Trisomy 15 and other nonrandom chromosome changes in Rauscher murine leukemia virus-induced leukemia cell lines.

The cytogenetics of Rauscher murine leukemia virus-induced erythroid, myeloid, and lymphatic leukemias were studied in BALB/c and DBA/2 mice. In primary virus-induced leukemias, no chromosome abnormalities were found. However, in tumors derived from transplanted leukemia tissue and in cell lines obtained from these tumors, euploidy and mostly aneuploidy were observed that varied according to the type of tumor cells and the number of passages. Eleven erythroleukemia cell lines showed aneuploidy from the first in vivo transplant stage on. Nonrandom abnormalities were found: trisomy 15 in 9 of 11 lines, followed by trisomy 3 in 8 of 11 lines and monosomy 6 in 5 of 11 lines. Subsequent evolution of the karyotypes was frequent (10 of 11 lines) and rapid. In vivo cell lines established from these tumors showed many structural rearrangements. Three myeloid lines revealed a stable karyotype with no or only minor changes: trisomy 15 in 1 line and normal diploid in 2 lines. One lymphatic leukemia cell line established in vitro presented with a very stable karyotype: trisomies 14 and 15. Another in vivo transplantable line showed trisomies 11, 9, and 17. These results suggest that trisomy of chromosome 15 plays a significant role in tumors derived from different types of mouse leukemias.

Cytogenetic analysis of human renal carcinoma cell lines of common origin (NC 65).

Cytogenetic analyses were performed on 3 clonal cell lines derived from a human renal cell carcinoma and its lymph node metastasis, two long-term tissue culture cell lines (NC 65-Sp and NC 65-R) and a serially transplantable tumor line growing on nude mice and brought into culture at the fifth animal passage (NC 65-V). Karyotype were established using banding techniques. Most of the marker chromosomes could be identified and were derived by deletion, inversion, translocation, or isochromosome formation of Chromosomes 1, 3, 4, 5, 8, 9, and 17. These markers were different from HeLa markers. NC 65-Sp had a near diploid chromosome number, NC 65-R a hypotetraploid number, and NC 65-V had a bimodal chromosome number, and NC 65-V had a bimodal chromosome number. Three chromosome markers were shared by the three cell lines; NC 65-R and NC 65-V shared an additional set of four markers. Markers specific to each line were also observed; they demonstrated the independent derivation of the lines and eliminated laboratory cross-contamination. Common markers between the lines confirmed their common tumoral origin.

Establishment and characterization of a melanoma cell line from a xeroderma pigmentosum patient: activation of N-ras at a potential pyrimidine dimer site.

Patients suffering from the genetic disorder xeroderma pigmentosum (XP) display an extreme sensitivity of their skin to sun (UV) exposure and predisposition to skin cancer due to deficiencies in the excision DNA repair pathway. Here we describe the establishment and characterization of the first tumor cell line derived from an XP patient (belonging to complementation group C). The melanoma cell line designated XP44RO(Mel) has retained its tumorigenic and XP phenotype (UV sensitivity, reduced unscheduled DNA synthesis) and showed karyotypic abnormalities characteristic of melanomas. Transfection of XP44RO(Mel) DNA to NIH3T3 cells and oligonucleotide hybridization revealed that the N-ras oncogene was activated by an A.T to T.A or C.G transversion at the third position of codon 61. This mutation occurs at a dipyrimidine site. It is likely initiated by a UV-induced pyrimidine dimer and is of a type rarely observed in mammalian shuttle vector systems and endogenous genes after UV irradiation.

Split-signal FISH for detection of chromosome aberrations in acute lymphoblastic leukemia.

Chromosome aberrations are frequently observed in precursor-B-acute lymphoblastic leukemias (ALL) and T-cell acute lymphoblastic leukemias (T-ALL). These translocations can form leukemia-specific chimeric fusion proteins or they can deregulate expression of an (onco)gene, resulting in aberrant expression or overexpression. Detection of chromosome aberrations is an important tool for risk classification. We developed rapid and sensitive split-signal fluorescent in situ hybridization (FISH) assays for six of the most frequent chromosome aberrations in precursor-B-ALL and T-ALL. The split-signal FISH approach uses two differentially labeled probes, located in one gene at opposite sites of the breakpoint region. Probe sets were developed for the genes TCF3 (E2A) at 19p13, MLL at 11q23, ETV6 at 12p13, BCR at 22q11, SIL-TAL1 at 1q32 and TLX3 (HOX11L2) at 5q35. In normal karyotypes, two colocalized green/red signals are visible, but a translocation results in a split of one of the colocalized signals. Split-signal FISH has three main advantages over the classical fusion-signal FISH approach, which uses two labeled probes located in two genes. First, the detection of a chromosome aberration is independent of the involved partner gene. Second, split-signal FISH allows the identification of the partner gene or chromosome region if metaphase spreads are present, and finally it reduces false-positivity.

MDR1 expression in poor-risk acute myeloid leukemia with partial or complete monosomy 7.

Expression of the multidrug resistance (MDR1) phenotype, encoded by the MDR1 gene, is an adverse prognostic factor for CR and survival in acute myeloid leukemia (AML). Other prognostic factors, such as specific cytogenetic abnormalities, have been identified in AML. We have investigated the expression of the MDR1 gene in untreated AML patients with monosomy 7 (n = 12), and partial deletions (n = 7) of the long arm of chromosome 7 (respectively -7/7q-), because of the extremely bad prognosis associated with these cytogenetic abnormalities and because of the fact that the MDR1 gene is located on chromosome 7q21.1. The findings were compared with the level of MDR1 expression in a group of 42 other AML patients, matched for age with favourable, neutral or complex cytogenetic abberations. MDR1 mRNA expression, as measured by the RNase protection assay was significantly higher in the -7/7q- group vs other AML patients (median 1.3 vs 0.1 arbitrary units, P = 0.02). Protein expression of MDR1 in the -7/7q- group, as determined with the monoclonal antibody MRK16, was found to be similar to the levels found in the control group. With a functional rhodamine retention assay using the modulator PSC833, increased MDR1 activity was observed in the -7/7q- group as compared to the control group of patients (P = 0.05). Considering the higher MDR1 mRNA expression and equal or slightly elevated level of protein expression of MDR1, we studied the presence of MDR1 genes in this group of -7/7q- patients. Fluorescence in situ hybridization (FISH) studies, using a specific MDR1 probe revealed no loss of an MDR1 allele in any of the deleted q- arms of the seven patients with 7q-, whereas all monosomy 7 patients lacked one MDR1 gene homologue. To determine whether there was selective loss of the MDR1 gene in the -7/7q- patients, the genetic polymorphism of the MDR1 gene was used. Both allelic variants (G and T) were represented in the -7/7q- and in the control group, showing a predominance for GT at position 2677 of the MDR1 gene in the control group. In the 12 monosomy 7 patients loss of the MDR1 allele was random. Methylation studies of the CpG island of the MDR1 gene revealed no hypermethylation in any of the -7/7q- patients. We conclude that MDR1 expression in -7/7q- AML patients is upregulated at transcriptional, but not at translational level, suggesting that mechanisms other than MDR1 are responsible for the poor prognosis in these patients.

t(7;12)(q36;p13) and t(7;12)(q32;p13)--translocations involving ETV6 in children 18 months of age or younger with myeloid disorders.

Our retrospective karyotype review revealed two rare recurrent translocations affecting ETV6 (TEL): t(7;12)(q36;p13) and t(7;12)(q32;p13). Five patients with a t(7;12) were from a group of 125 successfully karyotyped pediatric patients enrolled in consecutive clinical AML trials of the Dutch Childhood Leukemia Study Group over a period of 7 years. During a search of available cytogenetic databases, we found 7q and 12p abnormalities in two additional Dutch patients and in three participants in Pediatric Oncology Group trials. A del(12p) had been initially identified in four of these patients and re-examination of the original karyograms revealed a t(7;12)(q36;p13) in two instances and a probable t(7;12) in the other two. FISH confirmed the presence of a t(7;12)(q36;p13) in the latter. Most (n = 7) also had trisomy 19. The t(7;12)(q36;p13) (n = 9) was more common than the t(7;12)(q32;p13) (n = 1). These subtle translocations were found only in children 18 months of age or younger. A literature search revealed that the t(7;12) with break-points at 7q31-q36 and 12p12-p13 had been reported in six children with myeloid disorders and in two with acute lymphoblastic leukemia; all were 12 months of age or younger. Only two of the 17 for whom survival data were available, were alive after at least 22 months of continuous complete remission. Our findings suggest that ETV6 rearrangements due to a t(7;12) may play an adverse role in myeloid disorders in children 18 months of age or younger. Therefore, children in this age group with myeloid disorders should be screened for both MLL and ETV6 rearrangements.

Isolation, properties and chromosomal localization of four closely linked hamster interferon-alpha-encoding genes.

Three recombinant phages containing hamster interferon-alpha-encoding genes (Ha Ifa) were isolated from a Ha genomic library, using a murine (Mu) Ifa probe. The phage inserts contained overlapping genomic fragments which span a total length of approx. 30 kb, on which four Ha Ifa genes are localized. The Ifa gene cluster could be assigned to hamster chromosome 2q. The nt sequences of the four Ifa genes were determined. Two of the genes are functional (Ha Ifa-1 and Ifa-3) and two are pseudogenes (Ifa-ps2 and Ifa-ps4). Ha Ifa-1 and -3 were transiently expressed in COS cells and they gave rise to protein products (A1 and A3, respectively) with antiviral properties on hamster CHO cells. In addition, Ha A1 revealed high antiviral activity on murine L929 cells.

Effect of physical exercise on blood lipids and adipose tissue composition in young healthy men.

In a prospective, controlled study, the influence of strenuous physical exercise on plasma total cholesterol, HDL cholesterol, apolipoprotein A-I, total triglycerides and fatty acid composition of adipose tissue was studied during 7 months of training in 15 senior oarsmen and 21 controls matched for age, smoking and drinking habits. Dietary intake was monitored. At the start of the study there were no differences in lipid parameters and adipose tissue composition between oarsmen and controls. A significant decrease in total cholesterol and total triglycerides and an increase in HDL cholesterol and apolipoprotein A-I was noted in the oarsmen after only 2 weeks of training. Body weight remained stable but skin-fold thickness decreased. Food consumption increased considerably but diet composition remained unchanged. Nevertheless the fatty acid pattern of adipose tissue changed significantly, as a result of altered preferential endogenous fatty acid synthesis and/or fatty acid oxidation. Since lipid parameters and adipose tissue composition were equal in oarsmen and controls before the start of the training, it is concluded that the changes induced by exercise only last for the duration of the training period.

Is the chromosomal region 9q34 always involved in variants of the Ph1 translocation?

Six variants of the Ph1 translocation are described. The clinical diagnoses were chronic myeloid leukemia (CML) in 5 cases (patients 1-5) and acute lymphocytic leukemia (ALL) in patient 6. Three Ph1 variants were clear complex translocations, involving chromosomes #9, #22, and a third chromosome, i.e., #16, #11, or #14. The other three Ph1 variants appeared as "simple" translocations between chromosome #22 and chromosome #19, #4, or #12 when G- or Q-banding were used. When studied with high resolution R-banding, a small deletion of the terminal part of one chromosome #9 was visible, strongly suggesting that these variants were also complex translocations, i.e., t(9;19;22)(q34;p13;q11),t(4;9;22) (p16;q34;q11), and t(9;12;22)(q34;p13;q11). In the latter two cases, using in situ hybridization techniques, we demonstrated the presence of c-abl sequences on the Ph1 chromosome. This proved the involvement of 9q34 in these two variants. Our proposal is that most, and probably all, variants of Ph1 are complex translocations involving part of 9q34 and that the conjunction of a specific region of 22q11 with a specific segment of 9q34 (carrying the c-abl protooncogene) is essential for the development of Ph1 + CML.

A new B-cell line showing a complex translocation (8;14;18) and BCL2 rearrangement.

A cell line named ROS-50 (Rotterdam suspension cell line no. 50) has been established from peripheral blood of a 69-year-old male with acute lymphoblastic leukemia (FAB type L3). Among the aberrations, cytogenetic analysis showed the presence of 14q+, 18q-, and two 8q- marker chromosomes. With fluorescence in situ hybridization (FISH) we characterized the chromosomal translocations, t(8;14) and t(14;18), in which the same chromosome 14 is involved. PCR analysis demonstrated the presence of on IGH-BCL2 rearrangement with a breakpoint in the minor cluster region (mcr) confirming the t(14;18) characteristic for follicular lymphoma. Additional studies showed high expression of BCL2 protein, an early B-cell immunophenotype, and an unusual pattern of IGH gene rearrangement.

Translocation of c-abl to "masked" Ph in chronic myeloid leukemia.

In two patients with chronic myeloid leukemia (CML), the nature of the chromosomal rearrangement giving rise to "masked" Ph has been studied by in situ hybridization of human c-abl sequences. The c-abl probes hybridized to the 22q11 region of the "masked" Ph, demonstrating that translocation of sequences from 9q34 to the Ph did occur exactly as in standard Ph or in other types of variants previously studied. These results provide additional evidence for the occurrence of a constant molecular rearrangement in Ph-positive CML.

An ultra-micro method for the determination of total nitrogen in biological fluids based on Kjeldahl digestion and enzymatic estimation of ammonia.

An ultra-micro method for the determination of the total nitrogen-content of biological fluids and suspensions is described, based on a digestion in sulphuric acid and a enzymatic determination of the ammonia formed with glutamate dehydrogenase (EC 1.4.1.3). The proposed method yields the same results as the classical Kjeldahl procedure, but is less time-consuming. The detection-limit of the nitrogen, without loss of precision and accuracy, is much lower than in the original Kjeldahl procedure, and is in the order of 35 ng N per sample.

Prediction of infarct size from serial CK determinations: evaluation by clinical studies and computer simulation.

To assess reduction of infarct size by therapeutic intervention, a high predictive accuracy is mandatory. The CK release in the circulation (CKr) was studied in 12 consecutive patients after uncomplicated myocardial infarction, admitted within 5 h after onset of symptoms. Despite improvement of existing methods, such as a more frequent sampling, CK-MB determination instead of total CK determination and use of a gamma-exponential instead of a log-normal curve-fitting technique, the correlation between CKr predicted from measurements within 7 h after the start of CK rise and CKr calculated after completion of the CK curve remained poor. Computer simulations were done to investigate measurement errors as a cause of this failure. Normally distributed noise, with standard deviations ranging from 0.2% to 8.0% of peak CK-MB, was added to the first points of an ideal gamma-exponential CK-MB curve and predictions were made from these "noisy" points. A small noise already produced a great variation in prediction: 0.8% noise resulted in a deviation of predicted CKr from calculated CKr ranging from --20 to +6%. It is concluded that adequate prediction of infarct size from serial CK determinations in the first 7 h after onset of the CK rise must fail if the precision of the biochemical determination is not less than 0.4%.

Hemodynamic effects and image quality of low-osmolar ionic and nonionic contrast media during pulmonary angiography.

RATIONALE AND OBJECTIVES: We compared the hemodynamic responses to ionic and nonionic low-osmolar contrast media of patients who underwent pulmonary angiography. METHODS: Ninety-nine consecutive patients with suspected pulmonary emboli were randomly assigned to receive either 40 ml iohexol or 40 ml ioxaglate in 2 sec at 600 psi (0.17 kg/m2). Mean pulmonary arterial pressure, pulse rate, and blood pressure were recorded before, immediately after, and 2, 5, and 10 min following injection. Image quality was assessed by readers who were unaware of drug assignment. RESULTS: Pulmonary arterial pressure increased to a maximum at 2 min and was higher in patients with pulmonary emboli (p = .06). There were no significant differences between the two contrast media used. The systolic blood pressure and pulse rate in patients with pulmonary emboli increased significantly more in the ioxaglate group (ps = .03 and .04, respectively). Image quality was excellent in 90% of both groups. CONCLUSION: Both contrast agents are safe for pulmonary angiography and yield similar image quality. There appears to be a positive inotropic effect of ioxaglate.

In vitro adsorption of possible aetiological factors of hepatic encephalopathy.

Four different adsorbents (activated charcoal, XAD-4, a strong base anion and a strong acid cation-exchange resin) were tested in vitro for their capacity to remove substances that may be important in the development of hepatic encephalopathy. Separate columns packed with one of these adsorbents were perfused for three hours with a reconstituted plasma solution containing simultaneously high concentrations of amino-acids, ammoniumchloride, short-chain fatty acids, octopamine and bile salts. Effective removal of all these substances was only obtained when either activated charcoal, or XAD-4, were combined with the cation-exchange resin. Possible implications for the treatment of hepatic coma are discussed.

Maternal stress during hospitalization of the adopted child.

PURPOSE: To identify and describe the experiences of mothers whose adopted children are hospitalized and to compare those experiences to those of mothers whose biological children are hospitalized. DESIGN: Comparative descriptive design. METHODS: Mothers of hospitalized children (n = 33 adopted; n = 19 biological) completed a slightly revised version of the Parental Stressor Scale: Pediatric Intensive Scale Unit (PSS:PICU). Adoptive mothers also completed a questionnaire related to their perceptions of the impact of their child's adoption on the hospitalization experience. RESULTS: Adoptive mothers perceived statistically significantly higher levels of stress related to their child's behavioral/emotional response to hospitalization. Children who had been with the family for less time perceived the hospitalization as more stressful. When both groups of mothers were considered, mothers of younger children perceived a higher level of stress. A majority of adoptive mothers felt the adoption had an impact on the hospitalization experience, especially related to their limited information about their child's medical history, staff lack of knowledge about legal issues, and concern about attachment to the child. CLINICAL IMPLICATIONS: Nurses need to be aware of adoptive mothers' concerns related to attachment issues, limited family medical history, and legal rights in order to provide sensitive and effective care. Inservice education programs could be designed to help teach all staff about these important issues.

Unique issues of the adopted child: helping parents talk openly and honestly with their child and the community.

1. Due to the psychological impact of losses associated with adoption, adopted children are at risk for developing adjustment problems. Children raised in families that are comfortable talking about adoption are more likely to be emotionally healthy. 2. The child's perception and understanding of adoption changes as the child develops and matures. 3. Health care professionals can teach parents about typical concerns related to adoption and ways to provide information and support consistent with the child's emotional maturity and learning ability. 4. Parents can also learn strategies to counter critical social attitudes and promote a more positive view of adoption.