Antiretrovirals inhibit arginase in human microglia

Lisi, Lucia; Laudati, Emilia; Miscioscia, Teresa Fabiola; Russo, Cinzia Dello; Topai, Alessandra; Navarra, Pierluigi

doi:10.1111/jnc.13393

Cited by 15 publications

(16 citation statements)

References 31 publications

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“…This is not surprising as it has been characterized that CHME-5 cells require a cytomix formula (e.g. interferon-c, TNF-a, IL1-b) to produce proinflammatory substances such as nitric oxide [25]. Furthermore, by using rat-specific gRNAs and PCR assays in our experiments, we confirmed that CHME-5 cells are derived from rat.…”

Section: Discussionsupporting

confidence: 83%

In vitro modeling of HIV proviral activity in microglia

et al. 2017

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Microglia, the resident macrophages of the brain, play a key role in the pathogenesis of Human Immunodeficiency Virus (HIV)-associated neurocognitive disorders (HAND) due to their productive infection by HIV. This results in the release of neurotoxic viral proteins and pro-inflammatory compounds which negatively affect the functionality of surrounding neurons. Because models of HIV infection within the brain are limited, we aimed to create a novel microglia cell line with an integrated HIV-provirus capable of recreating several hallmarks of HIV infection. We utilized CRISPR/Cas9 gene editing technology and integrated a modified HIV provirus into CHME-5 immortalized microglia to create HIV-NanoLuc CHME-5. In the modified provirus, the Gag-Pol region is replaced with the coding region for Nanoluciferase (NanoLuc), which allows for the rapid assay of HIV LTR activity using a luminescent substrate, while still containing the necessary genetic material to produce established neurotoxic viral proteins (e.g. tat, nef, gp120). We confirmed that HIV-NanoLuc CHME-5 microglia express NanoLuc, along with the HIV viral protein Nef. We subsequently exposed these cells to a battery of experiments to modulate the activity of the provirus. Proviral activity was enhanced by treating the cells with pro-inflammatory factors LPS and TNF-α and by overexpressing the viral regulatory protein Tat. Conversely, genetic modification of the TLR-4 gene by CRISPR/Cas9 reduced LPS-mediated proviral activation, and pharmacological application of NF-κB inhibitor sulfasalazine similarly diminished proviral activity. Overall these data suggest that HIV-NanoLuc CHME-5 may be a useful tool in the study of HIV-mediated neuropathology and proviral regulation.

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Section: Discussionsupporting

confidence: 83%

In vitro modeling of HIV proviral activity in microglia

et al. 2017

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“…In contrast to the HMC3/CHME3 aliquots, increased iNOS expression and NO production was detected also after overexpression of the endogenous retrovirus W envelope protein [88] and in response to a mixture of pro-inflammatory cytokines, including TNFα, IL-1β, and IFNγ [106]. Furthermore, the CHME-5 cells displayed increased arginase (ARG) activity in response to IL-4, which was significantly reduced by several antiretroviral drugs [106].…”

Section: The Human Microglial Chme-5 Cell Linementioning

confidence: 99%

The human microglial HMC3 cell line: where do we stand? A systematic literature review

Russo

Cappoli

Coletta

et al. 2018

J Neuroinflammation

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162

130

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Microglia, unique myeloid cells residing in the brain parenchyma, represent the first line of immune defense within the central nervous system. In addition to their immune functions, microglial cells play an important role in other cerebral processes, including the regulation of synaptic architecture and neurogenesis. Chronic microglial activation is regarded as detrimental, and it is considered a pathogenic mechanism common to several neurological disorders. Microglial activation and function have been extensively studied in rodent experimental models, whereas the characterization of human cells has been limited due to the restricted availability of primary sources of human microglia. To overcome this problem, human immortalized microglial cell lines have been developed. The human microglial clone 3 cell line, HMC3, was established in 1995, through SV40-dependent immortalization of human embryonic microglial cells. It has been recently authenticated by the American Type Culture Collection (ATCC®) and distributed under the name of HMC3 (ATCC®CRL-3304). The HMC3 cells have been used in six research studies, two of which also indicated by ATCC® as reference articles. However, a more accurate literature revision suggests that clone 3 was initially distributed under the name of CHME3. In this regard, several studies have been published, thus contributing to a more extensive characterization of this cell line. Remarkably, the same cell line has been used in different laboratories with other denominations, i.e., CHME-5 cells and C13-NJ cells. In view of the fact that “being now authenticated by ATCC®” may imply a wider distribution of the cells, we aimed at reviewing data obtained with the human microglia cell line clone 3, making the readers aware of this complicated nomenclature. In addition, we also included original data, generated in our laboratory with the HMC3 (ATCC®CRL-3304) cells, providing information on the current state of the culture together with supplementary details on the culturing procedures to obtain and maintain viable cells.

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“…LPSinduced NF-κB p65 activation was consistent with other microglial cell lines [28,[37][38][39] . Given the fact that many studies investigating microglial responses used the same dose of LPS (1 μg/mL), we are confident that CHME-5 cells were adequately treated to observe inflammatory responses [36,40,41] ; unlike the use of a lower dose of LPS (1 ng/mL), which failed to activate and release nitric oxide in CHME-5 cells [42] . Additionally, other studies using human primary microglia and CHME-5 cells have used similar LPS dose range (0.1-1 μg/mL) and time points (6 h for RT-PCR) to investigate inflammatory responses [26,36,37,40,41] .…”

Section: Discussionmentioning

confidence: 91%

LPS-induced TLR4 neuroinflammatory signaling in CHME-5 microglial cells

Figueroa-Hall¹,

Anderson²,

Das³

et al. 2017

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Aim:In the field of neuroinflammation, identifying specific effects of pharmacological agents and other factors is problematic given the relative difficulty and expense in obtaining and culturing primary microglia. Immortalized microglial cell lines are very useful, but only a limited number have been characterized for inflammatory signaling. Therefore, characterization of lipopolysaccharide (LPS)-induced toll-like receptor 4 (TLR4) signaling in CHME-5, a microglial cell line, is expected to be of value as an experimental model of inflammatory signaling in the central nervous system (CNS). Methods: It was recently suggested that CHME-5 cells are of rat origin, not human, hence, verification of this claim using short tandem repeat genotype sequencing, along with immunoblotting, reverse transcription-polymerase chain reaction, and immunocytochemistry techniques to validate that CHME-5 retain morphological, phenotypical, and functional characteristics of primary microglia were undertaken. Results: LPS induced inhibitor kappa B-alpha and nuclear factorkappa B (NF-κB) p65 activation, NF-κB p65 binding activity, and tumor necrosis factor alpha gene expression. Additionally, results also confirmed the maintenance of microglial phenotype as seen with increased cluster of differentiation 68 gene and protein expression, immunofluorescence, and the absence of glial fibrillary acidic protein-immunoreactivity. Key words:Escherichia coli lipopolysaccharide, nuclear factor-kappa B p65, cluster of differentiation 68, toll-like receptor 4, neuroinflammation ABSTRACTArticle history:

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Antiretrovirals inhibit arginase in human microglia

Cited by 15 publications

References 31 publications

In vitro modeling of HIV proviral activity in microglia

In vitro modeling of HIV proviral activity in microglia

The human microglial HMC3 cell line: where do we stand? A systematic literature review

LPS-induced TLR4 neuroinflammatory signaling in CHME-5 microglial cells

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