Silicon-doped calcium phosphates; the critical effect of synthesis routes on the biological performance.

In this study, the effect of using different types of fuel and various amounts of Si4+ ions on the biological properties of silicon-doped calcium phosphates (CaPs), which were synthesized using solution combustion method were investigated. X-ray diffraction (XRD) patterns showed that hydroxyapatite/beta-tricalcium phosphate (HA/ßTCP) was crystallized in all synthesized samples. The synthesized sample using glycine as fuel, which doped with 0.1 mol Si4+ ions exhibited the most desirable properties. Consecutively, the zeta potential and specific surface area were enhanced from -20 to -27 mV and 38 to 146 m2/g, respectively, by increasing the amount of Si4+ ions from 0 to 0.1 mol. The bioactivity of the samples immersed in simulated body fluid (SBF) was innovatively determined by the joint analyses of the tensiometer, inductively coupled plasma (ICP), field emission scanning electron microscopy (FESEM), and XRD data. These findings plus theoretical calculations demonstrate, for the first time, that the Si4+ doping could improve the bioactivity of the powders up to ~155%. The results of in vitro cell-based experiments, including cell viability, alizarin red staining, and cell attachment, confirmed the positive effects of Si-doped powders in the biological systems. Furthermore, Si-doped powders were able to improve the migration ability of mammalian cells in vitro; they could be considered good candidates in angiogenesis-based therapeutic strategies.

Effect of supervision on ultrasonographic measurements. A blinded randomized cross-over study.

BACKGROUND: Musculoskeletal ultrasound is becoming an increasingly popular clinical tool in the hands of physiatrists. Herewith, although the role of direct supervision (by an expert) is universally recognized as the core element for appropriate ultrasound training, to our best notice, its impact on ultrasonographic measurements has not been studied quantitatively in the hitherto literature. AIM: To quantify the effect of supervision in the early period of musculoskeletal ultrasound training by using three different tissues (muscle-cartilage-tendon) as models. DESIGN: A blinded randomized cross-over study. SETTING: Physical Medicine and Rehabilitation Department of a University Hospital. POPULATION: A total of 9 sonographers (8 novice and 1 expert) were involved whereby the novice sonographers were randomly divided into two groups; Group A (N.=4) and Group B (N.=4). METHODS: All sonographers performed three thickness measurements; medial head of the gastrocnemius muscle, patellar tendon and femoral cartilage on the left lower limb of the same subject. The expert supervised Group A in the first half of the study (9 days), and Group B in the second half (9 days). Throughout the study period, all the participants were blinded to the data. Relative effectiveness, expert effect, order effect, treatment-period effect were studied for cross-over variance analysis. RESULTS: For all the three sites, measurements under the supervision of the expert were significantly different than those without him -p values pertaining to relative effectiveness, expert effect were 0.014, 0.013 for femoral cartilage; <0.001, <0.001 for gastrocnemius and <0.001, <0.001 for patellar tendon, respectively). For gastrocnemius muscle measurements, studying with the expert in the second half of the study (vs in the first half) was better concerning the precision of the measurements (order effect P<0.001). CONCLUSION: Supervision during measurements of novice sonographers is crucial and their data should otherwise be interpreted attentively.

Multivariate optimization on flow-injection electrochemical hydride generation atomic absorption spectrometry of cadmium.

A novel electrochemical hydride generation (ECHG) system working in flow-injection (FI) mode was developed for determination of cadmium coupled to an electrically heated quartz tube atomizer (QTA) by atomic absorption spectrometry (AAS). A Plackett-Burman experimental design for screening has been used to evaluate the influence of several variables on the analytical response. Then, the significant parameters such as the concentration of NaCl in catholyte, applied electrolytic current and flow rate of carrier gas have been simultaneously optimized using a central composite design (CCD). Under the optimized conditions, the detection limit (3σ(b), n=9) was found to be 0.51 ng mL(-1) Cd and the relative standard deviation (RSD) for nine replicate analyses of 20 ng mL(-1) Cd was 6.5%. The calibration curve was linear in the range of 2-50 ng mL(-1) of Cd. The potential interferences from various ions were also evaluated. The analysis of a reference material showed good agreement with the certified value. The proposed method was successfully applied to the determination of Cd in tap water sample.

Mechanistic aspects of electrochemical hydride generation for cadmium.

The mechanistic aspects of a novel methodology for the electrochemical hydride generation (EcHG) of cadmium, entitled as the catholyte variation, have been studied. The hydrogen overvoltage of different cathode materials was determined in dilute electrolyte. The lead-tin alloy used as the cathode material for the EcHG of Cd had the highest hydrogen overpotential. In this technique, both electrolyte medium and cathode surface would change during electrolysis. The catholyte variation EcHG coupled with a flame heated quartz tube atomizer (QTA) atomic absorption spectrometer (AAS) produced peak-shaped transient signal of Cd. The analytical signal was significantly enhanced at neutralization time. The simultaneous production of lead and tin hydrides as well as the hydrogen at that time may speed up the desorption of cadmium species from the surface of the cathode. Also the high hydrogen overvoltage and the neutral medium aided the formation and separation of cadmium hydride molecules. The generated CdH2 vapour was kinetically stable.

Electrochemical hydride generation atomic absorption spectrometry for determination of cadmium.

An electrolytic hydride generation system for determination of another hydride forming element, cadmium, by catholyte variation electrochemical hydride generation (EcHG) atomic absorption spectrometry is described. A laboratory-made electrolytic cell with lead-tin alloy as cathode material is designed as electrolytic generator of molecular hydride. The influences of several parameters on the analytical signal have been evaluated using a Plackett-Burman experimental design. The significant parameters such as cathode surface area, electrolytic current, carrier gas flow rate and catholyte concentration have been optimized using univariate method. The analytical figures of merit of procedure developed were determined. The calibration curve was linear up to 20ngml-1of cadmium. The concentration detection limit (3σ, n=8) of 0.2ngml-1 and repeatability (relative standard deviation, n=7) of 3.1% were achieved at 10.0ngml-1. It was shown that interferences from major constituents at high concentrations were significant. The accuracy of method was verified using a real sample (spiked tap water) by standard addition calibration technique. Recovery of 104% was achieved for Cd in the spiked tap water sample.

Clastogenic factors as biomarkers of oxidative stress in chronic hepatitis C.

BACKGROUND/AIMS: Clastogenic factors (CFs) are composed of lipid peroxidation products, cytokines and other oxidants with chromosome-damaging properties. They are regularly observed after radiation exposure and in chronic inflammatory diseases, where they are supposed to be risk factors for carcinogenesis. It appeared of interest to investigate their presence in the plasma of patients with chronic hepatitis C. METHODS: CFs are detected by chromosomal breakage studies. They were compared to malondialdehyde (MDA), total plasma thiols (t-SH), alanine aminotransferase (ALT), viral load and histological data. RESULTS: CFs were increased in 19 of 20 patients, 16 had increased MDA levels and 15 had decreased t-SH levels. Mean values were significantly different from the 20 controls (p<0.001). After the first 3 months of interferon treatment, all three markers showed significant improvement, but were not completely normalized. There was a positive correlation between CFs and necroinflammatory activity (p<0.03), while MDA was correlated with fibrosis (p<0.03). Viral load was correlated with necrosis and inflammation (p<0.05). CONCLUSION: The presence of CFs in chronic hepatitis C confirms the occurrence of oxidative stress in this disease and could be useful in clinical trials for testing antioxidants. The CF test is a sensitive assay for the detection of oxidative stress and correlates with necroinflammatory activity.

Anticlastogenic effects of Ginkgo biloba extract (EGb 761) and some of its constituents in irradiated rats.

In a previous study we reported that radiation-induced clastogenic factors (CF) are found in the plasma of Chernobyl accident recovery workers and that their chromosome damaging effects are inhibited by antioxidant treatment with a Ginkgo biloba extract (EGb761). In the present study, we induced CF in rats with a radiation dose of 4.5 Gy. The protective effects of the complete extract were compared to those obtained with the extract devoid of its terpene fraction (CP205), with isolated ginkgolides A+B and bilobalide at the concentrations present in EGb761. The pretreatment samples were taken at day 22 postirradiation, the posttreatment samples the day following arrest of the 3-week treatment. The adjusted clastogenic score (ACS) were reduced from 11.71+/-3.55 to 2.00+/-2.83 after treatment with 100 mg/kg and from 13.43+/-2.23 to 4.29+/-2.14 with 50 mg/kg of the complete extract (p<0.0001). Similar protective effects were observed with CP205, ginkgolides and bilobalide (p<0. 001), while the reduction of ACS in placebo-treated rats was not statistically significant (12.80+/-1.79 and 9.20+/-2.68). However, if the efficacy of the treatment was compared to placebo, only the complete extract was significantly protective. While all components exerted anticlastogenic effects at the concentrations present in the complete extract, the comparison of the different groups by analysis of variance did not reveal significant differences. This may be due to to the small number of animals available in each treatment group. The complete extract reduced the ACS by 83% at the dose of 100 mg/kg, while the lower dose of 50 mg/kg and the three components reached only 66%-68% reduction. The better protection provided by the complete extract is due to synergistic rather than to additive effects.

Multiparameter analysis of clastogenic factors, pro-oxidant cytokines, and inflammatory markers in HIV-1-infected patients with asymptomatic disease, opportunistic infections, and malignancies.

HIV-1-infected patients are in chronic oxidative stress and clastogenic factors (CFs) are present in their plasma. CFs from patients with HIV are formed via superoxide anion radical and stimulate further superoxide production. The pathophysiolgic significance and the exact composition of the circulating clastogenic material in patients with HIV is unknown. Cytokines, such as tumor necrosis factor-alpha (TNF-alpha), are increased in the plasma of patients with HIV and TNF-alpha shows clastogenic activity in vitro. The aim of this clinical study was to compare levels of CF in HIV-1-positive patients with asymptomatic disease, opportunistic infections, and malignancies with those in HIV-1-negative control groups and to correlate CF activity with CD4+ T cell numbers, the cytokines (TNF-alpha, interleukin-2 [IL-2], IL-6), and the inflammatory markers (C-reactive protein [CRP], neopterin, granulocyte elastase). CFs were significantly increased in all HIV-1-positive patients and in HIV-1-negative patients with malignant tumors. HIV-1-positive patients with Kaposi's sarcoma showed the highest CF activity in their plasma (p < 0.08). CFs appear very early in HIV infection, and they correlate negatively with CD4+ T cells, which are an indicator of disease activity. The presence of CF in the plasma of HIV-infected patients is not a general response to a viral infection because these factors are not increased in HIV-1-negative patients with viral infection (zoster). CFs are not specific for the HIV-1 infection; they also occur in HIV-1-negative patients with malignant tumors. There was a tendency towards a positive correlation (p < 0.14) between CF and TNF-alpha but there was no positive correlation of CF with IL-2, IL-6, CRP, elastase, and neopterin levels. This indicates that TNF-alpha may be among the components of CF in HIV-1-infected patients. In addition, other unidentified components may contribute to the clastogenic activity of the plasma or the composition of CF may vary from patient to patient. Further clinical studies with larger sample populations are necessary to analyze the composition of CF in HIV-1-positive patients.

Oxyradical-mediated clastogenic plasma factors in psoriasis: increase in clastogenic activity after PUVA.

Psoriasis is a common skin disorder characterized by hyperproliferation and incomplete differentiation of epidermal keratinocytes. Psoralen plus UVA (PUVA) is one of the treatments proposed for this disease. We had reported previously that exposure of regular blood cultures from healthy donors to PUVA leads to chromosomal breakage via the formation of transferable clastogenic materials, a phenomenon inhibitable by superoxide dismutase. In the present paper we show that these clastogenic factors (CF) are also formed in vivo. The CF were found in about 50% of the psoriasis patients studied (14 out of 31). In PUVA-treated psoriasis patients, the clastogenic activity of the plasma increased significantly between the first and the last (16th) exposure to PUVA. We hypothesize that CF formation in psoriasis is similar to that in other diseases accompanied by oxidative stress, in particular chronic inflammatory diseases with autoimmune reactions such as lupus erythematosus, progressive systemic sclerosis, rheumatoid arthritis and others. Increased superoxide production by phagocytes, formation of lipid peroxidation products and release of cytokines are considered to be responsible for the superoxide-stimulating and chromosome-damaging properties of patients' plasma. During PUVA therapy, superoxide generated via the interaction of psoralen with UVA may contribute to CF formation in addition to superoxide from inflammatory cells. An increased risk of cancer and leukemia is observed in diseases accompanied by CF formation. Therefore CF may contribute to the well-known risk of photocarcinogenesis by PUVA therapy. This additional risk may be preventable by antioxidants and superoxide scavengers.

Clastogenic factors in the plasma of children exposed at Chernobyl.

Clastogenic factors (CFs), as they were described previously in accidentally or therapeutically irradiated persons, in A-bomb survivors and in liquidators of the Chernobyl nuclear power plant, were also detected in the plasma of Chernobyl-exposed children. A high percentage of plasma ultrafiltrates from 170 children, immigrated to Israel in 1990, exerted clastogenic effects in test cultures set up with blood from healthy donors. The differences were highly significant in comparison to children immigrated from 'clean' cities of the former Soviet Union or children born in Israel. The percentage of CF-positive children and the mean values of the adjusted clastogenic scores (ACS) were higher for those coming from Gomel and Mozyr, which are high exposure sites (IAEA measurements), compared to those coming from Kiev. There was no correlation between residual 137-Caesium body burden and presence of CFs. However, both measurements were not done at the same time (in 1990 and 1992-1994, respectively). Also no relationship could be revealed between enlargement of the thyroid gland and CF-positivity. CFs are not only observed after irradiation, but in a variety of chronic inflammatory diseases with autoimmune reactions. They were also described in the congenital breakage syndromes, which are hereditary diseases with the highest cancer incidence in humans. Whether the clastogenic effects continuously produced by circulating CFs represent a risk factor for malignant late effects deserves further study and follow-up. Since CF formation and CF action are mediated by superoxide radicals, prophylactic treatment with antioxidants may be suggested for Chernobyl-exposed children, whose plasma induces a strongly positive CF-test.

Chromosome damage in PUVA-treated human lymphocytes is related to active oxygen species and clastogenic factors.

Besides the direct interaction of psoralens with DNA and other macromolecules, the role of reactive oxygen species in the PUVA-induced cellular injury has been stressed. The present study shows that treatment of human blood cultures with 5-methoxypsoralen or 8-methoxypsoralen, followed by UVA exposure, results in chromosome damage. The supernatant of these cultures contains secondarily formed chromosome damaging material, called clastogenic factor (CFs). Not only CF formation, but also CF action is inhibited by superoxide dismutase (SOD), suggesting that superoxide is formed on the pathway to chromosome aberration. CF is detectable in the cell culture supernatants after a minimal delay of 18 h, and reaches a plateau at 24 h of cultivation. SOD is no longer protective if added after 24 h, i.e., the enzyme can prevent, but not repair the oxyradical-induced damage.

Non-specific incision of DNA due to the presence of 8-methoxypsoralen photoinduced interstrand cross-links in Saccharomyces cerevisiae.

The repair of DNA interstrand cross-links (CL) induced by 8-methoxypsoralen (8-MOP) plus UVA irradiation was analyzed by the alkaline step elution technique. A double-exposure protocol was used with 8-MOP, starting with exposure to monochromatic 405-nm radiation inducing only DNA monoadducts (MA), followed, after washing out of unbound 8-MOP molecules, by a second exposure to 365-nm radiation inducing varying relative amounts of CL at a constant level of total photoadducts. In the range of doses used for the second exposure, repair of CL took place; however, in the presence of increased relative amounts of CL induced non-specific incision of DNA occurred. This endonucleolytic cleavage appears to be related to the increased mutagenic and recombinogenic effects observed at increased levels of CL.