Tissue-engineered cultured periosteum used with platelet-rich plasma and hydroxyapatite in treating human osseous defects

BACKGROUND: The aim of the present controlled clinical study was to compare the clinical response of human cultured periosteum (HCP) sheets in combination with platelet-rich plasma (PRP) and porous hydroxyapatite (HA) granules to a mixture of PRP and HA in the treatment of human infrabony periodontal defects. METHODS: Thirty interproximal infrabony osseous defects in 30 healthy, non-smoking subjects diagnosed with chronic periodontitis were included in this study. The subjects were randomly assigned to the test group (HCP sheets combined with PRP and HA) or the control group (PRP with HA). Clinical and radiographic measurements were made at baseline and the 12-month post-surgical evaluation. RESULTS: Compared to baseline, the 12-month results indicated that both treatment modalities resulted in statistically significant changes (P <0.01) in the gingival index, bleeding on probing, probing depth, clinical attachment level, and radiographic infrabony defect depth. Compared to the control group, the test group exhibited a statistically significantly more favorable change in clinical attachment gain (3.9 +/- 1.6 mm versus 2.7 +/- 1.3 mm; P <0.05), vertical relative attachment gain (83.5% +/- 31.7% versus 55.0% +/- 21.9%; P <0.05), and radiographic infrabony defect fill (4.9 +/- 1.2 mm versus 3.2 +/- 1.1 mm; P <0.01). CONCLUSIONS: Compared to PRP with HA, treatment with a combination of HCP sheets, PRP, and HA led to a significantly more favorable clinical improvement in infrabony periodontal defects. A factor likely contributing to these favorable clinical results is the presence of osteogenic cells in the HCP sheets, which provided greater regeneration potential.     

A third-generation, long-acting, dihydropyridine calcium antagonist, azelnidipine, attenuates stent-associated neointimal formation in non-human primates

BACKGROUND: Calcium antagonists have been shown to reduce atherogenesis and improve clinical outcomes in atherosclerotic vascular disease. No study has so far, however, addressed the effects of calcium antagonists on stent-associated neointimal formation. We therefore investigated whether a third-generation calcium antagonist, azelnidipine, attenuates in-stent neointimal formation in non-human primates. METHOD: Male cynomolgus monkeys were fed a high cholesterol diet for 4 weeks, and were randomly assigned to three groups: a vehicle group and two other groups treated with azelnidipine at 3 and 10 mg/kg per day for an additional 24 weeks (n = 12 each). Multi-link stents were then implanted in the iliac artery. RESULTS: Azelnidipine at the high dose reduced neointimal thickness (0.25 +/- 0.02 versus 0.19 +/- 0.02 mm; P < 0.05). Azelnidipine also reduced local oxidative stress and monocyte chemoattractant protein 1 (MCP-1) expression. No difference was found between the three groups in the degrees of injury score, inflammation score, plaque neovascularization, or plasma lipid levels. Azelnidipine also reduced MCP-1-induced proliferation/migration of vascular smooth muscle cells in vitro. CONCLUSIONS: This study demonstrated for the first time that azelnidipine attenuates in-stent neointimal formation associated with the reduced expression of MCP-1 and smooth muscle proliferation/migration in the neointima. These data in non-human primates suggest potential clinical benefits of azelnidipine as a 'vasculoprotective calcium antagonist' in patients undergoing vascular interventions.     

Secure and effective gene delivery system of plasmid DNA coated by polynucleotide

Polynucleotides are anionic macromolecules which are expected to transfer into the targeted cells through specific uptake mechanisms. So, we developed polynucleotides coating complexes of plasmid DNA (pDNA) and polyethylenimine (PEI) for a secure and efficient gene delivery system and evaluated their usefulness. Polyadenylic acid (polyA), polyuridylic acid (polyU), polycytidylic acid (polyC), and polyguanylic acid (polyG) were examined as the coating materials. pDNA/PEI/polyA, pDNA/PEI/polyU, and pDNA/PEI/polyC complexes formed nanoparticles with a negative surface charge although pDNA/PEI/polyG was aggregated. The pDNA/PEI/polyC complex showed high transgene efficiency in B16-F10 cells although there was little efficiency in pDNA/PEI/polyA and pDNA/PEI/polyU complexes. An inhibition study strongly indicated the specific uptake mechanism of pDNA/PEI/polyC complex. Polynucleotide coating complexes had lower cytotoxicity than pDNA/PEI complex. The pDNA/PEI/polyC complex showed high gene expression selectively in the spleen after intravenous injection into mice. The pDNA/PEI/polyC complex showed no agglutination with erythrocytes and no acute toxicity although these were observed in pDNA/PEI complex. Thus, we developed polynucleotide coating complexes as novel vectors for clinical gene therapy, and the pDNA/PEI/polyC complex as a useful candidate for a gene delivery system.     

Direct evidence for increased hydroxyl radicals originating from superoxide in the failing myocardium

Experimental and clinical studies have suggested an increased production of reactive oxygen species (ROS) in the failing myocardium. The present study aimed to obtain direct evidence for increased ROS and to determine the contribution of superoxide anion (*O(2)(-)), H(2)O(2), and hydroxy radical (*OH) in failing myocardial tissue. Heart failure was produced in adult mongrel dogs by rapid ventricular pacing at 240 bpm for 4 weeks. To assess the production of ROS directly, freeze-clamped myocardial tissue homogenates were reacted with the nitroxide radical, 4-hydroxy-2,2,6, 6,-tetramethyl-piperidine-N-oxyl, and its spin signals were detected by electron spin resonance spectroscopy. The rate of electron spin resonance signal decay, proportional to *OH level, was significantly increased in heart failure, which was inhibited by the addition of dimethylthiourea (*OH scavenger) into the reaction mixture. Increased *OH in the failing heart was abolished to the same extent in the presence of desferrioxamine (iron chelator), catalase (H(2)O(2) scavenger), and 4,5-dihydroxy-1,3-benzene disulfonic acid (Tiron; LaMotte) (*O(2)(-) scavenger), indicating that *OH originated from H(2)O(2) and *O(2)(-). Further, *O(2)(-) produced in normal myocardium in the presence of antimycin A (mitochondrial complex III inhibitor) could reproduce the increase of H(2)O(2) and *OH seen in the failing tissue. There was a significant positive relation between myocardial ROS level and left ventricular contractile dysfunction. In conclusion, in the failing myocardium, *OH was produced as a reactive product of *O(2)(-) and H(2)O(2), which might play an important role in left ventricular failure.     

Effect of leukocytapheresis therapy using a leukocyte removal filter in Crohn's disease

Eighteen patients with active Crohn's disease were treated with one leukocytapheresis session per week for a five-week intensive therapy, decreasing to one leukocytapheresis session per month for five sessions of initial maintenance therapy. Nutritional indices, inflammatory reactions, flow cytometry profiles, and cytokine production were also assessed before and after the intensive and initial maintenance therapy. Nine of the patients (50%) attained remission at the end of the intensive therapy. The nine non-remission patients had exhibited longer periods of suffering and more severely affected sites prior to the therapy. In 14 of 18 patients (77.8%), the nutritional indices, Internal Organization of Inflammatory Bowel Disease (IOIBD) score and Crohn's Disease Activity Index (CDAI) improved from the pretherapy levels, but only the remission group (50%) showed improvement in C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR). The remission group showed significantly higher pretherapy CD4+ CD45+ cell ratios and interleukin-2 (IL-2) production than the non-remission group, and significantly lower activated cells.     

Monocyte chemoattractant protein-1 activates a regional Th1 immunoresponse in nephritis of MRL/lpr mice

OBJECTIVE: Monocyte chemoattractant protein-1 (MCP-1) is upregulated and recruits and activates inflammatory cells in nephritis of MRL lpr mice. It has been shown that anti-MCP-1 gene therapy is specifically effective in nephritis, while it was apparent that an imbalance towards Th1 predominance accelerates nephritis in MRL/lpr mice. The aim of this study was to clarify whether blockade of the MCP-1 signal by anti-MCP-1 gene therapy influences the Th1/Th2 balance in MRL/lpr mice. METHOD: An NH2-terminal deletion mutant of the MCP-1 gene (7ND) was injected into the skeletal muscles of MRL/Ipr mice with advanced stage nephritis to suppress MCP-1 and its receptor (CCR2) signaling pathway. We evaluated the local tissue production of cytokines in splenocytes and microdissected infiltrating cells within the glomeruli or interstitium. RESULT: Although the production of cytokines in splenocytes was not influenced by anti-MCP-1 gene therapy, kidney glomeruli IL-12 mRNA production and interstitium-infiltrating cell production of IL-12 and IFN-gamma mRNA were significantly reduced. CONCLUSION: The blockade of MCP-1 gene therapy does not influence helper T cell polarization, but acts directly on the regional Th1 immunoreaction in MRL/lpr mice.     

Polymorphisms of apolipoprotein e and methylenetetrahydrofolate reductase in the Japanese population

AIM: The aim of this study is to analyze the effect of apolipoprotein E (apo E) and methylenetetrahydrofolate reductase (MTHFR) gene polymorphisms on serum lipid and homocysteine levels in the general Japanese population. METHODS: We analyzed the polymorphisms in individuals randomly selected from among participants of Serum Lipid Survey 2000. RESULTS: The frequency of the epsilon2, epsilon3, and epsilon4 alleles of APOE was 4.2, 85.3, and 10.5%, respectively. Individuals with the genotype epsilon4/epsilon4 had the highest total and low-density lipoprotein (LDL) cholesterol levels, while those with epsilon2/epsilon2 had the lowest. Individuals with the epsilon2/epsilon2 and epsilon2/epsilon4 genotypes had higher remnant-like particles (RLP)-cholesterol levels than those with epsilon2epsilon3, epsilon3epsilon3, and epsilon3epsilon4. There was a trend for individuals with the epsilon2/epsilon4 and epsilon2/epsilon2 genotypes to have higher triglyceride levels, although the difference was not significant. The presence of the T allele in a MTHFR polymorphism (C667T) was associated with higher homocysteine levels, which is more prominent in men than in women. CONCLUSION: Thus in our large-scale analysis we have shown that RLP-cholesterol is better associated with, APOE genotype than triglyceride and the effect of the T allele on MTHFR polymorphism (C667T) homocysteine levels is more prominent in men than in women among Japanese.