High glutamic acid decarboxylase antibody titers may be associated with a decline in ß-cell function over time and future insulin deficiency in latent autoimmune diabetes in adults.

AIMS: Latent autoimmune diabetes in adults (LADA) is characterized by positive islet-associated autoantibodies including glutamic acid decarboxylase antibody (GADA), and gradual decline in insulin secretion, progressing to insulin dependency. This cross-sectional study aimed to determine whether GADA by enzyme-linked immunosorbent assay (GADA-ELISA) titer of ≥180 U/mL could be associated with decline in ß-cell function in participants with LADA. METHODS: Sixty-three participants with LADA were recruited and an association between insulin secretion capacity and disease duration was investigated. Insulin peptide-specific inflammatory immunoreactivity was investigated to determine the disease's activity. RESULTS: There was a significant inverse correlation between disease duration and C-peptide index in participants with GADA-ELISA titer of ≥180 U/mL (Spearman's r (rs) = -0.516, p < 0.01). The positivity rate of insulin peptide-specific inflammatory immunoreactivity was significantly higher in those with ≥180 U/mL than in those with <180 U/mL (p < 0.05). In participants with human leukocyte antigen (HLA)-DRB1*04:05, a significant inverse correlation was observed between disease duration and C-peptide index in those with ≥180 U/mL (rs = -0.751, p < 0.01). CONCLUSIONS: GADA-ELISA titer of ≥180 U/mL, especially with HLA-DRB1*04:05, might reflect higher disease activity and may be associated with decline in ß-cell function over time and future insulin dependency in LADA.

Immune reconstitution and cidofovir administration rescue human adenovirus hepatitis after allogeneic hematopoietic cell transplantation.

Human adenovirus infection (HAdV) may be fatal in patients undergoing allogeneic hematopoietic cell transplantation (HCT). Cidofovir is effective in only a part of the post-HCT HAdV infection. Therefore, posttransplant immune reconstitution is important for HAdV clearance. We describe the detailed immune reconstitution and response of adenovirus-specific T cells in a patient with inborn errors of immunity who had disseminated HAdV infection with hepatitis post-HCT and was treated with cidofovir. Though the patient received cidofovir for only 19 days starting from Day 72 after HCT because of renal dysfunction, we observed T-cell reconstitution, a decrease in HAdV copy number, and amelioration of the symptoms of HAdV infection after Day 90. We initially observed expanded NK and CD8+CD45RO+ memory subsets and later gradual increase of naïve T cells eveloped after cessation of cidofovir treatment. An increase in adenovirus-specific IFN-γ secretion from 2 to 4 months after HCT was confirmed by ELISpot assay. The progression of immune reconstitution and cidofovir treatment are considered to have contributed to survival in this patient. Optimization of transplantation methods, prompt appropriate antiviral medication, and virus-specific T-cell therapy would be necessary as the better strategy for systemic HAdV infection.

In Silico Analyses, Experimental Verification and Application in DNA Vaccines of Ebolavirus GP-Derived pan-MHC-II-Restricted Epitopes.

(1) Background and Purpose: Ebola virus (EBOV) is the causative agent of Ebola virus disease (EVD), which causes extremely high mortality and widespread epidemics. The only glycoprotein (GP) on the surface of EBOV particles is the key to mediating viral invasion into host cells. DNA vaccines for EBOV are in development, but their effectiveness is unclear. The lack of immune characteristics resides in antigenic MHC class II reactivity. (2) Methods: We selected MHC-II molecules from four human leukocyte antigen II (HLA-II) superfamilies with 98% population coverage and eight mouse H2-I alleles. IEDB, NetMHCIIpan, SYFPEITHI, and Rankpep were used to screen MHC-II-restricted epitopes with high affinity for EBOV GP. Further immunogenicity and conservation analyses were performed using VaxiJen and BLASTp, respectively. EpiDock was used to simulate molecular docking. Cluster analysis and binding affinity analysis of EBOV GP epitopes and selected MHC-II molecules were performed using data from NetMHCIIpan. The selective GP epitopes were verified by the enzyme-linked immunospot (ELISpot) assay using splenocytes of BALB/c (H2d), C3H, and C57 mice after DNA vaccine pVAX-GPEBO immunization. Subsequently, BALB/c mice were immunized with Protein-GPEBO, plasmid pVAX-GPEBO, and pVAX-LAMP/GPEBO, which encoded EBOV GP. The dominant epitopes of BALB/c (H-2-I-AdEd genotype) mice were verified by the enzyme-linked immunospot (ELISpot) assay. It is also used to evaluate and explore the advantages of pVAX-LAMP/GPEBO and the reasons behind them. (3) Results: Thirty-one HLA-II-restricted and 68 H2-I-restricted selective epitopes were confirmed to have high affinity, immunogenicity, and conservation. Nineteen selective epitopes have cross-species reactivity with good performance in MHC-II molecular docking. The ELISpot results showed that pVAX-GPEBO could induce a cellular immune response to the synthesized selective peptides. The better immunoprotection of the DNA vaccines pVAX-LAMP/GPEBO coincides with the enhancement of the MHC class II response. (4) Conclusions: Promising MHC-II-restricted candidate epitopes of EBOV GP were identified in humans and mice, which is of great significance for the development and evaluation of Ebola vaccines.

PP19128R, a Multiepitope Vaccine Designed to Prevent Latent Tuberculosis Infection, Induced Immune Responses In Silico and In Vitro Assays.

Background: Latent tuberculosis infection (LTBI) is the primary source of active tuberculosis (ATB), but a preventive vaccine against LTBI is lacking. Methods: In this study, dominant helper T lymphocyte (HTL), cytotoxic T lymphocyte (CTL), and B-cell epitopes were identified from nine antigens related to LTBI and regions of difference (RDs). These epitopes were used to construct a novel multiepitope vaccine (MEV) based on their antigenicity, immunogenicity, sensitization, and toxicity. The immunological characteristics of the MEV were analyzed with immunoinformatics technology and verified by enzyme-linked immunospot assay and Th1/Th2/Th17 cytokine assay in vitro. Results: A novel MEV, designated PP19128R, containing 19 HTL epitopes, 12 CTL epitopes, 8 B-cell epitopes, toll-like receptor (TLR) agonists, and helper peptides, was successfully constructed. Bioinformatics analysis showed that the antigenicity, immunogenicity, and solubility of PP19128R were 0.8067, 9.29811, and 0.900675, respectively. The global population coverage of PP19128R in HLA class I and II alleles reached 82.24% and 93.71%, respectively. The binding energies of the PP19128R-TLR2 and PP19128R-TLR4 complexes were -1324.77 kcal/mol and -1278 kcal/mol, respectively. In vitro experiments showed that the PP19128R vaccine significantly increased the number of interferon gamma-positive (IFN-γ+) T lymphocytes and the levels of cytokines, such as IFN-γ, tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and IL-10. Furthermore, positive correlations were observed between PP19128R-specific cytokines in ATB patients and individuals with LTBI. Conclusions: The PP19128R vaccine is a promising MEV with excellent antigenicity and immunogenicity and no toxicity or sensitization that can induce robust immune responses in silico and in vitro. This study provides a vaccine candidate for the prevention of LTBI in the future.

Analysis of cytomegalovirus-specific T-cell responses in patients with hypertension: comparison of assay methods and antigens.

BACKGROUND: Recent studies suggest an association between cytomegalovirus (CMV) infection and hypertension. In the present study, we used a variety of antigens and different assay methods to investigate the relationship between CMV-specific T-cell responses and arterial stiffness in patients with hypertension. METHODS: To evaluate arterial stiffness, pulse wave velocity (PWV) was measured in 207 hypertensive patients (average age, 63 ± 8 years). To measure CMV pp65 and IE-1-specific T-cell responses, we performed intracellular cytokine staining (ICS) and enzyme-linked immunospot (ELISPOT) assays. We also analyzed CMV-specific T-cell responses against 10 different CMV antigens using ELISPOT assays. RESULTS: In patients with hypertension, senescent CD8+ T-cell frequencies were significantly correlated with arterial stiffness. Moreover, arterial stiffness was independently associated with CMV pp65-specific CD8+ T-cell responses as measured by ICS. CMV-specific CD8+ T-cell responses measured by ICS and ELISPOT assays showed good agreement and significant correlation with each other. ELISPOT analyses against 10 different CMV antigens revealed a consistent response pattern irrespective of age, gender, and diabetes. CONCLUSIONS: CMV pp65-specific CD8+ T-cell responses were independently correlated with arterial stiffness in patients with hypertension. Additionally, the results of ICS and ELISPOT assays showed a significant correlation and good agreement with each other. These findings are important for guiding choices regarding the broad clinical application of CMV-specific T-cell response assays in this patient population.