| Full text | |
| Author(s): Show less - |
Batista, Camila F.
[1]
;
Souza, Fernando N.
[1]
;
Santos, Kamila R.
[1]
;
Ramos Sanchez, Eduardo M.
[2]
;
Reis, Luiza Campos
[2]
;
Bertagnon, Heloisa G.
[1]
;
Blagitz, Maiara G.
[1]
;
Gomes, Renata C.
[1]
;
Lage, Andrey P.
[3]
;
Heinemann, Marcos B.
[4]
;
Della Libera, Alice M. M. P.
[1]
Total Authors: 11
|
| Affiliation: | [1] Univ Sao Paulo, Fac Med Vet & Zootecnia, Dept Clin Med, Vet Clin Immunol Res Grp, Av Prof Dr Orlando Marques de Paiva 87, BR-05508270 Sao Paulo, SP - Brazil
[2] Univ Sao Paulo, Lab Sorol & Imunobiol, Inst Med Trop Sao Paulo, Av Prof Dr Eneas de Carvalho Aguiar 470, BR-05403000 Sao Paulo, SP - Brazil
[3] Univ Fed Minas Gerais, Escola Vet, Dept Med Vet Prevent, Av Presidente Antonio Carlos 6627, BR-31270010 Belo Horizonte, MG - Brazil
[4] Univ Sao Paulo, Fac Med Vet & Zootecnia, Dept Med Vet Prevent & Saude Anim, Av Prof Dr Orlando Marques de Paiva 87, BR-05508270 Sao Paulo, SP - Brazil
Total Affiliations: 4
|
| Document type: | Journal article |
| Source: | VETERINARY IMMUNOLOGY AND IMMUNOPATHOLOGY; v. 196, p. 53-59, FEB 2018. |
| Web of Science Citations: | 0 |
| Abstract | |
The present study aimed to validate the use of R-phycoerythrin (R-PE)-labeled Mannheimia haemolytica to simultaneously stimulate phagocytosis and intracellular production of reactive oxygen species (ROS) by blood phagocytes in bronchoalveolar lavage (BAL) fluid. Initially, R-PE-labeled M. haemolytica was inactivated using a water bath at 60 degrees C for 60 min. Afterwards, R-PE labelling of bacteria was confirmed by flow cytometry. The geometric mean fluorescence intensity of R-PE-labeled bacteria (FL2 detector, 585 +/- 42 nm) was analyzed by flow cytometry and was 41.5-fold higher than the respective unlabeled controls, confirming the success of bacterial conjugation to R-PE. Phagocytosis and intracellular production of ROS by blood neutrophils and monocytes, and by BAL CD14 macrophages, in 12 healthy 6-month-old male calves were then performed using R-PE-labeled bacteria and 2',7'-dichlorofluoresceindiacetate (DCFH-DA) as probes. Confocal microscopy was used to confirm phagocytosis of R-PE-labeled M. haemolytica by phagocytes and to exclude erroneous measurements of bacteria adhering to the leukocyte membrane. The present study showed that there is no difference in the ROS production without stimulus and in the presence of M. haemolytica by peripheral blood neutrophils and monocytes, in contrast to the increased ROS production by local alveolar macrophages upon stimulation by M. haemolytica. This emphasizes the importance of alveolar macrophages in the maintenance of homeostasis and health of the respiratory system, which can be supported during the inflammatory process by the rapid recruitment of neutrophils with high microbicidal and phagocytic capacity. The method described here provides an easy and feasible tool to measure phagocytosis and intracellular ROS production by phagocytes, especially when commonly used probes for intracellular ROS production were used, such as DCFH-DA and dihydrorhodamine 123. (AU) | |
| FAPESP's process: | 14/23189-4 - Intramammary infections by Staphylococcus aureus: crosstalk between mammary epithelial cells and peripheral blood mononuclear cells, and how it governs immunity in the mammary gland of dairy cows |
| Grantee: | Fernando Nogueira de Souza |
| Support Opportunities: | Scholarships in Brazil - Post-Doctoral |
| FAPESP's process: | 11/23936-6 - Dynamics of blood and alveolar phagocytes in calves with mannheimiosis |
| Grantee: | Camila Freitas Batista |
| Support Opportunities: | Scholarships in Brazil - Doctorate |
| FAPESP's process: | 12/00749-9 - Dynamics of blood and alveolar phagocytes in calves with mannhemiosis |
| Grantee: | Alice Maria Melville Paiva Della Libera |
| Support Opportunities: | Regular Research Grants |