Identification of endoglin as a functional marker that defines long-term repopulating hematopoietic stem cells.
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We describe a strategy to obtain highly enriched long-term repopulating (LTR) hematopoietic stem cells (HSCs) from bone marrow side-population (SP) cells by using a transgenic reporter gene driven by a stem cell enhancer. To analyze the gene-expression profile of the rare HSC population, we developed an amplification protocol termed "constant-ratio PCR," in which sample and control cDNAs are amplified in the same PCR. This protocol allowed us to identify genes differentially expressed in the enriched LTR-HSC population by oligonucleotide microarray analysis using as little as 1 ng of total RNA. Endoglin, an ancillary transforming growth factor beta receptor, was differentially expressed by the enriched HSCs. Importantly, endoglin-positive cells, which account for 20% of total SP cells, contain all the LTR-HSC activity within bone marrow SP. Our results demonstrate that endoglin, which plays important roles in angiogenesis and hematopoiesis, is a functional marker that defines LTR HSCs. Our overall strategy may be applicable for the identification of markers for other tissue-specific stem cells. (+info)
Differential expression profiling of head and neck squamous carcinoma: significance in their phenotypic and biological classification.
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The genetic events associated with the development and progression of head and neck squamous carcinoma (HNSC) are largely unknown. We analysed 12 matched pairs of histologically normal squamous mucosa and tumor specimens from six conventional and six phenotypic variants HNSC to define the differentially expressed genes in these tumors. Parallel expression analysis of 8055 unique genes was performed, and the level of the hybridization signal for each gene was measured after normalization. Hierarchical cluster analysis of the expressed genes showed distinct inter- and intra-tumoral patterns in and between conventional squamous carcinoma and squamous carcinoma variants. We also identified 26 (0.32%) differentially expressed genes that were consistently different between matched pairs of normal and tumor specimens; a selected set of the overexpressed genes was validated using real-time quantitative RT-PCR. The majority of the genes were associated with differentiation and proliferation. Our study defines a set of genes that could form the basis for the construction of limited HNSC targeted expression array and in-depth studies and further highlights gene profile differences that may be useful in pathobiologic classification of HNSC. (+info)
Altered mRNA expression of Pax5 and Blimp-1 in B cells in multiple myeloma.
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Multiple myeloma (MM) is a plasma cell disorder that potentially initiates during an early stage of B-cell development. We encountered an unidentified isoform of B cell-specific activator protein (BSAP, or Pax5) in MM cells while performing differential analyses to compare mRNA expression in malignant and normal plasma cells. Pax5 is a transcription factor that plays a central role throughout B-cell development until the point of terminal differentiation. Our finding of this unique isoform prompted us to investigate Pax5 isoform usage in plasma cells and B-cell populations in other MM and healthy subjects. In contrast to normal Pax5 expression, we observed multiple isoforms of Pax5 in conjunction with low levels of expression of the full-length Pax5 in B cells from MM patients. The expressed isoforms in MM varied considerably from patient to patient, with no clear pattern. We also performed semiquantitative analyses of the mRNA expression levels of B lymphocyte-induced maturation protein (Blimp-1), because expression levels of Pax5 and Blimp-1 have been shown to be inversely correlated. We observed the expression of Blimp-1 in the B-cell populations in all 11 MM patients but in none of 11 healthy subjects. We hypothesize that premature Blimp-1 expression coupled to altered and deficient Pax5 expression causes some proliferating B cells to prematurely differentiate to plasma cells in MM. (+info)
Identification of genes regulated by 2-methoxyestradiol (2ME2) in multiple myeloma cells using oligonucleotide arrays.
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Our previous study demonstrated that 2-methoxyestradiol (2ME2), an estrogen derivative, induces apoptosis in multiple myeloma (MM) cells; however, the related transcriptional events are unclear. In the present study, we used oligonucleotide microarrays to identify genes altered during 2ME2-induced apoptosis in MM cells. 2ME2 triggers an early transient induction of genes known to trigger cell death and repression of growth/survival-related genes. Many genes regulating cell defense/repair machinery also were transiently induced. Since 2ME2 also induces apoptosis in MM cells resistant to conventional therapies such as dexamethasone (Dex), we compared the gene profiles of 2ME2-treated and Dex-resistant MM cells. Our results suggest that 2ME2 overcomes Dex resistance by modulating genes that confer chemoresistance in MM cells. Microarray results were confirmed by Northern and Western blot analyses. A comparative analysis of selected genes from freshly isolated MM patient cells and 2ME2-treated MM.1S MM cells further provides an in vivo relevance of our in vitro studies. Collectively, these findings suggest genetic events mediating anti-MM activity of 2ME2, as well as mechanisms whereby 2ME2 overcomes Dex resistance in MM cells. These studies may therefore allow improved therapeutic use of 2ME2, based upon targeting genes that regulate MM cell growth and survival. (+info)
Noninvasive diagnosis of partial atrial standstill using magnetocardiograms.
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A 59-year-old woman with partial atrial standstill was studied using magnetocardiograms (MCGs), which revealed through QRS-T subtraction and time-frequency analysis that there was a high-frequency (6 Hz) magnetic source at the low atrial septum. MCGs are useful for noninvasively evaluating the clinical course of patients with atrial fibrillation. (+info)
Uptake in supraclavicular area fat ("USA-Fat"): description on 18F-FDG PET/CT.
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The supraclavicular region is a common site for lymph node metastases. A commonly reported type of nonmalignant (18)F-FDG uptake on PET imaging in the supraclavicular region is "muscle uptake" purportedly due to muscle contraction in tense patients during the (18)F-FDG uptake phase. PET/CT offers the unique opportunity to correlate PET findings with CT anatomy in the supraclavicular region. METHODS: Images from the first 359 consecutive clinical whole-body studies (in 347 patients) using (18)F-FDG and a PET/CT scanner (with CT attenuation correction and ordered-subsets expectation maximization [OSEM] reconstruction) were retrospectively reviewed. The supraclavicular region was evaluated for the presence of abnormal uptake on PET images, and the corresponding CT findings were assessed. Three distinct patterns of abnormal (18)F-FDG uptake were noted: pattern A (uptake localizing to supraclavicular area fat [USA-fat], i.e., without corresponding lymph node or muscle uptake on CT), pattern B (uptake localizing to muscle on CT), and pattern C (uptake localizing to lymph nodes or soft-tissue masses on CT). RESULTS: Forty-nine patients (14.1%) (32 female, 17 male; mean age, 51.4 +/- 15.6 y; age range, 12-77 y) showed abnormal (18)F-FDG uptake in the supraclavicular region. Twenty patients (5.8%) had muscle uptake (group B); 15 (4.3%) had definite abnormal lymph nodes (group C). However, 14 patients (4.0%) had USA-fat (group A) and foci of very low Hounsfield units on CT. These foci were also present on (68)Ge attenuation-corrected images (when obtained) and non-attenuation-corrected images. Uptake in USA-fat was typically bilateral and symmetric, intense, more often multifocal than linear, and located in fat on PET/CT. Age was not significantly different for group C versus the 2 other groups. Intensity; mean standardized uptake value, lean (SUV(L MEAN)); or maximum standardized uptake value, lean (SUV(L MAX)), did not allow differentiation between patterns A and C (P > 0.05). Standardized uptake values (SUV(L MAX), 3.1; SUV(L MEAN), 2.1) were significantly lower in group B than in the 2 other groups (P < 0.005). CONCLUSION: So-called muscle uptake in the supraclavicular region may be caused in a significant proportion of cases by an unrelated process we call the USA-fat finding, with (18)F-FDG uptake in tissues of low-Hounsfield (fat) density. This finding most likely reflects an underlying nonpathologic process that we hypothesize to be in foci of brown fat. This intense supraclavicular uptake should be recognized and should not be misinterpreted as a malignant metastatic process or as muscle uptake. (+info)
Clinically significant inaccurate localization of lesions with PET/CT: frequency in 300 patients.
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This study evaluated lesion mislocalization between PET and CT on PET/CT studies when CT instead of germanium is used for attenuation correction (AC). METHODS: PET/CT scans were obtained for 300 clinical patients. Both CT and germanium scans were used to correct PET emission data. Cases were noted of suspected inaccurate localization of lesions on any of the 5 sets of images (PET using germanium AC [GeAC] fused and not fused with CT, PET using CT AC fused and not fused with CT, and PET with no AC [NAC]). Independent CT or MRI was used to determine true lesion locations. RESULTS: Six of 300 patients (2%) had lesion mislocalization when CT was used for AC or fusion. True liver dome lesions were mislocalized to the right lung base on PET/CT, likely because of a respiratory motion difference between PET and CT. No mislocalization was present on NAC PET or non-CT-fused GeAC PET images. CONCLUSION: Serious lesion mislocalization on PET/CT studies may occur, albeit very infrequently, when CT is used for either AC or fusion. (+info)
Subtraction helical CT angiography of intra- and extracranial vessels: technical considerations and preliminary experience.
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BACKGROUND AND PURPOSE: Reported CT angiographic (CTA) subtraction methods are not simple, robust, or real time. We investigated a novel technique for semiautomated digital subtraction CTA of the intracranial and extracranial arteries. METHODS: Thirty patients underwent precontrast (low milliampere-seconds) and postcontrast (pitch, 1.5; collimation, 1-2.5 mm) helical imaging with a vacuum-type head holder to facilitate image registration and minimize movement. A reconstructed three-dimensional model of the precontrast bone dataset was subtracted from the postcontrast dataset to produce subtracted maximum-intensity-projection angiograms. Experienced (operator 1) and less-experienced (operator 2) staff performed the standard and subtraction reconstructions, and image generation time and quality (graded 1-5) were compared. A third operator blinded to the method assessed the hard-copy image quality. RESULTS: Image quality with subtraction postprocessing was significantly better with both operators (operator 1, mean improvement of 0.87 grade, median improvement of 1 grade, P <.001; operator 2, mean improvement of 0.63 grade, median improvement of 1 grade, P <.001). Hard-copy image quality was better with the subtraction method (operator 1, P >.001; operator 2, P <.001). Blood vessels at the base of the brain were better demonstrated on subtraction images in 13 of 14 examinations. For the less experienced operator, the reconstruction time was significantly less with the subtraction method than with the conventional method (mean, 7.5 vs 10.1 minutes; P =.001). CONCLUSION: When separation of the vasculature from bone is important and technically difficult, digital subtraction CTA offers a potential advantage. This semiautomated technique is fast and easy to learn, and variably experienced staff can use it. (+info)