BackgroundThe therapeutic potential of mesenchymal stem cells (MSCs) may be attributed partly to humoral factors such as growth factors, cytokines, and chemokines. Human term placental tissue-derived MSCs (PlaMSCs), or conditioned medium left over from cultures of these cells, have been reported to enhance angiogenesis. Recently, the exosome, which can transport a diverse suite of macromolecules, has gained attention as a novel intercellular communication tool. However, the potential role of the exosome in PlaMSC therapeutic action is not well understood. The purpose of this study was to evaluate PlaMSC-derived exosome angiogenesis promotion in vitro and in vivo.MethodsMSCs were isolated from human term placental tissue by enzymatic digestion. Conditioned medium was collected after 48-h incubation in serum-free medium (PlaMSC-CM). Angiogenic factors present in PlaMSC-CM were screened by a growth factor array. Exosomes were prepared by ultracentrifugation of PlaMSC-CM, and confirmed by transmission electron microscopy, dynamic light scattering, and western blot analyses. The proangiogenic activity of PlaMSC-derived exosomes (PlaMSC-exo) was assessed using an endothelial tube formation assay, a cell migration assay, and reverse transcription-PCR analysis. The in-vivo angiogenic activity of PlaMSC-exo was evaluated using a murine auricle ischemic injury model.ResultsPlaMSC-CM contained both angiogenic and angiostatic factors, which enhanced endothelial tube formation. PlaMSC-exo were incorporated into endothelial cells; these exosomes stimulated both endothelial tube formation and migration, and enhanced angiogenesis-related gene expression. Laser Doppler blood flow analysis showed that PlaMSC-exo infusion also enhanced angiogenesis in an in-vivo murine auricle ischemic injury model.ConclusionsPlaMSC-exo enhanced angiogenesis in vitro and in vivo, suggesting that exosomes play a role in the proangiogenic activity of PlaMSCs. PlaMSC-exo may be a novel therapeutic approach for treating ischemic diseases.Electronic supplementary materialThe online version of this article (doi:10.1186/s13287-017-0660-9) contains supplementary material, which is available to authorized users.

Background: Peripheral lung lesions are increasing in numbers. Endoscopic diagnosis is essential for the prevention of unnecessary operations. Conventional diagnostic procedures have limitations in availability and results. Objectives: Endobronchial ultrasonography (EBUS) was investigated as a means to guide transbronchial lung biopsy, to reduce the discomfort during the procedure and to improve diagnostic accuracy. Methods: In 50 cases, we performed transbronchial lung biopsy combined with EBUS and fluoroscopic guidance. The results were compared to 42 controls assessed by fluoroscopy only. Results: In 38 cases (76%), EBUS could describe the peripheral lesion (33 from inside, including 9 cases with difficulties in fluoroscopic observation, and 5 from an adjacent bronchus, indicating the correct location of the lesion). If successfully placed inside, a change in the patient’s position was not required, which helped to reduce patient discomfort. Lung cancer was diagnosed in 24 patients and benign disease in 25 patients; in 1 case diagnosis remained unknown. When the EBUS probe could be introduced inside the lesion, the sensitivity for cancer diagnosis and specificity for cancer exclusion were 100%, respectively (15/15, 18/18). Compared to the controls in whom the biopsy site was determined by fluoroscopy only, the sensitivity tended to be superior by EBUS, although it did not reach statistical significance (p = 0.06). However, specificity and accuracy were statistically significant (both p = 0.02). Conclusions: When the lesion can be correctly described by EBUS from inside the lesion, EBUS is useful to guide transbronchial lung biopsy, can contribute to a reduction in patient discomfort and improves the accuracy of diagnosis. Additional navigation tools to increase correct positioning of the EBUS probe are desirable.

Periodontal disease is characterized by the destruction of tooth supporting tissues. Regeneration of periodontal tissues using ex vivo expanded cells has been introduced and studied, although appropriate methodology has not yet been established. We developed a novel cell transplant method for periodontal regeneration using periodontal ligament stem cell (PDLSC)-transferred amniotic membrane (PDLSC-amnion). The aim of this study was to investigate the regenerative potential of PDLSC-amnion in a rat periodontal defect model. Cultured PDLSCs were transferred onto amniotic membranes using a glass substrate treated with polyethylene glycol and photolithography. The properties of PDLSCs were investigated by flow cytometry and in vitro differentiation. PDLSC-amnion was transplanted into surgically created periodontal defects in rat maxillary molars. Periodontal regeneration was evaluated by microcomputed tomography (micro-CT) and histological analysis. PDLSCs showed mesenchymal stem cell-like characteristics such as cell surface marker expression (CD90, CD44, CD73, CD105, CD146, and STRO-1) and trilineage differentiation ability (i.e., into osteoblasts, adipocytes, and chondrocytes). PDLSC-amnion exhibited a single layer of PDLSCs on the amniotic membrane and stability of the sheet even with movement and deformation caused by surgical instruments. We observed that the PDLSC-amnion enhanced periodontal tissue regeneration as determined by micro-CT and histology by 4 weeks after transplantation. These data suggest that PDLSC-amnion has therapeutic potential as a novel cell-based regenerative periodontal therapy.

To investigate the mechanisms by which phorbol esters potentiate transmitter release from mossy fibre terminals we used fura dextran to measure the intraterminal Ca2+ concentration in mouse hippocampal slices. A phorbol ester, phorbol 12,13‐diacetate (PDAc), potentiated the field excitatory postsynaptic potential (fEPSP) slope. PDAc also enhanced the stimulation‐dependent increase of [Ca2+]i in the mossy fibre terminal (Δ[Ca2+]pre). The magnitude of the PDAc‐induced fEPSP potentiation (463 ± 57 % at 10 μM) was larger than that expected from the enhancement of Δ[Ca2+]pre (153 ± 5 %). The Δ[Ca2+]pre was suppressed by ω‐agatoxin IVA (ω‐AgTxIVA, 200 nM), a P/Q‐type Ca2+ channel‐specific blocker, by 31 %. The effect of PDAc did not select between ω‐AgTxIVA‐sensitive and ‐resistant components. The PDAc‐induced potentiation of the fEPSP slope was partially antagonized by the protein kinase C (PKC) inhibitor bisindolylmaleimide I (BIS‐I, 10 μM), whereas the Δ[Ca2+]pre was completely blocked by BIS‐I. Although the BIS‐I‐sensitive fEPSP potentiation was accompanied by a reduction of the paired‐pulse ratio (PPR), the BIS‐I‐resistant component was not. Whole‐cell patch clamp recording from a CA3 pyramidal neuron in a BIS‐I‐treated slice demonstrated that PDAc (10 μM) increased the frequency of miniature excitatory postsynaptic currents (mEPSCs, 259 ± 33 % of control) without a noticeable change in their amplitude (102 ± 5 % of control). These results suggest that PKC potentiates transmitter release by at least two distinct mechanisms, one Δ[Ca2+]pre dependent and the other Δ[Ca2+]pre independent. In addition, some phorbol ester‐mediated potentiation of synaptic transmission appears to occur without activating PKC.

Mesenchymal stem cell (MSC)-conditioned medium (MSC-CM) has been reported to enhance wound healing. Exosomes contain nucleic acids, proteins, and lipids, and function as an intercellular communication vehicle for mediating some paracrine effects. However, the function of MSC-derived exosomes (MSC-exo) remains elusive. In this study, we isolated human placenta MSC (PlaMSC)-derived exosomes (PlaMSC-exo) and examined their function in vitro. PlaMSCs were isolated from human term placenta using enzymatic digestion. PlaMSC-exo were prepared from the conditioned medium of PlaMSC (PlaMSC-CM) by ultracentrifugation. The expression of stemness-related genes, such as OCT4 and NANOG, in normal adult human dermal fibroblasts (NHDF) after incubation with PlaMSC-exo was measured by real-time reverse transcriptase PCR analysis (real-time PCR). The effect of PlaMSC-exo on OCT4 transcription activity was assessed using Oct4-EGFP reporter mice-derived dermal fibroblasts. The stimulating effects of PlaMSC-exo on osteoblastic and adipocyte-differentiation of NHDF were evaluated by alkaline phosphatase (ALP), and Alizarin red S- and oil red O-staining, respectively. The expression of osteoblast- and adipocyte-related genes was also assessed by real-time PCR. The treatment of NHDF with PlaMSC-exo significantly upregulated OCT4 and NANOG mRNA expression. PlaMSC-exo also enhanced OCT4 transcription. The NHDF treated with PlaMSC-exo exhibited osteoblastic and adipocyte-differentiation in osteogenic and adipogenic induction media. PlaMSC-exo increase the expression of OCT4 and NANOG mRNA in fibroblasts. As a result, PlaMSC-exo influence the differentiation competence of fibroblasts to both osteoblastic and adipocyte-differentiation. It shows a new feature of MSCs and the possibility of clinical application of MSC-exo. J. Cell. Biochem. 117: 1658-1670, 2016. © 2015 Wiley Periodicals, Inc.