During the last decade, a novel mechanism of protein release has been recognized that involves small (30-100 nm) membrane vesicles termed exosomes. [1][2][3] Exosomal vesicles are secreted following the fusion of multivesicular late endosomes with the plasma membrane. While the range of exosomal proteins depends on cell type, these vesicles commonly carry cell-surface proteins and cytoskeletal proteins. Several physiologic roles have been assigned to exosomes, including the expulsion of obsolete membrane constituents, exchange of cellular material and intercellular communication. Exosome production has been observed in a variety of cell types in vitro, including reticulocytes, 4) cytotoxic T lymphocytes, 5)B lymphocytes, 6) dendritic cells 7) and neoplastic intestinal epithelial cells.8) Recent studies have reported that such vesicles are present in some physiological fluids, such as bronchoalveolar lavage 9) or urine. 10) However, there is still little evidence of whether exosomes are produced in vivo.Saliva is considered to provide the first line of oral cavity defense against bacterial and viral attack. Human whole saliva contains a potent mixture of diverse components such as mucin, immunoglobulin A (IgA), proline-rich proteins and defensins, which are produced in three major paired salivary glands (parotid, submadibular and sublingual) and several minor glands. Although comprehensive proteome analyses of whole saliva have been reported recently, 12,13) the protein constituents of saliva are not fully understood.The membrane-associated serine protease dipeptidyl peptidase IV (DPP IV), which is identical to the lymphocyte surface glycoprotein CD26, cleaves dipeptides from the N-terminus of peptides with a proline or alanine residue in the penultimate position.14) CD26/DPP IV is highly expressed on fibroblasts, epithelial and endothelial cells, and specific leukocyte subsets. The extracellular protease domain of CD26/DPP IV, which is produced by proteolytic cleavage of the membrane-bound form of CD26/DPP IV, also exists in a soluble form in plasma.15) Recently, it was reported that DPP IV is released from intestinal epithelial cells into the extracellular milieu as a constituent of exosome-like vesicles. 8)More recently, our previous report revealed that DPP IV is released into snake venom in an unprocessed form, 16) suggesting that the DPP IV is associated with membrane. In addition, we have found exosome-like vesicles that carry DPP IV in snake venom.17) DPP IV activity has been found in human saliva, 18) but the mechanism of its release has not been elucidated. Snake venom is highly modified saliva that is produced by special glands of certain species of snakes. Therefore, we hypothesized that exosome-like vesicles carrying DPP IV could be present in human saliva. We demonstrate for the first time a population of vesicles in human whole saliva similar in size to the previously described exosomes. We found that these vesicles contain DPP IV, galectin-3 and IgA. MATERIALS AND METHODS MaterialsGly-Pro-4-methyl...

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Interleukin-6 (IL-6) is known to be involved in the pathogenesis of various inflammatory diseases, but its role in bleomycin (BLM)-induced lung injury and subsequent fibrotic changes remains to be determined. We evaluated the role of IL-6 in the lung inflammatory changes induced by BLM using wild-type (WT) and IL-6-deficient (IL-6(-/-)) mice. The mice were treated intratracheally with 1 mg/kg BLM and killed 2, 7, or 21 days later. Lung Inflammation in the acute phase (Days 2 and 7) was assessed by differential cell counts in bronchoalveolar lavage (BAL) fluid and cytokine levels in the lung. Lung fibrotic changes were evaluated on Day 21 by histopathology and collagen assay. On Day 2, BLM administration induced significant increases in the numbers of total cells, macrophages, and neutrophils in BAL fluid, which were attenuated in IL-6(-/-) mice (P < 0.05). Lung pathology also showed inflammatory cell accumulation, which was attenuated in the IL-6(-/-) mice compared with WT mice. In WT mice, elevated levels of TGF-beta(1) and CCL3 were observed 2 and 7 days after BLM challenge, respectively. On Day 7, BLM-induced inflammatory cell accumulation did not differ between the genotypes. Lung pathology 21 days after BLM challenge revealed significant fibrotic changes with increased collagen content, which was attenuated in IL-6(-/-) mice. Although the TGF-beta(1) level in the lung did not differ between the genotypes on Day 21, CCL3 was significantly lower in IL-6(-/-) mice. These results indicate that IL-6 may play an important role in the pathogenesis of BLM-induced lung injury and subsequent fibrotic changes.

Small non-coding RNAs, such as microRNAs (miRNAs), are involved in diverse processes, including organ development and tissue differentiation. Exosomes are small membrane vesicles (30-100 nm in diameter) produced by numerous cells. Recently, exosomes have been shown to contain miRNAs. However, the small RNAs contained in exosomes are not fully characterized. In a previous study, we found at least two types of salivary exosome that are different in size and have different proteomes. Studies of salivary exosomal small RNAs are limited to miRNAs. In this study, we examined small RNA transcriptomes using next generation sequencing technology to elucidate a full transcriptome set of small RNAs expressed in the two types of salivary exosomes and in whole saliva (WS). Many types of small RNA, such as miRNA, piwi-interacting RNA (piRNA), small nucleolar RNA (snoRNA) and other small RNAs are contained in salivary exosomes and WS. Among these small RNAs we identified novel miRNA candidates.

RAGE plays an important role in the pathogenesis of LPS-induced lung injury in mice. It was suggested that sRAGE should be tested as a treatment modality in other models of acute lung injury.