Development and validation of a nomogram incorporating gene expression profiling and clinical factors for accurate prediction of metastasis in patients with cutaneous melanoma following Mohs micrographic surgery.

BACKGROUND: There is a need to improve prognostic accuracy for patients with cutaneous melanoma. A 31-gene expression profile (31-GEP) test uses the molecular biology of primary tumors to identify individual patient metastatic risk. OBJECTIVE: Develop a nomogram incorporating 31-GEP with relevant clinical factors to improve prognostic accuracy. METHODS: In an IRB-approved study, 1124 patients from 9 Mohs micrographic surgery centers were prospectively enrolled, treated with Mohs micrographic surgery, and underwent 31-GEP testing. Data from 684 of those patients with at least 1-year follow-up or a metastatic event were included in nomogram development to predict metastatic risk. RESULTS: Logistic regression modeling of 31-GEP results and T stage provided the simplest nomogram with the lowest Bayesian information criteria score. Validation in an archival cohort (n = 901) demonstrated a significant linear correlation between observed and nomogram-predicted risk of metastasis. The resulting nomogram more accurately predicts the risk for cutaneous melanoma metastasis than T stage or 31-GEP alone. LIMITATIONS: The patient population is representative of Mohs micrographic surgery centers. Sentinel lymph node biopsy was not performed for most patients and could not be used in the nomogram. CONCLUSIONS: Integration of 31-GEP and T stage can gain clinically useful prognostic information from data obtained noninvasively.

Mitotic rate for thin melanomas: should a single mitotic figure warrant a sentinel lymph node biopsy?

BACKGROUND: The seventh edition of the American Joint Committee on Cancer guidelines recognize mitotic rate (MR) as a component of the staging criteria for cutaneous melanomas with a Breslow depth ≤1 mm. OBJECTIVE: This review discusses the evidence behind the threshold of 1 mitosis per square millimeter as a prognostic variable in thin melanomas, particularly because it relates to the decision to pursue a sentinel lymph node biopsy (SLNB). MATERIALS AND METHODS: We performed a systematic review using the PubMed database to identify articles that contain prognostic information for thin melanomas based on MR and sentinel lymph node (SLN) status. RESULTS: Although the threshold of a single mitosis correlates with a statistically significant decrease in survival rates for patients with thin melanomas, the clinical relevance remains questionable particularly because it relates to the decision to pursue an SLNB. CONCLUSION: A single mitosis in thin melanomas does not increase the risk of a positive SLN so much that SLN biopsy should be routinely performed for this cohort.

Microdermabrasion: molecular mechanisms unraveled, part 2.

Microdermabrasion (MDA) remains a common in-office procedure for many dermatologic practices. The procedure offers minimal downtime with a low incidence of side effects, making it a relatively desirable option for skin rejuvenation. Investigators have identified many of the molecular mechanisms behind this technology in an attempt to optimize clinical results. In particular, activation of the wound healing response plays a key role in the remodeling of post-MDA treated skin, although this response varies based on the type of MDA employed. In addition, advances in MDA technology offer new and promising ways to enhance transcutaneous penetration of active ingredients to improve clinical outcomes. Our review addresses innovative applications of MDA in the last 10 years of research.

Microdermabrasion: molecular mechanisms unraveled, part 1.

Microdermabrasion (MDA) remains a common in-office procedure for many dermatologic practices.The procedure offers minimal downtime with a low incidence of side effects, making it a relatively desirable option for skin rejuvenation. Investigators have identified many of the molecular mechanisms behind this technology in an attempt to optimize clinical results. In particular, activation of the wound healing response plays a key role in the remodeling of post-MDA treated skin, although this response varies based on the type of MDA employed.While many studies discuss the clinical applications of MDA and their relation to histologic changes found after treatment, few address the basic science behind the technology.Our review covers progress made in the last 10 years of research, with an emphasis on the molecular mechanisms.

The evolving conception and management challenges of malignant fibrous histiocytoma.

BACKGROUND: Malignant fibrous histiocytoma (MFH) is a rare and aggressive tumor. Mohs micrographic surgery (MMS) has been reported as an effective treatment, although most cases were published before advances in cytopathologic techniques led to reclassification of many tumors. OBJECTIVE: To evaluate a contemporary cohort of individuals with MFH and analyze management practices. METHODS: We reviewed all cases of MFH diagnosed at our institution from January 1995 to December 2010, evaluating 839 records to identify 36 patients undergoing management of tumors of the head and neck. RESULTS: Seventeen of the total 36 patients (47%; mean age 67) experienced tumor recurrence, and 10 (28%) developed metastases. Seven of nine patients initially treated with MMS (78%), and 10 of 24 (42%) treated with WLE experienced recurrence (p = .06). Patients treated with MMS had smaller tissue defects after surgery. The mean contemporary recurrence rate of MFH treated with MMS is significantly higher (58.8%) than the cumulative recurrence rate reported before 2000 (7.4%) (p < .001). CONCLUSIONS: Our study is consistent with reports of MFH as an aggressive neoplasm and describes the largest population treated with MMS in 3 decades. The changing conception of MFH, along with a propensity for in-transit metastases, may explain higher contemporary recurrence rates.

What's new in skin resurfacing and rejuvenation?

Skin resurfacing and rejuvenation techniques have experienced significant advances in the last few decades, and new devices are continuously being introduced into the marketplace. The large number of available modalities such as fractional lasers, radiofrequency systems, microdermabrasion, laser-assisted lipolysis, and ultrasound provides practitioners with a wide array of choices to address the needs of cosmetic patients. Many of the newer technologies, such as devices that integrate a fractional delivery system, provide excellent results with few adverse side effects. However, proper perioperative management is essential to achieving the desired effects. Furthermore, practitioners must be fluent with the operation and expected complications of these new technologies. While there are many overlapping treatment indications for each of the devices mentioned above, some modalities offer distinct advantages making proper patient selection essential. In certain cases, utilizing various combinations of mechanical, optic, acoustic, and electrical energies may be necessary to achieve the desired outcomes. This review discusses the application of new and existing skin rejuvenation techniques to clinical practice. A particular emphasis is placed on the use of fractional, radiofrequency, microdermabrasion, laser-assisted lipolysis, and ultrasound devices.

Effects of amorphous layers on ADF-STEM imaging.

A study of high-resolution ADF imaging in uncorrected and aberration-corrected STEMs was carried out by multislice simulation. The presence of amorphous layers at the surface of a crystalline specimen is shown to significantly alter the visibility of the atomic columns. After propagating through an amorphous layer a portion of the beam passes without any alteration while scattered electrons introduce a Gaussian background. The dependence of the image contrast on the crystal structure, orientation and the types of the atoms present in the crystal was studied. In the case of uncorrected probes an amorphous layer thicker than 200 A is necessary to achieve considerable reduction of the visibility of the atomic columns, but with aberration-corrected probes only 60 A is necessary. With changes in defocus, crystalline specimens with amorphous layers on the top can also be imaged and high-resolution ADF images can be obtained. An amorphous layer at the beam entry surface affects the ADF image more than that of an amorphous layer at the exit surface. Approximately linear reduction of the contrast (with a slop of 1) is expected with increased thickness of amorphous layer.

Effects of tilt on high-resolution ADF-STEM imaging.

A study of the effects of small-angle specimen tilt on high-resolution annular dark field images was carried out for scanning transmission electron microscopes with uncorrected and aberration-corrected probes using multislice simulations. The results indicate that even in the cases of specimen tilts of the order of 1 degree a factor of 2 reduction in the contrast of the high-resolution image should be expected. The effect holds for different orientations of the crystal. Calculations also indicate that as the tilted specimen gets thicker the contrast reduction increases. Images simulated with a low-angle annular dark field detector show that tilt effects are more pronounced in this case and suggest that these low-angle detectors can be used to correct specimen tilt during scanning transmission electron microscopes operation.

Separation of bulk and surface-losses in low-loss EELS measurements in STEM.

To identify major features in low electron energy loss spectra, the different excitations (bulk plasmons, interband transitions, surface plasmons, Cherenkov and surface guided modes) must be delineated from each other. In this paper, this process is achieved by noting the linear thickness dependence of bulk processes contrasted with the constant thickness behavior of surface excitations. An alternative approach of analyzing bulk plasmon-loss is also introduced. Using a new algorithm, the parameters of plasma generation-plasmon energy E(P,0), a damping parameter DeltaE(P) and the coefficient of the dispersion relation gamma were obtained from a single curve fitting on the example of Si. The ability to separate surface-losses from the rest of the data permitted identification of the fine structure of the surface-losses. The strong peak at 8.2 eV characteristic of non-radiative surface plasmon excitations was measured for Si. Analysis of surface excitations indicates that a 10ASiO2 surface coating layer is still present despite careful cleaning the specimen. Dielectric functions deduced from the EELS data prove to be considerably affected by the presence of the surface-losses for samples as thick as 800A.

Prevalence of viable Toxoplasma gondii in beef, chicken, and pork from retail meat stores in the United States: risk assessment to consumers.

The prevalence of viable Toxoplasma gondii was determined in 6,282 samples (2,094 each of beef, chicken, and pork) obtained from 698 retail meat stores from 28 major geographic areas of the United States. Each sample consisted of a minimum of 1 kg of meat purchased from the retail meat case. To detect viable T. gondii, meat samples were fed to T. gondii-free cats and feces of cats were examined for oocyst shedding. Initially, 100 g of meat from 6 individual samples of a given species were pooled (total, 600 g), fed to a cat over a period of 3 days, and feces were examined for oocysts for 14 days; the remaining meat samples were stored at 4 C for 14 days (until results of the initial cat fecal examination were known). When a cat fed pooled samples had shed oocysts, 6 individual meat samples from each pool were bioassayed for T. gondii in cats and mice. Toxoplasma gondii isolates were then genetically characterized using the SAG2 locus and 5 hypervariable microsatellite loci. In all, 7 cats fed pooled pork samples shed oocysts. Toxoplasma gondii oocysts were detected microscopically in the feces of 2 of the cats; 1 isolate was Type II and the second was Type III. Analyzed individually, T. gondii was detected by bioassay in 3 of the 12 associated samples with genetic data indicating T. gondii isolates present in 2. The remaining 5 pooled pork samples had so few oocysts that they were not initially detected by microscopic examination, but rather by mouse bioassay of cat feces. Two were Type I, 1 was Type II, and 2 were Type III. None of the cats fed chicken or beef samples shed oocysts. Overall, the prevalence of viable T. gondii in retail meat was very low. Nevertheless, consumers, especially pregnant women, should be aware that they can acquire T. gondii infection from ingestion of undercooked meat, and in particular, pork. Cooking meat to an internal temperature of 66 C kills T. gondii.

Depth-dependent imaging of individual dopant atoms in silicon.

We have achieved atomic-resolution imaging of single dopant atoms buried inside a crystal, a key goal for microelectronic device characterization, in Sb-doped Si using annular dark-field scanning transmission electron microscopy. In an amorphous material, the dopant signal is largely independent of depth, but in a crystal, channeling of the electron probe causes the image intensity of the atomic columns to vary with the depths of the dopants in each column. We can determine the average dopant concentration in small volumes, and, at low concentrations, the depth in a column of a single dopant. Dopant atoms can also serve as tags for experimental measurements of probe spreading and channeling. Both effects remain crucial even with spherical aberration correction of the probe. Parameters are given for a corrected Bloch-wave model that qualitatively describes the channeling at thicknesses 20 nm, but does not account for probe spreading at larger thicknesses. In thick samples, column-to-column coupling of the probe can make a dopant atom appear in the image in a different atom column than its physical position.

Spatial distribution of competing ions around DNA in solution.

The competition of monovalent and divalent cations for proximity to negatively charged DNA is of biological importance and can provide strong constraints for theoretical treatments of polyelectrolytes. Resonant x-ray scattering experiments have allowed us to monitor the number and distribution of each cation in a mixed ion cloud around DNA. These measurements provide experimental evidence to support a general theoretical prediction: the normalized distribution of each ion around polyelectrolytes remains constant when ions are mixed at different ratios. In addition, the amplitudes of the scattering signals throughout the competition provide a measurement of the surface concentration parameter that predicts the competition behavior of these cations. The data suggest that ion size needs to be taken into account in applying Poisson-Boltzmann treatments to polyelectrolytes such as DNA.

Simulation of thermal diffuse scattering including a detailed phonon dispersion curve.

Thermal vibration of the atoms in a crystal give rise to a diffuse background in the diffraction pattern (in between the normal allowed Bragg reflections). The Einstein model for phonon vibrations in a crystal leads to Gaussian statistics for the phonons. However, the Einstein model ignores the possibility of correlation between the atoms. An accurate model of the phonon dispersion curves for silicon is used to generate a set of more accurate random atomic displacements. These displacements are used in a multislice-style simulation to gauge the validity of the Einstein approximation. The phonon dispersion curve yields a small additional oscillatory structure in the thermal diffuse scattering (TDS) pattern. This does not produce significant changes in the annular dark field scanning transmission electron microscope (ADF-STEM) image signal, but could have a large impact on convergent beam measurements of bond charges.

A high efficiency annular dark field detector for STEM.

A new high efficiency annular dark field (ADF) detector for an HB501 STEM (Scanning Transmission Electron Microscope) has been constructed and tested. This detector uses a single crystal YAP scintillator and a solid quartz light pipe extending from the scintillator (inside the vacuum) to the photomultiplier tube (outside the vacuum). A factor of approximately 100 improvement in signal relative to the original detector has been obtained. This has substantially improved the signal to noise ratio in the recorded high resolution ADF-STEM images.

Superoxide anion production from human neutrophils measured with an improved kinetic and endpoint microassay.

Superoxide dismutase (SOD)-inhibitable reduction of cytochrome c is the basis for a widely used assay to measure superoxide production. We report novel modifications leading to a dual wavelength, high throughput simultaneous kinetic and endpoint microplate assay with high reproducibility. Neutrophils were isolated using a modified elutriation procedure to minimize priming and adherence during isolation. Cytochrome c reduction was measured in a microplate reader using 96-well polystyrene plates with a modified (Plastek A*) surface to prevent the adherence and consequent activation of PMNs. Comparison of Plastek A* treated and untreated plates revealed a statistically significant difference in basal as well as stimulated levels of superoxide production. Absorption measurements were made at both 550 nm, the absorption maximum of reduced cytochrome c, and 557 nm, an isosbestic point. A significant increase in both well-to-well reproducibility and sensitivity (detection limit) was realized by using the normalized 550-557 nm difference values compared to the 550 nm absorbance values alone. These modifications represent an improved method for handling and assessing the function of superoxide production, providing greater experimental reproducibility and lessening the perturbations caused by the microplate.

Digital reconstruction of bright field phase contrast images from high resolution electron micrographs.

The theory of bright-field image formation of thin specimens in a conventional transmission electron microscope is presented. The recorded image contrast is shown to be predominantly linear in the electron atom scattering amplitudes which are in general complex (possessing phase and amplitude). A linearized image model describing multiple images of varying defocus (defocus series) is derived. Image degradation is characterized by an instrumental transfer function (including spherical aberration, defocus and partial coherence), a finite signal-to-noise ratio and a Debye--Waller temperature factor. Using the minimum mean square error criterion, a new method of image reconstruction to recover the real and imaginary parts of the ideal phase contrast image from a defocus series is derived. This new method of image reconstruction reduces to the well known Wiener filter in the appropriate limiting conditions. A defocus series of micrographs taken on the Kyoto 500 keV electron microscope using a radiation damage resistant specimen of chlorinated copper phthalocyanine is processed. The signal-to-noise ratio of this series is found to be approximately 10. A resolution of approximately 2 A is apparent in the unprocessed images. The complex image reconstructed from this defocus series shows increased resolution in the real part of the image (approximately 1.4 A) and increased heavy/light atom contrast in the imaginary part of the image.