A comparison of biotin and isotope-labeled ribonucleic acid probes for in situ detection of HPV-16 ribonucleic acid in genital precancers

Genital precancers were analyzed by in situ hybridization for the presence of HPV RNA using single-stranded RNA probes. To compare the sensitivity of biotin- and 35S-labeled probes, serial sections from three biopsies containing variable amounts of HPV 16 RNA were analyzed with probes containing each label. The probe template was a cloned fragment of the HPV 16 genome spanning portions of the E2, E5 and L2 orf's. When a focus contained a strong hybridization signal with 35S-labeled probes (i.e., target/background ratio greater than 40) serial sections of the same area usually demonstrated a strongly positive signal with biotin-labeled probes. When the signal with 35S in a given focus was weak (target/background ratio less than 20), serial sections contained either a very weak or no detectable signal with the biotinylated probe. From this study it appears than 35S-labeled RNA probes exceed the sensitivity of biotin-labeled probes by nearly an order of magnitude with short exposures (4 days), and with longer exposures (1 to 2 weeks), approach the sensitivity of that reported for tritiated probes and exposures of 1 month or longer.     

Neurobehavioral phenotype in carriers of the fragile X premutation

There have been contradictory findings in the fragile X (fraX) literature about possible neurocognitive and psychological symptoms due to the fraX premutation (pM). The purpose of the present study was to investigate the relationship between CGG repeat length and neurobehavioral functioning in carriers of the fraX pM. Eighty‐five female carriers of the pM with allele sizes ranging from 59–166 were administered a comprehensive IQ test (WAIS‐III) and completed a questionnaire designed to measure psychopathology (Symptom Checklist (SCL)‐90‐R). No relationship between allele size and cognition was identified. A significant negative relationship between allele size and age was found, as well as a positive relationship between allele size and depression. Follow‐up analyses separating small and large allele sizes (below and above 100 CGG repeats) indicated that individuals with larger allele sizes scored significantly higher on the Interpersonal Sensitivity and Depression subscales of the SCL‐90‐R. Despite the limitation of few individuals with high CGG repeat lengths, our findings suggest that females with larger premutated alleles (≥ 100 repeats) display some clinical manifestations of fraX syndrome. © 2001 Wiley‐Liss, Inc.     

The tissue microarray data exchange specification: A community-based,open source tool for sharing tissue microarray data

Background  

Tissue Microarrays (TMAs) allow researchers to examine hundreds of small tissue samples on a single glass slide. The information held in a single TMA slide may easily involve Gigabytes of data. To benefit from TMA technology, the scientific community needs an open source TMA data exchange specification that will convey all of the data in a TMA experiment in a format that is understandable to both humans and computers. A data exchange specification for TMAs allows researchers to submit their data to journals and to public data repositories and to share or merge data from different laboratories. In May 2001, the Association of Pathology Informatics (API) hosted the first in a series of four workshops, co-sponsored by the National Cancer Institute, to develop an open, community-supported TMA data exchange specification.     

Bursitis due to Mycobacterium goodii, a recently described, rapidly growing mycobacterium

We report a case of olecranon bursitis due to Mycobacterium goodii in a 60-year-old man. Prior to recognition of his infection, he received intrabursal steroids and underwent olecranon bursectomy. His infection was cured with antimicrobial therapy consisting of doxycycline and ciprofloxacin. This case illustrates that previously unrecognized members of the Mycobacterium smegmatis group of mycobacteria have pathogenic potential.     

Rat hepatic lipocytes express smooth muscle actin upon activation in vivo and in culture.

Myofibroblasts are mesenchymal cells that are prominent in liver injury. The origin of myofibroblasts in liver is debated, although morphologic evidence to date has suggested that these cells are derived from lipocytes (fat-storing cells, Ito cells). In the present study, we have utilized smooth muscle alpha actin antibody--a marker of myofibroblasts and smooth muscle cells--to study lipocytes in situ in normal and fibrotic rat liver as well as during their 'activation' in culture. Dual immunofluorescence studies on tissue sections from normal liver identified lipocytes as perisinusoidal desmin-positive, smooth muscle alpha actin-negative cells. In bile duct obstructed fibrotic liver, desmin-positive cells were numerous in areas of fibrosis and these cells also exhibited smooth muscle alpha actin. In carbon tetrachloride-induced fibrosis, cells expressing both desmin and smooth muscle alpha actin were present in fibrotic bands and in regenerating nodules. These results suggested that lipocytes had acquired characteristics of myofibroblasts during liver injury. To further address this issue we examined lipocytes immediately after isolation and also in primary culture. In freshly isolated lipocytes from normal liver, smooth muscle alpha actin was absent. In contrast, freshly isolated lipocytes from CCl4-treated animals expressed this smooth muscle marker immediately after isolation. In primary culture on plastic, lipocytes from normal liver began to express smooth muscle alpha actin coincident with culture-induced activation; at 14 days, smooth muscle alpha actin was identified in all cells. Electron microscopy demonstrated a highly developed array of microfilament bundles characteristic of actin filaments. Immunoblot of culture-activated lipocytes using the smooth muscle alpha actin antibody demonstrated the expected 42 kD protein (corresponding to the molecular size of smooth muscle alpha actin). Although smooth muscle alpha actin was readily detectable in culture-activated cells, it was not expressed in cells in which a quiescent phenotype was preserved by maintenance in culture on a laminin-rich gel. These findings demonstrate that the acquisition by lipocytes of a smooth muscle marker accompanies their 'activation', and are consistent with the hypothesis that lipocytes transform to myofibroblasts during liver injury.     

Neuroblastoma in a child with Wiedemann-Beckwith syndrome

We report on a patient with Wiedemann-Beckwith syndrome (WBS) who developed abdominal neuroblastoma. Although WBS patients are known to have a higher incidence of embryonal tumors, this is only the 4th known case of neuroblastoma associated with this syndrome. Chromosomes on peripheral lymphocytes and tumor cells were normal. Children with WBS should be screened for a variety of embryonal neoplasms, not only Wilms tumor.     

Viral integration and fragile sites in human papillomavirus-immortalized human keratinocyte cell lines.

Human papillomavirus (HPV) types 16 and 18 integration sites were mapped in six HPV-immortalized human keratinocyte cell lines by fluorescence in situ hybridization (FISH). Mapping of HPV sequences in these cell lines revealed that HPV integration varied in copy number and location but that integration sites were stable over extended passages in culture. Integration occurred at different sites throughout the genome and did not correspond to the location of specific cellular genes. However, integration sites were consistent with integration near or within known fragile sites in five of the six cell lines. Induction of aphidicolin-sensitive fragile sites in one cell line prior to in situ hybridization revealed that integrated HPV DNA was disrupted by fragile-site expression, suggesting that integration occurred within a fragile site.