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The progestin-only contraceptive medroxyprogesterone acetate, but not norethisterone acetate, enhances HIV-1 Vpr-mediated apoptosis in human CD4+ T cells through the glucocorticoid receptor.

Tomasicchio M, Avenant C, Du Toit A, Ray RM, Hapgood JP - PLoS ONE (2013)

Bottom Line: These results, together with the findings that RU486, a GR antagonist, prevents Dex-, MPA- and Vpr-mediated apoptosis, provide evidence for the first time that GR agonists or partial agonists increase apoptosis in primary CD4(+) T-cells via the GR.This work suggests that contraceptive doses of MPA but not NET-A or physiological doses of progesterone could potentially accelerate depletion of CD4(+) T-cells in a GR-dependent fashion in HIV-1 positive women, thereby contributing to immunodeficiency.The results imply that choice of progestin used in contraception may be critical to susceptibility and progression of diseases such as HIV-1.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cell Biology, University of Cape Town, Cape Town, Western Province, South Africa.

ABSTRACT
The glucocorticoid receptor (GR) regulates several physiological functions, including immune function and apoptosis. The HIV-1 virus accessory protein, viral protein R (Vpr), can modulate the transcriptional response of the GR. Glucocorticoids (GCs) and Vpr have been reported to induce apoptosis in various cells, including T-cells. We have previously shown that the injectable contraceptive, medroxyprogesterone acetate (MPA) is a partial to full agonist for the GR, unlike norethisterone acetate (NET-A). We investigated the functional cross talk between the GR and Vpr in inducing apoptosis in CD4(+) T-cells, in the absence and presence of GCs and these progestins, as well as progesterone. By using flow cytometry, we show that, in contrast to NET-A and progesterone, the synthetic GR ligand dexamethasone (Dex), cortisol and MPA induce apoptosis in primary CD4(+) T-cells. Furthermore, the C-terminal part of the Vpr peptide, or HIV-1 pseudovirus, together with Dex or MPA further increased the apoptotic phenotype, unlike NET-A and progesterone. By a combination of Western blotting, PCR and the use of receptor- selective agonists, we provide evidence that the GR and the estrogen receptor are the only steroid receptors expressed in peripheral blood mononuclear cells. These results, together with the findings that RU486, a GR antagonist, prevents Dex-, MPA- and Vpr-mediated apoptosis, provide evidence for the first time that GR agonists or partial agonists increase apoptosis in primary CD4(+) T-cells via the GR. We show that apoptotic induction involves differential expression of key apoptotic genes by both Vpr and GCs/MPA. This work suggests that contraceptive doses of MPA but not NET-A or physiological doses of progesterone could potentially accelerate depletion of CD4(+) T-cells in a GR-dependent fashion in HIV-1 positive women, thereby contributing to immunodeficiency. The results imply that choice of progestin used in contraception may be critical to susceptibility and progression of diseases such as HIV-1.

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The progestin MPA, but not NET-A or P4, induces apoptosis in CD4+ T-cells.Cells were treated with or without 100 nM Dex, 100 nM F, 1 µM MPA, 10 µM NET-A, 1 µM P4 or vehicle control (EtOH) for 24 hrs. Cells were stained with anti-CD4, anti-CD14, annexin V and 7-AAD using the Apoptosis Detection kit I (BD biosciences). (A) Gating strategy and representative zebra plots of untreated (EtOH), MPA or Dex treated PBMCs. (B) The histogram shows pooled results from two independent experiments with samples from three donors. Data were acquired on a FACS calibur system (BD Biosciences) and analyzed using Flo-Jo software (Tree Star Inc., San Carlos, CA, USA). Statistical significance was determined by one-way ANOVA with Dunnett’s post-test, where *, **, and *** indicate p<0.05, 0.01 and 0.005 respectively. Error bars represent standard deviation.
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pone-0062895-g001: The progestin MPA, but not NET-A or P4, induces apoptosis in CD4+ T-cells.Cells were treated with or without 100 nM Dex, 100 nM F, 1 µM MPA, 10 µM NET-A, 1 µM P4 or vehicle control (EtOH) for 24 hrs. Cells were stained with anti-CD4, anti-CD14, annexin V and 7-AAD using the Apoptosis Detection kit I (BD biosciences). (A) Gating strategy and representative zebra plots of untreated (EtOH), MPA or Dex treated PBMCs. (B) The histogram shows pooled results from two independent experiments with samples from three donors. Data were acquired on a FACS calibur system (BD Biosciences) and analyzed using Flo-Jo software (Tree Star Inc., San Carlos, CA, USA). Statistical significance was determined by one-way ANOVA with Dunnett’s post-test, where *, **, and *** indicate p<0.05, 0.01 and 0.005 respectively. Error bars represent standard deviation.

Mentions: GCs have been shown to induce apoptosis in several different cell lines, including CD4+ T-cells [35]. The progestin MPA is a partial to full GR agonist, unlike NET-A and P4 which have weak to no GR activity [30]–[33]. We investigated the relative capability of MPA and NET-A to induce apoptosis in CD4+ T-cells and CD14+ monocytes, as compared to the endogenous GC agonist F, the synthetic GR agonist Dex and P4. Briefly, PBMCs were isolated and treated with 100 nM Dex, 100 nM MPA, 10 µM NET-A, 1 µM P4 or vehicle control (EtOH) for 24 hrs. Cells were stained with anti-CD4 (T-cells), anti-CD14 (monocytes), 7-aminoactinomycin D (7-AAD), annexin V and the data were acquired using the Becton Dickinson FACS Calibur. 7-AAD was included to discriminate between live and dead cells. CD4+ T-cells and CD14+ monocytes were gated from the total PBMC population as indicated and the apoptotic cells were detected with the apoptosis marker annexin V (Figure 1A). As expected Dex and F induced apoptosis in a statistically significant manner in CD4+ T-cells by about 2-fold and 1.6-fold, respectively, compared to untreated cells (Figure 1B). Importantly, MPA also statistically significantly induced apoptosis in these cells (1.5-fold), yet to a lesser extent than Dex. By contrast, when cells were treated with NET-A or P4 no increase in apoptosis compared to control, was detected in CD4+ T-cells (Figure 1B). The apoptotic effect of Dex, F and MPA was however not observed in CD14+ monocytes (data not shown) and therefore the following experiments were carried out in CD4+ T-cells.


The progestin-only contraceptive medroxyprogesterone acetate, but not norethisterone acetate, enhances HIV-1 Vpr-mediated apoptosis in human CD4+ T cells through the glucocorticoid receptor.

Tomasicchio M, Avenant C, Du Toit A, Ray RM, Hapgood JP - PLoS ONE (2013)

The progestin MPA, but not NET-A or P4, induces apoptosis in CD4+ T-cells.Cells were treated with or without 100 nM Dex, 100 nM F, 1 µM MPA, 10 µM NET-A, 1 µM P4 or vehicle control (EtOH) for 24 hrs. Cells were stained with anti-CD4, anti-CD14, annexin V and 7-AAD using the Apoptosis Detection kit I (BD biosciences). (A) Gating strategy and representative zebra plots of untreated (EtOH), MPA or Dex treated PBMCs. (B) The histogram shows pooled results from two independent experiments with samples from three donors. Data were acquired on a FACS calibur system (BD Biosciences) and analyzed using Flo-Jo software (Tree Star Inc., San Carlos, CA, USA). Statistical significance was determined by one-way ANOVA with Dunnett’s post-test, where *, **, and *** indicate p<0.05, 0.01 and 0.005 respectively. Error bars represent standard deviation.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3643923&req=5

pone-0062895-g001: The progestin MPA, but not NET-A or P4, induces apoptosis in CD4+ T-cells.Cells were treated with or without 100 nM Dex, 100 nM F, 1 µM MPA, 10 µM NET-A, 1 µM P4 or vehicle control (EtOH) for 24 hrs. Cells were stained with anti-CD4, anti-CD14, annexin V and 7-AAD using the Apoptosis Detection kit I (BD biosciences). (A) Gating strategy and representative zebra plots of untreated (EtOH), MPA or Dex treated PBMCs. (B) The histogram shows pooled results from two independent experiments with samples from three donors. Data were acquired on a FACS calibur system (BD Biosciences) and analyzed using Flo-Jo software (Tree Star Inc., San Carlos, CA, USA). Statistical significance was determined by one-way ANOVA with Dunnett’s post-test, where *, **, and *** indicate p<0.05, 0.01 and 0.005 respectively. Error bars represent standard deviation.
Mentions: GCs have been shown to induce apoptosis in several different cell lines, including CD4+ T-cells [35]. The progestin MPA is a partial to full GR agonist, unlike NET-A and P4 which have weak to no GR activity [30]–[33]. We investigated the relative capability of MPA and NET-A to induce apoptosis in CD4+ T-cells and CD14+ monocytes, as compared to the endogenous GC agonist F, the synthetic GR agonist Dex and P4. Briefly, PBMCs were isolated and treated with 100 nM Dex, 100 nM MPA, 10 µM NET-A, 1 µM P4 or vehicle control (EtOH) for 24 hrs. Cells were stained with anti-CD4 (T-cells), anti-CD14 (monocytes), 7-aminoactinomycin D (7-AAD), annexin V and the data were acquired using the Becton Dickinson FACS Calibur. 7-AAD was included to discriminate between live and dead cells. CD4+ T-cells and CD14+ monocytes were gated from the total PBMC population as indicated and the apoptotic cells were detected with the apoptosis marker annexin V (Figure 1A). As expected Dex and F induced apoptosis in a statistically significant manner in CD4+ T-cells by about 2-fold and 1.6-fold, respectively, compared to untreated cells (Figure 1B). Importantly, MPA also statistically significantly induced apoptosis in these cells (1.5-fold), yet to a lesser extent than Dex. By contrast, when cells were treated with NET-A or P4 no increase in apoptosis compared to control, was detected in CD4+ T-cells (Figure 1B). The apoptotic effect of Dex, F and MPA was however not observed in CD14+ monocytes (data not shown) and therefore the following experiments were carried out in CD4+ T-cells.

Bottom Line: These results, together with the findings that RU486, a GR antagonist, prevents Dex-, MPA- and Vpr-mediated apoptosis, provide evidence for the first time that GR agonists or partial agonists increase apoptosis in primary CD4(+) T-cells via the GR.This work suggests that contraceptive doses of MPA but not NET-A or physiological doses of progesterone could potentially accelerate depletion of CD4(+) T-cells in a GR-dependent fashion in HIV-1 positive women, thereby contributing to immunodeficiency.The results imply that choice of progestin used in contraception may be critical to susceptibility and progression of diseases such as HIV-1.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cell Biology, University of Cape Town, Cape Town, Western Province, South Africa.

ABSTRACT
The glucocorticoid receptor (GR) regulates several physiological functions, including immune function and apoptosis. The HIV-1 virus accessory protein, viral protein R (Vpr), can modulate the transcriptional response of the GR. Glucocorticoids (GCs) and Vpr have been reported to induce apoptosis in various cells, including T-cells. We have previously shown that the injectable contraceptive, medroxyprogesterone acetate (MPA) is a partial to full agonist for the GR, unlike norethisterone acetate (NET-A). We investigated the functional cross talk between the GR and Vpr in inducing apoptosis in CD4(+) T-cells, in the absence and presence of GCs and these progestins, as well as progesterone. By using flow cytometry, we show that, in contrast to NET-A and progesterone, the synthetic GR ligand dexamethasone (Dex), cortisol and MPA induce apoptosis in primary CD4(+) T-cells. Furthermore, the C-terminal part of the Vpr peptide, or HIV-1 pseudovirus, together with Dex or MPA further increased the apoptotic phenotype, unlike NET-A and progesterone. By a combination of Western blotting, PCR and the use of receptor- selective agonists, we provide evidence that the GR and the estrogen receptor are the only steroid receptors expressed in peripheral blood mononuclear cells. These results, together with the findings that RU486, a GR antagonist, prevents Dex-, MPA- and Vpr-mediated apoptosis, provide evidence for the first time that GR agonists or partial agonists increase apoptosis in primary CD4(+) T-cells via the GR. We show that apoptotic induction involves differential expression of key apoptotic genes by both Vpr and GCs/MPA. This work suggests that contraceptive doses of MPA but not NET-A or physiological doses of progesterone could potentially accelerate depletion of CD4(+) T-cells in a GR-dependent fashion in HIV-1 positive women, thereby contributing to immunodeficiency. The results imply that choice of progestin used in contraception may be critical to susceptibility and progression of diseases such as HIV-1.

Show MeSH
Related in: MedlinePlus