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Although prostaglandin I2 is used to treat pulmonary hypertension (PH), continuous intravenous administration is necessary. We investigated whether human PGIS (hPGIS) gene transfer using adeno-associated virus (AAV) vector was effective in treating an animal model of PH. PH was induced by subjecting mice to 10% O(2). Type 1-AAV-hPGIS was injected into the left thigh muscle after 24h. Significant PH was induced at 8 weeks, but AAV-hPGIS administration significantly inhibited the increase in RV systolic pressure. PH-induced BNP up-regulation in the RV was reduced to the control level. The severe medial thickening of pulmonary arteries in PH was significantly suppressed by AAV-hPGIS. The hPGIS gene was detected only on the injected side. No pathological changes were observed at the injected site. At 24 weeks, all PH mice were deceased, but 47% of AAV-hPGIS-treated mice survived. This study demonstrated that AAV-hPGIS administration was effective in treating PH and prolonging survival.

The administration of granulocyte colony-stimulating factor (G-CSF) after myocardial infarction (MI) improves cardiac function and survival rates in mice. It was also reported recently that bone marrow (BM)-derived c-kit(+) cells or macrophages in the infarcted heart are associated with improvement of cardiac remodeling and function. These observations prompted us to examine whether BM-derived hematopoietic cells mobilized by G-CSF administration after MI play a beneficial role in the infarct region. A single hematopoietic stem cell from green fluorescent protein (GFP)-transgenic mice was used to reconstitute hematopoiesis in each experimental mouse. MI was then induced, and the mice received G-CSF for 10 days. In the acute phase, a number of GFP(+) cells showing the elongated morphology were found in the infarcted area. Most of these cells were positive for vimentin and alpha-smooth muscle actin but negative for CD45, indicating that they were myofibroblasts. The number of these cells was markedly enhanced by G-CSF administration, and the enhanced myofibroblast-rich repair was considered to lead to improvements of cardiac remodeling, function, and survival rate. Next, G-CSF-mobilized monocytes were harvested from the peripheral blood of GFP-transgenic mice and injected intravenously into the infarcted mice. Following this procedure, GFP(+) myofibroblasts were observed in the infarcted myocardium. These results indicate that cardiac myofibroblasts are hematopoietic in origin and could arise from monocytes/macrophages. MI leads to the recruitment of monocytes, which differentiate into myofibroblasts in the infarct region. Administration of G-CSF promotes this recruitment and enhances cardiac protection.

Inhalation and oral steroids therapy has been used in the management of asthma, chronic obstructive pulmonary disease (COPD) and interstitial lung disease. This article described the effects of inhaled corticosteroids on bone. Inhaled corticosteroids are the standard of therapy in asthma and are commonly prescribed for chronic obstructive pulmonary disease. Accumulating evidence suggests that the effect of inhaled corticosteroids on bone is not small. Recent studies have shown that long-term use of inhaled corticosteroid at moderate or high doses is also associated with increased risk for the effects of inhaled corticosteroids on bone. Older age, inmobility and steroid treatment are major risk factors of osteoporosis. Treatment with bisphosphonate will be recommended in these patients, so earlier prevention for corticosteroids induced osteoporosis is necessary at administration of inhaled corticosteroids.

Sympathetic innervation is critical for effective cardiac function. However, the developmental and regulatory mechanisms determining the density and patterning of cardiac sympathetic innervation remain unclear, as does the role of this innervation in arrhythmogenesis. Here we show that a neural chemorepellent, Sema3a, establishes cardiac sympathetic innervation patterning. Sema3a is abundantly expressed in the trabecular layer in early-stage embryos but is restricted to Purkinje fibers after birth, forming an epicardial-to-endocardial transmural sympathetic innervation patterning. Sema3a(-/-) mice lacked a cardiac sympathetic innervation gradient and exhibited stellate ganglia malformation, which led to marked sinus bradycardia due to sympathetic dysfunction. Cardiac-specific overexpression of Sema3a in transgenic mice (SemaTG) was associated with reduced sympathetic innervation and attenuation of the epicardial-to-endocardial innervation gradient. SemaTG mice demonstrated sudden death and susceptibility to ventricular tachycardia, due to catecholamine supersensitivity and prolongation of the action potential duration. We conclude that appropriate cardiac Sema3a expression is needed for sympathetic innervation patterning and is critical for heart rate control.

Granulocyte colony-stimulating factor (G-CSF) is known to mobilize bone marrow stem cells into the peripheral circulation. This study was designed to investigate whether G-CSF by itself or in combination with hepatocyte growth factor (HGF) can promote vasculogenesis and angiogenesis in murine hind limb ischemia. Hind limb ischemia was induced in BALB/c nude or C57/BL6 mice that received bone marrow transplantation from green fluorescent protein (GFP)-transgenic mice. In the HGF group, hHGF expression plasmid was injected into the ischemic muscles. In the G-CSF group, G-CSF was administered subcutaneously for 10 days. The G-CSF+HGF group was concomitantly treated with G-CSF and HGF, and the control group received no treatment. All effects were confirmed at 4 weeks. The G-CSF+HGF group had a higher laser Doppler blood perfusion index, higher microvessel density, and a lower incidence of hind limb necrosis than the other groups. Confocal laser microscopy revealed that a number of GFP-positive cells infiltrated to the vasculature of the ischemic area. Some of the GFP positive cells were clearly co-immunostained with alpha-smooth muscle actin as well as von Willebrand factor. G-CSF-mobilized stem cells co-expressed CD49d and CD34, which would have promoted their adhesion to cells in the ischemic muscle that expressed HGF-induced vascular cell adhesion molecule-1. The combination of G-CSF and HGF had a significant synergistic effect, suggesting that the combination of mobilization of stem cells from bone marrow to peripheral circulation and their recruitment to the ischemic area might potentiate angiogenesis and vasculogenesis.

Background: Molecular mechanisms regulating the cardiac sensory nervous system remain poorly understood. Cardiac sensory nerve impairment causes silent myocardial ischemia, a main cause of sudden death in diabetes mellitus (DM). The present study focused on the roles of nerve growth factor (NGF) in the regulation of the cardiac sensory nervous system and analyzed the mechanism of silent myocardial ischemia in DM., Methods and Results: We screened neurotrophic factors and found that cardiac sensory nerves developed in parallel with NGF synthesized in the heart. Cardiac nociceptive sensory nerves that were immunopositive for calcitonin gene-related peptide, dorsal root ganglia (DRG), and the dorsal horn were markedly retarded in NGF-deficient mice, whereas cardiac-specific overexpression of NGF rescued these deficits. DM was induced with streptozotocin in wild-type and transgenic mice overexpressing NGF in the heart. Downregulation of NGF, calcitonin gene-related peptide-immunopositive cardiac sensory denervation, and atrophic changes in DRG were observed in DM-induced wild-type mice, whereas these deteriorations were reversed in DM-induced NGF transgenic mice. Cardiac sensory function, measured by myocardial ischemia-induced c-Fos expression in DRG, was also downregulated by DM in the wild-type mice but not in NGF transgenic mice. Direct gene transfer of NGF in the diabetic rat hearts improved impaired cardiac sensory innervation and function, determined by electrophysiological activity of cardiac afferent nerves during myocardial ischemia., Conclusions: These findings demonstrate that the development and regulation of the cardiac sensory nervous system are dependent on NGF synthesized in the heart and that DM-induced NGF reduction causes cardiac sensory neuropathy.

To develop a novel nonviral gene carrier, three types of polyamine-dialkyl phosphates conjugates were synthesized via an unprecedented synthetic intermediate, dimerized dicetyl phosphate (DCP) anhydride, and the transfection efficiency and the complexation properties of the conjugate-DNA were evaluated. Condensation of DCP by 1,3,5-triisopropylbenzenesulfonyl chloride, TPSCl, gives the dimerized anhydride, which is stable enough to isolate by column chromatography in approximately 90% yield. The anhydride is reactive with various amines, i.e., spermidine, spermine, and polyethylenimine (PEI(1800)), providing corresponding polyamine-DCP conjugates via phosphoramidate linkage. The polyamine-DCP conjugates exhibited moderate transfection efficacy evaluated by beta-galactosidase assay. The conjugate-DNA complex was observed by using an atomic force microscope (AFM), revealing that the PEI(1800)-DCP conjugate, which showed the most efficient transfection, enables the formation of the more compact complex with DNA.

The plica synovialis mediopatellaris, a so-called shelf, is found in 20%-50% of normal knees. A shelf syndrome with pain or functional injury is uncommon, however, and cases that occur during the adolescent growth phase are rare. We report a rare case of a bilateral shelf syndrome that developed simultaneously in both knees. The patient was a 13-year-old girl who presented with bilateral knee pain and severely decreased range of motion. The symptoms were not alleviated after internal therapy, so arthroscopy was performed. It showed a white hypertrophied synovialis mediopatellaris, a shelf from the suprapatellar pouch, in both knees. The shelf had an opening between the free margin and the insertion point, giving rise to a synovialis mediopatellaris with a detached structure (type D according to the Sakakibara classification). The cause of the pain was thought to be impingement of the synovialis mediopatellaris. The shelf was ablated by arthroscopy using a holmium-YAG laser. Since the patient was a 13-year-old girl in the growth phase, it is possible that the growth resulted in tension on the shelf remnant bilaterally, intruding into the medial aspect of the patellofemoral joint and increasing the tension, resulting in the symptoms reported.

We investigated the effects of erythropoietin (Epo) in glial cell development, especially the maturation of late stage immature oligodendrocytes and the proliferation of astrocytes. Epo mRNA level in oligodendrocytes was much more prominent than those in neurons or astrocytes, which were the same as those in the young adult kidney, while Epo receptor (Epo-R) mRNA level were almost the same among neural cells, kidney and liver tissues. On immunohistochemical examination, Epo-R expression was also detected in O4-positive immature oligodendrocytes and glial fibrillary acidic protein positive astrocytes. These results suggested that types of both glial cells are responsive to Epo. The numbers of mature oligodendrocytes, which are characterized by myelin basic protein and process development, were increased by treatment with recombinant human Epo (rhEpo) (0.001-0.1 U/ml). The maturation of oligodendrocytes was also enhanced by coculture with astrocytes in vitro. However, when mixed cultured cells (oligodendrocytes+astrocytes) were treated with anti-Epo antibody and/or soluble Epo-R, the differentiation of oligodendrocytes was partially inhibited. Interestingly, high dose rhEpo (1, 3, 10 U/ml) markedly enhanced the proliferation of astrocytes. These results suggested that Epo not only promotes the differentiation and/or maturation in oligodendrocytes, but also enhances the proliferation of astrocytes. It is generally accepted that astrocytes produce Epo, and therefore Epo might act on astrocytes in an autocrine manner. The astrocytes stimulated with Epo may further accelerate the maturation of oligodendrocytes. These comprehensive effects of Epo might also affect the ability of oligodendrocyte lineage cells to promote myelin repair in the normal and damaged adult central nervous system.

The purpose of this study was to examine how chondrocytes are involved in the molecular mechanism of inflammation in rheumatoid arthritis (RA). A chondrosarcoma cell line (OUMS-27) was cultured and treated with interleukin-1beta (IL-1beta). Changes in the expression levels of matrix metalloproteinase-1 (MMP-1), metalloproteinase-13 (MMP-13), and gliostatin/platelet-derived endothelial cell growth factor (GLS/PD-ECGF) were assessed by reverse transcription polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assays. IL-1beta induced the expressions of MMP-1, MMP-13, and GLS mRNAs and proteins in a dose-dependent manner. Selective inhibition of the p38 mitogen-activated protein kinase (p38 MAPK) pathway with SB 203580 and SB 202190 blocked the expression of MMP-1, MMP-13, and GLS more strongly than selective in hibition of the extracellular signal-regulated kinase 1 and 2 (ERK1/2) pathway by PD 98059. These findings suggest that chondrocytes may intensify cartilage destruction and inflammation in RA by the induction of MMP-1, MMP-13, and GLS by IL-1beta and that the p38 MAPK pathway plays an important role in these inductions.