Integrated YAC Contig Map of the Prader–Willi/Angelman Region on Chromosome 15q11–q13 with Average STS Spacing of 35 kb

Prader–Willi syndrome and Angelman syndrome are associated with parent-of-origin-specific abnormalities of chromosome 15q11–q13, most frequently a deletion of an ~4-Mb region. Because of genomic imprinting, paternal deficiency of this region leads to PWS and maternal deficiency to AS. Additionally, this region is frequently involved in other chromosomal rearrangements including duplications, triplications, or supernumerary marker formation. A detailed physical map of this region is important for elucidating the genes and mechanisms involved in genomic imprinting, as well as for understanding the mechanism of recurrent chromosomal rearrangments. An initial YAC contig extended from D15S18 to D15S12 and was comprised of 23 YACs and 21 STSs providing an average resolution of about one STS per 200 kb. To close two gaps in this contig, YAC screening was performed using two STSs that flank the gap between D15S18 and 254B5R and three STSs located distal to the GABRA5–149A9L gap. Additionally, we developed 11 new STSs, including seven polymorphic markers. Although several groups have developed whole-genome genetic and radiation hybrid maps, the depth of coverage for 15q11–q13 has been somewhat limited and discrepancies in marker order exist between the maps. To resolve the inconsistencies and to provide a more detailed map order of STSs in this region, we have constructed an integrated YAC STS-based physical map of chromosome 15q11–q13 containing 118 YACs and 118 STSs, including 38 STRs and 49 genes/ESTs. Using an estimate of 4 Mb for the size of this region, the map provides an average STS spacing of 35 kb. This map provides a valuable resource for identification of disease genes localized to this region as well as a framework for complete DNA sequencing.     

Molecular dissection of the Prader-Willi/Angelman syndrome region (15q11-13) by YAC cloning and FISH analysis.

Prader-Willi syndrome (PWS) and Angelman syndrome (AS) are distinct mental retardation disorders associated with deletions of proximal 15q (q11-q13) of different parental origin. Yeast artificial chromosome (YAC) clones were isolated for 9 previously mapped DNA probes from this region, and for one newly derived marker, LS6-1 (D15S113). A YAC contig of 1-1.5 Mb encompassing four markers (ML34, IR4-3R, PW71, and TD189-1) was constructed. Multi-color fluorescence in situ hybridization (FISH) analysis of interphase nuclei was combined with YAC contig information to provide the following order of markers: cen-IR39-ML34-IR4-3R-PW71-TD189-1-LS6++ +-1-TD3-21-GABRB3-IR10-1-CMW1-tel. FISH analysis was performed on 8 cases of PWS and 3 cases of AS, including 5 patients with normal karyotypes. All eleven patients were deleted for YACs in the interval from IR4-3R to GABRB3. On the proximal side of the deletion interval, 10/10 breakpoints fell within a single ML34 YAC of 370 kb. On the distal side, 8/9 breakpoints fell within a single IR10-1 YAC of 200 kb. These results indicate a striking consistency in the location of the proximal and distal breakpoints in PWS and AS patients. FISH analysis on a previously reported case of familial AS confirmed a submicroscopic deletion including YACs corresponding to LS6-1, TD3-21 and GABRB3 and supports the separation of the PWS and AS critical regions. Since these three YACs do not overlap each other, the minimum size of the AS critical region is > or = 650 kb.     

Molecular characterisation of four cases of intrachromosomal triplication of chromosome 15q11-q14

CONTEXT—Chromosomal abnormalities that involve the proximal region of chromosome 15q occur relatively frequently in the human population. However, interstitial triplications involving one 15 homologue are very rare with three cases reported to date.
OBJECTIVE—To provide a detailed molecular characterisation of four additional patients with interstitial triplications of chromosome 15q11-q14.
DESIGN—Molecular analyses were performed using DNA markers and probes specific for the 15q11-q14 region.
SETTING—Molecular cytogenetics laboratory at the University of Chicago.
SUBJECTS—Four patients with mild to severe mental retardation and features of Prader-Willi syndrome (PWS) or Angelman syndrome (AS) were referred for molecular cytogenetic analysis following identification of a suspected duplication/triplication of chromosome 15q11-q14 by routine cytogenetic analysis.
MAIN OUTCOME MEASURES—Fluorescence in situ hybridisation (FISH) was performed to determine the type of chromosomal abnormality present, the extent of the abnormal region, and the orientation of the extra chromosomal segments. Molecular polymorphism analysis was performed to determine the parental origin of the abnormality. Methylation and northern blot analyses of the SNRPN gene were performed to determine the effect of extra copies of the SNRPN gene on its methylation pattern and expression.
RESULTS—Fluorescence in situ hybridisation (FISH) using probes within and flanking the Prader-Willi/Angelman syndrome critical region indicated that all patients carried an intrachromosomal triplication of proximal 15q11-q14 in one of the two chromosome 15 homologues (trip(15)). In all patients the orientation of the triplicated segments was normal-inverted-normal, suggesting that a common mechanism of rearrangement may have been involved. Microsatellite analysis showed the parental origin of the trip(15) to be maternal in three cases and paternal in one case. The paternal triplication patient had features similar to PWS, one maternal triplication patient had features similar to AS, and the other two maternal triplication patients had non-specific findings including hypotonia and mental retardation. Methylation analysis at exon 1 of the SNRPN locus showed increased dosage of either the paternal or maternal bands in the paternal or maternal triplication patients, respectively, suggesting that the methylation pattern shows a dose dependent increase that correlates with the parental origin of the triplication. In addition, the expression of SNRPN was analysed by northern blotting and expression levels were consistent with dosage and parental origin of the triplication.
CONCLUSIONS—These four additional cases of trip(15) will provide additional information towards understanding the phenotypic effects of this abnormality and aid in understanding the mechanism of formation of other chromosome 15 rearrangements.


Keywords: chromosome 15 triplication; Prader-Willi syndrome; Angelman syndrome; autism     

Telomerase activity and human papillomavirus in malignant, premalignant and benign cervical lesions.

The purpose of this study was to define a correlation between telomerase activity and human papillomavirus (HPV) in normal control tissue and in benign, premalignant and malignant cervical lesions. Telomerase activity was detectable in 33 out of 34 cases of squamous-cell carcinoma, five out of six cases of microinvasive carcinoma, 8 out of 20 cases and two out of six cases of high- and low-grade squamous intraepithelial lesions (SILs) respectively. The higher frequency of positive telomerase in invasive carcinoma compared with SILs was observed in both HPV-associated and non-associated groups. Whereas 92.6% of HPV-positive and 100% of HPV-negative invasive lesions expressed telomerase, only 50% of HPV-positive and 25% of HPV-negative SILs did. Interestingly, telomerase activity was also detectable in 13 out of 28 cases of benign lesions regardless of the presence of HPV. In conclusion, there may be two roles of telomerase in the cervix. The first one would present in benign lesions; the second is associated with cancer development and activated during the late stage of multistep carcinogenesis in both HPV-positive and -negative groups.     

Borderline phyllodes tumor arising in accessory breast tissue at the axilla

Accessory breast tissue is an anatomical variation which occurs during embryogenic development. It appears most frequently at the axilla. Benign and malignant processes in general breast tissue can occur in accessory breast tissue. We report a case of 76-year-old female presented with palpable, huge mass at the right axilla which pathology of the mass was borderline phyllodes tumor. Phyllodes tumors arising in accessory breast tissue is an extremely rare condition. And this case study showed more detail on phyllodes tumor which would encourage the advance in management of the disease.     

The validity and reliability of the Thai version of the Kujala score for patients with patellofemoral pain syndrome

Purpose: To develop a Thai version of the Kujala score and show the evaluation of the validity and reliability of the score.

Method: The Thai version of the Kujala score was developed using the forward–backward translation protocol. The 49 PFPS patients answered the Thai version of questionnaires including the Kujala score, Short Form-36 (SF-36) and International Knee Documentation Committee (IKDC) Subjective Knee Form. The validity between the scores has been tested. The reliability was assessed using test–retest reliability and internal consistency.

Results: The Thai version of the Kujala score showed a good correlation with Thai IKDC Subjective Knee Form (Pearson’s correlation coefficient; r?=?0.74: p?r?=?0.586, 0.571 and 0.524, respectively: p?p?p?Conclusion: The Thai version of the Kujala score has shown good validity and reliability. This score can be effectively used for evaluating Thai patients with patellofemoral pain syndrome.

• Implications for Rehabilitation

• The Kujala score is a self-administered questionnaire for patients with patellofemoral pain syndrome (PFPS).

• The validity and reliability of the Thai version of Kujala are compatible with other versions (Turkish, Chinese and Persian version).

• The Thai version of Kujala has been shown to have validity and reliability in Thai PFPS patients and can be used for clinical evaluation and also in the research work.

     

Epidemiologic Analysis of Proximal Deep Vein Thrombosis in Thai Patients: Malignancy,the Predominant Etiologic Factor

From January 1995 to December 2000, 1555 patients (585 males, 970 females, with a mean age of 52.3 years) with acute limb swelling underwent diagnostic duplex venous ultrasonagraphy at the Vascular Surgery Unit, Department of Surgery, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok, Thailand. Of these 1555 patients, 514 (188 males, 326 females, with a mean age of 53 years) were diagnosed with deep vein thrombosis (DVT). DVT was found mainly in the lower extremities, occurring in 506 of these 514 cases (98.4%). The affected vein in the lower extremity was found more commonly in the left side (359 limbs) than in the right side (184 limbs) (p < 0.001). DVT also occurred in the upper extremities in 8 of the 514 cases (1.6%). In this study, out of 308 patients with DVT in their lower limbs, DVT was associated with malignancy in 123 cases (39.9%), with postoperation in 48 cases (15.6%), and with limb immobilization in 38 cases (12.3%). Malignancy-related DVT was found to be more extensive and virulent and was associated with high morbidity and mortality. Further study should be conducted to investigate the role of anticoagulant drugs for the prevention of DVT in cancer patients. Perhaps the spectrum of DVT as presented in this series is characteristic not only of the Thai population but also of the populations of other Asian countries.     

Contiguous gene syndrome of holoprosencephaly and hypotrichosis simplex: association with an 18p11.3 deletion

We report a patient with a unique combination of features, including microcephaly; mental retardation; poorly developed frontal lobes; hypoplastic pituitary gland; hypothyroidism; alopecia universalis; single maxillary central incisor; taurodontism; median palatal ridge; longitudinally grooved nails; and scoliosis. His unbalanced karyotype was found to be 45,XY,der(15;18)(q10;q10). The constellation of anomalies appears to represent a contiguous gene syndrome caused, at least in part, by deletion of TGIF and the gene responsible for hereditary hypotrichosis simplex. The phenotype of our patient differs other reported patients with del(18p). Possible explanations include (1) the effects of a different deleted region, (2) a positional effect caused by a gene close by, or (3) by interruption of a different gene resulting from chromosomal translocation.     

Distribution of the Epstein-Barr virus in the normal stomach and gastric lesions in Thai population

The Epstein-Barr virus (EBV) is one of the infectious agents found in stomach tissue. Recently, EBV-associated gastric carcinoma (EBVaGC) was classified as a new subtype of gastric carcinoma. To date, there is a lack of knowledge about the distribution and prevalence of EBV infection in both the normal stomach and various gastric lesions, including EBVaGC, in the Thai population. In this study, we detected EBV in the normal stomach (NS; n = 19), chronic gastritis (CG; n = 36), intestinal metaplasia (IM; n = 40), gastric dysplasia (GD; n = 15), and gastric adenocarcinoma (GC; n = 33) by polymerase chain reaction (PCR) amplification of the latent membrane protein (LMP1) gene of EBV. EBV-PCR amplification was positive in 42.1%, 36.1%, 22.5%, 13.3%, and 33.3% of NS, CG, IM, GD, and GC, respectively. For further clarification in EBVaGC, we performed EBV-encoded small RNA in situ hybridization (EBER-ISH) in PCR-positive cases of GD and GC. Four GC cases were EBER-ISH positive (12.1%), while both GD cases were EBER-ISH negative. In addition, we determined the distribution of the EBV strain (type A or B) based on EBNA3C sequence and EBV variants based on LMP1 variation (wild-type and 30-bp deletion variants; wt-LMP1 or del-LMP1). The results showed that type A and wt-LMP1 were the most prevalent in all lesions. In conclusion, EBV is common in both the NS and gastric lesions, and the frequency of EBVaGC was 12.1% in Thai patients.     

SHP-1 promoter 2 methylation in cerebrospinal fluid for diagnosis of leptomeningeal epithelial-derived malignancy (carcinomatous meningitis)

Current diagnostic methods for leptomeningeal metastasis (LM) from epithelial-derived malignancy (EDM) have limited sensitivity. Here, we explored SHP-1 promoter 2 methylation (SHP1P2)—an epithelial-specific methylation marker previously proven as risk stratification and potential diagnostic marker in non-small cell lung cancer—for EDM with LM. We prospectively recruited 136 patients who were diagnosed EDM with LM (n?=?25), EDM without LM (n?=?14), non-EDM with LM (n?=?8), and benign meningeal diseases (n?=?89). The primary cancer sites for EDM with LM were lung (n?=?17), breast (n?=?5), and colon (n?=?3). We performed quantitative analyses of cell-free (cfSHP1P2) and whole fraction (wSHP1P2) from cerebrospinal fluid (CSF); results were correlated with the clinicopathological data, including CSF cytology. Median cfSHP1P2 and wSHP1P2 were 3.08 [range: 0–163.5] and 9.35 [0.69–91.63] ng/ml, respectively, in EDM with LM; 0 [0–0.08] and 0.23 [0–7.84] ng/ml in EDM without LM; and were undetectable in most cases of benign meningeal diseases and non-EDM with LM. The cut-off values of 0.22 ng/ml for methylated cfSHP1P2 and 0.59 ng/ml for wSHP1P2 were the best to discriminate EDM with LM from EDM without LM (sensitivity: 79–100?%; specificity: 83–100?%), as well as from other benign conditions (sensitivity: 85–100?% specificity: 78–100?%). CSF cytology yielded 76?% sensitivity for diagnosing EDM with LM. Further validation of CSF SHP1P2 methylation detection as a role of adjunctive tool for LM from EDM should be interested based on our study.