TY - JOUR
T1 - Specific Silencing of Microglial Gene Expression in the Rat Brain by Nanoparticle-Based Small Interfering RNA Delivery
AU - Guo, Shanshan
AU - Cázarez-Márquez, Fernando
AU - Jiao, Han
AU - Foppen, Ewout
AU - Korpel, Nikita L.
AU - Grootemaat, Anita E.
AU - Liv, Nalan
AU - Gao, Yuanqing
AU - van der Wel, Nicole
AU - Zhou, Bing
AU - Nie, Guangjun
AU - Yi, Chun-Xia
N1 - Funding Information: This work was supported by a joint grant by The Netherlands Organisation for Health Research and Development (ZonMw) and the Dutch Diabetes Research Foundation (C.-X.Y., 459001004). We thank Andries Kalsbeek (Netherlands Institute for Neuroscience) for his intellectual input, and Tineke Veenendaa (Cell Microscopy Core, University Medical Center Utrecht) for her technical support with synthesizing the Au-555. Publisher Copyright: ©
PY - 2022/2/2
Y1 - 2022/2/2
N2 - Microglia are the major innate immune cells in the brain and are essential for maintaining homeostasis in a neuronal microenvironment. Currently, a genetic tool to modify microglial gene expression in specific brain regions is not available. In this report, we introduce a tailor-designed method that uses lipid and polymer hybridized nanoparticles (LPNPs) for the local delivery of small interfering RNAs (siRNAs), allowing the silencing of specific microglial genes in the hypothalamus. Our physical characterization proved that this LPNP-siRNA was uniform and stable. We demonstrated that, due to their natural phagocytic behavior, microglial cells are the dominant cell type taking up these LPNPs in the hypothalamus of rats. We then tested the silencing efficiency of LPNPs carrying a cluster of differentiation molecule 11b (CD11b) or Toll-like receptor 4 (TLR4) siRNA using different in vivo and in vitro approaches. In cultured microglial cells treated with LPNP-CD11b siRNA or LPNP-TLR4 siRNA, we found a silencing efficiency at protein expression levels of 65 or 77%, respectively. In line with this finding, immunohistochemistry and western blotting results from in vivo experiments showed that LPNP-CD11b siRNA significantly inhibited microglial CD11b protein expression in the hypothalamus. Furthermore, following lipopolysaccharide (LPS) stimulation of cultured microglial cells, gene expression of the TLR4 downstream signaling component myeloid differentiation factor 88 and its associated cytokines was significantly inhibited in LPNP-TLR4 siRNA-treated microglial cells compared with cells treated with LPNP-scrambled siRNA. Finally, after LPNP-TLR4 siRNA injection into the rat hypothalamus, we observed a significant reduction in microglial activation in response to LPS compared with the control rats injected with LPNP-scrambled siRNA. Our results indicate that LPNP-siRNA is a promising tool to manipulate microglial activity locally in the brain and may serve as a prophylactic approach to prevent microglial dysfunction-associated diseases.
AB - Microglia are the major innate immune cells in the brain and are essential for maintaining homeostasis in a neuronal microenvironment. Currently, a genetic tool to modify microglial gene expression in specific brain regions is not available. In this report, we introduce a tailor-designed method that uses lipid and polymer hybridized nanoparticles (LPNPs) for the local delivery of small interfering RNAs (siRNAs), allowing the silencing of specific microglial genes in the hypothalamus. Our physical characterization proved that this LPNP-siRNA was uniform and stable. We demonstrated that, due to their natural phagocytic behavior, microglial cells are the dominant cell type taking up these LPNPs in the hypothalamus of rats. We then tested the silencing efficiency of LPNPs carrying a cluster of differentiation molecule 11b (CD11b) or Toll-like receptor 4 (TLR4) siRNA using different in vivo and in vitro approaches. In cultured microglial cells treated with LPNP-CD11b siRNA or LPNP-TLR4 siRNA, we found a silencing efficiency at protein expression levels of 65 or 77%, respectively. In line with this finding, immunohistochemistry and western blotting results from in vivo experiments showed that LPNP-CD11b siRNA significantly inhibited microglial CD11b protein expression in the hypothalamus. Furthermore, following lipopolysaccharide (LPS) stimulation of cultured microglial cells, gene expression of the TLR4 downstream signaling component myeloid differentiation factor 88 and its associated cytokines was significantly inhibited in LPNP-TLR4 siRNA-treated microglial cells compared with cells treated with LPNP-scrambled siRNA. Finally, after LPNP-TLR4 siRNA injection into the rat hypothalamus, we observed a significant reduction in microglial activation in response to LPS compared with the control rats injected with LPNP-scrambled siRNA. Our results indicate that LPNP-siRNA is a promising tool to manipulate microglial activity locally in the brain and may serve as a prophylactic approach to prevent microglial dysfunction-associated diseases.
KW - CD11b
KW - TLR4
KW - hypothalamus
KW - microglia
KW - nanoparticles
KW - phagocytosis
KW - siRNA
UR - http://www.scopus.com/inward/record.url?scp=85123947377&partnerID=8YFLogxK
U2 - https://doi.org/10.1021/acsami.1c22434
DO - https://doi.org/10.1021/acsami.1c22434
M3 - Article
C2 - 35041392
SN - 1944-8244
VL - 14
SP - 5066
EP - 5079
JO - ACS applied materials & interfaces
JF - ACS applied materials & interfaces
IS - 4
ER -