No abstract

Endoscopic submucosal dissection (ESD) permits en bloc removal of superficial oesophageal squamous cell carcinoma (ESCC). However, post-procedure stricture is common after ESD for widespread tumours, and multiple endoscopic balloon dilation (EBD) procedures are required. We aimed to evaluate the safety and effectiveness of endoscopic transplantation of tissue-engineered autologous oral mucosal epithelial cell sheets that had been transported by air over a distance of 1200 km in controlling postprocedural oesophageal stricture. Ten patients who underwent complete circular or semicircular ESD for ESCC were transplanted with cell sheets. The safety of the entire process including cell sheet preparation, transport, ESD and cell sheet transplantation was assessed. The incidence of oesophageal stricture, number of EBD sessions, and time until epithelialization were investigated. Each ESD was successfully performed, with subsequent cell sheet engrafting carried out safely. Following cell sheet transplantation, the luminal stenosis rate was 40%, while the median number of EBD sessions was 0. The median post-ESD ulcer healing period was rather short at 36 days. There were no significant complications at any stage of the process. Cell sheet transplantation and preparation at distant sites and transportation by air could be a safe and promising regenerative medicine technology.

No abstract

The oral mucosa exhibits unique regenerative properties, sometimes referred to as foetal-like wound healing. Researchers from our institute have used sheets of oral mucosa epithelial cells (OMECs) for regenerative medicine applications including cornea replacement and oesophageal epithelial regeneration for stricture prevention. Here, we have isolated exosomes from clinical-grade production of OMEC sheets from healthy human donors (n = 8), aiming to evaluate the clinical potential of the exosomes to stimulate epithelial regeneration and to improve understanding of the mode-of-action of the cells. Exosomes were isolated from conditioned (cExo) and non-conditioned (ncExo) media. Characterization was performed using Western blot for common exosomal-markers: CD9 and flotillin were positive while annexin V, EpCam and contaminating marker GRP94 were negative. Nanoparticle tracking analysis revealed a diameter of ~120 nm and transmission electron microscopy showed a corresponding size and spherical appearance. Human skin fibroblasts exposed to exosomes showed dose-dependent reduction of proliferation and a considerable increase of growth factor gene expression (HGF, VEGFA, FGF2 and CTGF). The results were similar for both groups, but with a trend towards a larger effect from cExo. To study adhesion, fluorescently labelled exosomes were topically applied to pig oesophageal wound-beds ex vivo and subsequently washed. Positive signal could be detected after as little as 1 min of adhesion, but increased adhesion time produced a stronger signal. Next, labelled exosomes were added to full-thickness skin wounds in rats and signal was detected up to 5 days after application. cExo significantly reduced the wound size at days 6 and 17. In conclusion, exosomes from OMEC sheets showed pro-regenerative effects on skin wound healing. This is the first time that the healing capacity of the oral mucosa is studied from an exosome perspective. These findings might lead to a combinational therapy of cell sheets and exosomes for future patients with early oesophageal cancer.

No abstract

No abstract

Emerging evidence has revealed the pivotal roles of C-type lectin-like receptors (CTLRs) in the regulation of a wide range of immune responses. Human natural killer cell receptor-P1A (NKRP1A) is one of the CTLRs and recognizes another CTLR, lectin-like transcript 1 (LLT1) on target cells to control NK, NKT and Th17 cells. The structural basis for the NKRP1A-LLT1 interaction was limitedly understood. Here, we report the crystal structure of the ectodomain of LLT1. The plausible receptor-binding face of the C-type lectin-like domain is flat, and forms an extended β-sheet. The residues of this face are relatively conserved with another CTLR, keratinocyte-associated C-type lectin, which binds to the CTLR member, NKp65. A LLT1-NKRP1A complex model, prepared using the crystal structures of LLT1 and the keratinocyte-associated C-type lectin-NKp65 complex, reasonably satisfies the charge consistency and the conformational complementarity to explain a previous mutagenesis study. Furthermore, crystal packing and analytical ultracentrifugation revealed dimer formation, which supports a complex model. Our results provide structural insights for understanding the binding modes and signal transduction mechanisms, which are likely to be conserved in the CTLR family, and for further rational drug design towards regulating the LLT1 function.Keywords: Cell surface receptor r Crystal structure r C-type lectin-like receptor r Natural killer cell r Protein-protein interactions Additional supporting information may be found in the online version of this article at the publisher's web-site Correspondence: Prof. Katsumi Maenaka e-mail: maenaka@pharm.hokudai.ac.jp * Equal contribution.C 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.eji-journal.eu 1606 S. Kita et al. Eur. J. Immunol. 2015. 45: 1605-1613 Introduction Natural killer (NK) cells are a component of innate immunity, and display cytotoxic activities against tumor cells and virally infected cells [1,2]. The activities of NK cells are precisely regulated by activating and inhibitory signals, through diverse receptors on the cell surface [3][4][5]. These receptors are encoded in two genomic regions, the leukocyte receptor complex (chromosome 19 in human) [6] and the NK gene complex (chromosome 12 in human) [7,8]. The leukocyte receptor complex encodes the NK receptors of the immunoglobulin (Ig) superfamily, such as killer Ig-like receptors and leukocyte Ig-like receptors, whereas the NK gene complex region encodes the NK receptors of the C-type lectinlike receptors (CTLRs). The CTLRs are subdivided into two groups, according to their expression patterns. Killer cell lectin-like receptors (KLRs) are expressed in NK cells, and C-type lectin receptors (CLECs) are expressed in non-NK cells [9]. The ligands of KLRs are (1) major histocompatibility complex (MHC) class I molecules and relatives, and (2) CLECs. The former members include CD94, the NKG2 subfamily [10], and the Ly49 subfamily [11], and the latter members are the NK receptor-P1 (NKRP1) subfamily [12,13]. The NKRP1 su...

No abstract