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Biomedical subjects

M Yoshinari

Publications and source records attributed to M Yoshinari.

At least 19 recordsLinked to original sources

Quartz-crystal microbalance-dissipation technique for the study of initial adsorption of fibronectin onto tresyl chloride-activated titanium.

The immobilization of cell-adhesive proteins onto titanium implants improves biological response at the implant-tissue interface. Previous studies demonstrated the easy and direct attachment of fibronectin onto titanium with the use of a 2,2,2-trifluoroethanesulfonyl chloride (tresyl chloride) activation technique. The present study investigated the initial adsorption behavior of fibronectin on tresyl chloride-activated titanium by the quartz-crystal microbalance-dissipation (QCM-D) technique. The crystal resonant frequency and the dissipation shift of the oscillator were simultaneously measured by the injection of fibronectin/phosphate-buffered saline solution (pH = 7.4). The tresyl chloride-activated titanium surface showed a faster and greater decrease in frequency than that of untreated titanium, indicating that a greater amount of fibronectin was adsorbed in the former case during a 120-min adsorption. The dissipation-frequency plots revealed that, during the initial stage of adsorption, the bond between fibronectin and tresyl chloride-activated titanium is stronger than that between fibronectin and untreated titanium. The QCM-D technique can provide new insights into the adsorption mechanism of fibronectin.

Adsorption↗

Cloning and functional characterization of MEF2D/DAZAP1 and DAZAP1/MEF2D fusion proteins created by a variant t(1;19)(q23;p13.3) in acute lymphoblastic leukemia.

We analyzed the TS-2 acute lymphoblastic leukemia (ALL) cell line that contains a t(1;19)(q23;p13.3) but lacks E2A-PBX1 fusion typically present in leukemias with this translocation. We found that the t(1;19) in TS-2 fuses the 19p13 gene DAZAP1 (Deleted in Azoospermia-Associated Protein 1) to the 1q23 gene MEF2D (Myocyte Enhancer Factor 2D), leading to expression of reciprocal in-frame DAZAP1/MEF2D and MEF2D/DAZAP1 transcripts. MEF2D is a member of the MEF2 family of DNA binding proteins that activate transcription of genes involved in control of muscle cell differentiation, and signaling pathways that mediate response to mitogenic signals and survival of neurons and T-lymphocytes. DAZAP1 is a novel RNA binding protein expressed most abundantly in the testis. We demonstrate that MEF2D/DAZAP1 binds avidly and specifically to DNA in a manner indistinguishable from that of native MEF2D and is a substantially more potent transcriptional activator than MEF2D. We also show that DAZAP1/MEF2D is a sequence-specific RNA-binding protein. MEF2D has been identified as a candidate oncogene in murine retroviral insertional mutagenesis studies. Our data implicate MEF2D in human cancer and suggest that MEF2D/DAZAP1 and/or DAZAP1/MEF2D contribute to leukemogenesis by altering signaling pathways normally regulated by wild-type MEF2D and DAZAP1.

Cell Line, Tumor↗

Combination treatment of vitamin C and desferrioxamine suppresses glomerular superoxide and prostaglandin E production in diabetic rats.

AIMS: Increased oxidative stress may contribute to the development of diabetic nephropathy. Conversely, it has been proposed that enhanced glomerular production of prostaglandin E(2) (PGE(2)) may be the cause of glomerular hyperfiltration in streptozotocin (STZ)-induced diabetic rats. As the role of superoxide anion (O(2-)) production in early diabetic nephropathy is not fully understood, we investigated the effect of vitamin C and desferrioxamine treatment on glomerular O(2-) and PGE(2) production in diabetic rats. METHODS: STZ-induced diabetic rats were given drinking water containing 1 g/l of vitamin C and desferrioxamine for 10 days, and glomerular O(2-) production, glomerular PGE(2) synthesis and creatinine clearance were examined. RESULTS: Glomerular O(2-) production increased in untreated diabetic rats compared to non-diabetic controls (142.2 +/- 12.4 vs. 65.4 +/- 3.6 counts/mg protein/min). Treatment with vitamin C and desferrioxamine significantly decreased glomerular O(2-) production (93.7 +/- 6.7 counts/mg protein/min). Glomerular PGE(2) synthesis and creatinine clearance were significantly increased in untreated diabetic rats compared to controls and PGE(2) synthesis was reduced and creatinine clearance tended to decrease by the treatment. CONCLUSIONS: Our results demonstrated that vitamin C and desferrioxamine suppressed the enhanced glomerular O(2-) production with subsequent decrease in PGE(2) production. Antioxidant therapy may be beneficial in preventing the development of diabetic nephropathy.

Animals↗

Effects of multigrooved surfaces on osteoblast-like cells in vitro: scanning electron microscopic observation and mRNA expression of osteopontin and osteocalcin.

This study evaluated the behavior of osteoblast-like cells on multigrooved surfaces consisting of a combination of microgrooves and macrogrooves. A polystyrene substrate was fabricated with multigrooves with 90-degree, V-shaped microgrooves with a 2-microm pitch cut on trapezoidal macrogrooves, which had a 50-microm ridge width, a 50-microm wall width, a 50-microm bottom width, and 25-microm depth. Smooth polystyrene substrates were also prepared as controls. Rat bone marrow cells were cultured as osteoblast-like cells on the substrates for morphological evaluation using a scanning electron microscope, and for biochemical evaluation using the quantitative reverse transcriptase-polymerase chain reaction technique for osteopontin and osteocalcin mRNA expression. After 8 days of incubation, the osteoblast-like cells were aligned parallel to the surface grooves on the multigrooved substrates. After 16 days of incubation, a dense mineralized extracellular matrix (ECM) was produced along the multigrooves. The ECM on the multigrooved surface appeared oriented more in the direction of the grooves than on the smooth surface, and trapezoid-shaped macrogrooves of the ECM were cast upside down. Although there were not significant differences, the osteopontin and osteocalcin mRNA expressions of the osteoblast-like cells on the multigrooved surfaces tended to be higher than on smooth surfaces. These results suggest that multigrooves could be used to control the orientation of mineralized ECM as well as of cells, and also to enhance the production of mineralized ECM.

Animals↗

Calcium phosphate formation on the phosphorylated dental bonding agent in electrolyte solution.

The aim of the present study was to study the mineral formation on a phosphorylated dental bonding agent using a mineralization inductive solution. Clearfil Photobond, which contained phosphate monomer, was cured by photo-irradiation and heat treated, and was then immersed in Hanks' balanced salt solution (HBSS) with pH = 7.4 for 1, 3, 5, 7, 14, and 28 days at 37 degrees C. The white substances were deposited on the phosphorylated polymer, i.e. cured Photobond disk, after the immersion in HBSS. The white substances become visible after 3 days immersion. After 7 days immersion, surface of the phosphorylated polymer disk was almost covered with white substance layers. The measurement of white substances by means of X-ray diffraction, Fourier-transform infrared and electron probe microanalysis revealed that their main component was carbonate-containing hydroxyapatite. Scanning electron microscopy pictures showed that a large number of globules of hydroxyapatite were fused together, and that each globule was composed of a group of numerous thin-film form flakes uniting and/or clustering together. The results obtained in this study concluded that the presence of phosphonic acid and phosphate group of phosphorylated dental bonding agent enhanced the nucleation and growth of hydroxyapatite crystals on its surface.

Biocompatible Materials↗

Effects of multigrooved surfaces on fibroblast behavior.

Microgrooves have been investigated as substrates for the control of cell alignment. However, they are relatively too narrow and shallow for controlling the orientation of extracellular matrices (ECM) such as collagen. Multigrooves, a combination of microgrooves and macrogrooves, are expected to be able to control the orientation of both cells and ECM. This study investigated a method for fabricating multigrooves and evaluated fibroblast behavior on these novel surfaces. Multigrooved patterns were fabricated on a gold-alloy metal die, in which 90-degree V-shaped microgrooves with a 2-microm pitch were cut on trapezoidal macrogrooves. The macrogrooves had a 50- microm ridge width, a 50-microm wall width, a 50-microm bottom width, and a 25-microm depth. The grooves were made by an ultraprecision micromachine using a single crystal diamond. This metal die served as a template for making surface replicas from polystyrene. Microgrooved and smooth polystyrene replicas also were prepared as comparative substrates. Mouse fibroblast L929 cells were cultured in each type of replica substrate for 7 to 21 days. After these periods, the cells were fixed with 2.5% glutaraldehyde, treated with conventional methods, and, finally, observed by SEM. Confocal laser scanning microscopy was performed to investigate ECM formation. The multigrooved metal die exhibited the desired sharp configuration without defects. The dimensional values of the multigrooves on the polystyrene replicas were almost the same as the designed values. The fibroblasts on the multigrooved and microgrooved substrates were aligned parallel to the surface grooves after 7 days of incubation. In contrast to the microgrooved and flat surfaces, a dense extracellular matrix was produced along the multigrooves after 21 days of incubation. These results suggest that multigrooves can control the orientation of ECM as well as cells and thus enhance the production of ECM.

Animals↗

Bone response to calcium phosphate-coated and bisphosphonate-immobilized titanium implants.

Thin calcium phosphate (Ca-P) coatings have been introduced to overcome the shortcomings of plasma-sprayed Ca-P coatings. In our previous experiments, thin Ca-P coatings also enabled the immobilization of bisphosphonate, which is a drug used to treat osteoporosis. The present study was designed to evaluate the bone response to titanium implants treated with a thin Ca-P coating and bisphosphonate. Forty cylindrical commercially pure titanium implants with a length of 7 mm and a diameter of 3 mm were used as test implant fixtures. Three groups of surface-treated implants were prepared: (1) blasted with titanium powder and etched with a solution of 10% HF + 5% HNO3 (control); (2) modified with 0.5-microm thick Ca-P coatings and rapid heat-treating, and (3) immobilized with bisphosphonate by immersion in pamidronate disodium solution (10(-2) M) for 24 h at 37 degrees C. These surface-treated implants were inserted into edentulous areas in the mandibular molar region of five beagle dogs. After implantation periods of 4 and 12 weeks, the bone implant interface was evaluated histologically and histomorphometrically. All measurements were statistically evaluated using a one-way ANOVA and Fisher PLSD test for multiple comparisons among the means. Four weeks after the implantation, higher percentage of bone contact was found around the thin Ca-P-coated implants compared to that of the control group. The highest percentage of bone contact was found around the bisphosphonate-immobilized implants after 12 weeks of implantation. These data suggest that a thin coating of calcium phosphate followed by bisphosphonate-immobilization is effective in the promotion of osteogenesis on surfaces of dental implants.

Animals↗

Sodium-coupled glucose transporter as a functional glucose sensor of retinal microvascular circulation.

To clarify the function of the Na(+)-coupled glucose transporter in the regulation of cellular tone of cultured retinal pericytes, we investigated the effects of extracellular glucose concentration on cell size. The surface area and diameter of cultured bovine retinal pericytes under different glucose concentrations were measured by using a light microscope with a digital camera. We also examined the effects of extracellular Na(+) and Ca(2+), inhibitors of the Na(+)-coupled glucose transporter and Na(+)-Ca(2+) exchanger, a Ca(2+) channel blocker, and nonmetabolizable sugars on cell size. The surface area and diameter of the cells changed according to extracellular glucose concentrations. alpha-Methyl glucoside, which enters the cell through the Na(+)-coupled glucose transporter, induced cellular contraction. However, the cells did not contract in response to 2-deoxyglucose, which enters the cell through a facilitated glucose transporter. Glucose-induced cellular contraction was abolished in the absence of extracellular Na(+) and Ca(2+). Moreover, phlorizin, an inhibitor of the Na(+)-coupled glucose transporter, and 2',4'-dichlorobenzamil-HCl, an inhibitor of the Na(+)-Ca(2+) exchanger, also abolished glucose-induced cellular contraction, whereas nicardipine, a Ca(2+) channel blocker, did not. Our results indicate that high extracellular glucose concentrations induce contraction of bovine retinal pericytes via Na(+) entry through a Na(+)-coupled glucose transporter, suggesting that the Na(+)-coupled glucose transporter may act as a functional glucose sensor of retinal microvascular circulation.>

Amiloride↗

Immobilization of bisphosphonates on surface modified titanium.

The efficiency of surface modifications on the immobilization of bisphosphonates on titanium was investigated with Ca-ion implantation and thin hydroxyapatite coatings. The ALP activity of osteoblastic cells and the inhibitory effects on the initial adherence of P. gingivalis were also evaluated using bisphosphonate-immobilized titanium. X-ray photoelectron spectroscopy analysis suggested that titanium surfaces modified with Ca-ion implantation and thin hydroxyapatite coatings caused the immobilization of bisphosphonate on titanium plates. The ALP activity of osteoblastic cells cultured on plates immobilized with bisphosphonate was almost the same as that of cells cultured on titanium plates, indicating that the bisphosphonate-immobilization showed no toxic effect on osteoblastic cells, and that it provides a favorable micro-environment with osteogenetic ability. Data of the adherence of oral bacteria showed that a bisphosphonate-immobilized titanium surface inhibited the initial adherence of P. gingivalis. These results indicate that the immobilization of bisphosphonates on titanium modified with Ca-ion implantation and thin hydroxyapatite coatings are useful for dental implants.

Animals↗

Influence of surface modifications to titanium on antibacterial activity in vitro.

The antibacterial effect of surface modifications to titanium on Porphyromonas gingivalis ATCC 33277 and Actinobacillus actinomycetemcomitans ATCC 43718 was evaluated. Surface modifications were performed with dry processes including ion implantation (Ca+, N+, F+), oxidation (anode oxidation, titania spraying), ion plating (TiN, alumina), and ion beam mixing (Ag, Sn, Zn, Pt) with Ar+ on polished pure titanium plates. F+-implanted specimens significantly inhibited the growth of both P. gingivalis and A. actinomycetemcomitans than the polished titanium. The other surface-modified specimens did not exhibit effective antibacterial activity against both bacteria. No release of the fluorine ion was detected from F-implanted specimens under dissolution testing. This result and the characterization of the F+-implanted surfaces suggested that the possible antibacterial mechanism of the F+-implanted specimen was caused by the formation of a metal fluoride complex on the surfaces. In addition, F+-implanted surfaces did not inhibit the proliferation of fibroblast L929-cells. These findings indicate that surface modification by means of a dry process is useful in providing antibacterial activity of oral bacteria to titanium implants exposed to the oral cavity.

Aggregatibacter actinomycetemcomitans↗

Substrate affects the initial attachment and subsequent behavior of human osteoblastic cells (Saos-2).

Interaction between implant materials and bone cells contributes to the clinical success of dental implants. The object of this study was to investigate the initial attachment and subsequent behavior of human osteoblastic cells (Saos-2) to pure titanium (Ti), hydroxyapatite (HA), and glass. We, therefore, performed a time-course study for examining the area, attachment rate, distribution of focal adhesion kinase (FAK) vinculin, and actin, and the motility of Saos-2 cells on the materials. On Ti, cell area increased gradually, whereas on HA, cells spread quickly, but quitted spreading at 12 h after cell seeding. The number of cells on HA was greater than on the other materials. On Ti, the numbers of FAK- and vinculin-positive focal adhesions increased continuously. On HA, although the number of FAK-positive focal adhesions also increased continuously, the number of vinculin-positive focal adhesions decreased. Furthermore, actin staining showed that the cells on HA poorly formed stress fibers with weak polarity, whereas the cell on Ti possessed well-defined polarized stress fibers. On HA, cells started extension earlier than on Ti, motility was inactive, and the cells settled on the materials. These results suggest that the earlier settling of osteoblasts on HA might result in earlier osteogenesis on HA than other materials.

Actins↗

Pancreatic islet blood flow in conscious rats during hyperglycemia and hypoglycemia.

Anesthesia affects general hemodynamics and regulation of organ perfusion. We used colored microspheres to measure pancreatic islet blood flow in conscious rats at two time points, during either hyperglycemia or hypoglycemia. This method, using black and green microspheres, was validated by comparison with previous microsphere experiments and by lack of effect of a nonmetabolizable glucose analog, 3-O-methylglucose, on islet perfusion. Basal and glucose-stimulated islet blood flow levels were similar in pentobarbital sodium-anesthetized and conscious rats. However, the basal distribution of pancreatic blood flow was altered by anesthesia (fractional islet blood flow 5.8 +/- 0.4% in conscious rats, 7.9 +/- 0.8% in pentobarbital-anesthetized rats, P < 0.05). Insulin-induced hypoglycemia significantly increased whole pancreatic blood flow in conscious rats, whereas islet blood flow remained unchanged and fractional islet blood flow was decreased (5.8 +/- 0.5% in the basal state, 4.2 +/- 0.4% during hypoglycemia, P < 0.001). Methylatropine pretreatment significantly increased islet blood flow during hypoglycemia by 181%. This result suggests that prevention of hypoglycemia-induced increase in islet perfusion may be mediated, at least in part, by a cholinergic, vagal muscarinic mechanism.

3-O-Methylglucose↗

Monitoring of minimal residual disease in children with acute promyelocytic leukemia by RT-PCR detecting PML/RARalpha chimeric gene: a retrospective study of clinical feasibility.

We studied retrospectively the clinical feasibility of minimal residual disease (MRD) monitoring by reverse transcription-polymerase chain reaction (RT-PCR) detecting the PML/retinoic acid receptor alpha (RARalpha) chimeric gene in children with acute promyelocytic leukemia (APL). MRD monitoring of APL was performed with standard and nested RT-PCR for PML/RARalpha gene, the sensitivity of which was 1 leukemic cell in 10(3)-10(4) and 1 in 10(4)-10(5) cells, respectively. Patients were nine children with APL (average age: 8.3 year; average period of follow-up: 69.2 months) who, after achieving remission with all-trans retinoic acid (ATRA), received treatment either with multidrug chemotherapy or with a combination of chemotherapy and ATRA. Out of six patients treated with multidrug-combined chemotherapy, two patients exhibited PCR positivity after six months of post- remission therapy, which shifted from the detectable range of the nested PCR to that of the standard PCR. These two patients subsequently relapsed and, together with two of the other patients receiving multidrug-combined chemotherapy, underwent allogeneic bone marrow transplantation. No MRD was detected in these patients after transplantation. In the remaining three patients who underwent cyclic treatment with alternative chemotherapy and ATRA, two showed positive RT-PCR at the nested or standard level, respectively, after six months of combined therapy, and one of them relapsed. Overall, three of four patients with MRD detected in post-remission period ultimately relapsed, while all of five patients without detectable MRD had a good prognosis. These findings suggest that impending relapse may be predicted by the detection of preceding PCR positivity with an increasing quantity of the PML/RARalpha mRNA that appears beyond six months of post-remission chemotherapy, with or without combined ATRA therapy.

Adolescent↗

Application of sintered titanium alloys to metal denture bases: a study of titanium powder sheets for complete denture base.

The purpose of this study was the fabrication of titanium powder sheets to enable the application of sintered titanium alloys as metal denture bases. The effects of titanium particle shape and size, binder content, and plasticizer content on the surface smoothness, tensile strength and elongation of titanium powder sheets was investigated. To select a suitable ratio of powdered metal contents for application as a metal denture base, the effects of aluminum content in Ti sheets and various other powder metal contents in Ti-Al sheets on the density, sintering shrinkage, and bending strength were evaluated. Based on the results of the above experiments, we developed a mixed powder sheet composed of 83Ti-7Al-10Cr with TA45 titanium powder (atomized, -45 microm), and 8 mass% binder content. This titanium alloy sheet had good formability and ductility. Its sintered titanium alloy had a density of 3.2 g/cm3, sintering shrinkage of 3.8%, and bending strength of 403 MPa. The titanium alloy sheet is clinically acceptable for fabricating denture bases.

Aluminum↗

Orthostatic hypertension in patients with type 2 diabetes.

OBJECTIVE: The prevalence and clinical importance of orthostatic hypertension (OHT) in diabetic patients has not been elucidated, in contrast to orthostatic hypotension, which is occasionally found in diabetic patients with autonomic neuropathy. RESEARCH DESIGN AND METHODS: The prevalence and severity of orthostatic hypertension was investigated in 277 Japanese male patients with type 2 diabetes, including 90 hypertensive patients and 128 nondiabetic age-matched male subjects. Patients treated with antihypertensive drugs were excluded from the study. OHT was defined as an increase in diastolic blood pressure (DBP) from <90 to >or=90 mmHg and/or an increase in systolic blood pressure (SBP) from <140 to >or=140 mmHg after standing from supine position. Clinical profiles and several serum biochemical parameters were determined in addition to chest X-rays and electrocardiograms. RESULTS: The prevalence of OHT in normotensive and hypertensive diabetic patients was significantly higher than in control subjects (12.8 vs. 1.8%, P < 0.01, for normotensive patients; 12.6 vs. 11.1%, not significant, for hypertensive patients). Orthostasis induced a mean increase of 6.8 +/- 11.4 mmHg in SBP and 9.1 +/- 5.2 mmHg in DBP in diabetic patients with OHT compared with those without OHT (-1.0 +/- 9.0 and 3.8 +/- 6.6 mmHg, respectively). Vibration sensation in the lower limb was reduced in diabetic patients with OHT, but the percent coefficient of variation of RR interval, cardio-to-thoracic ratio on chest X-ray, and serum triglyceride levels were higher in these patients compared with normotensive diabetic patients without OHT. CONCLUSIONS: Orthostatic hypertension is a novel complication in normotensive diabetic patients and may associate with early stage neuropathy and development of sustained hypertension.

Diabetes Mellitus, Type 2↗