Extensive protein expression changes induced by pamidronate in RAW 264.7 cells as determined by IP-HPLC.
Bisphosphonate
Global protein expressions
IP-HPLC
Molecular signaling
Pamidronate
RAW 264.7 cells
Journal
PeerJ
ISSN: 2167-8359
Titre abrégé: PeerJ
Pays: United States
ID NLM: 101603425
Informations de publication
Date de publication:
2020
2020
Historique:
received:
29
01
2020
accepted:
24
04
2020
entrez:
9
6
2020
pubmed:
9
6
2020
medline:
9
6
2020
Statut:
epublish
Résumé
Bisphosphonate therapy has become a popular treatment for osteoporosis, Paget's disease, multiple myeloma, osteogenesis imperfecta, myocardial infarction, and cancer despite its serious side effects. Bisphosphonate-induced molecular signaling changes in cells are still not clearly elucidated. As bisphosphonates are primarily engulfed by macrophages, we treated RAW 264.7 cells (a murine macrophage cell line) with pamidronate and investigated global protein expressional changes in cells by immunoprecipitation high performance liquid chromatography (IP-HPLC) using 218 antisera. Pamidronate upregulated proliferation-activating proteins associated with p53/Rb/E2F and Wnt/β-catenin pathways, but downregulated the downstream of RAS signaling, pAKT1/2/3, ERK-1, and p-ERK-1, and subsequently suppressed cMyc/MAX/MAD network. However, in situ proliferation index of pamidronate-treated RAW264.7 cells was slightly increased by 3.2% vs. non-treated controls. Pamidronate-treated cells showed increase in the expressions of histone- and DNA methylation-related proteins but decrease of protein translation-related proteins. NFkB signaling was also suppressed as indicated by the down-regulations of p38 and p-p38 and the up-regulation of mTOR, while the protein expressions related to cellular protection, HSP-70, NRF2, JNK-1, and LC3 were upregulated. Consequently, pamidronate downregulated the protein expressions related to immediate inflammation,cellular differentiation, survival, angiogenesis, and osteoclastogenesis, but upregulated PARP-1 and FAS-mediated apoptosis proteins. These observations suggest pamidronate affects global protein expressions in RAW 264.7 cells by stimulating cellular proliferation, protection, and apoptosis but suppressing immediate inflammation, differentiation, osteoclastogenesis, and angiogenesis. Accordingly, pamidronate appears to affect macrophages in several ways eliciting not only its therapeutic effects but also atypical epigenetic modification, protein translation, RAS and NFkB signalings. Therefore, our observations suggest pamidronate-induced protein expressions are dynamic, and the affected proteins should be monitored by IP-HPLC to achieve the therapeutic goals during treatment.
Sections du résumé
BACKGROUND
BACKGROUND
Bisphosphonate therapy has become a popular treatment for osteoporosis, Paget's disease, multiple myeloma, osteogenesis imperfecta, myocardial infarction, and cancer despite its serious side effects. Bisphosphonate-induced molecular signaling changes in cells are still not clearly elucidated.
METHODS
METHODS
As bisphosphonates are primarily engulfed by macrophages, we treated RAW 264.7 cells (a murine macrophage cell line) with pamidronate and investigated global protein expressional changes in cells by immunoprecipitation high performance liquid chromatography (IP-HPLC) using 218 antisera.
RESULTS
RESULTS
Pamidronate upregulated proliferation-activating proteins associated with p53/Rb/E2F and Wnt/β-catenin pathways, but downregulated the downstream of RAS signaling, pAKT1/2/3, ERK-1, and p-ERK-1, and subsequently suppressed cMyc/MAX/MAD network. However, in situ proliferation index of pamidronate-treated RAW264.7 cells was slightly increased by 3.2% vs. non-treated controls. Pamidronate-treated cells showed increase in the expressions of histone- and DNA methylation-related proteins but decrease of protein translation-related proteins. NFkB signaling was also suppressed as indicated by the down-regulations of p38 and p-p38 and the up-regulation of mTOR, while the protein expressions related to cellular protection, HSP-70, NRF2, JNK-1, and LC3 were upregulated. Consequently, pamidronate downregulated the protein expressions related to immediate inflammation,cellular differentiation, survival, angiogenesis, and osteoclastogenesis, but upregulated PARP-1 and FAS-mediated apoptosis proteins. These observations suggest pamidronate affects global protein expressions in RAW 264.7 cells by stimulating cellular proliferation, protection, and apoptosis but suppressing immediate inflammation, differentiation, osteoclastogenesis, and angiogenesis. Accordingly, pamidronate appears to affect macrophages in several ways eliciting not only its therapeutic effects but also atypical epigenetic modification, protein translation, RAS and NFkB signalings. Therefore, our observations suggest pamidronate-induced protein expressions are dynamic, and the affected proteins should be monitored by IP-HPLC to achieve the therapeutic goals during treatment.
Identifiants
pubmed: 32509464
doi: 10.7717/peerj.9202
pii: 9202
pmc: PMC7246033
doi:
Types de publication
Journal Article
Langues
eng
Pagination
e9202Informations de copyright
© 2020 Lee et al.
Déclaration de conflit d'intérêts
The authors declare that they have no competing interests.
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