Shigefumi Suehiro

Dose-Dependent Contribution of CD34-Positive Cell Transplantation to Concurrent Vasculogenesis and Cardiomyogenesis for Functional Regenerative Recovery After Myocardial Infarction

Background— Multilineage developmental capacity of the CD34+ cells, especially into cardiomyocytes and smooth muscle cells (SMCs), is still controversial. In the present study we performed a series of experiments to prove our hypothesis that vasculogenesis and cardiomyogenesis after myocardial infarction (MI) may be dose-dependently enhanced after CD34+ cell transplantation. Methods and Results— Peripheral blood CD34+ cells were isolated from total mononuclear cells of patients with limb ischemia by apheresis after 5-day administration of granulocyte colony-stimulating factor. PBS and 1×103 (low), 1×105 (mid), or 5×105 (high) CD34+ cells were intramyocardially transplanted after ligation of the left anterior descending coronary artery of nude rats. Functional assessments with the use of echocardiography and a microtip conductance catheter at day 28 revealed dose-dependent preservation of left ventricular function by CD34+ cell transplantation. Necropsy examination disclosed dose-dependent augmentation of capillary density and dose-dependent inhibition of left ventricular fibrosis. Immunohistochemistry for human-specific brain natriuretic peptide demonstrated that human cardiomyocytes were dose-dependently observed in ischemic myocardium at day 28 (high, 2480±149; mid, 1860±141; low, 423±9; PBS, 0±0/mm2; P<0.05 for high versus mid and mid versus low). Immunostaining for smooth muscle actin and human leukocyte antigen or Ulex europaeus lectin type 1 also revealed dose-dependent vasculogenesis by endothelial cell and SMC development after CD34+ cell transplantation. Reverse transcriptase–polymerase chain reaction indicated that human-specific gene expression of cardiomyocyte (brain natriuretic peptide, cardiac troponin-I, myosin heavy chain, and Nkx 2.5), SMC (smooth muscle actin and sm22&agr;), and endothelial cell (CD31 and KDR) markers were dose-dependently augmented in MI tissue. Conclusions— Human CD34+ cell transplantation may have significant and dose-dependent potential for vasculogenesis and cardiomyogenesis with functional recovery from MI.

Calorie Restriction Improves Cardiovascular Risk Factors via Reduction of Mitochondrial Reactive Oxygen Species in Type II Diabetic Rats

Uncoupling protein 2 (UCP2) is an important regulator of intracellular reactive oxygen species (ROS) production. We determined the effects of calorie restriction (CR) on the dynamic aspects of mitochondrial ROS production, UCP2, and the nitric oxide (NO)-cGMP pathway in the cardiovascular tissues of type II diabetic Otsuka Long-Evans Tokushima Fatty (OLETF) rats. Some rats were on restricted diets (30% reduction from free intake) from age 29 to 42 weeks. Blood glucose, hemoglobin A1c, plasma levels of free fatty acid, triacylglycerol, and plasminogen activator inhibitor-1 in OLETF rats were significantly higher than those in nondiabetic control [Long-Evans Tokushima Otsuka (LETO)] rats at 29 weeks. Mitochondrial ROS production and UCP2 expression significantly increased in the heart and aorta of OLETF rats compared with those in LETO rats. A fibrogenic growth factor, transforming growth factor (TGF)-β1 in the coronary vessels, endothelial nitric-oxide synthase, and aortic nitrotyrosine were increased in OLETF rats at 42 weeks. In contrast, an index of the NO-cGMP pathway, phosphorylated vasodilator-stimulated phosphoprotein, and superoxide dismutase activity in the aorta were significantly diminished. The relationship between UCP2 and ROS production in the cardiovascular function of diabetic rats being fed a calorie-restricted diet is unknown. These abnormalities in OLETF rats were reversed to normal levels by CR. CR significantly improved the NO-cGMP pathway via normalizing ROS generation in OLETF rats. A decrease in UCP2 expression by CR may be a compensatory mechanism to counteract decreased intracellular oxidative stress. The data suggest that CR may prevent cardiovascular tissues from oxidative stress provoked by diabetes mellitus.

S-allyl cysteine prevents CCl4-induced acute liver injury in rats

Aged garlic extract (AGE) possesses multiple biological activities. We evaluated the protective effect of S-allyl cysteine (SAC), one of the organosulfur compounds of AGE, against carbon tetrachloride (CCl4)-induced acute liver injury in rats. SAC was administrated intraperitoneally (50–200 mg/kg). SAC significantly suppressed the increases of plasma ALT and LDH levels. SAC also attenuated histological liver damage. CCl4 administration induced lipid peroxidation accompanied by increases in the plasma malondialdehyde and hepatic 4-hydroxy-2-nonenal levels, and SAC dose-dependently attenuated these increases. The hepatic total level of hydroxyoctadecadienoic acid (HODE), a new oxidative stress biomarker, was closely correlated with the amount of liver damage. These results suggest that SAC decreased CCl4-induced liver injury by attenuation of oxidative stress, and may be a better therapeutic tool for chronic liver disease.

CORM-3-derived CO modulates polymorphonuclear leukocyte migration across the vascular endothelium by reducing levels of cell surface-bound elastase.

Recently, it has been shown that carbon monoxide (CO)-releasing molecule (CORM)-released CO can suppress inflammation. In this study, we assessed the effects and potential mechanisms of a ruthenium-based water-soluble CO carrier [tricarbonylchloroglycinate-ruthenium(II) (CORM-3)] in the modulation of polymorphonuclear leukocyte (PMN) inflammatory responses in an experimental model of sepsis. Sepsis in mice was induced by cecal ligation and puncture. CORM-3 (3 mg/kg iv) was administered 15 min after the induction of cecal ligation and puncture. PMN accumulation in the lung (myeloperoxidase assay), bronchoalveolar lavage (BAL) fluid, and lung vascular permeability (protein content in BAL fluid) were assessed 6 h later. In in vitro experiments, human PMNs were primed with LPS (10 ng/ml) and subsequently stimulated with formyl-methionyl-leucylphenylalanine (fMLP; 100 nM). PMN production of ROS (L-012/dihydrorhodamine-123 oxidation), degranulation (release of elastase), and PMN rolling, adhesion, and migration to/across human umbilical vein endothelial cells (HUVECs) were assessed in the presence or absence of CORM-3 (1-100 muM). The obtained results indicated that systemically administered CORM-3 attenuates PMN accumulation and vascular permeability in the septic lung. Surprisingly, in in vitro experiments, treatment of PMNs with CORM-3 further augmented LPS/fMLP-induced ROS production and the release of elastase. The latter effects, however, were accompanied by an inability of PMNs to mobilize elastase to the cell surface (plasma membrane), an event required for efficient PMN transendothelial migration. The CORM-3-induced decrease in cell surface levels of elastase was followed by decreased PMN rolling/adhesion to HUVECs and complete prevention of PMN migration across HUVECs. In contrast, treatment of HUVECs with CORM-3 had no effect on PMN transendothelial migration. Taken together, these findings indicate that, in sepsis, CORM3-released CO, while further amplifying ROS production and degranulation of PMNs, concurrently reduces the levels of cell surface-bound elastase, which contributes to suppressed PMN transendothelial migration.

PlGF Repairs Myocardial Ischemia through Mechanisms of Angiogenesis, Cardioprotection and Recruitment of Myo-Angiogenic Competent Marrow Progenitors

Rationale Despite preclinical success in regenerating and revascularizing the infarcted heart using angiogenic growth factors or bone marrow (BM) cells, recent clinical trials have revealed less benefit from these therapies than expected. Objective We explored the therapeutic potential of myocardial gene therapy of placental growth factor (PlGF), a VEGF-related angiogenic growth factor, with progenitor-mobilizing activity. Methods and Results Myocardial PlGF gene therapy improves cardiac performance after myocardial infarction, by inducing cardiac repair and reparative myoangiogenesis, via upregulation of paracrine anti-apoptotic and angiogenic factors. In addition, PlGF therapy stimulated Sca-1+/Lin− (SL) BM progenitor proliferation, enhanced their mobilization into peripheral blood, and promoted their recruitment into the peri-infarct borders. Moreover, PlGF enhanced endothelial progenitor colony formation of BM-derived SL cells, and induced a phenotypic switch of BM-SL cells, recruited in the infarct, to the endothelial, smooth muscle and cardiomyocyte lineage. Conclusions Such pleiotropic effects of PlGF on cardiac repair and regeneration offer novel opportunities in the treatment of ischemic heart disease.

Risk Factor Analysis in Patients With Liver Cirrhosis Undergoing Cardiovascular Operations

BACKGROUND Variable outcomes of cardiac operations have been reported in cirrhotic patients, but no definitive predictive prognostic factors have been established. This retrospective study assessed operative results to identify risk factors associated with morbidity after cardiovascular operations in cirrhotic patients. METHODS The study comprised 42 cirrhotic patients who underwent cardiovascular operations from January 1991 to January 2009. Thirty patients were Child-Turcotte-Pugh class A, and 12 were class B. Hospital morbidity occurred in 13 patients (31.0%; M group), including 4 who died in-hospital. Patients without severe complications (N group) were compared with the M group patients. The Model for End-Stage Liver Disease (MELD) score was evaluated in 25 patients. RESULTS Significant differences in hospital morbidity between the M vs N groups were identified for platelet count (8.7 +/- 3.8 vs 12.1 +/- 4.2 x 10(4)/microL), MELD score (17.8 +/- 5.3 vs 9.8 +/- 4.9), operation time (370 +/- 88 vs 313 +/- 94 minutes), and cardiopulmonary bypass time (174 +/- 46 vs 149 +/- 53 minutes) in univariate analyses (p < 0.005). Platelet count, operation time, and age were significantly associated with hospital morbidity in multivariate analyses (p < 0.005). Platelet count of 9.6 x 10(4)/microL and MELD score of 13 were cutoff values for hospital morbidity. CONCLUSIONS Careful consideration of operative indications and methods are necessary in cirrhotic patients with low platelet counts or high MELD scores. A high incidence of hospital morbidity is predicted in patients with platelet counts of less than 9.6 x 10(4)/microL or MELD scores exceeding 13.

Repair of postinfarction ventricular septal defect with joined endocardial patches.

We describe a technique for repair of ventricular septal defect with two bovine pericardial patches joined to make a single pouch. The size of the finished pouch can be adjusted as desired after both patches are sutured to the myocardium, unlike when one patch is used. Suturing is easier than when a single patch is used.

Does overgrowth of costal cartilage cause pectus excavatum? A study on the lengths of ribs and costal cartilages in asymmetric patients

PURPOSE The cause of pectus excavatum has been hypothesized to be overgrowth of the costal cartilage. According to this theory, the length of costal cartilages must be longer in the side of deep depression in asymmetric patients. To challenge this hypothesis, we measured the lengths of ribs and costal cartilages and investigated lateral differences. SUBJECTS AND METHODS Twenty-four adolescent and adult patients with asymmetric pectus excavatum (14-30 years of age) with no history of surgery were investigated in this study. The fifth and sixth ribs and costal cartilages were individually traced to measure their full lengths on 3-dimensional computed tomographic (CT) images. As an index of asymmetry, sternal rotation angle was measured in the chest CT images. Patients with a 21 degrees or greater angle of sternal twist were designated as an asymmetric group and those with an angle of smaller than 20 degrees as a symmetric group. Lateral differences in the fifth and sixth costal and costal cartilage lengths were compared between the groups. RESULTS On comparison of the costal and costal cartilage lengths in the asymmetric group, the right fifth ribs and costal cartilages were significantly shorter than the left (P = .02 and .03, respectively), and right sixth ribs were also significantly shorter than the left (P = .004), but right sixth costal cartilages were not (P = .31). In the symmetric group, the lengths of the left and right fifth ribs and costal cartilages were showing no significant difference (P = .20 and P = .80, respectively), and those of the sixth ribs and costal cartilage were also showing no significant difference (P = .97 and P = .64, respectively). DISCUSSION The ribs and costal cartilages on the right side with severer depression were significantly shorter or not different than those on the contralateral side. Based on these findings, the theory of costal cartilage overgrowth is contradictory. The etiology of asymmetric chest deformity should be reevaluated.

Correlation between Dynamic Computed Tomographic and Histopathological Findings in the Diagnosis of Small Hepatocellular Carcinoma

Background/Aims: To determine the characteristic image findings on dynamic computed tomography (CT) for small hepatocellular carcinoma (HCC), we evaluated the correlation of histopathological and radiological findings with respect to the angioarchitecture in small HCCs. Methods: CT and early- and late-phase dynamic CT findings of 80 small HCCs (≤3 cm) were divided into iso-, high, low, and mixed density. We studied the correlation between the imaging findings and the histopathological findings as follows: differentiation grade; presence of fibrous capsule; presence of Glisson’s sheath, and growth pattern. Results: High-density early-phase CT and low-density late-phase CT correlated significantly with moderately/poorly differentiated HCCs, which have a fibrous capsule, no Glisson’s sheath, and an expansive growth pattern. In contrast, well-differentiated HCCs with a Glisson’s sheath and a replacing pattern (early HCC) appeared as iso-dense lesions in the early and late phases. Well-differentiated HCCs (non-early wHCC) demonstrated various density images in the early phase and low-density images in the late phase. Conclusions: Dynamic CT is an economic and simple diagnostic tool for planning treatment of small HCC lesions because of the multistep nature of HCC carcinogenesis and the hemodynamic changes of tumor blood flow.

Supplementation of α-tocopherol improves cardiovascular risk factors via the insulin signalling pathway and reduction of mitochondrial reactive oxygen species in type II diabetic rats

This study determined the effects of α- and γ-tocopherol supplementation on metabolic control and oxidative stress in type 2 diabetic Otsuka Long–Evans Tokushima Fatty (OLETF) rats. Blood glucose, haemoglobin A1c (HbA1c), urinary protein, plasma free fatty acid, triacylglycerol and plasminogen activator inhibitor-1 (PAI-1) levels in OLETF rats were significantly higher than in non-diabetic control Long–Evans Tokushima Otsuka (LETO) rats. α-Tocopherol inhibited the increase in urinary protein, blood glucose, HbA1c and PAI-1 levels, but γ-tocopherol did not. Plasma and hepatic lipid peroxidation and hepatic steatosis were increased in OLETF rats. α-Tocopherol decreased lipid peroxidation. Mitochondrial reactive oxygen species production and uncoupling protein 2 (UCP2) expression were significantly increased in the heart and aorta of OLETF rats compared with LETO rats. Endothelial NO synthase and aortic nitrotyrosine were increased in OLETF rats. In contrast, the expression of phosphorylated vasodilator-stimulated phosphoprotein and glucose transporter 4 in the aorta was significantly decreased in OLETF rats. These abnormalities were reversed by α-tocopherol. These findings suggest that α-tocopherol may prevent cardiovascular tissues from oxidative stress and insulin signalling disorder resulting from diabetes mellitus.