Interventional cardiovascular procedures among HIV-infected individuals on antiretroviral therapy 1995-2000.

OBJECTIVE: To calculate the rate of interventional cardiac procedures (ICP) among HIV-infected individuals ever treated with antiretroviral therapy (ART) and to describe clinical and sociodemographic characteristics associated with ICP. METHODS: Since 1992, ART in British Columbia (BC) has been centrally distributed by the BC Centre for Excellence in HIV/AIDS. The BC Cardiac Registry maintains information regarding all cardiac procedures performed in BC. The two databases were linked to determine the number of HIV-positive individuals on ART who underwent ICP. Age-adjusted analyses were conducted using direct standardization, and linear regression to test for trend over time. Logistic regression was used to identify patient and treatment characteristics independently associated with having an interventional cardiac procedure. RESULTS: Of the 5082 individuals who have ever received ART, 63 (< 1%) were captured in the Cardiac Registry. There were 97 events: 70 (72%) since 1999. The age-adjusted event rate per 1000 HIV-positive individuals on ART increased significantly over time (P = 0.015) whereas that for the general BC population did not increase over time (P = 0.191). In multivariate analysis, age at baseline per 10 year increase [adjusted odds ratio (AOR) 2.5; 95% confidence interval (CI), 1.8-3.2), and months on ART (AOR 1.3; 95% CI, 1.1-1.4) remained significant. CONCLUSIONS: The rate of ICP among HIV-positive individuals on ART appears to be increasing; in addition, the duration of time on ART is independently associated with ICP after adjustment for patient demographic characteristics.

Co-morbidity data in outcomes research: are clinical data derived from administrative databases a reliable alternative to chart review?

Evaluation of co-morbidity data is essential in health outcomes research. Co-morbidity data derived from administrative databases has been criticized for lacking the accuracy required for clinical research. We compared co-morbidity data derived from a Canadian provincial hospitalization database with chart review in 817 adults treated with a percutaneous coronary intervention at a single tertiary care hospital between 1994 and 1995. While the administrative database tended to under-estimate the prevalence of some co-morbid conditions, the agreement between chart review and administrative data was good to very good for most conditions. Asymptomatic conditions were noted to have lower levels of agreement. Multivariate risk models for all-cause mortality constructed from both data sources were almost identical, suggesting minimal misclassification. The results indicate that clinical data abstracted from most Canadian hospitalization databases can provide reliable information regarding baseline co-morbid conditions believed to influence survival in a population undergoing percutaneous coronary interventions.

Molecular analysis of simple variant translocations in acute promyelocytic leukemia.

The primary cytogenetic abnormality in acute promyelocytic leukemia (APL; FAB M3) is a reciprocal translocation, t(15;17)(q22;q12), which serves to fuse the PML gene on chromosome 15 to the retinoic acid receptor alpha (RARA) gene on chromosome 17. A PML-RARA fusion message transcribed from the der(15) is thought to mediate leukemogenesis. Two APL patients with simple variants of this translocation, t(3;15)(q21;q22) and t(X;15)(p11;q22), have previously been reported who lack cytogenetic involvement of chromosome 17, although their breakpoint positions on chromosome 15 still suggest the involvement of the PML gene. Here we report on a combined analysis by molecular genetics and in situ hybridization of these two patients, in which we wanted to determine whether the PML gene has alternative fusion partners or whether cryptic rearrangement of the RARA locus has occurred instead. A cryptic involvement of RARA was demonstrated in both patients by a combination of Southern analysis, reverse transcription coupled to PCR (RT-PCR), and fluorescence in situ hybridization. The results indicate an absolute requirement for the rearrangement of the RARA gene in the pathogenesis of APL and underline the importance of RARA during normal myeloid differentiation.

Ordering of probes surrounding the Ewing's sarcoma breakpoint on chromosome 22 using fluorescent in situ hybridization to interphase nuclei.

Eight probes were localized by fluorescent in situ hybridization to the region surrounding the Ewing's sarcoma breakpoint on chromosome 22. Three of these were initially ordered by pair-wise hybridization to metaphase chromosomes with differential detection of the probes. These and the remaining probes were then ordered by hybridizing two or three probes simultaneously to interphase nuclei. In the two probe experiments and some of the three probe experiments, the order was derived by comparing mean interphase distances between signals from the probes. In the three probe experiments, either two probes were detected with one fluorochrome and the third with another or all three probes were individually distinguished by detecting one probe with fluorescein isothiocyanate (FITC), one with Texas Red, and one with both fluorochromes to give a mixed color. The order of the signals was then noted. Greater than 60 percent of configurations with a discrete order were shown or deduced to be correct. These approaches are assessed and we demonstrate a more obvious predominating order when all three probes are differentially detected. The order of the probes was deduced to be centromere: D22S271: D22S260: lambda S1: D22S262: cosK1831: Ewing's sarcoma breakpoint: cosLIF: D22S261: lambda S15: telomere.

Diagnosis of Ewing's sarcoma and peripheral neuroectodermal tumour based on the detection of t(11;22) using fluorescence in situ hybridisation.

Fluorescence in situ hybridisation (FISH) has been used increasingly for gene mapping and ordering probes on interphase and metaphase preparations. The association of consistent chromosomal aberrations with certain malignancies allows the possibility of using interphase cytogenetics as a diagnostic tool. In small round cell tumours of children accurate diagnosis may be difficult using existing methods. We have therefore evaluated the diagnostic potential of this technique when applied to the characteristic t(11;22) found in Ewing's sarcoma and peripheral neuroectodermal tumour (ES and PNET). Interphase nuclei were prepared from normal human foreskin fibroblasts (HFF), two Ewing's sarcoma cell lines and several fresh tumour biopsies. DNA probes each side of the breakpoint at 22q12 were labelled with biotin and digoxygenin, hybridised to chromosomes in interphase and detected in different colours. Measurements between pairs of signals arising from each copy of chromosome 22 were taken and statistical analysis performed. There was a highly significant difference (P < 0.0001) between the two populations of measurements obtained (from nuclei with and without the t(11;22)). Studying four tumours and one further ES line (blind) it was found that median values from 30 nuclei could correctly identify which samples contained the t(11;22). This application of interphase cytogenetics contributes a reliable, accurate and conceptually simple diagnostic test for ES and PNET. It may now be applied to other tumours with characteristic translocations, amplifications or deletions when suitable probes are available. This approach is likely to become a routine in clinical diagnosis.

The 200/220 kDa antigen recognized by monoclonal antibody (MAb) UJ127.11 on neural tissues and tumors is the human L1 adhesion molecule.

MAb UJ127.11, raised against 16 week human fetal brain, recognizes an antigen present primarily on normal and tumor tissues derived from the neuroectoderm. The antigen has previously been identified as a 220/240 kDa cell surface glycoprotein as determined by immunoprecipitation studies. We show here, that the 220/240 kDa antigen is the human L1 cell adhesion molecule and by Western blot analysis actually has a calculated molecular weight of between 200-220 kDa. Immunocytochemical studies with UJ127.11 and an antibody (5G3) recently utilized to isolate human L1 from brain indicate that both reagents have very similar binding profiles. The binding of radiolabelled UJ127.11 to its target antigen can be blocked by the addition of a rabbit anti-human L1 antiserum. Furthermore, sequential immunoprecipitation and Western blot analysis shows that UJ127.11 and the rabbit anti-human L1 antiserum recognize identical proteins.

A novel and rapid method for isolating sequences adjacent to rare cutting sites and their use in physical mapping.

We describe a simple PCR based technique which can be used to isolate sequences adjacent to rare cutter sites and can subsequently be employed for the construction of long range physical maps. The method involves the ligation of an adaptor to rare cutter sequences and its use as a target for forward priming in PCR. Primers to Alu repeat elements initiate synthesis of the reverse strand. Using this technique any rare cutter site which has a repeat element within amplification range can be cloned. We have isolated six unique sequences around NotI sites from an irradiation reduced hybrid containing a fragment of human chromosome 22 and are using these for physical mapping around the Ewing's sarcoma translocation breakpoint on chromosome 22.

Expression of neural cell adhesion molecule (NCAM) isoforms in neuroblastoma.

A comparative study on the expression of the neural cell adhesion molecule (NCAM) in human neuroblastoma cell lines and tissues was undertaken. NCAMs are a family of closely related cell surface glycoproteins involved in cell-cell interactions. Using antibodies that recognise distinct epitopes on NCAM, their presence was shown in neuroblastoma, but these studies do not yield any information on the specific NCAM isoforms associated with the tumour. Western and Northern blot analyses were therefore carried out to characterise the NCAM isoforms in this neuroectodermal tumour. Western blot studies using the monoclonal antibody ERIC-1 showed that all human neuroblastoma cell lines tested expressed the 140 and 120 kilodalton isoforms of NCAM in their desialo state. Some of the cell lines also expressed NCAM-180. The data are corroborated by Northern blotting where a transcript of 7.4 kilobase pairs was identified only in lines expressing NCAM-180; the 6.7 and 5.4 kilobase pair transcripts coding for 140 and 120 kilodalton isoforms, respectively, were present in all the cell lines tested. The NCAM isoforms identified in neuroblastoma were also different from those found in adult and fetal brain tissue, suggesting that aberrations are expressed in the molecule during tumorigenesis.

Expression of the cluster 1 antigen (neural cell adhesion molecule) in neuroectodermal tumours.

In this study, we have investigated the expression of the neural cell adhesion molecule (NCAM) in the human brain, primary brain tumours and neuroblastoma. Adult brain was found to express discrete isoforms of 180, 170, 140 and 120 kDa, which on neuraminidase treatment resolved into bands of 180, 170, 140, 120 and 95 kDa. Primary brain tumours such as Schwannoma and medulloblastoma expressed embryonic NCAM characterised by a high level of glycosylation, whereas other tumours, e.g. astrocytoma, meningioma, glioma and oligodendroglioma expressed adult NCAM. Post-neuraminidase treatment, differential expression of the 180, 170, 140, 120 and 95 kDa isoforms were noted in these various tumour types. On the other hand, neuroblastoma cell lines were found to express only embryonic NCAM, which after neuraminidase treatment resulted in differential presence of only 180, 140 and 120 kDa proteins.