Cervus elpahus elaphus).The cause of the bovine spongiform encephalopathy (BSE) epidemic in the United Kingdom (UK) was the inclusion of contaminated meat and bone meal in the protein rations fed to cattle. Those rations were not restricted to cattle but were also fed to other livestock including farmed and free living deer. Although there are no reported cases to date of natural BSE in European deer, BSE has been shown to be naturally or experimentally transmissible to a wide range of different ungulate species. Moreover, several species of North America's cervids are highly susceptible to chronic wasting disease (CWD), a transmissible spongiform encephalopathy (TSE) that has become endemic. Should BSE infection have been introduced into the UK deer population, the CWD precedent could suggest that there is a danger for spread and maintenance of the disease in both free living and captive UK deer populations. This study compares the immunohistochemical and biochemical characteristics of BSE and CWD in experimentally-infected European red deer (After intracerebral or alimentary challenge, BSE in red deer more closely resembled natural infection in cattle rather than experimental BSE in small ruminants, due to the lack of accumulation of abnormal PrP in lymphoid tissues. In this respect it was different from CWD, and although the neuropathological features of both diseases were similar, BSE could be clearly differentiated from CWD by immunohistochemical and Western blotting methods currently in routine use.Red deer are susceptible to both BSE and CWD infection, but the resulting disease phenotypes are distinct and clearly distinguishable. These invariably fatal neuro-degenerative disorders typically show lesions such as spongiform change, gliosis and deposition of the disease-associated form (PrPCervus elaphus nelsoni), which is closely related to the red deer (Cervus elaphus elaphus), the most frequently farmed cervid in the UK. Unlike other TSEs, CWD is the only TSE that is maintained in free-ranging animals with low population densities (2–3 deer per square mile), and surveillance data suggests an increasing prevalence of infection [Contaminated meat and bone meal (MBM) within concentrated rations fed to cattle was the cause of the BSE epidemic in the United Kingdom (UK ) and elsnfection . It is hnfection , perhapsnfection .Should BSE infection have been introduced to the UK red deer population, both the stability of the BSE agent through the species barriers and the high transmissibility of CWD, could account for spread and maintenance of BSE in free-living and captive UK deer. Due to the potential risk for human health from the consumption of contaminated deer products, it was deemed appropriate to determine the susceptibility or resistance of UK red deer to BSE and, if susceptible, to characterize the resulting disease phenotype in comparison with that of CWD in the same subspecies.We previously described the susceptibility of UK red deer to BSE infection by intracerebral injection , and the3.5 RIII mouse  units ID50/g; as with the BSE inoculum for i.c. challenge, this one also was provided by the TSE archive at VLA-Weybridge. Two and four negative control animals underwent identical procedures to the i.c and orally dosed deer, respectively, with sterile normal saline instead of brain homogenate. All experimental animal procedures were approved by the Moredun Research Institute Animal Experiments Ethical Committee and authorised under the UK Animals (Scientific Procedures) Act 1986.Six one year-old red deer were intra-cerebrally (i.c.) inoculated with 0.5 ml of a 10% homogenate of a brain pool of five BSE-positive bovine brains (BBP12/92); this experiment has already been described . A furthThis was carried out by the Canadian Food Inspection Agency . Four European red deer were challenged with 5 g of a pooled brain homogenate from four elk with clinical CWD. The inoculum was administered in three doses three days apart and although it was not titrated, it gave an optical density reading of >3.5 (upper limit of measurement) in the Bio-Rad TsSeE test.Deer challenged with BSE were clinically monitored daily and weighed monthly, until they reached a clinical end point established at the beginning of the experiment, as previously described . After dd. The tissues examined by IHC from BSE inoculated deer have been described in detail previously [Fixed samples were post-fixed in fresh 10% neutral buffered formalin and routinely processed for embedding in paraffin wax and light microscope examinations, either after staining with haematoxylin and eosin or immunolabelling for PrPeviously , and incThe IHC method has been detailed elsewhere ,12. Brieres-concentrated. After heating, 20 mg equivalent of samples were loaded in duplicate lanes onto pre-cast 12% bis-tris gels , and subjected to electrophoresis. The proteins were then transferred onto PVDF membranes, blocked (Bio-Rad blocking solution) and probed for 30 mins at room temperature with each of a panel of five antibodies: 6H4 , P4 (aa 156–164 ), and 12res bands, and for the affinities for each PrP antibody. For comparison, brain samples from CWD positive and negative elk, BSE positive cattle and scrapie positive sheep were used.The blots were assessed for molecular weights and proportions of the three PrPd profiling", and the discriminatory WB allow differentiation between sheep BSE and natural and experimental scrapie in sheep and in other species. The IHC "epitope mapping" and the discriminatory WB are based on the availability of specific epitopes on PrPd subjected to enzymatic attack "in vivo" (in intracellular lysosomes) and "in vitro" (by protease digestion), respectively [d are not labelled by 12B2 or P4. In contrast, in CWD and scrapie, the PrPd molecule is truncated further towards the N terminus and the labelling with those antibodies is maintained after "in vivo" or "in vitro" digestion. "PrPd profiling" is the most sophisticated technique for IHC strain differentiation: using a single antibody directed at the globular or C-terminal domains of the protein, this approach entails the subjective scoring (0–3) of different PrPd types  across different areas of the brain to produce a graphic profile, which is representative of the IHC phenotype [The IHC "epitope mapping" and "PrPectively ,20. It hhenotype ,12.d.All six deer challenged i.c. with BSE developed clinical disease between 794 and 1260 days post-inoculation with a mean incubation period of 1027 days. A detailed description of the clinical signs was provided in an earlier report . Brieflyd accumulation in the secondary follicles of rectal biopsies taken at 7 months post-infection.All four deer orally challenged with CWD started to show behavioural changes between 577 and 586 days post challenge; these progressed to definite neurological disease between 742 and 760 days post-challenge Table . ClinicaIn haematoxylin and eosin stained brain sections, spongiform change was widespread in all seven deer that developed clinical signs after BSE challenge regardless of the route. Vacuolation was seen in the neuronal perikaryon and neuropil of the brainstem, the molecular layer of the cerebellum, the thalamus, and the layers V and VI of the cerebral cortex Fig . Spongy Accumulation of PrPd in the brain. Widespread diffuse particulate labelling of the neuropil and intense intra-neuronal accumulations of PrPd were observed throughout the brains of BSE affected deer in sections labelled with BAR224 antibody. Peri-neuronal labelling was prominent in the dorsal motor nucleus of the vagus nerve (DMNV) and in the corpus striatum. Intraneuronal granular accumulations were prominent in nuclei of the medulla oblongata, such as the accessory cuneate, spinal trigeminal and posterior olivary, but were also present elsewhere in the brain with the exception of Purkinje cells. Notably, there was also intense labelling of the Golgi neurones in the granular cell layer of the cerebellum  and to globular domain (6H4 and Sha31) and no signal with the antibodies raised to the N-terminus of PrP , intracerebral cattle BSE (100%) and oral deer CWD (100%) included in this study suggest that there is a strong species barrier between cattle and deer in the case of BSE. Although it needs to be pointed out that the remaining five deer orally dosed are still alive and that infection cannot be ruled out, this species barrier would agree with the low attack rate found in intracerebral infections of cattle with the CWD agent .d in BSE infected deer is truncated at an epitope further towards the C terminus of the PrP molecule than it does in CWD. This differential truncation also takes place when exogenous enzymes are applied to brain samples as indicated by the discriminatory WB results.The examinations conducted on this experimental material show that infections of red deer with BSE and at least one source of CWD can be clearly differentiated by the same laboratory methods previously used to discriminate between BSE and scrapie in sheep ,25,26 and accumulation -the PrPd profile-, while these were highly consistent amongst BSE infected deer, some variability was observed in the CWD cases, with one deer showing a slightly divergent profile to the other three and also a considerably higher magnitude PrPd signal in all brain areas. This variability was also observed in the electrophoretic migration speed of PrPres in the red deer CWD samples, as previously described for elk, mule deer and white tail deer [d observed in those samples were indistinguishable from those in the obex of the CWD red deer recipients. Similarly, all four elk used in the inoculum demonstrated almost identical molecular profiles, in contrast with the CWD recipient red deer where some variability was observed. It is thus possible that the host species may have an effect on molecular and pathological phenotypes.As for the subjectively assessed patterns of IHC PrPail deer , raisingd found between the BSE orally and i.c. challenged deer may be due to differences in the route of access of infectivity to the brain, as proposed for sheep infected with BSE [d accumulates in the porencephalic lesion resulting from the repair process at the site of inoculation, and appears to spread to other neighbouring areas of the cerebral cortex [The minor differences in the topographical distribution of PrPwith BSE . In thisl cortex . This paThe fact that the circumventricular organs, which have leaky capillaries and appear to be involved in the neuroinvasion process associated with infectivity present in blood , were afd was detected in the enteric nervous system and in structures of the peripheral nervous system of the BSE infected deer, regardless of the route of challenge. It is difficult to tell whether that peripheral PrPd accumulation preceded neuroinvasion or resulted from centrifugal spread, but the lack of LRS involvement in those same deer rather supports the later possibility. This is unlikely to be due to the i.c. route of challenge, as the only orally-dosed deer succumbing to BSE was also negative for PrPd in lymphoid tissues, and as lymphoid involvement has been shown in BSE i.c. challenged sheep of several PRNP genotypes [PRNP allele [PrPenotypes . The absenotypes , and of P allele ,30.d in LRS tissues of BSE affected deer, as in natural cattle BSE, might be indicative of a low level of circulating infectivity and suggests that BSE in deer may not be contagious under conditions of natural exposure. Conversely, red deer infected with CWD showed widespread lymphoid involvement, which would be in agreement with the highly contagious nature of this infection in native North American cervids.The absence of PrPd truncation pattern both "in vivo" and "in vitro", and the predominantly intracellular accumulation of PrPd are features of deer BSE that are in contrast with those of deer CWD. However, only one of six deer developed disease after alimentary exposure to 25 g of a BSE brain pool homogenate after an incubation period of nearly 5 years; this suggests a strong species barrier but if a TSE in European red deer should ever be identified then BSE/CWD discrimination would be an urgent priority. To determine whether there are potential naturally occurring BSE-like strains and to determine the degree to which there is strain variation, it would be necessary to examine many more naturally occurring CWD cases. These results will support the ongoing European surveillance for natural TSEs in red deer and the further assessment of potential risk to human health.European red deer are susceptible to infection with the cattle BSE agent, not only by the intra-cerebral but also by the oral route, and although the clinical signs and spongiform change are similar to those of CWD in the same species, these two infections can be easily differentiated. The lack of lymphoid involvement, the PrPSM collated the experimental data and wrote the paper. MJ, LG and AB designed the study. SM, SS, LG, MJ and AB examined the tissues. MS, MC and PS performed the biochemical analyses. MD and HR challenged, managed and monitored the deer. All authors have read and approved the final manuscript.


VK600-1E mutation, the molecular hallmark of the solid variant of PTC. Histological and molecular evaluation of the surgical specimen confirmed this pre-operative diagnosis. Thus combining cytology to BRAF molecular analysis is useful to refine the cytological diagnosis of this variant also on FNC specimens.Papillary thyroid carcinoma (PTC) composed by predominant solid areas is diagnosed as a distinct variant on histological samples. Here we present a case of PTC recognized preoperatively by fine needle cytology as a solid variant. This diagnosis was made by combining cytology with the detection of the BRAF Appropriate positive (DNA from the neoplastic FRO cell line) and negative controls were included. The PCR product underwent elctrophoresis in agarose gel at the 100 V for 30 minutes Fig.  and it wMacroscopic examination of the resected specimen showed in the right lobe a 19 mm, well-delimited, non encapsulated lesion, nearly exclusively composed by solid sheets of tumour cells with typical nuclear features of papillary carcinoma Fig. . NuclearBRAF molecular analysis. This is an ideal marker in thyroid cytology since it is highly specific for PTC [BRAF mutation and the histological type of PTC was useful to predict on cytological samples the final precise diagnosis. The BRAF triplet deletion TGA 1799–1801 is the hallmark of the solid variant of PTC, having been reported only in this variant [BRAF triplet detection is of value as a predictive marker of solid variant on subsequently resected PTC.Modern cytology can exploit reliable molecular markers to refine the microscopic diagnosis. In the present study we report a combined cytological and molecular diagnosis of solid variant of papillary thyroid carcinoma. The cytology alone was not conclusive; the little amount of colloid, the elevated cellularity raised the suspect of a neoplasm; however only a slight degree of nuclear changes were observed: grooves and micronucleoli were sporadic, whereas intranuclear inclusions were not observed. The on-site evaluation of the Diff-Quick stained smear was very useful to specify the need of an additional pass to obtain material for  for PTC . Further variant ,13. HereThe author(s) declare that they have no competing interests.GT conceived the study and wrote the manuscript; RM and UM carried out the molecular study; IC evaluated the cytological sample; AF evaluated the results; MA evaluated the histological sample; LP participated in the design of the study. All authors read and approved the final manuscript.Written informed consent was obtained from the patient for publication of this case report and any accompanying images. A copy of the written consent is available for review by the Editor-in-Chief of this journal.


Some electron-affinic drugs, developed as hypoxic cell radiosensitizers, become selectively bound to the molecules of hypoxic cells by metabolism. This technique has been used to identify zones of chronically hypoxic cells in multicellular spheroids and animal tumours. Tritiated-misonidazole was administered to a patient with advanced melanoma 22 h prior to the surgical resection of a large metastatic s.c. lesion growing on the face. Autoradiographic analysis of histological sections revealed zones of intense labelling by the radioactive drug, indicative of tumour cells which were chronically hypoxic. This technique appears to provide an indirect measurement of tissue pO2 at the cellular level from which estimates of the tumour hypoxic fraction can be made. These data are encouraging as regards the development of 'sensitizer-adduct' procedures for the invasive and non-invasive measurement of hypoxia in both tumours and normal tissues.


The ever expanding epidemic of end-stage heart failure represents one of the greatest challenges of modern cardiovascular medicine. With medical treatments hampered by significant limitations, physicians caring for patients with advanced heart disease have turned to cardiac transplantation and durable mechanical circulatory assist devices as definitive therapies. These advanced therapeutic modalities are not widely available outside the United States and Europe, but nevertheless offer enormous potential for patients in the Arab Gulf suffering from end-stage heart failure. This review will discuss the management of end-stage heart failure in the Gulf States, with an emphasis on therapies best utilized within a framework of regional cooperation and coordination. The management of patients with end-stage heart failure has emerged as one of the major challenges in cardiovascular medicine. There has been considerable progress in the understanding of heart failure pathophysiology leading to medical therapies that impact the prognosis and symptoms of chronic heart failure.52 and diabetes mellitus in 23.7%.1517The incidence of cardiovascular disease and heart failure is projected to increase substantially in the Arab Gulf States as the region completes an epidemiological transition fueled by socioeconomic change.It is estimated that 5-10% of heart failure patients have end-stage, refractory disease.1927th century, digitalis glycosides such as digoxin reduce hospitalizations for heart failure and provide modest symptomatic relief, but they do not prolonged survival.4041The major advances in heart failure therapies have been primarily seen with systolic heart failure with a reduced ejection fraction. There is no debate that beta-adrenergic blockers, angiotensin converting enzyme (ACE) inhibitors, and angiotensin-receptor blockers (ARBs) offer an improved survival, and the frequency of their administration is emerging as a quality of care benchmark.In patients with diastolic heart failure, the most effective therapies will include diuresis, control of blood pressure, and maintenance of sinus rhythm.34344Most medical therapies have been directed toward NYHA Functional Class II-III or AHA/ACC Stage C heart failure and have a limited role once the evolution of heart failure have moved to Class IV NYHA or Stage D Tables  and 2. IMany heart failure patients with a low ejection fraction often also have interventricular conduction delay, as manifest by a widened QRS complex (>120 msec) on electrocardiogram from either a bundle branch block or nonspecific conduction delay. This delay creates dyssynchronous contraction of the left ventricle and can contribute to remodeling, mitral regurgitation and symptoms of congestion.26The primary goals in the management of end-stage heart failure are to reduce hospitalizations and to improve quality of life. Disease management programs have been shown to be effective in this regard, providing a structured continuity between the hospital discharge and the home setting.46Many patients with refractory congestive symptoms and low cardiac output despite conventional therapy will benefit from infusion of a positive inotrope.53Orthotopic cardiac transplantation remains the definitive therapy for terminal heart failure.59The primary barrier to cardiac transplantation in the Gulf States is the ad hoc organization of donor supply. While a limited number of centers have been able to perform the intermittent transplant, no program in the Gulf has achieved sufficient experience that would meet program certification goals of national regulatory bodies. Saudi Arabia is the only country within the Gulf region with a solid organ transplant organization, the Saudi Center for Organ Transplantation (SCOT), and King Faisal Specialist Hospital in Riyadh is the only heart transplant center. In 2008, the last year for which data is available, there were 19 cardiac transplantations in SCOT, preceded by 12 transplants in the year prior.Sadly, mortality is rapidly rising in the Gulf states from motor vehicle collisions as well as cerebral vascular accidents from poorly controlled hypertension—the most common conditions causes brain death while preserving the potential for organ donation. While automobile accidents and strokes present a significant challenge to public health authorities, they also represent an opportunity to coordinate donor management, procurement and distribution. The Gulf Cooperative Countries (GCC) have already established political, travel and trade alliances. Established in 1981, the six GCC countries include Saudi Arabia, United Arab Emirates, Kuwait, Oman, Qatar and Bahrain with a population base approaching 41 million and a unifying culture. Organization of solid organ transplantation dovetails with the GCC charter which specifically outlines a goal of integration and inter-connection between members states in all fields to encourage cooperation and joint ventures.Many existing organ procurement organizations cross political boundaries to serve the collective transplant need. In the United States, the New England Organ Bank represents 6 states and Bermuda, with a population base of 11.2 million. The Eurotransplant Interntational Foundation serves the transplant needs of Austria, Belgium, Germany, Luxembourg, the Netherlands, Slovenia and Croatia, with a population of 124.5 million and coordinated 6,600 solid organ donations. In order to address the current and future needs of all solid organ transplantation for the GCC, a coordinated transnational organ procurement organization representing the interests of the populations of all the Gulf States must be considered.And if any one saved a life, it would be as if he saved the life of the whole people. 64It is beyond the scope of this article to discuss the many religious and cultural issues surrounding solid organ transplantation. Organ donation, harvest and transplantation are a relatively new cultural concept in the Islamic and Arabic world and subject to a variety of interpretations. Renal transplantation, given its longer history, has wider acceptance than other solid organ programs.Given the immediate limitations to cardiac transplantation in the Gulf States and ultimate limitations of donor supply, ventricular assist devices (VAD) may be the most reasonable option for long-term hemodynamic support.665The clear survival advantage and quality of life for patients receiving VADs compared to medical therapy now justifies their use for circulatory support well beyond the transplant setting. In REMATCH, the implantation of a left-sided VAD was associated with a relative reduction in the risk of death of 48 percent during the entire follow-up period of 4 years and an absolute reduction in the mortality rate of 27 percent at one year.6972Prior to VAD implantation, all patients should be evaluated at cardiac centers with specific expertise in advanced heart failure.The ample costs associated with VAD implantation are contingent on the type of device used, length of hospital stay, country-specific health economics, and the nature of the underlying disease.Any cost-effectiveness analysis must also take into account the rapid evolution of technology used. Since the axial flow devices are smaller, simpler to use, and more durable, it can be anticipated that they will also be more cost effective. In addition, the cost of a new medical technology usually declines as it becomes more widespread and management issues standardized. Continued improvements to the next generation of devices can be anticipated to extend durability and reduce complication rates.80The management of heart failure is both a challenge and an opportunity for the health care systems of the Gulf region. Advanced therapies for terminal heart failure remain elusive. While cardiac transplantation remains a potential option, there are logistical limitations in the Arabic world at the present time that limit widespread applicability. The new technologies of mechanical circulatory assistance may be a better option for end-stage patients, and requires thoughtful analysis and candid discussion about the priorities of health care resource allocation. Heart failure patients in the Gulf are deserving of these advanced therapies that will improve the quality and length of their lives.


The role of the inferior parietal lobule (IPL) and superior temporal gyrus (STG) or subcortical pathways as possible anatomical correlates of spatial neglect is currently intensely discussed. Some of the conflicting results might have arisen because patients were examined in the acute stage of disease.We examined the anatomical basis of spatial neglect in a sample of patients examined in the post-acute stage following right-hemispheric vascular brain damage. Lesions of 28 patients with chronic spatial neglect were contrasted to lesions of 22 control patients without neglect using lesion subtraction techniques and voxel-wise comparisons.The comparisons identified the temporo-parietal junction (TPJ) with underlying white matter, the supramarginal gyrus, the posterior STG, and the insula as brain regions damaged significantly more often in neglect compared to non-neglect patients. In a subgroup of neglect patients showing particularly large cancellation bias together with small errors on line bisection damage was prevalent deep in the frontal lobe while damage of patients with the reverse pattern was located in the white matter of the TPJ.Considering our results and the findings of previous studies, spatial neglect appears to be associated with a network of regions involving the TPJ, inferior IPL, posterior STG, the insular cortex, and posterior-frontal projections. Frontal structures or projections may be of particular relevance for spatial exploration, while the IPL may be important for object-based attention as required for line bisection. Damage to the right cerebral hemisphere may lead to a severe impairment in spatial orienting towards the left side of space: spatial neglect ,2. The eSeveral factors other than differences in sample size and statistical power could explain these conflicting results. One important difference was the time of examination respective to lesion onset. Whereas in the study of Mort et al , patienta priori anatomical or etiological exclusion criteria. In addition, we evaluated brain lesions of a subgroup of neglect patients particularly impaired in line bisection with patients particularly impaired in spatial exploration and found evidence for an anatomical dissociation between these two groups of neglect patients.The present study evaluated the frequency of damage of the IPL and STG in 50 patients with right-hemispheric vascular damage, 28 presenting spatial neglect two months post-injury. In order to avoid a selection bias in our sample, we did not apply any 50 patients with recent vascular damage to the right cerebral hemisphere admitted to our rehabilitation service were studied. All patients gave written consent according to the Declaration of Helsinki, and the study was approved by the Ethical Committee of the University Hospital Geneva. We included patients with a recent unilateral brain injury, the only exclusion criteria being the presence of previous brain damage or absence of a visible brain lesion on MRI or CT. The presence of motor and somatosensory impairment was assessed using standardized neurological examination. Visual fields were examined using standard confrontation testing. Patients were attributed to the neglect group if they a) showed at least some signs of contralesional unawareness in everyday actions  and b) had scores indicative of spatial neglect in at least two of three formal neglect tests. The three tests used to assess neglect were the following:participants were required to cancel out small bells dispersed among other symbols printed on a horizontally presented sheet of A4 paper. Five or more (of 15) contralesional omissions were considered as a sign of spatial neglect .patients marked the middle of 18 lines ranging from 12–20 cm aligned horizontally on an A4 sheet of paper. The deviation from true midline was calculated in percent of the line length. A mean ipsilesional bias of more than 10% differentiated the patients' score significantly (significance level: α < .05) from the mean of neurologically healthy participants , and waspatients were asked to copy simple forms . Their drawings were rated with respect to the presence of contralesional omissions, with 0 indicating absence of elements on the left, 1 lack of precision or distortion of the left side, and 2 a complete drawing. Scores below 2 were considered consistent with spatial neglect.Using these criteria, 28 patients were attributed to the neglect group and 22 patients to the control group. The clinical characteristics of these two patient groups are shown in Table Lesion mappings were based on manual transposition of lesions from individual MRI-scans or CT-scans into the template brain of the Montreal Neurological Institute (MNI). Since this is a retrospective study, we used the brain scans that were available for individual participants at the time of the clinical examination. T2-weighted MRI had been performed in 52%, T1-weighted MRI in 12% and CT in 36% of patients. The clinical MRI protocol comprised a 25-slice acquisition (slice-thickness: 6 mm). In most patients (74%) brain images were acquired at least 5 days post-injury .In order to minimize errors of transfer from the brain scan to the template image, we proceeded in the following steps : i) slic2-test comparing the frequency of involvement  in each group  independently for each voxel. The position of significant differences is indicated in x-, y-, and z-coordinates of the Talairach-space  and IPL damage  was significantly higher in neglect compared to control patients. However, a limitation of this approach is that the STG has a greater anterior-posterior extension and is located more centrally within the vascular territory of the middle cerebral artery (MCA) compared to the IPL. Therefore, neglect patients may have damage to the STG simply because there is a high probability that MCA infarcts damage at least some portion of the STG. This hypothesis may be tested by computing the sensitivity  and specificity (i.e. the probability that no neglect occurs when the region is spared) of STG and IPL damage as predictors of the occurrence of spatial neglect  between neglect and control patients. All voxels shown in the figure were damaged more often in the neglect than the control group. There were essentially two clusters of voxels for which the test was statistically significant, an anterior and a posterior cluster. The anterior cluster reached from the inferior to the superior insular cortex and further into the white matter of the frontal lobe anterior to the horn of the lateral ventricle. Within this cluster a group of voxels differentiating best between both groups  was situated slightly anterior to the head of the caudate nucleus . The posterior cluster was located at the TPJ and reached into the supramarginal gyrus and the inferior postcentral gyrus. The STG was also involved, but only with its most posterior part neighbouring at the TPJ. Within the posterior cluster, a group of voxels in the white matter beneath the TPJ  differentiated best between neglect and control patients .We further evaluated the incidence of damage to specific cortical and subcortical areas with a voxel-wise analysis. In this analysis, a χparisons . Figure A voxel-wise comparison including only patients with infarction showed a similar result Figure , the notThus, our comparison between neglect and non-neglect patients revealed an anterior and a posterior cluster differentiating best between these two groups. Both, the IPL and the STG were damaged significantly more often in neglect than in control patients, although only the posterior part of the STG was involved.Previous studies have suggested that patients with a strong bias on cancellation tasks may anatomically dissociate from patients with a large line bisection error. While frontal or subcortical injury has been reported in the former , the latFigure Using a lesion comparison method previously applied by several authors the present study confirms the findings of previous studies evaluating the anatomy of spatial neglect. Certainly, several aspects of the methodology of our study may be criticized. Thus, lesion analysis based on CT-scans is limited because of the low resolution of CT-images in the z-plane. Though in most patients MRI sequences were acquired, which have better resolution and better sensitivity for cerebral infarction than CT-scans, in some patients only T1-weighted MRI had been performed which does not always differentiate well between intact and injured tissue. In addition, CT- and MRI-scans were acquired for clinical purposes and consequently differed with respect to the orientation of axial slices as well as the imaging protocol. Although we tried to minimize these problems as possible by including only those patients in whom damaged brain tissue could clearly be identified, they might have influenced the lesion mappings.Considering these difficulties it is remarkable how consistent our results are with the findings of previous studies. When comparing the lesions of chronic patients carefully selected for the presence of behavioural and test criteria of spatial neglect with non-neglect patients, we identified three sites that were significantly more often damaged in neglect patients: a region centred around the TPJ reaching into the supramarginal gyrus and the posterior STG; the white matter anterior to the caudate nucleus; and the insula. Several older ,8 and moThe second region that was significantly more often damaged in neglect compared to non-neglect patients was the posterior insular cortex. Spatial neglect after isolated insular lesions has occasionally been reported , and freWhile brain tissue surrounding the TPJ and insular cortex were reliable predictors of spatial neglect in our study, the association of the frontal white matter rostral to the caudate nucleus with spatial neglect was less reliable. This region was very small and was significantly more often damaged in neglect than non-neglect patients only when patients with haemorrhagic injury were included. Thus, though damage to caudate-frontal projections might lead to hypoperfusion of the prefrontal cortex  the impoOur results are of particular relevance in view of the recent controversy concerning the brain region whose damage is critical for the occurrence of spatial neglect. While Mort and collaborators  identifiSeveral factors might explain why the critical lesion associated with spatial neglect was localized more anteriorly in the study of Karnath et al.  comparedAn alternative possibility is that differences in patient selection resulted in contrasting anatomical findings. The patients of Karnath et al.  were exaFinally, it is appealing to consider the controversy about the critical brain damage leading to spatial neglect in light of the hypothesis that neglect might result from intra-hemispheric disconnection. A recent study using electrical stimulation of patients undergoing surgery of right temporo-parietal glioma reported that stimulation of the superior longitudinal fasciculus (SLF) resulted in greater line bisection errors than stimulation of the right IPL or posterior STG . The SLFNeurophysiological, neuropsychological, and functional imaging studies indicate that spatial attention is subserved by a distributed network of cortical and subcortical regions involving the IPL, STG, prefrontal cortex, and the insula ,29,40. IThe authors declare that they have no competing interests.LG and RP acquired clinical data and performed the lesion analyses; RP analyzed the data; LG, RP and AS drafted the manuscript. All authors read and approved the final manuscript.


The emerging epidemic of overweight/obesity in adolescents in Ho Chi Minh City, Vietnam underlines the importance of studying the metabolic syndrome in Vietnamese adolescents who are becoming progressively more inactive. No study in Vietnam has examined the association of metabolic syndrome with moderate to vigorous physical activity (PA) levels among adolescents. We aimed to examine this association in a sample of urban adolescents from Ho Chi Minh City.A cross-sectional assessment was conducted in 2007 on a representative sample of 693 high-school students from urban districts in Ho Chi Minh City. Metabolic syndrome was defined according to the International Diabetes Federation criteria and physical activity was measured with Actigraph accelerometers. The association between physical activity and metabolic syndrome was assessed by using multiple logistic regression models.Overall 4.6% of the adolescents and 11.8% of the overweight/obese adolescents had metabolic syndrome. Elevated BP was the most common individual component of the metabolic syndrome (21.5%), followed by hypertriglyceridemia (11.1%). After adjusting for other study factors, the odds of metabolic syndrome among youth in the lowest physical activity group  were five times higher than those in the highest physical activity group (>103 minutes/day) . Metabolic syndrome was also positively associated with socioeconomic status .A more physically active lifestyle appears to be associated with a lower odds of metabolic syndrome in Vietnamese adolescents. Socio-economic status should be taken into account when planning interventions to prevent adolescent metabolic syndrome. Metabolic syndrome is a clustering of cardiovascular disease risk factors that includes glucose intolerance, hypertension, elevated triglycerides, low HDL cholesterol, and obesity . This clThis syndrome continues to increase in both developed and developing countries, but has already become a major threat to global public health. It is especially of concern when it affects children and adolescents, as a consequence of increasing rates of childhood obesity and more sedentary lifestyles -11. The Studies in adults and also in adolescents reveal that physical activity (PA) - a modifiable lifestyle factor - is strongly associated with clustering of metabolic syndrome components -18. ReceVietnam is undergoing a socio-economic and nutrition transition especially in large cities, such as Ho Chi Minh City, where lifestyles are becoming more sedentary with lower levels of PA, and diets more energy dense with higher fat content. The nutrition transition has lead to a threefold increase in the prevalence of overweight/obesity over the last five years among adolescents in Ho Chi Minh City . The emeAlthough the association of metabolic syndrome with different levels of objectively assessed PA -16 and sA cross-sectional assessment was conducted in 2007 on a representative sample of high-school students from urban areas of Ho Chi Minh City. These students had been recruited into a cohort study from a sub-sample of students from urban districts identified in a survey in 2004 . The samTwo trained data collectors took the anthropometric measurements of the students using standard methods . Weight 2). The subjects were classified as overweight/obese by applying the age and sex specific International Obesity Task Force (IOTF) BMI cut off points  aUsing the same criteria, the prevalence of metabolic syndrome found among overweight and obese Vietnamese adolescents  was higher than that were reported for South Korean adolescents (1.5% and 14.7% for overweight and obesity in 2005) , and forA number of difficulties were encountered when trying to compare the findings from our study with the results from other countries. The estimates of the prevalence of metabolic syndrome from different countries were based on different reference data, different cut off values, or different criteria making cross country comparisons difficult. A further difficulty was the limited number of surveys assessing metabolic syndrome in adolescents from Asia, especially in urban populations as in our study.The most common individual component of the metabolic syndrome in Vietnamese adolescents was elevated BP, followed by hypertriglyceridemia. This contrasts with the Korean studies that found glucose intolerance was the dominant feature of the metabolic syndrome . Among TThe overweight/obese children in our study had significantly higher odds of having the metabolic syndrome compared to the non-overweight/obese counterpart. This finding is consistent with the observation that in children, obesity is strongly associated with metabolic syndrome ,11. AfteThe beneficial effect of physical activity for metabolic syndrome could be explained by its influence on body composition. Skeletal muscle is the most insulin sensitive tissue in the body. Physical activity has been shown to improve skeletal muscle insulin sensitivity and reduce insulin resistance by improving glucose transport in muscle cells and improving peripheral microcirculation ,37. AddiIt has been emphasized that a sustained amount of moderate to vigorous physical activity on a regular basis can induce and maintain beneficial effects on metabolic syndrome in adolescents . FurtherMany studies have revealed that the prevalence of the metabolic syndrome significantly increases with increasing socio-economic status in both developing ,42 and dIt was also mentioned that the prevalence of metabolic syndrome increased when age increased regardless of gender ,45. In tThere are conflicting findings from previous studies about the association of pubertal status and metabolic syndrome in adolescents. While studies have found no relationship between pubertal status and the likelihood of metabolic syndrome in adolescents ,16, otheSimilarly, we also did not find an association between gender and metabolic syndrome. Although, there seemed to be some gender differences with regard to the different components of the metabolic syndrome, these were all non significant, which might have been due to the somewhat limited statistical power of the study.The key strengths of this study included the representative study sample of adolescents from Ho Chi Minh City that improved external validity of the study and the use of an objective measure of physical activity with accelerometers. We also used an age-appropriate measurement tool to identify metabolic syndrome in these adolescents.A major limitation was the lack of any time dimension in assessing factors associated with metabolic syndrome due to the cross-sectional design of the study. Because both the outcome and the risk factors were examined at one point in time, we do not know whether these factors preceded or followed the onset of metabolic syndrome. An example of the inability of the study design to determine between cause and effect is the relationship between obesity and physical inactivity. Another limitation of this study was that the association between sedentary behaviors, including the time spent watching television and playing video and computer games, and the metabolic syndrome was not examined. Furthermore, the sample size of the study might have been a limitation as well, evidenced by the large confidence intervals for example of the moderate/vigorous physical activity odds-ratios for metabolic syndrome. Despite these limitations, our findings contribute to understanding the association between physical activity and the metabolic syndrome. These findings suggest that many adolescents in Ho Chi Minh City might need to spend more time on moderate to vigorous physical activity to minimize their risk of developing the metabolic syndrome.Physical activity is important for metabolic health in children and adolescents. Increased levels of physical activity at a young age may help reduce levels of chronic non-communicable diseases in the future. Resources should be allocated to provide support to develop effective campaigns for physical activity promotion to reduce the prevalence of metabolic syndrome during childhood in Ho Chi Minh City. These efforts to promote increased levels of physical activity in youth should engage all levels of society including families, schools, communities, and government agencies.BP: Blood pressure; BMI: Body mass index; AOR: Adjusted odds ratio; LDL-c: Low density lipoprotein cholesterol; HDL-c: High density lipoprotein cholesterol; TG: Triglyceride; IDF: International Diabetes Federation.The authors declare that they have no competing interests.THHDN, HKT participated in the design, carried out the study, performed the statistical analysis and drafted the manuscript. HvdP carried out the transformation of the accelerometer data and advised about analysis of the measures of physical activity. PK contributed to the statistical analyses. MJD participated in the design of the study and the analytical strategy and contributed drafting the manuscript. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2458/10/141/prepub


Transgenic modification of plants is a key enabling technology for developing sustainable biofeedstocks for biofuels production. Regulatory decisions and the wider acceptance and development of transgenic biofeedstock crops are considered from the context of science-based risk assessment. The risk assessment paradigm for transgenic biofeedstock crops is fundamentally no different from that of current generation transgenic crops, except that the focus of the assessment must consider the unique attributes of a given biofeedstock crop and its environmental release. For currently envisioned biofeedstock crops, particular emphasis in risk assessment will be given to characterization of altered metabolic profiles and their implications relative to non-target environmental effects and food safety; weediness and invasiveness when plants are modified for abiotic stress tolerance or are domesticated; and aggregate risk when plants are platforms for multi-product production. Robust risk assessments for transgenic biofeedstock crops are case-specific, initiated through problem formulation, and use tiered approaches for risk characterization. Agricultural biofeedstocks are critical for decreasing petroleum dependence through sustainable biofuels production. Continued rapid improvements in both biofuel resources and processes are needed if agricultural biofeedstock crops are to significantly address concerns about the depletion of fossil fuel reserves, domestic energy security and greenhouse gas emissions as contributors to climate change . VariousImplicit in the successful movement towards a biofuel future is the use of modern biotechnology, including transgenics, as an enabling technology ,5,6. TraAs for any technological innovation, the use of a particular transgenic concept for biofeedstocks production must undergo commercial, regulatory and public scrutiny in terms of its impact on the environment, human and animal health and sustainability . Such coDespite their more recent development, risk assessment frameworks for transgenic plants are being widely implemented ,16-20 anIn consideration of the fact that risk is the joint probability of exposure and the consequence of exposure, the conventional risk assessment process describes exposure and its consequence (an adverse effect or harm) in four steps: hazard identification, dose-response, exposure characterization and risk characterization . For traBacillus thuringiensis [Bt crops [Evidence of hazard - the recognized potential of a substance to cause harm to human health or the environment - is not common for transgenic plants and, therefore, the broader consideration of potential harms warranting evaluation is identified through problem formulation (see following discussion). This approach is in keeping with the recognition that problem formulation and scoping (inclusive of hazard identification) needs increased emphasis in risk assessment . Similarngiensis ,32. ThusBt crops ).The transgenic plant risk assessment adheres to a weight-of-evidence approach that considers comparative risks. The weight-of-evidence approach allows for the consideration of how, for a particular case, the preponderance of evidence identifies risk to human health or the environment. This directs decision-making away from a narrowly focused evaluation of a specific endpoint and, instead, considers risk within a fuller ecological context. A sequential lines-of-evidence approach  providesImplicit to transgenic plant risk assessment is a case-specific consideration developed on the basis of comparability . The priFor biofeedstock crops equivalence for many attributes of the plant is not intended. For instance, desirable changes may represent altered profiles of oils or enzymes, altered metabolic profiles for stress tolerance, or widely altered phenotypes of wild or semi-domestic species so as to better respond to agronomic management. For any of these concepts the consideration of equivalence still holds with a focus on the intended changes and establishing for all other plant attributes their equivalence to the non-modified comparator.The phenotype is generally recognized as the key to understanding comparability on the basis of familiarity and equivalence relative to a baseline. Phenotype refers to observable characteristics of an organism described as physical or biochemical traits. What constitutes a phenotype should reflect a biologically relevant level of detail consistent with the particular risk comparison being made. Through a consideration of a specific transgenic plant (an event) with respect to appropriate comparators, comparability describes the intended change brought about by the transformation and establishes that the transformation is limited to the intended phenotypic change. Thus, the transgenic plant phenotype is compared to baseline phenotypes with a focus on the difference found through the comparison. When the degree of equivalence is established, the subsequent risk assessment can focus on the change. The nature of phenotyping and the way that change is represented through genetic engineering of plants will be an area for continued clarification as scientists anticipate novel innovations such as transcription factors, changes in metabolic profiles, or RNA interference.Zea mays) stover recovery and use for energy production or through crop use for multi-product production . However, it will also entail substantial shifts in production systems, as when annual row crop production shifts to perennial biofeedstock production. In these instances, the risk assessment process can be applied within a decision-making context to better understand the risks (and/or benefits) arising from replacement of conventional crops and cropping systems with new technologies, including plants transformed through transgenics. The principle of comparability will consider biofeedstock crops and cropping systems relative to the baseline of existing crops and their management as the comparator.A further aspect of the comparative risk assessment for biofeedstock crops is the consideration of alterations in agroecosystems and their management when current generation food crops are supplanted by biofuel crops. This will initially represent gradual changes, for instance with respect to maize  protein arises from the toxicity of the expressed protein to a specific lepidopteron pest, European corn borer, which is of economic importance in corn production. Because this plant-expressed Bt protein is active against lepidopteron species, an assessment of the risk to non-target lepidopteran species inhabiting corn production systems is warranted.' The problem statement focuses a broad-based concern (impact of Bt maize pollen) to a specific consideration (maize expressing Cry1A(b) protein and non-target lepidopteron species inhabiting maize production systems) and, thus, distills the problem to an analytically tractable form.Problem context represents the activity that establishes the parameters for the risk assessment . The conet al.  in formuProblem definition proceeds from the problem statement to the identification of postulated significant risks that warrant further analysis for a particular case and leads to a specific analysis plan . The proThe problem formulation will, ideally, develop a concise problem statement, a risk hypothesis, a conceptual model and an analysis plan. Together, these elements represent the design for the risk assessment and set the bounds for the risk characterization and the way that risk findings will be reported and considered.The problem statement is an outcome of the consideration of problem context and translates a broad regulatory need to a specific statement of need/intent for the risk assessment which is further clarified and articulated as a risk hypothesis. A risk hypothesis represents an assumption regarding the cause-effect relationships among attributes of the risk characterization, including sources, exposure routes, endpoints, responses, and measures relevant to the risk assessment . The risThe conceptual model and analytical plan are critical products coming from the problem formulation . The con50) or a chronic no observed adverse effects concentration (NOAEC) may be specified as the relevant effect measurement. Secondly, measures of exposure in the environment are described through a causal pathway to contact or co-occurrence of the transgenic plant with the entity of value. These measures of stressor exposure are most often expressed as estimated environmental concentrations (EEC) that are developed on the basis of the casual pathway for exposure. Oftentimes, the analysis plan will also be concerned with measures of system or receptor characteristics. These are characteristics of the ecosystem that influence the behaviour, location, life history characteristics or system attributes relevant to the entity of value. For instance, in the case of non-target risks, this may involve the determination of the abundance and distribution of the entity of value  at a relevant life stage within a landscape or region.The analysis plan is a plan of work that outlines the analytic and interpretative approaches which will be used to characterize risk. Measures of effect (assessment endpoints) should be established that represent measurable changes relevant to the risk characterization. For instance, if the effect measure relates to a toxicological hazard, an acute effect concentration (EC50). Stochastic risk assessments more frequently compare exposure exceedance probabilities to an effect measurement; as for instance the percent of occasions where exposure exceeds the NOAEC for the entity of value. In many instances, there may be an established regulatory threshold for concern that defines both the appropriate risk formulation as well as the result that would trigger regulatory concern. For instance, in the case of a first tier assessment for transgenic plant effects on non-target arthropods, an RQ < 0.1 would be considered evidence of no likely harm [Finally, the analysis plan will establish the appropriate risk formulation to be considered in the risk characterization. The risk formulation represents the way in which the exposure measurement is related to the effect measurement. In its simplest form the risk formulation may be a ratio  of exposure to effect measurements . If, instead, they had focused on the pathway of exposure to define the specific stressor - Cry1A(b) protein in their study - and its environmental concentration, a conclusion as to no probable harm would have been more readily obtained.The analytical phase of any risk assessment is initiated with lower tiered testing which conservatively addresses broad questions using simple experimental designs. Any subsequent tests at higher tiers are more realistic and complex. Higher tiers of testing are triggered within the risk assessment process when there is recognition that the degree of potential harm or analytical uncertainties requires more exacting determination of probable risk. Since higher tiered tests are only prompted by the risk assessment process, an iterative approach effectively focuses consideration to the most relevant concerns and conserves time resources. Furthermore, to effectively address risk assessments which may initiate from a variety of concerns, the tiered scheme must allow for a non-linear approach, so as to allow the most relevant lines of evidence to be brought forward. For instance, Rosi-Marshall et al.  developeThe analytical plan and subsequent risk findings need to consider uncertainty as an inherent part of the risk assessment. Both lack of knowledge and variability need to be specifically considered with respect to the risk characterization. Lack of knowledge can be reduced through data acquisition, but can never be eliminated as an unresolved residual in the determination of risk. Variability, on the other hand, is inherent to risk analysis which is comprised of measurements of parameters; it cannot be reduced but it can be better described with improved information . For newAddressing the uncertainty of the risk characterization should reflect the degree of understanding needed to describe behaviour of the transgenic crop within the bounds of a conservatively cast threshold for harm. Given that there is sufficient certainty that the behaviour of the transgenic crop does not exceed a conservative threshold that could lead to harm, greater precision is unnecessary for the purposes of risk assessment. For example, if we are sufficiently certain that a RQ is < 0.1, and this represents a threshold for concern, a more exact value of the RQ does not represent greater certainty in the risk finding for the purposes of decision-making.Second order, or 'iatrogenic' risk,  can contThe concept of using a tiered system and recognized defaults for risk assessment is an important method for dealing with uncertainties . Within Use of comparative risk assessment is a further means whereby uncertainty is reduced by the consideration of carefully matched comparators  in order to focus on aspects of the comparison that represent substantive, consequential changes in the crop or system under consideration. By focusing on aspects of the crop or system that are changed for the transgenic biofeedstock crop relative to a baseline comparator, there is a reduction in the number of parameters and assumptions that must be considered. For instance, if the change brought about via plant transformation is to express a single unique protein in a plant, then the focus can be largely on the protein and its expression system, given that the principle of comparability  has been addressed.Increasing transparency in the risk assessment process by establishing problem context is integral to the relevance of risk assessments to both narrowly defined regulatory considerations and broader-based societal decision-making . TranspaOnce the standard for decision-making is understood, the strength of risk assessment as originally detailed by the Red Book paradigm  can be min planta production of key bioprocessing enzymes as a means to improve production efficiency and economy  [a priori reason to anticipate that environmentally stable enzymes will have direct effects to non-target organisms, but rather they may have indirect effects on critical ecological services such as nutrient cycling and decomposition. Thus, the need for environmental risk assessment is initiated from an ecological value context (Appendix 1). In terms of human health, confinement conditions and consequences of acute episodes of low-level occurrence in the food supply provide the relevant context for initiation of a food safety risk assessment, even though these products are intended for non-food uses [Early generation innovation for bioenergy crops has emphasized rations) ,43. For rations) -48, laccrations)  and amylrations)  for use ood uses .5 hectares could be devoted to high amylase maize production in the USA, with environmental loadings of thermostable alpha-amylase expressed in grain conservatively estimated as 0.14 kg/ha for residues of high amylase maize production [The need for case-by-case approaches to risk assessment is shown for contrasting cases where bioprocessing enzymes are expressed in maize as aids for ethanol production. Highly thermostable forms of alpha-amylase and endoglucanase have been expressed in maize kernel starch endosperm , ubiquitoduction . In contoduction , there ioduction . The difoduction , the resFor plant recombinant enzymes, as represented by the forgoing cases, a tiered process for assessing environmental exposure and effect provides an efficient means to characterize risk in a manner proportional to the reasonably anticipated impact. For instance, a tiered process for discerning novel enzyme impacts to soil processes could proceed in a hierarchical fashion from (i) characterization of fundamental attributes of the molecule and its interaction with soil colloids; to (ii) characterization of chemical reactivity; (iii) determination of environmental degradation and persistence; and - as needed - culminate with (iv) impact assessment for critical soil processes. The overarching goal of such a tiered approach is to provide regulators and data developers with effective means whereby - through a stepwise process - the environmental fate and effects of diverse types of bioactive transgenic proteins introduced to the environment can be assessed with regard to impact on soil processes. Similar tiered processes also can be implemented for other aspects of the risk assessment. For instance, from the perspective of human health, an early tier focus on exposure characterization can contribute to the determination of the degree to which the consequence of admixture of the industrial product within the food supply should be evaluated in a higher tier assessment .For these industrial enzyme cases, exposure characterization is a useful first step in problem scoping. However, since for these cases the problem formulation initiates from uncertainty regarding the impact to an ecological value , exposure analysis alone will be insufficient to establish risk. Further problem definition can determine the appropriate analytical plan for risk characterization. The analytical plan for more fully characterizing risk in this instance would focus on measurements of the ecological value .A limitation of many current studies ostensibly designed to address risks of transgenic plants to the environment is a tendency to focus on structural elements of the ecosystem, such as a specific non-target species, rather than on functional endpoints such as ecological services . ThorougThe use of transgenics to confer plant tolerance to environmental stresses, such as drought, chilling or salts, has been envisioned as a means to improve crop productivity  and numeet al. [The risk assessment paradigm for stress tolerant transgenic crops is similar to that for existing transgenic crops. However, there is a recognized need for increased emphasis on aspects of the risk assessment that deal with invasiveness potential and the environmental and human health aspects of the alteration of plant composition . Appropret al. . Nicksonet al.  developeet al.  would leet al. . In thisEucalyptus genetically engineered for freeze tolerance and targeted for introduction as a biomass crop in forest plantations in the Southeastern USA exemplifies a transgenic biofeedstock crop where the risk assessment focuses on the invasiveness of the whole plant [Eucaliptus grandis × E. urophylla) has been cultivated in southern Florida where it is a familiar, non-invasive species. This finding, however, is not consistent with the assignment of E. grandis to an invasiveness watch list for Florida [Arabidopsis C-Repeat Binding Factor [Eucalyptus plantations to supplant existing tree plantations may raise questions regarding the ecological community structure [le plant . The pre Florida . As a prg Factor , the trag Factor . In additructure . These iNumerous life-cycle analyses have considered the various costs of biofuel (ethanol and biodiesel) production ,65. And Co-production and extraction of heterologous proteins from biofeedstock crops is a critical consideration for the sustainability of cost-effective biofuels production . HeterolHuman health and environmental risk assessment are critical to the development of plant multi-product concepts, in view of the consequences arising from their in-field confinement and downstream processing, segregation and channelling . Again, Panicum virgatum) and poplar (Populus) for ethanol production [Jatropha curcas) for biodiesel production [Glycine max) [The long-term success in developing sustainable bioenergy resources is frequently tied to perennial herbaceous and woody plants such as switchgrass  . In thosine max) . GeneticIssues regarding the weediness and invasiveness of the transgenic plant, as well as of gene flow into wild populations, will be of paramount concern in risk assessments of forest and range plants which are domesticated and otherwise modified for use as biofeedstock crops. Given the reality of gene flow as a natural phenomenon, emphasis in the risk assessment should be placed on the consequences of transgene occurrence within native populations. The consequences will vary, dependent on the receiving environment and the ecological value of receptor populations. For considerations of weediness, invasiveness and gene flow there are effective models for analysis of plant introductions , invasivIssues of gene flow are not restricted to the transgenic biofeedstock crop. For instance, for switchgrass introduced into the upper midwestern USA many of the same questions of gene flow impact to local ecotypes exist for a non-transgenic southern variety of switchgrass as there will for a transgenic switchgrass variety. Regulatory statutes differ, however, such that, while the consequences of gene flow from the transgenic variety will always be explicitly considered in a regulatory-compliant environmental risk assessments, the need to do a similar assessment for a newly introduced non-transgenic biofeedstock crop will vary depending on the regulatory jurisdiction. Furthermore, consequences of crop-to-crop gene flow may be relevant in certain risk assessments and not others, depending on the operative statute. Nevertheless, the fore-mentioned analytical models are able to address gene flow and its consequences in any of these situations.The regulatory safety and public acceptance of a given transgenic biofeedstock crop are informed through the science-based process of risk assessment. Frameworks for the risk assessment of transgenic plants are becoming well-established and provide a logical approach for considering the human health and environmental consequences of various sustainable biofeedstock crops. In applying these frameworks to biofeedstock crops, special emphasis is needed regarding comparative risks of new versus established agronomic systems and for environmental release within a given region. In addition, aggregate risks arising from use of crops as multi-product production platforms will need to be more comprehensively considered and developed. The strength of risk assessments for transgenic biofeedstock crops depends on flexibility for case-specific determinations which are designed on the basis of problem formulation and use sequential approaches for data development and risk characterization.EC: effect concentration; EEC: estimated environmental concentration; NOAEC: no observed adverse effects; RQ: risk quotient.The author declares that they have no competing interests.Bacillus thuringiensis are selectively active on Lepidoptera. Cry1A(b) protein is expressed in maize for the targeted control of European maize borer ) and may harm sensitive non-target Lepidoptera if exposed in and around fields of Cry1A(b) maize. Monarch butterfly (Danaus plexippus) occurs in and around maize fields where it may be exposed to Cry1A(b) maize pollen in its diet.Proteins from the Cry1 class of delta-endotoxins of Dietary exposure to Cry1A(b) maize pollen will not adversely affect Monarch populations in and around maize fields to a greater extent than does non-transgenic maize pollen when exposure occurs to at environmentally relevant concentrations.50 for purified toxins incorporated into an artificial diet relative to control; Tier 2: Dose-response for larval weight for Cry1A(b) pollen present on food source (milkweed) relative to a non-transgenic control. Exposure characterization - Estimated environmental concentration (EEC) for Cry1A(b) present in pollen contaminating milkweed leaves. Risk characterization - Risk is formulated as a risk quotient (RQ = EEC/EC50) with a regulatory level of concern being RQ > 0.1.Hazard is established on the basis of stressor class. Effect characterization - Tier 1: ECColeomegilla maculata (De Geer)) may be adversely affected when feeding on prey that have ingested toxins expressed in potatoes expressing Cry3A toxin. Development and fecundity may additionally be related to feeding on pollen as a supplemental food source.Where transgenic insect-resistant (IR) crops are grown, field monitoring shows shifts in beneficial insect abundance relative to non-transgenic fields. Agroecosystems where transgenic IR crops are grown may have trophic level effects affecting occurrence and distribution of beneficial insects relative to comparable agroecosystems where transgenic IR crops do not occur. Ladybird beetle (Leptinotarsa decemlineata (Say)) and Cry3A potato pollen can be eaten by ladybird beetle without adverse effects on their survival or predation potential relative to eating non-intoxicated Colorado potato beetle and non-transgenic potato pollen.Cry3A-intoxicated Colorado potato beetle  may introduce novel RNA sequences into the environments where RNAi crops occur. Persistence of these RNA sequences may create off-target effects in receiving environments.Persistence of plant-expressed RNAi in the soil environment is no different than for wild-type short length RNA sequences that are introduced to the soil environment.There is uncertainty as to the nature of off-target unintended effects that could occur. Determination as to whether there is a causal pathway for exposure to soil biota addresses the extent to which unintended effects are possible. Lack of persistence will significantly reduce probable harm to off-target biota.Extracellular soil enzymes provide important ecological services relative to the dynamics of organic matter mineralization and carbon substrate stabilization. Soil cellulase enzymes, in particular, are important for degradation of plant cellulose residues and therefore impact organic matter mineralization rates which, in turn, influence soil carbon stabilization. Thermostable endo-1,4-β-glucanase expressed in maize may increase activity in the degradation of plant residues incorporated to soil to affect the dynamics of soil carbon stabilization.Rates of organic matter mineralization in soil receiving plant residues from high glucanase maize will not differ from that observed for wild type enzymes when considered under environmentally relevant conditions and concentrations.Uncertainty as to consequence, so seek a determination of no difference between the transgenic and wild-type enzyme, in order to be conservatively protective of ecological-value.


Class V cavity preparations were prepared on the buccal and lingual surfaces of the extracted premolars with occlusal margins in the enamel and gingival margins in the cementum/dentin. The teeth were divided into five groups of 10 teeth each and 20 cavity preparations per group. In the experimental groups, cavities were treated with Single-Bond, Adper Prompt, i-Bond, Clearfil S3, and G-Bond as the dentin bonding agents. After the application of the dentin bonding agents, the cavity preparations were restored with resin composite (Clearfil APX). The specimens were thermocycled, stained with methylene blue dye, and sectioned to evaluate the dye penetration. Data were analyzed using the one-way ANOVA test  and Dunn’s procedure for pairwise comparison of the data.This study showed that at the coronal margin and the apical margins, the preparations treated with Clearfil S3 showed significantly less leakage than the other groups. Enamel margins provided better marginal sealing than dentin/ cementum margins.The study demonstrated that Clearfil S3 bond had a better sealing ability at both coronal(enamel) and apical (dentin/cementum) margins compared with the other dentin bonding agents used. Vital dentin is an extension of certain pulpal tissues presenting the first line of defense against patient hypersensitivity and long-term bacterial microleakage that can lead to recurrent caries and pulpal irritation. The most significant factor in determining resistance to recurrent caries, postoperative sensitivity, marginal staining, and pulpal damage is the ability of the restorative material/ adhesive to seal the restorative interface with the adjacent tooth surface.2th generation bonding systems. The claim of the manufacturer is that this bonding system will eliminate mixing and etching, while, at the same time, accomplishing the priming and the bonding of the dental surfaces. and self-etching primer technique (6th and 7th generation DBA).[Although enamel adhesion is a predictable and established entity, an adequate bond to dentin is more difficult to achieve. To overcome this challenge, technological advancements of dentin adhesives have, at this time, evolved into two trends: total acid-etching techniques [5ion DBA).Self-etch adhesives, composed of aqueous mixture of acidic functional monomers that are generally phosphoric acid esters, do not require a separate acid-etch component and subsequent rinsing procedures. Self-etch systems also purportedly enhance marginal integrity and reduce or eliminate patient symptoms.The results showed that at the coronal (enamel) margins of the cavity, Clearfil S3 exhibited less microleakage than the other bonding systems, followed closely by Single- Bond and G-bond. The i-Bond and Adper Prompt showed significantly greater microleakage at the coronal margins and Adper Prompt showed the highest microleakage among all the adhesive systems that were compared.The probable explanations for these results of Clearfil S3 include:9The type of solvent used in the adhesive systems. Instead of acetone, Clearfil S3 Bond uses alcohol as the primer component solvent. It has been proposed that acetone-based adhesives require the use of a wet bonding technique with a relatively small window of opportunity to achieve optimal hybridization.The Clearfil S3 Bond formulation includes a proprietary “Molecular Dispersion Technology,” enabling a twophase liquid, hydrophilic/hydrophobic component homogenous state at the molecular level, reportedly resulting in reduction and/or loss of water droplets at the adhesive interface and, therein, a superior bond.Also, the 10-methacryloyloxydecyl dihydrogen phosphate adhesive monomer molecular structure allows for decalcification and penetration into the tooth structure, creating a chemical bond to calcium.There was a significant difference in the microleakage between Single Bond with Adper Prompt and i-Bond.The probable explanations for these results of Single Bond include:Single bond combines the functions of the primer and adhesive components of the conventional three-step adhesive system and has alcohol and water as solvents in its composition.The presence of alcohol in the adhesive increases the diffusion into the dentin, thereby enhancing the adhesion. The moisture of dentin tubules pulls the alcohol into and in between the tubules, taking the resin with it. The alcohol and moisture then vaporize from the substrate leaving behind the resin.There was significant difference in the microleakage between G-Bond when compared with Adper Prompt and i-Bond.The probable explanations for these results of G-Bond are:The interface formed is totally different from that of the interface formed by earlier bonding materials. The surface of the dentin is decalcified only slightly and there is almost no exposure of the collagen fibers. This suggests that an extremely thin (300 nm or less) interface is formed and that in this area, functional monomers contained in the bonding material react with hydroxyapatite at the “nano” level to form insoluble calcium. Therefore, the interface formed by G-Bond is expected to be stronger and more durable than that formed by other bonding materials.It would appear appropriate to call the interface exhibiting this a nano interaction zone, or a reacted layer at the “nano” level, as opposed to the traditional hybrid layer appellation.G-Bond’s 5% filler further seals the tubules and decreases pulpal sensitivity. There are fewer voids between the thin adhesive layer and the dentin, which minimizes microleakage and prevents microbial invasion.The results showed that i-Bond showed significantly greater microleakage at the coronal margins when compared with ClearfilS3, G-Bond and Single-Bond and less microleakage than Adper Prompt.13The probable explanations for these results of i-Bond include:The technique protocols (drying procedures) associated with i-Bond proved to be labor-intensive and may not be realistic for some practitioners in a clinical setting.Possible explanations for this occurrence include multiple applications (manufacturer’s instructions) of the i-Bond adhesive onto the preparation surfaces and increased waiting periods prior to light polymerization.The most extensive microleakage of the dye was seen with Adper Prompt as compared with all the other adhesives.The probable explanations for these results of Adper Prompt include:th generation dentin bonding agents can explain the poor results in terms of microleakage.It is claimed that the absence of the fillers in the 6It was also stated that there is a possibility that the lack of a separate primer may reduce the infiltration depth or the wettability of the dentin adhesives, thereby reducing the sealing adhesion and the sealing capacity.None of the groups of the dentin bonding agents prevented the microleakage at the restoration–tooth interface.8158The results also showed that at the apical (dentin/cementum) margins of the cavity, again, Clearfil S3 exhibited less microleakage than the other bonding systems followed closely by G-Bond. The i-Bond, Single Bond, and Adper Prompt showed significantly more microleakage at the apical margins, and Adper Prompt showed the highest microleakage among all the adhesive systems that were compared.The results also showed that the microleakage at the enamel margins is less compared to that of the microleakage at the dentin/cementum margins.16These results were in accordance with the previous results.7The most probable reasons for the less microleakage at the enamel margins compared to that of the microleakage at the dentin/cementum margins are:The hypermineralization of the dentin surface and subsequent collapse of the collagen fibrillar network, cavity configuration (C Factor), dentin tubule orientation to the cervical wall (CEJ), organic content of dentin substrate and movement of dentinal tubular fluids, incomplete alteration/removal of the smear layer by acidic primers for adequate demineralization and hybrid layer formation, and inefficient infiltration/ penetration of primer components into the demineralized collagen fibrillar network. Solvent carriers  in the adhesive agent reacted differently with varying degrees of surface moisture present in the tubules.In class V cavities, located at approximately 1 mm from the CEJ, dentinal tubules are oriented parallel to the cervical wall. A classic hybrid layer formation is absent in the dentinal margins, and this absence is an important cause of leakage.[in vitro, this can be a screening apparatus for ensuing in vivo studies. Previous research indicates that data obtained from in vitro microleakage testing may be useful but not always necessarily reproducible in clinical in vivo settings.[Microleakage tests provide adequate screening methods, possibly determining clinical success and longevity of the adhesive systems. Although this study was conducted settings.The results of this study demonstrate that the dynamic nature of the dentin substrate morphology is indeed an important factor and, possibly, an insurmountable impediment for perfect adhesion of restorative materials to the tooth structure. Clinical trials should be performed for carious and noncarious Class V cervical lesions to assess the performance of these new adhesive systems before definite conclusions can be formulated.


Giant cell tumor (GCT) of bone is a locally destructive tumor that occurs predominantly in long bones of post-pubertal adolescents and young adults, where it occurs in the epiphysis. The majority are treated by aggressive curettage or resection. Vascular invasion outside the boundary of the tumor can be seen. Metastasis, with identical morphology to the primary tumor, occurs in a few percent of cases, usually to the lung. On occasion GCTs of bone undergo frank malignant transformation to undifferentiated sarcomas. Here we report a case of GCT of bone that at the time of recurrence was found to have undergone malignant transformation. Concurrent metastases were found in the lung, but these were non-transformed GCT. Giant cell tumor of bone is a locally destructive tumor that occurs predominantly in long bones of post pubertal adolescents and young adults, where it occurs in the epiphysis. The majority are treated by aggressive curettage or resection. Histologically, giant cell tumor of bone classically shows many large multinucleated giant cells with interspersed haphazardly arranged mononuclear cells, and the nuclear features of both elements are described as similar. Some tumors also have areas with a fascicular or storiform pattern devoid of giant cells resembling a benign fibrous histiocytoma. Vascular invasion outside the boundary of the tumor can be seen. The rate of local recurrence varies among centers and is influenced by the completeness of surgical treatment, with high speed burring, adjuvants, and bone cement adding to the effectiveness of curettage treatment. UnresecThis study was performed with the approval of the Rush University IRB, ORA#: 09092501-CA01. The study included detailed clinical information, imaging and pathology. Tissue was fixed in 10% buffered formalin at room temperature and dehydrated and paraffin embedded in overnight processing. Immunohistochemical stains were performed as follows: The following antibodies were used on Ventana Benchmark and the manufacturer's solution CC1 for antigen retrieval.: CD4  1:10 dilution, CD43  prediluted, P63  1:500. Ki-67 immunostaining was performed on the Dako autostainer PLUS with FLEX Envision chemistry using clone MIB-1 , at 1:400 dilution after citrate buffer antigen retrieval under pressure in a microwave oven. Images were captured on an Olympus BX41 microscope with a Spot Insight color camera with Spot Advanced software.A 29 year-old male was diagnosed with a giant cell tumor of the left proximal tibia at an outside institution in January 2005. He underwent intralesional curettage followed by heat cauterization and methacrylation. Eight months later, in September 2005, the patient had a local recurrence of the tumor which was treated with cement removal and repeat curettage followed by argon beam ablation and repeat cementing.The patient presented at this institution in June 2009 with severe pain and disability of the left leg at the site of the previous excisions. Radiographs demonstrated a large lucency around the cement Figure . ConcurrHistological sections of the tumor from presentation showed classic giant cell tumor of bone with the typical histological patterns Figure . Most arThe gross specimen from the tibial excision consisted of fragments of pink-tan and red-brown tissue with pieces of cement and bone fragments. The metastatic tumor nodules resected from the lung Figure  grossly Giant cell tumors of bone are neoplasms of stromal cells that recruit a mononuclear population of hematopoietic origin . These mIn karyotypic analysis of giant cell tumors of bone, end-to-end fusions of various chromosomes, termed telomeric associations, are seen in most tumors in a subset of cells, and these have been localized by FISH studies to the CD68 negative spindled stromal component. At loweThis case illustrates a sarcoma arising at the site of a locally recurrent GCT of bone, with concurrent non-sarcomatous metastases. Local recurrence of GCT of bone is common, occurring 10-20% of the time-18, and Diagnosis of transformation relies on overt malignant cytological features, as necrosis and scattered mitoses can be seen in the usual "benign" giant cell tumor of bone. Immunohistochemical studies are not required for the diagnosis of transformation. Usually there is no evident lineage of maturation, but cases of osteosarcoma have been reported,20-24. TWe report here the case of a giant cell tumor of the proximal tibia of a 29 year old man treated with aggressive curettage that recurred 4 years later with malignant transformation and "benign" metastases to the lung. Recurrence, malignant transformation, and metastasis with "benign" morphology all occur in giant cell tumor of bone. This is the first report of all three occurring in a single patient. The influence of local recurrence on malignant transformation and pulmonary metastases is largely unknown. Malignant transformation has been reported most frequently with radiation even in nonrecurrent tumors. Similarly, benign pulmonary metastases have been reported to occur in recurrent and nonrecurrent tumors. The current report adds little to the discussion of the fate of locally recurrent giant cell tumor of bone. The molecular pathways leading to giant cell tumors of bone are still largely uncharacterized.The authors declare that they have no competing interests.IM reviewed the pathology and wrote the discussion of the molecular biology. AB formatted the article. SY and AM reviewed the pertinent literature. RG obtained the approval of the institutional review board and patient's consent. SG prepared the clinical case report. All authors read and approved the final manuscript.Written informed consent was obtained from the patient for publication of this case report and accompanying images. A copy of the written consent is available for review by the Editor-in Chief of this Journal.


Each year in North America, approximately 6 million individuals suffer a fracture of whom 5–10% demonstrate delayed healing or nonunions.Bone healing can be promoted by a number of novel therapeutics including bone stimulators. The most common bone stimulators include low-intensity pulsed ultrasound (LIPUS) and pulsed electromagnetic field therapy.The use of electric stimulation in the treatment of fractures nonunion dates back to the mid-1800s.Given the increased morbidity and costs associated with nonunions in our health care system, the identification of alternative bone healing therapies is warranted. LIPUS and electrical stimulation offer two noninvasive approaches to the promotion of healing of nonunions (and fresh fractures). The use of bone stimulators represents a multimillion dollar market globally; however, indications and evidence for each type of bone stimulator is not always transparent.et al. is an overview on the currently available fundamental, preclinical, and clinical evidence on the biologic rationale and therapeutic efficacy of electrical stimulation devices in long-bone nonunions. A clinical series by Gupta et al. shows a clinical evidence of the use of pulse electromagnetic stimulation in nonunion tibial diaphyseal fractures. Petrisor et al. present a systematic review on extracorporeal shock wave therapy and its use in fracture management. The symposium ends with a systematic review on the economic evaluation of bone stimulation modalities and a discussion on available therapies, indications, and protocols for fracture healing in India.The current symposium provides an in-depth review of the current evidence for the two most common bone stimulators, LIPUS and electrical stimulation. We further provide new data for a lesser known bone stimulator, extracorporeal shock waves. Our symposium brings together experts in the field to provide insight into the basic cellular mechanisms of action as well as the clinical evidence base for fresh fracture healing and nonunions. The manuscript by George Methews extentively covers the newer developments in biology and biomechanics, fixation devices to affect the care of musculoskeletal injuries. Galkowski highlights the need for effective non-invasive therapies of bone stimulation to shorten the fracture healing time by decreasing the use of invasive procedures. Ultrasound therapy is covered in detail in three articles. The article by Gregory J. Della Rocca reviews the scientific evidence of cellular effects of ultrasound to accelerate normal fracture healing and promote fracture healing in a compromised tissue bed. The systematic reviews on LIPUS in fresh fractures and nonunions attempts to evolve the evidence of its use in the management of fresh fractures and nonunions. The article by Kuzyk RT and Schemitsch EH provides insight into the basic science of electrical stimulation. The article by Kooistra Over the last several years, the concepts and ideas attributed to and labeled collectively as evidence based orthopedics have become a part of daily clinical lives, and clinicians increasingly hear about evidence based guidelines, care paths, and questions and solutions. This symposium of papers supports the new paradigm of evidence based orthopedics by providing an up-to-date critical appraisal of bone stimulators in 2009. Structured randomized controlled trials to compare the efficacy and safety, and a cost-effective analysis of bone stimulation therapies in comparison to invasive implant surgery are needed.