Dihydroartemisinin Increases the Sensitivity of Photodynamic Therapy Via NF-κB/HIF-1α/VEGF Pathway in Esophageal Cancer Cell in vitro and in vivo

Background/Aims: Although photodynamic therapy (PDT) can relieve esophageal obstruction and prolong survival time of patients with esophageal cancer, it can induce nuclear factor-kappa B (NF-κB) activation in many cancers, which plays a negative role in PDT. Dihydroartemisinin (DHA), the most potent artemisinin derivative, can enhance the effect of PDT on esophageal cancer cells. However, the mechanism is still unclear. Methods: We generated stable cell lines expressing the super-repressor form of the NF-κB inhibitor IκBα and cell lines with lentivirus vector-mediated silencing of the HIF-1α gene. Esophageal xenograft tumors were created by subcutaneous injection of Eca109 cells into BALB/c nude mice. Four treatment groups were analyzed: a control group, photosensitizer alone group, light alone group, and PDT group. NF-κB expression was detected by an electrophoretic mobility shift assay, hypoxia-inducible factor α (HIF-1α) and vascular endothelial growth factor (VEGF) by real-time PCR, NF-κB, HIF-1α, and VEGF protein by western blot, and Ki-67, HIF-1α, VEGF, and NF-κB protein by immunohistochemistry. Results: PDT increased NF-κB activity and the gene expression of HIF-1α and VEGF in vitro and in vivo. In contrast, the DHA groups, particularly the combined DHA and PDT treatment group, abolished the effect. The combined treatment significantly inhibited tumor growth in vitro and in vivo. NF-κB activity and HIF-1α expression were also reduced in the stable IκBα expression group, whereas the former showed no change in HIF-1α-silenced cells. Conclusion: DHA might increase the sensitivity of esophageal cancer cells to PDT by inhibiting the NF-κB/HIF-1α/VEGF pathway

Responses of Murine and Human Macrophages to Leptospiral Infection: A Study Using Comparative Array Analysis

<div><p>Leptospirosis is a re-emerging tropical infectious disease caused by pathogenic <i>Leptospira</i> spp. The different host innate immune responses are partially related to the different severities of leptospirosis. In this study, we employed transcriptomics and cytokine arrays to comparatively calculate the responses of murine peritoneal macrophages (MPMs) and human peripheral blood monocytes (HBMs) to leptospiral infection. We uncovered a series of different expression profiles of these two immune cells. The percentages of regulated genes in several biological processes of MPMs, such as antigen processing and presentation, membrane potential regulation, and the innate immune response, etc., were much greater than those of HBMs (>2-fold). In MPMs and HBMs, the caspase-8 and Fas-associated protein with death domain (FADD)-like apoptosis regulator genes were significantly up-regulated, which supported previous results that the caspase-8 and caspase-3 pathways play an important role in macrophage apoptosis during leptospiral infection. In addition, the key component of the complement pathway, C3, was only up-regulated in MPMs. Furthermore, several cytokines, e.g. interleukin 10 (IL-10) and tumor necrosis factor alpha (TNF-alpha), were differentially expressed at both mRNA and protein levels in MPMs and HBMs. Some of the differential expressions were proved to be pathogenic <i>Leptospira</i>-specific regulations at mRNA level or protein level. Though it is still unclear why some animals are resistant and others are susceptible to leptospiral infection, this comparative study based on transcriptomics and cytokine arrays partially uncovered the differences of murine resistance and human susceptibility to leptospirosis. Taken together, these findings will facilitate further molecular studies on the innate immune response to leptospiral infection.</p></div

Comparisons of major regulated chemokines and receptors in murine peritoneal macrophages (MPMs) and human peripheral blood monocytes (HBMs) infected by <i>L. interrogans</i>.

<p>MPM-1 h, -2 h, and -4 h show the gene expression fold changes of the leptospiral-infected MPMs at 1-, 2-, and 4-h, respectively; HBM-1 h, -2 h, and -4 h show the gene expression fold changes (Ln(microarray folds)) of the leptospiral-infected HBMs at 1-, 2-, and 4-h, respectively. Average fold changes are listed in the affiliated table below.</p

The gene regulations of ECM components, synthesis enzymes, and degrading enzymes in human peripheral blood monocytes (HBMs) after 4-h <i>L. interrogans</i> infection.

<p>The gene regulations of ECM components, synthesis enzymes, and degrading enzymes in human peripheral blood monocytes (HBMs) after 4-h <i>L. interrogans</i> infection.</p

Comparison of cytokine regulation folds of murine peritoneal macrophages (MPMs) and human peripheral blood monocytes (HBMs) at mRNA and protein levels.

<p>Hierarchical cluster analyses of the average microarray data and the average cytokine array data of 62 cytokines of murine peritoneal macrophages (MPMs) and 60 cytokines of human peripheral blood monocytes (HBMs) were performed using Cluster3.0 software and visualized by using TreeView software. MPMs and HBMs were infected by <i>L. interrogans</i> for 4-h or infected by <i>L. biflexa</i> for 4-h, respectively.</p

Immunization with a Prefusion SARS-CoV-2 Spike Protein Vaccine (RBMRNA-176) Protects against Viral Challenge in Mice and Nonhuman Primates

There is an urgent need for a broad-spectrum and protective vaccine due to the emergence and rapid spreading of more contagious SARS-CoV-2 strains. We report the development of RBMRNA-176, a pseudouridine (Ψ) nucleoside-modified mRNA-LNP vaccine encoding pre-fusion stabilized trimeric SARS-CoV-2 spike protein ectodomain, and evaluate its immunogenicity and protection against virus challenge in mice and nonhuman primates. A prime-boost immunization with RBMRNA-176 at intervals of 21 days resulted in high IgG titers (over 1:819,000 endpoint dilution) and a CD4+ Th1-biased immune response in mice. RBMRNA-176 vaccination induced pseudovirus-neutralizing antibodies with IC50 ranging from 1:1020 to 1:2894 against SARS-CoV-2 spike pseudotyped wild-type and variant viruses, including Alpha, Beta, Gamma, and Kappa. Moreover, significant control of viral replication and histopathology in lungs was observed in vaccinated mice. In nonhuman primates, a boost given by RBMRNA-176 on day 21 after the prime induced a persistent and sustained IgG response. RBMRNA-176 vaccination also protected macaques against upper and lower respiratory tract infection, as well as lung injury. Altogether, these findings support RBMRNA-176 as a vaccine candidate for prevention of COVID-19

The differences of up-regulated and down-regulated signaling pathways of murine peritoneal macrophages (MPMs) and human peripheral blood monocytes (HBMs) infected by <i>L. interrogans</i>.

<p>The persistent regulations during the 4-h leptospiral infection, not the instantaneous regulations at the 1-h and the 2-h time points, were included in the statistical analysis using CapitalBio MAS (Molecule Annotation System version 3.0) software. The values above the bars show the percentages of total regulations, including up-regulations and down-regulations, of each KEGG signaling pathway.</p

Comparisons of major regulated inflammatory cytokines and receptors in murine peritoneal macrophages (MPMs) and human peripheral blood monocytes (HBMs) infected by <i>L. interrogans</i>.

<p>MPM-1 h, -2 h, and -4 h show the gene expression fold changes of the leptospiral-infected MPMs at 1-, 2-, and 4-h, respectively; HBM-1 h, -2 h, and -4 h show the gene expression fold changes (Ln(microarray folds)) of the leptospiral-infected HBMs at 1-, 2-, and 4-h, respectively. Average fold changes are listed in the affiliated table below.</p

The gene regulations of ECM components, synthesis enzymes, and degrading enzymes in murine peritoneal macrophages (MPMs) after 4-h <i>L. interrogans</i> infection.

<p>The gene regulations of ECM components, synthesis enzymes, and degrading enzymes in murine peritoneal macrophages (MPMs) after 4-h <i>L. interrogans</i> infection.</p