (Basel) ; 9(11)2021 Nov 09.
Artigo em Inglês | MEDLINE | ID: mdl-34835231

RESUMO

Staphylococcus aureus is a causative agent of bovine mastitis, capable of causing significant economic losses to the dairy industry worldwide. This study focuses on obtaining single-chain fragment variables (scFvs) against the virulence factors of S. aureus and evaluates the protective effect of scFvs on bovine mammary epithelial (MAC-T) cells and mice mammary gland tissues infected by S. aureus. After five rounds of bio-panning, four scFvs targeting four virulence factors of S. aureus were obtained. The complementarity-determining regions (CDRs) of these scFvs exhibited significant diversities, especially CDR3 of the VH domain. In vitro, each of scFvs was capable of inhibiting S. aureus growth and reducing the damage of MAC-T cells infected by S. aureus. Preincubation of MAC-T cells with scFvs could significantly attenuate the effect of apoptosis and necrosis compared with the negative control group. In vivo, the qPCR and ELISA results demonstrated that scFvs reduced the transcription and expression of Tumor Necrosis Factor alpha (TNF-α), interleukin-1ß (IL-1ß), IL-6, IL-8, and IL-18. Histopathology and myeloperoxidase (MPO) results showed that scFvs ameliorated the histopathological damages and reduced the inflammatory cells infiltration. The overall results demonstrated the positive anti-inflammatory effect of scFvs, revealing the potential role of scFvs in the prevention and treatment of S. aureus infections.

3.
Int J Mol Sci ; 23(1)2021 Dec 29.
Artigo em Inglês | MEDLINE | ID: mdl-35008761

RESUMO

Staphylococcus aureus is a vital bovine mastitis pathogen causing huge economic losses to the dairy industry worldwide. In our previous studies, leukotoxin ED (LukED) was detected in most S. aureus strains isolated from bovine mastitis. Here, four single-chain fragment variables (scFvs) (ZL8 and ZL42 targeting LukE, ZL22 and ZL23 targeting LukD) were obtained using purified LukE and LukD proteins as the antigens after five rounds of bio-panning. The complementarity-determining region 3 (CDR3) of the VH domain of these scFvs exhibited significant diversities. In vitro, the scFvs significantly decreased LukED-induced cell killing by inhibiting the binding of LukED to chemokine receptors (CCR5 and CXCR2) and reduced the death rates of bovine neutrophils and MAC-T cells caused by LukED and S. aureus (p < 0.05). In an S. aureus-induced mouse mastitis model, histopathology and MPO results revealed that scFvs ameliorated the histopathological damages and reduced the infiltration of inflammatory cells (p < 0.05). The ELISA and qPCR assays showed that scFvs reduced the transcription and expression levels of Tumor Necrosis Factor-alpha (TNF-α), interleukin-1ß (IL-1ß), IL-6, IL-8 and IL-18 (p < 0.05). The overall results demonstrated the protective anti-inflammatory effect of scFvs in vitro and in vivo, enlightening the potential role of scFvs in the prevention and treatment of S. aureus-induced mastitis.


Assuntos
Inflamação/patologia , Leucocidinas/metabolismo , Mastite/microbiologia , Anticorpos de Cadeia Única/imunologia , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/fisiologia , Animais , Bovinos , Morte Celular , Citocinas/metabolismo , Feminino , Células HEK293 , Humanos , Mediadores da Inflamação/metabolismo , Leucocidinas/isolamento & purificação , Glândulas Mamárias Animais/patologia , Camundongos , Camundongos Endogâmicos BALB C , Neutrófilos/metabolismo , Biblioteca de Peptídeos , Peroxidase/metabolismo , Receptores de Quimiocinas/metabolismo , Anticorpos de Cadeia Única/isolamento & purificação , Infecções Estafilocócicas/patologia , Staphylococcus aureus/crescimento & desenvolvimento
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