Detection of surface and cytoplasmic CD4 on blood monocytes from normal and HIV-1 infected individuals

doi:10.1016/0022-1759(90)90256-U

Journal of Immunological Methods

Volume 135, Issues 1–2, 31 December 1990, Pages 59-69

https://doi.org/10.1016/0022-1759(90)90256-U Get rights and content

Abstract

CD4 on blood monocytes is generally regarded as being found on a subset of blood monocytes. However, our results show that all monocytes are CD4⁺ but the number of molecules per cell is lower than T cells. We have performed immunofluorescent (flow cytometry, microscopy) analysis of monocytes from normal donors and HIV-1-infected patients using anti-CD4 (Leu-3a) and the monocyte-specific marker anti-CD14 (Leu-M3) monoclonal antibodies. Specifically: (1) 91% of monocytes from normal individuals show dual positivity for CD14 and CD4 (n = 14) as measured by flow cytometry; (2) 76% of monocytes expressed surface bound CD4 and CD14 when an enhanced two colour immunofluorescence microscopic technique was employed; (3) all CD14⁺ monocytes stained with an intensity of 3⁺−4⁺ for cytoplasmic CD4. Few monocytes were CD14⁻ and CD4⁺. Cell surface CD4 expression was blocked with unconjugated anti-CD4 prior to staining; (4) the staining intensity for cytoplasmic CD4 in T cells was negligible; (5) CD4 expression on monocytes obtained from patients was as observed with normal individuals. The conclusion drawn is that all monocytes are CD4⁺ and the CD4 expression in monocytes is mainly cytoplasmic. Thus, all monocytes are potentially infectable with HIV-1.

References (15)

B. Freundlich et al.
Use of gelatin/plasma coated flasks for isolating human peripheral blood monocytes
J. Immunol. Methods
(1983)
S.Z. Salahuddin et al.
Human T lymphotropic virus type III infection of human alveolar macrophages
Blood
(1986)
M. Busch et al.
Detection of human immunodeficiency virus infection of myeloid precursors in bone marrow samples from AIDS patients
Blood
(1986)
S. Crowe et al.
Quantitative immunocytofluorographic analysis of CD4 antigen expression and HIV infection of human peripheral blood monocyte/macrophages
Aids Res. Human Retroviruses
(1987)
S. Crowe et al.
Human monocyte/macrophage are the major reservoir of HIV in vivo
C.R. Faltynek et al.
Treatment with recombinant IFN-γ decreases cell surface CD4 levels on peripheral blood and on myelomonocytic cell lines
J. Immunol.
(1989)
T.M. Folks et al.
Infection and replication of HIV-1 in purified progenitor cells of normal human bone marrow
Science
(1988)

There are more references available in the full text version of this article.

Cited by (48)

Depletion of high-content CD14+ cells from apheresis products is critical for successful transduction and expansion of CAR T cells during large-scale cGMP manufacturing
2021, Molecular Therapy Methods and Clinical Development
Citation Excerpt :
Indeed, Shah et al.30 also reported recently that monocyte frequencies negatively affected CAR T cell manufacturing by inhibiting transduction and expansion of anti-CD22 CAR T cells, and that upfront incorporation of CD4/8-T cell selection effectively salvaged apheresis material unable to be used for CAR T cell manufacturing using their previous selection method. One potential complication with upfront CD4/CD8 selection, though, is that monocytes also express CD4 on their surface;31 therefore, it still may be worthwhile to monitor the monocyte content in incoming apheresis and post-selected products to ensure the CD14 content is within the acceptable manufacturing threshold of ≤40% as established here. In conclusion, high monocyte content poses a serious challenge for CAR T cell manufacturing starting from unmanipulated apheresis products.
With the US Food and Drug Administration (FDA) approval of four CD19- and one BCMA-targeted chimeric antigen receptor (CAR) therapy for B cell malignancies, CAR T cell therapy has finally reached the status of a medicinal product. The successful manufacturing of autologous CAR T cell products is a key requirement for this promising treatment modality. By analyzing the composition of 214 apheresis products from 210 subjects across eight disease indications, we found that high CD14⁺ cell content poses a challenge for manufacturing CAR T cells, especially in patients with non-Hodgkin’s lymphoma and multiple myeloma caused by the non-specific phagocytosis of the magnetic beads used to activate CD3⁺ T cells. We demonstrated that monocyte depletion via rapid plastic surface adhesion significantly reduces the CD14⁺ monocyte content in the apheresis products and simultaneously boosts the CD3⁺ content. We established a 40% CD14⁺ threshold for the stratification of apheresis products across nine clinical trials and demonstrated the effectiveness of this procedure by comparing manufacturing runs in two phase 1 clinical trials. Our study suggests that CD14⁺ content should be monitored in apheresis products, and that the manufacturing of CAR T cells should incorporate a step that lessens the CD14⁺ cell content in apheresis products containing more than 40% to maximize the production success.
Immunomodulation by cannabinoids: Current uses, mechanisms, and identification of data gaps to be addressed for additional therapeutic application
2021, Advances in Pharmacology
Citation Excerpt :
HIV is a disease transmitted by two species of lentivirus that target the immune system. HIV gains entry into cells by initially binding via gp120 to CD4 (Fanales-Belasio, Raimondo, Suligoi, & Butto, 2010), which is expressed on T helper cells and also at significantly lower levels on monocytes/macrophages, DCs, eosinophils and microglial cells (Filion, Izaguirre, Garber, Huebsh, & Aye, 1990; Lucey, Dorsky, Nicholson-Weller, & Weller, 1989; Patterson et al., 1995; Perry & Gordon, 1987). Upon binding of viral gp120 to CD4, gp120 then binds to chemokine receptors, such as CXCR4 and CCR5.
The endocannabinoid system plays a critical role in immunity and therefore its components, including cannabinoid receptors 1 and 2 (CB₁ and CB₂), are putative druggable targets for immune-mediated diseases. Whether modulating endogenous cannabinoid levels or interacting with CB₁ or CB₂ receptors directly, cannabinoids or cannabinoid-based therapeutics (CBTs) show promise as anti-inflammatory or immune suppressive agents. Herein we provide an overview of cannabinoid effects in animals and humans that provide support for the use of CBTs in immune-mediated disease such as multiple sclerosis (MS), inflammatory bowel disease (IBD), asthma, arthritis, diabetes, human immunodeficiency virus (HIV), and HIV-associated neurocognitive disorder (HAND). This is not an exhaustive review of cannabinoid effects on immune responses, but rather provides: (1) key studies in which initial and/or novel observations were made in animal studies; (2) critical human studies including meta-analyses and randomized clinical trials (RCTs) in which CBTs have been assessed; and (3) evidence for the role of CB₁ or CB₂ receptors in immune-mediated diseases through genetic analyses of single nucleotide polymorphisms (SNPs) in the CNR1 and CNR2 genes that encode CB₁ or CB₂ receptors, respectively. Perhaps most importantly, we provide our view of data gaps that exist, which if addressed, would allow for more rigorous evaluation of the efficacy and risk to benefit ratio of the use of cannabinoids and/or CBTs for immune-mediated diseases.
HIV Entry and Its Inhibition by Bifunctional Antiviral Proteins
2018, Molecular Therapy Nucleic Acids
HIV entry is a highly specific and time-sensitive process that can be divided into receptor binding, coreceptor binding, and membrane fusion. Bifunctional antiviral proteins (bAVPs) exploit the multi-step nature of the HIV entry process by binding to two different extracellular targets. They are generated by expressing a fusion protein containing two entry inhibitors with a flexible linker. The resulting fusion proteins exhibit exceptional neutralization potency and broad cross-clade inhibition. In this review, we summarize the HIV entry process and provide an overview of the design, antiviral potency, and methods of delivery of bAVPs. Additionally, we discuss the advantages and limitations of bAVPs for HIV prevention and treatment.
Multilineage polyclonal engraftment of Cal-1 gene-modified cells and in vivo selection after SHIV infection in a nonhuman primate model of AIDS
2016, Molecular Therapy Methods and Clinical Development
Citation Excerpt :
These findings suggest that only infection-susceptible cells (those that permit virus entry, i.e. CD4+CCR5+ and/or CD4+CXCR4+ subsets) undergo virus-mediated positive selection, which is more difficult to measure in a mixed population of susceptible and nonsusceptible cells. Although CD14+ cells can be infected, the lack of selection for Cal-1-marked cells in this subset is consistent with the limited proportions that express sufficient levels of CD4 and CCR5/CXCR4 to permit virus entry.48,49 The selective upregulation of mC46 in infection-susceptible cells highlights a potential use of Cal-1 to better understand viral pathogenesis in our SHIV-infection model.
We have focused on gene therapy approaches to induce functional cure/remission of HIV-1 infection. Here, we evaluated the safety and efficacy of the clinical grade anti-HIV lentiviral vector, Cal-1, in pigtailed macaques (Macaca nemestrina). Cal-1 animals exhibit robust levels of gene marking in myeloid and lymphoid lineages without measurable adverse events, suggesting that Cal-1 transduction and autologous transplantation of hematopoietic stem cells are safe, and lead to long-term, multilineage engraftment following myeloablative conditioning. Ex vivo, CD4+ cells from transplanted animals undergo positive selection in the presence of simian/human immunodeficiency virus (SHIV). In vivo, Cal-1 gene-marked cells are evident in the peripheral blood and in HIV-relevant tissue sites such as the gastrointestinal tract. Positive selection for gene-marked cells is observed in blood and tissues following SHIV challenge, leading to maintenance of peripheral blood CD4+ T-cell counts in a normal range. Analysis of Cal-1 lentivirus integration sites confirms polyclonal engraftment of gene-marked cells. Following infection, a polyclonal, SHIV-resistant clonal repertoire is established. These findings offer strong preclinical evidence for safety and efficacy of Cal-1, present a new method for tracking protected cells over the course of virus-mediated selective pressure in vivo, and reveal previously unobserved dynamics of virus-dependent T-cell selection.
CD4+/CD8+ macrophages infiltrating at inflammatory sites: A population of monocytes/macrophages with a cytotoxic phenotype
2006, Blood
Citation Excerpt :
In addition, we noted that CD4 and CD8 were distributed not only on the cell surface but also in the cytoplasm of DP monocytes. These findings are consistent with the previous observation that CD4 is also expressed in the cytoplasm of human monocytes.29 The majority of CD8 molecules are heterodimers composed of α- and β-chains.
We found a population of nonlymphoid cells expressing both CD4 and CD8 in peripheral blood mononuclear cells (PBMCs) of human T-cell leukemia virus type-I pX transgenic rats with autoimmune diseases. These cells, which showed a monocytic phenotype, were also found in wild-type rats, and their number increased by adjuvant-assisted immunization. GM-CSF increased the number of these double-positive (DP) monocytes in PBMCs. Consistent with the idea that DP monocytes differentiate into DP macrophages at sites of inflammation, we found infiltration of DP macrophages at the site of myosin-induced myocarditis in wild-type rats; these cells exhibited a T-helper 1 (Th1)-type cytokine/chemokine profile and expressed high levels of Fas ligand, perforin, granzyme B, and NKR-P2 (rat orthologue of human NKG2D). Adoptive transfer of GFP-positive spleen cells confirmed hematogenous origin of DP macrophages. DP monocytes had a cytotoxic phenotype similar to DP macrophages, indicating that this phenotypic specialization occurred before entry into a tissue. In line with this, DP monocytes killed tumor cells in vitro. Combined evidence indicates that certain inflammatory stimuli that induce GM-CSF trigger the expansion of a population of DP monocytes with a cytotoxic phenotype and that these cells differentiate into macrophages at inflammatory sites. Interestingly, human PBMCs also contain DP monocytes.
Enolase 1 of Candida albicans binds human CD4+ T cells and modulates naïve and memory responses
2023, European Journal of Immunology

View all citing articles on Scopus

View full text

Article preview

Journal of Immunological Methods

Abstract

References (15)

Use of gelatin/plasma coated flasks for isolating human peripheral blood monocytes

J. Immunol. Methods

Human T lymphotropic virus type III infection of human alveolar macrophages

Blood

Detection of human immunodeficiency virus infection of myeloid precursors in bone marrow samples from AIDS patients

Blood

Quantitative immunocytofluorographic analysis of CD4 antigen expression and HIV infection of human peripheral blood monocyte/macrophages

Aids Res. Human Retroviruses

Human monocyte/macrophage are the major reservoir of HIV in vivo

Treatment with recombinant IFN-γ decreases cell surface CD4 levels on peripheral blood and on myelomonocytic cell lines

J. Immunol.

Infection and replication of HIV-1 in purified progenitor cells of normal human bone marrow

Science

Cited by (48)

Depletion of high-content CD14<sup>+</sup> cells from apheresis products is critical for successful transduction and expansion of CAR T cells during large-scale cGMP manufacturing

Immunomodulation by cannabinoids: Current uses, mechanisms, and identification of data gaps to be addressed for additional therapeutic application

HIV Entry and Its Inhibition by Bifunctional Antiviral Proteins

Multilineage polyclonal engraftment of Cal-1 gene-modified cells and in vivo selection after SHIV infection in a nonhuman primate model of AIDS

CD4<sup>+</sup>/CD8<sup>+</sup> macrophages infiltrating at inflammatory sites: A population of monocytes/macrophages with a cytotoxic phenotype

Enolase 1 of Candida albicans binds human CD4<sup>+</sup> T cells and modulates naïve and memory responses