Gender differences in distal femoral morphology and the role of gender specific implants in total knee replacement: a prospective clinical study.

Gender differences in distal femoral morphology may affect femoral component fit using a standard range of prostheses. The clinical relevance of this is controversial. Standardised measurements were taken from the distal femora of 50 males and 50 females during total knee replacement (TKR). Corresponding measurements were taken from the respective gender specific and standard femoral components. No demographic differences were noted. Significant differences in both frequency and magnitude existed in the medial-lateral femoral component overhang between the sexes. In females, standard implants overhung at the anterior flange width (AFW) by >2mm in 24/50 (48%) and by >3mm in 17/50 (34%) (p<0.001). Also at the anterior medial-lateral width (MLA) 29/50 (58%) overhung by >2mm and 24/50 (48%) by >3mm (p<0.001). In males, standard implants overhung by >2mm in 1/50 (2%). In females, gender specific implants overhung by >2mm in 3/50 (6%). Females had a mean aspect ratio of 1.02 (0.82 to 1.35) and men 0.98 (0.79 to 1.19). Femoral component overhang can occur in females undergoing TKR and a gender specific implant would reduce the potential for medial-lateral overhang. Long term studies are awaited to quantify the clinical implications of overhang.

Minimal contribution of desmethyl-gefitinib, the major human plasma metabolite of gefitinib, to epidermal growth factor receptor (EGFR)-mediated tumour growth inhibition.

Desmethyl-gefitinib is a major metabolite of gefitinib observed in human plasma at concentrations similar to those of gefitinib. The epidermal growth factor receptor (EGFR)-related inhibitory effects of gefitinib and desmethyl-gefitinib have been compared both in vitro, using enzyme kinase assays and tumour cell growth inhibition, and in vivo by assessment of tumour xenografts growth inhibition in the mouse. Both gefitinib (IC(50) = 0.022 microM) and its desmethyl metabolite (0.036 microM) inhibited subcellular EGFR tyrosine kinase activity with a similar potency and selectivity. However, desmethyl-gefitinib (IC(50) = 0.76 microM) was 15 times less active than gefitinib (0.049 microM) against EGF-stimulated KB cell growth in a whole cell assay. Following a preliminary pharmacokinetic study to compare apparent oral bioavailability, gefitinib (75 mg kg(-1)) and desmethyl-gefitinib (150 mg kg(-1)) were administered orally for 15 days to female nude mice bearing LoVo tumour xenografts. Tumour concentrations of gefitinib (AUC = 300 microg h g(-1)) were much higher than those of desmethyl-gefitinib (44.3 microg h g(-1)), although plasma concentrations of gefitinib (48.4 microg h ml(-1)) and desmethyl-gefitinib (39.0 microg h ml(-1)) were quite similar at these dose levels. Gefitinib produced significant tumour growth inhibition throughout the course of the study ultimately resulting in a 50% decrease (compared with controls) by day 15. In contrast, although present at comparable plasma levels, desmethyl-gefitinib had little effect on tumour growth and is, therefore, considered unlikely to contribute significantly to the therapeutic activity of gefitinib in the clinical situation.

The prevalence of bacteriuria in older institutionalized patients.

We set out to determine the prevalence of bacteriuria in elderly patients institutionalised in 10 homes in Northumberland. Single mid-stream urine (MSU) specimens were collected from 161 asymptomatic patients aged 64-101 years. Thirty-four (21%) samples had > 100 white blood cells (WBC/microliter) on microscopy as well as a significant bacteriuria of > 10(8) bacteria/l, being suggestive of a true urinary tract infection. A total of 115(71%) samples had < 100 WBC. Seventy-five (46%) samples could be described as 'contaminated' or 'infection unlikely' but of these only 31 (19.2% of all samples) were reported as 'contaminated' by the laboratory. This study confirms previous work in America to indicate the high level of bacteriuria in institutionalised elderly patients as well as the very high number (46%) who may show bacteriuria but no rise in the number of WBC suggesting contamination only. Nurses and GPs must be very cautious in interpreting MSU results from such patients.

Detection of PCR products using self-probing amplicons and fluorescence.

Molecular diagnostics is progressing from low-throughput, heterogeneous, mostly manual technologies to higher throughput, closed-tube, and automated methods. Fluorescence is the favored signaling technology for such assays, and a number of techniques rely on energy transfer between a fluorophore and a proximal quencher molecule. In these methods, dual-labeled probes hybridize to an amplicon and changes in the quenching of the fluorophore are detected. We describe a new technology that is simple to use, gives highly specific information, and avoids the major difficulties of the alternative methods. It uses a primer with an integral tail that is used to probe an extension product of the primer. The probing of a target sequence is thereby converted into a unimolecular event, which has substantial benefits in terms of kinetics, thermodynamics, assay design, and probe reliability.

Appearances can be deceptive: an APC 1893del4 mutation with unusual properities. Mutations in brief no. 171. Online.

During a systematic search for germ-line APC mutations causative of familial adenomatous polyposis, we discovered what appeared to be an insertion mutation while simply checking exon 14PCR products by agarose gel electrophoresis (AGE). On AGE, exon 14PCR product from the known affected member of this family gave two bands: one of normal length, the other retarded on the gel equivalent to an increase in length of some 20-25 bp. Direct sequencing of DNA purified from the two bands gave identical results, and was consistent with amplification from the same two alleles: one wild-type, and the other having an 1893del4 mutation. This suggested that the normal length band on AGE consisted of DNA homoduplexes (normal:normal and mutant:mutant) and the retarded band consisted of DNA heteroduplexes (normal:mutant and mutant:normal). This hypothesis was tested by subjecting purified material from each of the two bands alone to a single cycle of heat denaturation and annealing, which showed that either band was equally capable of regenerating both bands. Because the anomalous migration of the heteroduplexes is observed in the presence of ethidium bromide, it implies that they have a cruciform of cruciform-like structure. This case illustrates the necessity to be aware of anomalous DNA migration and always sequence all putative mutations.

APC mutations in familial adenomatous polyposis families in the Northwest of England.

We have investigated a series of FAP patients in the Northwest of England in order to identify and characterise the specific APC mutations. Using SSCP, we found 27 mutations in a total of 50 families investigated. The mutations were predominantly frameshift or nonsense mutations and there were two splice site changes. We have described two patients with severe Gardner's phenotype from different ethnic backgrounds who share the same mutation at codon 1537. Although the frequency of the most common mutation appears low, it is not dissimilar to that reported by other groups.

Expression of cytokine messenger RNA after heart transplantation: relationship with rejection and serum cytokines.

Different groups of cytokines may initiate or inhibit the rejection process. We used the polymerase chain reaction to study the expression of cytokine mRNA for interleukin (IL)-2, -4, -6 and -10, tumor necrosis factor-alpha, and interferon-gamma in 187 biopsy specimens from 24 human cardiac transplant recipients 5-555 days after transplantation. Cytokine levels in the serum were also measured. Cytokine mRNA was detected in 38.5% of biopsy specimens. IL-10 mRNA was detected more frequently with mild or absent rejection (11.6% in grades 0 and 1 - vs. 1.4% in grades 2 and 3, P=0.01). Up to 90 days after transplantation, IL-2 mRNA was detected more frequently with moderate rejection (13% in grades 2 and 3 vs. 0% in grades 0 and 1, P=0.076), and IL-4 mRNA was detected more frequently with mild or absent rejection (16% in grades 0 and 1 - vs. 0% in grades 2 and 3, P=0.061). More than 90 days after transplantation, IL-2 mRNA was detected more frequently with mild or absent rejection (10% in grades 0 and 1 vs. 0% in grades 2 and 3, P=0.078). Serum IL-4 levels corresponding to biopsy specimens positive for IL-4 mRNA were higher than those corresponding to specimens negative for IL-4 mRNA (59 pg/ml vs. 32 pg/ml medians, P=0.028). Our results suggest that IL-10 and possibly IL-4 (T helper 2 cytokines) may suppress graft rejection, whereas IL-2 (T helper 1 cytokine) may promote cellular rejection. In addition, cytokine profiles may change with length of time after transplantation. The association of elevated serum levels of IL-4 with increased expression of intragraft IL-4 mRNA may suggest release of this cytokine from the graft into the circulation.

Severe Gardner syndrome in families with mutations restricted to a specific region of the APC gene.

Familial adenomatous polyposis (FAP) is associated with a number of extraintestinal manifestations, which include osteomas, epidermoid cysts, and desmoid tumors, often referred to as "Gardner syndrome." Recent studies have suggested that some of the phenotypic features of FAP are dependent on the position of the mutation within the APC gene. In particular, the correlation between congenital hypertrophy of the retinal pigment epithelium (CHRPE) and APC genotype indicates that affected families may be divided into distinct groups. We have investigated the association between the dentoosseous features of GS on dental panoramic radiographs (DPRs) and APC genotype in a regional cohort of FAP families. DPRs were performed on 84 affected individuals from 36 families, and the dento-osseous features of FAP were quantified by a weighted scoring system. Significant DPR abnormalities were present in 69% of affected individuals. The APC gene mutation was identified in 27 of these families, and for statistical analysis these were subdivided into three groups. Group 1 comprised 18 affected individuals from seven families with mutations 5' of exon 9; these families (except one) did not express CHRPE. Groups 2 comprised 38 individuals from 16 families with mutations between exon 9 and codon 1444, all of whom expressed CHRPE. Group 3 comprised 11 individuals from four families with mutations 3' of codon 1444, none of whom expressed CHRPE. Families with mutations 3' of codon 1444 had significantly more lesions on DPRs (P < .001) and appeared to have a higher incidence of desmoid tumors. These results suggest that the severity of some of the features of Gardner syndrome may correlate with genotype in FAP.

Non-penetrance and late appearance of polyps in families with familial adenomatous polyposis.

One case of non-penetrance of the familial adenomatous polyposis (FAP) gene at 59 years of age and late onset of polyps on endoscopy and biopsy in this and two other families is described. Screening protocols should include dental screening as well as indirect ophthalmoscopy and endoscopy to detect minimal manifestations of the gene. In the absence of a specific DNA predictive test, bowel screening should continue well beyond 30 years of age.

Human neuroblastoma cells treated with aluminium express an epitope associated with Alzheimer's disease neurofibrillary tangles.

A number of studies have implicated aluminium as a possible factor in the pathogenesis of Alzheimer's disease (AD). Following an examination of the uptake of aluminium by human neuroblastoma cells in culture, treated with a range of concentrations of aluminium complexed with ethylene-diaminetetra-acetic acid (EDTA), we have now carried out an immunocytochemical study. Using an antibody to phosphorylated tau protein, which reacts specifically with AD neurofibrillary tangles (NFT), we have found that after treatment periods of 16 days to 8 weeks with aluminium-EDTA, the cells show positive staining with this antibody. No such reaction was detected in cells grown in medium alone, nor in aluminium-EDTA-treated cells subjected to the same immunocytochemical procedure but without added primary antibody. Cells grown in medium plus EDTA, which contains a low level of aluminium contamination, showed a slight reaction. Our system may provide a suitable model for studying the early changes which lead to NFT formation.

DNA single-strand breaks in adult and embryonic avian erythrocytes.

We have investigated the possibility that the reactivation rate of adult avian erythrocytes, which is slower than that of embryonic erythrocytes, after fusion with metabolically active cells, is due to a greater number of single-strand breaks (ssb) in the DNA of the former. We have assayed ssb by measuring the template activity of the erythrocyte nuclei for added Escherichia coli DNA polymerase. We have found that differences in the numbers of ssb within polymerase-accessible regions between adult and embryonic cells are within experimental error. We conclude that, unless very localized clusters of damage exist within the DNA (which would not be detectable by this or other techniques), the difference in reactivation rate is not attributable to differences in ssb numbers.

Uptake of aluminium by human neuroblastoma cells.

We have treated human neuroblastoma cells with various complexes of aluminium, over a range of concentrations, and have measured the amount of aluminium entering the cells after one week, using atomic absorption spectroscopy. We have found that the cells incorporate much higher levels of the element after treatment with aluminium-EDTA than with aluminium-citrate or aluminium-maltol. With aluminium-EDTA, initially the uptake increases with increasing concentration in the medium but eventually reaches a plateau; the concentration within the cells is then higher than that in the medium. No obvious change in morphology of the cells occurs after treatment. The number of cells present after a week's treatment with aluminium-EDTA is lower than that of untreated cells but does not vary over a wide range of aluminium concentrations.

Effect of N-methyl-N-nitrosourea on avian erythrocyte nuclei reactivation.

Avian erythrocytes are terminally differentiated cells but they can be reactivated by fusion with actively metabolising cells. We have examined the effects of treating the erythrocytes with a carcinogenic methylating agent, N-methyl-N-Nitrosourea (MNU), on the process of reactivation of adult and embryonic nuclei. We have found that the rate of nuclear enlargement is slightly lower in nuclei from MNU-treated cells than from control cells and that there is a marked delay of about 24 h in the appearance of nucleoli in both adult and embryonic cells. This is not due to an effect of MNU on ribosomal (r)DNA: the number of rDNA genes appears to be similar in treated and control cells. Also, the number of rDNA genes appears to be similar in adult and embryonic cells and in unreactivated and reactivated embryonic nuclei: thus, differences in reactivation rate between adult and embryonic cells, observed by us and others, can not be attributed to a gross difference in their ribosomal DNA contents, and reappearance of nucleoli on reactivation can not be due to an amplification of rDNA (i.e., to recovery of such genes if lost on terminal differentiation). We suggest that MNU, although a monofunctional alkylating agent, may cause increased association--possibly cross-linkage--between DNA and protein in chromatin, thereby hindering access of host cell reactivating proteins, especially to the nucleolar regions.

Chromatin characteristics of carcinogen-treated avian erythrocyte nuclei and effects of reactivation.

We have examined nucleosome repeat length and accessibility of erythrocyte nuclei to micrococcal nuclease before and after reactivation of the erythrocytes by fusion with A9 cells, and also after treatment with the alkylating agent N-methyl-N-nitrosourea (MNU). We have found that the repeat length is higher for adult than for embryonic nuclei (227 and 202 base pairs, respectively) and after reactivation the value decreases (to 183 +/- 2 bp and 172 +/- 4 bp, respectively). Values for MNU-treated, reactivated nuclei are slightly (but probably not significantly) higher (188 bp and 176 +/- 1 bp) than for corresponding untreated nuclei. However, the rate of digestion is lower in MNU-treated erythrocytes, both unreactivated and reactivated, than in the corresponding untreated nuclei.