Explore the vast range of titles available.

Background. Individuals infected with human immunodeficiency virus (HIV) have higher risk than HIV-negative individuals for diseases associated with aging. T-cell senescence, characterized by expansion of cells lacking the costimulatory molecule CD28, has been hypothesized to mediate these risks. Methods. We measured the percentage of CD28 - CD4 + and CD8 + T cells from HIV-infected treatment-naive adults from 5 Adult Clinical Trials Group (ACTG) antiretroviral therapy (ART) studies and the ALLRT (ACTG Longitudinal Linked Randomized Trials) cohort, and from 48 HIV-negative adults. Pretreatment and 96-week posttreatment %CD28 - cells were assessed using linear regression for associations with age, sex, race/ethnicity, CD4 count, HIV RNA, ART regimen, and hepatitis C virus (HCV) infection. Results. In total, 1291 chronically HIV-infected adults were studied. Pretreatment, lower CD4 count was associated with higher %CD28 - CD4 + and %CD28 - CD8 + cells. For CD8 + cells, younger age and HCV infection were associated with a lower %CD28 - . ART reduced %CD28 - levels at week 96 among virally suppressed individuals. Older age was strongly predictive of higher %CD28 - CD8 + . Compared to HIV-uninfected individuals, HIV-infected individuals maintained significantly higher %CD28 - . Conclusions. Effective ART reduced the proportion of CD28 - T cells. However, levels remained abnormally high and closer to levels in older HIV-uninfected individuals. This finding may inform future research of increased rates of age-associated disease in HIV-infected adults.

Background. Individuals infected with human immunodeficiency virus (HIV) have increased risk of cardiovascular events. It is unknown whether T cell activation and senescence, 2 immunologic sequelae of HIV infection, are associated with vascular disease among HIV-infected adults. Methods. T cell phenotyping and carotid ultrasound were assessed among 115 HIV-infected women and 43 age- and race/ethnicity-matched HIV-uninfected controls participating in the Women's Interagency HIV Study. Multivariate analyses were used to assess the association of T cell activation (CD38⁺HLA-DR⁺) and senescence (CD28⁻CD57⁺) with subclinical carotid artery disease. Results. Compared with HIV-uninfected women, frequencies of CD4⁺CD38⁺HLA-DR⁺, CD8⁺CD38⁺HLA-DR⁺, and CD8⁺CD28⁻CD57⁺ T cells were higher among HIV-infected women, including those who achieved viral suppression while receiving antiretroviral treatment. Among HIV-infected women, adjusted for age, antiretroviral medications, and viral load, higher frequencies of activated CD4⁺ and CD8⁺ T cells and immunosenescent CD8⁺ T cells were associated with increased prevalence of carotid artery lesions (prevalence ratio lesions associated with activated CD4⁺ T cells, 1.6 per SD [95% confidence interval {CI}, 1.1–2.2]; P = .02; prevalence ratio lesions associated with activated CD8⁺ T cells, 2.0 per SD [95% CI, 1.2–3.3]; P < .01; prevalence ratio lesions associated with senescent CD8⁺ T cells, 1.9 per SD [95% CI, 1.1–3.1]; P = .01). Conclusions. HIV-associated T cell changes are associated with subclinical carotid artery abnormalities, which may be observed even among those patients achieving viral suppression with effective antiretroviral therapy.

The role of CD8(+) T lymphocytes in chronic hepatitis C virus (HCV) infection and in liver injury with subsequent development of fibrosis and cirrhosis is poorly understood. To address this question, we performed a follow-up study including 27 chronically HCV-infected individuals. We determined clonality and phenotypes of circulating CD8(+) T cells employing TCRBV spectratyping. Antigen specificity was tested by rMHC-peptide tetramer staining and stimulation with recombinant HCV antigens. In addition, T-cell clonality and phenotypes were followed during the variable clinical response of interferon- (IFN) alpha treatment. We could demonstrate that CD8(+) T-cell expansions were significantly associated with liver fibrosis and cirrhosis. Likewise, increased oligoclonality of circulating CD8(+) T cells in chronic HCV infection was identified as an indicator for poor clinical response to IFN-alpha therapy. Moreover, we also found that IFN-alpha therapy enhanced the differentiation of CD8(+) T cells towards a late differentiation phenotype (CD28(-) CD57(+)). In cases of virus elimination the disappearance of expanded terminally differentiated CD8(+) cells was observed. Thus, this study identifies an association of clonal expansions of circulating CD8(+) T cells with liver pathology and provides a possible explanation for the fact that response to IFN-alpha therapy diminishes with the duration of infection.

Abstract Background Cancer immunotherapy research, immune microenvironment exploration, and biomarker discovery are key application areas of immune checkpoint analysis. PD-1 and CD28 are crucial receptors expressed on the surface of T cells, playing vital roles in regulating T cell activation and immune response. Accurate detection of these immune checkpoints, such as PD-1 and CD28 on lymphocytes, is essential for understanding immune responses, particularly in clinical contexts such as cancer immunotherapy. Flow cytometry offers a precise approach to detect these markers in whole blood samples. Methods This study developed and optimized a flow cytometry-based detection method utilizing the CYTEK NL-CLC flow cytometer to quantitatively assess the expression of PD-1 and CD28 on lymphocytes. A detailed protocol was established and validated, focusing on key performance parameters. Results Our method showed high sensitivity and specificity, providing a powerful tool for immune monitoring and treatment decision-making. Validation results, including precision, dilution linearity, fluorescence stability, reference interval, and accuracy, all met acceptable criteria and have been reviewed and approved for clinical testing. Conclusions The CYTEK NL-CLC flow cytometer is positioned as a reliable and effective platform for immune checkpoint analysis in both clinical and research settings, supporting its integration into cancer immunotherapy workflows and personalized medicine strategies.

Background Adaptive immune cells, including CD4+CD69+ and CD4+CD25+FoxP3+ regulatory T (Treg) cells, are important for maintaining immunological tolerance. In human systemic lupus erythematosus (SLE), CD4+CD25+FoxP3+ Treg cells are reduced, whereas CD69 expression is increased, resulting in a homeostatic immune imbalance that may intensify autoreactive T cell activity. To analyze the mechanisms implicated in autotolerance failure, we evaluated CD4+CD69+ and CD4+CD25+FoxP3+ T cells and interleukin profiles in a pristane-induced SLE experimental model. Methods For lupus induction, 26 female Balb/c mice received a single intraperitoneal 0.5 ml dose of pristane, and 16 mice received the same dose of saline. Blood and spleen samples were collected from euthanized mice 90 and 120 days after pristane or saline inoculation. Mononuclear cells from peripheral blood (PBMC), peritoneal lavage (PL) and splenocytes were obtained by erythrocyte lysis and cryopreserved for further evaluation by flow cytometry using the GuavaEasyCyte TM HT. After thawing, cells were washed and stained with monoclonal antibodies against CD3, CD4, CD8, CD25, CD28, CD69, FoxP3, CD14 and Ly6C (BD Pharmingen TM). Interleukins were quantified using Multiplex® MAP. The Mann-Whitney test and the Pearson coefficient were used for statistical analysis, and p < 0.05 considered significant. Results Compared with the controls, SLE-induced animals presented increased numbers of CD4+CD69+ T cells in the blood on T90 and T120 (p = 0.022 and p = 0.008) and in the spleen on T120 (p = 0.049), but there were decreased numbers in the PL (p = 0.049) on T120. The percentage of Treg was lower in blood (p < 0.005 and p < 0.012) on T90 and T120, in spleen (p = 0.043) on T120 and in PL (p = 0.001) on T90. Increased numbers of CD4 + CD69+ T cells in the PL were positively associated with high IL-2 (p = 0.486) and IFN-γ (p = 0.017) levels, whereas reduced Treg cells in the blood were negatively correlated with TNFα levels (p = 0.043) and positively correlated with TGFβ1 (p = 0.038). Conclusion Increased numbers of CD4+CD69+ T cells and reduced numbers of CD4+CD25+FoxP3+ Treg cells with an altered interleukin profile suggests loss of autotolerance in pristane-induced lupus mice, which is similar to human lupus. Therefore, this model is useful in evaluating mechanisms of cellular activation, peripheral tolerance and homeostatic immune imbalance involved in human SLE.

Background Although therapeutic human papillomavirus vaccines could offer a noninvasive treatment for patients with cervical intraepithelial neoplasia, none has been clinically implemented. Oral administration of the therapeutic human papillomavirus vaccine IGMKK16E7 results in the histological regression of human papillomavirus 16–positive cervical intraepithelial neoplasia 2/3 to normal (complete response). We investigated biomarkers that could predict complete response after oral administration of IGMKK16E7. Methods Forty-two patients administered high-dose oral IGMKK16E7 in a phase I/II trial were included. Cervix-exfoliated cells were collected before vaccine administration. Gene expression of CD4, CD8, FOXP3, programmed cell death 1 protein, CTLA4, CD103, CD28, CD80, CD86, and programmed cell death 1 ligand 1 in the cells was measured by quantitative reverse transcriptase–polymerase chain reaction. Receiver operating characteristic curve analysis and Mann-Whitney tests were used to explore potential biomarkers. Pearson correlation coefficient analysis was used to correlate gene expression profiles with clinical outcome. Results The only predictive biomarker of vaccine response for which receiver operating characteristic curve analysis showed significant diagnostic performance with histological complete response was CD86 (area under the curve = 0.71, 95% confidence interval = 0.53 to 0.88, P = .020). Patients with complete response had significantly lower CD86 expression (CD86-low) than patients with no complete response (P = .035). The complete response rates for CD86-low and CD86-high patients were 50% and 19%, respectively, and CD86-low patients had a significantly higher complete response rate (P = .047). Compared with all patients, the CD86-low group had a 1.5-fold increase in the complete response rate. Gene expression of CD86 and CTLA4 showed the strongest positive correlation with clinical outcomes in the incomplete response group (P < .001). Conclusion Low expression of CD86 in exfoliated cervical cells can be used as a pretreatment biomarker to predict histological complete response after IGMKK16E7 administration.

Cytomegalovirus (CMV) seropositivity is associated with increased risk for atherosclerotic disease in patients with end-stage renal disease. This association is due to a unique peripheral blood CD4+ T cell population which lack CD28 (CD4+CD28− T cells). Here we found that this patient population has a significant age-dependent increase of CD4+CD28− T cells that comprise over half of the circulating CD4 T cells in some. Patients over 50 years of age have a 50-fold higher percentage of CD4+CD28− T cells compared to seronegative patients and a 5-fold higher percentage when compared to seropositive healthy controls. Stimulation by CMV-antigen or by polyclonal stimulation using PMA and ionomycin showed that CD4+CD28− cells in patients with end stage renal disease degranulated and secreted interferon γ thus indicating that they are cytolytic. The average anti-CMV IgG titer displayed a remarkable age-dependent increase only in patients with end stage renal disease. These findings are highly suggestive of repetitive antigenic stimulation of the immune system in patients with end stage disease by subclinical CMV reactivation which might contribute to increased atherosclerotic risk.

CD4+CD25+ regulatory T cells (Tregs) are crucial for the maintenance of immunological tolerance. Recent data indicate that Tregs not only develop in the thymus during ontogeny but can also differentiate from naive T cells in the periphery. The following protocol describes a method by which Tregs are generated in vitro by stimulation of naive T cells in the presence of transforming growth factor β (Ti-Tregs). In vitro -induced regulatory T cells express markers of conventional Treg such as CD25 and the genetic program committing transcription factor FoxP3. Functionally the in vitro -generated Ti-Tregs suppress T-cell activation and proliferation while in vivo these cells have been proven to control inflammation in different animal models, suggesting a potential use of these cells for immunotherapy. The protocol can be completed within 5 days.

Plasma cell myeloma (PCM) exhibits immunophenotypic aberrancies that can be used for minimal residual disease (MRD) detection after therapy. The authors sought to determine whether non-neoplastic plasma cells, especially in the bone marrow (BM) post various therapies, would exhibit immunophenotypic variations interfering PCM MRD detection. The authors studied the flow cytometric immunophenotypes of non-neoplastic plasma cells from 50 BM specimens, including 12 untreated BM and 38 BM specimens from patients with non-plasmacytic haematological malignancies undergoing various therapies, and compared with 59 BM specimens positive for PCM MRD. Non-neoplastic plasma cells showed heterogeneous expressions of CD45 (78% (41–100)) and CD19 (80% (52–97)), and were negative for CD20 and CD117. CD56 was observed in a small subset (6% (0–37)) and CD28 in a larger subset (15% (0–59)) of non-neoplastic plasma cells, with the latter more frequently expressed in post-treatment BMs (p=0.01). However, despite a partial immunophenotypic overlap, PCM cells could be reliably discriminated from non-neoplastic plasma cells by the presence of a higher number of aberrancies (3 (1–6) vs 0 (0–2)) and stronger intensity and uniformity of aberrant expression (p<0.001 in each marker using a cut-off value). In addition, simultaneous assessment of cytoplasmic κ/λ with surface markers detected light chain restriction in all 59 PCM cases. In conclusion, non-neoplastic plasma cells in BM are more immunophenotypically heterogeneous than previously understood; however, these immunophenotypic variations differ from those of PCM. With advances in multicolour flow cytometry and application of recently validated markers, PCM MRD may still be reliably distinguished from non-neoplastic plasma cells.

CD28−, CD57+ and KLRG1+ are cell surface markers that have been used to describe senescent T-lymphocytes in humans. However, the relationship among these phenotypes during aging, and their relationship with the concept of in vitro cellular aging have not been well established. Using five-colour flow cytometry, we analyzed peripheral blood T-lymphocytes for their expression of CD28, CD57 and KLRG1 in 11 young (Y) and 11 old (O) apparently healthy human subjects. The proportions of CD28− and CD57+ cells were significantly higher among the T-cell populations of O compared to Y subjects; the proportion of KLRG1+ cells was significantly higher only among CD8+ cells. Populations that were more frequent in the elderly participants were characterised as CD28+ CD57+, CD28− CD57+ or CD28− CD57−. The expression of p16 and p21, considered as markers for in vitro senescence, was higher in CD28+ CD57+ cells than in other subpopulations in both age groups. The expression of p21 was age-related, which was not the case for p16. Thus, although both p16 and p21 are involved in T-cell senescence, they appear to behave differently. CMV infection and shifts in subpopulations are unlikely as explanations of the observed differences. Their higher levels of p16 and p21 expression, coupled with their higher prevalence in the elderly participants make CD28+ CD57+ cells the subpopulation of T-cells most closely corresponding to the concept of senescent cells.