In utero activation of fetal memory T cells alters host regulatory gene expression and affects HIV susceptibility

Abstract

In utero priming to malaria antigens renders cord blood mononuclear cells (CBMC) more susceptible to productive HIV infection in vitro in the absence of exogenous stimulation. This provides a unique model to better understand mechanisms affecting lymphocyte susceptibility to HIV infection in vivo. Effector memory CD3(+)CD4(+) T cells (T(EM)) were the exclusive initial targets of HIV with rapid spread to central memory cells. HIV susceptibility correlated with increased expression of CD25 and HLA-DR on T(EM). Virus entered all samples equally, however gag/pol RNA was only detected in HIV susceptible samples, suggesting regulation of proviral gene transcription. Targeted analysis of human genes in memory T cells showed greater expression of IFNG, NFATc1, IRF1, FOS, and PPIA and decreased expression YY1 and TFCP2 in HIV susceptible samples. Thus fetal priming to exogenous antigens enhances specific proviral gene transcription pathways in effector memory cells that may increase risk of vertical transmission of HIV.

Figure 1. HIV_BaL minus strand strong-stop DNA is detectable in CD3⁺CD4⁺ cells and absent in non CD3⁺CD4⁺ cells twenty-four hours post virus exposure

HIV –ssDNA normalized to β-globin. Specific conditions indicated in the figure: i) CBMC cultured without virus, ii) Non CD3⁺CD4⁺ and iii) CD3⁺CD4⁺ conditions represent cell fractions following magnetic bead separation twenty-four hours after whole CBMC were exposed to virus, iv) PHA+Virus condition represents whole CBMC cultured in the presence of PHA for three days prior to virus exposure. n= 8 HIV susceptible samples. Lines represent median for each condition.

Figure 2. Frequency of central memory or effector memory CD4⁺ cell subsets does not significantly vary between HIV susceptible, HIV not susceptible, and North American samples

Frequency of CD45RO⁺CD27⁺ (central memory) and CD45RO⁺CD27⁻ (effector memory) cells were determined after gating on viable CD3⁺CD4⁺ lymphocytes. Lines represent median values. N=8 for each group.

Figure 3. Expression of activation markers on T_CM and T_EM cells correlates with HIV susceptibility

Integrated mean fluorescent indices (iMFI) for CD25 (top) and HLA-DR (bottom) gated on viable CD3⁺CD4⁺ lymphocytes and appropriate CD45RO and CD27 expression. N=6 for each group. Lines indicate geometric mean of iMFI. Statistics calculated by t test.

Figure 4. Detection of minus strand strong-stop DNA is similar between HIV susceptible and HIV not susceptible CBMC twenty-four hours post virus exposure, but gag/pol RNA is only detected in HIV susceptible CBMC

A) HIV –ssDNA normalized to β-globin. Sample sizes: HIV not susceptible, n=31; HIV susceptible, n=8. B) gag/pol RNA normalized to R18. Sample sizes: HIV not susceptible, n=13; HIV susceptible, n=8. Lines indicate median.

Figure 5. Differential expression of host genes in total memory CD3⁺CD4⁺ cells from HIV susceptible, HIV not susceptible and North American CBMC

A: HIV susceptible vs North American. B: HIV not susceptible vs North American. C: HIV susceptible vs HIV not susceptible. N=4 for all groups. * p=0.07-0.14, ** p=0.05.