PMID- 31689421
OWN - NLM
STAT- MEDLINE
DCOM- 20200929
LR  - 20200929
IS  - 1872-7905 (Electronic)
IS  - 0022-1759 (Linking)
VI  - 477
DP  - 2020 Feb
TI  - rM-CSF efficiently replaces L929 in generating mouse and rat bone marrow-derived 
      macrophages for in vitro functional studies of immunity to intracellular 
      bacteria.
PG  - 112693
LID - S0022-1759(19)30291-1 [pii]
LID - 10.1016/j.jim.2019.112693 [doi]
AB  - Methods used to prepare bone marrow-derived macrophages (BMDMs) may influence the 
      outcomes of immunological assays in which they are used. Supernatant conditioned 
      by growth of L929 cells has often been used to generate mouse macrophages from 
      bone marrow in vitro but is subject to lot-to-lot variability. To reduce 
      experimental variability and to standardize techniques across laboratories, we 
      investigated recombinant M-CSF (rM-CSF) as an alternative supplement for BMDM 
      maturation in the context of macrophage infection, using the intracellular 
      bacterium Live Vaccine Strain (LVS) of Francisella tularensis as a prototype. We 
      compared rM-CSF with L929 supernatant in terms of their effects on mouse and rat 
      macrophage growth, maturation patterns, surface marker expression, and the 
      expression of selected genes. Further, we compared macrophage infectivity and 
      bacterial replication using LVS. Finally, we compared the in vitro function of 
      BMDMs co-cultured with splenocytes from vaccinated animals in terms of their 
      control of intramacrophage bacterial replication, as well as production of 
      cytokines and nitric oxide. We demonstrated that rM-CSF produced BMDMs with 
      similar, or minimal, phenotypic and gene expression outcomes compared to those 
      generated with media containing L929 supernatant. Most importantly, functional 
      outcomes were similar. Taken together, our data support the use of the rM-CSF in 
      cell culture media as an alternative to L929-supplemented media for functional 
      bioassays that use C57BL/6J mouse or Fischer 344 rat BMDMs to study intracellular 
      infections. This comparison therefore facilitates future protocol 
      standardization.
CI  - Published by Elsevier B.V.
FAU - Rice, Helen M
AU  - Rice HM
AD  - Laboratory of Mucosal Pathogens and Cellular Immunology, Division of Bacterial, 
      Parasitic and Allergenic Products, Center for Biologics Evaluation and Research, 
      Food and Drug Administration, Silver Spring, MD, USA.
FAU - Rossi, Amy P
AU  - Rossi AP
AD  - Laboratory of Mucosal Pathogens and Cellular Immunology, Division of Bacterial, 
      Parasitic and Allergenic Products, Center for Biologics Evaluation and Research, 
      Food and Drug Administration, Silver Spring, MD, USA.
FAU - Bradford, Mary Katherine
AU  - Bradford MK
AD  - Laboratory of Mucosal Pathogens and Cellular Immunology, Division of Bacterial, 
      Parasitic and Allergenic Products, Center for Biologics Evaluation and Research, 
      Food and Drug Administration, Silver Spring, MD, USA.
FAU - Elkins, Karen L
AU  - Elkins KL
AD  - Laboratory of Mucosal Pathogens and Cellular Immunology, Division of Bacterial, 
      Parasitic and Allergenic Products, Center for Biologics Evaluation and Research, 
      Food and Drug Administration, Silver Spring, MD, USA. Electronic address: 
      karen.elkins@fda.hhs.gov.
FAU - De Pascalis, Roberto
AU  - De Pascalis R
AD  - Laboratory of Mucosal Pathogens and Cellular Immunology, Division of Bacterial, 
      Parasitic and Allergenic Products, Center for Biologics Evaluation and Research, 
      Food and Drug Administration, Silver Spring, MD, USA. Electronic address: 
      roberto.depascalis@fda.hhs.gov.
LA  - eng
PT  - Journal Article
DEP - 20191102
PL  - Netherlands
TA  - J Immunol Methods
JT  - Journal of immunological methods
JID - 1305440
RN  - 0 (Bacterial Vaccines)
RN  - 0 (CSF1 protein, human)
RN  - 0 (Culture Media)
RN  - 0 (Recombinant Proteins)
RN  - 0 (Vaccines, Attenuated)
RN  - 81627-83-0 (Macrophage Colony-Stimulating Factor)
SB  - IM
MH  - Animals
MH  - Bacterial Infections/immunology
MH  - Bacterial Vaccines/immunology
MH  - Biological Assay/methods
MH  - Cell Culture Techniques/*methods
MH  - Cell Differentiation/drug effects
MH  - Cell Line
MH  - Coculture Techniques/methods
MH  - Culture Media/*pharmacology
MH  - Female
MH  - Fibroblasts
MH  - Francisella tularensis/immunology
MH  - Gene Expression Regulation/immunology
MH  - Immunoassay/methods
MH  - Lymphocytes
MH  - Macrophage Colony-Stimulating Factor/*pharmacology
MH  - Macrophages/*drug effects/immunology/metabolism/microbiology
MH  - Male
MH  - Mice
MH  - Mice, Inbred C57BL
MH  - Rats
MH  - Rats, Inbred F344
MH  - Recombinant Proteins/metabolism
MH  - Vaccines, Attenuated/immunology
OTO - NOTNLM
OT  - Francisella tularensis
OT  - L929
OT  - LVS
OT  - Macrophage culture
OT  - in vitro testing
OT  - rM-CSF
EDAT- 2019/11/07 06:00
MHDA- 2020/09/30 06:00
CRDT- 2019/11/06 06:00
PHST- 2019/08/01 00:00 [received]
PHST- 2019/10/30 00:00 [revised]
PHST- 2019/10/30 00:00 [accepted]
PHST- 2019/11/07 06:00 [pubmed]
PHST- 2020/09/30 06:00 [medline]
PHST- 2019/11/06 06:00 [entrez]
AID - S0022-1759(19)30291-1 [pii]
AID - 10.1016/j.jim.2019.112693 [doi]
PST - ppublish
SO  - J Immunol Methods. 2020 Feb;477:112693. doi: 10.1016/j.jim.2019.112693. Epub 2019 
      Nov 2.