Tohru Tanigawa

Vildagliptin Stimulates Endothelial Cell Network Formation and Ischemia-induced Revascularization via an Endothelial Nitric-oxide Synthase-dependent Mechanism

Background: DPP-4 inhibitors exert pleiotropic effects that modulate cardiovascular disease. Results: The DPP-4 inhibitor vildagliptin stimulates ischemia-induced revascularization through eNOS signaling. The angiogenic actions of vildagliptin are mediated by both GLP-1-dependent and -independent mechanisms. Conclusion: DPP-4 inhibitor promotes endothelial cell function via eNOS signaling. Significance: DPP-4 inhibitor could be beneficial in patients with diabetes-related vascular complications. Dipeptidyl peptidase-4 inhibitors are known to lower glucose levels and are also beneficial in the management of cardiovascular disease. Here, we investigated whether a dipeptidyl peptidase-4 inhibitor, vildagliptin, modulates endothelial cell network formation and revascularization processes in vitro and in vivo. Treatment with vildagliptin enhanced blood flow recovery and capillary density in the ischemic limbs of wild-type mice, with accompanying increases in phosphorylation of Akt and endothelial nitric-oxide synthase (eNOS). In contrast to wild-type mice, treatment with vildagliptin did not improve blood flow in ischemic muscles of eNOS-deficient mice. Treatment with vildagliptin increased the levels of glucagon-like peptide-1 (GLP-1) and adiponectin, which have protective effects on the vasculature. Both vildagliptin and GLP-1 increased the differentiation of cultured human umbilical vein endothelial cells (HUVECs) into vascular-like structures, although vildagliptin was less effective than GLP-1. GLP-1 and vildagliptin also stimulated the phosphorylation of Akt and eNOS in HUVECs. Pretreatment with a PI3 kinase or NOS inhibitor blocked the stimulatory effects of both vildagliptin and GLP-1 on HUVEC differentiation. Furthermore, treatment with vildagliptin only partially increased the limb flow of ischemic muscle in adiponectin-deficient mice in vivo. GLP-1, but not vildagliptin, significantly increased adiponectin expression in differentiated 3T3-L1 adipocytes in vitro. These data indicate that vildagliptin promotes endothelial cell function via eNOS signaling, an effect that may be mediated by both GLP-1-dependent and GLP-1-independent mechanisms. The beneficial activity of GLP-1 for revascularization may also be partially mediated by its ability to increase adiponectin production.

iPS cell sheets created by a novel magnetite tissue engineering method for reparative angiogenesis

Angiogenic cell therapy represents a novel strategy for ischemic diseases, but some patients show poor responses. We investigated the therapeutic potential of an induced pluripotent stem (iPS) cell sheet created by a novel magnetite tissue engineering technology (Mag-TE) for reparative angiogenesis. Mouse iPS cell-derived Flk-1+ cells were incubated with magnetic nanoparticle-containing liposomes (MCLs). MCL-labeled Flk-1+ cells were mixed with diluted extracellular matrix (ECM) precursor and a magnet was placed on the reverse side. Magnetized Flk-1+ cells formed multi-layered cell sheets according to magnetic force. Implantation of the Flk-1+ cell sheet accelerated revascularization of ischemic hindlimbs relative to the contralateral limbs in nude mice as measured by laser Doppler blood flow and capillary density analyses. The Flk-1+ cell sheet also increased the expressions of VEGF and bFGF in ischemic tissue. iPS cell-derived Flk-1+ cell sheets created by this novel Mag-TE method represent a promising new modality for therapeutic angiogenesis.

Dental pulp-derived stem cell conditioned medium reduces cardiac injury following ischemia-reperfusion.

Stem cells from human exfoliated deciduous teeth (SHEDs) can regenerate various tissues. We investigated the impact of SHED-conditioned medium (SHED-CM) on myocardial injury in a mouse model of ischemia-reperfusion (I/R). Wild-type (WT) mice were subjected to myocardial ischemia followed by reperfusion. SHED-CM was intravenously injected at 5 min after reperfusion. Administration of SHED-CM reduced myocardial infarct size as well as decreased apoptosis and inflammatory cytokine levels, such as TNF-α, IL-6, and IL-β, in the myocardium following I/R. In cultured cardiac myocytes, SHED-CM significantly suppressed apoptosis under hypoxia/serum-deprivation and reduced LPS-induced expression of pro-inflammatory genes. Furthermore, anti-apoptotic action of SHED-CM was stronger than bone marrow-derived stem cell (BMSC)-CM or adipose-derived stem cell (ADSC)-CM in cardiac myocytes. SHED-CM contains a higher concentration of hepatocyte growth factor (HGF) than BMSC-CM and ADSC-CM, and neutralization of HGF attenuated the inhibitory actions of SHED-CM on apoptosis in cardiac myocytes. Finally, WT mice were intravenously treated with an HGF-depleted SHED-CM, followed by myocardial I/R. HGF depletion significantly attenuated the inhibitory actions of SHED-CM on myocardial infarct size and apoptosis after I/R. SHED-CM protects the heart from acute ischemic injury because it suppresses inflammation and apoptosis. SHED-CM could be a useful treatment option for acute myocardial infarction.

Multilayered adipose-derived regenerative cell sheets created by a novel magnetite tissue engineering method for myocardial infarction

BACKGROUND Adipose-derived regenerative cells (ADRCs) are a promising source of autologous stem cells for regeneration and repair of damaged tissue. Herein, we investigated the therapeutic potential of ADRC sheets created by a magnetite tissue engineering technology (Mag-TE) for myocardial infarction. METHODS AND RESULTS Adipose tissue was obtained from wild-type (WT) mice and ADRCs were isolated. ADRCs incubated with magnetic nanoparticle-containing liposomes (MCLs) were cultured. MCL-labeled ADRCs were mixed with a diluted extracellular matrix (ECM) precursor, and a magnet was placed on the reverse side. Magnetized ADRCs formed multilayered cell sheets after a 24-h incubation. WT mice were subjected to myocardial infarction by permanent ligation of the left anterior descending artery. We then transplanted the constructed ADRC sheet or a cell-free collagen gel sheet, as a control, onto the infarcted myocardium using an Alnico magnet before skin closure. Cardiac parameters were measured by echocardiogram, and angiogenesis was determined by tissue capillary density. ADRC sheet-treated mice showed significant improvements in systolic function, infarct wall thinning, and fibrotic length after myocardial infarction. ADRC sheet implantation also promoted angiogenesis in both the infarct area and the border zone in WT mice after myocardial infarction. The angiogenic effects of ADRC sheets were attributed to an increased expression of VEGF and bFGF mRNA in ischemic hearts. CONCLUSIONS ADRC sheets created by this Mag-TE method protect the heart against pathological cardiac remodeling. Our ADRC sheets have the potential to be a novel regenerative strategy for ischemic heart disease.

Therapeutic Reendothelialization by Induced Pluripotent Stem Cells After Vascular Injury—Brief Report

Objective—Endothelial damage is an early requisite step for atherosclerosis after vascular injury. It has been reported that vascular wall cells can develop from induced pluripotent stem (iPS) cell–derived fetal liver kinase-1–positive (Flk-1+) cells. Here, we investigated the efficacies of intravenously administered iPS cell–derived Flk-1+ cells on reendothelialization and neointimal hyperplasia in a mouse model of vascular injury. Approach and Results—Femoral arteries of KSN nude mice were injured using a steel wire. Mouse iPS cell–derived Flk-1+ or Flk-1− cells were intravenously injected into those mice at 24 hours after vascular injury. Delivery of iPS cell–derived Flk-1+ cells significantly attenuated neointimal hyperplasia compared with controls. Evans blue staining of the injured vessel revealed that administration of iPS cell–derived Flk-1+ significantly enhanced reendothelialization compared with the Flk-1− cell control group. Recruitment of PKH26-labeled iPS cell–derived Flk-1+ cells to the site of injury was also detectable. Expression level of CXCR4 in iPS cell–derived Flk-1+ cells was 7.5-fold higher than that of iPS cell–derived Flk-1− cells. Stromal cell-derived factor-1&agr; treatment significantly enhanced adhesion and migration of iPS cell–derived Flk-1+ cells to the endothelia, but these were not observed in Flk-1− cells. Conclusions—Intravenously administered iPS cell–derived Flk-1+ cells are recruited to the site of vascular injury, thereby enhancing reendothelialization followed by suppression of neointimal hyperplasia. Administration of iPS cell–derived Flk-1+ cells is a potentially useful therapeutic means for vascular dysfunction and prevention of restenosis after angioplasty.

Evidence for bilateral endolymphatic hydrops in ipsilateral delayed endolymphatic hydrops: preliminary results from examination of five cases.

Abstract Conclusion: After the administration of a standard dose of gadodiamide, an intravenous gadolinium-based contrast agent (GBCA), magnetic resonance imaging (MRI) evaluation of endolymphatic hydrops (EH) became possible in patients with ipsilateral delayed endolymphatic hydrops (DEH). We found that patients with ipsilateral DEH may also have bilateral EH. Objective: MRI evaluation contributes to understanding of the pathological conditions in patients with EH. However, double or triple the standard dose of GBCA is often required to obtain images of high quality. We attempted to examine EH bilaterally in patients with ipsilateral DEH after routine administration of an intravenous GBCA. Methods: GBCA (gadodiamide, 0.2 ml/kg) was administered intravenously to five patients with ipsilateral DEH. Three-dimensional fluid attenuated inversion recovery (3D-FLAIR) MRI was performed with a 3-T MRI scanner 4 h after GBCA administration. Results: In all five patients, EH was observed in the affected vestibules. Moreover, EH was observed bilaterally in four (80%) of five patients with ipsilateral DEH. The region of the deaf ear affected by EH was considerably larger compared with the normal ear in three patients. However, observed regions of EH were of approximately the same size in both ears in patients 4 and 5.

3D-FLAIR MRI findings in patients with low-tone sudden deafness.

Abstract Conclusion: The findings suggest that alterations in the composition of inner ear fluid play important roles in the development of low-tone sudden deafness (LTSD). High-intensity signals on three-dimensional fluid attenuated inversion recovery (3D-FLAIR) may reflect an increased concentration of protein in the inner ear due to the increased permeability of blood vessels. Disordered blood flow in the inner ear is associated with an increased permeability of the blood–labyrinth barrier. Therefore, the disordered blood flow in the cochlea may be closely related to the pathophysiological mechanisms of LTSD. Objectives: The 3D-FLAIR sequence has been used to detect alterations in the composition of inner ear fluid. The purpose of this study was to report imaging findings in cases of LTSD. Methods: 3D-FLAIR magnetic resonance imaging was performed in five women with nonrecurrent-type LTSD. Results: Three of the five patients (60%) showed high-intensity signals in the cochlear basal turn on precontrast 3D-FLAIR. Postcontrast enhancement was not prominent in any patient. In patient 1, the cochlea of the unaffected side showed high-intensity signals. No patients had such signals in the vestibulae or the semicircular canals.

Adiponectin deficiency exacerbates age-related hearing impairment

Obesity-related disorders are closely associated with the development of age-related hearing impairment (ARHI). Adiponectin (APN) exerts protective effects against obesity-related conditions including endothelial dysfunction and atherosclerosis. Here, we investigated the impact of APN on ARHI. APN-knockout (APN-KO) mice developed exacerbation of hearing impairment, particularly in the high frequency range, compared with wild-type (WT) mice. Supplementation with APN prevented the hearing impairment in APN-KO mice. At 2 months of age, the cochlear blood flow and capillary density of the stria vascularis (SV) were significantly reduced in APN-KO mice as compared with WT mice. APN-KO mice also showed a significant increase in terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL)-positive apoptotic cells in the organ of Corti in the cochlea at 2 months of age. At the age of 6 months, hair cells were lost at the organ of Corti in APN-KO mice. In cultured auditory HEI-OC1 cells, APN reduced apoptotic activity under hypoxic conditions. Clinically, plasma APN levels were significantly lower in humans with ARHI. Multiple logistic regression analysis identified APN as a significant and independent predictor of ARHI. Our observations indicate that APN has an important role in preventing ARHI.

Visualization of endolymphatic hydrops after administration of a standard dose of an intravenous gadolinium-based contrast agent

Abstract Conclusion: Even after the administration of a standard dose of an intravenous gadolinium-based contrast agent (GBCA), visualization of endolymphatic hydrops (ELH) became possible in patients with Menieres disease. The next step would be to consistently visualize ELH in the upper part of the cochlea. Objective: To visualize ELH after routine administration of an intravenous GBCA. Methods: An intravenous GBCA (gadodiamide; 0.2 ml/kg) was administered to three patients with unilateral Menieres disease and two healthy volunteers. Three-dimensional fluid attenuated inversion recovery (3D-FLAIR) magnetic resonance imaging (MRI) was performed with a 3 T MRI scanner 4 h later. Results: In all three patients, ELH was observed in the affected vestibules. In contrast, the endolymphatic space of both vestibules was the same size in healthy volunteers. ELH of the cochlea was not observed in any of the subjects. Gadolinium enhancement was insufficient in the upper turns of both cochleae in patients 1 and 3.

Effects of MRI contrast agents (Omniscan™) on vestibular end organs

Conclusion: These findings indicate that application of 4–16-fold diluted Omniscan™ could have depressant effects on the frog vestibular end organs. Based on cochlear studies on cytotoxicity using 8-fold diluted Omniscan, 16-fold diluted Omniscan is optimal for the inner ear and its application could be useful for visualization of endolymphatic hydrops as well as the control of vertiginous attacks in Menieres disease. Objectives: Endolymphatic hydrops could be visualized recently by intratympanic injection of Omniscan diluted 8- or 16-fold with saline using three-dimensional fluid attenuated inversion recovery (3-D FLAIR) MRI. However, the effects of the Omniscan on vestibular end organs are not clearly understood. The purpose of this study was to investigate the effects of Omniscan on vestibular end organs both morphologically and physiologically. Materials and methods: Vestibular hair cells isolated from bullfrogs were loaded with 4–32-fold diluted Omniscan (gadolinium hydrate) and morphological changes were observed. Moreover, compound action potentials of posterior semicircular canals were also observed after application of Omniscan. Results: Application of 4–16-fold diluted Omniscan caused morphological damage of isolated vestibular hair cells and decreased action potentials of the posterior semicircular canal. After application of 32-fold diluted Omniscan, no apparent changes were noted even after 20 min, both morphologically and physiologically.