Epstein-Barr virus-related dacryocystitis: a case report.

BACKGROUND AND OBJECTIVE: Acute dacryocystitis is an atypical and rare manifestation of pediatric mononucleosis still widely underdiagnosed in clinical practice. We report this rare condition and describe challenges in its diagnosis and treatment on the basis of a presented case. CASE PRESENTATION: A 6-year-old Caucasian girl without any ophthalmic history was admitted for right preseptal cellulitis requiring intravenous antibiotic therapy. During hospitalization, she developed a fluctuating lump in the nasolacrimal region which resembled an abscess, both clinically and radiologically. There was no spontaneous purulent discharge. Serology was positive for acute mononucleosis and Epstein-Barr virus-related dacryocystitis was diagnosed. Following multidisciplinary discussion, she was treated conservatively with digital lacrimal sac massages and intravenous antibiotic therapy with an excellent outcome. DISCUSSION: This rare form of Epstein-Barr virus is poorly documented in the literature, and thus barely known. As initial symptoms are nonspecific (rhinitis, fever, eyelid edema and erythema lack of purulent discharge, and moderate bilateral cervical lymphadenopathy), diagnosis is often difficult. Nevertheless, differentiating between dacryocystitis and abscess is crucial to select the appropriate treatment and avoid unnecessary, potentially harmful surgery. Conservative management of dacryocystitis appears to be the gold standard of treatment. CONCLUSION: Acute dacryocystitis in children free of ophthalmic history should raise suspicion of primary Epstein-Barr virus infection. With conservative treatment, prognosis appears to be excellent; therefore, surgery should be avoided as much as possible.

Student perception of a new integrated anatomy practical program: does students' prior learning make a difference?

While there is evidence that science and non-science background students display small differences in performance in basic and clinical sciences, early in a 4-year, graduate entry medical program, this lessens with time. With respect to anatomy knowledge, there are no comparable data as to the impact previous anatomy experience has on the student perception of the anatomy practical learning environment. A study survey was designed to evaluate student perception of the anatomy practical program and its impact on student learning, for the initial cohort of a new medical school. The survey comprised 19 statements requiring a response using a 5-point Likert scale, in addition to a free text opportunity to provide opinion of the perceived educational value of the anatomy practical program. The response rate for a total cohort of 82 students was 89%. The anatomy practical program was highly valued by the students in aiding their learning of anatomy, as indicated by the high mean scores for all statements (range: 4.04-4.7). There was a significant difference between the students who had and had not studied a science course prior to entering medicine, with respect to statements that addressed aspects of the course related to its structure, organization, variety of resources, linkage to problem-based learning cases, and fairness of assessment. Nonscience students were more positive compared to those who had studied science before (P levels ranging from 0.004 to 0.035). Students less experienced in anatomy were more challenged in prioritizing core curricular knowledge. Clin. Anat. 24:664-670, 2011. © 2011 Wiley-Liss, Inc.

Effects of unilateral ovariectomy with compensatory hypertrophy on ovarian blood flow, oxygen consumption and progestin secretion in the 16-day pregnant rat.

Stimulated ovulation with resultant multiple corpora lutea (CL) can result in lower progesterone levels than expected from the increased luteal tissue mass. Unilateral ovariectomy (ULO) was used to increase CL number in rats and determine whether this would compromise luteal tissue blood flow, oxygen consumption and progestin secretion. All investigations were performed in vivo, using a venous outflow technique on Day 16 of gestation, when progesterone secretion is maximal. ULO, performed before pregnancy, doubled CL number and total CL mass in the remaining ovary of six treated compared to five control rats. Growth of CL was not affected. The rate of ovarian blood flow (microL min(-1) mg CL(-1)) fell to 47% of control levels in ULO animals and progesterone secretion (microg h(-1) mg CL(-1)) to 68%. Secretion of the minor progestin, 20alpha-hydroxypregn-4-en-one was not affected. Tissue oxygen consumption was maintained despite the reduction in blood flow by an increase in oxygen extraction from arterial blood. These results suggest that overcrowding of CL in ULO-stimulated rat ovaries compromises luteal tissue blood flow and subsequently progesterone secretion.

Proximal arterial vasoconstriction precedes regression of the hyaloid vasculature.

PURPOSE: To determine whether constriction of proximal arterial vessels precedes involution of the distal hyaloid vasculature in the mouse, under normal conditions, and whether this vasoconstriction is less pronounced when the distal hyaloid network persists, as it does in oxygen-induced retinopathy (OIR). METHODS: Photomicrographs of the vasa hyaloidea propria were analysed from pre-term pups (1-2 days prior to birth), and on Days 1-11 post-birth. The OIR model involved exposing pups to approximately 90% O(2) from D1-5, followed by return to ambient air. At sampling times pups were anaesthetised and perfused with india ink. Retinal flatmounts were also incubated with FITC-lectin (BS-1, G. simplicifolia,); this labels all vessels, allowing identification of vessels not patent to the perfusate. RESULTS: Mean diameter of proximal hyaloid vessels in pre-term pups was 25.44 +/- 1.98 microm; +/- 1 SEM). Within 3-12 hrs of birth, significant vasoconstriction was evident (diameter:12.45 +/- 0.88 microm), and normal hyaloid regression subsequently occurred. Similar vasoconstriction occurred in the O(2)-treated group, but this was reversed upon return to room air, with significant dilation of proximal vessels by D7 (diameter: 31.75 +/- 11.99 microm) and distal hyaloid vessels subsequently became enlarged and tortuous. CONCLUSIONS: Under normal conditions, vasoconstriction of proximal hyaloid vessels occurs at birth, preceding attenuation of distal hyaloid vessels. Vasoconstriction also occurs in O(2)-treated pups during treatment, but upon return to room air, the remaining hyaloid vessels dilate proximally, and the distal vessels become dilated and tortuous. These observations support the contention that regression of the hyaloid network is dependent, in the first instance, on proximal arterial vasoconstriction.

Effects of noradrenaline on blood flow, progesterone secretion and oxygen consumption in the intact ovary of rats on day 16 of pregnancy.

The effects of noradrenaline on the rates of secretion of ovarian progesterone and 20 alpha-hydroxypregn-4-en-3-one (20 alpha-OHP), blood flow and oxygen consumption were examined in rats on day 16 of pregnancy. A modified venous outflow technique was used to infuse noradrenaline directly into the ovary, without recirculation, and to monitor subsequent changes in the ovary. Noradrenaline was infused for periods of 10 min at a low and a high concentration, which achieved effective blood concentrations of about 6.25 and 25 ng ml-1, respectively. Each period of noradrenaline infusion was interspersed by a 10 min period of infusion of its ascorbic acid carrier. Two series of infusions of low and high concentrations of noradrenaline were carried out on each rat. Neither the infusion of the ascorbic acid carrier nor of the low concentration of noradrenaline had any effect on ovarian progestin secretion. The high concentration of noradrenaline reduced blood flow by 30% but had no apparent effect on progestin secretion or oxygen consumption. Collectively, these findings question the generally accepted view that noradrenaline has a physiological role in the regulation of progesterone secretion. Further, putative luteotrophins need to be examined in the intact ovary as well as under in vitro and indirect in vivo conditions to determine their physiological role.

Direction of blood flow and changes in resistance of major arteries supplying the ovary of the pregnant rat.

In this study, we examined changes in vascular resistance contributing to increased ovarian blood flow in the pregnant rat. Ovarian blood flow was monitored in vivo using a venous outflow cannulation technique in nonpregnant rats and in pregnant rats at Day 16 and Day 22, and increased from 0.18 +/- 0.02 to 0.81 +/- 0.09 and 1.02 +/- 0.08 ml min(-1) ovary(-1) (mean +/- SEM; n = 7, 7, 6), respectively. Intrinsic vessels within the ovarian complex accounted for 81%, 73%, and 70% of total resistance to ovarian blood flow. Of the two major extrinsic supply vessels, from one-half to two-thirds of the ovarian blood was derived from the uterine artery, and the ovarian artery never contributed to uterine blood flow. These results indicate that the major supply vessels are unlikely to limit ovarian blood flow, even near term when competing demand by the gravid uterus reaches a peak. The finding that ovarian blood flow is derived predominantly from the uterine artery may relate to local mechanisms that influence ovarian function and fetal growth.

Fluorescence in situ hybridization establishes the order cen-DXS28(C7)-DXS67(B24)-DXS68(L1)-tel in human chromosome Xp21.3.

We report here on the order of three DNA markers, C7, B24, and L1, based on the arrangement of their fluorescently labeled hybridization sites in interphase cell nuclei. The three markers map distal to the Duchenne muscular dystrophy (DMD), glycerol kinase deficiency (GKD), and adrenal hypoplasia (AHC) loci on human chromosome Xp21.3. Their order has been a matter of controversy. In interphase chromatin, B24 maps between C7 and L1. We estimate from interphase distance that C7 and L1 are 300-500 kb apart. When the three markers are hybridized to interphase cells of Nijmegen1, a patient with DMD, GKD, and AHC, only C7 appears to be deleted, rather than both C7 and L1, as had been reported elsewhere. C7 is also the only one of the three markers deleted in several other DMD patients studied by others. The deletion results indicate that C7 is the most proximal of the three markers and allow the trio of ordered probes to be oriented on the chromosome: cen-C7(DXS28)-B24(DXS67)-L1(DXS68)-tel.

Mapping of human chromosome Xq28 by two-color fluorescence in situ hybridization of DNA sequences to interphase cell nuclei.

We have used the proximity of probe hybridization sites in interphase chromatin to derive the order of DNA sequences in a 2-3-Mbp region of human chromosome Xq28. The map generated bridges the results of genetic and pulsed-field gel electrophoresis mapping to produce a more complete map of Xq28 than possible with either of these other techniques alone. Two-color fluorescence in situ hybridization (FISH) was used to detect the positions of two or more probes in G1 male interphase nuclei. We show that cosmids that are 50 kbp to 2-3 Mbp apart can be ordered rapidly with two alternative approaches: (1) by comparing the average measured distance between two probes and (2) simply by scoring the order of red and green fluorescent dots after detection of three or more probes with two fluorochromes. The validity of these approaches is demonstrated using five cosmids from a region spanning approximately 800 kbp that includes the factor VIII (F8), glucose-6-phosphate dehydrogenase (G6PD), and color-vision pigment (CV) genes. The cosmid map derived from interphase mapping is consistent with the map determined by restriction-fragment analysis. The two interphase mapping approaches were then used (1) to orient the F8/CV cluster relative to two markers, c1A1 and st14c, which we show by metaphase mapping to be proximal to the F8/CV cluster, (2) to position st14c (DXS52) between c1A1 and F8, and (3) to orient the CV gene cluster relative to G6PD by using two CV-flanking cosmids, 18b41 and fr7. The probe order in Xq28 derived from interphase proximity is cen-c1A1-st14c-5'F8 (p624-p542-p625)-G6PD-18b41-3' green-green-red-fr7-tel. We also show that, to determine their order by using metaphase chromosomes, sequences must be at least 1 Mbp apart, an order of magnitude greater than required in interphase chromatin. The data show that FISH mapping is a simple way to order sequences separated by greater than or equal to 50 kbp for the construction of long-range maps of mammalian genomes.

Identification and localization of the gene for EXTL, a third member of the multiple exostoses gene family.

Hereditary multiple exostoses (EXT) is an autosomal dominant disorder characterized by multiple bony outgrowths from the juxtaepiphyseal region of long bones. In a small proportion of cases, these exostoses progress to malignant chondrosarcomas. Genetic linkage of this disorder has been described to three independent loci on chromosomes 8q24.1 (EXT1), 11p11-13 (EXT2), and 19p (EXT-3). The EXT1 and EXT2 genes were isolated recently and show extensive sequence homology to each other. These genes are deleted in exostoses-derived tumors, supporting the hypothesis that they encode tumor suppressors. We have identified a third gene that shows striking sequence similarity to both EXT1 and EXT2 at the nucleotide and amino acid sequence levels, and have derived its entire coding sequence. Although the mRNA transcribed from this gene is similar in size to that from EXT1 and EXT2, its pattern of expression is quite different. We have localized this gene by fluorescence in situ hybridization to metaphase chromosomes and by whole genome radiation hybrid mapping to chromosome 1p36.1 between DIS458 and DIS511, region that frequently shows loss of heterozygosity in a variety of tumor types. This gene, EXTL (for EXT-like), is therefore a new member of the EXT gene family and is a potential candidate for several disease phenotypes.

Gene structure, sequence, and chromosomal localization of the human red cell-type low-molecular-weight acid phosphotyrosyl phosphatase gene, ACP1.

Two distinct isoenzymes of the human red cell-type acid phosphatase (RCAP) have been known to exist for some time, but the genetic basis of this phenomenon was uncertain. We previously reported the isolation and characterization of two cDNA clones for human RCAP. We showed that the coding regions of the two cDNAs for the human isoenzymes were identical except for a divergent segment spanning nucleotides 176-274, called the variable region. We have now cloned, characterized, and mapped the gene encoding the red cell-type acid phosphatase isoenzymes. The human ACP1 gene is shown to span about 18 kb and to consist of seven exons. The promoter region of ACP1 is very GC-rich and has no apparent TATA or CCAAT boxes. The sequence information confirms that the variable regions of the isoenzymes exist in the gDNA sequence as separate and distinct exons, apparently subject to mutually exclusive alternative splicing. Using a genomic ACP1 clone, we have established the chromosomal localization of the gene to the distal portion of 2p25 by fluorescence in situ hybridization.

Comparative mapping of the region of human chromosome 7 deleted in williams syndrome.

Williams syndrome (WS) is a complex developmental disorder resulting from the deletion of a large (approximately 1.5-2 Mb) segment of human chromosome 7q11.23. Physical mapping studies have revealed that this deleted region, which contains a number of known genes, is flanked by several large, nearly identical blocks of DNA. The presence of such highly related DNA segments in close physical proximity to one another has hampered efforts to elucidate the precise long-range organization of this segment of chromosome 7. To gain insight about the structure and evolutionary origins of this important and complex genomic region, we have constructed a fully contiguous bacterial artificial chromosome (BAC) and P1-derived artificial chromosome (PAC) contig map encompassing the corresponding region on mouse chromosome 5. In contrast to the difficulties encountered in constructing a clone-based physical map of the human WS region, the BAC/PAC-based map of the mouse WS region was straightforward to construct, with no evidence of large duplicated segments, such as those encountered in the human WS region. To confirm this difference, representative human and mouse BACs were used as probes for performing fluorescence in situ hybridization (FISH) to metaphase and interphase chromosomes. Human BACs derived from the nonunique portion of the WS region hybridized to multiple, closely spaced regions on human chromosome 7q11.23. In contrast, corresponding mouse BACs hybridized to a single site on mouse chromosome 5. Furthermore, FISH analysis revealed the presence of duplicated segments within the WS region of various nonhuman primates (chimpanzee, gorilla, orangutan, and gibbon). Hybridization was also noted at the genomic locations corresponding to human chromosome 7p22 and 7q22 in human, chimpanzee, and gorilla, but not in the other animal species examined. Together, these results indicate that the WS region is associated with large, duplicated blocks of DNA on human chromosome 7q11.23 as well as the corresponding genomic regions of other nonhuman primates. However, such duplications are not present in the mouse.

Stimulation of p53-mediated transcriptional activation by the p53-binding proteins, 53BP1 and 53BP2.

p53 is a tumor suppressor protein that controls cell proliferation by regulating the expression of growth control genes. In a previous study, we identified two proteins, 53BP1 and 53BP2, that are able to bind to wild type but not to mutant p53 via the DNA-binding domain of p53. We isolated cDNAs expressing a full-length human 53BP1 clone, which predicts a protein of 1972 residues that can be detected in the H358 human lung carcinoma cell line. The 53BP1 and 53BP2 genes were mapped to chromosomes 15q15-21 and 1q41-42, respectively. Immunofluorescence studies showed three types of staining patterns for 53BP1 as follows: both cytoplasmic and nuclear, homogeneous nuclear, and a nuclear dot pattern. In contrast, 53BP2 localized exclusively to the cytoplasm, and this pattern did not change upon coexpression of wild type p53. Although our previous study revealed that p53 is not able to bind simultaneously to either 53BP1 or 53BP2 and to DNA carrying a consensus binding site, both 53BP1 and 53BP2 enhanced p53-mediated transcriptional activation and induced the expression of a p53-dependent protein, suggesting that these proteins might function in signal transduction pathways to promote p53 activity.

Studies of metaphase and interphase chromosomes using fluorescence in situ hybridization.

Our measurements have bearing on the resolution with which maps can be constructed and abnormalities can be detected by studying the proximity of DNA sequences in metaphase and interphase chromosomes. The results of our analyses are summarized in Figure 8. Metaphase chromosomes are compacted sufficiently that it is impractical to order sequences separated by less than approximately 1 Mbp. In contrast, 100-kbp resolution can be obtained in interphase chromosomes. Distance measurements reveal that interphase chromatin behaves as a random polymer over distances up to 1-2 Mbp. At greater distances, higher order constraints, perhaps the dimensions of the individual chromosome domains, come into play. A caveat remains: Because the effect of the FISH procedure on native chromosome organization is not well understood, these conclusions may not be applicable to native chromatin. We have illustrated that FISH, with appropriately chosen probes, can supplement conventional cytogenetics in the study of chromosome abnormalities. The technique is increasingly being applied in research laboratories to detect and characterize chromosome abnormalities and point the way to the location of genes involved in human disease.

Characterization of somatic cell hybrids by bivariate flow karyotyping and fluorescence in situ hybridization.

We report on the use of flow karyotyping and fluorescence in situ hybridization (FISH) to characterize the human chromosomes in somatic cell hybrids. The identity, DNA content, and relative frequency of human chromosomes are derived from flow karyotypes, i.e., measurements of Hoechst and chromomycin fluorescence intensities of chromosomes by dual beam flow cytometry. Chromosome integrity is assessed by comparing the peak position of a human chromosome in the flow karyotypes of a hybrid cell line and its human donor. When human donor cells are unavailable, the peak position of a human chromosome in a hybrid line is compared to the range of peak positions among normal individuals. The relative frequency of human chromosomes in subclones or hybrids grown in culture is monitored using the volumes of peaks in flow karyotypes. FISH with biotinylated human genomic DNA or chromosome-specific repeat sequence as probe is used in conjunction with flow karyotyping to confirm the number of human chromosomes in hybrids. Some small rearrangements are detected by flow karyotyping and not by FISH. On the other hand, translocations between human and rodent chromosomes are detected by FISH and not always by flow karyotyping. Flow karyotyping and FISH were used to characterize over 100 hybrid lines donated by other laboratories. A hybrid set useful for the construction of chromosome-enriched gene libraries is presented. In this set, each of the 24 human chromosome types is present and intact, as judged by these techniques, in a line containing little or no other human material.

Segmental duplications: organization and impact within the current human genome project assembly.

Segmental duplications play fundamental roles in both genomic disease and gene evolution. To understand their organization within the human genome, we have developed the computational tools and methods necessary to detect identity between long stretches of genomic sequence despite the presence of high copy repeats and large insertion-deletions. Here we present our analysis of the most recent genome assembly (January 2001) in which we focus on the global organization of these segments and the role they play in the whole-genome assembly process. Initially, we considered only large recent duplication events that fell well-below levels of draft sequencing error (alignments 90%-98% similar and > or =1 kb in length). Duplications (90%-98%; > or =1 kb) comprise 3.6% of all human sequence. These duplications show clustering and up to 10-fold enrichment within pericentromeric and subtelomeric regions. In terms of assembly, duplicated sequences were found to be over-represented in unordered and unassigned contigs indicating that duplicated sequences are difficult to assign to their proper position. To assess coverage of these regions within the genome, we selected BACs containing interchromosomal duplications and characterized their duplication pattern by FISH. Only 47% (106/224) of chromosomes positive by FISH had a corresponding chromosomal position by comparison. We present data that indicate that this is attributable to misassembly, misassignment, and/or decreased sequencing coverage within duplicated regions. Surprisingly, if we consider putative duplications >98% identity, we identify 10.6% (286 Mb) of the current assembly as paralogous. The majority of these alignments, we believe, represent unmerged overlaps within unique regions. Taken together the above data indicate that segmental duplications represent a significant impediment to accurate human genome assembly, requiring the development of specialized techniques to finish these exceptional regions of the genome. The identification and characterization of these highly duplicated regions represents an important step in the complete sequencing of a human reference genome.

A murine ortholog of the human serpin SCCA2 maps to chromosome 1 and inhibits chymotrypsin-like serine proteinases.

Squamous cell carcinoma antigens (SCCA) 1 and 2 are inhibitory members of the high-molecular-weight serine proteinase inhibitor (serpin) family. The biological functions of SCCA1 and 2 are unknown. One approach to determining the function of human proteins is to study orthologs in other species, such as the mouse. The purpose of this study was to determine whether orthologs to human SCCA1 or 2 exist in the mouse. We report the identification and characterization of a novel serpin, sqn5 (now designated Scca2). Comparative amino acid sequence analysis suggests that Scca2 is a member of the ov-serpin subfamily of serpins with highest homology to SCCA1 and SCCA2. Fluorescence in situ hybridization revealed that the Scca2 mapped near Bcl2 on mouse chromosome 1. This region is syntenic with the human locus for SCCA1 and SCCA2 on 18q21.3. The tissue expression patterns as determined by RT-PCR showed a restricted distribution. Scca2 was detected in the lung, thymus, skin, and uterus, as are SCCA1 and SCCA2. Unlike the SCCAs, however, Scca2 was detected also in the gastrointestinal tract. Enzyme-inhibition assays using a GST-SCCA2 fusion protein revealed that SCCA2 inhibited chymotrypsin-like serine proteinases, but not papain-like cysteine proteinases. SCCA2 inhibited CTSG at 1:1 stoichiometry and with a second-order rate constant of kass = 1.7 x 10(5) M-1 s-1. SCCA2 also inhibited human mast cell chymase but the stoichiometry was 2:1, and the second-order rate constant was kass = 0.9 x 10(4) M-1 s-1. This inhibitory profile is identical to that observed for human SCCA2. Based on these findings, Scca2 appears to be the murine ortholog of human SCCA2.

Cosmids map two incontinentia pigmenti type 1 (IP1) translocation breakpoints to a 180-kb region within a 1.2-Mb YAC contig.

Incontinentia pigmenti (IP) is an X-linked dominant disorder of neuroectodermal development. Based on the observation of six unrelated females with clinical features of nonfamilial IP with constitutional de novo reciprocal X;autosome translocations, a putative incontinentia pigmenti type 1 locus (IP1; MIM No. 308300) was localized to region Xp11.21. Using available regional DNA markers, we constructed a yeast artificial chromosome (YAC) contig that contained 1.2 Mb of distal Xp11.21 and spanned two IP1 X-chromosomal breakpoints. This contig was used to generate a detailed molecular map of the region and identify three regional CpG islands. YAC-derived cosmids were used to clone and map the IP1 breakpoints to a 180-kb interval that was flanked by DNA markers DXS705 and DXS741. The physical map and genomic clones should facilitate the isolation and characterization of transcripts associated with the IP1 translocation breakpoints.