201143a  (fluidigm)

Journal: Cell Reports Medicine

Article Title: BACH2 promotes seeding and establishment of long-lived HIV-1 reservoir in memory CD4 + T cells

doi: 10.1016/j.xcrm.2025.102311

Figure Lengend Snippet: Memory differentiation of CCR5 + CD4 + T cells is impeded by heightened inflammation (A) Sample collection scheme. (B) Plasma IL-18 concentration determined by ELISA. Indicated pre- and post-ART samples from 83 study participants were included. (C and D) IL-18Rα expression in activated CD4 + T cells. CD4 + T cells from four healthy blood donors were used. (E and F) Activated CCR5 + cells were purified as described in A and were further differentiated under the Th1 polarization condition with or without the presence of IL-18 (5 ng/mL). BACH2 transcripts were measured by RT-qPCR and normalized by POLR2A . Conversion to MP cells was determined by flow cytometry. CD4 + T cells from seven healthy blood donors were used. (G–L) The impact of ART on CD4 recovery (G), TCF1 expression (H–K), and BACH2 transcription (L). The percentage of CD4 + T cells and TCF1 expression in total and memory (CD45RA − ) CD4 + T cells from 10 PLWH were analyzed by flow cytometry. BACH2 transcripts in CD4 + T cells from five PLWH were measured by qPCR. Error bars indicate mean standard error of the mean (SEM). (M and N) The role of BACH2 in HIV-1 transcription in latent reservoir cells. CD4 + T cells from eight PLWH on suppressive ART were collected to generate Cas9 ctrl and BACH2 -KO cells. After electroporation, cells were rested for 3 days before treatment with either PBS or anti-CD3 and anti-CD28 antibodies for 24 h. Cellular RNA was extracted to measure cell-associated HIV-1 RNA. In (G), (J), (K), and (L), p values were calculated using the one-way ANOVA and Holm-Sidak multiple comparisons test. Other p values were calculated using the paired t test. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001.

Article Snippet: Anti-human CD45RA −143ND , Fluidigm , Cat# 3143006B; RRID: AB_2651156.

Techniques: Clinical Proteomics, Concentration Assay, Enzyme-linked Immunosorbent Assay, Expressing, Purification, Quantitative RT-PCR, Flow Cytometry, Electroporation

Journal: Cell Reports Medicine

Article Title: BACH2 promotes seeding and establishment of long-lived HIV-1 reservoir in memory CD4 + T cells

doi: 10.1016/j.xcrm.2025.102311

Figure Lengend Snippet: Generation of BACH2 -KO humanized mice for HIV-1 infection (A) Strategy for generating BACH2 -edited humanized mice. (B) Human immune cell reconstitution and T cell development in mice engrafted with BACH2 -KO or Cas9 ctrl CD34 + cells. Blood samples were collected 10–11 weeks after transplantation. (C and D) Human T cell and macrophage development in mice engrafted with BACH2 -KO or Cas9 ctrl CD34 + cells. Spleen samples were collected 12 weeks after transplantation. The total naive (CD45RA + CCR7 + ), central memory (CD45RA − CCR7 + ), effector memory (CD45RA − CCR7 − ), and regulatory (CD25 + Foxp3 + ) CD4 + T cells as well as macrophages were determined by flow cytometry. (E and F) HIV-1 infection in mice engrafted with BACH2 -KO or Cas9 ctrl CD34 + cells. Plasma viral loads (E) and loss of CD4 + T cells (F) were determined at indicated time points. (G–I) Quantification of HIV-1 infection. The frequency of p24 + of the CD3 + CD8 − population was measured by flow cytometry. The frequency of HIV-1 DNA + cells was determined by qPCR. In (F), p values were calculated using the two-way ANOVA and Holm-Sidak multiple comparisons test. Other p values were calculated using the unpaired t test. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001.

Article Snippet: Anti-human CD45RA −143ND , Fluidigm , Cat# 3143006B; RRID: AB_2651156.

Techniques: Infection, Transplantation Assay, Flow Cytometry, Clinical Proteomics

Journal: Cell Reports Medicine

Article Title: BACH2 promotes seeding and establishment of long-lived HIV-1 reservoir in memory CD4 + T cells

doi: 10.1016/j.xcrm.2025.102311

Figure Lengend Snippet: BACH2 is required for the seeding and establishment of long-lived HIV-1 reservoir in memory CD4 + T cells (A) HIV-1 infection and ART in the control and BACH2 -KO groups. ART was provided 2–3 weeks post HIV BaL infection. Plasma HIV-1 RNA levels determined by RT-qPCR. Cas9 ctrl, n = 10; BACH2 -KO, n = 9. (B–D) Viral rebound. CD8-depleted total splenocytes from ART-treated mice are adoptively transferred into the HIV-1-negative unengrafted MISTRG-6-15 mice. Each recipient mouse receives the whole splenocyte population from one ART mice. Blood samples were collected once per week to monitor virus rebound. Virus rebound is confirmed when plasma viral load is >1,000 copies per mL. Cas9 ctrl, n = 10; BACH2 -KO, n = 14. In (D), time to rebound is set at 12 weeks (open circles) for mice without viral rebound. (E and F) Frequency of HIV-1 DNA + cells in tissues from aviremic mice on suppressive ART. In (F), T CM cells (CD3 + CD4 + CD45RA − CCR7 + ), non-T CM cells (CD3 + CD4 + CD45RA − CCR7 − ), and macrophages (CD14 + ) were purified by sorting before HIV-1 DNA measurement. Open circle: undetected. In (E), Cas9 ctrl, n = 10; BACH2 -KO, n = 14. In (F), Cas9 ctrl, n = 10; BACH2 -KO, n = 9. In (C), the p value was calculated by the log rank (Mantel-Cox) test. Other p values were calculated using the unpaired t test. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001.

Article Snippet: Anti-human CD45RA −143ND , Fluidigm , Cat# 3143006B; RRID: AB_2651156.

Techniques: Infection, Control, Clinical Proteomics, Quantitative RT-PCR, Virus, Purification