Genetic reporter system for oncogenic Igh-Myc translocations in mice.

The Myc-deregulating chromosomal T(12;15)(Igh-Myc) translocation, the hallmark mutation of inflammation- and interleukin 6-dependent mouse plasmacytoma (PCT), is the premier model of cancer-associated chromosomal translocations because it is the only translocation in mice that occurs spontaneously (B lymphocyte lineage) and with predictably high incidence (approximately 85% of PCT), and has a direct counterpart in humans: Burkitt lymphoma t(8;14)(q24;q32) translocation. Here, we report on the development of a genetic system for the detection of T(12;15)(Igh-Myc) translocations in plasma cells of a mouse strain in which an enhanced green fluorescent protein (GFP)-encoding reporter gene has been targeted to Myc. Four of the PCTs that developed in the newly generated translocation reporter mice, designated iGFP(5'Myc), expressed GFP consequent to naturally occurring T(12;15) translocation. GFP expression did not interfere with tumor development or the deregulation of Myc on derivative 12 of translocation, der (12), because the reporter gene was allocated to the reciprocal product of translocation, der (15). Although the described reporter gene approach requires refinement before T(12;15) translocations can be quantitatively detected in vivo, including in B lymphocyte lineage cells that have not yet completed malignant transformation, our findings provide proof of principle that reporter gene tagging of oncogenes in gene-targeted mice can be used to elucidate unresolved questions on the occurrence, distribution and trafficking of cells that have acquired cancer-causing chromosomal translocations of great relevance for humans.

Bullying in Pattani primary schools in southern Thailand.

BACKGROUND: The purpose of this research was to examine risk factors that affect the likelihood of students in Pattani primary schools bullying other children. Risk factors investigated include school rural/urban location, age, gender, religion, family physical abuse and preference of cartoon type. Identifying students who are at high risk of bullying could assist educational authorities to introduce better strategies for reducing the problem. METHODS: A total of 1440 students at public and private primary schools in Pattani province were interviewed to collect relevant data. Pearson's chi-square test was used to assess the associations between the likelihood of bullying and possible risk factors. Logistic regression was used to investigate independent associations between the predictor variables and the outcome. RESULTS: We found that 32.9% reported that they had (ever) bullied other children. Bullying was significant associated with age (odds ratio 1.56 for 11+ years, 95% confidence intervals 1.23, 2.18) and family physical abuse (odds ratio 4.50, 95% confidence intervals 3.40, 5.89). In addition, Those students who preferred action cartoons tended to bully others 1.87 times more than those who preferred watching comedy cartoons. CONCLUSIONS: There are significant differences in our population in rates of bullying others that vary according to age, preferred cartoon type and whether or not family (parental) physical abuse has been witnessed. The factor 'preference for cartoon type', not examined in previous research, remained significant after multivariable adjustment. Although there is an association, the cause of this is not clear but merits further examination.

Plasma cell tumour progression in iMycEmu gene-insertion mice.

The authors have recently reported that gene-targeted iMyc(Emu) mice that carry a His(6)-tagged mouse Myc cDNA, Myc(His), just 5' of the immunoglobulin heavy-chain enhancer, Emu, are prone to 'spontaneous' neoplasms of the B-lymphocyte lineage. The present study has used histological, immunohistochemical, and molecular genetic methods to investigate a subset of these neoplasms referred to as extraosseous plasmacytomas (PCTs). It is shown that 20.8% (20/96) of tumour-bearing iMyc(Emu) mice on a mixed genetic background of segregating C57BL/6 and 129/SvJ alleles develop PCT by 500 days. The Myc(His)-induced PCTs produced monoclonal immunoglobulin and developed in the gut-associated lymphoid tissue (GALT), particularly the mesenteric node and Peyer's patches. The PCTs overexpressed Myc(His), at the expense of normal Myc, and exhibited gene expression changes on cDNA macroarrays that were consistent with Myc(His)-driven neoplasia. Surprisingly, in one of three PCT-derived cell lines, Myc(His) was 'replaced' by a naturally occurring T(12;15) translocation, which changed the mode of Myc deregulation from gene insertion (Myc(His) transgene) to chromosomal translocation (juxtaposition of normal Myc to the immunoglobulin heavy-chain locus Igh). These findings provide evidence that recreation of the mouse PCT-associated T(12;15)(Igh(Emu)-Myc) translocation by gene insertion in mice results in the predictable development of PCTs in approximately one-fifth of the tumour-bearing mice. Myc(His)-driven PCTs recapitulate aspects of human plasma cell neoplasms, for which relatively few models exist in mice. For example, PCT development in the iMyc(Emu) mice may provide a good system to study the mechanism by which human MYC facilitates the progression of plasma cell myeloma (multiple myeloma) in humans.

Spectral karyotyping and fluorescence in situ hybridization detect novel chromosomal aberrations, a recurring involvement of chromosome 21 and amplification of the MYC oncogene in acute myeloid leukaemia M2.

Recurring chromosomal aberrations are of aetiological, diagnostic, prognostic and therapeutic importance in acute myeloid leukaemia (AML). However, aberrations are detected in only two thirds of AML cases at diagnosis and recurrent balanced translocations in only 50%. Spectral karyotyping (SKY) enables simultaneous visualization of all human chromosomes in different colours, facilitating the comprehensive evaluation of chromosomal abnormalities. Therefore, SKY was used to characterize 37 cases of newly diagnosed AML-M2, previously analysed using G-banding. In 15/23 patients it was possible to obtain metaphases from viably frozen cells; in 22 additional cases, fixed-cell suspensions were used. Of the 70 chromosomal aberrations identified by SKY, 30 aberrations were detected for the first time, 18 aberrations were redefined and 22 were confirmed. SKY detected two reciprocal translocations, t(X;3) and t(11;19). In five cases, eight structural aberrations resulted in partial gains of chromosome 21, six of which were undetected by G-banding. In 4/5 cases, these resulted in copy number increases for AML1. Amplification of MYC was detected in three cases. Using SKY and FISH, clonal aberrations were identified in 5/18 cases with a presumed normal karyotype; 3/5 aberrations were of known unfavourable prognostic significance. Karyotypes were entered into a custom-designed SKY database, which will be integrated with other cytogenetic and genomic databases.

Novel molecular cytogenetic techniques for identifying complex chromosomal rearrangements: technology and applications in molecular medicine.

Molecular cytogenetic techniques that are based on fluorescence in situ hybridisation (FISH) have become invaluable tools for the diagnosis and identification of the numerous chromosomal aberrations that are associated with neoplastic disease, including both haematological malignancies and solid tumours. FISH can be used to identify chromosomal rearrangements, by detecting specific DNA sequences with fluorescently labelled DNA probes. The technique of comparative genomic hybridisation (CGH) involves two-colour FISH. It can be used to establish ratios of fluorescence intensity values between tumour DNA and control DNA along normal reference metaphase chromosomes, and thereby to detect DNA copy-number changes such as gains and losses of specific chromosomal regions and gene amplifications. Spectral karyotyping (SKY) is a novel molecular cytogenetic method for characterising numerical and structural chromosomal aberrations. SKY involves the simultaneous hybridisation of 24 differentially labelled chromosome-painting probes, followed by spectral imaging and chromosome classification, and produces a colour karyotype of the entire genome. The use of SKY has contributed significantly to the identification of chromosomal anomalies that are associated with constitutional and cancer cytogenetics, and has revealed many aberrations that go undetected by traditional banding techniques. In this article, we have reviewed these new molecular cytogenetic techniques and described their various applications in molecular medicine.

Cytogenetic analysis of the bipotential murine pre-B cell lymphoma, P388, and its derivative macrophage-like tumor, P388D1, using SKY and CGH.

Spectral karyotyping (SKY) and comparative genomic hybridization (CGH) were used to elucidate the divergent cytogenetic make-up of the prototypical bilineage lymphoblastic pre-B lymphoma, P388, and its progenitor macrophage-like tumor, P388D1. P388 was found to be diploid and genomically stable. P388D1 was triploid, highly unstable and characterized by numerous marker chromosomes (Chrs) and composite rearrangements. The karyotype of P388D1 was so complex that its clonal relatedness to P388 would have remained questionable without confirmation by molecular analysis of the clonotypic immunoglobulin heavy-chain and light-chain gene recombinations that coexisted in both tumors. The intrinsic instability of the P388D1 genome was indicated by the observation that only four out of 42 aberrations uncovered by SKY (in a total of 27 metaphases) occurred consistently (100% incidence), whereas 27 changes occurred non-randomly (27 to 96% incidence) and 11 alterations randomly (4 to 11% incidence). Persistent cytogenetic instability was also observed in P388 'macrophages' after phorbol ester- and ionomycin-induced conversion in vitro of P388 lymphoma cells. The 'cytogenetic noise' in these cells was manifested by a multiplicity of sporadic chromosomal aberrations; ie 25 distinct changes were identified by SKY in 40 metaphases. The results in P388D1 and P388 'macrophages' were interpreted to indicate that the myeloid differentiation program in the bipotential pre-B cell lymphoma P388 is invariably characterized by karyotypic instability. The study presented here demonstrates the power of the combined SKY and CGH approach to resolve complicated karyotypes of important and widely used mouse tumors.

Histamine, xanthine oxidase generated oxygen-derived free radicals and Helicobacter pylori in gastroduodenal inflammation and ulceration.

OBJECTIVE: The relationship between Helicobacter pylori (H. pylori), xanthine oxidase (XO)-induced oxygen derived free radicals (ODFR) and histamine in the induction of human gastroduodenal disorders was investigated. PATIENTS AND METHODS: Histamine concentration, XO and xanthine dehydrogenase (XD) activities were measured in endoscopically obtained biopsies from 66 symptomatic patients. RESULTS: H. pylori infection was associated with lower oxyntic and duodenal histamine in 'normal' controls (group N) (p < 0.002 and p < 0.05, respectively). Patients with gastroduodenal disease tended to have reduced mucosal concentration of histamine, but comparing H. pylori positive and negative patients, infection did not lead to a further fall in histamine concentration. H. pylori positive duodenal ulcer (DU) patients tended to have higher XO activity than group N (p = 0.051) and had a significantly lowered activity of XD, the precursor of XO (p' < 0.05). Histamine concentration at the ulcer-edge was lower while XO activity was higher than in the distant normal mucosa (p < 0.05, respectively). Gastritis (group GL) with H. pylori also had lower XD than H. pylori positive group N (p' < 0.025) but no corresponding rise in XO activity. In group N, duodenal mucosal histamine and XD activity were inversely related (Rs = -0.51, p < 0.025). CONCLUSIONS: These findings support the hypothesis that histamine, xanthine oxidase related ODFR, and H. pylori may be closely associated in the manifestations of chronic duodenal ulcer.

Ultrasonography and developmental dysplasia of the hip: a cost-benefit analysis of three delivery systems.

The role of ultrasound (US) in the diagnosis and management of infants with developmental dysplasia of the hips (DDH) is becoming widely accepted. In our community, there exist three delivery systems for US-DDH: the radiology based, the combined radiology/orthopaedic based, and the orthopaedic office based. This study reviewed the costs and benefits of each delivery system and found that once expertise had been gained and start-up costs were met, the orthopaedic office-based system was the most convenient, efficient, and cost effective for the patient/family and treating physicians. This mirrors the experience of cardiologists, obstetricians, and family practitioners, fields in which the utility of office-based ultrasonography is widely recognized and has become the standard.

The effect of lactulose, pectin, arabinogalactan and cellulose on the production of organic acids and metabolism of ammonia by intestinal bacteria in a faecal incubation system.

An in vitro faecal incubation system was used to study the metabolism of complex carbohydrates by intestinal bacteria. Homogenates of human faeces were incubated anaerobically with added lactulose, pectin, the hemicellulose arabinogalactan, and cellulose, both before and after subjects had been pre-fed each carbohydrate. Fermentation of added substrate was assessed by the production of short-chain fatty acids (SCFA) and suppression of net ammonia generation over 48 h of incubation. Control faecal homogenates to which carbohydrate was not added yielded an average increment of SCFA of 43 mmol/l, equivalent to 172 mmol/kg in the original stool. The addition of lactulose, pectin and arabinogalactan each increased the yield of SCFA by a similar amount, averaging 6.5 mmol/g carbohydrate or 1.05 mol/mol hexose equivalent; organic acid yield was not increased by pre-feeding these substances for up to 2 weeks. Acetate was the major SCFA in all samples at all times and, after pre-feeding with extra carbohydrate, butyrate concentrations exceeded propionate in all samples. Faecal homogenates incubated with cellulose showed no greater SCFA production than controls over the first 48 h, but there was a slight increase when samples from two subjects pre-fed cellulose were incubated for 14 d. Net ammonia generation was markedly suppressed by addition of lactulose to faecal incubates with an initial period of net bacterial uptake of ammonia. Pectin and arabinogalactan also decreased ammonia generation, but the reductions were not significant unless subjects were pre-fed these materials; cellulose had no effect on ammonia generation.

The interpretation of retrograde pancreatography in the elderly.

Endoscopic retrograde pancreatography is regarded as the most specific technique in the diagnosis of chronic pancreatitis. In the elderly the relevance of endoscopic retrograde pancreatography changes in establishing the diagnosis is disputed. The pancreatograms of 101 patients aged seventy-five years or more, who had endoscopic retrograde pancreatography for suspected biliary or pancreatic pathology, were reviewed. Only four patients subsequently proved to have unequivocal chronic pancreatitis. There was no significant difference in the size or contour of the main pancreatic duct, or in side branch changes between those patients presenting with common bile duct stones, incidental biliary pathology and pain of probable pancreatic origin. Three definite diagnostic criteria for the diagnosis of chronic pancreatitis emerged--duct obstruction with a stricture, gross irregularity of the main pancreatic duct and the presence of large cavities.

Mucosal surface pH of the large intestine of the rat and of normal and inflamed large intestine in man.

The surface pH of rat distal colonic mucosa and human rectal mucosa was measured in vitro using first a small pH electrode with a flattened tip. In buffer with pH 7.56 the mean rat colonic surface pH was 6.72. Lowering the buffer pH in steps resulted in a small fall in surface pH, the values being buffer pH 7.06 surface pH 6.64, buffer pH 6.58 surface pH 6.61 and finally buffer pH 6.09 surface pH 6.39. Similar results were obtained with a buffer where butyrate, 30 mmol/l replaced chloride and when a CO2/bicarbonate buffer was used. During the time taken for the study transmural potential difference only changed by 1-2 mV. Serosal surface pH changed with buffer pH, suggesting that the maintained surface pH is a property of the mucosal surface only. The surface pH of human rectal mucosa was similar to that of rat distal colonic mucosa. As buffer pH fell from pH 7.51 to 5.96 mucosal surface pH only fell from pH 6.80 to 6.26. The values obtained in ulcerative proctitis did not differ from normal mucosa. Secondly pH microelectrodes were used to measure the juxta mucosal pH and the pH-microclimate thickness when luminal pH was controlled. The microclimate had a pH 6.63 adjacent to the mucosa with a thickness of 840 micron. The importance of mucus in maintaining the microclimate was shown by n-acetyl cysteine thinning and prostaglandin E2 thickening the layer. These results describe a surface microclimate in the large intestine of appreciable thickness and a constant juxta mucosal pH. Luminal pH changes produce only a small change in microclimate pH.

The intermediate filament cytoskeleton of macrophages.

This study characterizes two- and three-dimensional ultrastructure and surface topography of polymerized networks of intermediate filaments (IF) isolated from mouse peritoneal macrophages. Isolated IF bound to monoclonal anti-IF antibodies in enzyme-linked immunosorbent assays. Immunogold labeling of IF with specific antibodies revealed that epitopes are distributed along filaments particularly at junctions where filaments interconnect. Networks of IF, viewed by scanning electron microscopy, organized as ropelike groups of interconnecting filaments which swirl and encircle each other to form three-dimensional lattices containing ellipsoidal-, circular-, and vacuole-shaped cavities. Cavity diameters were similar in size to organelles and vacuoles; diameters were grouped as small (12-288 nm), medium (0.3-1.7 micron), and large (2-3 micron). The walls of the cavities appeared as beaded structures with alternating globular and linear regions. Linear regions were 14 nm. Repeat distances taken from the central axis of globular regions were 23-27 nm. The lattice organization of IF observed in vitro was similar to images seen in vivo in Triton-insoluble cytoskeletons immunofluorescently labeled with specific antibodies. In whole cells processed for TEM, swirling bundles of IF were found encircling membranous vacuoles. Based on the lattice architecture of IF, cavity dimensions, and IF location, we postulate that intermediate filaments may function in the mechanical and spatial distribution of vacuoles in the cell cytoplasm.

The effect of luminal pH on large intestinal absorption in ulcerative proctocolitis.

The sensitivity of large intestinal absorption to pH and the absorption of short-chain fatty acids were measured in ulcerative proctocolitis by a dialysis bag method. Lowering luminal pH from 7 to 5.5 significantly reduced absorption of salt and water in proctocolitis, but not in normal controls. Short-chain fatty acid absorption was not affected by inflammation or change in pH when patients with ulcerative proctocolitis were compared with controls.

Meningitis caused by Streptococcus suis type II.

A 49 year old head porter developed meningitis shown to be caused by Streptococcus suis type II. The rare human infections with the organism, a pathogen of pigs, usually occur in those employed handling pig meat and are associated with auditory or vestibular dysfunction.

Spontaneous bacterial peritonitis due to Campylobacter jejuni.

A patient with alcoholic cirrhosis complicated by ascites developed spontaneous bacterial peritonitis. Campylobacter jejuni--(a microaerophilic bacillus)--was isolated from ascitic fluid and blood.

Domestic metallic mercury poisoning.

In a family exposed to metallic mercury vapour two patients had acrodynia, one had the nephrotic syndrome, and one person remained well. Recognition of the variable manifestations of the disease and prevention of further exposure were the most important aspects of management. Recovery appeared to be complete as blood mercury levels fell to normal. Urinary mercury levels were too variable to be reliable as indications of progress.