ssRNA40 sodium (R-1075 sodium) is a single-stranded RNA40 derived from HIV-1. ssRNA40 sodium activates the TLR7, TLR8, TLR2, RIG-I, MDA5, MyD88, Caspase-3, IRE1α, NLRP3 inflammasome and IRF7 signaling pathways. ssRNA40 sodium alters mRNA expression in neutrophils, induces pro-inflammatory cytokines, ROS, autophagy ( autophagy ), pyroptosis ( pyroptosis ), neuronal death, neurodegeneration, aggregate formation and NK cell activation. ssRNA40 sodium activates the expression of CD62L, CD11b, CD69, MX1, OAS1, ATG7, LC3B and XBP1 in immune cell and neuronal populations. ssRNA40 sodium causes cortical neuron loss and axonal damage in mice in a TLR7 -dependent manner. ssRNA40 sodium can be used in research on HIV-1 infection, neurodegeneration, COVID-19 and HIV-associated neurological disorders [1] [2] [3] [4] [5] [6].
In Vivo, ssRNA40 sodium (10 µg/mL; intratracheal administration; single injection) induces approximately 20% loss of cortical neurons and axonal damage in wild-type mice in a TLR7-dependent manner[2].
In Vitro, ssRNA40 sodium (3 ng/μL; 3 h) downregulates the mRNA expression level of TLR2, upregulates the mRNA expression levels of TLR7, TLR8, RIG-I and MDA5, but has no effect on the mRNA expression level of TLR4 in purified human neutrophils[1]. Co-administration of ssRNA40 sodium (3 ng/μL; 3 h) with TLR agonists significantly upregulates the mRNA expression of TLR2, TLR4, TLR7, TLR8, RIG-I and MDA5 in purified human neutrophils from healthy donors in a TLR agonist-dependent manner[1]. ssRNA40 sodium (3 ng/μL; 8 h) significantly reduces the expression level of CD62L, promotes the secretion of IL-6 and TNF-α, and increases the production of ROS in purified human neutrophils, whereas it exerts no significant effect on the expression of CD11b[1]. ssRNA40 sodium (0-20 μg/mL; 0-6 days) induces dose-dependent and time-dependent cell death in primary cortical neurons from wild-type mice in a TLR7-dependent, cell-autonomous, and MyD88-dependent manner (treatment with 10 μg/mL for 4 days reduces cell viability by approximately 30%), whereas primary cortical neurons from TLR7-knockout mice are completely protected[2]. ssRNA40 sodium (10 μg/mL; 36 h) induces cell death in N1E-115 mouse neuroblastoma cells[2]. ssRNA40 sodium (10 μg/mL; 3 days) activates caspase-3 in primary mouse cortical neurons, and inhibition of caspase-3 blocks ssRNA40-induced neuronal cell death[2]. ssRNA40 sodium (1-25 μg/mL; 6-24 h) induces dose-dependent and time-dependent secretion of TNF-α, IL-6, MCP-1 and RANTES (but not IL-10) from primary microglia of wild-type mice in a TLR7-dependent manner, whereas primary microglia from TLR7-knockout mice do not secrete these cytokines upon ssRNA40 stimulation[2]. ssRNA40 sodium (2 μg/mL; 1-24 h) activates the TLR8/IRE1α-XBP1 pathway in human MDDCs, induces XBP1 splicing and sXBP1 promoter binding, and significantly promotes the expression of proinflammatory cytokines (IL-1β, IL-6, TNFα) at both mRNA and protein levels, with all these effects dependent on the activities of TLR8 and IRE1α[3]. ssRNA40 sodium (6 h) activates CD3−CD56+/− NK cells in HIV-1-negative peripheral blood mononuclear cells (PBMCs); when ssRNA40 is complexed with Dotap, the activation effect is significantly enhanced, with the median percentage of CD69+ NK cells reaching 28.8% after 6 h of stimulation[4]. ssRNA40 sodium (5 μg/mL; 24-48 h) activates the NLRP3 inflammasome in human primary monocyte-derived microglia (HMG), induces the substantial expression and secretion of pro-inflammatory cytokines (IL-1β, IL-18) and neurotoxic cytokines (C1q, IL-1α, TNF-α), and the release of IL-1β depends on the activities of NLRP3 and caspase-1[5]. Supernatant from HMG activated by ssRNA40 sodium (5 μg/mL; 48 h) induces significant neurotoxicity in human primary neurons (HPN), including reduced cell viability and neurite damage[5]. ssRNA40 sodium (5 μg/mL; 24 h) induces NLRP3-dependent mitochondrial damage in HMG, including increased ROS production and loss of mitochondrial membrane potential[5]. ssRNA40 sodium (5 μg/mL; 24 h) upregulates the expression of autophagy/mitophagy receptors (SQSTM1, OPTN, PINK1, Parkin) and induces their mitochondrial recruitment in HMG and HFMG[5]. ssRNA40 sodium (5 μg/mL; 24-48 h) impairs autophagic flux in HMG, which in turn leads to persistent activation of the NLRP3 inflammasome, activation of caspase-1, and pyroptosis[5]. ssRNA40 sodium (2 μg/mL; 16 h) induces robust IFN-α production in primary human plasmacytoid dendritic cells (pDC), and this response is significantly dependent on the TLR7 signaling pathway and autophagy; inhibition of either pathway drastically reduces IFN-α secretion[6]. ssRNA40 sodium (2 μg/mL; 16 h) upregulates the expression of autophagy-related proteins (ATG7 and LC3-II) and induces the formation of a large number of autophagosomes in primary human pDCs[6]. ssRNA40 sodium (2 μg/mL; 12 h) induces IRF7 activation (nuclear translocation and phosphorylation) in primary human pDCs and promotes the co-expression of activated IRF7 with autophagic LC3B puncta; inhibition of autophagy or the mTOR signaling pathway attenuates this activation[6].
References:
[1]. Giraldo DM, et al. HIV-1-derived single-stranded RNA acts as activator of human neutrophils. Immunol Res. 2016;64(5-6):1185-1194.
[2]. Lehmann SM, et al. Extracellularly delivered single-stranded viral RNA causes neurodegeneration dependent on TLR7. J Immunol. 2012;189(3):1448-1458.
[3]. Fernández JJ, et al. Innate IRE1α-XBP1 activation by viral single-stranded RNA and its influence on lung cytokine production during SARS-CoV-2 pneumonia. Genes Immun. 2024;25(1):43-54.
[4]. Alter G, et al. Single-stranded RNA derived from HIV-1 serves as a potent activator of NK cells. J Immunol. 2007 Jun 15;178(12):7658-66.
[5]. Rawat P, et al. Human immunodeficiency virus Type-1 single-stranded RNA activates the NLRP3 inflammasome and impairs autophagic clearance of damaged mitochondria in human microglia. Glia. 2019;67(5):802-824.
[6]. Zhou D, et al. Production of interferon α by human immunodeficiency virus type 1 in human plasmacytoid dendritic cells is dependent on induction of autophagy. J Infect Dis. 2012;205(8):1258-1267.
In Vivo, ssRNA40 sodium (10 µg/mL; intratracheal administration; single injection) induces approximately 20% loss of cortical neurons and axonal damage in wild-type mice in a TLR7-dependent manner[2].
In Vitro, ssRNA40 sodium (3 ng/μL; 3 h) downregulates the mRNA expression level of TLR2, upregulates the mRNA expression levels of TLR7, TLR8, RIG-I and MDA5, but has no effect on the mRNA expression level of TLR4 in purified human neutrophils[1]. Co-administration of ssRNA40 sodium (3 ng/μL; 3 h) with TLR agonists significantly upregulates the mRNA expression of TLR2, TLR4, TLR7, TLR8, RIG-I and MDA5 in purified human neutrophils from healthy donors in a TLR agonist-dependent manner[1]. ssRNA40 sodium (3 ng/μL; 8 h) significantly reduces the expression level of CD62L, promotes the secretion of IL-6 and TNF-α, and increases the production of ROS in purified human neutrophils, whereas it exerts no significant effect on the expression of CD11b[1]. ssRNA40 sodium (0-20 μg/mL; 0-6 days) induces dose-dependent and time-dependent cell death in primary cortical neurons from wild-type mice in a TLR7-dependent, cell-autonomous, and MyD88-dependent manner (treatment with 10 μg/mL for 4 days reduces cell viability by approximately 30%), whereas primary cortical neurons from TLR7-knockout mice are completely protected[2]. ssRNA40 sodium (10 μg/mL; 36 h) induces cell death in N1E-115 mouse neuroblastoma cells[2]. ssRNA40 sodium (10 μg/mL; 3 days) activates caspase-3 in primary mouse cortical neurons, and inhibition of caspase-3 blocks ssRNA40-induced neuronal cell death[2]. ssRNA40 sodium (1-25 μg/mL; 6-24 h) induces dose-dependent and time-dependent secretion of TNF-α, IL-6, MCP-1 and RANTES (but not IL-10) from primary microglia of wild-type mice in a TLR7-dependent manner, whereas primary microglia from TLR7-knockout mice do not secrete these cytokines upon ssRNA40 stimulation[2]. ssRNA40 sodium (2 μg/mL; 1-24 h) activates the TLR8/IRE1α-XBP1 pathway in human MDDCs, induces XBP1 splicing and sXBP1 promoter binding, and significantly promotes the expression of proinflammatory cytokines (IL-1β, IL-6, TNFα) at both mRNA and protein levels, with all these effects dependent on the activities of TLR8 and IRE1α[3]. ssRNA40 sodium (6 h) activates CD3−CD56+/− NK cells in HIV-1-negative peripheral blood mononuclear cells (PBMCs); when ssRNA40 is complexed with Dotap, the activation effect is significantly enhanced, with the median percentage of CD69+ NK cells reaching 28.8% after 6 h of stimulation[4]. ssRNA40 sodium (5 μg/mL; 24-48 h) activates the NLRP3 inflammasome in human primary monocyte-derived microglia (HMG), induces the substantial expression and secretion of pro-inflammatory cytokines (IL-1β, IL-18) and neurotoxic cytokines (C1q, IL-1α, TNF-α), and the release of IL-1β depends on the activities of NLRP3 and caspase-1[5]. Supernatant from HMG activated by ssRNA40 sodium (5 μg/mL; 48 h) induces significant neurotoxicity in human primary neurons (HPN), including reduced cell viability and neurite damage[5]. ssRNA40 sodium (5 μg/mL; 24 h) induces NLRP3-dependent mitochondrial damage in HMG, including increased ROS production and loss of mitochondrial membrane potential[5]. ssRNA40 sodium (5 μg/mL; 24 h) upregulates the expression of autophagy/mitophagy receptors (SQSTM1, OPTN, PINK1, Parkin) and induces their mitochondrial recruitment in HMG and HFMG[5]. ssRNA40 sodium (5 μg/mL; 24-48 h) impairs autophagic flux in HMG, which in turn leads to persistent activation of the NLRP3 inflammasome, activation of caspase-1, and pyroptosis[5]. ssRNA40 sodium (2 μg/mL; 16 h) induces robust IFN-α production in primary human plasmacytoid dendritic cells (pDC), and this response is significantly dependent on the TLR7 signaling pathway and autophagy; inhibition of either pathway drastically reduces IFN-α secretion[6]. ssRNA40 sodium (2 μg/mL; 16 h) upregulates the expression of autophagy-related proteins (ATG7 and LC3-II) and induces the formation of a large number of autophagosomes in primary human pDCs[6]. ssRNA40 sodium (2 μg/mL; 12 h) induces IRF7 activation (nuclear translocation and phosphorylation) in primary human pDCs and promotes the co-expression of activated IRF7 with autophagic LC3B puncta; inhibition of autophagy or the mTOR signaling pathway attenuates this activation[6].
References:
[1]. Giraldo DM, et al. HIV-1-derived single-stranded RNA acts as activator of human neutrophils. Immunol Res. 2016;64(5-6):1185-1194.
[2]. Lehmann SM, et al. Extracellularly delivered single-stranded viral RNA causes neurodegeneration dependent on TLR7. J Immunol. 2012;189(3):1448-1458.
[3]. Fernández JJ, et al. Innate IRE1α-XBP1 activation by viral single-stranded RNA and its influence on lung cytokine production during SARS-CoV-2 pneumonia. Genes Immun. 2024;25(1):43-54.
[4]. Alter G, et al. Single-stranded RNA derived from HIV-1 serves as a potent activator of NK cells. J Immunol. 2007 Jun 15;178(12):7658-66.
[5]. Rawat P, et al. Human immunodeficiency virus Type-1 single-stranded RNA activates the NLRP3 inflammasome and impairs autophagic clearance of damaged mitochondria in human microglia. Glia. 2019;67(5):802-824.
[6]. Zhou D, et al. Production of interferon α by human immunodeficiency virus type 1 in human plasmacytoid dendritic cells is dependent on induction of autophagy. J Infect Dis. 2012;205(8):1258-1267.