The immunogenicity of HLA-A2-restricted T-cell epitopes in the S protein of the Severe acute respiratory syndrome coronavirus (SARS-CoV) and of human coronavirus strain 229e (HCoV-229e) was analyzed for the elicitation of a T-cell immune response in donors who had fully recovered from SARS-CoV infection. We employed online database analysis to compare the differences in the amino acid sequences of the homologous T epitopes of HCoV-229e and SARS-CoV. The identified T-cell epitope peptides were synthesized, and their binding affinities for HLA-A2 were validated and compared in the T2 cell system. The immunogenicity of all these peptides was assessed by using T cells obtained from donors who had fully recovered from SARS-CoV infection and from healthy donors with no history of SARS-CoV infection. HLA-A2 typing by indirect immunofluorescent antibody staining showed that 51.6% of SARS-CoV-infected patients were HLA-A2 positive. Online database analysis and the T2 cell binding test disclosed that the number of HLA-A2-restricted immunogenic epitopes of the S protein of SARS-CoV was decreased or even lost in comparison with the homologous sequences of the S protein of HCoV-229e. Among the peptides used in the study, the affinity of peptides from HCoV-229e (H77 and H881) and peptides from SARS-CoV (S978 and S1203) for binding to HLA-A2 was higher than that of other sequences. The gamma interferon (IFN-␥) release Elispot assay revealed that only SARS-CoV-specific peptides S1203 and S978 induced a high frequency of IFN-␥-secreting T-cell response in HLA-A2؉ donors who had fully recovered from SARS-CoV infection; such a T-cell epitope-specific response was not observed in HLA-A2؉ healthy donors or in HLA-A2 ؊ donors who had been infected with SARS-CoV after full recovery. Thus, T-cell epitopes S1203 and S978 are immunogenic and elicit an overt specific T-cell response in HLA-A2 ؉ SARS-CoV-infected patients.
Increasing evidence indicates the immunosuppressive nature of the local environment in tumor. The present study was focused on analyzing the immune status within hepatocellular carcinoma. In contrast to the increasing number of CD4(+) T cells, CD8(+), CD3(-)CD56(+), CD3(+)CD56(+), and gammadeltaT cells were all found to be under-represented in tumor infiltrating lymphocytes. Notably, the relative abundance of CD3(+)CD56(+) cells appeared to be correlated with patient survival. Functional analysis demonstrated that CD4(+) cells in the tumor tended to produce more IL-10 but less IFN-gamma, whereas CD8(+) cells showed impaired capacity for the production of both IFN-gamma and perforin. Consistent with previous reports, we observed a significant increase of Foxp3(+) cells in the tumor tissue. Intriguingly, although over 90% of CD4(+)CD25(high) cells were found to be Foxp3(+), the majority of Foxp3(+) cells were identified in the CD4(+)CD25(medium) and CD4(+)CD25(-) subsets. In support of its role as a negative regulator, CD4(+)CD25(high) cells suppressed the proliferation of CD4(+)CD25(-) cells isolated from the same tissues in an APC dependent manner. In conclusion, the tumor microenvironment of hepatocellular carcinoma is featured by the presence of multiple immunosuppressive factors.
The MAGE-A3 protein, one of the promising tumor antigens for immunotherapy, is highly expressed in human hepatocellular carcinoma (HCC). In this study, we estimated the specific CD8(+) T cell immune response to MAGE-A3 p271-279 peptide (M3(271)) in the peripheral blood of HCC patients without antigen vaccination in order to evaluate its immunotherapeutic potential in these patients. After expansion in vitro, the functional IFN-gamma producing M3(271) specific CD8(+) T cells were detected in 30.8% (8/26) of HLA-A2(+)MAGE-A3(+) HCC patients. The effector CD8(+ )T cells could release cytotoxic molecules of granzyme B and perforin after restimulation with natural HLA-A2(+)MAGE-A3(+) HCC cell lines in the samples tested. The functional supertype of HLA-A2 in the presentation of HLA-A*0201 restricted M3(271) peptide has been identified in the Chinese HCC patients of Han ethnicity, that widely expanded the applicability of this tumor peptide vaccine in Chinese HCC patients. Thus, the functionally detectable pre-existence of M3(271)-specific CD8(+) T cells in HCC patients makes M3(271) a potential target for immunotherapy in these patients. The responsive CD8(+ )T cells to both NY-ESO-1 and MAGE-A3 antigens provide a rationale for the application of a bivalent vaccine in HCC patients with tumors expressing both antigens.
Induction of cardiomyocyte proliferation, the most promising approach to reverse myocardial attrition, has been gaining importance as a therapy for cardiovascular disease. Hypoxia and macrophages were previously independently reported to promote cardiomyocyte proliferation in mice. However, whether hypoxia promotes cardiomyocyte proliferation in humans, and the association between hypoxia and macrophages in cardiomyocyte proliferation, have not to the best of our knowledge been previously investigated. The present study investigated the cardiomyocyte proliferation in 22 acyanotic and 29 cyanotic patients. Cardiomyocyte proliferation in a hypoxic mouse model (15% O2) was subsequently performed and the macrophage subsets were analyzed. A C-C chemokine receptor type 2 (CCR2) inhibitor was used to increase the number of resident macrophages in order to investigate the effect of macrophages on cardiomyocyte proliferation. The results demonstrated that cardiomyocyte proliferation in the cyanotic infant group was significantly increased compared with the acyanotic infant group and the hypoxia-treated C57BL/6J neonates confirmed the hypoxia-induced cardiomyocyte proliferation. However, hypoxia did not induce the proliferation of isolated cardiomyocytes. Notably, hypoxia treatment increased the number of cardiac resident macrophages in neonate hearts. Furthermore, increasing the number of resident macrophages significantly enhanced cardiomyocyte proliferation. In conclusion, postnatal hypoxia promoted cardiomyocyte proliferation in humans and animals, and cardiac resident macrophages may be involved in this process. Therefore, this novel mechanism may provide a promising strategy for cardiovascular disease treatment.
There is a large subpopulation of multinucleated polyploid cardiomyocytes (M*Pc CMs) in the adult mammalian heart. However, the pathophysiological significance of increased M*Pc CMs in heart disease is poorly understood. We sought to determine the pathophysiological significance of increased M*Pc CMs during hypoxia adaptation. Methods and results: A model of hypoxia-induced cardiomyocyte (CM) multinucleation and polyploidization was established and found to be associated with less apoptosis and less reactive oxygen species (ROS) production. Compared to mononucleated diploid CMs (1*2c CMs), tetraploid CMs (4c CMs) exhibited better mitochondria quality control via increased mitochondrial autophagy (mitophagy). RNA-seq revealed Prkaa2, the gene for AMPKα2, was the most obviously up-regulated autophagy-related gene. Knockdown of AMPKα2 increased apoptosis and ROS production and suppressed mitophagy in 4c CMs compared to 1*2c CMs. Rapamycin, an autophagy activator, alleviated the adverse effect of AMPKα2 knockdown. Furthermore, silencing PINK1 also increased apoptosis and ROS in 4c CMs and weakened the adaptive superiority of 4c CMs. Finally, AMPKα2 −/− mutant mice exhibited exacerbation of apoptosis and ROS production via decreases in AMPKα2-mediated mitophagy in 4c CMs compared to 1*2c CMs during hypoxia. Conclusions: Compared to 1*2c CMs, hypoxia-induced 4c CMs exhibited enhanced mitochondria quality control and less apoptosis via AMPKα2-mediated mitophagy. These results suggest that multinucleation and polyploidization allow CM to better adapt to stress via enhanced mitophagy. In addition, activation of AMPKα2 may be a promising target for myocardial hypoxia-related diseases.
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