PMC:7025468 / 36712-41669 JSONTXT

{"project":"TEST0","denotations":[{"id":"32117283-59-64-3683930","span":{"begin":70,"end":71},"obj":"[\"18458093\"]"},{"id":"32117283-62-68-3683931","span":{"begin":73,"end":75},"obj":"[\"20932494\"]"},{"id":"32117283-71-77-3683932","span":{"begin":275,"end":277},"obj":"[\"23518444\", \"25678251\", \"20953917\"]"},{"id":"32117283-110-116-3683933","span":{"begin":393,"end":395},"obj":"[\"16884330\"]"},{"id":"32117283-114-120-3683934","span":{"begin":397,"end":399},"obj":"[\"1922954\"]"},{"id":"32117283-105-110-3683935","span":{"begin":507,"end":508},"obj":"[\"14673760\", \"18458095\", \"18458093\"]"},{"id":"32117283-110-115-3683936","span":{"begin":512,"end":513},"obj":"[\"18923691\"]"},{"id":"32117283-113-119-3683937","span":{"begin":515,"end":517},"obj":"[\"16314451\"]"},{"id":"32117283-117-123-3683938","span":{"begin":519,"end":521},"obj":"[\"22541426\"]"},{"id":"32117283-121-127-3683939","span":{"begin":523,"end":525},"obj":"[\"18204468\"]"},{"id":"32117283-80-86-3683940","span":{"begin":608,"end":610},"obj":"[\"22541426\"]"},{"id":"32117283-84-90-3683941","span":{"begin":612,"end":614},"obj":"[\"30150282\"]"},{"id":"32117283-141-147-3683942","span":{"begin":758,"end":760},"obj":"[\"25896300\"]"},{"id":"32117283-145-151-3683943","span":{"begin":762,"end":764},"obj":"[\"8413223\"]"},{"id":"32117283-79-85-3683944","span":{"begin":846,"end":848},"obj":"[\"22140520\"]"},{"id":"32117283-83-89-3683945","span":{"begin":850,"end":852},"obj":"[\"17890055\"]"},{"id":"32117283-87-93-3683946","span":{"begin":854,"end":856},"obj":"[\"25705792\"]"},{"id":"32117283-109-115-3683947","span":{"begin":968,"end":970},"obj":"[\"22453030\"]"},{"id":"32117283-113-119-3683948","span":{"begin":972,"end":974},"obj":"[\"23685241\"]"},{"id":"32117283-117-123-3683949","span":{"begin":976,"end":978},"obj":"[\"17890055\"]"},{"id":"32117283-121-127-3683950","span":{"begin":980,"end":982},"obj":"[\"16921377\"]"},{"id":"32117283-168-174-3683951","span":{"begin":1153,"end":1155},"obj":"[\"24939850\"]"},{"id":"32117283-172-178-3683952","span":{"begin":1157,"end":1159},"obj":"[\"20181935\"]"},{"id":"32117283-179-185-3683953","span":{"begin":3327,"end":3329},"obj":"[\"19148178\", \"27832707\", \"29611111\"]"},{"id":"32117283-101-107-3683954","span":{"begin":3573,"end":3575},"obj":"[\"22140520\"]"},{"id":"32117283-101-107-3683955","span":{"begin":3853,"end":3855},"obj":"[\"22140520\"]"},{"id":"32117283-105-111-3683956","span":{"begin":3857,"end":3859},"obj":"[\"25472883\"]"}],"text":"Discussion\nMeth abuse is a well-established risk factor for HIV/AIDS (5, 57). The drug can enhance virus replication and clinical progression, often promoting poor adherence to anti-retroviral therapies. Meth has also been shown to modulate inflammatory cytokine expression (10–12). Specifically, Meth has been reported to enhance IL-1β expression in dendritic cells and the rat hypothalamus (13, 14). The effects of Meth on host defenses and the miRNAs that regulate these processes, are not well-defined (3–5, 9, 18, 23, 58). In addition, miRNA targets are highly sensitive to changes in their expression (23, 24). IL-1β has been shown to participate in an auto-regulatory loop, and stimulate NFκB-dependent miR-146a to disrupt key inflammatory responses (31, 43). Several direct targets of miR-146a have been implicated in HIV-1 pathobiology (47, 48, 59). Notably, miR-146a targets both TRAF6 and IRAK1, signaling molecules that facilitate innate immune responses (26, 27, 48, 60). HIV-1 can induce expression of IL-1β, which is associated with progression of HIV/AIDS, and in microglial cells HIV-1 infection resulted in overexpression of miR-146a (16, 61).\nHere, we demonstrate that Meth can induce expression of IL-1β and miR-146a in CD4+ T-cells, with overexpression of the cytokine leading to increased expression of miR-146a. Meth mediated miR-146a overexpression targeted TRAF6 to modulate innate immune signaling pathways. Based on our findings, we hypothesized that IL-1β signaling results in increased IL-1β mRNA levels and increased miR-146a expression. We further hypothesized that IL-1β-dependent induction of miR-146a would target key innate immune pathways by decreasing TRAF6 expression (Figure S2). Although there are several known targets of miR-146a, only TRAF6 was significantly affected during Meth treatment. We also observed that induction of an IL-1β auto-regulatory loop contributed to Meth mediated increases in HIV-1 replication.\nMeth treatment of CD4+ T-cells augmented Caspase-1 activation and enhanced IL-1β release. Subsequently, both IL-1β mRNA and miR-146a levels rose, indicating that IL-1β increased expression of these transcripts. We also explored changes in TRAF6 and IRAK1 proteins, both direct targets of miR-146a. While we found no change in IRAK1 protein expression, there was decreased TRAF6 expression, indicating selective inhibition of TRAF6 by Meth via miR-146a.\nTo further elucidate the role of Meth in IL-1β and miR-146a overexpression, we blocked IL-1β signaling at two steps. Cells were treated with either IL-1RA to block binding of IL-1β to its receptor, or exogenous IFNα, a type I IFN known to inhibit Caspase-1 activation and antagonize mature IL-1β release. We observed decreased IL-1β release after exogenous IFNα treatment, along with decreased Caspase-1 activation. Blocking either the IL-1 receptor with IL-1RA, or release of IL-1β with IFNα, abrogated Meth mediated effects on miR-146a and IL-1β mRNA. Our data demonstrate that Caspase-1 activation, and increased IL-1β release and signaling are critical for Meth mediated miR-146a overexpression and enhanced IL-1β mRNA expression. Notably, Caspase-1 and IL-1β are associated with pryoptosis, or inflammation mediated apoptosis; pryoptosis is linked to progression of HIV-1 infection via CD4+ T-cell depletion (62–64).\nAt day 1 of HIV-1 infection, we observed enhanced release of IL-1β, followed by increased IL-1β mRNA and miR-146a expression at day two. Decreased TRAF6 expression, a known miR-146a target, is associated with increased HIV-1 replication (47). We observed decreased TRAF6 expression upon infection with HIV-1 at the gene expression level. IRAK1 protein expression levels were also decreased following HIV-1 infection. These findings are consistent with previous reports that HIV-1 decreases IRAK1 and TRAF6 expression (47, 55).\nIn sum, we found that Meth enhanced HIV-1 replication through an IL-1β positive auto-regulatory loop. By blocking IL-1 signaling using IL-1RA, we observed dose dependent decreased HIV-1 replication after Meth treatment. We also observed that Meth enhanced IL-1β mRNA levels were abrogated in a dose dependent manner upon treatment with IL-1RA. These results support the involvement of an IL-1β auto-regulatory loop in Meth mediated enhanced HIV-1 replication. Alternatively, when HIV-1 replication was inhibited using IFNα, we found decreased IL-1β and miR-146a expression. Our results indicate that increased levels of IL-1β directly contribute to Meth mediated increased HIV-1 replication in CD4+ T-cells. Based on our results, it appears that Meth mediated increased IL-1β expression acts to prime cells to be more susceptible to infection with HIV-1.\nThese studies unveil unique effects of Meth on IL-1β to dysregulate innate immune signaling pathways and enhance HIV-1 infection. This novel mechanism of action of Meth points to potential therapeutic targets relevant to drug abusing hosts."}

{"project":"2_test","denotations":[{"id":"32117283-18458093-35221736","span":{"begin":70,"end":71},"obj":"18458093"},{"id":"32117283-20932494-35221737","span":{"begin":73,"end":75},"obj":"20932494"},{"id":"32117283-23518444-35221738","span":{"begin":275,"end":277},"obj":"23518444"},{"id":"32117283-25678251-35221738","span":{"begin":275,"end":277},"obj":"25678251"},{"id":"32117283-20953917-35221738","span":{"begin":275,"end":277},"obj":"20953917"},{"id":"32117283-16884330-35221739","span":{"begin":393,"end":395},"obj":"16884330"},{"id":"32117283-1922954-35221740","span":{"begin":397,"end":399},"obj":"1922954"},{"id":"32117283-14673760-35221741","span":{"begin":507,"end":508},"obj":"14673760"},{"id":"32117283-18458095-35221741","span":{"begin":507,"end":508},"obj":"18458095"},{"id":"32117283-18458093-35221741","span":{"begin":507,"end":508},"obj":"18458093"},{"id":"32117283-18923691-35221742","span":{"begin":512,"end":513},"obj":"18923691"},{"id":"32117283-16314451-35221743","span":{"begin":515,"end":517},"obj":"16314451"},{"id":"32117283-22541426-35221744","span":{"begin":519,"end":521},"obj":"22541426"},{"id":"32117283-18204468-35221745","span":{"begin":523,"end":525},"obj":"18204468"},{"id":"32117283-22541426-35221746","span":{"begin":608,"end":610},"obj":"22541426"},{"id":"32117283-30150282-35221747","span":{"begin":612,"end":614},"obj":"30150282"},{"id":"32117283-25896300-35221748","span":{"begin":758,"end":760},"obj":"25896300"},{"id":"32117283-8413223-35221749","span":{"begin":762,"end":764},"obj":"8413223"},{"id":"32117283-22140520-35221750","span":{"begin":846,"end":848},"obj":"22140520"},{"id":"32117283-17890055-35221751","span":{"begin":850,"end":852},"obj":"17890055"},{"id":"32117283-25705792-35221752","span":{"begin":854,"end":856},"obj":"25705792"},{"id":"32117283-22453030-35221753","span":{"begin":968,"end":970},"obj":"22453030"},{"id":"32117283-23685241-35221754","span":{"begin":972,"end":974},"obj":"23685241"},{"id":"32117283-17890055-35221755","span":{"begin":976,"end":978},"obj":"17890055"},{"id":"32117283-16921377-35221756","span":{"begin":980,"end":982},"obj":"16921377"},{"id":"32117283-24939850-35221757","span":{"begin":1153,"end":1155},"obj":"24939850"},{"id":"32117283-20181935-35221758","span":{"begin":1157,"end":1159},"obj":"20181935"},{"id":"32117283-19148178-35221759","span":{"begin":3327,"end":3329},"obj":"19148178"},{"id":"32117283-27832707-35221759","span":{"begin":3327,"end":3329},"obj":"27832707"},{"id":"32117283-29611111-35221759","span":{"begin":3327,"end":3329},"obj":"29611111"},{"id":"32117283-22140520-35221760","span":{"begin":3573,"end":3575},"obj":"22140520"},{"id":"32117283-22140520-35221761","span":{"begin":3853,"end":3855},"obj":"22140520"},{"id":"32117283-25472883-35221762","span":{"begin":3857,"end":3859},"obj":"25472883"}],"text":"Discussion\nMeth abuse is a well-established risk factor for HIV/AIDS (5, 57). The drug can enhance virus replication and clinical progression, often promoting poor adherence to anti-retroviral therapies. Meth has also been shown to modulate inflammatory cytokine expression (10–12). Specifically, Meth has been reported to enhance IL-1β expression in dendritic cells and the rat hypothalamus (13, 14). The effects of Meth on host defenses and the miRNAs that regulate these processes, are not well-defined (3–5, 9, 18, 23, 58). In addition, miRNA targets are highly sensitive to changes in their expression (23, 24). IL-1β has been shown to participate in an auto-regulatory loop, and stimulate NFκB-dependent miR-146a to disrupt key inflammatory responses (31, 43). Several direct targets of miR-146a have been implicated in HIV-1 pathobiology (47, 48, 59). Notably, miR-146a targets both TRAF6 and IRAK1, signaling molecules that facilitate innate immune responses (26, 27, 48, 60). HIV-1 can induce expression of IL-1β, which is associated with progression of HIV/AIDS, and in microglial cells HIV-1 infection resulted in overexpression of miR-146a (16, 61).\nHere, we demonstrate that Meth can induce expression of IL-1β and miR-146a in CD4+ T-cells, with overexpression of the cytokine leading to increased expression of miR-146a. Meth mediated miR-146a overexpression targeted TRAF6 to modulate innate immune signaling pathways. Based on our findings, we hypothesized that IL-1β signaling results in increased IL-1β mRNA levels and increased miR-146a expression. We further hypothesized that IL-1β-dependent induction of miR-146a would target key innate immune pathways by decreasing TRAF6 expression (Figure S2). Although there are several known targets of miR-146a, only TRAF6 was significantly affected during Meth treatment. We also observed that induction of an IL-1β auto-regulatory loop contributed to Meth mediated increases in HIV-1 replication.\nMeth treatment of CD4+ T-cells augmented Caspase-1 activation and enhanced IL-1β release. Subsequently, both IL-1β mRNA and miR-146a levels rose, indicating that IL-1β increased expression of these transcripts. We also explored changes in TRAF6 and IRAK1 proteins, both direct targets of miR-146a. While we found no change in IRAK1 protein expression, there was decreased TRAF6 expression, indicating selective inhibition of TRAF6 by Meth via miR-146a.\nTo further elucidate the role of Meth in IL-1β and miR-146a overexpression, we blocked IL-1β signaling at two steps. Cells were treated with either IL-1RA to block binding of IL-1β to its receptor, or exogenous IFNα, a type I IFN known to inhibit Caspase-1 activation and antagonize mature IL-1β release. We observed decreased IL-1β release after exogenous IFNα treatment, along with decreased Caspase-1 activation. Blocking either the IL-1 receptor with IL-1RA, or release of IL-1β with IFNα, abrogated Meth mediated effects on miR-146a and IL-1β mRNA. Our data demonstrate that Caspase-1 activation, and increased IL-1β release and signaling are critical for Meth mediated miR-146a overexpression and enhanced IL-1β mRNA expression. Notably, Caspase-1 and IL-1β are associated with pryoptosis, or inflammation mediated apoptosis; pryoptosis is linked to progression of HIV-1 infection via CD4+ T-cell depletion (62–64).\nAt day 1 of HIV-1 infection, we observed enhanced release of IL-1β, followed by increased IL-1β mRNA and miR-146a expression at day two. Decreased TRAF6 expression, a known miR-146a target, is associated with increased HIV-1 replication (47). We observed decreased TRAF6 expression upon infection with HIV-1 at the gene expression level. IRAK1 protein expression levels were also decreased following HIV-1 infection. These findings are consistent with previous reports that HIV-1 decreases IRAK1 and TRAF6 expression (47, 55).\nIn sum, we found that Meth enhanced HIV-1 replication through an IL-1β positive auto-regulatory loop. By blocking IL-1 signaling using IL-1RA, we observed dose dependent decreased HIV-1 replication after Meth treatment. We also observed that Meth enhanced IL-1β mRNA levels were abrogated in a dose dependent manner upon treatment with IL-1RA. These results support the involvement of an IL-1β auto-regulatory loop in Meth mediated enhanced HIV-1 replication. Alternatively, when HIV-1 replication was inhibited using IFNα, we found decreased IL-1β and miR-146a expression. Our results indicate that increased levels of IL-1β directly contribute to Meth mediated increased HIV-1 replication in CD4+ T-cells. Based on our results, it appears that Meth mediated increased IL-1β expression acts to prime cells to be more susceptible to infection with HIV-1.\nThese studies unveil unique effects of Meth on IL-1β to dysregulate innate immune signaling pathways and enhance HIV-1 infection. This novel mechanism of action of Meth points to potential therapeutic targets relevant to drug abusing hosts."}

{"project":"MyTest","denotations":[{"id":"32117283-18458093-35221736","span":{"begin":70,"end":71},"obj":"18458093"},{"id":"32117283-20932494-35221737","span":{"begin":73,"end":75},"obj":"20932494"},{"id":"32117283-23518444-35221738","span":{"begin":275,"end":277},"obj":"23518444"},{"id":"32117283-25678251-35221738","span":{"begin":275,"end":277},"obj":"25678251"},{"id":"32117283-20953917-35221738","span":{"begin":275,"end":277},"obj":"20953917"},{"id":"32117283-16884330-35221739","span":{"begin":393,"end":395},"obj":"16884330"},{"id":"32117283-1922954-35221740","span":{"begin":397,"end":399},"obj":"1922954"},{"id":"32117283-14673760-35221741","span":{"begin":507,"end":508},"obj":"14673760"},{"id":"32117283-18458095-35221741","span":{"begin":507,"end":508},"obj":"18458095"},{"id":"32117283-18458093-35221741","span":{"begin":507,"end":508},"obj":"18458093"},{"id":"32117283-18923691-35221742","span":{"begin":512,"end":513},"obj":"18923691"},{"id":"32117283-16314451-35221743","span":{"begin":515,"end":517},"obj":"16314451"},{"id":"32117283-22541426-35221744","span":{"begin":519,"end":521},"obj":"22541426"},{"id":"32117283-18204468-35221745","span":{"begin":523,"end":525},"obj":"18204468"},{"id":"32117283-22541426-35221746","span":{"begin":608,"end":610},"obj":"22541426"},{"id":"32117283-30150282-35221747","span":{"begin":612,"end":614},"obj":"30150282"},{"id":"32117283-25896300-35221748","span":{"begin":758,"end":760},"obj":"25896300"},{"id":"32117283-8413223-35221749","span":{"begin":762,"end":764},"obj":"8413223"},{"id":"32117283-22140520-35221750","span":{"begin":846,"end":848},"obj":"22140520"},{"id":"32117283-17890055-35221751","span":{"begin":850,"end":852},"obj":"17890055"},{"id":"32117283-25705792-35221752","span":{"begin":854,"end":856},"obj":"25705792"},{"id":"32117283-22453030-35221753","span":{"begin":968,"end":970},"obj":"22453030"},{"id":"32117283-23685241-35221754","span":{"begin":972,"end":974},"obj":"23685241"},{"id":"32117283-17890055-35221755","span":{"begin":976,"end":978},"obj":"17890055"},{"id":"32117283-16921377-35221756","span":{"begin":980,"end":982},"obj":"16921377"},{"id":"32117283-24939850-35221757","span":{"begin":1153,"end":1155},"obj":"24939850"},{"id":"32117283-20181935-35221758","span":{"begin":1157,"end":1159},"obj":"20181935"},{"id":"32117283-19148178-35221759","span":{"begin":3327,"end":3329},"obj":"19148178"},{"id":"32117283-27832707-35221759","span":{"begin":3327,"end":3329},"obj":"27832707"},{"id":"32117283-29611111-35221759","span":{"begin":3327,"end":3329},"obj":"29611111"},{"id":"32117283-22140520-35221760","span":{"begin":3573,"end":3575},"obj":"22140520"},{"id":"32117283-22140520-35221761","span":{"begin":3853,"end":3855},"obj":"22140520"},{"id":"32117283-25472883-35221762","span":{"begin":3857,"end":3859},"obj":"25472883"}],"namespaces":[{"prefix":"_base","uri":"https://www.uniprot.org/uniprot/testbase"},{"prefix":"UniProtKB","uri":"https://www.uniprot.org/uniprot/"},{"prefix":"uniprot","uri":"https://www.uniprot.org/uniprotkb/"}],"text":"Discussion\nMeth abuse is a well-established risk factor for HIV/AIDS (5, 57). The drug can enhance virus replication and clinical progression, often promoting poor adherence to anti-retroviral therapies. Meth has also been shown to modulate inflammatory cytokine expression (10–12). Specifically, Meth has been reported to enhance IL-1β expression in dendritic cells and the rat hypothalamus (13, 14). The effects of Meth on host defenses and the miRNAs that regulate these processes, are not well-defined (3–5, 9, 18, 23, 58). In addition, miRNA targets are highly sensitive to changes in their expression (23, 24). IL-1β has been shown to participate in an auto-regulatory loop, and stimulate NFκB-dependent miR-146a to disrupt key inflammatory responses (31, 43). Several direct targets of miR-146a have been implicated in HIV-1 pathobiology (47, 48, 59). Notably, miR-146a targets both TRAF6 and IRAK1, signaling molecules that facilitate innate immune responses (26, 27, 48, 60). HIV-1 can induce expression of IL-1β, which is associated with progression of HIV/AIDS, and in microglial cells HIV-1 infection resulted in overexpression of miR-146a (16, 61).\nHere, we demonstrate that Meth can induce expression of IL-1β and miR-146a in CD4+ T-cells, with overexpression of the cytokine leading to increased expression of miR-146a. Meth mediated miR-146a overexpression targeted TRAF6 to modulate innate immune signaling pathways. Based on our findings, we hypothesized that IL-1β signaling results in increased IL-1β mRNA levels and increased miR-146a expression. We further hypothesized that IL-1β-dependent induction of miR-146a would target key innate immune pathways by decreasing TRAF6 expression (Figure S2). Although there are several known targets of miR-146a, only TRAF6 was significantly affected during Meth treatment. We also observed that induction of an IL-1β auto-regulatory loop contributed to Meth mediated increases in HIV-1 replication.\nMeth treatment of CD4+ T-cells augmented Caspase-1 activation and enhanced IL-1β release. Subsequently, both IL-1β mRNA and miR-146a levels rose, indicating that IL-1β increased expression of these transcripts. We also explored changes in TRAF6 and IRAK1 proteins, both direct targets of miR-146a. While we found no change in IRAK1 protein expression, there was decreased TRAF6 expression, indicating selective inhibition of TRAF6 by Meth via miR-146a.\nTo further elucidate the role of Meth in IL-1β and miR-146a overexpression, we blocked IL-1β signaling at two steps. Cells were treated with either IL-1RA to block binding of IL-1β to its receptor, or exogenous IFNα, a type I IFN known to inhibit Caspase-1 activation and antagonize mature IL-1β release. We observed decreased IL-1β release after exogenous IFNα treatment, along with decreased Caspase-1 activation. Blocking either the IL-1 receptor with IL-1RA, or release of IL-1β with IFNα, abrogated Meth mediated effects on miR-146a and IL-1β mRNA. Our data demonstrate that Caspase-1 activation, and increased IL-1β release and signaling are critical for Meth mediated miR-146a overexpression and enhanced IL-1β mRNA expression. Notably, Caspase-1 and IL-1β are associated with pryoptosis, or inflammation mediated apoptosis; pryoptosis is linked to progression of HIV-1 infection via CD4+ T-cell depletion (62–64).\nAt day 1 of HIV-1 infection, we observed enhanced release of IL-1β, followed by increased IL-1β mRNA and miR-146a expression at day two. Decreased TRAF6 expression, a known miR-146a target, is associated with increased HIV-1 replication (47). We observed decreased TRAF6 expression upon infection with HIV-1 at the gene expression level. IRAK1 protein expression levels were also decreased following HIV-1 infection. These findings are consistent with previous reports that HIV-1 decreases IRAK1 and TRAF6 expression (47, 55).\nIn sum, we found that Meth enhanced HIV-1 replication through an IL-1β positive auto-regulatory loop. By blocking IL-1 signaling using IL-1RA, we observed dose dependent decreased HIV-1 replication after Meth treatment. We also observed that Meth enhanced IL-1β mRNA levels were abrogated in a dose dependent manner upon treatment with IL-1RA. These results support the involvement of an IL-1β auto-regulatory loop in Meth mediated enhanced HIV-1 replication. Alternatively, when HIV-1 replication was inhibited using IFNα, we found decreased IL-1β and miR-146a expression. Our results indicate that increased levels of IL-1β directly contribute to Meth mediated increased HIV-1 replication in CD4+ T-cells. Based on our results, it appears that Meth mediated increased IL-1β expression acts to prime cells to be more susceptible to infection with HIV-1.\nThese studies unveil unique effects of Meth on IL-1β to dysregulate innate immune signaling pathways and enhance HIV-1 infection. This novel mechanism of action of Meth points to potential therapeutic targets relevant to drug abusing hosts."}