TY - JOUR
T1 - Reduction of inflammation in a chronic periodontitis model in rats by TNF-α gene silencing with a topically applied siRNA-loaded calcium phosphate paste
AU - Tenkumo, Taichi
AU - Rojas-Sánchez, Leonardo
AU - Vanegas Sáenz, Juan Ramón
AU - Ogawa, Toru
AU - Miyashita, Makiko
AU - Yoda, Nobuhiro
AU - Prymak, Oleg
AU - Sokolova, Viktoriya
AU - Sasaki, Keiichi
AU - Epple, Matthias
N1 - Funding Information:
The authors would like to thank Ms. Satomi Oizumi (Tohoku University Graduate School of Dentistry) for her valuable work regarding the histological preparation. This work was financially supported by a Grant-in-Aid for Scientific Research from the Japan Society for the Promotion of Science and the Deutscher Akademischer Austauschdienst (DAAD). L.R.-S. thanks the Ministry of Science, Technology and Telecommunications of Costa Rica (MICITT, Costa Rica) for a Ph.D. scholarship. The funders had no role in the study design, data collection and analysis, decision to publish, or manuscript preparation.
Publisher Copyright:
© 2020
PY - 2020/3/15
Y1 - 2020/3/15
N2 - We developed a calcium phosphate-based paste containing siRNA against TNF-α and investigated its anti-inflammatory and bone-healing effects in vitro and in vivo in a rat periodontitis model. The bioactive spherical CaP/PEI/siRNA/SiO2 nanoparticles had a core diameter of 40–90 nm and a positive charge (+23 mV) that facilitated cellular uptake. The TNF- α gene silencing efficiency of the nanoparticles in J774.2 monocytes, gingival-derived cells, and bone marrow-derived cells was 12 ± 2%, 36 ± 8%, and 35 ± 22%, respectively. CaP/PEI/siRNA/SiO2 nanoparticles cancelled the suppression of alkaline phosphatase (ALP) activity in LPS-stimulated bone marrow-derived cells. In vivo, ALP mRNA was up-regulated, TNF-α mRNA was down-regulated, and the amount of released TNF-α was significantly reduced after topical application of the calcium phosphate-based paste containing siRNA-loaded nanoparticles. The number of TNF-α-positive cells in response to CaP/PEI/siRNA/SiO2 nanoparticle application was lower than that observed in the absence of siRNA. Elevated ALP activity and numerous TRAP-positive cells (osteoclasts) were observed in response to the application of all calcium phosphate pastes. These results demonstrate that local application of a paste consisting of siRNA-loaded calcium phosphate nanoparticles successfully induces TNF-α silencing in vitro and in vivo and removes the suppression of ALP activity stimulated by inflammation. Statement of significance: We developed a calcium phosphate-based paste containing nanoparticles loaded with siRNA against TNF-α. The nanoparticles had a core diameter of 40–90 nm and positive charge (+23 mV). The anti-inflammatory and osteoinductive effects of the paste were investigated in vitro and in vivo in a rat periodontitis model. In vitro, the TNF-α gene silencing efficiency of the nanoparticles in J774.2 monocytes, gingival-derived cells, and bone marrow-derived cells was 12 ± 2%, 36 ± 8%, and 35 ± 22%, respectively. The ALP activity of bone marrow-derived cells was recovered. In vivo, TNF-α mRNA was down-regulated and the amount of released TNF-α was significantly reduced, whereas the ALP mRNA was up-regulated. Elevated ALP activity and TRAP-positive cells were observed by immunohistochemistry.
AB - We developed a calcium phosphate-based paste containing siRNA against TNF-α and investigated its anti-inflammatory and bone-healing effects in vitro and in vivo in a rat periodontitis model. The bioactive spherical CaP/PEI/siRNA/SiO2 nanoparticles had a core diameter of 40–90 nm and a positive charge (+23 mV) that facilitated cellular uptake. The TNF- α gene silencing efficiency of the nanoparticles in J774.2 monocytes, gingival-derived cells, and bone marrow-derived cells was 12 ± 2%, 36 ± 8%, and 35 ± 22%, respectively. CaP/PEI/siRNA/SiO2 nanoparticles cancelled the suppression of alkaline phosphatase (ALP) activity in LPS-stimulated bone marrow-derived cells. In vivo, ALP mRNA was up-regulated, TNF-α mRNA was down-regulated, and the amount of released TNF-α was significantly reduced after topical application of the calcium phosphate-based paste containing siRNA-loaded nanoparticles. The number of TNF-α-positive cells in response to CaP/PEI/siRNA/SiO2 nanoparticle application was lower than that observed in the absence of siRNA. Elevated ALP activity and numerous TRAP-positive cells (osteoclasts) were observed in response to the application of all calcium phosphate pastes. These results demonstrate that local application of a paste consisting of siRNA-loaded calcium phosphate nanoparticles successfully induces TNF-α silencing in vitro and in vivo and removes the suppression of ALP activity stimulated by inflammation. Statement of significance: We developed a calcium phosphate-based paste containing nanoparticles loaded with siRNA against TNF-α. The nanoparticles had a core diameter of 40–90 nm and positive charge (+23 mV). The anti-inflammatory and osteoinductive effects of the paste were investigated in vitro and in vivo in a rat periodontitis model. In vitro, the TNF-α gene silencing efficiency of the nanoparticles in J774.2 monocytes, gingival-derived cells, and bone marrow-derived cells was 12 ± 2%, 36 ± 8%, and 35 ± 22%, respectively. The ALP activity of bone marrow-derived cells was recovered. In vivo, TNF-α mRNA was down-regulated and the amount of released TNF-α was significantly reduced, whereas the ALP mRNA was up-regulated. Elevated ALP activity and TRAP-positive cells were observed by immunohistochemistry.
KW - Anti-inflammation
KW - Calcium phosphate
KW - Gene silencing
KW - Nanoparticles
KW - Periodontitis
KW - TNF-α
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U2 - 10.1016/j.actbio.2020.01.031
DO - 10.1016/j.actbio.2020.01.031
M3 - Article
C2 - 31982590
AN - SCOPUS:85079154002
VL - 105
SP - 263
EP - 279
JO - Acta Biomaterialia
JF - Acta Biomaterialia
SN - 1742-7061
ER -