sepharose and Calcinosis

Calcific aortic valve disease (CAVD) is the most common valvular disorder. While fluid stresses are presumed to play a role in disease progression, the valvular hemodynamic changes experienced over the course of CAVD remain largely unknown. The study aim was to develop a laboratory protocol for the fabrication of tissue valve models mimicking mild and moderate calcific stenosis, for future use in flow studies.. Different hydroxyapatite (HA)-agarose mixtures were injected into porcine valve leaflets. Micro-computed tomography (micro-CT) was used to quantify HA deposition volume, area fraction and regional distribution, while von Kossa staining was performed to assess tissue mineralization. Particle image velocimetry measurements were carried out in intact and injected valves subjected to in vivo-like hemodynamics to characterize the degree of valvular stenosis in terms of geometric orifice area (GOA) and peak systolic velocity.. The 5% HA-1% agarose solution (solution 1) and the 5% HA-0.5% agarose solution (solution 2) maximized the HA deposition volume. Leaflet injections with solution 1 resulted in a significant 1.9-fold increase in HA area fraction relative to solution 2 injections. While solution 1 injections generated multiple sites of high HA concentration, solution 2 injections produced smaller, discrete spots. Injections of both solution 1 and solution 2 into whole valves generated significant 47% and 32% reductions, respectively, in GOA and 1.8-fold and 1.5-fold increases, respectively, in peak systolic velocity, relative to untreated valves.. Tissue valve models were generated that recapitulated the structure and hemodynamics of mild and moderate valvular calcification. Those models may be used for future investigations of the native valvular hemodynamic alterations that occur during CAVD.

Topics: Animals; Aortic Valve; Aortic Valve Stenosis; Blood Flow Velocity; Calcinosis; Disease Models, Animal; Durapatite; Hemodynamics; Regional Blood Flow; Sepharose; Severity of Illness Index; Swine; X-Ray Microtomography

Mammography is a widely used tool for the screening of breast cancer, and calcifications are a common finding in most mammograms. The location, size, number, morphology, and distribution of calcifications are an important information to differentiate a benign lesion from probably malignant pathologies. Calcifications are not detectable with a standard dynamic contrast enhanced breast MRI. The authors present a novel method for the detection and imaging of calcifications in breast tissue without ionizing radiation or contrast agents.. Measurements of localized tissue displacement in phantoms due to applied acoustic radiation force were performed. This displacement was imaged with a displacement sensitive spin-echo MRI sequence. Pieces of eggshell that represent calcifications were embedded in tissue-mimicking agarose phantoms. The sizes of the calcifications were 0.8 x 0.8 x 0.4, 1.5 x 1.5 x 0.4, and 2 x 3 x 0.4 mm3. The calcifications were scanned with ultrasound (U.S.) at 2.5 MHz and intensities up to I(spta) =7.18 W/cm2. The U.S. beam was moved inside the phantom by a computer-controlled three-dimensional hydraulic positioning system. The U.S. beam was scanned over the two smaller calcifications with the displacement sensitivity of the MRI sequence parallel to the U.S. beam path. Grayscale coded maps of the displacement scans are presented. For the 0.8 x 0.8 x 0.4 mm3 calcification, the U.S. intensities were varied. Finite element simulations were performed to verify if the experiments complied with theory.. The authors found that the displacement caused by the U.S. is increased at the position of the calcification. The area of increased displacement is at least twice as large as the calcification itself. The simulations show this increase in displacement and area at the position of the calcification. When changing the displacement sensitivity direction to perpendicular to the U.S. beam, a crossed black and white four-leaf clover is visible at the position of the calcification.. The U.S. is scattered and reflected by the calcifications. This leads to the increased displacement which is transmitted to the surrounding material because of the elastic coupling between the calcification and the agarose material. Due to the high differences in acoustic impedance and elastic properties between the surrounding tissue and the calcification, even the detection of pieces smaller than the resolution of the MRI scanner is possible. The acoustic radiation force contrast in MR phase-difference images offers a positive signal for calcifications from a smooth background in phantoms. This method offers a possibility of differentiating qualitatively and quantitatively hard calcifications from stiffer inclusions such as tumors.

Topics: Acoustics; Calcinosis; Finite Element Analysis; Hot Temperature; Magnetic Resonance Imaging; Phantoms, Imaging; Sepharose

Using the biomimetic method, we formed a hydroxyapatite (HAp) layer on/in certain types of nonionic hydrogels that contain hydroxyl groups. The hydrogels used were poly(vinyl alcohol) (PVA), poly(2-hydroxyethyl methacrylate) (PHEMA), poly(glucosyloxyethyl methacrylate) (PGEMA), and agarose. Under an optical microscope, we observed a thin, continuous HAp layer on the top surface of the PVA, PHEMA, and PGEMA gels. On the other hand, we only observed an intermittent HAp layer on the surface of the agarose gel. The swelling ratio and the bound water content of these hydrogels were measured as an essential character in HAp formation. There was some relation among the HAp formation, the swelling ratios, and the bound water content.

Topics: Calcinosis; Calorimetry, Differential Scanning; Durapatite; Glucosides; Hydrogels; Polyhydroxyethyl Methacrylate; Polymers; Polyvinyl Alcohol; Sepharose; Surface Properties; X-Ray Diffraction

Ros 17/2 clonal rat osteosarcoma cells calcify when cultured in the presence of 10 micrograms/ml beta-glycerol phosphate in an agarose gel. Culture in 1% agarose inhibited cell division while allowing cells to remain metabolically active and viable for over 21 days. Serial photography of the same microscopic field shows a progressive deposition of calcium phosphate during the course of the experiment. The deposition of calcium around cells was confirmed by calcium-specific stains, and by energy dispersive X-ray analysis (EDX) during scanning electron microscopy. Cells with high calcium content analyzed by EDX had Ca:P ratios similar to hydroxyapatite. Total calcium progressively increased in beta-glycerol phosphate-treated cultures whereas the control plates maintained a constant calcium content over 16 days. Alkaline phosphatase activity increased with time in culture whereas cells with beta-glycerol phosphate maintained the alkaline phosphatase values achieved at the time of initial calcification. Alkaline phosphatase staining revealed no correlation between the presence of the enzyme activity and calcification. Radioimmunoassay for the bone-specific vitamin K-dependent protein bone Gla protein showed that beta-glycerol phosphate-treated cells accumulate over sixfold greater amounts of this protein. Our studies show that ROS cells can calcify and accumulate bone-specific matrix components when cultured in a 3-dimensional agarose matrix.

Topics: Alkaline Phosphatase; Animals; Calcinosis; Cell Line; DNA, Neoplasm; Microscopy, Electron, Scanning; Osteoblasts; Osteosarcoma; Rats; Sepharose