CD9 is an integral membrane protein associated with integrins and other membrane proteins. Mice lacking CD9 were produced by homologous recombination. Both male and female CD9-/- mice were born healthy and grew normally. However, the litter size from CD9-/- females was less than 2% of that of the wild type. In vitro fertilization experiments indicated that the cause of this infertility was due to the failure of sperm-egg fusion. When sperm were injected into oocytes with assisted microfertilization techniques, however, the fertilized eggs developed to term. These results indicate that CD9 has a crucial role in sperm-egg fusion.

Paracrine signaling by bone-marrow-derived mesenchymal stem cells (MSCs) plays a major role in tissue repair. Although the production of regulatory cytokines by MSC transplantation is a critical modulator of tissue regeneration, we focused on exosomes, which are extracellular vesicles that contain proteins and nucleic acids, as a novel additional modulator of cell-to-cell communication and tissue regeneration. To address this, we used radiologic imaging, histological examination, and immunohistochemical analysis to evaluate the role of exosomes isolated from MSC-conditioned medium (CM) in the healing process in a femur fracture model of CD9 2/2 mice, a strain that is known to produce reduced levels of exosomes. We found that the bone union rate in CD9 2/2 mice was significantly lower than wild-type mice because of the retardation of callus formation. The retardation of fracture healing in CD9 2/2 mice was rescued by the injection of exosomes, but this was not the case after the injection of exosomes-free conditioned medium (CM-Exo). The levels of the bone repairrelated cytokines, monocyte chemotactic protein-1 (MCP-1), MCP-3, and stromal cell-derived factor-1 in exosomes were low compared with levels in CM and CM-Exo, suggesting that bone repair may be in part mediated by other exosome components, such as microRNAs. These results suggest that exosomes in CM facilitate the acceleration of fracture healing, and we conclude that exosomes are a novel factor of MSC paracrine signaling with an important role in the tissue repair process. STEM CELLS TRANSLATIONAL MEDICINE 2016;5:1620-1630 SIGNIFICANCEThis work focuses on exosomes, which are extracellular vesicles, as a novel additional modulator of cell-to-cell communication. This study evaluated the role of exosomes isolated from mesenchymal stem cell (MSC)-conditioned medium (MSC-CM) in the fracture-healing process of CD9 2/2 mice, a strain that is known to produce reduced levels of exosomes. Retardation of fracture healing in CD9 2/2 mice was rescued by the injection of MSC exosomes, but this was not the case after the injection of exosome-free CM. This study finds that MSC exosomes are a novel factor of MSC paracrine signaling, with an important role in the tissue repair process.

Stem cell therapy can help repair damaged heart tissue. Yet many of the suitable cells currently identified for human use are difficult to obtain and involve invasive procedures. In our search for novel stem cells with a higher cardiomyogenic potential than those available from bone marrow, we discovered that potent cardiac precursor-like cells can be harvested from human menstrual blood. This represents a new, noninvasive, and potent source of cardiac stem cell therapeutic material. We demonstrate that menstrual blood-derived mesenchymal cells (

The heparin-binding epidermal growth factor (EGF)-like growth factor (HB-EGF) is a member of the EGF family of growth factors that binds to and activates the EGF receptor (EGFR) and the related receptor tyrosine kinase, ErbB4. HB-EGF-null mice (HB del/del ) were generated to examine the role of HB-EGF in vivo. More than half of the HB del/del mice died in the first postnatal week. The survivors developed severe heart failure with grossly enlarged ventricular chambers. Echocardiographic examination showed that the ventricular chambers were dilated and that cardiac function was diminished. Moreover, HB del/del mice developed grossly enlarged cardiac valves. The cardiac valve and the ventricular chamber phenotypes resembled those displayed by mice lacking EGFR, a receptor for HB-EGF, and by mice conditionally lacking ErbB2, respectively. HB-EGF-ErbB interactions in the heart were examined in vivo by administering HB-EGF to WT mice. HB-EGF induced tyrosine phosphorylation of ErbB2 and ErbB4, and to a lesser degree, of EGFR in cardiac myocytes. In addition, constitutive tyrosine phosphorylation of both ErbB2 and ErbB4 was significantly reduced in HB del/del hearts. It was concluded that HB-EGF activation of receptor tyrosine kinases is essential for normal heart function. The ErbB family of receptor tyrosine kinases have fundamental roles in development, proliferation, and differentiation (1). There are four members of the receptor tyrosine kinase ErbB family, EGFR͞ErbB1͞HER1, ErbB2͞HER2͞neu, ErbB3͞HER3, and ErbB4͞HER4. Epidermal growth factor (EGF) family ligands bind to and activate their receptors by inducing the formation of homodimers and heterodimers, resulting in autophosphorylation of specific tyrosine residues within the cytoplasmic domain. The phosphorylated tyrosine residues bind adapter proteins, which are instrumental in mediating downstream signaling pathways that determine the biological activity of the ErbB family of ligands.In vertebrates, the EGF family of ligands bind to ErbB receptors with some degree of preference. EGF, transforming growth factor-␣, and amphiregulin bind to EGF receptor (EGFR); heparin-binding EGF-like growth factor (HB-EGF), epiregulin, and betacellulin bind to both EGFR and ErbB4; NRG-1 (neuregulin͞heregulin͞NDF) and NRG-2 bind to ErbB3 and ErbB4; and NRG-3 and NRG-4 bind to ErbB4 but not to ErbB3. Although no ligand for ErbB2 has yet been described, ErbB2 is active as a signaling receptor by forming heterodimers with other ErbB receptors (2).HB-EGF is synthesized as a type I transmembrane protein (proHB-EGF) composed of signal peptide, heparin-binding, EGF-like, juxtamembrane, transmembrane, and cytoplasmic domains (3, 4). The membrane-bound proHB-EGF is cleaved at the juxtamembrane domain, resulting in the shedding of soluble HB-EGF (5). The full-length proHB-EGF is biologically active as a juxtacrine growth factor that signals neighboring cells in a nondiffusible manner (6-8). ProHB-EGF forms complexes with CD9 (9) and integrin ␣3␤1 (10) on the cell membrane. ProHB-EGF is al...

Membrane fusion is an essential step in the encounter of two nuclei from sex cells-sperm and egg-in fertilization. However, aside from the involvement of two molecules, CD9 and Izumo, the mechanism of fusion remains unclear. Here, we show that spermegg fusion is mediated by vesicles containing CD9 that are released from the egg and interact with sperm. We demonstrate that the CD9 ؊/؊ eggs, which have a defective sperm-fusing ability, have impaired release of CD9-containing vesicles. We investigate the fusion-facilitating activity of CD9-containing vesicles by examining the fusion of sperm to CD9 ؊/؊ eggs with the aid of exogenous CD9-containing vesicles. Moreover, we show, by examining the fusion of sperm to CD9 ؊/؊ eggs, that hamster eggs have a similar fusing ability as mouse eggs. The CD9-containing vesicle release from unfertilized eggs provides insight into the mechanism required for fusion with sperm.