Publications by authors named "Arlene Lim"

8 Publications

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Inhibition of HSP90 and Activation of HSF1 Diminish Macrophage NLRP3 Inflammasome Activity in Alcohol-Associated Liver Injury.

Alcohol Clin Exp Res 2020 06 18;44(6):1300-1311. Epub 2020 May 18.

From the, Department of Medicine, (AC, DB, AL, PM), University of Massachusetts Medical School, Worcester, Massachusetts.

Background: Activation of NLRP3 in liver macrophages contributes to alcohol-associated liver disease (ALD). Molecular chaperone heat shock protein (HSP) 90 facilitates NLRP3 inflammasome activity during infections and inflammatory diseases. We previously reported that HSP90 is induced in ALD and regulates proinflammatory cytokines, tumor necrosis factor alpha, and IL-6. Whether HSP90 affects IL-1β and IL-18 regulated by NLRP3 inflammasome in ALD is unknown. Here, we hypothesize that HSP90 modulated NLRP3 inflammasome activity and affects IL-1β and IL-18 secretion in ALD.

Methods: The expression of HSP90AA1 and NLRP3 inflammasome genes was evaluated in human alcoholic livers and in mouse model of ALD. The importance of HSP90 on NLRP3 inflammasome activation in ALD was evaluated by administering HSP90 inhibitor, 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG) to mice subjected to ALD, and in vitro to bone marrow-derived macrophages (BMDM) stimulated with LPS and ATP. The effect of activation of HSF1/HSPA1A axis during HSP90 inhibition or direct activation during heat shock of BMDMs on NLRP3 activity and secretion of downstream cytokines was evaluated.

Results: We found positive correlation between induction of HSP90 and NLRP3 inflammasome genes in human alcoholic cirrhotic livers. Administration of 17-DMAG in mouse model of ALD significantly down-regulated NLRP3 inflammasome-mediated caspase-1 (CASP-1) activity and cytokine secretion, with reduction in ALD. 17-DMAG-mediated decrease in NLRP3 was restricted to liver macrophages. Using BMDMs, we show that inhibition of HSP90 prevented CASP-1 activity, and Gasdermin D (GSDMD) cleavage, important in release of active IL-1β and IL-18. Interestingly, activation of the heat shock factor 1 (HSF1)/HSPA1A axis, either during HSP90 inhibition or by heat shock, decreased NLRP3 inflammasome activity and reduced secretion of cytokines.

Conclusion: Our studies indicate that inhibition of HSP90 and activation of HSF1/HSPA1A reduce IL-1β and IL-18 via decrease in NLRP3/CASP-1 and GSDMD activity in ALD.
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http://dx.doi.org/10.1111/acer.14338DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7328660PMC
June 2020

Chronic alcohol-induced liver injury correlates with memory deficits: Role for neuroinflammation.

Alcohol 2020 03 6;83:75-81. Epub 2019 Aug 6.

Department of Medicine, University of Massachusetts Medical School, Worcester, MA, United States. Electronic address:

Alcohol use disorder (AUD) affects over 15 million adults over age 18 in the United States, with estimated costs of 220 billion dollars annually - mainly due to poor quality of life and lost productivity, which in turn is intricately linked to cognitive dysfunction. AUD-induced neuroinflammation in the brain, notably the hippocampus, is likely to contribute to cognitive impairments. The neuroinflammatory mechanisms mediating the impact of chronic alcohol on the central nervous system, specifically cognition, require further study. We hypothesized that chronic alcohol consumption impairs memory and increases the inflammatory cytokines TNFα, IL6, MCP1, and IL1β in the hippocampus and prefrontal cortex regions in the brain. Using the chronic-binge Gao-NIAAA alcohol mouse model of liver disease, representative of the drinking pattern common to human alcoholics, we investigated behavioral and neuroinflammatory parameters. Our data show that chronic alcohol intake elevated peripheral and brain alcohol levels, induced serum alanine aminotransferase (ALT, a marker of liver injury), impaired memory and sensorimotor coordination, and increased inflammatory gene expression in the hippocampus and prefrontal cortex. Interestingly, serum ALT and hippocampal IL6 correlated with memory impairment, suggesting an intrinsic relationship between neuroinflammation, cognitive decline, and liver disease. Overall, our results point to a likely liver-brain functional partnership and suggest that future strategies to alleviate hepatic and/or neuroinflammatory impacts of chronic AUD may result in improved cognitive outcomes.
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http://dx.doi.org/10.1016/j.alcohol.2019.07.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7002207PMC
March 2020

Human Binge Alcohol Intake Inhibits TLR4-MyD88 and TLR4-TRIF Responses but Not the TLR3-TRIF Pathway: HspA1A and PP1 Play Selective Regulatory Roles.

J Immunol 2018 04 14;200(7):2291-2303. Epub 2018 Feb 14.

Department of Medicine, University of Massachusetts Medical School, Worcester, MA 01605

Binge/moderate alcohol suppresses TLR4-MyD88 proinflammatory cytokines; however, alcohol's effects on TLR-TRIF signaling, especially after in vivo exposure in humans, are unclear. We performed a comparative analysis of the TLR4-MyD88, TLR4-TRIF, and TLR3-TRIF pathways in human monocytes following binge alcohol exposure. Mechanistic regulation of TLR-TRIF signaling by binge alcohol was evaluated by analyzing IRF3 and TBK1, upstream regulator protein phosphatase 1 (PP1), and immunoregulatory stress proteins HspA1A and XBP-1 in alcohol-treated human and mouse monocytes/macrophages. Two approaches for alcohol exposure were used: in vivo exposure of primary monocytes in binge alcohol-consuming human volunteers or in vitro exposure of human monocytes/murine macrophages to physiological alcohol concentrations (25-50 mM ethanol), followed by LPS (TLR4) or polyinosinic-polycytidylic acid (TLR3) stimulation ex vivo. In vivo and in vitro binge alcohol exposure significantly inhibited the TLR4-MyD88 cytokines TNF-α and IL-6, as well as the TLR4-TRIF cytokines/chemokines IFN-β, IP-10, and RANTES, in human monocytes, but not TLR3-TRIF-induced cytokines/chemokines, as detected by quantitative PCR and ELISA. Mechanistic analyses revealed TBK-1-independent inhibition of the TLR4-TRIF effector IRF3 in alcohol-treated macrophages. Although stress protein XBP-1, which is known to regulate IRF3-mediated IFN-β induction, was not affected by alcohol, HspA1A was induced by in vivo alcohol in human monocytes. Alcohol-induced HspA1A was required for inhibition of TLR4-MyD88 signaling but not TLR4-TRIF cytokines in macrophages. In contrast, inhibition of PP1 prevented alcohol-mediated TLR4-TRIF tolerance in macrophages. Collectively, our results demonstrate that in vivo and in vitro binge alcohol exposure in humans suppresses TLR4-MyD88 and TLR4-TRIF, but not TLR3-TRIF, responses. Whereas alcohol-mediated effects on the PP1-IRF3 axis inhibit the TLR4-TRIF pathway, HspA1A selectively suppresses the TLR4-MyD88 pathway in monocytes/macrophages.
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http://dx.doi.org/10.4049/jimmunol.1600924DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5860983PMC
April 2018

Inhibition of heat shock protein 90 alleviates steatosis and macrophage activation in murine alcoholic liver injury.

J Hepatol 2014 Oct 22;61(4):903-11. Epub 2014 May 22.

Department of Medicine, University of Massachusetts Medical School, 364 Plantation Street, Worcester, MA 01605, United States. Electronic address:

Background & Aims: Heat shock protein 90 (hsp90) is an emerging therapeutic target in chronic liver diseases. Hsp90 plays an important role in liver immune cell activation; however its role in alcoholic liver disease (ALD) remains elusive. Here we hypothesize that hsp90 is crucial in alcohol induced steatosis and pro-inflammatory cytokine production. To test this hypothesis, we employed a pharmacological inhibitor of hsp90, 17-DMAG (17-Dimethylamino-ethylamino-17-demethoxygeldanamycin) in an in vivo mouse model of acute and chronic alcoholic liver injury.

Methods: C57BL/6 mice were given either a single dose of ethanol via oral gavage (acute) or chronically fed alcohol for 2 weeks followed by oral gavage (chronic-binge). 17-DMAG was administered during or at the end of feeding. Liver injury parameters, inflammatory cytokines and lipid metabolism genes were analysed.

Results: Our results reveal increased expression of hsp90 in human and mouse alcoholic livers. In vivo inhibition of hsp90, using 17-DMAG, not only prevented but also alleviated alcoholic liver injury, determined by lower serum ALT, AST and reduced hepatic triglycerides. Mechanistic analysis showed that 17-DMAG decreased alcohol mediated oxidative stress, reduced serum endotoxin, decreased inflammatory cells, and diminished sensitization of liver macrophages to LPS, resulting in downregulation of CD14, NFκB inhibition, and decreased pro-inflammatory cytokine production. Hsp90 inhibition decreased fatty acid synthesis genes via reduced nuclear SREBP-1 and favoured fatty acid oxidation genes via PPARα.

Conclusions: Inhibition of hsp90 decreased alcohol induced steatosis and pro-inflammatory cytokines and inhibited alcoholic liver injury. Hsp90 is therefore relevant in human alcoholic cirrhosis and a promising therapeutic target in ALD.
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http://dx.doi.org/10.1016/j.jhep.2014.05.024DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4169725PMC
October 2014

Neuroinflammation and α-synuclein accumulation in response to glucocerebrosidase deficiency are accompanied by synaptic dysfunction.

Mol Genet Metab 2014 Feb 11;111(2):152-62. Epub 2013 Dec 11.

Center for Health Sciences, Biosciences Division, SRI International, Menlo Park, CA 94025, USA. Electronic address:

Clinical, epidemiological and experimental studies confirm a connection between the common degenerative movement disorder Parkinson's disease (PD) that affects over 1 million individuals, and Gaucher disease, the most prevalent lysosomal storage disorder. Recently, human imaging studies have implicated impaired striatal dopaminergic neurotransmission in early PD pathogenesis in the context of Gaucher disease mutations, but the underlying mechanisms have yet to be characterized. In this report we describe and characterize two novel long-lived transgenic mouse models of Gba deficiency, along with a subchronic conduritol-ß-epoxide (CBE) exposure paradigm. All three murine models revealed striking glial activation within nigrostriatal pathways, accompanied by abnormal α-synuclein accumulation. Importantly, the CBE-induced, pharmacological Gaucher mouse model replicated this change in dopamine neurotransmission, revealing a markedly reduced evoked striatal dopamine release (approximately 2-fold) that indicates synaptic dysfunction. Other changes in synaptic plasticity markers, including microRNA profile and a 24.9% reduction in post-synaptic density size, were concomitant with diminished evoked dopamine release following CBE exposure. These studies afford new insights into the mechanisms underlying the Parkinson's-Gaucher disease connection, and into the physiological impact of related abnormal α-synuclein accumulation and neuroinflammation on nigrostriatal dopaminergic neurotransmission.
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http://dx.doi.org/10.1016/j.ymgme.2013.12.003DOI Listing
February 2014

Inhibition of heat shock protein (molecular weight 90 kDa) attenuates proinflammatory cytokines and prevents lipopolysaccharide-induced liver injury in mice.

Hepatology 2012 May 18;55(5):1585-95. Epub 2012 Mar 18.

Department of Medicine, University of Massachusetts Medical School, Worcester, MA 01605, USA.

Unlabelled: Endotoxin-mediated proinflammatory cytokines play a significant role in the pathogenesis of acute and chronic liver diseases. Heat shock protein 90 (molecular weight, 90 kDa) (hsp90) functions as an important chaperone of lipopolysaccharide (LPS) signaling and is required for the production of proinflammatory cytokines. We hypothesized that inhibition of hsp90 would prevent LPS-induced liver injury by decreasing proinflammatory cytokines. C57BL/6 mice were injected intraperitoneally with an hsp90 inhibitor, 17-dimethylamino-ethylamino-17-demethoxygeldanamycin (17-DMAG), and LPS. Parameters of liver injury, proinflammatory cytokines, and associated mechanisms were studied by in vivo and in vitro experiments. Inhibition of hsp90 by 17-DMAG prevented LPS-induced increases in serum alanine aminotransferase activity and significantly reduced serum tumor necrosis factor alpha (TNFα) and interleukin-6 (IL-6) protein as well as messenger RNA (mRNA) in liver. Enhanced DNA-binding activity of heat shock transcription factor 1 (HSF1) and induction of target gene heat shock protein 70 (molecular weight, 70 kDa) confirmed hsp90 inhibition in liver. 17-DMAG treatment decreased cluster of differentiation 14 mRNA and LPS-induced nuclear factor kappa light-chain enhancer of activated B cells (NFκB) DNA binding without affecting Toll-like receptor 4 mRNA in liver. Mechanistic studies revealed that 17-DMAG-mediated inhibition of TNFα showed no effect on LPS-induced NFκB promoter-driven reporter activity, but significantly decreased TNFα promoter-driven reporter activity. Chromatin immunoprecipitation assays showed that 17-DMAG enhanced HSF1 binding to the TNFα promoter, but not the IL-6 promoter, suggesting HSF1 mediated direct inhibition of TNFα, but not IL-6. We show that HSF1 indirectly regulates IL-6 by the induction of another transcription factor, activating transcription factor 3. Inhibition of HSF1, using small interfering RNA, prevented 17-DMAG-mediated down-regulation of NFκB-binding activity, TNFα, and IL-6 induction, supporting a repressive role for HSF1 on proinflammatory cytokine genes during hsp90 inhibition.

Conclusion: Hsp90 inhibition in vivo reduces proinflammatory cytokines and prevents LPS-induced liver injury likely through repressive action of HSF1. Our results suggest a novel application for 17-DMAG in alleviating LPS-induced liver injury.
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http://dx.doi.org/10.1002/hep.24802DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3342823PMC
May 2012

An essential role for monocyte chemoattractant protein-1 in alcoholic liver injury: regulation of proinflammatory cytokines and hepatic steatosis in mice.

Hepatology 2011 Dec;54(6):2185-97

Department of Medicine, University of Massachusetts Medical School, Worcester, MA 01605, USA.

Unlabelled: The importance of chemokines in alcoholic liver injury has been implicated. The role of the chemokine, monocyte chemoattractant protein-1 (MCP-1), elevated in patients with alcoholic liver disease is not yet understood. Here, we evaluated the pathophysiological significance of MCP-1 and its receptor, chemokine (C-C motif) receptor 2 (CCR2), in alcoholic liver injury. The Leiber-DeCarli diet containing alcohol or isocaloric control diets were fed to wild-type (WT) and MCP-1-deficient knockout (KO) mice for 6 weeks. In vivo and in vitro assays were performed to study the role of MCP-1 in alcoholic liver injury. MCP-1 was increased in Kupffer cells (KCs) as well as hepatocytes of alcohol-fed mice. Alcohol feeding increased serum alanine aminotransferase in WT and CCR2KO, but not MCP-1KO, mice. Alcohol-induced liver steatosis and triglyceride were attenuated in alcohol-fed MCP-1KO, but high in CCR2KO mice, compared to WT, whereas serum endotoxin was high in alcohol-fed WT and MCP-1KO mice. Expression of liver proinflammatory cytokines tumor necrosis factor alpha, interleukin (IL)-1β, IL-6, KC/IL-8, intercellular adhesion molecule 1, and cluster of differentiation 68 was induced in alcohol-fed WT, but inhibited in MCP-1KO, mice independent of nuclear factor kappa light-chain enhancer of activated B cell activation in KCs. Oxidative stress, but not cytochrome P450 2E1, was prevented in chronic alcohol-fed MCP-1KO mice, compared to WT. Increased expression of peroxisome proliferator-activated receptor (PPAR)α and PPARγ was accompanied by nuclear translocation, DNA binding, and induction of fatty acid metabolism genes acyl coenzyme A oxidase and carnitine palmitoyltransferase 1A in livers of alcohol-fed MCP-1KO mice, compared to WT controls. In vitro assays uncovered an inhibitory effect of recombinant MCP-1 on PPARα messenger RNA and peroxisome proliferator response element binding in hepatocytes independent of CCR2.

Conclusion: Deficiency of MCP-1 protects mice against alcoholic liver injury, independent of CCR2, by inhibition of proinflammatory cytokines and induction of genes related to fatty acid oxidation, linking chemokines to hepatic lipid metabolism.
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http://dx.doi.org/10.1002/hep.24599DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3342822PMC
December 2011

Genetics of P-element transposition into Drosophila melanogaster centric heterochromatin.

Genetics 2003 Dec;165(4):2039-53

Molecular and Cell Biology Laboratory, The Salk Institute for Biological Studies, San Diego, CA 92037, USA.

Heterochromatin is a major component of higher eukaryotic genomes, but progress in understanding the molecular structure and composition of heterochromatin has lagged behind the production of relatively complete euchromatic genome sequences. The introduction of single-copy molecular-genetic entry points can greatly facilitate structure and sequence analysis of heterochromatic regions that are rich in repeated DNA. In this study, we report the isolation of 502 new P-element insertions into Drosophila melanogaster centric heterochromatin, generated in nine different genetic screens that relied on mosaic silencing (position-effect variegation, or PEV) of the yellow gene present in the transposon. The highest frequencies of recovery of variegating insertions were observed when centric insertions were used as the source for mobilization. We propose that the increased recovery of variegating insertions from heterochromatic starting sites may result from the physical proximity of different heterochromatic regions in germline nuclei or from the association of mobilizing elements with heterochromatin proteins. High frequencies of variegating insertions were also recovered when a potent suppressor of PEV (an extra Y chromosome) was present in both the mobilization and selection generations, presumably due to the effects of chromatin structure on P-element mobilization, insertion, and phenotypic selection. Finally, fewer variegating insertions were recovered after mobilization in females, in comparison to males, which may reflect differences in heterochromatin structure in the female and male germlines. FISH localization of a subset of the insertions confirmed that 98% of the variegating lines contain heterochromatic insertions and that these schemes produce a broader distribution of insertion sites. The results of these schemes have identified the most efficient methods for generating centric heterochromatin P insertions. In addition, the large collection of insertions produced by these screens provides molecular-genetic entry points for mapping, sequencing, and functional analysis of Drosophila heterochromatin.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1462875PMC
December 2003