Homeostatic Expansion of CD4+ T Cells Promotes Cortical and Trabecular Bone Loss, Whereas CD8+ T Cells Induce Trabecular Bone Loss Only.
ABSTRACT: Background:Bone loss occurs in human immunodeficiency virus (HIV) infection but paradoxically is intensified by HIV-associated antiretroviral therapy (ART), resulting in an increased fracture incidence that is largely independent of ART regimen. Inflammation in the bone microenvironment associated with T-cell repopulation following ART initiation may explain ART-induced bone loss. Indeed, we have reported that reconstitution of CD3+ T cells in immunodeficient mice mimics ART-induced bone loss observed in humans. In this study, we quantified the relative effects of CD4+ and CD8+ T-cell subsets on bone. Methods:T-cell subsets in T-cell receptor ? knockout mice were reconstituted by adoptive transfer with CD4+ or CD8+ T-cells subsets were reconstituted in T-cell receptor ? knockout mice by adoptive transfer, and bone turnover, bone mineral density, and indices of bone structure and turnover were quantified. Results:Repopulating CD4+ but not CD8+ T cells significantly diminished bone mineral density. However, micro-computed tomography revealed robust deterioration of trabecular bone volume by both subsets, while CD4+ T cells additionally induced cortical bone loss. Conclusions:CD4+ T-cell reconstitution, a key function of ART, causes significant cortical and trabecular bone loss. CD8+ T cells may further contribute to trabecular bone loss in some patients with advanced AIDS, in whom CD8+ T cells may also be depleted. Our data suggest that bone densitometry used for assessment of the condition of bone in humans may significantly underestimate trabecular bone damage sustained by ART.
Project description:<h4>Background</h4>Although effective antiretroviral therapy(ART) increases CD4+ T-cell count, responses to ART vary considerably and only a minority of patients normalise their CD4+/CD8+ ratio. Although retention of naïve CD4+ T-cells is thought to predict better immune responses, relationships between CD4+ and CD8+ T-cell subsets and CD4+/CD8+ ratio have not been well described.<h4>Methods</h4>A cross-sectional study in a cohort of ambulatory HIV+ patients. We used flow cytometry on fresh blood to determine expanded CD4+ and CD8+ T-cell subsets; CD45RO+CD62L+(central memory), CD45RO+CD62L-(effector memory) and CD45RO-CD62L+(naïve) alongside routine T-cell subsets(absolute, percentage CD4+ and CD8+ counts), HIVRNA and collected demographic and treatment data. Relationship between CD4+/CD8+ T-cell ratio and expanded T-cell subsets was determined using linear regression analysis. Results are median[IQR] and regression coefficients unless stated.<h4>Results</h4>We recruited 190 subjects, age 42(36-48) years, 65% male, 65.3% Caucasian, 91% on ART(52.6% on protease inhibitors), 78.4% with HIVRNA<40cps/ml and median ART duration 6.8(2.6-10.2) years. Nadir and current CD4+ counts were 200(112-309) and 465(335-607) cells/mm3 respectively. Median CD4+/CD8+ ratio was 0.6(0.4-1.0), with 26.3% of subjects achieving CD4+/CD8+ ratio>1. Of the expanded CD4+ T-cell subsets, 27.3(18.0-38.3)% were naïve, 36.8(29.0-40.0)% central memory and 27.4(20.0-38.5)% effector memory. Of the CD8+ T-cells subsets, 16.5(10.2-25.5)% were naïve, 19.9(12.7-26.6)% central memory and 41.0(31.8-52.5)% effector memory. In the multivariable adjusted analysis, total cumulative-ART exposure(+0.15,p?=?0.007), higher nadir CD4+ count(+0.011,p<0.001) and higher %CD8+ naive T-cells(+0.0085,p<0.001) were associated with higher CD4+/CD8+ ratio, higher absolute CD8+ T-cell(-0.0044,p<0.001) and higher %CD4+ effector memory T-cells(-0.004,p?=?0.0036) were associated with lower CD4+/CD8+ ratio. Those with CD4+/CD8+ ratio>1 had significantly higher median %CD8+ naive T-cells; 25.4(14.0-36.0)% versus 14.4(9.4-21.6)%, p<0.0001, but significantly lower absolute CD8+ count; 464(384.5-567) versus 765(603-1084) cells/mm3, p<0.001.<h4>Conclusions</h4>Study suggests important role for naïve CD8+ T-cell populations in normalisation of the immune response to HIV-infection. How these findings relate to persistent immune activation on ART requires further study.
Project description:<h4>Objective</h4>To estimate the prognostic value of T-cell subsets in Zambian patients initiating antiretroviral therapy (ART), and to assess the impact of a nutritional intervention on T-cell subsets.<h4>Methods</h4>This was a sub-study of a randomised clinical trial of a nutritional intervention for malnourished adults initiating ART. Participants in a randomised controlled trial (NUSTART trial) were enrolled between April and December 2012. Participants received lipid-based nutritional supplement either with or without additional vitamins and minerals. Immunophenotyping was undertaken at baseline and, in survivors, after 12 weeks of ART to characterize T-cell subsets using the markers CD3, CD4, CD8, CD45RA, CCR7, CD28, CD57, CD31, ?4?7, Ki67, CD25 and HLA-DR. Univariate and multivariate survival analysis was performed, and responses to treatment were analysed using the Wicoxon rank-sum test.<h4>Results</h4>Among 181 adults, 36 (20%) died by 12 weeks after starting ART. In univariate analysis, patients who died had fewer proliferating, more naïve and fewer gut homing CD4+ T-cells compared to survivors; and more senescent and fewer proliferating CD8+ T-cells. In a multivariate Cox regression model high naïve CD4+, low proliferating CD4+, high senescent CD8+ and low proliferating CD8+ subsets were independently associated with increased risk of death. Recent CD4+ thymic emigrants increased less between recruitment and 12 weeks of ART in the intervention group compared to the control group.<h4>Conclusions</h4>Specific CD4+ T-cell subsets are of considerable prognostic significance for patients initiating ART in Zambia, but only thymic output responded to this nutritional intervention.
Project description:While it is known that irradiation can induce local and systemic bone loss over time, how focal irradiation induces systemic bone complications remains unclear. Immune cells are thought to be crucial to bone homeostasis, and abnormal immune cells lead to serious disruption of bone homeostasis, such as in acute lymphoblastic leukaemia. This disruption primarily occurs due to inhibition of the osteogenic differentiation of bone mesenchymal stem cells (BMSCs).In this study, we detected local and systemic bone loss in trabecular bone by micro-computed tomography (micro-CT) and measurement of peroxisome proliferator-activated receptor gamma (PPAR?) and runt-related transcription factor 2 (RUNX2) expression in BMSCs using real-time polymerase chain reaction and western blotting. Additionally, changes in lymphocytes (B cells and CD4+ and CD8+ T cells) in the peripheral blood and bone marrow were analysed by flow cytometry. BMSC-derived osteoblasts and adipocytes, cultured in osteogenic or adipogenic media or co-cultured with lymphocytes, were detected by BCIP/NBT, Alizarin Red S and Oil Red O staining.Focal irradiation induced local and systemic bone loss in trabecular bone. Increased PPAR? expression and decreased RUNX2 expression were observed, accompanied by upregulated adipogenesis and downregulated osteogenesis of BMSCs. B cells and CD8+ T lymphocytes were increased in the blood and bone marrow after irradiation, while CD4+ T lymphocytes were decreased in the blood. Inhibition of RUNX2 expression and reduction of alkaline phosphatase activity and mineralization deposits were observed in lymphocyte-co-cultured BMSCs, accompanied by an increase in PPAR? expression and in the number of lipid droplets.Focal irradiation induced local and systemic bone loss in trabecular bone. Increased B cells and CD8+ T lymphocytes led to systemic bone loss by decreasing BMSC osteogenesis.
Project description:Despite improvement in the prognosis of HIV/AIDS (human immunodeficiency virus/acquired immune deficiency syndrome) patients on antiretroviral therapy (ART), cryptococcal meningitis (CM) still causes 10-15% mortality among HIV-infected patients. The immunological impact of ART on the CD4+ and CD8+ T cell repertoire during cryptococcal co-infection is unclear. We determined longitudinal phenotypic changes in T cell subsets among patients with CM after they initiated ART. We hypothesized that ART alters the clonotypic phenotype and structural composition of CD4+ and CD8+ T cells during CM co-infection. For this substudy, peripheral blood mononuclear cells (PBMC) were isolated at four time points from CM patients following ART initiation during the parent study (ClinicalTrials.gov number, NCT01075152). Phenotypic characterization of CD4+ and CD8+ T cells was done using T cell surface marker monoclonal antibodies by flow cytometry. There was variation in the expression of immunophenotypic markers defining central memory (CD27+CD45R0+), effector memory (CD45R0+CD27-), immune activation (CD38+ and Human Leucocyte Antigen DR (HLA-DR+), and exhaustion (Programmed cell death protein one (PD-1) in the CD4+ T cell subset. In comparison to the CD4+ T cell population, the CD8+ central memory subset declined gradually with minimal increase in the effector memory subset. Both CD4+ and CD8+ T cell immune exhaustion and activation markers remained elevated over 12 weeks. The relative surge and decline in the expression of T cell surface markers outlines a variation in the differentiation of CD4+ T cells during ART treatment during CM co-infection.
Project description:Understanding the cytokine/chemokine networks in CD4(+) and CD8(+) T cells during the acute phase of infection is crucial to design therapies for the control of early human immunodeficiency virus (HIV)/simian immunodeficiency virus (SIV) replication. Here, we measured early changes in CD4(+) and CD8(+) T cells in the peripheral blood (PB), bone marrow (BM), and axillary lymph node (ALN) tissue of rhesus macaques infected with SIVMAC251. At 21 days after infection, all tissues showed a statistically significant loss of CD4(+) T cells along with immune activation of CD8(+) T cells in PB and ALN tissue. Twenty-eight different cytokines/chemokines were quantified in either anti-CD3/28 antibody- or staphylococcal enterotoxin B-stimulated single-positive CD4(+) and CD8(+) T cells. PB CD4(+) T cells produced predominantly interleukin-2 (IL-2), whereas CD4(+) and CD8(+) T-cell subsets in tissues produced ?-chemokines both before and 21 days after SIV infection. Tissues generally exhibited massive upregulation of many cytokines/chemokines following infection, possibly in an attempt to mitigate the loss of CD4(+) T cells. There was no evidence of a T-helper 1 (TH1)-to-TH2 shift in CD4(+) T cells or a T-cytotoxic 1 (TC1)-to-TC2 cytokine shift in CD8(+) T cells in PB, BM, and ALN T-cell subsets during the acute phase of SIV infection. Despite the upregulation of several important effector cytokines/chemokines (IL-2, IL-12, IL-17, gamma interferon, granulocyte-macrophage colony-stimulating factor) by CD4(+) and CD8(+) T cells, upregulation of ?-chemokines (CCL2 and CCL22), basic fibroblast growth factor (FGF-basic), hepatocyte growth factor (HGF), and migration inhibition factor (MIF) may provide a poor prognosis either by inducing increased virus replication or by other unknown mechanisms. Therefore, drugs targeting ?-chemokines (CCL2 and CCL22), FGF-basic, HGF, or MIF might be important for developing effective vaccines and therapeutics against HIV.Human immunodeficiency virus (HIV)/simian immunodeficiency virus (SIV) infection results in early depletion of CD4(+) T cells and dysregulation of protective immune responses. Therefore, understanding the cytokine/chemokine networks in CD4(+) and CD8(+) T cells in different tissues during the acute phase of infection is crucial to the design of therapies for the control of early viral replication. Here, we measured early changes in CD4(+) and CD8(+) T cells in peripheral blood (PB), bone marrow (BM), and axillary lymph node (ALN) tissue of rhesus macaques infected with SIVMAC251. There was no evidence of a T-helper 1 (TH1)-to-TH2 shift in CD4(+) T cells or a T-cytotoxic 1 (TC1)-to-TC2 cytokine shift in CD8(+) T cells in PB, BM, and ALN T-cell subsets during the acute phase of SIV infection. Despite the upregulation of several important effector cytokines/chemokines by CD4(+) and CD8(+) T cells, upregulation of ?-chemokines, fibroblast growth factor-basic, hepatocyte growth factor, and migration inhibition factor may provide a poor prognosis.
Project description:Antiretroviral therapy (ART) is generally prescribed to patients with human immunodeficiency virus (HIV) infection with vaccination introduced to prevent disease complications. However, little is known about the influence of immunization on T cell subsets' distribution during the course of infection. This study aims to identify the impact of viral replication and immunization on naïve, effector, effector memory, and central memory T cell subpopulations in ART-treated HIV-infected children. Fifty patients were recruited and injected intramuscularly with influenza A (H1N1) 2009 vaccine on the day of enrollment (day 0) and day 28. Blood samples were collected for pre- and postvaccination on days 0 and 56 for analyzing T cell phenotypes by flow cytometry. Phenotypes of all T cell subsets remained the same after vaccination, except for a reduction in effector CD8+ T cells. Moreover, T cell subsets from patients with controllable viral load showed similar patterns to those with virological failure. Absolute CD4 count was also found to have a positive relationship with naïve CD4+ and CD8+ T cells. In conclusion, vaccination and viral replication have a little effect on the distribution of T cell subpopulations. The CD4 count can be used for prediction of naïve T cell level in HIV-infected patients responding to ART.
Project description:Osteoporotic hip fracture is associated with significant trabecular bone loss, which is typically characterized as low bone density by dual-energy X-ray absorptiometry (DXA) and altered microstructure by micro-computed tomography (μCT). Emerging morphological analysis techniques, e.g. individual trabecula segmentation (ITS), can provide additional insights into changes in plate-like and rod-like trabeculae, two major microstructural types serving different roles in determining bone strength. Using ITS, we evaluated trabecular microstructure of intertrochanteric bone cores obtained from 23 patients undergoing hip replacement surgery for intertrochanteric fracture and 22 cadaveric controls. Micro-finite element (μFE) analyses were performed to further understand how the abnormalities seen by ITS might translate into effects on bone strength. ITS analyses revealed that, near fracture site, plate-like trabeculae were seriously depleted in fracture patients, but trabecular rod volume was maintained. Besides, decreased plate area and rod length were observed in fracture patients. Fracture patients also showed decreased elastic moduli and shear moduli of trabecular bone. These results provided evidence that in intertrochanteric hip fracture, preferential loss of plate-like trabeculae led to more rod-like microstructure and deteriorated mechanical competence adjacent to the fracture site, which increased our understanding of the biomechanical pathogenesis of hip fracture in osteoporosis.
Project description:HIV infection causes bone loss. We previously reported that immunosuppression-mediated B-cell production of receptor activator of NF-?B ligand (RANKL) coupled with decline in osteoprotegerin correlate with decreased bone mineral density (BMD) in untreated HIV infection. Paradoxically, antiretroviral therapy (ART) worsens bone loss although existing data suggest that such loss is largely independent of specific antiretroviral regimen. This led us to hypothesize that skeletal deterioration following HIV disease reversal with ART may be related to T-cell repopulation and/or immune reconstitution. Here we transplant T cells into immunocompromised mice to mimic ART-induced T-cell expansion. T-cell reconstitution elicits RANKL and TNF? production by B cells and/or T cells, accompanied by enhanced bone resorption and BMD loss. Reconstitution of TNF?- or RANKL-null T-cells and pharmacological TNF? antagonist all protect cortical, but not trabecular bone, revealing complex effects of T-cell reconstitution on bone turnover. These findings suggest T-cell repopulation and/or immune reconstitution as putative mechanisms for bone loss following ART initiation.
Project description:OBJECTIVES:To evaluate the immunological and viral consequences of planned treatment interruptions (PTI) in children with HIV. DESIGN:This was an immunological and virological sub-study of the Paediatric European Network for Treatment of AIDS (PENTA) 11 trial, which compared CD4-guided PTI of antiretroviral therapy (ART) with continuous therapy (CT) in children. METHODS:HIV-1 RNA and lymphocyte subsets, including CD4 and CD8 cells, were quantified on fresh samples collected during the study; CD45RA, CD45RO and CD31 subpopulations were evaluated in some centres. For 36 (18 PTI, 18 CT) children, immunophenotyping was performed and cell-associated HIV-1 DNA analysed on stored samples to 48 weeks. RESULTS:In the PTI group, CD4 cell count fell rapidly in the first 12 weeks off ART, with decreases in both naïve and memory cells. However, the proportion of CD4 cells expressing CD45RA and CD45RO remained constant in both groups. The increase in CD8 cells in the first 12 weeks off ART in the PTI group was predominantly due to increases in RO-expressing cells. PTI was associated with a rapid and sustained increase in CD4 cells expressing Ki67 and HLA-DR, and increased levels of HIV-1 DNA. CONCLUSIONS:PTI in children is associated with rapid changes in CD4 and CD8 cells, likely due to increased cell turnover and immune activation. However, children off treatment may be able to maintain stable levels of naïve CD4 cells, at least in proportion to the memory cell pool, which may in part explain the observed excellent CD4 cell recovery with re-introduction of ART.
Project description:Bone tissues have trabecular bone with a high bone turnover and cortical bone with a low turnover. The mechanisms by which the turnover rate of these bone tissues is determined remain unclear. Osteocytes secrete sclerostin, a Wnt/?-catenin signaling antagonist, and inhibit bone formation. We found that sclerostin expression in cortical bone is more marked than in trabecular bone in Sost reporter mice. Leukemia inhibitory factor (LIF) secreted from osteoclasts reportedly suppressed sclerostin expression and promoted bone formation. Here, we report that osteoclasts downregulate sclerostin expression in trabecular bone and promote bone turnover. Treatment of C57BL/6 mice with an anti-RANKL antibody eliminated the number of osteoclasts and LIF-positive cells in trabecular bone. The number of sclerostin-positive cells was increased in trabecular bone, while the number of ?-catenin-positive cells and bone formation were decreased in trabecular bone. Besides, Tnfsf11 heterozygous (Rankl+/-) mice exhibited a decreased number of LIF-positive cells and increased number of sclerostin-positive cells in trabecular bone. Rankl+/- mice exhibited a decreased number of ?-catenin-positive cells and reduced bone formation in trabecular bone. Furthermore, in cultured osteoclasts, RANKL stimulation increased Lif mRNA expression, suggesting that RANKL signal increased LIF expression. In conclusion, osteoclasts downregulate sclerostin expression and promote trabecular bone turnover.