Synthesis and activity of the salicylic acid ester of bakuchiol in psoriasis-surrogate keratinocytes and skin substitutes.

BACKGROUND: Topical retinoids are effective in retarding skin ageing and restoring homeostasis in skin conditions such as psoriasis. However their adverse effects (AEs), which include irritation (retinoid dermatitis), photosensitivity and teratogenicity, limit their use and patient compliance. Development of retinoid analogues with minimal AEs would allow a broader and more compliant use. AIM: To synthesise a novel molecule, bakuchiol salicylate (bakusylan), with a modulatory gene expression profile similar to retinoids, using as reference three prescription retinoids: tretinoin, tazarotene and adapalene. METHODS: We hypothesized that because bakuchiol salicylate has a structure entirely different from existing retinoids, there would be at least a partial uncoupling of AEs from the skin-normalizing activity of this retinoid. This hypothesis was tested at the transcriptional level in psoriatic cytokine-treated cultures of keratinocytes and organotypic skin substitutes, using DNA microarrays and custom PCR arrays. RESULTS: Evaluation of the gene expression profile of bakuchiol salicylate revealed elimination of several components of the retinoid-like proinflammatory response and teratogenic signature, without a substantial loss of normalizing potential. A possible mechanism of action, consisting of keratinocyte desensitization to psoriatic cytokine signalling through inhibition of the signal transducer and regulator of transcription (STAT)1/3/interferon inflammatory signal transduction axis was also identified. CONCLUSION: Bipartite materials obtained by merging two skin-active entities with specific, complementary bioactivities, such as bakuchiol and salicylic acid, may yield a new class of functional retinoids.

The EGF receptor ligand amphiregulin controls cell division via FoxM1.

Epidermal growth factor receptor (EGFR) is central to epithelial cell physiology, and deregulated EGFR signaling has an important role in a variety of human carcinomas. Here we show that silencing of the EGF-related factor amphiregulin (AREG) markedly inhibits the expansion of human keratinocytes through mitotic failure and accumulation of cells with ⩾ 4n DNA content. RNA-sequencing-based transcriptome analysis revealed that tetracycline-mediated AREG silencing significantly altered the expression of 2331 genes, 623 of which were not normalized by treatment with EGF. Interestingly, genes irreversibly upregulated by suppression of AREG overlapped with genes involved in keratinocyte differentiation. Moreover, a significant proportion of the irreversibly downregulated genes featured upstream binding sites recognized by forkhead box protein M1 (FoxM1), a key transcription factor in the control of mitosis that is widely dysregulated in cancer. The downregulation of FoxM1 and its target genes preceded mitotic arrest. Constitutive expression of FoxM1 in AREG knockdown cells normalized cell proliferation, reduced the number of cells with ⩾ 4n DNA content and rescued expression of FoxM1 target genes. These results demonstrate that AREG controls G2/M progression and cytokinesis in keratinocytes via activation of a FoxM1-dependent transcriptional program, suggesting new avenues for treatment of epithelial cancer.

Early tissue responses in psoriasis to the antitumour necrosis factor-α biologic etanercept suggest reduced interleukin-17 receptor expression and signalling.

BACKGROUND: Antitumour necrosis factor (anti-TNF)-α therapy has made a significant impact on the treatment of psoriasis. Despite these agents being designed to neutralize TNF-α activity, their mechanism of action in the resolution of psoriasis remains unclear. OBJECTIVES: To understand better the mechanism of action of etanercept by examining very early changes in the lesional skin of patients with psoriasis responding to etanercept. METHODS: Twenty patients with chronic plaque psoriasis were enrolled and received etanercept 50 mg twice weekly. Skin biopsies were obtained before treatment and on days 1, 3, 7 and 14 post-treatment. Skin mRNA expression was analysed by quantitative reverse-transcription polymerase chain reaction and microarray; cytokine and phosphoprotein levels were assessed using multiplexed bead arrays. RESULTS: In etanercept responders, we observed no significant changes in interleukin (IL)-17A, IL-22 or interferon-γ mRNA or protein in the first week of treatment; however, there was a 2·5-fold downregulation of IL-17 receptor C (IL-17RC) mRNA (P < 0·05) after day 1, accompanied by decreased extracellular signal-regulated kinase-1/2 phosphorylation. Transcriptional analysis revealed that genes suppressed by etanercept significantly overlapped with IL-17A-induced genes, and a marked overlap was also observed between the genes suppressed by etanercept and by the anti-IL-17A agent ixekizumab. Finally we show that TNF-α enhances the expression of IL-17RC, and short hairpin RNA inhibition of IL-17R expression abrogates synergistic gene induction by TNF and IL-17A. CONCLUSIONS: These results suggest that the early responses of psoriasis plaques to etanercept may be due to decreased tissue responsiveness to IL-17A due to suppressed IL-17RC expression in keratinocytes, blunting the strong synergy between TNF-α and IL-17, which contributes to the maintenance of psoriasis lesions.

A scanned, focused, multiple transducer ultrasonic system for localized hyperthermia treatments. 1987.

A commercial diagnostic ultrasound scanner (Octoson) was modified for performing hyperthermia treatments. The temperature elevations were induced in tissues by four large, focused ultrasonic transducers whose common focal zone was scanned along a computer controlled path as determined from B-scan images. The system is described and the results of preliminary tests demonstrating some of its capabilities are given. Extensive tests with canine thighs and kidneys were performed. The blood flow to the kidneys was controllable, and thus tumours having different blood perfusion rates could be simulated. The results showed that the system is capable of inducing a local temperature maximum deep in tissues (up to 10 cm was tested) and that tissues with high perfusion rates could be heated.

Plastic and adaptive gene expression patterns associated with temperature stress in Arabidopsis thaliana.

Transcriptional profiling using DNA microarrays has become a widely used approach for identifying genes with important roles in stress-regulatory networks. In previous studies, genes exhibiting a plastic expression pattern with respect to stress and control treatments have been identified as candidates with putative roles in stress-response pathways. This approach, however, often identifies numerous genes, and it is difficult to discern which genes have major effects that impact the fitness of individuals under stress. In this study, we investigated the impacts of temperature stress (cold and heat) on gene expression in the Arabidopsis thaliana model system. We identified genes exhibiting plastic patterns of gene expression with respect to temperature stress, but in contrast to previous studies, we also considered the adaptive significance of genes by examining their expression patterns among 10 Arabidopsis ecotypes indigenous to a range of latitudes. Our findings support a general association between plasticity of gene expression and adaptive value. In comparison to non-plastic genes, genes exhibiting plastic expression patterns were associated with greater among-ecotype variation in expression levels, and such variation was more strongly correlated with geographical temperature gradients. Surprisingly, while more than 16,000 genes were associated with plastic expression patterns, significant evidence of both expression plasticity and adaptive value was obtained for only 43 genes. These selected genes represent strong candidates for future experimental investigations into the molecular basis of temperature acclimation in the A. thaliana model system.

Reduced inbreeding depression due to historical inbreeding in Drosophila melanogaster: evidence for purging.

An important issue in conservation biology and the study of evolution is the extent to which inbreeding depression can be reduced or reversed by natural selection. If the deleterious recessive alleles causing inbreeding depression can be 'purged' by natural selection, outbred populations that have a history of inbreeding are expected to be less susceptible to inbreeding depression. This expectation, however, has not been realized in previous laboratory experiments. In the present study, we used Drosophila melanogaster as a model system to test for an association between inbreeding history and inbreeding depression. We created six 'purged' populations from experimental lineages that had been maintained at a population size of 10 male-female pairs for 19 generations. We then measured the inbreeding depression that resulted from one generation of full-sib mating in the purged populations and in the original base population. The magnitude of inbreeding depression in the purged populations was approximately one-third of that observed in the original base population. In contrast to previous laboratory experiments, therefore, we found that inbreeding depression was reduced in populations that have a history of inbreeding. The large purging effects observed in this study may be attributable to the rate of historical inbreeding examined, which was slower than that considered in previous experiments.

Optimization of the scintillation detector in a combined 3D megavoltage CT scanner and portal imager.

A parametric study is described leading to the optimization of a custom-made scintillation detector with a relatively high quantum efficiency (QE) for megavoltage photons and light output toward a remote lens. This detector allows low-dose portal imaging and continuous cone-beam megavoltage CT acquisition. The EGS4 Monte Carlo code was used to simulate the x-ray and electron transport in the detector. A Monte Carlo model of optical photon transport in a detector element was devised and used as well as various irradiation experiments on scintillators. Different detector materials and configurations were compared in terms of the optical photon irradiance on the lens from on- and off-axis detector elements and the practical constraints regarding detector construction and weight. Effects of scintillator material, detector element size, crystal coating type, and reflectivity, combinations of different coatings on detector faces, scintillator doping level, and crystal transparency were studied. With scintillator thickness adjusted to give an 18% x-ray QE at 6 MV, the light output of CsI(Tl) was at least eight times higher than ZnWO4, BGO and NE118 plastic. Further, CsI(Tl) showed the smallest decrease in QE going from 6 to 24 MV. The off-axis reduction in emittance from the periphery of the detector was relatively small with a slight dependence on the type and reflectivity of the coating and the crystal thickness for a fixed detector element cross section. Light output was more strongly dependent on the reflectivity of lambertian coatings than specular ones. For a fixed detector element cross section, optimum coating type depended on crystal thickness. Typical CsI(Tl) crystals showed a relatively small variation in light output with changes in optical attenuation length. The optimum detector element was found to be CsI(Tl) coated on five faces with TiO2-loaded epoxy resin offering about a ten-fold improvement in light output per incident photon compared to typical metal/phosphor screens.

A cone-beam megavoltage CT scanner for treatment verification in conformal radiotherapy.

PURPOSE: A prototype scanner for large-volume megavoltage computed tomography (MVCT) in a clinical set-up is described. The ultimate aim is to improve treatment accuracy in conformal radiotherapy through patient set-up error reduction and transit dosimetry. MATERIALS AND METHODS: The scanner consists of a custom-built 2D CsI(Tl) crystal array viewed by a lens and a CCD camera. Image acquisition is synchronized with radiation pulses. The 2D projections resulting from a single continuous 360 degrees gantry rotation are reconstructed using a cone-beam tomography algorithm. Prior to reconstruction, the raw projections are calibrated and corrected for centre of rotation movement and accelerator output fluctuation. The performance of the system has been evaluated by reconstructing projections of open fields, test objects and a humanoid phantom. RESULTS: Hundreds of 2D projections can be acquired with a clinically-acceptable data collection time (about 2 min) and dose (approximately 40 cGy, with a possible four-fold reduction). A maximum density resolution of about 2% is achieved offering some soft tissue discrimination without using image enhancement tools. A spatial resolution of 2.5 mm is obtained. The reconstructed image intensity is linear with electron density over the range of interest. Coronal or sagittal slices through the 3D reconstruction of the humanoid phantom show a better delineation of structures than the corresponding portal images taken at the same orientation. CONCLUSIONS: A similar image quality to our current single-slice MVCT scanner is achieved with the advantage of providing tens of tomographic slices for a single gantry rotation. This work demonstrates the feasibility of clinical cone-beam MVCT and indicates how this prototype can be improved.

Rapid portal imaging with a high-efficiency, large field-of-view detector.

The design, construction, and performance evaluation of an electronic portal imaging device (EPID) are described. The EPID has the same imaging geometry as the current mirror-based systems except for the x-ray detection stage, where a two-dimensional (2D) array of 1 cm thick CsI(Tl) detector elements are utilized. The approximately 18% x-ray quantum efficiency of the scintillation detector and its 30 x 40 cm2 field-of-view at the isocenter are greater than other area-imaging EPIDs. The imaging issues addressed are theoretical and measured signal-to-noise ratio, linearity of the imaging chain, influence of frame-summing on image quality and image calibration. Portal images of test objects and a humanoid phantom are used to measure the performance of the system. An image quality similar to the current devices is achieved but with a lower dose. With approximately 1 cGy dose delivered by a 6 MV beam, a 2 mm diam structure of 1.3% contrast and an 18 mm diam object of 0.125% contrast can be resolved without using image-enhancement methods. A spatial resolution of about 2 mm at the isocenter is demonstrated. The capability of the system to perform fast sequential imaging, synchronized with the radiation pulses, makes it suitable for patient motion studies and verification of intensity-modulated beams as well as its application in cone-beam megavoltage computed tomography.

Extraction of primary signal from EPIDs using only forward convolution.

A model is presented in which the scatter signal in images obtained obtained by electronic portal imaging devices (EPIDs) is removed by a forward convolution method. The convolution kernel, kt(r) is a cylindrically symmetric kernel, generated by Monte Carlo, representing the scattered signal of a pencil beam at the image plane after the photons have gone through an object of thickness, t. A set of the kernels is presented and used to extract the primary signal. The signal from primary photons in the image, P(r), is extracted by an iterative method in which the essential assumption is that the scatter signal S(r) can be described by a superposition of the signal that would be obtained with the object removed from the beam, O(r), and the kernel kt(r). The thickness, t, that is used to choose the kernel, is directly related to P(r) by a simple exponential relationship; hence the thickness, t, of the object and the primary signal, P(r), are both iterated to better estimates through this procedure. The model is tested on Monte Carlo simulated data, where the extracted primary signal is compared with the "true" primary signal. Results are presented for a set of phantoms of uniform thicknesses up to 35 cm, and for field areas up to 320 cm(2), and for an inhomogeneous phantom containing a sphere of different density. The primary signal can be extracted to better than 1.5%, even when the original Scatter-to-Primary Ratio (SPR) is more than 25%. Finally, we have tested the model on EPID images, a nonuniform (breast) phantom is presented here. The breast phantom both have a curved external contour and contains a structure of a different density (lung). The radiological thickness of this breast phantom, as extracted using the above convolution model, was found to be within 2.8 mm (1 sd) of the true radiological thickness.

The optimum intensities for multiple static multileaf collimator field compensation.

A method of determining the optimum beam intensities for compensation using multiple static multileaf collimator fields is presented. In this method a histogram of the number of beam pixels against beam intensity is generated for the intensity-modulated beam (IMB). The intensity of each beam to be used is chosen to minimize the mean square deviation between each bin in the histogram and the closest beam intensity. This method has been applied to sample IMBs possessing one maximum and two maxima. For both cases, the use of uniform beam intensity increments is shown to be close to optimal. In the case with two maxima, the efficacy of irradiating both peaks simultaneously, rather than separately, has been studied and shown to be of potential benefit. The optimum intensities for an IMB for breast radiotherapy are also presented.

Dosimetric evaluation of compensation in radiotherapy of the breast: MLC intensity modulation and physical compensators.

BACKGROUND AND PURPOSE: Electronic portal images may be used to design the compensation required to maximise dose uniformity in the breast from opposed tangential beams. MATERIALS AND METHODS: Four methods of implementing the desired compensation have been studied: a simple wedge, a physical compensator in conjunction with a wedge; one open field plus four shaped multi-leaf-collimated (MLC) fields, and one wedged field in conjunction with three shaped MLC fields. Evaluation was performed using thermoluminescent dosimeters (TLDs) placed inside a phantom which was designed to mimic the human breast. The measured results are compared with both the prediction of the in-house compensation design software and with the dose predicted by the GE Target II planning system. The implications of each method for the time taken to plan and deliver treatment were analysed. RESULTS: The dose inhomogeneity, as measured at seven points in the central plane was greatest for the simple wedge (root mean square (rms) = 4.5%) compared to an open field plus four shaped MLC fields (rms = 2.2%), a wedged field plus three shaped MLC fields (rms = 3.3%), and the physical compensator (rms = 2.4%). The times required to plan and prepare these treatments varied considerably. The standard wedged treatment required under 15 min; both MLC-based and the physical compensator treatments required approximately 50 min. Differences of treatment delivery times were up to 8 min. CONCLUSIONS: These results indicate that the dose inhomogeneity can be reduced by beam intensity modulation designed using EPIDs.

The application of transit dosimetry to precision radiotherapy.

A method of using electronic portal imaging (EPI) for transit dosimetry is described. In this method, a portal image of the treatment field is first aligned with a digitally reconstructed radiograph (DRR) to geometrically relate the computed tomography (CT) scan, used to generate the DRR, with the EPI. Then the EPI is corrected for scatter within the patient to yield a map of primary fluence striking the detector. This is backprojected through the planning CT data set to yield a distribution of primary fluence within the patient. This distribution is then convolved with dose deposition kemels to yield a map of dose delivery within the patient. Such a distribution may be compared with the dose distribution resulting from the original treatment plan in order to evaluate the adequacy of the treatment. This method has been evaluated using a humanoid phantom. We find the transit dosimetry relative dose distribution when compared with film and thermoluminescent dosimeter (TLD) measurements and compared with our planning system to agree within 2% in the pelvic region of a humanoid phantom.

Scattered radiation in portal images: a Monte Carlo simulation and a simple physical model.

The scattered radiation in 6 MV radiotherapy portal images is analyzed. First, a quantity SPR* is studied, by means of Monte Carlo (MC) modeling. SPR* is defined as the ratio, on the central axis, of the signal due to scattered radiation to that due to the primary radiation. The detector model mimics a high-energy photon detector in the context of transit dosimetry. Second, a physical model of SPR* has been developed from first principles. For a cylindrical phantom, placed symmetrically about the isocenter, it predicts that SPR* depends on the area A at the isocenter of the circular field and the phantom thickness d as follows. SPR* = k0Ad(1 + k1d)(1 + k2A), where k0 = 0.0266(L1 + L2)2/(L1L2)2, k2 = - [L1(-2) + L2(-2) + (L1(-1) + L2(-1))2((2/3) + (3 kappa/2))]/2pi, L1 is the source-to-isocenter distance, L2 is the isocenter-to-detector distance, and kappa is the mean energy of the radiation beam (MeV/0.511). Constant k1, for which there is no simple expression, depends on L2. Comparison to the MC data shows that for 60 or= 50 cm. Third, experimental measurements of the scatter-to-primary ratio were obtained using our custom built imaging system mounted on a Philips SL25 linear accelerator. In the first experiment, A was varied from 40 to 400 cm2 with L1 = L2 = 100 cm with d = 20 cm. In the second experiment water depth d was varied from 0 to 28 cm with L1 = L2 = 100 cm and A = 200 cm2. The rms agreements between the MC data and the experiments were 0.0015 and 0.0045, respectively.

Design of compensators for breast radiotherapy using electronic portal imaging.

A novel method of designing intensity modulated beams (IMBs) to achieve compensation in external beam radiotherapy of the breast, without the need for CT scans, is presented. The design method comprises three parts: (1) an electronic portal image is used to generate a map of radiological thickness; (2) this map is then used to obtain an estimate of the breast and lung outline; (3) a TMR-based dose calculation algorithm is then used to determine the optimum beam profile to achieve the best dose distribution. The dose distributions calculated for IMBs were compared with those calculated for the use of simple wedges. The results for two patients studied indicate that the dose inhomogeneity for IMBs is +/- 5%, compared with a value of +/- 10% for a wedged plan. The uncertainty in radiological thickness measurement corresponds to a dosimetric error of +/- 2%. Other errors associated with outline estimation are typically less than 2%, with a largest value of +5% for one of the patients who had a large and highly asymmetrical breast. The results for the two patients studied suggest that the uncertainties in the method are significantly smaller than the improvement in dose uniformity produced.

Noise reduction by frame averaging: a numerical simulation for portal imaging systems.

We have studied the usefulness of both pre- and post-ADC frame summing for the purpose of reducing the effect of quantum noise and digitization noise in portal imaging systems. The study is based on the fluorescent-screen video-camera type of system. The study predicts the not-surprising result that provided the noise level at the ADC input is sufficiently large, the overall SNR can be increased by a factor of square root of M1M2, where M1 and M2 are the number of frames summed before and after the ADC. The study also predicts, somewhat unexpectedly, that there is an operating region in which increasing M1 actually decreases the SNR in the final image. To avoid this region M1 must be less than approximately 6 x 2(2B) (1 + -delta-1)1/2/(iaccf), where B is the number of ADC bits, -delta is the mean number of optical photons detected by the video camera per detected x-ray photon, iacc is the open-field number of detected x-ray photons per accelerator pulse per pixel, and f is the patient transmission factor. An equivalent statement is that the rms noise at the input to the ADC, sigma in, must exceed approximately 0.4q where q is the quantization interval of the ADC. It is possible that some systems operate in or close to this region. A second feature of this anomalous behavior is that the final image is not necessarily improved by increasing the number M2 of post-ADC-summed frames. For example, when sigma in/q = 0.2, there is no improvement in the overall rms error for M2 > 32.(ABSTRACT TRUNCATED AT 250 WORDS)

Reproducibility of patient positioning during routine radiotherapy, as assessed by an integrated megavoltage imaging system.

A portal imaging system has been used, in conjunction with a movie measurement technique to measure set-up errors for 15 patients treated with radiotherapy of the pelvis and for 12 patients treated with radiotherapy of the brain. The pelvic patients were treated without fixation devices and the brain patients were treated with individually-moulded plastic shells. As would be expected the brain treatments were found to be more accurate than the pelvic treatments. Results are presented in terms of five error types: random error from treatment to treatment, error between mean treatment position and simulation position, random simulation error, systematic simulator-to-treatment errors and total treatment error. For the brain patients the simulation-to-treatment error predominates and random treatment errors were small (95% < or = 3 mm, 77% < or = 1.5 mm). Vector components of the systematic simulation-to-treatment errors were 1-2 mm with maximal random simulation error of +/- 5 mm (2 S.D.). There is much interest in the number of verification films necessary to evaluate treatment accuracy. These results indicate that one check film performed at the first treatment is likely to be sufficient for set-up evaluation. For the pelvis the random treatment error is larger (95% < or = 4.5 mm, 87% < or = 3 mm). The systematic simulation-to-treatment error is up to 3 mm and the maximal random simulation error is +/- 6 mm (2 S.D.). Thus corrections made solely on the basis of a first day check film may not be sufficient for adequate set-up evaluation.

A proof that uniform dose gives the greatest TCP for fixed integral dose in the planning target volume.

In this note it is shown that for a fixed integral dose to the planning target volume, the highest tumour control probability (TCP) arises when the dose is spatially uniform. This 'uniform dose theorem' is proved both for (i) a specific TCP model based on Poisson/independent voxel statistics, and (ii) any model for voxel control probability having a specific shape with respect to increasing dose.

A randomised trial of patient repositioning during radiotherapy using a megavoltage imaging system.

Effectiveness of radiotherapy is dependent on the accuracy of beam alignment. Recent developments in megavoltage imaging allow real-time monitoring of beam placement. Maximum gains from this new technology can only be made if the information is utilised to correct patient positioning prospectively before the majority of a treatment fraction is delivered. We have developed and utilised an integrated megavoltage imaging system to perform a randomised trial demonstrating significant improvements in accuracy using treatment intervention techniques for pelvic radiotherapy. The mean field-placement accuracy improved from 4.3 mm to 2 mm and the proportion of treatments given with a field-placement error of > or = 5 mm decreased from 69% to 7%. This improvement in accuracy may enable smaller margins around the target volume to be chosen whilst ensuring complete target coverage at each treatment fraction.

Preliminary clinical performance of a scanning detector for rapid portal imaging.

A scanning megavoltage imaging detector, with associated image storage and analysis facilities has been developed. This produces images of the treatment portals in under 10 seconds, in a digital format, facilitating rapid, quantitative image analysis. Image quality is comparable to, and at some sites improves upon, that available from film. Clinical problems in the use of megavoltage imaging include limited field of view, loss of information at the field edge due to penumbra effects, degradation of the image by bowel gas, and difficulties in the detection of soft tissue-air interfaces. Possible solutions to these problems are discussed. The imaging system has been used to assess the random errors occurring during routine para-aortic nodal irradiation. The errors detected are small, with over 95% of set-ups lying within +/- 4.5 mm of the mean daily position. No differences were detected in the magnitude of random errors between anterior and posterior treatment fields.