The clinical utility of conventional karyotyping in the detection of cytogenetic abnormalities in soft tissue tumours: an Asian institutional experience.

OBJECTIVES: To assess the clinical utility of conventional karyotyping as a diagnostic tool in soft tissue tumours amidst the increasing use of molecular cytogenetics. DESIGN: Case series. SETTING: Singapore General Hospital, an Asian institution. PARTICIPANTS: A total of 35 participants (18 male and 17 female) aged 15 to 81 years were included in this study. Conventional karyotyping of 35 consecutive fresh soft tissue tumour specimens was performed over 4 years and the results were analysed. RESULTS: Of the 35 cases of soft tissue tumours reviewed, chromosome abnormalities were detected in 22 (63%) cases, 11 (31%) showed a normal karyotype, and 2 (6%) had culture failure. Of the 22 cases with abnormal karyotype, nine (41%) cases showed recurring aberrations: Ewing's sarcomas (n=2), desmoplastic small round cell tumour (n=1), synovial sarcomas (n=3), myxoid liposarcomas (n=2), and lipoma (n=1). One lipoma case had a t(2;12)(q23;q15) in which 2q23 breakpoint was not reported before. Chromosomal aberration involving 12q15 breakpoint has been shown in a previous study to be indicative of a lipoma-like liposarcoma. Another lipoma case had addition of 5q15 and 9p13 together with a balanced aberration of t(12;13) (q13;q12) which were novel aberrations. One synovial sarcoma case showed t(3;7)(q21;p13) which was an uncharacteristic aberration. CONCLUSION: Conventional karyotyping demonstrated utility as a genome-wide screening tool for soft tissue tumours and an adjunct diagnostic tool in the event histopathology results were doubtful. With the more widespread use of karyotyping, novel recurring chromosomal aberrations may be discovered.

An abnormal nonhyperdiploid karyotype is a significant adverse prognostic factor for multiple myeloma in the bortezomib era.

Multiple myeloma is clinically heterogeneous and risk stratification is vital for prognostication and informing treatment decisions. As bortezomib is able to overcome several high-risk features of myeloma, the validity of conventional risk-stratification and prognostication systems needs to be reevaluated. We study the survival data of 261 previously untreated myeloma patients managed at our institution, where bortezomib became available from 2004 for the treatment of relapse disease. Patient and disease characteristics, and survival data were evaluated overall, and with respect to bortezomib exposure. Overall, the international staging system (ISS), metaphase karyotyping and interphase fluorescence in situ hybridization (FISH) were discerning of survival outcomes, where the median for the entire cohort was 5.2 years. However, when stratified by bortezomib exposure, only metaphase karyotyping was still discriminating of long-term prognosis. The presence of an abnormal nonhyperdiploid karyotype overrides all other clinical and laboratory parameters in predicting for a worse outcome on multivariate analysis (median survival 2.6 years, P = 0.001), suggesting that bortezomib used at relapse is better able to overcome adverse risk related to high tumor burden (as measured by the ISS) than adverse cytogenetics on conventional karyotyping. Metaphase karyotyping provides additional prognostic information on tumor kinetics where the presence of a normal diploid karyotype in the absence of any high-risk FISH markers correlated with superior survival and could act as a surrogate for lower plasma cell proliferation.

Production of globules in mouse L cells penetrated with hamster sperms.

Globule formation has been observed in long-term cultures of mouse L cells penetrated with hamster spermatozoa. These numerous, uniform, and spherical structures are approximately 10 micrometers in diameter when formed, and are positive to stains specific for proteins and nucleic acids. They may be produced either by sperms within their target cells or by the cells in response to penetration by the spermatozoa.

Sequential spine-hand radiography for assessing skeletal maturity with low radiation EOS imaging system for bracing treatment recommendation in adolescent idiopathic scoliosis: a feasibility and validity study.

PURPOSE: The EOS-imaging system is increasingly adopted for clinical follow-up in scoliosis with the advantages of simultaneous biplanar imaging of the spine in an erect position. Skeletal maturity assessment using a hand radiograph is an essential adjunct to spinal radiography in scoliosis follow-up. This study aims at testing the feasibility and validity of a newly proposed EOS workflow with sequential spine-hand radiography for skeletal maturity assessment and bracing recommendation. METHODS: EOS spine-hand radiographs from patients with diagnosis of idiopathic scoliosis, including both sexes and an age range of ten to 14 years, were scored using the Thumb Ossification Composite Index (TOCI), Sanders and Risser methods. Intraclass correlation coefficients (ICCs) were calculated for inter/intraobserver agreement and were tested with Cronbach's alpha values. RESULTS: In all, 60 EOS-spine hand radiographs selected from subjects with diagnosis of adolescent idiopathic scoliosis (AIS), including 32 male patients (mean age 11.53 years; 10 to 14) and 28 female patients (mean age 11.50 years; 10 to 13) who underwent sequential spine-hand low dose EOS imaging were generated for analysis. The overall interobserver (ICC = 0.997) and intraobserver agreement (α > 0.9) demonstrated excellent agreement for TOCI staging; ICC > 0.994 for both TOCI and Sanders staging comparing traditional digital versus EOS hand radiography; ICC ≥ 0.841 for agreement on bracing recommendation among TOCI versus the Risser and Sanders system. CONCLUSION: With the proposed new EOS workflow it was feasible to produce high image quality for skeletal maturity assessment with excellent reliability and validity to inform consistent bracing recommendation in AIS. The workflow is applicable for busy daily clinic settings in tertiary scoliosis centres with reduced time cost, improved efficiency and throughput of the radiology department. LEVEL OF EVIDENCE: III.

Enhancement of neutrophil function by the bronchial epithelium stimulated by epidermal growth factor.

The bronchial epithelium is an important physical barrier that regulates physiological processes including leukocyte trafficking. The aim of the present study was to elucidate the mechanisms whereby the bronchial epithelium, stimulated by epidermal growth factor (EGF) as part of a response to acute or chronic injury, could activate and chemoattract human neutrophils. Subconfluent human bronchial epithelial (16HBE) cells were stimulated with EGF to mimic the in vivo events after injury. The effect of the resulting EGF-conditioned media (CM) was compared with that of basal-CM with respect to neutrophil activation and chemotaxis. Such findings were then confirmed using primary bronchial epithelial cells (PBECs) from healthy volunteers. EGF-CM from 16HBE cells caused increased expression of CD11b/CD66b and CD62L loss on neutrophils when compared with basal-CM. EGF-CM contained significant neutrophil chemotactic activity involving granulocyte-macrophage colony-stimulating factor and interleukin-8 that was potentiated by leukotriene B(4). This was dependent on neutrophil phosphatidylinositol-3-kinase activation and Akt phosphorylation, with partial regulation by phospholipase D, but not mammalian target of rapamycin. Consistent with these observations, EGF-CM derived from PBECs displayed increased chemotactic activity. The present results suggest that the enhanced chemotactic activity of the epidermal growth factor-conditioned epithelium can enhance neutrophil-mediated immunity during acute injury, while during continued injury and repair, as in chronic asthma, this could contribute to persistent neutrophilic inflammation.

Relationship between the pharmacokinetics and the analgesic and respiratory pharmacodynamics of epidural sufentanil.

To establish the relationships between epidural sufentanil analgesia and respiratory effects and to determine the pharmacokinetics of the drug, 22 adult patients undergoing thoracotomy were put into a randomized, double-blind study and received either 30, 50, or 75 micrograms per dose in 20 ml normal saline solution. Repeated doses were given on request for the 24-hour study period. There was a weak but significant nonlinear correlation between length of effective analgesia and the cumulative dose of the drug (r = 0.26, p less than 0.001). In 12 of 22 patients, the maximal length of effective analgesia was reached before the last dose and the effect tended to taper off thereafter. The mean maximal length of effective analgesia was 4.69 +/- 0.32 hours (mean +/- SEM), whereas the length of effective analgesia with the last dose was only 3.34 +/- 0.46 hours (p less than 0.0005). There was a significant correlation between the peak serum concentrations of sufentanil during the dose interval and the length of effective analgesia (r = 0.44, p less than 0.0001). Area under the concentration-time curve was proportional to the size of the epidural dose, and with all three doses tested there was a gradual accumulation of sufentanil in the serum. Mean time-to-peak concentration (tmax) increased with repeated doses (p less than 0.05). Mean serum concentration of sufentanil during periods of slow respiratory rate (0.47 +/- 0.05 ng/ml) was significantly higher than during episodes without adverse respiratory effects (0.37 +/- 0.05 ng/ml, p less than 0.05). The above data suggest that an important part of the analgesic and adverse effects of sufentanil are mediated centrally, after this opioid is absorbed systemically.

Nasal effects of bradykinin and capsaicin: influence on plasma protein leakage and role of sensory neurons.

Nasal insufflation with bradykinin induces nasal discomfort, rhinorrhea, and nasal blockage, all features of rhinitis. We recently showed these effects to be mediated by the B2-receptor subtype, which has been identified at neural and vascular sites. To investigate the relative contribution of capsaicin-sensitive sensory neural stimulation to the action(s) of bradykinin, two randomized double-blind placebo-controlled studies have been undertaken comparing the nasal effects of single-dose administrations of bradykinin (1.88 x 10(-3) M) and capsaicin (3.28 x 10(-5) M). In comparison with placebo, both bradykinin and capsaicin induced nasal pain/discomfort (P less than 0.01) and rhinorrhea (P less than 0.02). Bradykinin significantly increased nasal airways resistance (P less than 0.005) and plasma protein exudation (P less than 0.02). No such changes were identified after nasal challenge with capsaicin. These findings suggest that bradykinin-induced nasal discomfort and rhinorrhea are neurally mediated, whereas the effects on nasal airways resistance and plasma protein exudation are due to a direct vascular action. In addition, these findings question the role of capsaicin-sensitive sensory neurons in nasal vasculature responses, because no vascular effects of capsaicin could be identified in the human nasal mucosa.

Biological activity and anti-prostaglandin effects of prostaglandin analogue, ent-11-epi-15-epi PGE2 methyl ester.

The biological activity and anti-prostaglandin property of the prostaglandin analogue, ent-11-epi-15-epi PGE2 methyl ester, were studied on isolated guinea pig ileum. Ent-11-epi-15-epi PGE2 methyl ester contracted guinea pig ileum and produced a concentration-response curve parallel to that of PGE2. However, the former exhibited a lower maximal effect than PGE2. At concentrations greater than 10(-6)M, ent-11-epi-15-epi PGE2 methyl ester selectively antagonized contractile actions of PGE2 and PGE2 alpha without affecting contractions induced by acetylcholine. These observations suggest that the PG analogue acted like a competitive antagonist to PGE2 and PGF2 alpha on guinea pig ileum in vitro.

Highly complex chromosomal rearrangement of chromosome 9 in a case of chronic myeloid leukemia.

A 63-year-old man with chronic myeloid leukemia (CML) was found to have a new complex Philadelphia translocation. All of the bone marrow cells had a rearrangement of a five-way translocation, t(9;22;10;12;1), involving a single chromosome 9. The patient went into blast crisis two years after initial diagnosis and the karyotype remained unchanged. He died in blast crisis 10 months later. We believe this case is a unique 5-way translocation in which four chromosomes were translocated to a single chromosome.

Pseudodicentric (16;12)(q11;p11.2) in a type AB (mixed) thymoma.

Genetic alterations of thymomas are rarely described in the literature. In this study, a previously unreported instance of aberrant karyotypic change consisting of 45,XX,pseu dic(16;12) (q11;p11.2) [cp23]/87-90,idemx2[cp4] in a Masaoka Stage II mixed thymoma or type AB thymoma affecting a 56-year-old Chinese woman is detailed. Abnormalities involving 12p containing important tumor suppressor-like genes have been documented especially in hematological malignancies. Recently, recurrent losses involving 16q, a locus known to harbor several tumor suppressor genes, have been described in type C thymomas (squamous cell carcinoma), suggesting a possible relationship between type AB thymoma and type C thymoma. Whether these genes are involved in the pathogenesis of type AB thymoma remain to be clarified and it is currently unclear if cytogenetic studies may eventually play a role in the classification of thymic tumors.

Cytogenetic analysis of invasive breast cancer: a study of 27 Asian patients.

Tumor cytogenetic analysis from 27 patients with breast cancer diagnosed at the Singapore General Hospital revealed complex karyotypic aberrations in 12 cases. The study group comprised 25 women and 2 men, ranging in age from 33 to 78 years (median 52 years). Ethnic distribution consisted of 22 Chinese, 3 Malaysian, and 2 Indian patients. Pathologic assessment disclosed 24 invasive ductal, 2 invasive mucinous, and 1 mixed invasive mucinous and ductal carcinomas. Histologic grading showed 3 grade 1, 10 grade 2, and 12 grade 3 tumors; 2 cancers were not graded, because they had been subjected to prior chemotherapy. Tumor sizes ranged from 1.5 to 10 cm (median 3 cm). Eleven cases were axillary node negative, whereas the remaining 16 node-positive cancers affected as many as 3 nodes in 8 cases and 4 or more nodes in another 8. Twenty cases demonstrated estrogen-receptor positivity, and 8 cases progesterone-receptor positivity. The spectrum of cytogenetic abnormalities involved chromosomes 1, 3, 6, 7, 8, 11, 16, and 17 and ranged from gains and deletions of both long and short arms, trisomy, monosomy, and other rearrangements. There was a trend toward the presence of karyotypic abnormalities in tumors of higher grade.

A case of t(8;14) with total and partial trisomy 3 in Waldenstrom macroglobulinemia.

A case of Waldenstrom macroglobulinemia with Burkitt-type t(8;14)(q24.1;q32) and total and partial trisomy 3 is reported. This is an unusual combination of chromosomal abnormalities in Waldenstrom macroglobulinemia.

Acute promyelocytic leukemia with a dicentric chromosome involving chromosomes 11, 17, and 18.

We report a case of acute promyelocytic leukemia with dicentric chromosome resulting from translocation of chromosomes 11, 17, and 18.

Interaction of kunjin virus with octyl-D-glucoside extracted Vero cell plasma membrane.

Initial experiments using whole cells have shown that there were specific and saturable interactions between kunjin (KUN) virus and receptor molecules on the Vero cell surfaces. Solubilisation of Vero cell plasma membranes with octyl-D-glucoside (OG) yielded an extract which also interacted specifically with KUN virus. This was proven using electron microscopy. When the virus-OG-extract complex was exposed onto Vero cell monolayers, no KUN virus was observed to enter into the whole cells. This would imply that there was virus-receptor interaction with the OG-extract leaving no free virus to attach to the whole cells. The attachment kinetics of KUN virus was studied further using the Scatchard analysis which indicated the involvement of more than one interactive macromolecule in the attachment event.

Inhibition of platelet aggregation and antagonism of vasopressin-induced ECG changes in primates by a carboprostacyclin analogue, ZK 36374.

A chemically stable carboprostacyclin analogue, ZK 36374 has been compared with two other prostacyclin derivatives with respect to ADP-induced in vitro aggregation of baboon and human platelets and ex vivo platelet aggregation in the baboon. ZK 36374 was also tested on the systemic arterial blood pressure of the baboon and against vasopressin-induced ECG changes in primates. Compared to the other two compounds, ZK 36374 displayed enhanced anti-platelet aggregating activity; there was dissociation between this property and its hypotensive potency. ZK 36374 antagonized the vasopressin-induced ECG changes. These results indicate that ZK 36374 possesses therapeutic potential in vascular disease including that affecting the coronary vessels.

Molecular and phenotypic spectrum of de novo Philadelphia positive acute leukemia.

The Philadelphia chromosome is present in a heterogeneous group of leukemias. It is most commonly associated with chronic myelogenous leukemia (CML) and B-lineage acute lymphoblastic leukemia (ALL) being found in more than 95% and 15-25% of cases respectively. We undertook a study to determine the morphologic, phenotypic and molecular diversity of Philadelphia positive de novo acute leukemia patients seen at our institution over the past 3 1/2 years. Twenty-one patients with de novo acute leukemia were found to have the Philadelphia chromosome by cytogenetic studies. They consisted of 3 patients with acute myelogenous leukemia (AML), 1 biphenotypic leukemia and 17 ALL patients. Of the patients with ALL, 16 were of B-lineage while 1 had a T-cell phenotype. Ten patients expressed the p210 BCR-ABL transcript alone and 10 expressed only the p190 BCR-ABL transcript. One patient had co-expression of p190 and p210 b3a2 BCR-ABL transcripts. Thus the Philadelphia chromosome can be found in a diverse cohort of morphologic and immunologic subtypes of de novo acute leukemia reflecting the heterogeneity of lineage involvement in this disease.

Effects of short-term exposure to 0.2 ppm ozone on biomarkers of inflammation in sputum, exhaled nitric oxide, and lung function in subjects with mild atopic asthma.

To gain further insight into the kinetics of airway inflammatory response and explore the possibility of nitric oxide as a surrogate marker of the lower airway inflammatory response to ozone, nine subjects with mild atopic asthma were exposed to filtered air or 0.2 ppm ozone for 2 hours with intermittent exercise. Lung function was measured at baseline and immediately after exposures. Sputum induction was performed at 6 hours and at 24 hours after exposures, and exhaled nitric oxide levels were measured at baseline, immediately, 6, and 24 hours after both exposures. A significant decline in forced expiratory volume in one second and inspiratory capacity was detectable following exposure to ozone. In addition, a 2-fold increase was observed in the percentage of polymorphonuclear leukocytes 6 hours after exposure to ozone, with no changes in other biomarkers at this time point. By 24 hours after ozone exposure, the neutrophilia had subsided but there was an increase in albumin, total protein, myeloperoxidase, and eosinophil cationic protein. Exhaled nitric oxide levels, histamine, interleukin-8, and growth-related oncogene-alpha in sputum did not change significantly following ozone exposure. It was concluded that short-term ozone exposure induces an acute inflammatory response in asthmatic airways, characterized by early polymorphonuclear leukocyte influx followed by plasma extravasation and activation of eosinophils and neutrophils. Exhaled nitric oxide is not a useful marker for detecting inflammatory response to ozone in persons with mild asthma.

Effects of bupivacaine and its isomers on guinea pig tracheal smooth muscle.

The effects of racemic bupivacaine and its (-)-(S)- and (+)-(R)-isomers on guinea pig tracheal smooth muscle tension were studied in vitro. Racemic bupivacaine had a dual action with contraction at low concentrations (6.9-55 x 10(-6) M) and relaxation at high concentrations (1.1-18 x 10(-4) M), (-)-(S)-Bupivacaine had dual action comparable to that of racemic bupivacaine, while (+)-(R)-bupivacaine had only weak contractile effects with more marked relaxant effects. The contractile effects of bupivacaine were abolished in Ca(2+)-free medium and by verapamil, while the relaxant effects were not influenced by verapamil or Ca(2+)-free medium. The (-)-(S)-isomer is responsible for the contractile effects of racemic bupivacaine, whereas both (-)-(S)- and (+)-(R)-isomers contribute to its relaxant effects. In preparations denuded of epithelium, the (maximal) responses were attenuated to 86% of maximum contraction and 48% of maximum relaxation, suggesting that the epithelium plays a larger role in relaxant than in contractile responses to bupivacaine. It is concluded that bupivacaine has a dual action on guinea pig tracheal smooth muscle with epithelium-independent. Ca(2+)-dependent contraction at low concentrations and epithelium-dependent, Ca(2+)-independent relaxation at higher concentrations.

Philadelphia positive acute lymphoblastic leukaemia (ALL): clinico-haematologic characteristics, molecular analyses and 3-year follow up--a single institution study.

INTRODUCTION: The Philadelphia chromosome is one of the commonest chromosomal aberration in adult acute lymphoblastic leukaemia (ALL) patients. We present the results of a 3-year prospective study to look at the clinico-haematologic, immunophenotypic, cytogenetic and molecular profile of 13 adult patients with Philadelphia (Ph) positive ALL out of 35 newly diagnosed ALL seen at our institution over the past 3 years. MATERIALS AND METHODS: Thirty-five adult ALL patients seen between 1996 and 1998 comprised the study group. Marrow samples were obtained for immunophenotyping and karyotypic analysis at diagnosis. Samples were also obtained simultaneously for molecular testing for Ph chromosome. RESULTS: Thirteen patients were found to be Ph positive by molecular analysis while cytogenetic studies identified the chromosomal abnormality in 9 of these patients. The median age of our Ph positive patients was similar to those without Ph chromosome. Pre-B phenotype appears to be common in this group of patients. In concordance with other studies, Ph positive ALL was associated with a poor prognosis in our patients. CONCLUSION: Identification of Ph chromosome is important in the management of patients with ALL as it is an important prognostic marker.