|
|
Blood, Vol. 109, Issue 6, 2505-2513, March 15, 2007
Increased mitochondrial mass characterizes the survival defect of HIV-specific CD8+ T cells
Blood Petrovas et al.
109: 2505
Supplemental materials for: Petrovas et al, Vol 109, Issue 6, 2505-2513
Files in this Data Supplement:
- Figure S1. HIV-specific CD8+ T cells exhibit increased NAO staining (PDF, 39.7 KB) -
(A) Representative flow cytometry showing ex vivo NAO staining in HIV-specific and CD8+ T cells from the same HIV+ patient and an HIV– donor. (B) Pooled data showing the percentages (%) of CD8+ T cells expressing high NAO levels in HIV-specific (n=4) and total CD8+ T cells from HIV+ (n=5) and healthy (n=3) donors.
- Figure S2. Mitochondrial mass is high in both early apoptotic (annexin V+ViVi–) and late apoptotic/dead (annexin V+ViVi+) CD8+ T cells and HIV-specific CD8+ T cells (PDF, 53.7 KB) -
Freshly isolated PBMCs from HIV-infected individuals were cultured either untreated or in the presence of 5 µg/mL plate-bound anti-CD95 antibody for 6 hours, harvested, stained, and analyzed on a FACSAria flow cytometry instrument. To distinguish between early apoptosis (intact plasma membrane) and late apoptosis (disrupted plasma membrane), annexin V (BD Bioscience, San Jose, CA; AnnV) and Violet Live/Dead Fixable Dead Cell Stain (Invitrogen, Carlsbad, CA; ViVi) were used. (A-D) Representative FACS plots of PBMCs from 1 HIV-infected individual are shown. For panels A and B, lymphocytes were gated using FSC and SSC and then total CD8+ T cells were gated using annexin V and ViVi. (A-B) Mitochondrial mass in live, early apoptotic, and late apoptotic/dead populations of total CD8+ T cells. FACS plot depicting live cells (AnnV–ViVi–), early apoptotic (AnnV+ViVi–), and late apoptotic/dead (AnnV+ViVi+) total CD8+ T cells in untreated (A) and anti-CD95–treated (B) PBMCs after 6-hour cultivation. (Right panel) FACS plots show MitoTracker staining in these subsets of total CD8+ T cells. For panels C and D, lymphocytes were gated using FSC and SSC then CD8 and HLA-A2–Gag tetramers. (C-D) FACS plot depicts live (AnnV–ViVi–), early apoptotic (AnnV+ViVi–), and late apoptotic/dead (AnnV+ViVi+) Gag-specific CD8+ T cells either untreated (C) or anti-CD95 treated (D) for 6 hours. (Right panel) FACS plots show MitoTracker staining for live, early apoptotic, and late apoptotic/dead Gag-specific CD8+ T cells. (E) Pooled data (n=6) showing mean + SE of percentage of high and low MitoTracker cells in live, early apoptotic, and late apoptotic/dead total CD8+ T cells after 6-hour anti-CD95 antibody stimulation. Student t test and Shapiro-Wilk W test for normality were used for statistical analysis with the JMP statistical analysis program (SAS, Cary, NC). P values less than .05 were considered significant.
|
|
