coinciding with biphasic immune reconstitution 146 Opposite effects of antiretroviral treatment on the activation markers CD38 and HLADR on CD8+ T cells, coinciding with biphasic immune reconstitution Enrique Espinosa, Dámaris P. Romero, Ricardo S. Vega-Barrientos, Alvaro Peña-Jiménez, Amy B. Peralta-Prado, M. Teresa Cantoral, Gustavo Reyes-Terán Instituto Nacional de Enfermedades Respiratorias, Mexico City, Mexico1 B Results Abstract Background and hypothesis Lymphocyte activation is involved in HIV disease, and is decreased by antiretroviral treatment. Coexpression of the activation markers CD38 and HLADR on lymphocytes is deemed an inequivocal indicator of activation. However, little is known of the functional role of these molecules in T cells. We hypothesized that differential changes in these markers during antiretroviral therapy (ART) could indicate different roles in the recovery of T cell functionality. Methods.- PBMCs from 36 HIV+ patients with adequate response to ART, participating in a prospective reconstitution study, were taken at weeks 0, 4, 8, 12, 24, 39, 52, and 104 after ART initiation, and analyzed for the expression of CD38 and HLADR on naive, central memory, and effector memory CD8 T cells by flow cytometry. Results.- RM-ANOVA showed a significant treatment effect in the expression of CD38, HLADR, and CD8 and HLADR coexpression in the three subpopulations (p<0.05). The frequencies of cells co-expressing CD38 and HLADR decreased consistently after a 4 week delay, staying lower than baseline (p<0.05). Frequency of cells expressing CD38 but not HLADR decreased significantly and steeply during the first two months of treatment (p<0.05 week 0 vs. Week 4, p<0.05 week 4 vs. week 8, Wilcoxon's), diminishing slowlier after week 8 (p>0.05 week 8 versus week 12), but staying lower than baseline thereafter (p<0.05). In contrast, cells expressing HLADR but not CD38 increased significantly and steeply during the first two treatment months, after which the change rate decreased, with the frequency of HLADR+ cells staying significantly higher than baseline. Conclusion.- Our results suggest that HLADR, as opposed to CD38, may have a role in the functional recovery of CD8 T cells. Figure 4.- Biphasic recuperation of CD4 T cell counts (see Fig. 3) coincides with decreases in CD38 expressing activated CD8 cells, but with a sustained increased in CD8 cells expressing only HLADR 50 50 50 Naive CD8 TC Naive CD8 TC Naive CD8 TC 45 45 45 CM CD8 TC CM CD8 TC CM CD8 TC 40 40 40 EM CD8 TC EM CD8 TC EM CD8 TC 35 35 35 30 30 30 % CD38+ HLADR- % CD38+ HLADR+ % CD38- HLADR+ 25 25 25 20 20 20 15 15 15 10 10 10 5 5 5 4 8 12 24 39 52 104 4 8 12 24 39 52 104 4 8 12 24 39 52 104 Weeks on ART Weeks on ART Weeks on ART IRIS onset time IQR Patients Flow cytometry read-outs HAART-naive adults, no evident inflammation or active OI; informed consent. Follow up and PBMC samples: weeks 0, 4, 8, 12, 24, 39, 52, 104 after ART initiation Figure 5.- Likewise, the dynamics of CD28+ CD8 cells is opposite to that of CD38+ activated CD8 T cells, but is parallel to CD8 T cell subpopulations expressing HLADR only. Activated CD8 TC subpopulations expressing CD38 corelate negatively with CD28 expression throughout reconstitution, but not the subpopulations expressing HLADR only 99 Reconstitution cohort 3 Confirmed basal hepatitis 5 Poor ART response or adherence 4 Lost to follow up 6 Immunomodulants during ART Recovery of CD28+ subpopulations 90 100 100 90 90 80 80 79 ART responders 80 70 70 70 60 60 60 50 % CD28+ Naive CD8 TC at week 8 50 % CD28+ Naive CD8 TC at week 8 50 - 3 Undecidable clinical events % CD28+ 40 40 40 30 30 30 76 Study population 20 20 20 10 10 10 Naive CD8 TC CM CD8 TC EM CD8 TC 5 10 15 20 25 30 35 40 45 50 10 20 30 40 50 60 70 80 4 8 12 24 39 52 104 % CD38+DR- Naive CD8TC at week 8 Weeks on ART % CD38+DR+ Naive CD8TC at week 8 Figure 6.- Tuberculosis-associated immune reconstitution inflammatory syndrome coincides with peak in CD38+ HLADR+ cell frequency, and with an increased frequency of single HLADR+ cells, not with single CD38+ cells. 53 (69.7%) IRIS free 23 (30.3 %) IRIS Gate: EM CD8 T cells Gate: EM CD8 T cells Gate: EM CD8 T cells 45 45 p=0.0474 37.5 IRIS-free 17 matched controls 40 40 16 selected cases IRIS-free * * TB-IRIS 32.5 35 35 p=0.003 TB-IRIS 30 30 27.5 25 25 % CD38- HLADR+ %CD38+ HLADR+ 22.5 % CD38+ HLADR- 20 20 Nested group (33) IRIS-free * 15 15 TB-IRIS 17.5 IRIS-free 10 10 TB-IRIS 12.5 5 5 Same effect in all subpops. Effect not found in naive CD8 TC Same effect in all subpops. 7.5 76 ART good responders in which IRIS was decidable, 92.1% were male, 85% had previous AIDS-related illness. Median nadir CD4= 50.5 cells/ml (22.5-100.3), Log VL=5.528 (5.02-5.87), age= 34.4 (29.9-41.4) years. 90.8% NRTI backbone+EFV , 9.2% NRTI backbone+PI -4 4 8 12 24 39 52 104 4 8 12 24 39 52 104 4 8 12 24 39 52 104 Weeks on ART Weeks on ART Weeks on HAART A particular population of activated CD8 cells, HLADR+ not expressing CD38, increases its frequency with long term immune recovery. Expression of HLADR in activated CD8 T cells coincides both with long term immune reconstitution (HLADR single positive cells), and with transient pathological inflammation during the initial phase of immune recovery (CD38+ HLADR+ and CD38- HLADR). This suggests that HLADR is more compatible with recovered, regulated, functionality. Viremia is controlled CD4 T cell count recovers biphasically 450 400 6 350 5 300 4 CD4 Tcells/ml blood 250 Log Viral load 3 200 150 2 100 1 50 4 8 12 24 39 52 104 Work funded by UC-Mexus, Conacyt grant P 50478 III, Mexico, and by H. Cámara de Diputados, Mexico 4 8 12 24 39 52 104 Weeks on ART Weeks on ART