Gravitational effects of Nix and Hydra in the external region of the Pluto-Charon system

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Diffusion of contrast medium after four different techniques for analgesia of the proximal metacarpal region: an in vivo and in vitro study

REASONS FOR PERFORMING STUDY: There is limited information on potential diffusion of local anaesthetic solution after various diagnostic analgesic techniques of the proximal metacarpal region. OBJECTIVE: To document potential distribution of local anaesthetic solution following 4 techniques used for diagnostic analgesia of the proximal metacarpal region. METHODS: Radiodense contrast medium was injected around the lateral palmar or medial and lateral palmar metacarpal nerves in 8 mature horses, using 4 different techniques. Radiographs were obtained 0, 10 and 20 min after injection and were analysed subjectively. A mixture of radiodense contrast medium and methylene blue was injected into 4 cadaver limbs; the location of the contrast medium and dye was determined by radiography and dissection. RESULTS: Following perineural injection of the palmar metacarpal nerves, most of the contrast medium was distributed in an elongated pattern axial to the second and fourth metacarpal bones. The carpometacarpal joint was inadvertently penetrated in 4/8 limbs after injections of the palmar metacarpal nerves from medial and lateral approaches, and in 1/8 limbs when both injections were performed from the lateral approach. Following perineural injection of the lateral palmar nerve using a lateral approach, the contrast medium was diffusely distributed in all but one limb, in which the carpal sheath was inadvertently penetrated. In 5/8 limbs, following perineural injection of the lateral palmar nerve using a medial approach, the contrast medium diffused proximally to the distal third of the antebrachium. CONCLUSIONS AND POTENTIAL RELEVANCE: Inadvertent penetration of the carpometacarpal joint is common after perineural injection of the palmar metacarpal nerves, but less so if both palmar metacarpal nerves are injected using a lateral approach. Following injection of the lateral palmar nerve using a medial approach, the entire palmar aspect of the carpus may be desensitised.

Recognition of Membrane-Bound Fusion-Peptide/MPER Complexes by the HIV-1 Neutralizing 2F5 Antibody : Implications for Anti-2F5 Immunogenicity

13 p.

The Ontogeny of Mucosal and Systemic Antibody Responses to HIV-1 Infection

The humoral immune system plays a critical role in the clearance of numerous pathogens. In the setting of HIV-1 infection, the virus infects, integrates its genome into the host's cells, replicates, and establishes a reservoir of virus-infected cells. The initial antibody response to HIV-1 infection is targeted to non-neutralizing epitopes on HIV-1 Env gp41, and when a neutralizing response does develop months after transmission, it is specific for the autologous founder virus and the virus escapes rapidly. After continuous waves of antibody mediated neutralization and viral escape, a small subset of infected individuals eventually develop broad and potent heterologous neutralizing antibodies years after infection. In this dissertation, I have studied the ontogeny of mucosal and systemic antibody responses to HIV-1 infection by means of three distinct aims: 1. Determine the origin of the initial antibody response to HIV-1 infection. 2. Characterize the role of restricted VH and VL gene segment usage in shaping the antibody response to HIV-1 infection. 3. Determine the role of persistence of B cell clonal lineages in shaping the mutation frequencies of HIV-1 reactive antibodies.

After the introduction (Chapter 1) and methods (Chapter 2), Chapter 3 of this dissertation describes a study of the antibody response of terminal ileum B cells to HIV-1 envelope (Env) in early and chronic HIV-1 infection and provides evidence for the role of environmental antigens in shaping the repertoire of B cells that respond to HIV-1 infection. Previous work by Liao et al. demonstrated that the initial plasma cell response in the blood to acute HIV-1 infection is to gp41 and is derived from a polyreactive memory B cell pool. Many of these antibodies cross-reacted with commensal bacteria, Therefore, in Chapter 3, the relationship of intestinal B cell reactivity with commensal bacteria to HIV-1 infection-induced antibody response was probed using single B cell sorting, reverse transcription and nested polymerase chain reaction (RT- PCR) methods, and recombinant antibody technology. The dominant B cell response in the terminal ileum was to HIV-1 envelope (Env) gp41, and 82% of gp41- reactive antibodies cross-reacted with commensal bacteria whole cell lysates. Pyrosequencing of blood B cells revealed HIV-1 antibody clonal lineages shared between ileum and blood. Mutated IgG antibodies cross-reactive with both Env gp41 and commensal bacteria could also be isolated from the terminal ileum of HIV-1 uninfected individuals. Thus, the antibody response to HIV-1 can be shaped by intestinal B cells stimulated by commensal bacteria prior to HIV-1 infection to develop a pre-infection pool of memory B cells cross-reactive with HIV-1 gp41.

Chapter 4 details the study of restricted VH and VL gene segment usage for gp41 and gp120 antibody induction following acute HIV-1 infection; mutations in gp41 lead to virus enhanced neutralization sensitivity. The B cell repertoire of antibodies induced in a HIV-1 infected African individual, CAP206, who developed broadly neutralizing antibodies (bnAbs) directed to the HIV-1 envelope gp41 membrane proximal external region (MPER), is characterized. Understanding the selection of virus mutants by neutralizing antibodies is critical to understanding the role of antibodies in control of HIV-1 replication and prevention from HIV-1 infection. Previously, an MPER neutralizing antibody, CAP206-CH12, with the binding footprint identical to that of MPER broadly neutralizing antibody 4E10, that like 4E10 utilized the VH1-69 and VK3-20 variable gene segments was isolated from this individual (Morris et al., 2011). Using single B cell sorting, RT- PCR methods, and recombinant antibody technology, Chapter 4 describes the isolation of a VH1-69, Vk3-20 glycan-dependent clonal lineage from CAP206, targeted to gp120, that has the property of neutralizing a neutralization sensitive CAP206 transmitted/founder (T/F) and heterologous viruses with mutations at amino acids 680 or 681 in the MPER 4E10/CH12 binding site. These data demonstrate sites within the MPER bnAb epitope (aa 680-681) in which mutations can be selected that lead to viruses with enhanced sensitivity to autologous and heterologous neutralizing antibodies.

In Chapter 5, I have completed a comparison of evolution of B cell clonal lineages in two HIV-1 infected individuals who have a predominant VH1-69 response to HIV-1 infection--one who produces broadly neutralizing MPER-reactive mAbs and one who does not. Autologous neutralization in the plasma takes ~12 weeks to develop (Gray et al., 2007; Tomaras et al., 2008b). Only a small subset of HIV-1 infected individuals develops high plasma levels of broad and potent heterologous neutralization, and when it does occur, it typically takes 3-4 years to develop (Euler et al., 2010; Gray et al., 2007; 2011; Tomaras et al., 2011). The HIV-1 bnAbs that have been isolated to date have a number of unusual characteristics including, autoreactivity and high levels of somatic hypermutations, which are typically tightly regulated by immune control mechanisms (Haynes et al., 2005; 2012b; Kwong and Mascola, 2012; Scheid et al., 2009a). The VH mutation frequencies of bnAbs average ~15% but have been shown to be as high as 32% (reviewed in Mascola and Haynes, 2013; Kwong and Mascola, 2012). The high frequency of somatic hypermutations suggests that the B cell clonal lineages that eventually produce bnAbs undergo high-levels of affinity maturation, implying prolonged germinal center (GC) reactions and high levels of T cell help. To study the duration of HIV-1- reactive B cell clonal persistence, HIV-1 reactive and non HIV-1- reactive B cell clonal lineages were isolated from an HIV-1 infected individual that produces bnAbs, CAP206, and an HIV-1 infected individual who does not produce bnAbs, 004-0. Single B cell sorting, RT-PCR and recombinant antibody technology was used to isolate and produce monoclonal antibodies from multiple time points from each individual. B cell sequences clonally related to mAbs isolated by single cell PCR were identified within pyrosequences of longitudinal samples of these two individuals. Both individuals produced long-lived B cell clones that persisted from 0-232 weeks in CAP206, and 0-238 weeks in 004-0. The average length of persistence of clones containing members isolated from two separate time points was 91.5 weeks both individuals. Examples of the continued evolution of clonal lineages were observed in both the bnAb and non-bnAb individual. These data indicated that the ability to generate persistent and evolving B cell clonal lineages occurs in both bnAb and non-bnAb individuals, suggesting that some alternative host or viral factor is critical for the generation of highly mutated broadly neutralizing antibodies.

Together the studies described in Chapter 3-5 show that multiple factors influence the antibody response to HIV-1 infection. The initial antibody response to HIV-1 Env gp41 can be shaped by a B cell response to intestinal commensal bacteria prior to HIV-1 infection. VH and VL gene segment restriction can impact the B cell response to multiple HIV-1 antigens, and virus escape mutations in the MPER can confer enhanced neutralization sensitivity to autologous and heterologous antibodies. Finally, the ability to generate long-lived HIV-1 clonal lineages in and of itself does not confer on the host the ability to produce bnAbs.

Comparison of two pressurisers for cementation of the proximal femur

PURPOSE: To compare pressures generated by 2 different cement pressurisers at various locations in the proximal femur. METHODS: Two groups of 5 synthetic femurs were used, and 6 pressure sensors were placed in the femur at 20-mm intervals proximally to distally. Cement was filled into the femoral canal retrogradely using a cement gun with either the half-moon pressuriser or the femoral canal pressuriser. Maximum pressures and pressure time integrals (cumulative pressure over time) of the 2 pressurisers were compared. RESULTS: At all sensors, the half-moon pressuriser produced higher maximum pressures and pressure time integrals than the femoral canal pressuriser, but the difference was significant only at sensor 1 (proximal femur). This may result in reduced cement interdigitation in the proximal femur. CONCLUSION: The half-moon pressuriser produced higher maximum cementation pressures and pressure time integrals than the femoral canal pressuriser in the proximal femur region, which is critical for rotational stability of the implant and prevention of implant fracture. KEYWORDS: arthroplasty, replacement, hip; bone cements; femur

Secular dynamics and family identification among highly inclined asteroids in the Euphrosyne region

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

BILATERAL ANOMALY IN EVANDROMYIA EVANDROI (DIPTERA: PSYCHODIDAE: PHLEBOTOMINAE) CAPTURED IN VICENCIA MUNICIPALITY, NORTHERN RAINFOREST REGION OF PERNAMBUCO STATE, BRAZIL

Approximately 800 species of phlebotomine sand flies, many of which are vectors of Leishmania, have been described. Besides morphological similarities within groups, the occurrence of anomalies within a species may lead to an erroneous description of new species. This paper describes one phlebotomine sand fly, Evandromyia evandroi, with a symmetrical bilateral anomaly in the number of spines on the gonostyle. In this specimen, the anomalous spine is located in the external region of gonostyle, inserted between the upper external and the lower external spines. It is important to document morphological anomalies, so as to avoid erroneous sand fly identifications.

Membrane associated transporter protein gene (SLC45A2) and the genetic basis of normal human pigmentation variation

This work is concerned with the genetic basis of normal human pigmentation variation. Specifically, the role of polymorphisms within the solute carrier family 45 member 2 (SLC45A2 or membrane associated transporter protein; MATP) gene were investigated with respect to variation in hair, skin and eye colour ― both between and within populations. SLC45A2 is an important regulator of melanin production and mutations in the gene underly the most recently identified form of oculocutaneous albinism. There is evidence to suggest that non-synonymous polymorphisms in SLC45A2 are associated with normal pigmentation variation between populations. Therefore, the underlying hypothesis of this thesis is that polymorphisms in SLC45A2 will alter the function or regulation of the protein, thereby altering the important role it plays in melanogenesis and providing a mechanism for normal pigmentation variation. In order to investigate the role that SLC45A2 polymorphisms play in human pigmentation variation, a DNA database was established which collected pigmentation phenotypic information and blood samples of more than 700 individuals. This database was used as the foundation for two association studies outlined in this thesis, the first of which involved genotyping two previously-described non-synonymous polymorphisms, p.Glu272Lys and p.Phe374Leu, in four different population groups. For both polymorphisms, allele frequencies were significantly different between population groups and the 272Lys and 374Leu alleles were strongly associated with black hair, brown eyes and olive skin colour in Caucasians. This was the first report to show that SLC45A2 polymorphisms were associated with normal human intra-population pigmentation variation. The second association study involved genotyping several SLC45A2 promoter polymorphisms to determine if they also played a role in pigmentation variation. Firstly, the transcription start site (TSS), and hence putative proximal promoter region, was identified using 5' RNA ligase mediated rapid amplification of cDNA ends (RLM-RACE). Two alternate TSSs were identified and the putative promoter region was screened for novel polymorphisms using denaturing high performance liquid chromatography (dHPLC). A novel duplication (c.–1176_–1174dupAAT) was identified along with other previously described single nucleotide polymorphisms (c.–1721C>G and c.–1169G>A). Strong linkage disequilibrium ensured that all three polymorphisms were associated with skin colour such that the –1721G, +dup and –1169A alleles were associated with olive skin in Caucasians. No linkage disequilibrium was observed between the promoter and coding region polymorphisms, suggesting independent effects. The association analyses were complemented with functional data, showing that the –1721G, +dup and –1169A alleles significantly decreased SLC45A2 transcriptional activity. Based on in silico bioinformatic analysis that showed these alleles remove a microphthalmia-associated transcription factor (MITF) binding site, and that MITF is a known regulator of SLC45A2 (Baxter and Pavan, 2002; Du and Fisher, 2002), it was postulated that SLC45A2 promoter polymorphisms could contribute to the regulation of pigmentation by altering MITF binding affinity. Further characterisation of the SLC45A2 promoter was carried out using luciferase reporter assays to determine the transcriptional activity of different regions of the promoter. Five constructs were designed of increasing length and their promoter activity evaluated. Constitutive promoter activity was observed within the first ~200 bp and promoter activity increased as the construct size increased. The functional impact of the –1721G, +dup and –1169A alleles, which removed a MITF consensus binding site, were assessed using electrophoretic mobility shift assays (EMSA) and expression analysis of genotyped melanoblast and melanocyte cell lines. EMSA results confirmed that the promoter polymorphisms affected DNA-protein binding. Interestingly, however, the protein/s involved were not MITF, or at least MITF was not the protein directly binding to the DNA. In an effort to more thoroughly characterise the functional consequences of SLC45A2 promoter polymorphisms, the mRNA expression levels of SLC45A2 and MITF were determined in melanocyte/melanoblast cell lines. Based on SLC45A2’s role in processing and trafficking TYRP1 from the trans-Golgi network to stage 2 melanosmes, the mRNA expression of TYRP1 was also investigated. Expression results suggested a coordinated expression of pigmentation genes. This thesis has substantially contributed to the field of pigmentation by showing that SLC45A2 polymorphisms not only show allele frequency differences between population groups, but also contribute to normal pigmentation variation within a Caucasian population. In addition, promoter polymorphisms have been shown to have functional consequences for SLC45A2 transcription and the expression of other pigmentation genes. Combined, the data presented in this work supports the notion that SLC45A2 is an important contributor to normal pigmentation variation and should be the target of further research to elucidate its role in determining pigmentation phenotypes. Understanding SLC45A2’s function may lead to the development of therapeutic interventions for oculocutaneous albinism and other disorders of pigmentation. It may also help in our understanding of skin cancer susceptibility and evolutionary adaptation to different UV environments, and contribute to the forensic application of pigmentation phenotype prediction.

Genes involved in complex adaptive processes tend to have highly conserved upstream regions in mammalian genomes

BACKGROUND:Recent advances in genome sequencing suggest a remarkable conservation in gene content of mammalian organisms. The similarity in gene repertoire present in different organisms has increased interest in studying regulatory mechanisms of gene expression aimed at elucidating the differences in phenotypes. In particular, a proximal promoter region contains a large number of regulatory elements that control the expression of its downstream gene. Although many studies have focused on identification of these elements, a broader picture on the complexity of transcriptional regulation of different biological processes has not been addressed in mammals. The regulatory complexity may strongly correlate with gene function, as different evolutionary forces must act on the regulatory systems under different biological conditions. We investigate this hypothesis by comparing the conservation of promoters upstream of genes classified in different functional categories.RESULTS:By conducting a rank correlation analysis between functional annotation and upstream sequence alignment scores obtained by human-mouse and human-dog comparison, we found a significantly greater conservation of the upstream sequence of genes involved in development, cell communication, neural functions and signaling processes than those involved in more basic processes shared with unicellular organisms such as metabolism and ribosomal function. This observation persists after controlling for G+C content. Considering conservation as a functional signature, we hypothesize a higher density of cis-regulatory elements upstream of genes participating in complex and adaptive processes.CONCLUSION:We identified a class of functions that are associated with either high or low promoter conservation in mammals. We detected a significant tendency that points to complex and adaptive processes were associated with higher promoter conservation, despite the fact that they have emerged relatively recently during evolution. We described and contrasted several hypotheses that provide a deeper insight into how transcriptional complexity might have been emerged during evolution.

cAMP stimulates transcription of the beta 2-adrenergic receptor gene in response to short-term agonist exposure.

In addition to conveying cellular responses to an effector molecule, receptors are often themselves regulated by their effectors. We have demonstrated that epinephrine modulates both the rate of transcription of the beta 2-adrenergic receptor (beta 2AR) gene and the steady-state level of beta 2AR mRNA in DDT1MF-2 cells. Short-term (30 min) exposure to epinephrine (100 nM) stimulates the rate of beta 2AR gene transcription, resulting in a 3- to 4-fold increase in steady-state beta 2AR mRNA levels. These effects are mimicked by 1 mM N6,O2'-dibutyryladenosine 3',5'-cyclic monophosphate (Bt2cAMP) or foskolin but not by phorbol esters. The half-life of the beta 2AR mRNA after addition of actinomycin D (46.7 +/- 10.2 min; mean +/- SEM; n = 5) remained unchanged after 30 min of epinephrine treatment (46.8 +/- 10.6 min; mean +/- SEM; n = 4), indicating that a change in transcription rate is the predominant factor responsible for the increase of beta 2AR mRNA. Whereas brief exposure to epinephrine or Bt2cAMP does not significantly affect the total number of cellular beta 2ARs (assessed by ligand binding), continued exposure results in a gradual decline in beta 2AR number to approximately 20% (epinephrine) or approximately 45% (Bt2cAMP) of the levels in control cells by 24 hr. Similar decreases in agonist-stimulated adenylyl cyclase activity are observed. This loss of receptors with prolonged agonist exposure is accompanied by a 50% reduction in beta 2AR mRNA. Transfection of the beta 2AR promoter region cloned onto a reporter gene (bacterial chloramphenicol acetyltransferase) allowed demonstration of a 2- to 4-fold induction of transcription by agents that elevate cAMP levels, such as forskolin or phosphodiesterase inhibitors. These results establish the presence of elements within the proximal promoter region of the beta 2AR gene responsible for the transcriptional enhancing activity of cAMP and demonstrate that beta 2AR gene expression is regulated by a type of feedback mechanism involving the second messenger cAMP.

Electron-impact excitation of Sc II: collision strengths and effective collision strengths for fine-structure transitions.

Accurate fine-structure atomic data for the Fe-peak elements are essential for interpreting astronomical spectra. There is a severe paucity of data available for Sc II, highlighted by the fact that no collision strengths are readily available for this ion. We present electron-impact excitation collision strengths and Maxwellian averaged effective collision strengths for Sc II. The collision strengths were calculated for all 3916 transitions amongst 89 jj levels (arising from the 3d4s, 3d2, 4s2, 3d4p, 4s4p, 3d5s, 3d4d, 3d5p, 4p2 and 3d4f configurations), resulting in a 944 coupled channel problem. The R-matrix package RMATRXII was utilized, along with the transformation code FINE and the external region code PSTGF, to calculate the collision strengths for a range of incident electron energies in the 0 to 8.3 Rydberg region. Maxwellian averaged effective collision strengths were then produced for 27 temperatures lying within the astrophysically significant range of 30 to 105 K.
The collision strengths and effective collision strengths were produced for two different target models. The purpose was to systematically examine the effect of including open 3p correlation terms into the configuration interaction expansion for the wavefunction. The first model consisted of all 36 CI terms that could be generated with the 3p core closed. The second model incorporated an additional six configurations which allowed for single-electron excitations from within the 3p core. Comparisons are made between the two models and the results of Bautista et al., obtained by private communication. It is concluded that the first model produced the most reliable set of collision and effective collision strengths for use in astrophysical and plasma applications.

S100A2 is a BRCA1/p63 coregulated tumour suppressor gene with roles in the regulation of mutant p53 stability

Here, we show for the first time that the familial breast/ovarian cancer susceptibility gene, BRCA1, along with interacting ΔNp63 proteins, transcriptionally upregulate the putative tumour suppressor protein, S100A2. Both BRCA1 and ΔNp63 proteins are required for S100A2 expression. BRCA1 requires ΔNp63 proteins for recruitment to the S100A2 proximal promoter region, while exogenous expression of individual ΔNp63 proteins cannot activate S100A2 transcription in the absence of a functional BRCA1. Consequently, mutation of the ΔNp63/p53 response element within the S100A2 promoter completely abrogates the ability of BRCA1 to upregulate S100A2. S100A2 shows growth control features in a range of cell models. Transient or stable exogenous S100A2 expression inhibits the growth of BRCA1 mutant and basal-like breast cancer cell lines, while short interfering RNA (siRNA) knockdown of S100A2 in non-tumorigenic cells results in enhanced proliferation. S100A2 modulates binding of mutant p53 to HSP90, which is required for efficient folding of mutant p53 proteins, by competing for binding to HSP70/HSP90 organising protein (HOP). HOP is a cochaperone that is required for the efficient transfer of proteins from HSP70 to HSP90. Loss of S100A2 leads to an HSP90-dependent stabilisation of mutant p53 with a concomitant loss of p63. Accordingly, S100A2-deficient cells are more sensitive to the HSP-90 inhibitor, 17-N-allylamino-17-demethoxygeldanamycin, potentially representing a novel therapeutic strategy for S100A2- and BRCA1-deficient cancers. Taken together, these data demonstrate the importance of S100A2 downstream of the BRCA1/ΔNp63 signalling axis in modulating transcriptional responses and enforcing growth control mechanisms through destabilisation of mutant p53.

Régulation de la proprotéine convertase subtilisine / kexine type 9 (PCSK9) dans les cellules intestinales Caco-2/15

La proprotéine convertase subtilisine/kexine type 9 (PCSK9) favorise la dégradation post-transcriptionnelle du récepteur des lipoprotéines de faible densité (LDLr) dans les hépatocytes et augmente le LDL-cholestérol dans le plasma. Cependant, il n’est pas clair si la PCSK9 joue un rôle dans l’intestin. Dans cette étude, nous caractérisons les variations de la PCSK9 et du LDLr dans les cellules Caco-2/15 différentiées en fonction d’une variété d’effecteurs potentiels. Le cholestérol (100 µM) lié à l’albumine ou présenté en micelles a réduit de façon significative l’expression génique (30%, p<0,05) et l’expression protéique (50%, p<0,05) de la PCSK9. Étonnamment, une diminution similaire dans le LDLr protéique a été enregistrée (45%, p<0,05). Les cellules traitées avec le 25-hydroxycholestérol (50 µM) présentent également des réductions significatives dans l’ARNm (37%, p<0,01) et la protéine (75%, p<0,001) de la PCSK9. Une baisse des expressions génique (30%, p<0,05) et protéique (57%, p<0,01) a également été constatée dans le LDLr. Des diminutions ont aussi été observées pour la HMG CoA réductase et la protéine liant l’élément de réponse aux stérols SREBP-2. Il a été démontré que le SREBP-2 peut activer transcriptionnellement la PCSK9 par le biais de la liaison de SREBP-2 à son élément de réponse aux stérols situé dans la région proximale du promoteur de la PCSK9. Inversement, la déplétion du contenu cellulaire en cholestérol par l’hydroxypropyl-β-cyclodextrine a augmenté l’expression génique de la PCSK9 (20%, p<0,05) et son contenu protéique (540%, p<0,001), en parallèle avec les niveaux protéiques de SREBP-2. L’ajout des acides biliaires taurocholate et déoxycholate dans le milieu apical des cellules intestinales Caco-2/15 a provoqué une baisse d’expression génique (30%, p<0,01) et une hausse d’expression protéique (43%, p<0,01) de la PCSK9 respectivement, probablement via la modulation du FXR (farnesoid X receptor). Ces données combinées semblent donc indiquer que la PCSK9 fonctionne comme un senseur de stérols dans le petit intestin.

The impact of different types of physical activity on total and regional bone mineral density in young Brazilian athletes

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Hemangioendothelioma of bone in a patient with a constitutional supernumerary marker

A 13-year old girl was diagnosed as having a bone hemangioendothelioma. Cytogenetic studies identified the presence of a small supernumerary marker chromosome in this patient. Classical cytogenetic methods using G-, C-, Ag-NOR-banding were supplemented by spectral karyotyping (SKY) and fluorescence in situ hybridization to reveal a karyotype 47,XX,+mar.ish der(22)(D22S543+) karyotype in cells derived from the tumor and lymphocytes. These findings suggest that the supernumerary marker chromosome originated from the proximal centromeric region of chromosome 22, and that trisomy of the region 22q11: was not associated with adverse phenotypic effects, but that the presence of trisomy 22q11 may be related to the development of this tumor. (C) Elsevier B.V., 1999. All rights reserved.