Biomedical subjects

A Pizzoferrato

Publications and source records attributed to A Pizzoferrato.

At least 37 records · Page 2Linked to original sources

PubMed1999

Cd 34+ cells and clonogenicity of peripheral blood stem cells during chemotherapy treatment in association with granulocyte colony stimulating factor in osteosarcoma.

The introduction of aggressive chemotherapy in the treatment of osteosarcoma has improved the long-term outcome for these patients. With the increasing aggressiveness of chemotherapy protocols, hematopoietic growth factors have emerged as useful adjuncts involving, in some cases, rescue by peripheral blood stem cell (PBSC) infusion to assist faster recovery and maintain relative dose intensity. To evaluate the number of PBSCs needed, we analyzed the number of CD34+ cells and hematopoietic progenitor cells in the peripheral blood of 16 patients with osteoblastic, condroblastic and fibroblastic osteosarcoma enrolled in an Istituto Ortopedico Rizzoli-Scandinavian Sarcoma Group (IOR-SSG) pilot study, consisting of two cycles of preoperative high dose chemotherapy. The blood samples were studied at different times. The CD34+ cells were analyzed by flow cytometry and the hematopoietic progenitor cells were analyzed by tissue culture clonogenic assay. In comparing the two courses of chemotherapy, we observed that modification of the mean values of WBC, CD34+ and CFU-GM were very similar. The second course of chemotherapy seemed to induce greater hematological toxicity. All three parameters showed good correlation. The results demonstrated that the best time to collect PBSC by means of leukapheresis is post G-CSF used as rescue after ifosfamide treatment. We verified the ability of G-CSF to mobilize PBSCs in patients with osteosarcoma through cytofluorimetric analysis of CD34+ cells and their clonogenic capability. Moreover, during this preoperative treatment, we identified the best time to collect a sufficient number of PBSCs, that is after 9-10 days of G-CSF treatment following the first cycle of ifosfamide.

Adolescent↗

PubMed1999

[Cytokines of bone turnover in postmenopause and old age].

BACKGROUND: The aim of the study was the assessment of the influence of cytokines on bone ageing by measuring their level in serum and their secretion in vitro by monocytes from women of different age. METHODS: The levels of cytokines in 34 postmenopausal subjects and 14 old subjects were compared to those measured in 13 cycling subjects who were considered as control group. Subjects suffering from diseases inducing secondary osteoporosis, subjects taking medications that affect bone metabolism and alcohol- or tobacco-consumers were excluded from the study. The levels in serum of (i) the bone stimulating peptide insulin-like growth factor-I (IGF-I), (ii) the inhibitor of bone resorption interferon-gamma (IFN gamma), (iii) the stimulators of bone resorption interleukin-1 beta (IL-1 beta) tumor necrosis factor alpha (TNF alpha) and interleukin 6 (IL-6) were evaluated by immunoassay. IL-1 beta, TNF alpha and IL-6 secreted by monocytes (MO) cultured in vitro from peripheral blood of the same subjects were measured, too. Bacterial endotoxin (LPS) was used as stimulator for cytokine secretion by monocytes. RESULTS: Unlike IFN gamma, which was unaltered, circulating IGF-I level was found significantly diminished in postmenopausal and old subjects compared to control group. Among stimulators of bone resorption, IL-6 was greatly increased in postmenopausal and old subjects, while TNF alpha was reduced in postmenopausal group. In the supernatants of unstimulated monocytes the level of IL-1 beta was consistently decreased in old subjects; TNF alpha was found to be decreased in postmenopausal and old subjects. The stimulation index (SI), calculated as the ratio between the level of cytokines secreted by LPS-stimulated MO and the level of cytokines secreted by unstimulated MO, was found to be significantly increased for IL-1 beta and TNF alpha in postmenopausal subjects vs control group. In the old subjects the SI for IL-6 was enhanced. CONCLUSIONS: The data collected suggest that the measurement of cytokines in serum and supernatants from monocytes may give a picture of the mechanisms regulating bone aging.

Adult↗

PubMed1998

Fluorescent microplate assay for respiratory burst of PMNs challenged in vitro with orthopedic metals.

This report describes a simple, rapid, automated microassay for measuring in vitro changes of oxidative burst of phagocytes following challenge with metals for orthopedic devices. The production of reactive oxygen species (ROS) by polymorphonuclear leukocytes (PMNs) was measured using 2',7'-dichlorofluorescin-diacetate (DCFH-DA) as fluorescent probe. DCFH-DA enters the cells and is oxidized by ROS to fluorescent DCF. The DCF generated was directly proportional to ROS produced intracellularly: The fluorescence intensity was read and converted to an index of ROS production by cells. In our experimental system, granulocytes (PMNs) were isolated from normal human blood and seeded in microplates. To verify if metals could influence ROS production, chromium, cobalt, nickel, molybdenum, titanium, aluminum, and vanadium prepared as aqueous extracts in phosphate-buffered saline were tested onto PMNs using phorbolmyristate acetate (PMA) as positive control. Molybdenum, aluminum, and vanadium increased ROS generation by PMNs, while signals not different from unstimulated PMNs were recorded for chromium, cobalt, nickel, and titanium. The DCFH-DA microplate-based assay provides an in vitro tool for the detection of oxygen-reactive species generated by PMNs as a response to metals.

Fluoresceins↗

PubMed1998

In vitro sister chromatid exchange induced by glass ionomer cements.

The genotoxicity of three glass ionomer cements used in dentistry, manufactured by American (Vitrebond), Japanese (Fuji I), and European (Ketac Cem) companies were examined. The cement components were mixed according to the manufacturers' instructions and allowed to set for two defined times: 1 h or 1 week, before extracting them, as established by ISO standard 10993 part 12. To highlight sister chromatid exchange during mitosis, the extracts then were tested with human peripheral lymphocytes in the presence or absence of metabolic activation with S9 mix. The test performed was a genotoxicity test as provided for in standard EN 30993 part 3. Vitrebond resulted in direct genotoxicity and was strongly cytotoxic both in the extracts performed at 1 h and those at 1 week if they were allowed to set without photoactivation. Fuji I was noncytotoxic and showed only uncertain indirect genotoxicity in the extracts at 1 h; genotoxicity was not present in the extracts at 1 week. Ketac Cem cement was not genotoxic nor was it cytotoxic either at 1 h or 1 week. The authors concluded that of the three cements tested the European cement Ketac Cem passed one of the tests suggested by the EEC standard for assessing genotoxicity.

DNA Damage↗

PubMed1998

Flow-cytometric analysis of leukocyte activation induced by polyethylene-terephthalate with and without pyrolytic carbon coating.

Leukocyte activation is one test for the evaluation of blood-materials interaction. The expression of adhesion molecules analyzed by flow cytometry provides a simple method to evaluate leukocyte activation by biomaterials: any change in these molecules can be predictive of the inflammatory activity of the materials. In this study the contact between leukocytes and uncoated polyethylene terephthalate or pyrolytic carbon-coated polyethylene terephthalate (PET and PET-PC, respectively) was inspected by analyzing whether the expression of some adhesion molecules involved in leukocyte activation, namely LFA-1 (CD11a/ CD18), Mac-1/CR3 (CD11b/CD18), and LECAM-1 (CD62L) can be modified. By flow cytometry expression of the adhesion molecules can be studied separately on lymphocytes and myeloid cells. The materials tested reduced the total numbers of both leukocytes and neutrophils, although not significantly. Neither PET nor PET-PC changed the expression of the adhesion molecules in lymphocytes: this suggests that no specific immune response is stimulated. On the contrary, statistically significant changes were observed for monocytes and granulocytes: the percentage of cells expressing Mac-1 and the density of such antigens on cell membranes increased while the percentage of LECAM-1 positive cells decreased. Similar changes were observed when the cells underwent the inflammatory stimulus provided by an in vitro challenge with bacterial endotoxin. Our results demonstrated that polyethylene terephthalate activates leukocytes by modifying the expression in neutrophils of the molecules involved in the early phase of the inflammatory response. Even after coating PET with pyrolytic carbon, the ability of this material to activate circulating leukocytes was maintained.

Biocompatible Materials↗

PubMed1998

False positive results in cytotoxicity testing due to unexpectedly volatile compounds.

We investigated the cytotoxicity of different dental materials according to the study protocol adopted by our lab for the screening of new materials. Experimental parameters used in such testing are addressed mainly in documents EN 30993 "Biological evaluation of medical devices, Part 5: Tests for cytotoxicity: in vitro methods" and "Biological evaluation of medical devices, Part 12: Sample preparation and reference materials." Cells were cultured in microplates and challenged with aqueous extracts of the materials. The assay methods were neutral red- and propidium iodide-uptake assays, both indicative of cell viability and able to provide quantitative data. The observation of contrasting results for one material using the above-mentioned methods raised some concern about the assay system used. With further experimentation, it appeared that a sustained release of volatile substances still present in one extract exerted a toxic effect in neighboring cultures. It is concluded that in the microenvironment of a microplate the distribution of samples cannot be disregarded, as it may be responsible for toxicity cross-contamination. Moreover, the use of more than one single method has to be recommended in cytotoxicity testing, in order to avoid false positive results due to experimental artifacts.

Animals↗

PubMed1998

The interface of bone microstructure and an innovative coating: an X-ray diffraction study.

The in vivo compatibility and degradation aspects of an innovative coating to be sprayed onto titanium implants were investigated. The surface of fluorinated apatite (fHA), consisting of fluorhydroxyapatite plasma sprayed in a vacuum atmosphere, was treated with carbonate to improve its biological compatibility. fHA coating was compared with titanium implants coated (a) with hydroxyapatite (HA) by the traditional plasma spraying, and (b) with titanium oxide (TiOx). Screw-shaped implants were inserted in the cortical bone of sheep tibiae. X-ray diffraction (XRD) analysis of bone tissue and coatings was carried out at 2, 4, 12 and 36 weeks after surgery. The crystallographic habit of the implant-facing bone, as well as the structural stability of the coating, were evaluated. For each time period and type of ceramic bone apatite lattice at the interface, no significantly different reference apatite lattice and no foreign peak were recorded. Two weeks after implantation, the bone at the interface was strongly unmineralized in all samples; after 4 weeks, poorly mineralized bone microareas decreased. At 12 weeks, the newly formed bone tissue at the interface with both the new coating and HA coating was shown to be fully mineralized; this crystallographic habit was retained at 36 weeks, when particle release from the tested material was lower compared to the controls. The XRD pattern of bone apatite surrounding the coating particles was unmodified. The innovative coating did not alter the mineralization process at the interface. It improved implant osteointegration, mainly due to a limited release of particles. Consequently, clinical performance of external fixation treatment could be improved by modifying the chemical composition of the implant surface.

Animals↗

PubMed1998

Lipid peroxidation and antioxidant status in adults receiving lipid-based home parenteral nutrition.

BACKGROUND: Infusion of lipid emulsions rich in polyunsaturated fatty acids (PUFAs) may increase lipid peroxidation, which is counteracted mainly by superoxide dismutase (SOD) (a zinc-, copper-, and manganese-dependent enzyme), selenium-dependent glutathione peroxidase (Se-GSHPx), and alpha-tocopherol. OBJECTIVE: We investigated lipid peroxidation and antioxidant status in patients receiving home parenteral nutrition (HPN) providing variable amounts of a lipid emulsion rich in PUFAs, and alpha-tocopherol, zinc, copper, and manganese as recommended by the American Medical Association, and no selenium. DESIGN: Serum malondialdehyde, plasma alpha-tocopherol, selenium, Se-GSHPx, PUFAs, and red blood cell Se-GSHPx and SOD were evaluated in 12 patients and in 25 healthy control subjects. Malondialdehyde was also assessed in a group of 40 healthy control subjects. RESULTS: Patients had significantly higher concentrations of malondialdehyde and SOD and lower alpha-tocopherol concentrations and selenium nutritional status. Linear regression analysis showed that malondialdehyde was associated with the daily PUFA load (r=0.69, P< 0.03) and with plasma alpha-tocopherol (r=-0.59, P< 0.05), but stepwise multiple regression analysis confirmed only the association between malondialdehyde and alpha-tocopherol; plasma alpha-tocopherol was associated with the daily PUFA load (r=-0.65, P< 0.04) and with the duration of HPN (r=-0.74, P< 0.02). CONCLUSIONS: In HPN patients, the peroxidative stress due to lipid emulsions rich in PUFAs is counteracted primarily by alpha-tocopherol. The dosages of alpha-tocopherol, zinc, copper, and manganese recommended by the American Medical Association appear sufficient to sustain SOD activity but inadequate to maintain alpha-tocopherol nutritional status. HPN formulations should be supplemented with selenium.

Adolescent↗

PubMed1997

CD62, thromboxane B2, and beta-thromboglobulin: a comparison between different markers of platelet activation after contact with biomaterials.

The authors examined the modifications of some markers of platelet activation after contact with biomaterials. Glycoprotein GMP-140 (CD62) was evaluated by flow cytometry; beta-thromboglobulin (beta-TG) and thromboxane B2 (TXB2) were determined by radioimmunoassay. Polyethylene terephthalate (PET) induced a remarkable platelet adhesion and a significant increase in beta-TG and TXB2, with no increase in CD62 on the nonadherent platelets. Pyrolytic carbon-coated PET (PC) did not induce platelet adhesion after 15 min of contact, but a significant increase in CD62 was detected. After 30 min a significant increase in platelet adhesion as well as the release of beta-TG and TXB2 were noted. The increase was lower than that observed for uncoated PET, and after 30 min of contact with PC the increase no longer was observed.

Biocompatible Materials↗

PubMed1997

High-performance liquid chromatography assay of N,N-dimethyl-p-toluidine released from bone cements: evidence for toxicity.

Five commercially available bone cements were analysed by high-performance liquid chromatography for detecting the residual content of an accelerator, the amine N,N-dimethyl-p-toluidine (DMPT), after curing. It was found that the concentration of DMPT in aqueous extracts decreases with time, being almost absent 7 days after curing. Differences were noticed among the cements; residual DMPT is higher in cements prepared with higher content of the amine. It is verified that DMPT's toxic effect on cell cultures is dose-related; a delay in the cell replication cycle is induced in vitro. Damage is reversible, thus justifying the low bone cement toxicity that is clinically ascertained.

Bone Cements↗

PubMed1997

Expression of adhesion molecules on endothelial cells after contact with knitted Dacron.

The aim of this study is to evaluate the expression of some adhesion molecules on the surface of endothelial cells cultured in contact with knitted Dacron. These molecules, as mediators of cell adhesion, could play a role in the modulation of adhesion on the biomaterials, therefore conditioning the response of tissues to implant. Twenty different cultures of human umbilical vein endothelial cells (HUVECs) were cultured in contact with knitted Dacron. Both HUVECs grown without the material and HUVECs incubated with endotoxin were used as control. After 24 h, the cell adhesion molecules PECAM-1, ELAM-1, ICAM-1 and VCAM-1 were evaluated on the cells by monoclonal antibodies and flow cytometry. After 24 h of contact with knitted Dacron, a significant decrease in the proportion of cells expressing PECAM-1 was observed, as well as a significant increase in the proportion of cells expressing ELAM-1. The contact with knitted Dacron did not induce significant variations of ICAM-1 and VACM-1. The incubation with endotoxin determined a significant increase in the proportion of ELAM-1-positive cells, a significant increase in ICAM-1 fluorescence intensity, and a significant increase both in fluorescence intensity and in the proportion of VCAM-1-positive cells. The results obtained with the endotoxin are in agreement with those reported in the literature. The ELAM-1 increase, observed after contact with knitted Dacron, could favour leucocyte adhesion, while the decrease in PECAM-1 expression could result from an inhibiting effect on the endothelial cell adhesion so as to hinder the mechanisms involved in the endothelialization of the material. The variations were interpreted as inhibiting endothelialization and favouring the leucocyte adhesion effect by knitted Dacron.

Analysis of Variance↗

PubMed1997

Determinants of functional status in healthy Italian nonagenarians and centenarians: a comprehensive functional assessment by the instruments of geriatric practice.

OBJECTIVE: To evaluate the physical ability and psychocognitive status of a population more than 90 years of age with regard to sociodemographic, behavioral, and biomedical variables known to affect functional status in old age. DESIGN: A survey design was used. SETTING: Emilia Romagna, Northern Italy. PARTICIPANTS: Eighty-four healthy community-dwelling subjects aged 90 to 106 years. MEASUREMENTS: Sociodemographic variables, health behavior, anthropometric indices, and serum DHEAS levels were recorded. Functional assessment was performed by instruments currently used in geriatric practice: the Mini-Mental State Examination (MMSE), Geriatric Depression Scale (GDS), and Activities of Daily Living (ADL) scale. A stepwise multiple regression analysis was performed. RESULTS: GDS scores correlated directly with MMSE scores and inversely with ADL severity scores. Poor education, institutionalization, sensory impairment, muscular mass loss, and lower DHEAS levels were the variables with the highest correlation to functional impairment. Smoking, alcohol consumption, and marital status were relatively unimportant. An inverse association was found between DHEAS levels and dependency scores of single ADLs (continence, mobility). CONCLUSION: Impaired cognitive and physical ability with no increase in depression prevalence was found in a sample of subjects more than 90 years of age free of major age-related disease. Muscular mass and DHEAS levels seem to play a role in maintaining physical independence. In turn, physical independence, as well as social and cultural factors, strongly affect the compliance of long-lived subjects with psychocognitive tests currently used in the clinical evaluation of younger old people, suggesting that these instruments are not reliable for screening for cognitive impairment and depression in the oldest old subjects.

Activities of Daily Living↗

PubMed1997

[In vitro production of endothelin-1 and prostacyclin by cultured endothelial cells in the presence of polymers].

The aim of this study was to evaluate endothelin-1 and prostacyclin production by human endothelial cells cultured in the presence of polyethylene terephthalate and collagen-coated PET. Cell counting and the assay of endothelin-1 and 6-keto-prostaglandin F1 alpha, stable metabolite of prostacyclin, were carried out after 48 hour contact of the cells with the examined materials. Endothelial cell contact with uncoated PET caused a significant reduction in cell number, a significant increase in the production of endothelin-1 and a not significant increase in 6-keto-prostaglandin F1 alpha. The endothelial cell contact with collagen-coated PET caused a highly significant decrease in cell number and a not significant decrease in endothelin-1 and 6-keto-prostaglandin F1 alpha. It was concluded that PET causes both a decrease in cell number and a remarkable increase in endothelin-1. On the contrary, collagen-coated PET determines a decrease in cell number and a slight reduction of endothelin-1 and 6-keto-prostaglandin F1 alpha.

Cells, Cultured↗

PubMed1997

[Plasma protein adsorption on variously treated +polybutylene terephthalate].

BACKGROUND: The aim of the research was the evaluation of plasmatic protein adsorption on untreated polybutylene terephthalate, corona treated polybutylene terephthalate and polyvinylacetate coated corona treated polybutylene terephthalate. METHODS: Total proteins, albumin, immunoglobulins, fibrinogen, insulin and ostecalcin were determined on plasma after contact with these polymers. RESULTS: Unsignificant variations were observed for the assayed proteins. CONCLUSIONS: The conclusions are drawn that the tested materials do not adsorbe significantly the most important plasmatic proteins.

Adsorption↗

PubMed1996

Activation of the plasma coagulation system induced by some biomaterials.

The ability of some biomaterials to activate plasma coagulation system was examined in vitro. After contact of platelet-rich plasma with biomaterials, some markers of the thrombin formation, i.e., fragment 1 + 2 and fibrinopeptide A, and some inhibitors of the blood coagulation mechanism were tested. Fragment 1 + 2 and fibrinopeptide A were found to be increased by all of the materials, though to a different extent. In particular, fragment 1 + 2 and fibrinopeptide A were significantly increased upon contact with polybutylene terephthalate and with collagen coated polyethylene terephthalate, respectively. Also antithrombin III was shown to decrease following exposure to biomaterials, but statistical significance was found only for polyethylene terephthalate and polyvinylacetate. As a results of this wide range of variability in the parameters, it is advisable to explore the plasma coagulation system with a multiparametric approach in which thrombin formation and coagulation inhibitors are thoroughly investigated.

Antithrombin III↗

PubMed1996

Assessment of metal extract toxicity on human lymphocytes cultured in vitro.

In this study the toxic effects of chromium, nickel, and cobalt extracts on in vitro cultured lymphocytes were evaluated. Graphite furnace atomic absorption spectrometry was used to measure the ion concentration. After serial dilution of the extracts, the viability of lymphocytes at 24, 48, and 72 h was estimated by flow cytometry, including propidium iodide staining and light scatter property assessment, and by MTT reduction test. The results of the investigation allowed us to conclude that 1) standardization of the procedure for preparing extracts is fundamental to obtaining repeatability of results; 2) the toxicity of an extract cannot be evaluated with a single viability assay; a combination of functional and structural tests is required; 3) when methods based on enzymatic reactions are performed, e.g. MTT test, it is advisable to replace the extract containing metal ions with fresh medium in order to avoid any interference with viability testing; 4) the amount of Co and Ni in the extract is similar, but the Cr release is very poor; 5) the lower toxicity of Cr extract probably is due to the lower ion concentration; 6) the assessment of 50% cytotoxic concentration (TC50) allows quantification of materials toxicity and comparison of various metals; and 7) the determination of a noncytotoxic concentration, i.e., a concentration lower than TC10, is required for subsequent investigation of cell functions because such studies can be carried out only on viable cell population.

Cell Survival↗

PubMed1996

Cytokine production and adhesive protein expression by endothelial cells after contact with polyethylene terephthalate.

The authors have evaluated adhesive protein expression and cytokine production by human umbilical vein endothelial cells cultured in contact with polyethylene terephthalate (PET). ELAM-1, ICAM-1 and VCAM-1 expression was determined by flow cytometry; the concentration of interleukin-1 alpha (IL-1 alpha), interleukin-6 (IL-6), granulocyte colony stimulating factor (G-CSF) and granulocyte-macrophage colony stimulating factor (GM-CSF) in the supernatant was determined by enzyme immunoassay. The contact with PET determined a significant increase in ELAM-1 expression and insignificant increase in cytokine production, demonstrating that PET had a limited capability to stimulate endothelial cells in a pro-inflammatory sense.

Cell Adhesion Molecules↗

PubMed1996

In vitro assessment of phagocytosis of bovine collagen by human monocytes/macrophages using a spectrophotometric method.

The use of a wound dressing with covering and haemostatic properties significantly improves wound healing. In this study, a lyophilized bovine collagen sponge used for the treatment of wounds and ulcerae has been tested in a cell culture system. Phagocytosis of collagen fragments by human blood monocytes/macrophages has been investigated. For the assessment of collagen ingestion by mononuclear phagocytes, a picrosirius dye specific for collagen molecules has been used. By adapting this histochemical technique to microplate cell culture system, replicate monocyte cultures are assayed. Collagen content is determined by evaluating spectrophotometrically at 540 nm the absorbance of a sirius red/picric acid solution. Using this simple and sensitive method, the phagocytosis of bovine collagen by LPS-stimulated monocytes/macrophages has been ascertained.

Analysis of Variance↗