Diagnostic accuracy and interobserver concordance: teledermoscopy of 600 suspicious skin lesions in Southern Denmark.

BACKGROUND: Skin cancer incidences are increasing and early diagnosis, especially of malignant melanoma, is crucial. Teledermatology including teledermoscopy (TDS) can be used to triage referrals of suspicious skin lesions, however, this is not currently recommended in Denmark. OBJECTIVES: To evaluate diagnostic accuracy, sensitivity, specificity and interobserver concordance of TDS, and to evaluate the number of incidental lesions potentially missed by TDS. METHODS: Fifty general practices were invited to send images of suspicious skin lesions for evaluation using smartphone TDS. Simultaneously, the patient was referred for a face-to-face (FTF) consultation. Images for TDS were independently evaluated by two dermatologists; a third dermatologist performed the FTF consultation. Diagnosis, management plan and level of diagnostic confidence were noted. For TDS photo quality was rated, and for FTF any incidental findings were described. RESULTS: Six hundred lesions in 519 patients were included. The diagnostic accuracy was significantly higher on FTF evaluation than on TDS (P < 0.01). However, this was associated with a significant difference in specificity (P ≤ 0.012) whereas no significant difference was found in sensitivity. The concordance between FTF and TDS, and the interobserver concordance of two TDS evaluations was moderate to substantial (AC1 = 0.57-0.71). Incidental melanomas were found in 0.6% of patients on FTF evaluation, adding an extra 13% of melanomas. However, on TDS these patients' photographed lesions all warranted FTF follow-up, where these melanomas would have been identified. CONCLUSION: In this large prospective study, no significant difference in sensitivity was observed between FTF and TDS, but specificity was lower on TDS than FTF. Taking management plans into account, we would, however, potentially have dismissed 2 of 23 melanomas, if only TDS had been used for assessment. One of these was a melanoma located on the scalp, an anatomic region less suitable for TDS.

Long term surgical and hearing outcomes in the management of tympanomastoid paragangliomas.

OBJECTIVE: To analyze the long term outcomes after surgery in tympanomastoid paragangliomas. STUDY DESIGN: Retrospective study. METHODS: The charts of 145 patients with tympanomastoid paragangliomas managed between 1988 and 2013 were reviewed. The clinical features, audiological data, pre- and postoperative notes were noted. The tumors were staged according to the modified Fish and Mattox classification. The surgical approaches for all patients were formulated according to the surgical algorithm developed at our center. RESULTS: 34 (23.5%), 46 (31.7%), 22 (15.2%), 18 (12.4%) and 25 (17.2%) patients were diagnosed to have TMP class A1, A2, B1, B2 and B3 tumors respectively. Gross tumor resection was achieved in 141 (97.2%) patients. The facial nerve was uncovered in four patients and infiltrated in three. The cochlea was found eroded in seven cases. The mean follow-up was 48.4 months. Recurrence was seen in one patient (0.7%). In the cases where the facial nerve was preserved (n=143), the nerve function was graded as HB grade 1 in 138 patients (97%). Postoperatively, the mean AC showed an improvement in all categories except in class B2 and B3, which corresponds to the classes that include patients who underwent subtotal petrosectomy. CONCLUSION: We report the long term surgical outcomes in tympanomastoid paragangliomas in the largest series published till date. It is possible to completely eradicate all types of tympanomastoid paragangliomas with minimum sequelae by choosing the correct surgical approach to achieve adequate exposure for individual tumor classes as described in our classification and algorithm. LEVEL OF EVIDENCE: IIb.

Long-term surgical and functional outcomes of the intact canal wall technique for middle ear cholesteatoma in the paediatric population.

In this paper, we report the postoperative outcomes in canal wall up procedures with second stage surgery in 40 children undergoing intervention for cholesteatoma of the middle ear. The residuals, recurrences and the hearing results were analysed. All 40 patients had a follow-up of at least five years. Of the 39 patients who underwent two staged surgery, 18 (46.1%) had a residual lesion that was identified and excised during the second surgery. Over a five year follow-up period, there were five (12.5%) patients with recurrences, all belonging to the group in whom a residual cholesteatoma was identified during the second staged surgery. The rate of residual cholesteatoma tends to decrease as age increases. The type of cholesteatoma, acquired or congenital middle ear, were not statistically related to the incidence of residual cholesteatoma. Hearing analysis showed that hearing recovery was excellent with canal wall up procedures and remained stable over five years.

Vocal cord hemangioma.

Laryngeal hemangiomas are relatively rare. Laryngeal hemangiomas occur in two main forms--infantile and adult laryngeal hemangiomas. While infantile hemangiomas are usually found to occur in the subglottis, adult hemangiomas occur commonly in the supraglottic regions of the larynx. Laryngeal hemangioma with cavernous features isolated to the free edge of the vocal fold is a very rare clinical finding. We present a case of hemangioma of the right vocal cord in an adult, which was managed successfully in our center.

Protein changes associated with ionizing radiation-induced apoptosis in human prostate epithelial tumor cells.

Ionizing radiation (IR) is an important component in the therapy of localized prostate cancer. Identification of protein alterations during IR-induced apoptosis prostate cancer cells is an important step toward understanding the new metabolic status of the dying cell. In the present study, we report changes in protein profile that define the execution phase of the apoptotic response in the in vitro model of tumorigenic radiation-transformed SV40-immortalized human prostate epithelial cells (267B1-XR), induced to undergo programmed cell death by IR. We employed an approach that involves use of analytical two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) coupled with Western blotting with specific antisera. Our results point out that apoptotic cells experience significant reduction in the levels of the intermediate filament proteins, keratins-18, 19, vimentin and the associated 14-3-3 adapter proteins. At the same time, molecular chaperones such as glucose-regulated protein 94, calreticulin, calnexin, and protein disulfide isomerase exhibit marked accumulation in these dying cells. The present data indicate that apoptosis-associated processes in prostate epithelial cells include solubilization of the rigid intermediate filament network by specific proteolysis as well as increased levels of endoplasmic reticulum (ER) proteins with chaperone functions.

Detection of heterogeneity of apoptotic fragments of poly (ADP-ribose) polymerase in MDA-MB-468 breast cancer cells: two-dimensional gel analysis.

Caspace-mediated proteolysis of the nuclear enzyme poly(ADP-ribose) polymerase (PARP) (EC 2.4, 2.30) is a biochemical marker of cell death in response to various apoptotic stimuli. Anti-PARP antibodies identifying the 89 kDa polypeptide from the C-terminus as well as the 113 kDa native enzyme are often used to demonstrate evidence of apoptosis-associated, interleukin converting enzyme (ICE)-mediated limited cleavage. Recent evidence points to redundancy of caspases, heterogeneity of their cleavage sites, and a possibility of generating distinct context-specific, and cell-specific PARP fragments. In the present study, we employed antibodies directed to multiple sites in PARP and probed two-dimensionally resolved proteins of the estrogen receptor negative MDA-MB-468 breast tumor cells, induced to undergo apoptosis by ionizing radiation (IR). Our results revealed that the 24 kDa apoptotic fragment of PARP, from the N-terminus, consists of at least three isoforms, located at a p/more basic than the full length enzyme. We also report a hitherto unrecognized feature of an anti-PARP antiserum, VIC-5, detecting both the 89 kDa and the 24 kDa caspase-generated fragments of PARP. Thus, application of two-dimensional electrophoresis combined with antisera directed to multiple sites would be valuable in distinguishing PARP cleavage site- and inhibitor specificities of proteases during apoptosis.

Differential expression of stathmin during neoplastic conversion of human prostate epithelial cells is reversed by hypomethylating agent, 5-azacytidine.

In a variety of human tumor tissues, including those of prostate and breast, CpG hypermethylation represents one of the mechanisms downregulating the expression of specific proteins, including tumor suppressor proteins. Using 267B1-XR cells generated by ionizing radiation-induced transformation of epithelial cells, derived from neonatal human prostate and immortalized by SV40 (267B1), we now report markedly low levels of expression of the cytoplasmic phosphoprotein stathmin, in addition to several proteins of the actin microfilaments and intermediate filaments that characterize the altered phenotype. Stathmin is emerging as a relay protein integrating signals from diverse pathways during differentiation and neoplastic progression. In this in vitro prostate carcinogenesis model system, where loss of specific-protein expression is a major feature of the transformed 267B1-XR cells, we employed 5-azacytidine treatment followed by 2D-PAGE to reveal if experimental genomic hypomethylation reinstated the levels of any of the differentially expressed proteins. Our data suggest that stathmin represents one such example.

Effects of collimator jaw setting on dose output for treatments with multileaf collimator.

The variations in dose output with collimator jaw setting have been investigated for treatment fields shaped by a multileaf collimator. Measurements have been made for 6 and 15 MV x-ray beams on a Varian Clinac 2100C machine. The results of our study show that the collimator jaw settings can affect the dose by about 5% for a small field shaped by a multileaf collimator. The effect is smaller for larger fields.

Apoptosis-associated proteolysis of vimentin in human prostate epithelial tumor cells.

Vimentin intermediate filaments (IF) are responsible for regulation of cell attachment and subcellular organization. Using an in vitro model system of human prostate epithelial cells (267B1-XR), we demonstrate that a series of vimentin proteolytic fragments represent some of the differentially expressed proteins in 2D-gel profiles of the apoptotic cells undergoing ionizing radiation-induced cell death. A caspase-sensitive motif search suggests that the type III IF protein (vimentin) is subject to proteolysis to promote the execution phase of apoptosis, in a manner similar to the well-established type V (lamins) and type I (keratins 18, 19) IF proteins. Furthermore, vimentin and a few of its derived polypeptides, reported to be specific to the apoptotic process, correspond to ubiquinated proteins, thus pointing to the complex interrelationships of protein ubiquination in solubilizing the IF network during apoptosis.

Two-dimensional gel electrophoresis analyses identify nucleophosmin as an estrogen regulated protein associated with acquired estrogen-independence in human breast cancer cells.

We have used two-dimensional gel electrophoresis to identify proteins associated with estrogen-induced proliferation in MCF-7 breast cancer cells and their progression to estrogen-independent proliferation. We compared the total cellular proteins from MCF-7 cells and an estrogen independent derivative of the MCF-7 cells MCF-7/LCC1 (Brünner et al. Cancer Research 1993, 53, 283-290), each grown with and without estradiol. These comparisons reveal seven estrogen-regulated proteins. Three of these proteins (HI-1: 36 kDa/pI 4.5, HI-10: 40 kDa/pI 5.5 and HI-19: 62 kDa/pI 5.0) exhibit a 'progression-like' pattern, being induced by estradiol in MCF-7 cells and constitutively present/upregulated in the MCF-7/LCC1 growing without estradiol. HI-11 (65 kDa/pI 5.5) is strongly induced by estradiol in MCF-7 cells but constitutively downregulated and unresponsive to estradiol in the MCF-7/LCC1 cells. Two proteins exhibit a suppressor pattern and are downregulated by estradiol in the estrogen-dependent MCF-7 cells (HI-3: 44 kDa/pI 4.4 and HI-4: 56 kDa/ pI 5.2) and present in MCF-7/LCC1 cells growing without estradiol at levels comparable to that seen in estrogen-treated MCF-7 cells. One protein (HI-9: 68 kDa/pI 5.5) exhibits a marked estrogen regulated pI shift, rather than changes in abundance. We purified and sequenced the HI-10 protein, which we identified as the nucleolar protein, nucleophosmin (NPM). One- and two-dimensional Western blot analyses of MCF-7/LCC1 cell lysates confirmed that HI-10 is immunoreactive with an antinucleophosmin antibody. Western blotting also confirmed the estrogenic regulation of NPM seen in the initial two-dimensional gel electrophoresis studies. Thus, NPM is induced by estradiol in the MCF-7 cells and upregulated in the MCF-7/LCC1 cells growing without estrogen, clearly associating its expression with an acquired estrogen-independent phenotype. NPM has several potentially important roles in regulating cell function and signaling. It is a substrate for phosphorylation by p34cdc2 kinase, protein kinase C and nuclear kinase II, and a repressor of the transcriptional regulating activities of both the IRF-1 tumor suppressor protein and the YY1 transcription factor. Studies are currently underway to determine which of these NPM functions may be involved in the hormonal progression of breast cancer.

Laser-based microfocused x-ray source for mammography: feasibility study.

A laser-produced plasma (LPP) x-ray source with possible application in mammography was created by focusing a laser beam on a Mo target. A Table-Top-Terawatt (TTT) laser operating at 1 J energy per pulse was employed. A dual pulse technique was used. Maximum energy transfer (approximately 10%) from laser light to hot electrons was reached at a 150 ps delay between pulses and the conversion efficiency (hard x-ray yield/laser energy input) was approximately 2 x 10(-4). The created LPP x-ray source is characterized by a very small focal spot size (tens of microns), Gaussian brightness distribution, and a very short pulse duration (a few ps). The spectral distribution of the generated x rays was measured. Images of the focal spot, using a pinhole camera, and images of a resolution pattern and a mammographic phantom were obtained. The LPP focal spot modulation transfer function for different magnification factors was calculated. We have shown that the LPP source in conjunction with a spherically bent, high throughput, crystal monochromator in a fixed-exit Rowland circle configuration can be used to created a narrow band tunable mammography system. Tunability to a specific patient breast tissue thickness and density would allow one to significantly improve contrast and resolution (exceeding 20 lp/mm) while lowering the exposure up to 50% for thicker breasts. The prospects for the LPP x-ray source for mammographic application are discussed.

Protein expression changes associated with radiation-induced neoplastic progression of human prostate epithelial cells.

Carcinogenic progression in most epithelial systems is a multistep process and presents as numerous (un)stable intermediate stages prior to the development of a fully malignant phenotype. Recently, we reported the neoplastic transformation of an SV40 immortalized, neonatal human prostate epithelial cell line (267B1) by multiple exposures to X-rays [1, 2]. The parental 267B1 cells acquired anchorage-independence and exhibited morphological transformation following exposure to two consecutive doses of 2 Gy. Exposure of either the parental 267B1 cells or the anchorage-independent derivatives (F3-SAC) to a total dose of 30 Gy of X-rays yielded tumorigenic transformants (267B1-XR and 267B1-SXR, respectively). All of these radiation-treated derivatives (F3-SAC, 267B1-XR, and 267B1-SXR) were characterized by reduced cell size and poorly organized actin stress fibers [2, 3]. The present study examines the protein expression changes associated with cytoskeletal alterations during the different steps of neoplastic progression induced by X-rays in the in vitro human prostate cell system. This analysis was achieved by using the high resolving power of two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) in the 267B1, F3-SAC, 267B1-XR, and 267B1-SXR cells. We report changes in the expression of gelsolin in the partially transformed, anchorage-independent, nontumorigenic (F3-SAC) cells and a progressive loss of expression of tropomyosin isoforms (TM-1 and TM-3), and myosin light chain-2 (MLC-2) in the tumorigenic (267B1-XR; 267B1-SXR) cells, respectively. In contrast, our results demonstrate that the levels of the small GTP-binding protein Rho-A, an active participant in the actin stress fiber organization, are not altered during neoplastic progression of these 267B1 cells. Thus the changes in synthesis of gelsolin, tropomyosins, and MLC-2 provide a rationale for the alterations in the actin stress fiber formation and reduction in cell size during the exposure of prostate epithelial cells to multiple doses of X-rays.

Scatter reduction in mammography with air gap.

Scatter reduction by air gaps in mammography was investigated. We have experimentally demonstrated that, independently of the imaging geometry, scatter in air-gap mammography can be well described by a virtual source of scatter (VSS) model. This model postulates that scatter radiation originates from a virtual point source of scatter placed on the central axis between the x-ray source and the exit surface of a patient at distance delta and utilizes only two parameters: delta and (S/P)0. The (S/P)0 parameter represents scatter-to-primary ratio without an air gap and delta is the distance from the exit surface of a patient to the virtual source of scatter. We have experimentally determined the analytical form of the two independent parameters of the VSS model; delta exhibits a linear increase proportional to the radiation field size, does not depend on patient thickness, and is in the 10-30 cm range, while (S/P)0 increases with the field size as a power function and is in the 0.4-1.3 range. In the framework of the VSS model the selectivity, the contrast improvement factor, and the signal-to-noise improvement factor were employed to evaluate performance of air-gap mammography systems. We have demonstrated that selectivity of an air gap rapidly deteriorates at some well-defined critical value of scatter fraction that has profound consequences on air-gap performance. Assuming fixed patient exposure, the results shows that, if a contrast limited detection system (such as film/screen mammography) is used, an air gap system can outperform a grid system only if a very large source-to-patient (SPD) distance is utilized, which might be possible with new laser-based x-ray sources. For the noise limited detection systems (such as digital mammography) even a small SPD (70 cm) and a small air-gap (20 cm) system will outperform a grid system.

Radiation-induced neoplastic transformation of human prostate epithelial cells.

We report the malignant transformation of human prostate epithelial cells (267B1) after multiple exposures to ionizing radiation. Carcinogenic progression of cells from immortal growth to anchorage-independent growth in soft agar to tumorigenicity in athymic mice resulted after a cumulative X-ray dose of 30 Gy. The tumors were characterized histologically as poorly differentiated adenocarcinomas, expressed prostate-specific antigen, and stained positive for keratin. No p53 or ras mutations were observed. Numerous chromosomal defects were noted on karyotypes after radiation exposure. However, chromosome 3 and 8 translocations were observed predominantly in the tumor outgrowths. These findings provide the first evidence of malignant transformation of human prostate epithelial cells exposed to ionizing radiation.

Cytoskeletal changes during radiation-induced neoplastic transformation of human prostate epithelial cells.

We recently reported tumorigenic transformation of SV40-immortalized neonatal human prostate epithelial cells (267B1) by exposure to fractionated doses of X-rays. Altered morphology and anchorage independence were observed following two successive fractions of 2 Gy each (F3-SAC). Additional 2 Gy treatments to these non-tumorigenic cells to a total dose of 30 Gy resulted in radiation-transformed tumorigenic colonies (267B1-SXR). Malignant transformation of parental 267B1 cells was also achieved by consecutive 2 Gy exposures to a total dose of 30 Gy (267B1-XR). This study discusses the cytoskeletal changes in the F3-SAC, 267B1-XR and 267B1-SXR derivatives of these human prostate epithelial cells. Confocal and conventional fluorescence microscopy of filamentous actin showed numerous, well organized, evenly distributed stress fibers in the parental cells prior to irradiation, while the anchorage-independent cells and several tumorigenic derivatives exhibited poor stress fiber organization after radiation exposure. This disorganization of actin microfilaments in the radiation-transformed cells was also accompanied by changes in the expression of selective tropomyosin isoforms as judged by two-dimensional gel electrophoresis. These changes in actin organization and tropomyosin expression appear to be coincidental with morphological transformation and acquisition of tumorigenicity in the 267B1 cells following radiation exposure.

Comparison of measured and calculated dose for asymmetric x-ray beams defined by independently movable collimators.

Linear accelerators with x-ray collimators that move independently are becoming increasingly common for treatment with asymmetric fields. In an asymmetric field, the center of the treatment field is away from the true central axis where dosimetric data are normally obtained. In this paper we present a simplified approach to the calculation of dose for asymmetric fields. We use central axis tissue-maximum ratio, off-axis factor in phantom and relative field-size factor in phantom to calculate dose. The accuracy of our calculations has been compared with ion-chamber measurements for 6 and 15 MV x-ray beams. Measurements were made at 5, 10, and 15 cm off-axis for a 20 cm x 20 cm asymmetric field at dmax and 6 cm depths in a solid-water phantom using a 0.6 cc Farmer chamber. Agreement within 3% was found at the measurement points.

A simple approach to the technical aspects of radiosurgery treatments.

An approach to radiosurgery treatment that can be readily adopted in most radiotherapy centers with linear accelerators is presented. In our institution, a Leksell-type of neurosurgical frame, a computed tomography scanner, locally fabricated cones, and 6 MV X-ray beams are used to perform radiosurgery treatments. Collimated arcs with dose distributions, that conform to the shape of the lesion in the transverse and the sagittal planes are used. It is argued that the uncertainties in the localization of the isocenter within a lesion and the specifications of the size of the target volume do not justify high precision mechanical devices for most radiosurgery treatments.

High-resolution two-dimensional electrophoresis of nuclear proteins: a comparison of HeLa nuclei prepared by three different methods.

A comparative analysis of HeLa cell nuclear proteins is presented using Iso-Dalt methods of protein resolution in two dimensions. The nuclear proteins were prepared by (1) spin through glycerol cushion, (2) spin through sucrose cushion, or (3) Triton wash. Improved resolution of total nuclear proteins in the range of pH 4.5-6.0 was achieved by substituting longer isotubes in combination with broad-range ampholines during the isoelectric focusing step. An attempt to indicate silver stainable protein spots common to total cellular extracts and nuclear preparations has been made. Also, proteins that appear to be well represented in all three nuclear preparations and remain undetectable in the total cellular protein pattern have been marked as probably being enriched nuclear proteins. Such a comparative analysis of whole nuclear protein preparations made it possible to document that the different preparations preserved the same set of proteins. The Triton-wash method of obtaining nuclei was identified as the preferred choice. Coomassie-stained gels and blots of these nuclear proteins could serve as a guide for accessing relevant protein spots for further biochemical analysis such as immunoblotting.

Comparison of absorbed doses in water and solid water for electron beams.

Solid water, as a substitute for water, has become commercially available for dosimetry measurements. A study was undertaken to compare the dose in water and solid water respectively for 6, 9, 12, 16 and 20 MeV electron beams. Measurements using ion chamber show that the dose in water is higher than the dose in solid water by 1% for 6, 9, and 12 MeV electrons. For 16 and 20 MeV electrons, the dose in water and solid water are the same within the uncertainty of our measurements.

Linear quadratic model and biologically equivalent dose for single fraction treatments.

The linear quadratic model has been used to calculate the biologically equivalent dose for single fraction treatments. Our calculations suggest that for late reacting tissue, such as the brain, a single fraction of 1440 cGy is equivalent to a conventional treatment of 5000 cGy in 25 fractions.