Polyelectrolyte multilayer-calcium phosphate composite coatings for metal implants.

The preparation of organic-inorganic composite coatings with the purpose to increase the bioactivity of bioinert metal implants was investigated. As substrates, glass plates and rough titanium surfaces (Ti-SLA) were employed. The method comprises the deposition of polyelectrolyte multilayers (PEMLs) followed by immersion of the coated substrate into a calcifying solution of low supersaturation (MCS). Single or mixed PEMLs were constructed from poly-L-lysine (PLL) alternating with poly-L-glutamate, (PGA), poly-L-aspartate (PAA), and/or chondroitin sulfate (CS). ATR-FTIR spectra reveal that (PLL/PGA)10 multilayers and mixed multilayers with a (PLL/PGA)5 base contain intermolecular ß-sheet structures, which are absent in pure (PLL/PAA)10 and (PLL/CS)10 assemblies. All PEML coatings had a grainy topography with aggregate sizes and size distributions increasing in the order: (PLL/PGA)n < (PLL/PAA)n < (PLL/CS)n. In mixed multilayers with a (PLL/PGA)n base and a (PLL/PAA)n or (PLL/CS)n top, the aggregate sizes were greatly reduced. The PEMLs promoted calcium phosphate nucleation and early crystal growth, the intensity of the effect depending on the composition of the terminal layer(s) of the polymer. In contrast, crystal morphology and structure depended on the supersaturation, pH, and ionic strength of the MCS, rather than on the composition of the organic matrix. Crystals grown on both uncoated and coated substrates were mostly platelets of calcium deficient carbonate apatite, with the Ca/P ratio depending on the precipitation conditions.

Preparation of a partially calcified gelatin membrane as a model for a soft-to-hard tissue interface.

Cartilage and/or bone tissue engineering is a very challenging area in modern medicine. Since cartilage is an avascular tissue with limited capacity for self-repair, using scaffolds provides a promising option for the repair of severe cartilage damage caused by trauma, age-related degeneration, and/or diseases. Our aim in this study was to design a model for a functional biomedical membrane to form the interface between a cartilage-forming scaffold and bone. To realize such a membrane gelatin gels containing calcium or phosphate ions were exposed from one side to a solution of the other constituent ion (i.e., a sodium phosphate solution was allowed to diffuse into a calcium-containing gel and vice versa). The partially calcified gels were analyzed by XRD, ATR-FTIR spectra, E-SEM, and EDX. Thus, we confirmed the existence of a gradient of crystals, with a dense top layer, extending several micrometers into the gel. XRD spectra and Ca/P atomic ratios confirmed the existence of calcium deficient apatites. The effect of different experimental parameters on the calcification process within the gelatin membranes has been elucidated. It was shown that increasing the gelatin concentration from 5 wt % to 10 wt % retards calcification. A similar effect was observed when glycerol, which is frequently used as plasticizer, was added to the system. With increasing calcium concentration within the organic matrix, the quantity and density of calcium phosphate crystals over/within the gel increased. The possible explanations for the above phenomena are discussed.

Evaluation of methods for urine inhibitory potential for precipitation of calcium oxalate.

Renal lithiasis is a significant medical and social problem. Worldwide recurrence is anywhere from 3% to 5%. Objective of this paper is to evaluate two methods for distinguishing between stone formers and non-stone formers. Urine samples were titrated with calcium and seed crystals were added to facilitate precipitation. Ionic calcium levels were monitored and compared between the two groups. Stone formers showed impaired tolerance to the calcium added and increased precipitation on seed crystals. Both methods discriminated between stone formers and non-stone formers. Further evaluations are needed to establish the better of the two for wider clinical use.

Complementary effects of multi-protein components on biomineralization in vitro.

The extracellular matrix (ECM) is composed of mixed protein fibers whose precise composition affects biomineralization. New methods are needed to probe the interactions of these proteins with calcium phosphate mineral and with each other. Here we follow calcium phosphate mineralization on protein fibers self-assembled in vitro from solutions of fibronectin, elastin and their mixture. We probe the surface morphology and mechanical properties of the protein fibers during the early stages. The development of mineral crystals on the protein matrices is also investigated. In physiological mineralization solution, the elastic modulus of the fibers in the fibronectin-elastin mixture increases to a greater extent than that of the fibers from either pure protein. In the presence of fibronectin, longer exposure in the mineral solution leads to the formation of amorphous calcium phosphate particles templated along the self-assembled fibers, while elastin fibers only collect calcium without any mineral observed during early stage. TEM images confirm that small needle-shape crystals are confined inside elastin fibers which suppress the release of mineral outside the fibers during late stage, while hydroxyapatite crystals form when fibronectin is present. These results demonstrate complementary actions of the two ECM proteins fibronectin and elastin to collect cations and template mineral, respectively.

Biomimetic organic-inorganic nanocomposite coatings for titanium implants.

A new class of organic-inorganic nanocomposites, to be used as coatings for surface enhancement of metal implants for bone replacement and repair, has been prepared by a biomimetic three-step procedure: (1) embedding amorphous calcium phosphate (ACP) particles between organic polyelectrolyte multilayers (PE MLs), (2) in situ transformation of ACP to octacalcium phospate (OCP) and/or poorly crystalline apatite nanocrystals by immersion of the material into a metastable calcifying solution (MCS) and (3) deposition of a final PE ML. The organic polyelectrolytes used were poly-L-glutamic acid and poly-L-lysine. The nanocomposites obtained by each successive step were characterized by scanning electron microscopy, energy dispersive X-ray analysis (EDS), and XRD, and their suitability as coatings for metal implants was examined by mechanical and in vitro biological tests. Coatings obtained by the first deposition step are mechanically unstable and therefore not suitable. During the second step, upon immersion into MCS, ACP particles were transformed into crystalline calcium phosphate, with large platelike OCP crystals as the top layer. After phase transformation, the nanocomposite was strongly attached to the titanium, but the top layer did not promote cell proliferation. However, when the coating was topped with an additional PE ML (step 3), smoother surfaces were obtained, which facilitated cell adhesion and proliferation as shown by in vitro biological tests using primary human osteoblasts (HO) directly seeded onto the nanocomposites. In fact, cell proliferation on nanocomposites with top PE MLs was far superior than on any of the individual components and was equivalent to proliferation on the golden standard (plastic).

Metabolic evaluation of urolithiasis patients from eastern Croatia.

Metabolic parameters were determined in fasting blood serum, fasting first morning urine, and 24-hour urine of male patients with recurrent calcium oxalate stones (N = 26, age 39.1 +/- 6.2 years) as well as in male healthy controls (N = 18, age 35.0 +/- 7.1 years), recruited from the eastern part of Croatia. The 24-hour urinary calcium excretion was significantly higher (p < 0.01) for patients (5.6 +/- 2.5 mmol) than for controls (3.7 +/- 1.9 mmol), but potassium excretion was higher (p < 0.01) for controls (74.5 +/- 33.8 mmol) than for patients (49.2 +/- 15.7 mmol). The mean ionic activity product of calcium and oxalate ions, IAP(CaOx), calculated from the fasting first morning urine parameters, was 25% higher for patients than for controls, but the difference was not statistically significant (p > 0.05). Very strong correlation (r = 0.97) was obtained between IAP(CaOx) values and calculated Ogawa indices that were recommended for estimating the potential risk for calcium oxalate stone formation.

Bone mineral density loss in patients with urolithiasis: a follow-up study.

BACKGROUND: Recurrent calcium urolithiasis is often associated with disorders of calcium metabolism. The purpose of this investigation was to assess bone mineral content (BMC) and bone mineral density (BMD) over a period of 1 year in patients with urolithiasis and to determine the factors that could have influenced the changes in bone density during that period. METHODS: The patient group comprised 34 men aged 41.2 plus minus 7.9 years with recurrent urolithiasis. A wide spectrum of biochemical measurements was performed. Bone mineral density (g/cm(2)), bone mineral content (BMC), and bone area (BA) were measured twice during a period of 1 year at the lumbar spine (L2-L4), femoral neck, Ward triangle, and trochanter, using dual energy absorptiometry. Patient results were compared to those obtained from 30 healthy male controls of a comparable age group. RESULTS: Nine patients were hypercalciuric, while the majority of the remaining metabolic parameters were within the reference values. Bone mineral content and bone areas at all regions were lower in patients comparing to controls, but not significantly. The greatest annual reduction of BMD was noticed at Ward triangle (-5.70% in patients and -2.36% in controls), followed by femoral neck (-4.06% patients, -2.03% controls) and trochanter (-3.06% patients, -1.39% controls). There was no significant decrease of the BMD of the spine. Analyzing the influence of age, body mass index (BMI), metabolic parameters, and dietary calcium intake on the annual reduction of bone density, we found that age, hyperuricosuria, and calcium intake were significantly associated with bone loss in that time period. CONCLUSIONS: Bone mass reduction in patients with urolithiasis over a 1-year period did not differ significantly from that in controls and was principally related to age, hyperuricosuria, and calcium dietary restriction but not to increased calcium excretion.