Biomedical subjects
E D Williams
Publications and source records attributed to E D Williams.
Quality assurance for MRI: practical experience.
The aim of this study is to propose guidelines for quality assurance (QA) in MRI, based on a comprehensive assessment of QA parameters undertaken on a busy clinical MRI scanner over the course of 1 year. QA phantoms supplied by the scanner manufacturer were used together with the Eurospin MRI phantom set. Signal-to-noise ratio (SNR) and image uniformity were measured daily from spin echo images acquired using a quadrature send-receive head coil and from a gradient echo sequence using the Helmholtz body coil. The voltage of the transmit radiofrequency (RF) amplifier was noted. Monthly measurements of slice thickness, geometric distortion, slice position, image resolution and image ghosting were acquired using the head coil. In addition, SNR was measured monthly on a selection of commonly used coils. Apart from some drift of the RF amplifier voltage, all measurements were within acceptable limits and were stable over the course of 1 year. Satisfactory measurements of SNR were possible using the simple phantom supplied with the scanner. The SNR, geometric distortion and RF amplifier voltage are simple to determine and can be measured in less than 15 min by the scanner operator, using the scanner software. Weekly recording of these parameters is recommended for busy clinical MRI scanners, as this should allow deviations from acceptable limits to be identified early. Such in-house checks can usefully be compared with the less frequent estimations performed by the service engineer. Comprehensive QA routines are discussed for systems used for quantitative measurements.
Development of a prostate cDNA microarray and statistical gene expression analysis package.
A cDNA microarray comprising 5184 different cDNAs spotted onto nylon membrane filters was developed for prostate gene expression studies. The clones used for arraying were identified by cluster analysis of > 35 000 prostate cDNA library-derived expressed sequence tags (ESTs) present in the dbEST database maintained by the National Center for Biotechnology Information. Total RNA from two cell lines, prostate line 8.4 and melanoma line UACC903, was used to make radiolabeled probe for filter hybridizations. The absolute intensity of each individual cDNA spot was determined by phosphorimager scanning and evaluated by a bioinformatics package developed specifically for analysis of cDNA microarray experimentation. Results indicated 89% of the genes showed intensity levels above background in prostate cells compared with only 28% in melanoma cells. Replicate probe preparations yielded results with correlation values ranging from r = 0.90 to 0.93 and coefficient of variation ranging from 16 to 28%. Findings indicate that among others, the keratin 5 and vimentin genes were differentially expressed between these two divergent cell lines. Follow-up northern blot analysis verified these two expression changes, thereby demonstrating the reliability of this system. We report the development of a cDNA microarray system that is sensitive and reliable, demonstrates a low degree of variability, and is capable of determining verifiable gene expression differences between two distinct human cell lines. This system will prove useful for differential gene expression analysis in prostate-derived cells and tissue.
Partial volume effects in MRI studies of multiple sclerosis.
We have estimated the accuracy of volume measurements of multiple sclerosis (MS) lesions made using magnetic resonance imaging (MRI) for lesions of comparable diameter to the image slice thickness. We used a phantom containing objects of known volume and obtained images using a range of slice thicknesses. Measurements on the phantom were used to assess a theoretical model, which was then employed to investigate the effects of image dimensions and geometry upon volume measurement accuracy. We observed measured volume to be dependent upon slice thickness. Thin slices gave the most accurate estimate of volume. As slice thickness increased relative to object diameter, the error in the volume measurement increased (to as much as 100%), the volume measured being dependent on the position of the object relative to the slice center. Using a signal intensity threshold value of 50% to outline objects gave results closest to the actual volume. As expected, a lower threshold value tended to give higher volume estimates (up to 100% larger), as did a semi-automated local edge detection technique. For accurate volume measurement, the slice thickness should be no more than a fifth of anticipated object diameter. For typical MS lesions (7 mm in diameter), this implies using a 1.5-mm slice thickness. For serial studies, a repositioning error of 1 mm could lead to differences in the volume measurement of individual lesions of up to 12% between studies for lesions of typical MS size and 5-mm slice thickness. These results emphasize the need for accurate patient repositioning, relatively thin slices, for regular quality assurance checks to ensure that pixel size and slice position are correct and stable over time, and that lesion outlining is performed in a consistent fashion. We would recommend the use of a 3D sequence with 1 mm cubic voxels for accurate measurements of MS lesions.
The effects of operator technique and bur design on temperature during osseous preparation for osteosynthesis self-tapping screws.
OBJECTIVE: Although the importance of atraumatic preparation of bone before the placement of osseointegrated implants has long been emphasized, the situation during placement of self-tapping screws has received little attention. Because the production of excessively high temperatures during osseous drilling is known to impair bony regeneration, the strict thermal criteria that are set for implant placement should theoretically be extended to trauma and orthognathic internal fixation. STUDY DESIGN: Temperatures achieved during the drilling of thawed human cadaveric bone before the insertion of osteosynthesis self-tapping screws were measured in vivo with thermocouples. Combinations of 2 drilling systems and 2 bur designs were studied. Variations in surgical technique were incorporated by altering both the force applied by the operator and the application of irrigant. Statistical significance was examined through use of a Student t test. RESULTS: One of the 2 bur designs and careful operator technique were associated with significantly smaller increases in temperature. Irrigation had the greatest effect on temperature recorded, the absence of irrigant resulting in temperatures in excess of 70 degrees C. CONCLUSIONS: These findings emphasize the importance of careful surgical technique and constant irrigation during osseous preparation if the potential for bone regeneration around osteosynthesis self-tapping screws is to be maximized.
Calcitonin receptor antibodies in the identification of osteoclasts.
Osteoclasts are the cells responsible for bone resorption, and their number and rate of formation are critical in determining bone mass. To identify and quantify osteoclasts, as well as to study their formation in bone and in osteoclastogenic cultures, osteoclast-specific cell markers are required. Only the calcitonin receptor (CTR) expression unambiguously identifies osteoclasts and distinguishes them from macrophage polykaryons. However, present autoradiographic methods for CTR detection are cumbersome and time consuming. We have developed rabbit polyclonal antibodies specific for the C-terminal intracellular domain of the mouse and rat Cla CTR. These antibodies labeled HEK-293 cells stably transfected with CTR (but not untransfected HEK-293 cells). This labeling is abrogated by preabsorbing the antibodies with the recombinant antigen. The antibodies immunostained primary mouse and rat osteoclasts as well as osteoclasts in sections of mouse bone. Osteoclasts (both mononuclear and multinucleated) formed from mouse bone marrow or spleen cells cocultured with osteoblasts in the presence of 1,25 dihydroxyvitamin D3 and prostaglandin E2 were also specifically immunostained by the CTR antibodies. Cocultures incubated under conditions that did not allow osteoclastogenesis (i.e., omission of mediators or osteoblasts, or culture for less than 4 days) were not immunostained by CTR antibodies. Autoradiographic detection of 125I-labeled salmon calcitonin combined with CTR immunohistochemistry showed that both methods labeled the same cells. A CTR polyclonal antibody and monoclonal antibody F4/80 were used in combination to show immunofluorescence labeling of murine osteoclasts and macrophage populations, respectively, in marrow/osteoblast cocultures. These results indicate that simple and rapid CTR antibody-based methods can be used to identify osteoclasts, and can be used to characterize the antigenic profile of osteoclasts by using double immunofluorescence analysis.
A comparison of two methods for measuring the signal to noise ratio on MR images.
The signal to noise ratio (SNR) is one of the important measures of the performance of a magnetic resonance imaging (MRI) system. The object of this study was to compare a single acquisition method, which estimates the noise from background pixels, with a dual acquisition method which estimates the noise from the subtraction of two sequentially acquired images. The dual acquisition method is more exact, but is slower to perform and requires image manipulation. A comparison between the two methods gave a good correlation, and a regression equation of SNRsingle = 1.1 + 0.94 SNRdual. The single acquisition method is therefore appropriate for use in a quality assurance programme, since it is quicker and simpler to perform and is a good indicator of the more exact measure.
A critical evaluation of methods for preparing pentavalent 99Tcm-DMSA.
Pentavalent 99Tcm-dimercaptosuccinic acid (99Tcm-(V)DMSA) has established uses in the detection and diagnosis of medullary thyroid carcinoma, osteosarcoma, amyloidosis and many soft tissue tumours, but is not readily available as a commercial kit. We have evaluated published methods for preparation of 99Tcm-(V)DMSA that are based on the modification of 99Tcm-(III)DMSA commercial kits. The criteria for assessment were achievement of satisfactory radiochemical purity and practical considerations regarding the ease of synthesis. Quantification of the desired product's activity was achieved via thin-layer chromatography (silica gel) with an n-butanol:acetic acid:water solvent system. The developed plates were imaged on a gamma camera and analysed using standard software. The methods investigated were designated as Birmingham, Cambridge and Canterbury: the latter two were very similar. With experience, the methods eventually produced similar radiochemical purities of > 90%. However, the Birmingham method was found to be more satisfactory in terms of its reliability and simplicity of synthesis.
High prevalence of RET/PTC rearrangements in Ukrainian and Belarussian post-Chernobyl thyroid papillary carcinomas: a strong correlation between RET/PTC3 and the solid-follicular variant.
A sharp increase in the incidence of pediatric thyroid papillary cancer was documented after the Chernobyl power plant explosion. An increased prevalence of rearrangements of the RET protooncogene (RET/PTC rearrangements) has been reported in Belarussian post-Chernobyl papillary carcinomas arising between 1990 and 1995. We analyzed 67 post-Chernobyl pediatric papillary carcinomas arising in 1995-1997 for RET/PTC activation: 28 were from Ukraine and 39 were from Belarus. The study, conducted by a combined immunohistochemistry and RT-PCR approach, demonstrated a high frequency (60.7% of the Ukrainian and 51.3% of the Belarussian cases) of RET/PTC activation. A strong correlation was observed between the solid-follicular subtype of papillary carcinoma and the RET/PTC3 isoform: 19 of the 24 RET/PTC-positive solid-follicular carcinomas harbored a RET/PTC3 rearrangement, whereas only 5 had a RET/PTC1 rearrangement. Taken together these results support the concept that RET/PTC activation plays a central role in the pathogenesis of thyroid papillary carcinomas in both Ukraine and Belarus after the Chernobyl accident.
Thyroid stimulating hormone (TSH)-associated follicular hypertrophy and hyperplasia as a mechanism of thyroid carcinogenesis in mice and rats.
Explore the source record for details and available documents.
Solitary, multiple, and familial oxyphil tumours of the thyroid gland.
A study of 169 oxyphil tumours of the thyroid, correlating histological features with age, sex, thyroiditis, and available clinical history, has shown two significant findings relevant to pathogenesis, diagnosis, and treatment. Oxyphil tumours with a papillary architecture can be divided into papillary carcinomas with secondary oxyphilia and true oxyphil tumours. The tumours of the former group are typically unencapsulated, invasive, show nuclei characteristic of papillary carcinoma generally, are associated with thyroiditis, and frequently show psammoma bodies. The tumours of the latter group are typically encapsulated, with nuclei similar to those found in other oxyphil tumours; usually lack psammoma bodies; and some show capsular and vascular invasion. Papillary architecture alone is not thought to justify a diagnosis of malignancy in this group of tumours, and further study with follow-up is needed to determine the appropriate treatment. It is suggested that oxyphilia in the first group is secondary to the accompanying thyroiditis, comparable to the oxyphilia in follicular cells in Hashimoto's thyroiditis, while the oxyphilia in the latter group, as in other oxyphil tumours, is due to a somatic mutation leading to an increase in mitochondrial number. About 20 per cent of all cases showed multiple oxyphil tumours. These were more frequently female, more often associated with thyroiditis, younger, and less often malignant than solitary tumours. The occurrence of multiple tumours of the same uncommon histological type in these patients is compatible with a germline mutation conferring a liability to oxyphil follicular cell tumours. This is supported by the occurrence of oxyphil tumours in families; the present series of cases included two mother-daughter pairs and similar examples have been reported, including multiple oxyphil tumours in identical twins. Further studies are needed to identify the gene involved and any co-operating genes. Oxyphil carcinoma in patients with multiple oxyphil tumours occurred at a mean age 10 years greater than that for multiple adenomas, suggesting that the carcinomas arose by progression from adenomas. It is important to complete thyroidectomy in a case with multiple oxyphil tumours if there is evidence of the presence of lesions in both lobes. It is necessary to consider the differences between primary and secondary oxyphilia, and between sporadic and multiple or familial oxyphil tumours in future studies of these lesions.
Thyroid cancer in patients with familial adenomatous polyposis.
The association between thyroid cancer and familial adenomatous polyposis (FAP), albeit rare, is well known. It has been suggested that the thyroid tumors have unique histologic characteristics and may be follicular in origin. Because of their rarity, treatment and long-term prognosis are uncertain. Twelve such patients (prevalence 399/100,000) seen during 1949-1995 were retrospectively reviewed. Histology was independently re-reviewed by two pathologists. There were 11 female patients (two sisters) and 1 male patient, with a mean age of 28 years (range 15-61 years). Eight patients (66%) had multicentric tumors and five (42%) bilateral disease. Average tumor diameter was 1.8 cm (range 0.2-5.0 cm). Regional nodal metastases were present in two patients. All 12 thyroid cancers in this series were papillary. The one male patient demonstrated "typical" histology with variable papillary and follicular architecture, whereas the 11 female patients had tumors with unusual histology as described by Harach. Five patients (41%) were treated by total thyroidectomy, five with near-total thyroidectomy, and two with lobectomy alone. Mean follow-up was 142 months (range 7 months to 30 years). Regional recurrent disease occurred in two patients, one of whom died of the disease. The 5- and 20-year survivals were 90% and 77%, respectively. The results indicated that all tumors in this study were papillary, although atypical histology was encountered in 91%. The mean age (28 years) is younger than that of patients with sporadic disease. Multicentricity and bilateral disease are common. In view of this finding, total thyroidectomy should be strongly considered. Long-term prognosis is excellent. The finding of unusual histology in a young patient with papillary thyroid carcinoma should arouse the suspicion of FAP.
Increased stem cell somatic mutation in the non-neoplastic colorectal mucosa of patients with familial adenomatous polyposis.
Colorectal tumorigenesis in familial adenomatous polyposis (FAP) results from somatic mutation of either the normal APC allele or another growth control gene in epithelial cells bearing a germline APC defect. The rate at which tumors develop is therefore dependent on the somatic mutation frequency; it is not known whether this is normal or elevated in FAP. We aimed to quantify stem cell somatic mutation in FAP, comparing it with hereditary nonpolyposis colorectal cancer (HNPCC) and Crohn's disease (CD). Stem cell somatic mutation frequency was studied in 47 FAP patients, 5 HNPCC patients, and 13 CD patients, all younger than 49 years, by quantifying crypt-restricted loss of O-acetyltransferase activity in sections of morphologically normal colonic mucosa from individuals heterozygous for this monogenically inherited polymorphism. Median stem cell somatic mutation frequency was significantly higher in FAP than HNPCC (4.2 x 10(-4) v 1.4 x 10(-4), Mann-Whitney U, P < .02). The level in CD (4.0 x 10(-4)) was similar to FAP. Mutated crypts occurred in groups more frequently in FAP (22%) than HNPCC (12%) or CD (10%), suggesting an increase in stem cell division associated with crypt fission in FAP. We conclude that stem cell somatic mutation frequency is raised in non-neoplastic colorectal mucosa in FAP. This is probably related to increased stem cell proliferation and contributes to the high rate of tumor formation in this condition.
Mutation of the RET proto-oncogene is correlated with RET immunostaining in subpopulations of cells in sporadic medullary thyroid carcinoma.
Mutations in the RET proto-oncogene, which encodes a receptor tyrosine kinase, are associated with the pathogenesis of medullary thyroid carcinoma (MTC). Somatic mutations in RET, predominantly at codon 918, and very rarely at codon 883, have been found in a proportion of sporadic MTC. We have previously shown that approximately 80% of sporadic MTCs had at least one subpopulation with a somatic RET mutation. Uneven distribution of somatic mutation within a single tumor or among metastases from a single individual was notable. In the present study, we sought to correlate RET expression, as demonstrated by RET immunohistochemistry, with mutation status in sporadic MTC for each tumor. Seventy evaluable subpopulations, belonging to 28 unrelated sporadic cases, comprising primary MTC and metastases, were immunostained with two different polyclonal antibodies raised against the C-terminus of RET. The regional presence of codon 918 or 883 seemed to coincide with increased RET immunopositivity in at least 62 of 70 (89%, P < 0.000001) tumor subpopulations. The reasons for this concordance are not entirely clear but could be related to either RNA or protein stability. Preliminary studies have suggested that the presence of somatic codon 918 mutation in MTC has a prognostic significance. If these preliminary results prove true, then given our data, we can further explore the feasibility of RET immunocytochemistry as a rapid assessment for the presence of somatic codon 918 for molecular diagnostic and prognostic purposes.
Effect of antagonism of the parathyroid hormone (PTH)/PTH-related protein receptor on decidualization in rat uterus.
Parathyroid hormone-related protein (PTHrP) was detected at 32.8 +/- 3.9 pmol 1-1 in uterine luminal fluid from immature rats treated with oestradiol. As mRNA encoding PTHrP has previously been localized to implantation sites in pregnant rats, the role of luminal PTHrP during pregnancy was explored. Infusion of a parathyroid hormone (PTH)/PTHrP receptor antagonist, [Asn10,Leu11]PTHrP(7-34) amide, into the uterine lumen during pregnancy in rats resulted in excessive decidualization. This effect was also observed after intrauterine infusion of a monoclonal antibody raised against PTHrP. The effect of infusion of PTH/PTHrP receptor antagonist was dependent upon successful implantation, was dose-dependent and confined to the treated horn. A decrease in the number of apoptotic decidual cells in antagonist-infused uterine horns compared with vehicle or non-infused horns was detected immunohistochemically at day 13 of pregnancy, and this decrease is likely to contribute to the 'over-decidualization' observed. In pseudopregnant rats, infusion of PTH/PTHrP receptor antagonist into the uterine lumen resulted in an increase in uterine wet weight of the infused horn compared with the non-infused horn, indicating a direct effect on deciduoma formation. Thus, activation of the PTH/PTHrP receptor by locally produced PTHrP appears to be crucial for normal decidualization during pregnancy in rats.
Somatic mutation of the glucose-6-phosphate dehydrogenase (g6pd) gene in colonic stem cells and crypt restricted loss of G6PD activity.
The study of somatic mutation frequency, particularly stem cell somatic mutation, is important to the understanding of mechanisms of carcinogenesis. The models currently in use for studies in stem cell tissues such as the colon infer the presence of stem cell somatic mutation from alteration in enzyme function, when this has shown to be mutagen dose dependent, restricted to the unit of clonal architecture, and persistent. The present study identifies and characterises somatic mutations in the g6pd gene in individual mouse colonic crypts showing histochemically demonstrable loss of G6PD activity. Microdissection of single crypts, showing either normal or low G6PD activity by histochemistry was performed in mice treated with ethylnitrosourea (ENU), and the presence of point mutations sought by PCR and direct sequencing. Because of the limitation of the small amount of partially degraded (due to fixation) DNA available from each crypt, only about 20% of the coding region of the g6pd gene could be sequenced. Despite this, somatic mutations were identified in 3 of the 9 crypts analysed which showed loss of G6PD activity, but in none of the crypts with normal activity. Each of the mutations identified would be predicted to lead to a decrease in enzyme activity. We conclude that we have confirmed that the crypt restricted loss of G6PD activity is indeed due to stem cell somatic mutation in the g6pd gene, and suggest that the G6PD model can be used as a paradigm for other models where somatic mutation is inferred from a change in histochemically identifiable gene expression.
A glucose-6-phosphate dehydrogenase (G6PD) splice site consensus sequence mutation associated with G6PD enzyme deficiency.
A glucose-6-phosphate dehydrogenase (G6PD) deficient strain of mouse (GPDX) which was developed using the ethylating agent ethylnitrosourea (ENU) has been used to study clonality in epithelial tissues. While the biochemical defect has been quantified, the genetic basis of the deficiency is unknown. The G6PD gene is composed of 13 exons. Exon 1 is not translated, and the ATG start site is near the 5' end of exon 2. Direct sequencing of the exonic regions of the gene from GPDX, C3H, 101, C57BL/6 and BALB/c mice was carried out. The coding region, in which (with a single exception) all mutations found to cause G6PD deficiency in man are situated, showed identical sequences in three of the four strains studied (101 coding region sequence was not examined). However, the G6PD gene in the GPDX mouse showed a single base difference from the other four strains and from the published mouse G6PD sequence (BALB/c) in the 5' splice site consensus sequence at the 3' end of exon 1, part of the untranslated region. The difference was confirmed in four different GPDX mice. This mutation was of the type (A to T transversion) that is known to be induced by ENU; its effect is likely to be exerted through a defect in transcription, splicing or translation, leading to a reduction in protein levels. By Western blot we have found a marked decrease in the G6PD protein levels in the GPDX mouse, with the C3H X GPDX heterozygote showing a lesser decrease. Recently, an increasing number of mutations in the untranslated regions of genes have been found which have effects on protein levels. We believe that the reduced enzyme activity in the GPDX mouse is due to the mutation in the 5' untranslated region (UTR), and that similar mutations may be relevant in other inherited conditions.
Papillary carcinoma of the thyroid with stromal lymphoid infiltrate.
Explore the source record for details and available documents.