Widespread mRNA association with cytoskeletal motor proteins and identification and dynamics of myosin-associated mRNAs in S. cerevisiae.

Programmed mRNA localization to specific subcellular compartments for localized translation is a fundamental mechanism of post-transcriptional regulation that affects many, and possibly all, mRNAs in eukaryotes. We describe here a systematic approach to identify the RNA cargoes associated with the cytoskeletal motor proteins of Saccharomyces cerevisiae in combination with live-cell 3D super-localization microscopy of endogenously tagged mRNAs. Our analysis identified widespread association of mRNAs with cytoskeletal motor proteins, including association of Myo3 with mRNAs encoding key regulators of actin branching and endocytosis such as WASP and WIP. Using conventional fluorescence microscopy and expression of MS2-tagged mRNAs from endogenous loci, we observed a strong bias for actin patch nucleator mRNAs to localize to the cell cortex and the actin patch in a Myo3- and F-actin dependent manner. Use of a double-helix point spread function (DH-PSF) microscope allowed super-localization measurements of single mRNPs at a spatial precision of 25 nm in x and y and 50 nm in z in live cells with 50 ms exposure times, allowing quantitative profiling of mRNP dynamics. The actin patch mRNA exhibited distinct and characteristic diffusion coefficients when compared to a control mRNA. In addition, disruption of F-actin significantly expanded the 3D confinement radius of an actin patch nucleator mRNA, providing a quantitative assessment of the contribution of the actin cytoskeleton to mRNP dynamic localization. Our results provide evidence for specific association of mRNAs with cytoskeletal motor proteins in yeast, suggest that different mRNPs have distinct and characteristic dynamics, and lend insight into the mechanism of actin patch nucleator mRNA localization to actin patches.

Neoadjuvant chemotherapy in breast cancer: early response prediction with quantitative MR imaging and spectroscopy.

A prospective study was undertaken in women undergoing neoadjuvant chemotherapy for locally advanced breast cancer in order to determine the ability of quantitative magnetic resonance imaging (MRI) and proton spectroscopy (MRS) to predict ultimate tumour response (percentage decrease in volume) or to detect early response. Magnetic resonance imaging and MRS were carried out before treatment and after the second of six treatment cycles. Pharmacokinetic parameters were derived from T1-weighted dynamic contrast-enhanced MRI, water apparent diffusion coefficient (ADC) was measured, and tissue water:fat peak area ratios and water T2 were measured using unsuppressed one-dimensional proton spectroscopic imaging (30 and 135 ms echo times). Pharmacokinetic parameters and ADC did not detect early response; however, early changes in water:fat ratios and water T2 (after cycle two) demonstrated substantial prognostic efficacy. Larger decreases in water T2 accurately predicted final volume response in 69% of cases (11/16) while maintaining 100% specificity and positive predictive value. Small/absent decreases in water:fat ratios accurately predicted final volume non-response in 50% of cases (3/6) while maintaining 100% sensitivity and negative predictive value. This level of accuracy might permit clinical application where early, accurate prediction of non-response would permit an early change to second-line treatment, thus sparing patients unnecessary toxicity, psychological morbidity and delay of initiation of effective treatment.

Differentiation of benign from malignant breast disease associated with screening detected microcalcifications using dynamic contrast enhanced magnetic resonance imaging.

Dynamic Contrast Enhanced Magnetic Resonance Imaging (DCE-MRI) is an effective diagnostic modality for symptomatic breast disease. However, its role in evaluating clinically occult disease associated with mammographically detected microcalcification remains unclear. Women recalled following screening mammography with microcalcification had DCE-MRI examination of the breast. The data were evaluated subjectively and objectively using both empirical and 2-compartment pharmacokinetic modelling techniques to evaluate signal intensity parameters. Eighty-eight patients aged 50-75 years (median 58) were recruited. Comparing malignant and benign lesions, the mean values in arbitrary units for the enhancement index at 1 min in the most enhancing 9-pixel square +/-1 standard deviation were 0.61+/-0.40 vs. 0.22+/-0.26 p=<0.001 with sensitivity, specificity, PPV, NPV and accuracy of 80.0%, 82.4%, 57.1%, 93.3% and 81.8%, respectively. The corresponding values attained by the radiologist were 75.0%, 89.7%, 68.2%, 92.4% and 86.4%. DCE-MRI is able to differentiate malignant from benign clinically occult lesions associated with microcalcification and may therefore offer an alternative to open surgical biopsy for women with equivocal findings following initial triple assessment for microcalcification in the breast.

Differentiation of benign and malignant sub-1 cm breast lesions using dynamic contrast enhanced MRI.

The purpose of this work is to assess the additional benefit of MRI-based morphology and quantification of contrast enhancement in the differential diagnosis of sub-1cm breast lesions. Forty-three women with suspected breast cancer were examined using X-ray mammography, ultrasound mammography, and MRI. Dynamic contrast imaging was performed and relative enhancement at various time-points was calculated. The dynamic data was also processed using a two-compartment pharmacokinetic model. Radiological interpretation of high-resolution post-contrast images revealed a similar accuracy (69%) compared to X-ray mammography (69%) and ultrasound mammography (67%). The best individual parameter calculated from the dynamic images was found to be the exchange rate constant which revealed a diagnostic accuracy of 0.74 +/- 0.08. When information from the post-contrast images and dynamic data was combined in a logistic regression model a diagnostic accuracy of 0.92 +/- 0.03 was achieved. In conclusion, MR imaging of small breast lesions is feasible and the incorporation of quantitative MR derived parameters is beneficial.

Anal gland carcinoma.

BACKGROUND: Anal gland carcinoma is a rare entity. The authors conducted a joint study of cases coded as definite or possible anal gland carcinoma from the archives of the Armed Forces Institute of Pathology and the Canadian Reference Center for Cancer Pathology. METHODS: Seven cases of potential anal gland carcinoma were identified from the Canadian files and 12 from the Armed Forces Institute of Pathology archives. Of these 19 cases, 14 had adequate material to allow clinical, histologic, and immunohistochemical analysis. RESULTS: Seven of these 14 cases met a modified World Health Organization (WHO) definition of anal gland carcinoma. The mean age of these patients was 66 years (range, 60-72 years), with a male-to-female ratio of 6:1. The tumors were composed of haphazardly dispersed, small glands with scant mucin production that invaded the wall of the anorectal area with no obvious intraluminal component observed clinically or microscopically. Immunohistochemical studies were performed on all seven of these cases, revealing cytokeratin (CK) 7+/CK 20- expression in six cases, and CK 7+/CK 20+ expression in one case. The remaining seven cases showed no intraluminal component but did not meet a modified WHO definition of anal gland carcinoma. This group included three mucinous adenocarcinomas (two clinically arising in anal fistulas), all of which were CK 7+/CK 20+, and a rectal-type adenocarcinoma that was CK 7-/CK 20+. There was also a tumor interpreted as probable rectal-type adenocarcinoma that was CK 7+/CK 20+, and a tumor interpreted as probable squamous cell carcinoma that was CK 7-/CK 20-. The seventh tumor in this group, which could not be classified, was CK 7+/CK 20-. CONCLUSIONS: A useful and discriminating definition of anal gland carcinoma is an anal canal tumor composed of haphazardly dispersed, small glands with scant mucin production invading the wall of the anorectal area without an intraluminal component. The glands are positive for CK 7.

Do adenoids regrow after excision?

OBJECTIVE: The goal was to determine the incidence of symptomatic adenoidal regrowth after adenoidectomy. STUDY DESIGN: A cross-sectional follow-up study was done in a randomly selected group of 175 children who had undergone adenoidectomy 2 to 5 years earlier. Nasopharyngoscopy was performed in those children who still had symptoms of nasal obstruction. SETTING: All surgery was performed at an academic hospital-based practice in the northeastern United States by a single surgeon using a consistent operative technique. RESULTS: Forty-six (26%) patients had nasal airway obstruction symptoms at follow-up. Of the 35 who agreed to undergo nasopharyngoscopy, not a single one had adenoids occupying more than 40% of the nasopharynx, and most (71%) were found to have either no or only trace amounts of adenoidal tissue (usually in the pharyngeal recess). CONCLUSION: Adenoids rarely, if ever, regrow enough to cause symptoms of nasal obstruction after adenoidectomy that includes visualization and electrocautery of the adenoid bed.

Fcgamma receptor-mediated phagocytosis in macrophages lacking the Src family tyrosine kinases Hck, Fgr, and Lyn.

Macrophage Fcgamma receptors (FcgammaRs) mediate the uptake and destruction of antibody-coated viruses, bacteria, and parasites. We examined FcgammaR signaling and phagocytic function in bone marrow-derived macrophages from mutant mice lacking the major Src family kinases expressed in these cells, Hck, Fgr, and Lyn. Many FcgammaR-induced functional responses and signaling events were diminished or delayed in these macrophages, including immunoglobulin (Ig)G-coated erythrocyte phagocytosis, respiratory burst, actin cup formation, and activation of Syk, phosphatidylinositol 3-kinase, and extracellular signal-regulated kinases 1 and 2. Significant reduction of IgG-dependent phagocytosis was not seen in hck(-)(/)-fgr(-)(/)- or lyn(-)(/)- cells, although the single mutant lyn(-)(/)- macrophages did manifest signaling defects. Thus, Src family kinases clearly have roles in two events leading to FcgammaR-mediated phagocytosis, one involving initiation of actin polymerization and the second involving activation of Syk and subsequent internalization. Since FcgammaR-mediated phagocytosis did occur at modest levels in a delayed fashion in triple mutant macrophages, these Src family kinases are not absolutely required for uptake of IgG-opsonized particles.

Combined proton MR spectroscopy and dynamic contrast enhanced MR imaging of human intracranial tumours in vivo.

A study was undertaken to determine if the vascular characteristics measured by dynamic contrast-enhanced magnetic resonance imaging (primarily permeability surface area product and extracellular-extravascular tissue volume fraction) would be beneficial in explaining the inter-lesion metabolic heterogeneity displayed by human intracranial tumours. Magnetic resonance spectroscopy was carried out using a single-voxel STEAM sequence and dynamic imaging was carried out using a combination of pre-contrast proton density-weighted FSPGR images (to remove the influence of native tissue T1), bolus injection of Gd-DTPA and subsequent T1-weighted FSPGR dynamic imaging. A two-compartment pharmacokinetic model was employed to determine vascular characteristics. Results obtained from 12 meningiomas suggest a possible correlation between the level of lipids/macromolecules and permeability surface area product, although the confounding issue of extra-voxel contamination arising from lipids in the scalp and skull marrow cannot be ruled out in the more superficial lesions. Results obtained from 11 gliomas (four low and seven high grade) demonstrate that permeability surface area product is not specific for the range of vascular characteristics and metabolite profiles observed in gliomas and is therefore unable to explain metabolic heterogeneity in these lesions.

Treatment of the macrophage-like P388D.1 cells with bacterial lipopolysaccharide and interferon-gamma causes long-term alterations in calcium metabolism.

The intracellular calcium concentrations ([Ca2+]i) of P338D.1 macrophage-like cells, activated with interferon-gamma (IFN-gamma) and/or bacterial lipopolysaccharide (LPS) were determined using fura-2/AM and ratiometric imaging techniques. Treatment of macrophages with IFN-gamma and LPS resulted in significant downward shift in [Ca2+]i, 8, 16 and 24 h but not at 1 and 4 h after treatment. The decrease in [Ca2+]i also occurred when macrophages were treated with LPS only, but not after exposure of the cells to recombinant IFN-gamma, indicating that LPS was an essential signal in the observed changes in [Ca2+]i of activated macrophages. The IFN-gamma and/or LPS alteration in the [Ca2+]i, paralleled the in vitro nitric oxide production of the activated macrophages, 8, 16 and 24 h after treatment. The decrease in the [Ca2+]i may be caused by vigorous buffering and storing of Ca2+ by macrophages to below the normal resting quantities, following the reported transient increase in Ca2+ during the priming stage of macrophage activation. Thus, the downward shift in [Ca2+]I may play a physiological role in the activation processes of macrophages for antimicrobial responses.

Primary malignant melanoma of the esophagus: radiographic findings in seven patients.

PURPOSE: To evaluate the radiographic findings in a series of patients with primary malignant melanoma of the esophagus. MATERIALS AND METHODS: Seven cases of primary esophageal melanoma were collected from the Hospital of the University of Pennsylvania and the radiologic archives of the Armed Forces Institute of Pathology. All patients underwent esophagography (double-contrast esophagograms in four patients, single-contrast esophagograms in three patients). Medical, endoscopic, and pathology reports were also reviewed. RESULTS: Six patients presented with dysphagia (average duration, 5 months). The remaining patient had recent onset of melena. In all patients, barium studies revealed bulky, polypoid intraluminal masses that focally expanded the esophagus without causing obstruction. The tumors were located in the distal third of the thoracic esophagus in four patients, the middle third in two, and the proximal third in one. Tumor pigmentation was noted at endoscopy in only one patient. All patients underwent extensive esophageal resection. Of four patients with clinical follow-up findings, two were alive and well 2 and 6 months after surgery; the other two had metastases to the lung and liver within 2 months of surgery. CONCLUSION: Primary esophageal melanomas have strikingly similar barium study findings, appearing as bulky, polypoid intraluminal masses that focally expand the esophagus without causing obstruction. Despite its rarity, primary esophageal melanoma should be considered when characteristic findings are present on barium studies.

Induction of nitric oxide (NO) synthesis in murine macrophages requires potassium channel activity.

The activation of macrophages for antimicrobial responses is a multistage event involving numerous intracellular signalling cascades that makes possible target cell destruction by these effector cells. This study examined the effects of different potassium channel inhibitors and activators on the NO production of murine macrophage-like cell lines P388D.1 and B10-4(S). We found that the potassium channel inhibitors tetraethylammonium, 4-aminopyridine, and quinine caused dose-dependent reductions in the NO production of macrophages, and that the potassium channel activator, minoxidol, caused a dose-dependent enhancement of NO production. The inhibition of NO production was due to involvement of potassium channels in the priming stage of macrophage activation, since pretreatment with the priming agent interferon-gamma partially restored the NO response of the macrophages. The results of this study demonstrate a link between potassium channel activity and the activation of anitimicrobial functions of murine macrophages.

In vivo quantification of citrate concentration and water T2 relaxation time of the pathologic prostate gland using 1H MRS and MRI.

We have previously reported a striking correlation between water T2 relaxation time and citrate concentration in the normal prostate (Liney G.P.; Lowry M.; Turnbull L.W.; Manton D.J.; Knowles A.J.; Blackband S.J.; Horsman A. Proton MR T2 maps correlate with the citrate concentration in the prostate. NMR Biomed. 9:59-64; 1996). In this study we present data from similar studies of the pathologic gland. The findings support the hypothesis that measurement of both citrate concentration and water T2 relaxation time in vivo may aid the differentiation of prostatic carcinoma from benign disease and normal tissue.

Successful treatment of non-familial haemophagocytic lymphohistiocytosis with interferon and gammaglobulin.

Two cases of non-familial haemophagocytic lymphohistiocytosis (HLH) are presented in which treatment with interferon alfa and gammaglobulin was associated with complete clinical remission. In one case, serological evidence of recent Epstein-Barr virus infection was found. Natural killer cell activity was within normal limits in both children, compatible with a secondary form of HLH. The combination of interferon alfa and intravenous gammaglobulin requires further evaluation in the treatment of non-familial HLH.

Small-cell carcinoma of the esophagus: radiographic findings.

PURPOSE: To determine the radiographic findings of small-cell carcinoma of the esophagus. MATERIALS AND METHODS: The authors retrospectively reviewed barium studies as well as medical and pathologic records for three cases of small-cell carcinoma of the esophagus contributed to the radiologic archives of the Armed Forces Institute of Pathology. RESULTS: Two patients presented with dysphagia and one with chest pain. In all three patients, barium studies revealed a smoothly marginated, sessile mass with a relatively flat central ulcer on the right postero-lateral wall of the midesophagus below the level of the carina. The masses all were 4-5 cm in diameter, and the ulcers were 2-3 cm in diameter. In all three patients, the results of endoscopy confirmed the presence of a sessile mass with central ulceration in the midesophagus. CONCLUSION: Small-cell carcinomas of the esophagus can have similar findings on barium studies. Although these findings are more likely to be caused by squamous-cell carcinoma, it is important to obtain endoscopic biopsy specimens, because preoperative histologic diagnosis of small-cell carcinoma can dramatically alter the management of these cases.

Esophageal leiomyomatosis.

PURPOSE: To reassess the clinical and radiologic findings in patients with esophageal leiomyomatosis. MATERIALS AND METHODS: A search of the authors' radiologic archives revealed six cases of esophageal leiomyomatosis in a 22-year period. The clinical findings and radiologic images were reviewed retrospectively. RESULTS: The average age of the patients was 10.8 years (range, 6-18 years). Five patients presented with slowly progressive dysphagia. Barium studies revealed smooth, tapered narrowing of the distal esophagus in five patients and characteristic defects on the superomedial aspect of the gastric fundus abutting the cardia, presumably due to bulging of this thickened mass of muscle into the stomach, in four patients. In two patients, computed tomography (CT) revealed marked thickening of the distal esophageal wall. CONCLUSION: Esophageal leiomyomatosis can be suggested in a pediatric patient with long-standing dysphagia in whom smooth, tapered distal esophageal narrowing is seen at barium study and circumferential esophageal wall thickening is seen at CT.

Proton MR T2 maps correlate with the citrate concentration in the prostate.

A significant correlation between water T2 relaxation time and citrate concentration in the normal prostate is demonstrated using spatially localized spectroscopy and water T2 maps. These data imply that MR images may be used to infer the concentration of citrate at high spatial resolution. Preliminary results in patients demonstrate that this relationship is maintained in benign prostatic hyperplasia (BPH) and prostatic carcinoma. The determination of citrate concentration and water T2 may aid the differentiation of prostate carcinoma from BPH and normal tissue.

Determination of proton metabolite concentrations and relaxation parameters in normal human brain and intracranial tumours.

Quantitative proton spectroscopic studies were performed on 39 volunteers and 16 patients with intracranial tumours. Estimates of T2 were obtained in white matter, grey matter, cerebellum, astrocytomas and meningiomas; T1 was determined in white matter only. White matter values of T2 for trimethylamines, creatine and N-acetyl aspartate (NAA) were 309 +/- 84, 195 +/- 41 and 369 +/- 124 ms, respectively (mean +/- SD, n = 20). Metabolite concentrations in white matter were 2.0 +/- 0.4 mumol/g wet weight for trimethylamines, 7.3 +/- 1.1 for creatine and 11.4 +/- 1.4 for NAA. The mean concentrations of creatine and NAA in grey matter and all of three metabolites in cerebellum were greater than those in white matter. Tumour spectra were characterized by increased trimethylamines, reduced creatine and NAA and occasionally the presence of lactate. Meningiomas were further characterized by the presence of alanine. The mean T2 and concentration of trimethylamines in tumours was significantly greater than in normal brain. Creatine and NAA concentrations were decreased in all tumours. The longer T2 of trimethylamines and presence of alanine in meningiomas indicate that important differences exist in membrane and glucose metabolism within these tumours when compared to either astrocytomas or normal brain tissue.

Phosphorus-31 metabolism of post-menopausal breast cancer studied in vivo by magnetic resonance spectroscopy.

We have studied the metabolism of 31P-containing metabolites of post-menopausal breast cancers in vivo using magnetic resonance spectroscopy (MRS) and a 5.5 cm surface coil. Spectra were acquired from 23 diameter. The spectra of the 19 previously untreated tumours had significantly higher phosphomonoester (PME) 31P relative peak areas than the normal breasts of eight post-menopausal women (11.7% and 7.7% respectively, P = 0.002). Although an increased PME relative peak area was characteristic of malignancy, PME relative peak area is similarly raised in lactating breast and, therefore, not a specific feature of cancer. An apparently lower nucleotide triphosphate (NTP) relative peak area in tumours than healthy postmenopausal breast was secondary to the differences in PME relative peak area; contamination by signal from chest wall muscle probably accounts for the ostensibly higher phosphocreatine (PCr) relative peak area of the tumours. Spectroscopy was repeated following chemotherapy in six women. An increase in PCr relative peak area was seen in all five patients who responded, but again this may represent increased contamination secondary to changes in tumour size. A fall in PME relative peak area was noted in four responders, but also one non-responder, so this finding may not be sufficiently specific to be of use clinically. Further studies are need to elucidate fully the role of MRS in breast cancer.