HMGB1 released by mesothelial cells drives the development of asbestos-induced mesothelioma.

Asbestos is the main cause of malignant mesothelioma. Previous studies have linked asbestos-induced mesothelioma to the release of HMGB1 from the nucleus to the cytoplasm, and from the cytoplasm to the extracellular space. In the cytoplasm, HMGB1 induces autophagy impairing asbestos-induced cell death. Extracellularly, HMGB1 stimulates the secretion of TNFα. Jointly, these two cytokines kick-start a chronic inflammatory process that over time promotes mesothelioma development. Whether the main source of extracellular HMGB1 were the mesothelial cells, the inflammatory cells, or both was unsolved. This information is critical to identify the targets and design preventive/therapeutic strategies to interfere with asbestos-induced mesothelioma. To address this issue, we developed the conditional mesothelial HMGB1-knockout (Hmgb1ΔpMeso) and the conditional myelomonocytic-lineage HMGB1-knockout (Hmgb1ΔMylc) mouse models. We establish here that HMGB1 is mainly produced and released by the mesothelial cells during the early phases of inflammation following asbestos exposure. The release of HMGB1 from mesothelial cells leads to atypical mesothelial hyperplasia, and in some animals, this evolves over the years into mesothelioma. We found that Hmgb1ΔpMeso, whose mesothelial cells cannot produce HMGB1, show a greatly reduced inflammatory response to asbestos, and their mesothelial cells express and secrete significantly reduced levels of TNFα. Moreover, the tissue microenvironment in areas of asbestos deposits displays an increased fraction of M1-polarized macrophages compared to M2 macrophages. Supporting the biological significance of these findings, Hmgb1ΔpMeso mice showed a delayed and reduced incidence of mesothelioma and an increased mesothelioma-specific survival. Altogether, our study provides a biological explanation for HMGB1 as a driver of asbestos-induced mesothelioma.

Mineral Trioxide Aggregate (MTA) Upregulates the Expression of DMP1 in Direct Pulp Capping in the Rat Molar.

Mineral trioxide aggregate (MTA) is an alternative endodontic material that predicts conductive or inductive calcified tissue formation from immature pulp mesenchymal stem cells (IPMSCs). The purpose of this study was to investigate whether MTA could promote reparative odontoblast differentiation via IPMSCs in the early phase of regeneration and compare with calcium hydroxide (CH). Direct pulp capping using calcium hydroxide (CH), MTA, and MTA with platelet-rich plasma (MTA + PRP) was performed on maxillary first molars of 8-week-old male Wistar rats (n = 36). After 3, 7, or 14 days, the teeth were analyzed for mineral density (MD) and volume of MD (VMD) via micro-focusing computed tomography (µCT), nestin, dentin matrix acidic phosphoprotein 1 (DMP1) immunohistochemistry, and real-time PCR for DMP1 mRNA expression. MTA stimulated the early phase differentiation of the IPMSCs into odontoblasts, with positive results for nestin and DMP1 compared with CH. Moreover, MTA + PRP stimulated calcified granule and dentin bridge formation through calcium mineral deposition, following the induction of DMP1 mRNA expression in IPMSCs. Our results suggested that the combination of MTA and PRP is an effective and clinically applicable method for activating endogenous dental pulp stem cells into odontoblasts in the early stages of pulp regeneration.

Pthlh, a promising cancer modifier gene in rat tongue carcinogenesis.

Susceptibly to the induction of rat tongue cancer (TC) by oral 4-nitroquinoline 1-oxide (4NQO) exposure is a polygenic trait. Among several quantitative trait loci identified by crosses between TC-susceptible Dark Agouti (DA) rats and TC-resistant Wistar-Furth (WF) rats, we focused on tongue cancer susceptibility locus (Tcas3) of chromosome 4. We examined tongue carcinogenesis in the reciprocal congenic strains DA.WF-Tcas3 and WF.DA-Tcas3 and in their parental strains. The Tcas3DA allele, and not the Tcas3WF allele, significantly favored tumor latency, incidence and TC number/size. In genomic DNA of TCs induced in (DA x WF) F1 rats, the resistant Tcas3WF allele was frequently and selectively lost, particularly in larger tumors. Thus, we searched the possible candidate genes in the Tcas3 region using microarray analysis of TCs in F1 rats and revealed significant upregulation of 2 cancer-related genes, parathyroid hormone-like hormone (Pthlh) and Kras2. The relevance of the WF allele of Pthlh as a cancer modifier was indicated by 3 single nucleotide polymorphisms specific to this strain. In contrast, no consistent strain-specific variations were found in Kras2. Moreover, the plasma Ca2+ level was consistently higher in DA rats when compared to the level in WF rats bearing TCs; moreover, the Pthlh-mRNA expression level was >30-fold higher in TCs when compared to this level in the normal tongue mucosa. Immunostaining experiments showed strong PTHrP protein expression in TCs of DA rats, and the signal was intensified in larger TCs. Kras2 was also upregulated in TCs, but to a lesser degree than PTHrP. Thus, Pthlh is a promising candidate modifier gene in the development and progression of rat TCs.

Induction of epithelial migration of lymphocytes by intercellular adhesion molecule-1 in a rat model of oral mucosal graft-versus-host disease.

To elucidate the involvement of intercellular adhesion molecule-1 (ICAM-1) in the migration of lymphocytes to the oral mucosal epithelium in a rat model of acute graft-versus-host disease (AGVHD), we investigated (1) ICAM-1 and major histocompatibility complex (MHC) class II expression by keratinocytes (KCs) and their role in the epithelial infiltration of CD8+ cells, (2) the tissue expression of interferon-γ (IFN-γ) mRNA and expression of IFN-γ receptor by KCs, and (3) the ability of KCs to direct CD8+ cells into the epithelial layers. We classified the oral mucosal lesions into three consecutive temporal phases on the basis of increased epithelial ICAM-1 expression: basal- (phase I), parabasal- (phase II), and pan-epithelial except for the cornified cell layer (phase III). Basal ICAM-1 expression by KCs preceded that of MHC class II molecules, infiltration of CD8+ cells and epithelial histological changes. Tissue expression of IFN-γ mRNA and expression of IFN-γ receptor on KCs evidenced by immunohistochemistry were detected in early lesions (phase I), indicating that locally produced IFN-γ induced ICAM-1 expression by KCs. CD8+ cells were bound to KCs in frozen sections of epithelial lesions, whereas no lymphocyte attachment was observed in normal KC. Adherence could be inhibited by pretreating CD8+ cells with lymphocyte function-associated antigen-1 (LFA-1) antibody and/or by pretreating sections with ICAM-1 antibody. Our data suggest that in the early phase of acute oral mucosal GVHD, the induction of ICAM-1 expression on KCs leads to the migration of CD8+ cells into the epithelium and that this is mediated in part by the ICAM-1/LFA-1 pathway.

Thymic alterations in GM2 gangliosidoses model mice.

BACKGROUND: Sandhoff disease is a lysosomal storage disorder characterized by the absence of ß-hexosaminidase and storage of GM2 ganglioside and related glycolipids. We have previously found that the progressive neurologic disease induced in Hexb(-/-) mice, an animal model for Sandhoff disease, is associated with the production of pathogenic anti-glycolipid autoantibodies. METHODOLOGY/PRINCIPAL FINDINGS: In our current study, we report on the alterations in the thymus during the development of mild to severe progressive neurologic disease. The thymus from Hexb(-/-) mice of greater than 15 weeks of age showed a marked decrease in the percentage of immature CD4(+)/CD8(+) T cells and a significantly increased number of CD4(+)/CD8(-) T cells. During involution, the levels of both apoptotic thymic cells and IgG deposits to T cells were found to have increased, whilst swollen macrophages were prominently observed, particularly in the cortex. We employed cDNA microarray analysis to monitor gene expression during the involution process and found that genes associated with the immune responses were upregulated, particularly those expressed in macrophages. CXCL13 was one of these upregulated genes and is expressed specifically in the thymus. B1 cells were also found to have increased in the thy mus. It is significant that these alterations in the thymus were reduced in FcRγ additionally disrupted Hexb(-/-) mice. CONCLUSIONS/SIGNIFICANCE: These results suggest that the FcRγ chain may render the usually poorly immunogenic thymus into an organ prone to autoimmune responses, including the chemotaxis of B1 cells toward CXCL13.