A new improved method for the concentration of HIV-1 infective particles

Kohno, Takehiro; Mohan, Sharad; Goto, Toshiyuki; Morita, Chizuko; Nakano, Takashi; Wu, Hong; Sangco, Joseph Carlo E; Morimatsu, Shinichi; Sano, Kohei

doi:10.1016/s0166-0934(02)00162-3

Cited by 31 publications

(28 citation statements)

References 19 publications

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“…For pseudotyped and internal promoter viruses, supernatants were concentrated using high–molecular weight polyethylene glycol (PEG) precipitation 18. Pellets were resuspended in StemSpan medium and MOIs were calculated using MOI = −Ln(1− p ) where p is the proportion of CEM T cells infected.…”

Section: Methodsmentioning

confidence: 99%

HIV-1 infects multipotent progenitor cells causing cell death and establishing latent cellular reservoirs

Carter

Onafuwa-Nuga

McNamara

et al. 2010

Nat Med

265

277

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HIV causes a chronic infection characterized by depletion of CD4+ T lymphocytes and development of opportunistic infections. Despite drugs that inhibit viral spread, HIV has been difficult to cure because of uncharacterized reservoirs of infected cells that are resistant to highly active antiretroviral therapy and the immune response. Here we used CD34+ cells from infected people as well as in vitro studies of wild type HIV to demonstrate infection and killing of CD34+ multipotent hematopoietic progenitor cells (HPCs). In some HPCs, we detected latent infection that stably persisted in cell culture until viral gene expression was activated by differentiation factors. A novel reporter HIV that directly detects latently infected cells in vitro confirmed the presence of distinct populations of active and latently infected HPCs. These findings have important implications for understanding HIV bone marrow pathology and the mechanisms by which HIV causes persistent infection.

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Section: Methodsmentioning

confidence: 99%

HIV-1 infects multipotent progenitor cells causing cell death and establishing latent cellular reservoirs

Carter

Onafuwa-Nuga

McNamara

et al. 2010

Nat Med

265

277

View full text Add to dashboard Cite

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“…HXB-ePLAP (15) or NL4-3-⌬GPE-GFP was cotransfected with a plasmid encoding either the vesicular stomatitis virus glycoprotein (VSV-G) or the HXB envelope; the helper plasmid pCMV-HIV-1 (28) was additionally cotransfected to permit infectious particles to be formed with NL4-3-⌬GPE-GFP. Viruses pseudotyped with the HXB envelope were concentrated using high-molecular-weight polyethylene glycol precipitation as described previously (39). Pellets were resuspended in 1/25 the original volume and were used immediately.…”

Section: Methodsmentioning

confidence: 99%

Latent HIV-1 Infection Occurs in Multiple Subsets of Hematopoietic Progenitor Cells and Is Reversed by NF-κB Activation

McNamara

Ganesh

Collins

2012

J Virol

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The ability of HIV-1 to establish a latent infection presents a barrier to curing HIV. The best-studied reservoir of latent virus in vivo is resting memory CD4 + T cells, but it has recently been shown that CD34 + hematopoietic progenitor cells (HPCs) can also become latently infected by HIV-1 in vitro and in vivo . CD34 + cells are not homogenous, however, and it is not yet known which types of CD34 + cells support a latent infection. Furthermore, the mechanisms through which latency is established in this cell type are not yet known. Here we report the development of a primary cell model for latent HIV-1 infection in HPCs. We demonstrate that in this model, latent infection can be established in all subsets of HPCs examined, including HPCs with cell surface markers consistent with immature hematopoietic stem and progenitor cells. We further show that the establishment of latent infection in these cells can be reversed by tumor necrosis factor alpha (TNF-α) through an NF-κB-dependent mechanism. In contrast, we do not find evidence for a role of positive transcription elongation factor b (P-TEFb) in the establishment of latent infection in HPCs. Finally, we demonstrate that prostratin and suberoylanilide hydroxamic acid (SAHA), but not hexamethylene bisacetamide (HMBA) or 5-aza-2′-deoxycytidine (Aza-CdR), reactivate latent HIV-1 in HPCs. These findings illuminate the mechanisms through which latent infection can be established in HPCs and suggest common pathways through which latent virus could be reactivated in both HPCs and resting memory T cells to eliminate latent reservoirs of HIV-1.

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“…Viral supernatants were collected, centrifuged at 2,000 rpm for 2 minutes to remove dead cells, and filtered using a 0.45 mm filter. Lentiviral supernatants were then concentrated by using PEG8000 and tittered by FACS analysis of GFP expression as described previously (24,25). Tittered lentiviral preparations were then stored at À80 C in appropriate aliquots.…”

Section: Lentivirus Productionmentioning

confidence: 99%

Inactivation of the Kinase Domain of CDK10 Prevents Tumor Growth in a Preclinical Model of Colorectal Cancer, and Is Accompanied by Downregulation of Bcl-2

Weiswald

Hasan

Wong

et al. 2017

Molecular Cancer Therapeutics

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Cyclin-dependent kinase 10 (CDK10), a CDC2-related kinase, is highly expressed in colorectal cancer. Its role in the pathogenesis of colorectal cancer is unknown. This study examines the function of CDK10 in colorectal cancer, and demonstrates its role in suppressing apoptosis and in promoting tumor growth and Modulation of CDK10 expression in colorectal cancer cell lines demonstrates that CDK10 promotes cell growth, reduces chemosensitivity and inhibits apoptosis by upregulating the expression of Bcl-2. This effect appears to depend on its kinase activity, as kinase-defective mutant colorectal cancer cell lines have an exaggerated apoptotic response and reduced proliferative capacity. , inhibiting CDK10 in colorectal cancer following intratumoral injections of lentivirus-mediated CDK10 siRNA in a patient-derived xenograft mouse model demonstrated its efficacy in suppressing tumor growth. Furthermore, using a tissue microarray of human colorectal cancer tissues, the potential for CDK10 to be a prognostic biomarker in colorectal cancer was explored. In tumors of individuals with colorectal cancer, high expression of CDK10 correlates with earlier relapse and shorter overall survival. The findings of this study indicate that CDK10 plays a role in the pathogenesis in colorectal cancer and may be a potential therapeutic target for treatment..

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A new improved method for the concentration of HIV-1 infective particles

Cited by 31 publications

References 19 publications

HIV-1 infects multipotent progenitor cells causing cell death and establishing latent cellular reservoirs

HIV-1 infects multipotent progenitor cells causing cell death and establishing latent cellular reservoirs

Latent HIV-1 Infection Occurs in Multiple Subsets of Hematopoietic Progenitor Cells and Is Reversed by NF-κB Activation

Inactivation of the Kinase Domain of CDK10 Prevents Tumor Growth in a Preclinical Model of Colorectal Cancer, and Is Accompanied by Downregulation of Bcl-2

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