Anna Dziedzic-Goclawska

Effect of radiation sterilization on the osteoinductive properties and the rate of remodeling of bone implants preserved by lyophilization and deep-freezing

The effect of various doses of ionizing radiation on the osteoinductive properties of decalcified bone matrices implanted heterotopically and on the rate of remodeling of nondecalcified bone grafts implanted orthotopically in allogeneic systems was studied. Decalcified bone matrices and nondecalcified bone grafts were preserved by lyophilization or by deep-freezing and were subsequently irradiated with appropriate doses at room temperature or at -72 degrees. Lyophilized matrices irradiated at room temperature with 35 and 50 kGy, respectively, were completely resorbed five weeks after heterotopic implantation into the muscles and did not induce osteogenesis, whereas the resorption of deep-frozen ones irradiated with the same doses at -72 degrees was slower and new bone formation was induced. The preservation of the osteoinductive capacity of irradiated, deep-frozen matrices may depend on two factors: reduction of radiation damage on the inducing agents and collagen irradiated in the presence of water, which may diminish the rate of matrix resorption. The rate of remodeling of undecalcified deep-frozen bone implants irradiated at -72 degrees and grafted orthotopically was higher than that of lyophilized ones irradiated at room temperature. It is possible that the temperature during irradiation plays a critical role in protection against radiation damage.

The EPR detection of foods preserved with the use of ionizing radiation

Abstract Solid constituents extracted from irradiated foods have been examined by the epr (esr) spectroscopy. It has been proved that some epr active species produced by radiation in foods are specific and stable enough to be used for the detection of irradiation treatment. The most promising results have been obtained with bones extracted from frozen raw meat (beef, pork, poultry and fish), with seeds of fruits (dates and figs), with dried mushrooms, gelatin and macaroni.

Decrease of the crystallinity of bone mineral in osteopetrotic rats

Abstract The membraneous and long bones of three groups of Fatty Orl-op rats i.e. phenotypically normal animals, osteopetrotic (op/op) mutants, and op/op mutants cured by transplantation of normal bone marrow were compared with regard to the amount and crystallinity of inorganic constituents. The electron spin resonance spectrometry based on measurements of stable paramagnetic centers induced by ionizing radiation in the crystalline lattice of bone hydroxyapatite was used for evaluation of crystallinity of bone mineral. The crystallinity coefficient defined as the ratio of spin concentration of these centers to the total ash content of the sample was calculated. In compact bone of op/op mutants a small increase (about 4%) of mineral content was observed, while the crystallinity of mineral was significantly lower, reaching about 80% of the value found in the phenotypically normal littermates. In op/op mutants cured by transplantation of normal bone marrow the trend for normalization was found. These observations demonstrate the possibility that the crystallinity of bone mineral can be used as a parameter in the diagnosis and monitoring of the therapy in osteopetrosis.

Initial studies on the crystallinity of the mineral fraction and ash content of isolated human and bovine osteons differing in their degree of calcification

SummarySeveral groups containing 10–15 isolated osteons differing in their degree of maturity were analysed. Samples were isolated from undecalcified human and bovine bone sections. The crystallinity coefficient, defined as the ratio of the number of radiation-induced paramagnetic defects in the crystalline lattice of hydroxyapatite to the total ash content, was calculated. The results were compared with measurements performed on fragments of total cortical bone, primary periosteal bone, and inner circumferential lamellar bone. The results show a higher crystallinity of fully calcified osteons as compared with that found at the initial stage of calcification. No differences in the ash content were observed between human osteons, from different stages of calcification evaluated morphologically and by X-ray absorption. These differences were evident when bovine osteons differing in their stage of calcification were compared. Human fully calcified osteons contain 60% ash and their crystallinity coefficient is 52.1. Human osteons at the initial stage of calcification contain 57% ash and their crystallinity coefficient is 40.6. The same parameters for fully calcified bovine osteons and for bovine osteons at the initial stage of calcification are 59% ash, 62.6 crystallinity coefficient and 46% ash, 43.0 crystallinity coefficient, respectively.

Radiation-induced paramagnetic centers in research on bone physiopathology.

It has been found that ionizing radiation induces stable paramagnetic centers in the crystalline lattice of bone hydroxyapatite (HA). These defects, which can be detected and measured by electron spin resonance (ESR) spectrometry, can be treated as a new kind of biologic marker and used for the following: (1) evaluation of crystallinity of bone mineral, because the yield of these centers depends on the size of HA crystals; (2) quantification of the rebuilding process of radiation-sterilized bone grafts, because of the stability of the centers; and (3) estimation of the dose of ionizing radiation absorbed by living organisms, because of the linear relation between the dose of ionizing radiation and the yield of radiation-induced paramagnetic centers in bone mineral.

Crystallinity of human pineal calcospherulites.

SummaryCrystallinity of mineral in human pineal calcospherulites was determined by electron spin resonance spectrometry after irradiation of the samples with gamma rays in a60Co-source. The radiation-induced stable paramagnetic centers in the crystalline lattice of hydroxyapatite crystals were used as a marker of the crystalline fraction and related to the total mineral content. The crystallinity of pineal sand is higher than that of compact bone. The numerical value of the crystallinity coefficient depends on both the average crystal size of hydroxyapatite and the percentage of the crystalline fraction in the total amount of mineral. Literature data show that the average size of hydroxyapatite crystals in pineal sand are smaller than in bone tissue. It is, therefore, concluded that the higher crystallinity of pineal acervuli is due to the lower percentage of the submicrocrystalline fraction in their mineral.

Analysis of the crystallinity of calcium phosphate deposits in rat liver mitochondria by electron spin resonance spectroscopy

The problem of the origin and nature of intramitochondrial mineral deposits is widely discussed because of the possible role of mitochondria in the calcium metabolism of the cell [ 1,2] These are sug gestions that physiologically calcium-loaded mitochondria are involved in the early stages of skeletal tissue mineralization [ 1,3] . At the First International Conference on Matrix Vesicle Calcification (New York 1974) evidence was given for a relationship between Ca ion transfer from mitochondria to the plasma membrane and the matrix vesicles associated with endochondrial ossification [4,5]. Intramitochondrial accumulation of inorganic Ca salts is observed in a variety of physiological conditions in vivo and also as the result of in vitro incubation of fragments of tissues, cells or isolated mitochondria in media with a high concentration of Ca’+ [6]. Amorphous granules as well as crystalline needle-shaped mineral deposits were described in mitochondria from various tissues [6]. On the other hand, only amorphous calcium phosphate has been observed to accumulate in isolated mitochondria [ 1,7,8]. Electron microscopy of blue crab hepatopancreas mitochondria shows bundles of needlelike crystals [9]. The way of preparation and preservation of mitochondria can influence the physico-chemical state of these deposits [ 1 O] Various methods as chemical analysis, electron microscopy, electron diffraction, X-ray diffraction as well as X-ray and electron probe analyses have been used to study intramitochondrial mineral deposits. In this paper a new approach is suggested which might help in the analysis of different kinds of intracellular tnineral deposits. It was shown [ 1 1 ] that with the use of electron spin resonance (ESR) spectrometry one can detect the crystalline fraction of the tissue mineral after irradiation of the sample with ionizing radiation. The irradiation produces paramagnetic defects in the crystalline lattice of hydroxyapatite [ 1 l131. The number of these paramagnetic centers detected and measured by the ESR technique related to the total ash content of the sample gives information on the crystallinity of the tissue mineral [ 141 . The advantage of ESR spectrometry in the detection of hydroxyapatite is connected with the fact that other forms of calcium salts present in the sample do not interfere with the detection of the signal derived from the paramagnetic centers induced in the hydroxyapatite crystals. The high sensitivity of this method allows the detection of 1 0e9 mole equivalent of spins in the sample.

Limitations of Bone Harvesting from Organ Donors in Poland

The demand for organ transplantations greatly exceeds the number of organ donors (OD) and it is likely that this discrepancy will continue. There is a continuously growing demand for biostatic allogenic bone transplantation mostly for orthopedic reconstructions. The bone material is predominantly harvested during postmortem examinations in forensic medicine departments. The collected amounts are not enough so the material need to be acquired from other sources. Bone collecting from OD seems to be the optimal solution. Between January 2000 and December 2005, 1,883 out of 2,601 organ donors fulfilled the criteria for bone donation, but only 42 bone harvesting (BH) were performed being only 2.22% of all suitable cases. The main reasons for so few bone procurements were: lack of acceptance of bone procurement by relevant part of donors’ families, insufficient cooperation between tissue banks and transplantation teams, lack of equipment for BH, and no experience in bone procurement.

Regional program for bone allograft harvesting following multiorgan procurement in Pomeranian Academy of Medicine in Szczecin.

In Poland, up to 1999, the bones for allograft preparations had been procured only in mortuaries of forensic medicine departments. The increasing demand for bone transplantations greatly exceeds the supply resulting in a long waiting time for bone allografts. In November 1999, for the first time in Poland, the group of orthopedic surgeons from the Pomeranian Academy of Medicine started the regional program for bone harvesting following vascularized organ procurements. The aim of this paper is to present the technical details and limitations of bone harvesting that occurred in 10 out of 25 multiorgan procurements.

THE EFFECT OF PRESERVATION PROCEDURES AND RADIATION STERILISATION CONDITIONS ON CONNECTIVE TISSUE GRAFTS AND THEIR CONSTITUENTS

ABSTRACT The Central Tissue Bank, established in 1963, and two other multi-tissue banks operating in Poland provide connective tissue allografts such as bone, cartilage, tendons, sclera, skin, acellular dermis and amnion. All grafts are radiation sterilised with a dose of 35 kGy in a 60Co source and/or with electron beam 10 MeV accelerator. Over 250,000 radiation-sterilised tissue grafts have been prepared and used in hospital departments throughout Poland and not one case of infectious disease transmission has been reported to date. High doses of ionising radiation can evoke numerous chemical and physical changes that may affect biological quality of tissue allografts, such as osteoinductive potential of bone, the mechanical properties of bone and other connective tissue grafts as well as the rate of their resorption in vivo. The origin and stability of free radicals and other paramagnetic entities radiation-induced in bone will be discussed. The effect of various preservation procedures (e.g. lyophilisation, deep-freezing) and radiation sterilisation conditions (doses, temperature of irradiation) on osteoinductive potential of bone as well as on degradation of collagen, a major constituent of connective tissue grafts, will be presented. The results of interdisciplinary research performed at the Central Tissue Bank in Warsaw, in collaboration with radiation chemists from the Institute of Nuclear Chemistry and Technology indicate, that radiation-induced changes can be diminished by modification of tissue preservation methods and that, to some extent it is possible to reduce undesired radiation-induced damage to the tissue grafts.