Induction of micronuclei in human fibroblasts and keratinocytes by 25 kV x-rays

A relative biological effectiveness (RBE) not much larger than unity is usually assumed for soft x-rays (up to approximately 50 keV) that are applied in diagnostic radiology such as mammography, in conventional radiotherapy and in novel radiotherapy approaches such as x-ray phototherapy. On the other hand, there have been recent claims of an RBE of more than 3 for mammography and respective conventional x-rays. Detailed data on the RBE of soft x-rays, however, are scarce. The aim of the present study was to determine the effect of low-energy x-rays on chromosomal damage in vitro, in terms of micronucleus induction. Experiments were performed with 25 kV x-rays and a 200 kV x-ray reference source. The studies were carried out on primary human epidermal keratinocytes (HEKn), human fibroblasts (HFIB) and NIH/3T3 mouse fibroblasts. Micronucleus (MN) induction was assayed after in vitro irradiation with doses ranging from 1 to 5.2 Gy. Compared to the effect of 200 kV x-rays, 25 kV x-rays resulted in moderately increased chromosomal damage in all cell lines studied. This increase was observed for the percentage of binucleated (BN) cells with micronuclei as well as for the number of micronuclei per BN cell. Moreover, the increased number of micronuclei per micronucleated BN cell in human keratinocytes and 3T3 mouse fibroblasts suggests that soft x-rays induce a different quality of damage. For all cell lines studied the analysis of micronucleus induction by 25 kV soft x-rays compared to 200 kV x-rays resulted in an RBE value of about 1.3. This indicates a somewhat enhanced potential of soft x-rays for induction of genetic effects.     

Experimentally induced chromosome abberrations in plants. II. The effect of cyanide and other heavy metal complexing agents on the production of chromosome aberrations by x-rays

The discovery of Lilly and Thoday, that the presence of potassium cyanide (KCN) increases the production of chromosome aberrations by x-rays in anoxia, but has no effect on the production of chromosome aberrations by x-rays in air, was confirmed. In the presence of cyanide, the effect of a given dose of x-rays in nitrogen was found to be even greater than the effect of the same dose of x-rays in air. The cyanide effect on x-ray breakage in nitrogen was obtained at cyanide concentrations as low as 2 x 10(-5)M. The breakage obtained after the combined x-ray-cyanide treatments was of the x-ray type, as evidenced by the distribution of breaks within and between the chromosomes. A number of other heavy metal complexing agents as well as some other compounds were tested for their ability to increase x-ray breakage in nitrogen and air. Of these compounds only cupferron proved to be effective. The results are discussed and it is concluded that the increased x-ray breakage in the presence of cyanide or cupferron cannot be due to an accumulation of peroxides. Instead it is suggested that the cyanide effect may be due to a complex formation between the active agents and heavy metals, presumably iron, within the chromosomes. The consequences of this hypothesis on the concept of the "oxygen effect," are discussed.     

EXPERIMENTALLY INDUCED CHROMOSOME ABERRATIONS IN PLANTS : II. THE EFFECT OF CYANIDE AND OTHER HEAVY METAL COMPLEXING AGENTS ON THE PRODUCTION OF CHROMOSOME ABERRATIONS BY X-RAYS

The discovery of Lilly and Thoday, that the presence of potassium cyanide (KCN) increases the production of chromosome aberrations by x-rays in anoxia, but has no effect on the production of chromosome aberrations by x-rays in air, was confirmed. In the presence of cyanide, the effect of a given dose of x-rays in nitrogen was found to be even greater than the effect of the same dose of x-rays in air. The cyanide effect on x-ray breakage in nitrogen was obtained at cyanide concentrations as low as 2 x 10^–5 M. The breakage obtained after the combined x-ray-cyanide treatments was of the x-ray type, as evidenced by the distribution of breaks within and between the chromosomes. A number of other heavy metal complexing agents as well as some other compounds were tested for their ability to increase x-ray breakage in nitrogen and air. Of these compounds only cupferron proved to be effective. The results are discussed and it is concluded that the increased x-ray breakage in the presence of cyanide or cupferron cannot be due to an accumulation of peroxides. Instead it is suggested that the cyanide effect may be due to a complex formation between the active agents and heavy metals, presumably iron, within the chromosomes. The consequences of this hypothesis on the concept of the "oxygen effect," are discussed.     

Clonogenic survival of human keratinocytes and rodent fibroblasts after irradiation with 25 kV x-rays

Low energy x-rays (E_ph 50 keV) are widely used in diagnostic radiology and radiotherapy. However, data on their relative biological effectiveness (RBE) are scarce. Of particular importance for risk estimation are the RBE values of x-rays in the range which is commonly used in mammography (10–30 keV). We have determined clonogenic survival after low-energy x-ray irradiation for three cell lines: primary human epidermal keratinocytes (HEKn), mouse fibroblasts (NIH/3T3) and Chinese hamster fibroblasts (V79). Experiments were performed with a 25 kV x-ray tube and compared to 200 kV x-rays as a reference. Compared to the effect of 200 kV x-rays, irradiation with 25 kV x-rays resulted in a decreased survival rate in the murine fibroblasts, but not in the human epithelial cell line. The RBE value was calculated for 10% surviving fraction. For HEKn cells, RBE was 1.33±0.27, for NIH/3T3 cells 1.25±0.07 and for V79 cells 1.10±0.09, respectively. No consistently increased RBE was observed in the various cell lines. Nevertheless, a potential of increased cytogenetic changes has to be considered for risk estimation of low-energy x-rays.     

Analysis of chromosome aberrations in human peripheral lymphocytes induced by 5.4 keV x-rays

Irradiation of human lymphocytes by x-rays has been seen, in past studies, to produce increasing frequencies of chromosome aberrations at lower x-ray energies. However, in one earlier irradiation experiment with chromium x-rays, the relative biological effectiveness (RBE) did not appear to be larger than that of hard x-rays, especially at higher doses. A possible reason for this unexpected result may have been the irradiation and culture conditions. We have, therefore, in the present study used a technique that has been developed in our laboratory to ensure uniformity of irradiation within lymphocytes and to avoid artefacts due to the cell cycle kinetics. Monolayers of 3-h-stimulated lymphocytes were exposed to 5.4 keV x-rays. A linear-quadratic dose-response was found for dicentrics. The comparison to an earlier finding with 220 kV x-rays shows the expected result of the RBE of the 5.4 keV x-rays to be above that of 220 kV x-rays. The intercellular distribution of dicentrics did not differ significantly from a Poisson distribution. Received: 17 January 1997 / Accepted in revised form: 17 July 1997     

Making sense of shadows: Dr. James Third and the introduction of x-rays, 1896 to 1902.

The discovery of x-rays was announced by German physicist Wilhelm Conrad Röntgen in December 1895. This review of the introduction of the use of x-rays in Kingston, Ont., shows the rapidity of their adoption in Canadian medicine. By February 1896 "x-ray photographs" were being taken by Captain John Cochrane of the Royal Military College of Canada in Kingston. Initially a scientific and popular curiosity, the new rays were quickly applied to medicine, and by the fall of 1896 the Kingston General Hospital had acquired its own x-ray apparatus. The hospital superintendent, Dr. James Third, became a leading practitioner and promoter of radiographic diagnosis and radiation therapy. He published, in 1902, the first comprehensive review of the diagnostic and therapeutic uses of x-rays by a Canadian physician. Third''s writings reveal his technical knowledge, his organized approach to the application of radiography to clinical medicine and his cautious attitude. Like other physicians who have witnessed the introduction of new diagnostic techniques, Third feared that the new technology would usurp the physician''s clinical skills.     

X-ray micrography and imaging of Escherichia coli cell shape using laser plasma pulsed point x-ray sources.

High-resolution x-ray microscopy is a relatively new technique and is performed mostly at a few large synchrotron x-ray sources that use exposure times of seconds. We utilized a bench-top source of single-shot laser (ns) plasma to generate x-rays similar to synchrotron facilities. A 5 microlitres suspension of Escherichia coli ATCC 25922 in 0.9% phosphate buffered saline was placed on polymethylmethyacrylate coated photoresist, covered with a thin (100 nm) SiN window and positioned in a vacuum chamber close to the x-ray source. The emission spectrum was tuned for optimal absorption by carbon-rich material. Atomic force microscope scans provided a surface and topographical image of differential x-ray absorption corresponding to specimen properties. By using this technique we observed a distinct layer around whole cells, possibly representing the Gram-negative envelope, darker stained areas inside the cell corresponding to chromosomal DNA as seen by thin section electron microscopy, and dent(s) midway through one cell, and 1/3- and 2/3-lengths in another cell, possibly representing one or more division septa. This quick and high resolution with depth-of-field microscopy technique is unmatched to image live hydrated ultrastructure, and has much potential for application in the study of fragile biological specimens.     

Stem cell engineering in bioreactors for large‐scale bioprocessing

Stem cells are promising cell sources for many biomedical applications including cell therapy, regenerative medicine, and drug discovery. However, the commonly used static tissue culture vessels can only generate a low number of cells. To provide an adequate number of stem cells for clinical applications, a scalable process based on bioreactors is needed. Stem cells can be either cultured as free cells/aggregates in suspension or as adherent cells on the solid substrates. Based on the cell property, different bioreactor configurations are developed to better expand stem cells while maintaining their differentiation capacity. In this review, several major types of bioreactor systems and their applications in stem cell engineering are discussed. Continued advancements in bioprocess and bioreactor research and development are important to engineer stem cells for their use in biomedical applications.     

DIFFERENTIAL SENSITIVITY OF PROPHASE POLLEN TUBE CHROMOSOMES TO X-RAYS AND ULTRAVIOLET RADIATION

1. Through use of the pollen tube technique it has been possible to study the sensitivity of prophase stages to x-rays and ultraviolet, and to correlate the varying sensitivity with changes in the generative nucleus of Tradescantia. 2. Sensitivity to ultraviolet decreases from the 2 hour stage until at 11 hours after germination there is no further production of breaks. The 0 and 1 hour stages show a decreased sensitivity over the 2 hour stage but it has been suggested that this is not due to a decreased sensitivity but to shielding by the pollen wall. 3. Sensitivity to x-rays rises to a peak at the 4 hour stage, but then subsides until no breaks are realized (at a dose of 370.8 r) after the 10 hour stage. In this respect the effects of x-rays and ultraviolet are similar. Each type of x-ray break shows its own individual trend. 4. Correlation of x-ray breaks with changes in the generative nucleus indicates that the important events determining the sensitivity of the chromosomes to breakage are the uptake of water at the time of germination and the movement involved in spiralization. The total absence of breaks after the 11 hour stage is not understood. 5. The changing sensitivity to ultraviolet may depend on any one or all of three factors: (a) the nucleic acid cycle, (b) changes in the matrix, and (c) the number of subdivisions in the chromosome. These are discussed although their relative importance is not known.     

THE INFLUENCE OF SPECIMEN TOPOGRAPHY ON X-RAY MICROANALYSIS ELEMENT MAPPING

The effect of specimen topography on x-ray microanalysis element mapping was studied with an electron microprobe and a scanning electron microscope equipped for x-ray detection. Using the lemma and palea of rice inflorescences as models, we determined that specimen topography can physically limit the detection of x-rays and thus lead to erroneous element mapping data. Any geometrical point on a specimen interfering with a straight line from the point of excitation to the detector will cause an absorptive shadow area on the element map. Electrons impinging on a sample surface cause emissions to occur in all directions. Emissions with sufficient energy (x-rays and backscattered electrons) can strike a topographical point different from the location of the focused electron beam, causing detectable x-ray excitation. This phenomenon will also result in erroneous element map data. Methods of recognition of specimen topographical effects on x-ray microanalysis are discussed.     

Current trends in photodynamic therapy of neoplasms and non-neoplastic diseases

The photodynamic therapy of tumors is a modern therapeutic modality for organ-preserving treatment of oncological diseases. The method is based on selective laser irradiation of tumor tissues previously sensitized by tumorotropic dyes. During the last decades, photodynamic therapy has become worldwide known as a proper approach to the treatment of the patients with malignant tumors of various locations and a number of nontumoral diseases. The characteristics of modern photosensitizers and light sources for photodynamic therapy and their clinical applications are reviewed.     

A field method for the cephalometric x-ray study of skulls in early Nubian cemeteries

The cephalometer, a clinical and research instrument employed by the orthodontic profession to record standardized x-rays of the human skull, has been modified for research work in remote areas where the usual laboratory facilities are not available. This cephalometer was specifically designed to record cephalograms (head radiographs) on the living Nubian population and their extensive skeletal record. The cephalometer consists of a light, portable tripod stand and an aluminum beam to which is attached at one end a Wehmer cephalostat (head holder), and on the other end an isotope radiation source. Hence, the radiation source and the cephalostat were held in the standard relationship of five feet from source to the mid-saggittal plane of the cranium. Ytterbium-169, with a half life of 32.5 days, was used to expose the film. The isotope and the use of the new Experimental Polaroid X-ray Film permitted the x-ray examination of Nubian skulls with no conventional power source or darkroom facilities.     

Principles and applications of dual source CT

This article describes the technical principles and clinical applications of dual source CT. A dual source CT (DSCT) is a CT system with two x-ray tubes and two detectors at an angle of approximately 90°. Both measurement systems acquire CT scan data simultaneously at the same anatomical level of the patient (same z-position). DSCT provides temporal resolution of approximately a quarter of the gantry rotation time for cardiac, cardio-thoracic and pediatric imaging. Successful imaging of the heart and the coronary arteries at high and variable heart rates has been demonstrated. DSCT systems can be operated at twice the spiral pitch of single source CT systems (up to pitch 3.2). The resulting high table speed is beneficial for pediatric applications and fast CT angiographic scans, e. g. of the aorta or the extremities. Operating both X-ray tubes at different tube potential (kV) enables the acquisition of dual energy data and the corresponding applications such as monoenergetic imaging and computation of material maps. Spectral separation can be improved by different filtration of the X-ray beams of both X-ray tubes. As a downside, DSCT systems have to cope with some challenges, among them the limited size of the second measurement system, and cross-scattered radiation.     

Gold nanostructures absorption capacities of various energy forms for thermal therapy applications

This mini-review has investigated the recent progress regarding gold nanostructures capacities of energy absorption for thermal therapy applications. Unselective thermal therapy of malignant and normal tissues could lead to irreversible damage to healthy tissues without effective treatment on target malignant tissues. In recent years, there has been a considerable progress in the field of cancer thermal therapy for treating target malignant tissues using nanostructures. Due to the remarkable physical properties of the gold nanoparticle, it has been considered as an exceptional element for thermal therapy techniques. Different types of gold nanoparticles have been used as energy absorbent for thermal therapy applications under several types of energy exposures. Electromagnetic, ultrasound, electric and magnetic field are examples for these energy sources. Well-known plasmonic photothermal therapy which applies electromagnetic radiation is under clinical investigation for the treatment of various medical conditions. However, there are many other techniques in this regard which should be explored.     

Re-evaluation of the RBE of 29 kV x-rays (mammography x-rays) relative to 220 kV x-rays using neoplastic transformation of human CGL1-hybrid cells

Neoplastic transformation of human CGL1-hybrid cells was examined after exposure to 29 kV x-rays (mammography x-rays) and conventional 220 kV x-rays. The study was designed to repeat, under well-defined irradiation and culture conditions, an earlier investigation by Frankenberg et al. (Radiat Res, 2002), and to assess the validity of the high RBE values of 29 kV x-rays that had been reported. The experiments with the two types of x-rays were performed simultaneously and shared the same controls. The transformation yields with both radiation qualities were fitted to the linear-quadratic dependence on absorbed dose, and a corresponding analysis was performed for the data earlier obtained by Frankenberg et al. The transformation yields in the present study exceed those in the earlier investigation substantially, and it appears that the difference reflects inadequate feeding conditions of the cell cultures in the early experiments. The standard error bands of the dose response curves are derived and are seen to be considerably more narrow in the present results. The lowest dose of the 29 kV x-rays was 1 Gy in both studies, and at this dose the RBE vs. the conventional x-rays has now been found to be 2 with a 95% confidence interval of 1.4-2.6. The previous result was about 3.2, but the 95% confidence is very broad for these data. The estimated limit at low doses is 3.4 in the present experiments with a confidence interval that extends from less than 2 to large values.     

Radiation protection of operators and patients in a hybrid Angio-MR suite

This work investigates the patient eye lens dose and x-ray scatter to the operator expected for a proposed hybrid Angio-MR concept. Two geometries were simulated for comparative assessment: a standard C-arm device for neuro-angiography applications and an innovative hybrid Angio-MR system concept, proposed by Siemens Healthineers. The latter concept is based on an over-couch x-ray tube and a detector inside an MRI system, with the aim of allowing combined, simultaneous MRI and x-ray imaging for procedures such as neurovascular interventions (including x-ray fluoroscopy and angiography imaging, 3D imaging, diffusion, and perfusion). To calculate the scattered radiation dose to the physician, Monte Carlo simulations were performed. Dose estimates of simplified models of the brain and eyes of both the patient and the physician and of the physician’s torso and legs have been calculated. A number of parameters were varied in the simulation including x-ray spectrum, field of view (FOV), x-ray tube angulation, presence of shielding material and position of the physician. Additionally, 3D dose distributions were calculated in the vertical and horizontal planes in both setups. The patient eye lens dose was also calculated using a detailed voxel phantom and measured by means of thermoluminescent dosimeters (TLDs) to obtain a more accurate estimate. Assuming the same number of x-rays and the same size of the irradiated area on the patient’s head, the results show a significant decrease in the scattered radiation to the physician for the Angio-MR system, while large increases, depending on setup, are expected to patient eye lens dose.     

Aging: A cell source limiting factor in tissue engineering

Tissue engineering has yet to reach its ideal goal, i.e. creating profitable off-the-shelf tissues and organs, designing scaffolds and three-dimensional tissue architectures that can maintain the blood supply, proper biomaterial selection, and identifying the most efficient cell source for use in cell therapy and tissue engineering. These are still the major challenges in this field. Regarding the identification of the most appropriate cell source, aging as a factor that affects both somatic and stem cells and limits their function and applications is a preventable and, at least to some extents, a reversible phenomenon. Here, we reviewed different stem cell types, namely embryonic stem cells, adult stem cells, induced pluripotent stem cells, and genetically modified stem cells, as well as their sources, i.e. autologous, allogeneic, and xenogeneic sources. Afterward, we approached aging by discussing the functional decline of aged stem cells and different intrinsic and extrinsic factors that are involved in stem cell aging including replicative senescence and Hayflick limit, autophagy, epigenetic changes, miRNAs, mTOR and AMPK pathways, and the role of mitochondria in stem cell senescence. Finally, various interventions for rejuvenation and geroprotection of stem cells are discussed. These interventions can be applied in cell therapy and tissue engineering methods to conquer aging as a limiting factor, both in original cell source and in the in vitro proliferated cells.     

Allogenic banking of dental pulp stem cells for innovative therapeutics

Medical research in regenerative medicine and cell-based therapy has brought encouraging perspectives for the use of stem cells in clinical trials. Multiple types of stem cells, from progenitors to pluripotent stem cells, have been investigated. Among these, dental pulp stem cells (DPSCs) are mesenchymal multipotent cells coming from the dental pulp, which is the soft tissue within teeth. They represent an interesting adult stem cell source because they are recovered in large amount in dental pulps with non-invasive techniques compared to other adult stem cell sources. DPSCs can be obtained from discarded teeth, especially wisdom teeth extracted for orthodontic reasons. To shift from promising preclinical results to therapeutic applications to human, DPSCs must be prepared in clinical grade lots and transformed into advanced therapy medicinal products (ATMP). As the production of patient-specific stem cells is costly and time-consuming, allogenic biobanking of clinical grade human leukocyte antigen (HLA)-typed DPSC lines provides efficient innovative therapeutic products. DPSC biobanks represent industrial and therapeutic innovations by using discarded biological tissues (dental pulps) as a source of mesenchymal stem cells to produce and store, in good manufacturing practice (GMP) conditions, DPSC therapeutic batches. In this review, we discuss about the challenges to transfer biological samples from a donor to HLA-typed DPSC therapeutic lots, following regulations, GMP guidelines and ethical principles. We also present some clinical applications, for which there is no efficient therapeutics so far, but that DPSCs-based ATMP could potentially treat.     

Intraoperative radiation therapy in uterine cervical cancer: A review

Locally advanced uterine cervical cancer continues to present a high number of pelvic relapses. Intraoperative radiation therapy (IORT) allows a precise therapeutic intensification in the surgical area in cases in which removal of the tumour recurrence is feasible. At the same time, IORT excludes the radiosensitive organs from the field of irradiation. While the first gynecological IORT took place in 1905, procedures have been limited over the years and the series are retrospective, including few patients. At the same, time recurrences are located at different pelvic areas. Both heterogeneity and the long recruiting time make it difficult to correctly interpret the published results. Despite this, we have reviewed the most relevant publications. Some institutions indicated IORT as a boost on the surgical bed of the excised tumor recurrence. In others, IORT permits an extra radiation dose after radical surgery of the primary tumor, usually in stage IIB. Most studies conclude that the addition of IORT increases the local control but probably with little impact on survival. On the other hand, there is a controversy in the indication of IORT in surgically resectable primary tumours. No clear advantage over the usual scheme of chemoradiation and brachytherapy has been detected. Randomized studies that allow a breakthrough in the conclusions are highly unlikely to be performed in this area.     

Omega-3 biotechnology: Thraustochytrids as a novel source of omega-3 oils

Thraustochytrids are large-celled marine heterokonts and classified as oleaginous microorganisms due to their production of docosahexaenoic (DHA) and eicosapentaenoic (EPA) ω-3-fatty acids. The applications of microbial DHA and EPA for human health are rapidly expanding, and a large number of clinical trials have been carried out to verify their efficacy. The development of refined isolation and identification techniques is important for the cultivation of thraustochytrids. With a high proportion of lipid biomass, thraustochytrids are also amenable to various production strategies which increase omega-3 oil output. Modifications to the existing lipid extraction methods and utilisation of sophisticated analytical instruments have increased extraction yields of DHA and EPA. Other metabolites such as enzymes, carotenoids and extracellular polysaccharides can also be obtained from these marine protists. Approaches such as the exploration for more diverse isolates having fast growth rates, metabolic engineering including gene cloning, and growing thraustochytrids on alternate low cost carbon source, will further enhance the biotechnological potential of thraustochytrids.