Stylophora pistillata were lopped of three different genets, their skeletons marked with alizarin red-S, and divided haphazardly into three morphometric treatment groups: (I) upright position; (II) horizontal position, intact tip; and (III) horizontal position, cut tip. After 1 y of in-situ growth, the 45 surviving ramets were brought to the laboratory, their tissues removed and their architectures analyzed by 22 morphological parameters (MPs). We found that within 1 y, isolated branches developed into small coral colonies by growing new branches from all branch termini, in all directions. No architectural dissimilarity was assigned among the three studied genets of treatment I colonies. However, a major architectural disparity between treatment I colonies and colonies of treatments II and III was documented as the development of mirror structures from both sides of treatments II and III settings as compared to tip-borne architectures in treatment I colonies. We did not observe apical dominance since fragments grew equally from all branch sides without documented dominant polarity along branch axis. In treatment II colonies, no MP for new branches originating either from tips or from branch bases differed significantly. In treatment III colonies, growth from the cut tip areas was significantly lower compared to the base, again, suggesting lack of apical dominance in this species. Changes in branch polarity revealed genet associated plasticity, which in one of the studied genets, led to enhanced growth. Different genets exhibited canalization flexibility of growth patterns towards either lateral growth, or branch axis extension . This study revealed that colony astogeny in S. pistillata is a regulated process expressed through programmed events and not directly related to simple energy trade-off principles or to environmental conditions, and that branch polarity and apical dominance do not dictate colony astogeny. Therefore, plasticity and astogenic disparities encompass a diversity of genetic (fixed and flexible) induced responses.The high morphological resemblance between branching corals and trees, can lead to comparative studies on pattern formation traits, best exemplified in plants and in some cnidarians. Here, 81 branches of similar size of the hermatypic coral  In multicellular organisms, the level of integration among bodily components dictates the final functional performance of the entire organism A further challenging topic is the study of organisms' architectures, made of multiple genetically identical modules, at several levels of organization, which are physiologically and structurally integrated Pseudopterogorgia bipinnata, Sànchez and Lasker In various modular organisms, including plants In branching corals, architectural characteristics can be deduced from traits at three hierarchical levels of organization, the individual polyps Stylophora pistillataS. pistillata, characterized by a continuum of architectural design with several distinct stages. Each stage was marked by its own characteristic morphometric parameters. We presupposed S. pistillata. Special attention is given to whether apical dominance and branch orientation are important in ruling colonial architectures of branching-corals'.To elucidate further the rules that govern colony development in branching forms, we studied plasticity of colony astogeny and branch to colony trajectories in the Indo-Pacific branching coral in situ growth, 45 of the 81 fragments (55.6%) survived and developed into colonies of various shapes. These included 15 colonies from treatment I , 14 colonies from treatment II  and 16 colonies from treatment III . Upon collection, each colony was dried, measured, and photographed from all angles and 22 morphometric parameters (MPs) were taken , treatment II (42±14%) and treatment III (23±3%) differed significantly from each other  as compare to genotype H (6.01±1.04gr) and J  between tips or bases of the two polarity manipulated treatments (II compared to III); and (2) between tip (either intact or cut-off) and base of the same branch. Five most relevant MPs for this comparisons were chosen: (1) nB ; (2) %nB (percentage of branches developed from tip (TBB) or from base (BBB) of the total number of branches in the colony); (3) TBL ; (4) EV ; and (5) %Nx, the number of branches from orders 2 to 4 as part of the total number of branches .Comparisons between the treatments revealed no significant differences in any tested MPs between treatments II and III for all the genets.3) compared to the base  revealed no significant differences between all five MPs studied in genets H and I  to coral colony astogeny and to address the possible existence of apical dominance, a well-documented phenomenon in the plant world, in shaping coral architectures. Results of this, and earlier studies on the branching coral Stylophora pistillata ramets, grown from similar size branches, and sub-cloned from three different coral genets. Ramets were haphazardly divided into three morphometric settings  or treatment II colonies , genotype-based differences emerged in treatments III . We found that altering branch orientation, in addition of trimming the branch tip (upper 0.5 cm), triggered species-specific and colony-specific architectural reactions. Genotype H exhibited an increase in percentage of down-facing branches (%DGB) as compared to the other two genotypes, genotype I gained more weight (W1) as compared to the other genotypes and genotype J showed an increase in percentage of branch order 2 (%N2) as compared to the other two genotypes. Other MPs, where differences were not yet resulted in significant values (because of high variation in the results) and were noticeable to the eye, were the developed total number of branches (nB) and ecological volume (EV). Genotype H showed 19% and 15% increase in total number of branches when comparing treatment I to II and I to III, respectively, and 45% and 7% in ecological volume, respectively. Genotype I exhibited increases of 115% and 143% in total number of branches when comparing treatment I to II and I to III, respectively, and 123% and 186% in ecological volume, respectively. Genotype J displayed 44% and 17% increase in nB, respectively, 27 and 32% and 39% in EV, respectively. As stated, these values however, were not significantly different from each other's.S. pistillata mode of astogeny S. pistillata branching system, as in the case of the soft coral Nephthea sp., where cutting the terminal polyps in young colonies did not change the way the colony developed While the traditional test for apical dominance in plants entails removal of the apical bud and measuring the effects on the dormant buds lower on the branch Stylophora settings II and III in the present study, Acropora branches developed terminal polyps at both sides of the branch. However, while in the Acropora treatment Stylophora treatment did not show any phototropism impact as new branches initiated up and down trajectories, without obvious preference for light.The impacts of branch orientation and polarity on morphometric parameters regulating colonial astogeny was revealed earlier by Kawaguti Stylophora species-specific colonial astogeny from vertical oriented branches, most new branch initiations developed along the branch.Changing branch orientation affected the way morphology developed, the complexity of the branching system (represented by numbers of branch generations), and the growth of new branches from branch tip or along the branch. When the initial branch orientation changed from horizontal to vertical, the percentage of high order of branches (N2 compare to N3 or N4) was higher than that of the horizontal branch. Altering the orientation of the branch from vertical to horizontal might result in shifting energy (evident by developing of new branches) from the original branch tip area (intact or cut-tip branch) to the opposite branch end, the base. It is of major interest to note that new branches developed from both tip ends rather than from the whole length of the branch, resembling induced positional information Stylophora pistillata is highly regulated as in whole colony scenarios Acropora millepora and Pocillopora damicornisAstogeny of cut branch in Stylophora pistillata, Meroz et al. It is evident that coral colonies, isolated branches, and spat not only ‘sense’ environmental cues but also ‘discern’ their special position. Working on two branching coral species, including Stylophora pistillata , were carefully detached from the substrates by chisel and hammer. Each colony represented only a single genet, as colonial fragments of this species in Eilat do not resume development in situ in clear plastic bags with alizarin Red S solution  sinTwenty-two morphometric parameters MPs;  were meaFirst step in the analysis carried out “Pearson Correlation”, in order to remove parameters that are related (p>0.9), therefore do not add information to the analysis. The second step of the analysis was to elucidate those MPs that provide the best discrimination between the groups, using Discriminant Analysis test. This analysis was preformed initially within each genotype between treatments and than, on each treatment between genotypes and for each test, three to four most discriminating MPs were chosen. Following that, an ANOVA was performed on the selected MPs checking for significance of differences, first in each genotype between treatments and than, in each treatment between genotypes. The level of significant was calculated using Bonferoni correction to avoid type I error. In the ANOVA tests, the preliminary assumption was the existence of homogeneity of variance and not normal distribution


A protective effect of calcium against colorectal cancer has been described in Anglo-Saxon but not in Latin communities, and no such effect has been observed regarding adenomas. We investigated the relationship between calcium, dairy products and the adenoma-carcinoma sequence in a French region by comparing small adenoma , large adenoma (n = 208) and polyp-free (n = 426) subjects, and cancer cases (n = 171) with population controls (n = 309). There was no protective effect of calcium against colorectal tumours except for low fat calcium and large adenomas in men . There was even a trend towards an increased risk of cancer with dairy calcium in men and non-dairy calcium in women. Vitamin D was inversely related to the risk of small adenomas in women . Regarding dairy products, only consumption of yoghurt displayed an inverse relationship with risk of large adenomas, in both men and women. These data failed to demonstrate a protective effect of calcium against colorectal carcinogenesis. They suggest that the type of dairy product might be the important factor with regard to prevention of colorectal tumours.


Somatic symptomatology is common in depression, and is often attributed to the Freudian-inspired concept of "somatization". While the same somatic symptoms and depression are common in mitochondrial disease, in cases with concurrent mood symptoms the diagnosis of a mitochondrial disorder and related therapy are typically delayed for many years. A short screening tool that can identify patients with depression at high risk for having underlying mitochondrial dysfunction is presented.Six items of the Karolinska Scales of Personality (KSP) were found to differentiate among 21 chronically-depressed Swedish subjects with low versus normal muscle ATP production rates. A screening tool consisting of the six KSP questions was validated in the relatives of American genetics clinic patients, including in 24 matrilineal relatives in families with maternally inherited mitochondrial disease and in 30 control relatives.-5).Among the depressed Swedish patients, the screening tool was positive in 13/14 with low and 1/7 with normal mitochondrial function (P = 0.0003). Applied to the American relatives of patients, the screening tool was positive in 13/24 matrilineal relatives and in 1/30 control relatives (P = 2 × 10Our preliminary data suggest that a small number of specific somatic-related questions can be constructed into a valid screening tool for cases at high risk for having a component of energy metabolism in their pathogenesis. Multiple medically-unexplained somatic complaints, usually associated with mood symptoms, is currently designated as "somatization" under the Diagnostic and Statistical Manual (DSM) system used to classify mental illness. This phenomenon has been observed by physicians for at least 3000 years and was known as "hysteria" until the previous DSM designation of "Briquet's syndrome". The French physician Briquet, in his 1859 treatise on hysteria, described mood as well as a long list of somatic symptoms among his 430 patients, including migraine, abdominal pain, muscle pain, palpitations, restlessness, hyperesthesias, anesthesias and fatigue [However, in 1895, Freud and Breuer in their classical study of hysteria  actuallyUnfortunately, considering the high prevalence and associated disability of depression with somatization, there has been little advancement in our understanding of its pathogenesis since the time of Freud. As the same somatic and mood symptoms are common in patients with mitochondrial disorders -8, recenThere is substantial overlap between patients identified with somatization and with mitochondrial disorders. In the latter conditions, symptoms typically wax and wane, and involve different tissues and body locations at different times, presumably in accordance with local energy supply and demand. These and other factors often lead the patient's symptoms to be dismissed as being "functional" or "psychosomatic", delaying typically by many years the specific testing that establishes the diagnosis of a mitochondrial disorder in patients with concurrent mood symptoms ,8.in vitro muscle ATP production rates using α-ketoglutarate as substrate in 21 adult Swedish subjects with longstanding major depression [Recently, we reported on pression . Consistpression , almost pression . HoweverInspired by the World Health Organization six-item, 5-point response format, self-report scale for the screening of adult Attention Deficit Hyperactivity Disorder (ADHD) , we haveversus with normal ATP production rates.We re-evaluated our data  from theIn order to validate our screening instrument, English translations  of the sThree of the 15 KSP scales, Somatic Anxiety, Muscular Tension and Psychasthenia, inquire regarding somatic symptomology or reactions to that symptomology ,13, and or the relative had been given a diagnosis of "depression" and treated for such by a medical or mental health professional.Among the American relatives, the presence of "depression" was defined as a probable diagnosis of major depressive disorder based upon meeting the DSM Version-IV-TR criteria for that condition from questionnaire data, In our 21 depressed subjects from Sweden, a score of 4 was found to be statistically more common (P < 0.05) in 10 of the 30 somatization-related KSP items [-5). This corresponds to 54% sensitivity and 97% specificity. Eleven of the 24 (46%) relatives in the mitochondrial group had a lifetime diagnosis of depression, versus 3 of 28 (11%) control relatives (P = 2 × 10-5). There was no correlation between the presence or absence of depression and the KSP screening results.In the American relatives of subjects, five of the six KSP items listed in Table Among our Swedish subjects with lower levels of mitochondrial function, the choice of the extreme "Applies Completely" response may reflect either more intense symptomatology and/or the tendency of a more dramatic account of the state, a "hysterical" disposition. The lower sensitivity of our proposed six-question screening battery in the American versus in the Swedish subjects may be due to a lower degree of symptomatology in the former, predominately reproductive-aged  relatives of patients versus in the our older patients  who themselves suffer from chronic mental illness. Applying the American data in a highly-selected population whereas 67% have some degree of energy depletion , our data corresponds to a 93% positive predictive value and a 72% negative predictive value for the screening battery, with a positive response defined as 2 to 6 questions answered as "Applies Completely". In a hypothetical population of lesser acuity whereas only 10% have some degree of energy depletion, our data corresponds to a 64% positive predictive value and a 95% negative predictive value.In the functional disorder of migraine, which demonstrates strong co-morbidity with depression , co-enzyWe propose that energy depletion constitutes at least part of the inherited biological predisposition towards the development of depression with somatization predicted by Freud. Furthermore, although our present findings require additional validation in varied groups of patients, our preliminary data suggest that a small number of specific somatic-related questions, inquiring for symptoms that likely would be instantly recognizable by Briquet and Freud, can be constructed into a valid screening instrument for cases at high risk for having a component of energy metabolism in their pathogenesis, or "mitosomatic" illness.KSP = Karolinska Scales of Personality; DSM = Diagnostic and Statistical Manual; ATP = Adenosine triphosphate; ADHD = Attention Deficit Hyperactivity Disorder; mtDNA = Mitochondrial DNA; P = Probability.The author(s) declare that they have no competing interests.AG conducted the Swedish study and RB conducted the American study. Both authors contributed to the study design, drafted the manuscript, and read and approved the final version.


The aim of the study was to determine if differences in clinical diagnosis versus autopsy findings concerning the cause of death in polytrauma fatalities would be detected in 19 cases of fatal polytrauma from a Level 1 trauma centre.Clinical diagnoses determining the cause of death in 19 cases of fatal polytrauma (2007 - 2008) from a Level 1 trauma centre were correlated with autopsy findings.In 13 cases (68%), the clinical cause of death and the cause of death as determined by autopsy were congruent. Marginal differences occurred in three (16%) patients while obvious differences in interpreting the cause of death were found in another three (16%) cases. Five fatalities  were remarked as early death (1-4 h after trauma) and one fatality with marginal differences as late death (>1 week after trauma). Obvious and marginal discrepancies mostly occurred in the early phase of treatment, especially when severely injured patients were admitted to the emergency room undergoing continued cardiopulmonary resuscitation, i. e. limiting diagnostic procedures, and thus the clinical cause of death was essentially determined by basic emergency diagnostics.Autopsy as golden standard to define the cause of death in fatal polytrauma varies from the clinical point of view, depending on the patient's pre-existing condition, mechanism of polytrauma, necessity of traumatic cardiopulmonary resuscitation, survival time, and thus the possibility to perform emergency diagnostics. An autopsy should be performed at least in cases of early fatal polytrauma to help establishing the definite cause of death. Moreover, autopsy data should be included in trauma registries as a quality assessment tool. Polytrauma, defined as a trauma pattern with an Injury Severity Score (ISS) >16 points and consisting of various injuries of which at least one is life-threatening, is associated with a mortality rate up to 23% .®-(ATLS)-Program have been set up  or Operation Room [OR]/1-4 h after trauma) and 6 32%) fatalities as late death (> 1 week after trauma). Four (21%) patients survived between 4 h and 1 week, thus not being classifiable in Trunkey's scheme .In summary, 5 fatalities presenting discrepancies at autopsy  were remarked as early death and 1 with marginal differences as late death. Obvious und marginal discrepancies concerning the cause of death mostly occurred in the early phase of treatment (n = 5/83%); 3 of the 6 discrepant fatalities arrived at the ER undergoing continued CPR or necessitated CPR in immediate clinical course.In the following, we present short case reports of the incongruent polytrauma fatalities in our study group.The three fatalities presenting obvious differences at autopsy when compared to clinical diagnoses, survived the traumatic incident between 2:10 h and 2:50 h (median 2:46 h) and reached the ER 1:19 h after accident (median). Median in-hospital survival time was 1:27 h, and 2 of these patients were brought to the ER undergoing continued CPR or necessitated CPR during ER diagnostic procedures.Table aO2 maximum 70 mmHg and PaCO2 of 70 mmHg). The patient was brought rapidly to the OR, ER diagnostics were based on physical examination, ultrasound performance, thoracic and pelvic x-ray scans and whole-body spiral computer tomography, showing sTBI, severe chest injury, pelvic trauma , serial fractures of the processi transversi of the vertebral column and a small retroperitoneal hematoma. During immediate damage control surgery for thoracic trauma, the man had to be resuscitated which remained unsuccessful; total survival time was 2:50 h, in-hospital survival time accounted for 1:37 h.The patient  sustained a high velocity car accident. He arrived intubated at the ER 1:13 h after the accident where sTBI and severe blunt chest trauma with bilateral lung contusions were diagnosed (ISS: 59 points). Chest drains were applied, but artificial ventilation remained difficult  was overrun by a car; 1:19 h later he was admitted to the ER. During immediate ER course, CPR efforts were necessary, and initial ISS was scored 43 points. Severe chest trauma with bilateral serial costal fractures, heart and lung contusions as well as hemorrhagic shock was diagnosed; chest drains were applied. Physical examination, X-ray scans, abdominal ultrasound and spiral computertomography were performed immediately, confirming the chest trauma and showing a hypertrophied right heart without signs of pericardial effusion. After extensive CPR, the man was pronounced dead 1:27 h after admission. Total survival time was 2:46 h; trauma surgeons blamed a hemorrhagic shock for the cardiac arrest.Autopsy confirmed the severe chest trauma with several bilateral rib fractures as well as heart and lung contusions, but furthermore yielded a pericardial tamponade with 280 ml of blood, caused by a 10 mm traumatic rupture of the V. cava inferior at the mouth of the right atrium which was considered forensically as competitive cause of death. It could not be clarified forensically to what extent the rupture of V. cava inferior was due to the original thoracic trauma or to ponderous CPR efforts with a chest drain applied to the right pleura which was probably moved during extensive closed chest CPR: the tip of the chest drain was found to be located near the mouth of the right atrium. Despite the non-diagnosed possible iatrogenic pericardial tamponade, death was considered unpreventable because of the severity of the chest trauma.The patient  was clamped by an 1800-kg-transformator that trapped him in a hoistway. After difficult extrication by the fire brigade he reached the ER 1:45 h after the accident endotracheally intubated and undergoing continued CPR. Initial ISS was scored 75 points, and emergency diagnostics consisted of basic emergency diagnostics  as extensive CPR was continued for another 0:25 h before the patient was pronounced dead. Survival time from the accident was 2:10 h. Based upon previous surgical examination and x-ray scans, ER physicians assumed as cause of death a blunt chest, abdominal and pelvic trauma, but autopsy solely showed - besides signs of extensive CPR - a fatal bleeding from the pelvic ring disruption with traumatic rupture of A. and V. iliaca communis dextra. Organs in thorax and upper abdomen were nearly unaltered. Although merely the pelvic injury led to death by exsanguination, the patient's demise was considered unavoidable as the blood loss already necessitated CPR on scene and the patient arrived at the ER without signs of life for a total time of 1:45 h.In the 3 fatal polytrauma issues presenting marginal differences concerning the cause of death, the patients reached the ER 1:00 h after the accident (median). 2 patients survived in-hospital for 0:12 h and 2:22 h respectively, one patient survived for 184 days. 1 patient was brought to the ER undergoing continued CPR.The patient f, 24 y) was assaulted with a knife and sustained 25 stab wounds to the ventral and dorsal chest wall; she was intubated on scene and brought to the ER 48 min after the attack, necessitating continued CPR already during transport. At admission to the ER, ISS was scored 75 points, and clinical procedures focussed on continuing extensive CPR. Emergency diagnostics were solely performed by means of physical examination, chest drains were not applied. She was pronounced dead 12 minutes after admission to the ER because of at least 45 min therapy-resistant asystolia; survival time from the traumatic incident was 1:00 h. Clinical cause of death was assumed as solely hemorrhagic shock due to multiple chest stab wounds with hypothesized cardiac and aortic injuries while autopsy showed besides the severe sharp thorax trauma  also a bilateral hematopneumothorax contributing to the woman's demise. Heart and aorta were unaltered. Although this was considered as marginal difference and it was clearly stated that death would not have been preventable, autopsy supported the recommendation that traumatic CPR should not be stopped until bilateral chest drains have been applied [ y was asA motorcycle driver  was hit by a truck; 1:12 h after the accident, he arrived endotracheally intubated at the ER where the following diagnoses were collected by physical examination, ultrasound performance, thoracic and pelvic x-ray scans as well as a whole-body CT: hemorrhagic shock, spine fracture Th12, bilateral serial costal fractures, dislocated pelvic fracture, right-sided talocalcanean joint fracture, and urinary bladder rupture (ISS: 50 points). After damage control surgery and hemodynamical stabilization, the patient was transferred to the Intensive Care Unit (ICU). The further clinical course was fraught with severe complications such as rhabdomyolysis, persisting Systemic Inflammatory Response Syndrome (SIRS), decubital ulcers, critical illness polyneuropathy, and respiratory failure. Clinical diagnostics throughout the prolonged ICU treatment period consisted of the whole bandwidth of medical possibilities. Nevertheless, the patient suffered a cardiac arrest on day 184 after admission, presenting a fulminant septic shock; CPR was carried out for 45 minutes. Clinical cause of death was determined as septic MOF with concomitant septic lung artery embolism. Autopsy confirmed besides the initial diagnoses a septic MOF as inevitable cause of death, but showed - as marginal difference - no hints for presence of lung artery embolism.The patient  was overrun by a train; at admission to the ER 1:00 h after trauma sTBI, left-sided proximal humerus fracture, left-sided open hip fracture, and traumatic amputation of the left lower leg were diagnosed (ISS: 75 points). Clinical diagnostics consisted of physical examination, ultrasound performance, thoracic and pelvic x-ray scans as well as a whole-body CT. During emergency neurosurgical intervention the patient had to be resuscitated; these efforts remained unsuccessful. Total survival time from the traumatic incident was 3:22 h, in-hospital survival time was 2:22 h. With respect to timely manners and rapid aggravation of vital parameters, orthopedic surgeons blamed a bleeding from the Sinus sagittalis superior with consecutive hemorrhagic shock as the main cause of death rather than sTBI. The particular injury pattern, especially the traumatic amputation of the left lower leg did not contribute clinically to the patient's assumed hemorrhagic shock as there was no significant bleeding from the wound because of vessel retraction.Autopsy showed - besides the correct diagnoses mentioned above - wide-spread, primarily non-survivable destructions of the entire brain tissue with comminute fractures of the basal skull and the calvarium. The cause of unpreventable death was forensically determined as sTBI, but not hemorrhagic shock, thus being qualified as marginal difference.Severely injured patients present a wide range of complex problems to the trauma leader in charge: diagnosis of life-threatening injuries and their therapy need to be set up rapidly. Depending on the severity of the trauma, and thus on the time available for establishing the diagnosis, the use of diagnostic means may be very limited. Yet, a recent study points out that the in-hospital survival time is inversely proportional to the discrepancy rate at autopsy, suggesting that lack of time for appropriate diagnostic tests have an important role in missed diagnoses . Especia• estimation of manner and cause of death,• accuracy/quality assurance of clinical diagnoses, and• verification/denial of causal relationship between trauma incident and death.Besides that, the cause of the traumatic incident can be clarified by autopsy in some cases, e.g. myocardial infarction, apoplectic insult or other severe pre-existing or acute diseases. Yet current literature has documented a decrease in autopsy frequency; the reasons are diversified, including technological advances in clinical testing and imaging and apprehension of potential medicolegal consequences resulting from discrepant findings. Nevertheless, disparity rates between clinical and post-mortem diagnoses concerning the cause of death are reported to reach up to 17.2% .In case of trauma-related death, diagnostic assessment in polytrauma patients might also be limited at autopsy, notably in patients requiring CPR following thorax trauma: iatrogenic CPR-injuries might be sometimes hard to differentiate from the original trauma incident - cardiac massage must be considered also as a blunt chest trauma . Here inEach diagnostic procedure has limitations depending on accessible data, time available for clinical diagnostics and incident-referred pattern of trauma, and no overall diagnostic forensic and clinical gold standard is to be defined or one diagnostic measure is considered to be superior to other diagnostic measures. When executing a data-based comparison of diagnostic accuracy between clinical and forensic findings, we conclude that final statement concerning the cause of death in early polytrauma death should be conducted in synopsis of all available methods including autopsy data. A single case analysis should be comprehended in each incongruent polytrauma fatality and is necessary for accuracy in determining the cause of death.We conducted a single centre study from a Level I trauma centre with a relatively small patient collective; generalization of our results may be difficult, because the management of severely injured patients in level I trauma centres might differ considerably from nontrauma centre hospitals. Furthermore, the temporal distribution of trauma deaths depends on the time intervals and classifications chosen in the given study. However, we consider our results as reliable, because review of the clinical records and death certificates as well as the final autopsy reports allowed a tight follow-up of each fatal outcome in our study by close personal interaction between surgical and forensic department. Verification of the study hypothesis was supported by the collected data. Although limitations have to be considered, we were able to demonstrate that unexpected findings concerning the cause of death are identified relatively frequently at autopsy in early polytrauma fatalities.Although preventable deaths (in terms of missed and thus non-addressed injuries) were not present, our study confirms that autopsy remains an important quality assessment tool for determining discrepant diagnoses concerning the cause of death in polytrauma fatalities, especially when autopsy is preceeded by a short ER stay and necessity of CPR during transport or in immediate ER course. A forensic autopsy in fatal polytrauma issues should standardly be conducted at least in each case of early/immediate death to help establishing the definite cause of the patient's demise - most likely already by proposal by the clinical physician in charge of the polytrauma patient. Autopsy provides important supplemental diagnosis, and inclusion of autopsy data from such polytrauma fatalities in national trauma registries for further research projects on this topic should be considered. Amplified interlocking between surgical and forensic departments ought to be strived for in the future.There is no conflict of interest. The corresponding author affirms that he has no relationships with a company whose product is mentioned in the article or with one that sells a competitive product. The presentation is impartial, and the content is independent of commercial influence.CTB, PG and JKF conceived of the study, and participated in its design and coordination. MT and WE participated in the design of the study. CTB drafted the manuscript. All authors read and approved the final manuscript.


Dinoflagellates are not only important marine primary producers and grazers, but also the major causative agents of harmful algal blooms. It has been reported that many dinoflagellate species can produce various natural toxins. These toxins can be extremely toxic and many of them are effective at far lower dosages than conventional chemical agents. Consumption of seafood contaminated by algal toxins results in various seafood poisoning syndromes: paralytic shellfish poisoning (PSP), neurotoxic shellfish poisoning (NSP), amnesic shellfish poisoning (ASP), diarrheic shellfish poisoning (DSP), ciguatera fish poisoning (CFP) and azaspiracid shellfish poisoning (ASP). Most of these poisonings are caused by neurotoxins which present themselves with highly specific effects on the nervous system of animals, including humans, by interfering with nerve impulse transmission. Neurotoxins are a varied group of compounds, both chemically and pharmacologically. They vary in both chemical structure and mechanism of action, and produce very distinct biological effects, which provides a potential application of these toxins in pharmacology and toxicology. This review summarizes the origin, structure and clinical symptoms of PSP, NSP, CFP, AZP, yessotoxin and palytoxin produced by marine dinoflagellates, as well as their molecular mechanisms of action on voltage-gated ion channels. Over the past few decades, the occurrence of harmful algal blooms (HABs) has increased both in frequency and in geographic distribution in many regions of the world. This has resulted in adverse impacts on public health and the economy, and has become a global concern –3. It isOf those causative organisms, dinoflagellates, a very large and diverse group of eukaryotic algae in the marine ecosystem, are the major group producing toxins that impact humans , 10. DinIt is known that most dinoflagellate toxins are neurotoxins, which interact with the specific receptors associated with neurotransmitter receptors, or voltage-sensitive ion channels , resultiA. catenella, which resulted in 102 people being ill and six deaths , indicating that brevetoxin analogues display a range of efficacies to neurotoxin site 2 ligands and are activators of neurotoxin site 5, with PbTx-1 being a full agonist and other derivatives acting as partial agonists i by activation of Ca2+ -release from internal stores and Ca2+-influx, while AZA3 induces only Ca2+-influx. AZA5 does not modify intracellular Ca2+ homeostasis. Recent investigation of the effect of AZA4 on cytosolic calcium concentration [Ca2+]i in fresh human lymphocytes demonstrated that AZA4 inhibits store-operated Ca2+ channels (SOC channels) and Ca2+ influx and that this process is reversible i may be activation of phospholipase C [+ conductive pathway which results in activation of the Na+/H+ exchanger [2+]i independently of the activity of voltage dependent Ca2+ channels and Na+/Ca’+ exchange [+ specific and Ca2+ specific channels. Overall, PTX might posses more than one site of action in excitable cells and act as an agonist for low conductance channels conducting Na+/K+, Ca2+ and H+ ions.Over the past few decades much effort has been devoted to define the action mechanisms of PTXs, however these have not been identified. Pharmacological and electrophysiological studies have demonstrated that PTXs act as a haemolysin and alter the function of excitable cells. PTX selectively binds to the Naof 20 pM  and tranof 20 pM –113. Prelipase C  and phoslipase C . There axchanger , 117. Otexchange . The lasThis paper briefly outlines the origin, structure, symptoms and molecular action mechanisms of neurotoxins produced by marine dinoflagellates. These toxins vary in chemical structure and mechanism of action, and produce very distinct biological effects, which provides a potential application of these toxins in pharmacology and toxicology. However, some of them have not been well studied due to the limited supply of pure toxins and their molecular action mechanisms are unknown. Moreover, novel species of neurotoxins produced by dinoflagellates have been found and identified, which provide a challenge for the characterization of their toxin mechanisms and their effects on marine organisms and humans. Further work using the cell-based approach is needed to determine the precise mode of action of these novel neurotoxins from marine dinoflagellates.


Ovarian cancer continues to be a major cause of morbidity and mortality in women. Antiangiogenic treatments have emerged as a promising strategy to treat ovarian cancer. This article reviews the rationale supporting the use of antiangiogenic treatments in ovarian cancer, the clinical development of this group of drugs and the toxicities specific to this modality of treatment. Ovarian cancer is the fourth most common cause of cancer death in women. Worldwide, there are more than 190 000 new cases of ovarian cancer each year, accounting for around 4% of all cancers diagnosed in women. Incidence rates vary considerably, with the highest rates in the United States and Northern Europe and the lowest rates in Africa and Asia.The majority of patients with ovarian cancer present late with advanced disease (FIGO stage III/IV) and in this group of patients, despite multimodality treatment with surgical debulking followed by platinum–taxane combination chemotherapy, median survival is only 3 years.New treatment approaches are therefore urgently required to improve outcome in this disease and one promising strategy to have emerged has been the study of angiogenesis in ovarian cancer and the role of modulators of angiogenesis in its treatment.μm in diameter as diffusion of nutrients and oxygen from nearby capillaries is inadequate beyond this point to sustain cell function  and may act as co-receptors to enhance VEGF signalling through VEGFR-1. Neuropilin expression is increased in ovarian carcinomas  for six cycles followed by 12 cycles of maintenance bevacizumab. GOG-218 is a three-arm placebo-controlled study – all patients receive carboplatin and paclitaxel for six cycles – in the first experimental arm, patients also receive concurrent and maintenance bevacizumab  for up to 16 doses, whereas the second experimental arm receives only concurrent bevacizumab followed by placebo maintenance.Bevacizumab, a humanised monoclonal antibody directed against VEGF, was the first antiangiogenic agent to be licensed in the treatment of cancer and has improved survival when administered in combination with chemotherapy in a broad range of epithelial malignancies , a fusion protein containing the VEGF-binding domains of both VEGFR-1 and -2 linked through the Fc region of human IgG1, is also under investigation as a single agent in relapsed platinum-resistant ovarian cancer with a response rate of 8% and acceptable toxicity being reported at a pooled interim analysis of a phase II study of two different dosing regimens  in 6 of 13 ovarian cancer patients recruited. Toxicity appears higher than that with single-agent anti-VEGF therapy, with two-thirds of the patients developing hypertension and 79% incidence of grade 1–3 hand–foot syndrome , proteinuria , haemorrhage, arterial and venous thrombotic events, impaired wound healing and, of particular concern in the initial phase II bevacizumab trials in ovarian cancer, gastrointestinal perforation.Although the role of VEGF in normal endothelial homoeostasis is not well understood, the rapid regression of capillaries in several different tissues such as pancreatic islets, thyroid, adrenal cortex, choroid plexus and small intestinal villi is seen within a few days of starting treatment with VEGF inhibitors in animal models   did not Reversible posterior leukoencephalopathy syndrome and tracheo-oesophageal fistulae are significant, but rare, complications of bevacizumab treatment. Vascular endothelial growth factor receptor tyrosine kinase inhibitors have also been associated with clinical hypothyroidism, which could be caused by the inhibition of iodine uptake in the thyroid  and metronomic oral cyclophosphamide (50 mg daily) in platinum-resistant/partially platinum-sensitive ovarian cancer with promising activity. Median time to progression was 7.2 months and median survival time was 16.9 months in this study involving 70 patients  target proliferating endothelial cells leading to their apoptosis with few side effects and no significant direct tumour cytotoxicity  is overexpressed in ovarian carcinoma, and EGFR signalling has been shown to upregulate VEGF expression. Although erlotinib (EGFR tyrosine kinase inhibitor) does not have any relevant single-agent activity in ovarian cancer, combined VEGF and EGFR inhibition is currently under investigation. The combination of erlotinib and bevacizumab has already been explored in a phase II study  signalling pathway is another potential target for antiangiogenic therapy, as PDGFR signalling is of key importance in maintaining the homoeostasis of pericytes, mesenchymally derived cells that surround capillaries and venules, and are vital for microvessel stability and function. Platelet-derived growth factor and PDGFR have also been identified in metastatic ovarian carcinoma, with overexpression of PDGFR associated with poor prognosis  and Notch-1. Delta-like ligand-4 is normally induced by VEGF as a negative-feedback regulator of vascular growth and is strongly expressed in tumour vasculature. Delta-like ligand-4 blockade in animal xenograft models interestingly results in increased tumour vasculature but reduced tumour growth . Temsirolimus is an inhibitor of mTOR and is currently under investigation in a phase II study in recurrent ovarian cancer (GOG-0170I).Vascular endothelial growth factor gene expression is influenced by several factors although hypoxia is vitally important. This leads to the activation of hypoxia-inducible factor-1β that has shown activity in phase I studies with minimal toxicity and is being studied further in ovarian cancer  family of serine/threonine kinases. Enzastaurin (LY317615) is an inhibitor of PKC-α5β1 integrin, which is part of a family of transmembrane proteins that play a critical role in vasculogenesis (α5 integrins) (Volociximab (M200) is a monoclonal antibody that specifically binds The pre-clinical and clinical studies show promise for the antiangiogenic approach in ovarian cancer treatment. Many unanswered questions, however, remain as to the best strategy to use with these treatments. For instance, is combination with chemotherapy the best approach and, if so, is concurrent antiangiogenic therapy sufficient or should maintenance treatment be pursued? Hopefully, answers to these questions will emerge for bevacizumab from the ongoing phase III studies (ICON7 and GOG-218). Given the plethora of antiangiogenic targets and drugs under development, future research will also need to concentrate on the best approaches to combine/sequence these agents.Currently, one of the biggest challenges in the clinical development of antiangiogenic agents is developing robust biomarkers for clinical use that will allow us to both ascertain the optimum biological dose of these agents in early clinical trials and also to select those patients who are most likely to benefit from antiangiogenic treatment. This is particularly important given the costs and not the insignificant toxicity profiles of these agents.in vivo (Dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) has been the most utilised pharmacodynamic imaging modality in early phase clinical trials of angiogenic inhibitors. This functional imaging technique is non-invasive and can be used to serially assess tumour vasculature 18F]fluorothymidine (FLT) PET imaging may be an early metabolic predictor of response to antiangiogenic therapy. In a phase II study of bevacizumab and irinotecan in recurrent glioblastoma, patients whose tumours demonstrated a metabolic response lived three times longer than non-responders  seems to be emerging from trials of VEGFR tyrosine kinase inhibitors  and endothelial progenitor cell (CEP) concentrations have also been shown to be promising surrogate markers in pre-clinical studies (Anti-VEGF drugs, in particular bevacizumab, have shown promising activity as single agents in chemotherapy-resistant ovarian cancer, supporting the strong pre-clinical rationale for the utility of this treatment strategy. Combinations with chemotherapy are currently being tested in clinical trials in the first-line setting and it is hoped that this approach may significantly improve patient outcome. Biomarkers of response, however, need to be studied to enable a better selection of patients who will benefit from treatment and to ascertain optimal dosing.As more targets for antiangiogenic therapies emerge from pre-clinical studies, the interactions between these different pro-angiogenic pathways, the concept of combination antiangiogenic treatments and the evaluation of mechanisms of ‘resistance’ to angiogenesis inhibitors will need to be addressed in the future.


Genetically engineered mice have been essential tools for elucidating the pathological mechanisms underlying human diseases. In the case of diseases caused by impaired desmosome function, mouse models have helped to establish causal links between mutations and disease phenotypes. This review focuses on mice that lack the desmosomal cadherins desmoglein 3 or desmocollin 3 in stratified epithelia. A comparison of the phenotypes observed in these mouse lines is provided and the relationship between the mutant mouse phenotypes and human diseases, in particular pemphigus vulgaris, is discussed. Furthermore, we will discuss the advantages and potential limitations of genetically engineered mouse lines in our ongoing quest to understand blistering skin diseases. In recent years, mouse models have become an essential tool for studying genetic diseases, especially in cases where the disease is caused by mutations in a single gene (monogenic disorders). It has been a challenge, however, to faithfully reproduce the pathophysiology of autoimmune disorders in mice, in part because the human disease is influenced by complex factors such as the genetic background, the nature of the autoantibodies (epitopes recognized and antibody subclass) and titers of the circulating autoantibodies. Pemphigus is a class of autoimmune blistering skin diseases which manifest itself in the form of painful lesions in the skin and oral mucosa and which, in the case of pemphigus vulgaris, can be lethal if left untreated. The sera of patients with pemphigus contain autoantibodies directed against desmosomal cadherins  , 2, a grHow does the binding of pathogenic pemphigus autoantibodies induce intraepithelial blistering, the characteristic histopathological feature of the disease? Do these antibodies inhibit the function of these proteins? A simple approach to test this hypothesis is to eliminate the target protein in genetically engineered mice and to determine whether the resulting loss-of-function phenotype replicates the disease. Pemphigus vulgaris (PV) patients develop autoantibodies that target desmoglein (DSG) 3 and under certain conditions desmoglein (DSG) 1 ).Dsc3 gene was embryonic lethal [Dsc3 null mutant mouse lines were generated that lacked DSC3 expression in basal keratinocytes of stratified epithelia, including skin and mucous membranes [Dsc3 null mice developed skin blisters , Dsc3 null skin blisters developed spontaneously and were present in all skin samples of newborn mice that were analyzed. Nevertheless, the extent of blister formation varied between individual mice, most likely due to different degrees of mechanical stress to which the animals were exposed prior to tissue harvesting. Even small trauma resulted in extensive skin blistering in Dsc3 null skin, a typical characteristic of PV in humans (Nikolsky's sign). Since germline deletion of the c lethal , conditiembranes . These t Figures  that werDsg3-null mutants . Given s tongue –13. The The data summarized above suggest that DSG3 and DSC3 have comparable functions in desmosomes. However, they have different roles in cell adhesion of specific tissues, due to the presence of different compensatory proteins in different tissues. Thus, whereas DSG3 appears to be essential for maintaining cell-cell adhesion of internal stratified epithelia, DSC3 plays a similar role in the epidermis of the skin.Dsg3 null and Dsc3 null mice  to telogen (resting phase of the hair growth cycle). Further, the wave of hair follicle cycling proceeds from head to tail of the mouse. In rare instances, hair loss has been reported in PF and PV patients , 14, sugDsg3 null mice do not develop an autoimmune disease; that is, they do not provide us with a tool to dissect autoantibody-mediated pathology. Nevertheless, these mice enabled us for the first time to link the loss of DSG3 function to PV histopathology. In other words, we were able to functionally identify DSG3 as a likely key target in a subset of pemphigus patients. Subsequent to the development of the Dsg3-null mouse model, Amagai and colleagues utilized this mouse line to develop an autoimmune model for PV in mice [The  in mice , demonst in mice , 17, see in mice , 18. Int in mice . AlthougDsc3 null mutation in stratified epithelia of mice suggest that loss of function of this protein, either due to mutations or due to autoantibodies, would result in PV-like lesions. Recent reports suggest that this might indeed be the case. Bolling and colleagues reported the case of a patient with PV-like symptoms who had developed DSC3- but not DSG3-autoantibodies [DSC3 mutation in a family in Afghanistan that was associated with hair loss and recurrent skin blistering [Dsc3 mutant mice. Nevertheless, this report did not provide histological data demonstrating intraepidermal blistering; that is, further evidence is required to convincingly link impaired DSC3 function to intraepidermal blistering in humans.Although it has been well established that DSG3 plays a key role in the development of PV, less is known about a potential role of desmocollins in autoimmune diseases. DSC1 has been linked to IgA pemphigus  are highly conserved between humans and mice. Although the ease of genetic manipulations and the resemblance to human physiology make mice an attractive choice to study skin disorders, there are potential pitfalls, such as the possibility of species-specific differences in the histology of the skin. Further, it is also not possible to exclude the existence of species-specific gene functions, requiring confirmation of experimental results in a human test system. Nevertheless, the mouse will remain a valuable tool for the analysis of human diseases and for the advancement in our understanding of basic epithelial cell biology.


Autism comprises a spectrum of behavioral and cognitive disturbances of childhood development and is known to be highly heritable. Although numerous approaches have been used to identify genes implicated in the development of autism, less than 10% of autism cases have been attributed to single gene disorders.We describe the use of high-resolution genome-wide tilepath microarrays and comparative genomic hybridization to identify copy number variants within 119 probands from multiplex autism families. We next carried out DNA methylation analysis by bisulfite sequencing in a proband and his family, expanding this analysis to methylation analysis of peripheral blood and temporal cortex DNA of autism cases and matched controls from independent datasets. We also assessed oxytocin receptor (OXTR) gene expression within the temporal cortex tissue by quantitative real-time polymerase chain reaction (PCR).OXTR (MIM accession no.: 167055), previously implicated in autism, was present in an autism proband and his mother who exhibits symptoms of obsessive-compulsive disorder. The proband's affected sibling did not harbor this deletion but instead may exhibit epigenetic misregulation of this gene through aberrant gene silencing by DNA methylation. Further DNA methylation analysis of the CpG island known to regulate OXTR expression identified several CpG dinucleotides that show independent statistically significant increases in the DNA methylation status in the peripheral blood cells and temporal cortex in independent datasets of individuals with autism as compared to control samples. Associated with the increase in methylation of these CpG dinucleotides is our finding that OXTR mRNA showed decreased expression in the temporal cortex tissue of autism cases matched for age and sex compared to controls.Our analysis revealed a genomic deletion containing the oxytocin receptor gene, OXTR in the development of the disorder.Together, these data provide further evidence for the role of OXTR and the oxytocin signaling pathway in the etiology of autism and, for the first time, implicate the epigenetic regulation of See the related commentary by Gurrieri and Neri:  The core autism phenotype includes deficits in social interaction, language development and patterns of repetitive behaviors and/or restricted interests. The population prevalence of the spectrum of autism disorders is estimated to range between 1/300 [o of 4:1 ,3. The do of 4:1  and Retto of 4:1 . Numerouo of 4:1 ,8. Howevin situ hybridization (FISH) [An additional approach to identify genes involved in autism is to characterize copy number variants (CNVs), that is, chromosomal deletions and duplications, that are known to be present within at least 5% of individuals with idiopathic autism . Autism n (FISH) , Southern (FISH) , loss ofn (FISH) -17,19, qn (FISH)  and, morn (FISH) .. The most significant finding thus far from our analysis is a heterozygous deletion of the oxytocin receptor gene (OXTR) (MIM accession no.: 167055) in an individual with autism and his mother with putative obsessive-compulsive disorder (OCD). We further investigated the relationship between OXTR and autism by carrying out epigenetic analysis of the promoter region of OXTR. We show that the gene is hypermethylated in independent cohorts with autism as compared to controls, in both peripheral blood mononuclear cells (PBMCs) and the temporal cortex. Additionally, our analysis of expression levels of OXTR in the temporal cortex shows decreased levels of expression in individuals with autism as compared to controls matched for age and sex.Here we describe the use of genome-wide tilepath microarrays and array comparative genomic hybridization (CGH) to identify CNVs in a dataset of 119 unrelated probands from multiplex autism families . The genOXTR in this complex disease.These data suggest that OXTR and the oxytocin signaling pathway play an important role in the etiology of autism and autism spectrum disorders and implicate epigenetic misregulation of Research diagnostic classification entailed the collection of detailed family, developmental, and medical history and systematic screening of each child with autism before proceeding with clinical assessments. The clinical diagnosis of autism was confirmed by standardized diagnostic assessments once a family had completed the screening process and was considered eligible. For research classification, a clinical diagnosis that conforms to Diagnostic and Statistical Manual of Mental Disorders, fourth edition (DSM-IV) criteria was made based on all available medical, clinical, and behavioral data. The clinical diagnosis was established as a research diagnosis on the basis of scores that exceed established cut-offs for autism on the Autism Diagnostic Interview-Revised (ADI-R) ,30. In aOXTR deletion, and phenotypically normal controls. Genomic DNA was extracted from whole blood using the Pure Gene method and standard protocols, as described previously [All aspects of the research study were approved by the Institutional Review Boards of the participating centers. DNA for copy number and methylation analyses was obtained from the peripheral blood of autism probands from multiplex families, the extended family of the proband with the eviously . DNA forMET and PRKCB1 genes [ and are summarized in Table Postmortem studies were performed using frozen brain tissue samples dissected from the superior temporal gyrus BA 41/42) of eight patient-control pairs, obtained through the Autism Tissue Program from the Maryland National Institute of Child Health and Human Development (NICHD) Brain Tissue Center and the Harvard Brain Tissue Resource Center  was used for data processing of the scanned images prior to porting into Nexus (BioDiscovery ) for analysis.High-resolution genome tilepath microarrays were constructed using large bacterial clones  clones) as previously described . Briefly2 ratios and combining BAC replicates. BioDiscovery's rank segmentation algorithm (RSA), which is similar to circular binary segmentation (CBS) [2 thresholds of ± 1.4 and 0.38 for two and one copy gain/loss, respectively. We applied a conservative cut-off of three BAC clones showing the same copy number change trend to define genomic deletion or duplication with the significance threshold set at 0.005. Array CGH dye-swap experiments were carried out on 21 samples to confirm 21 genomic changes, including novel and known CNVs within the database of genomic variants.Microarray data was preprocessed in Nexus by removing poor quality flagged spots (confidence <0.3 as defined by Bluefuse software), removing background by Lowess correction before normalization of logon (CBS) , was useOXTR: forward 5'-GCTGAACATCCCGAGGAACTG-3' and reverse 5'-GCAAATGAGCGGGAATCCTCTA-3'. DNA samples from autistic probands and their family members were used to determine if the CNV was familial or de novo. DNA isolated from tumor samples with known deletions at the regions containing OXTR were used as 1 × copy number controls and an unaffected individual identified in our normal-normal CGH experiment served as our 2 × copy number control at these loci. A primer and probe set designed to the single copy gene that encodes the RNA moiety for the RNase P enzyme, RPPH1 (MIM accession no.: 608513), a region of stable copy number (2 ×), was used to normalize each sample for equal input. Copy number was assessed in quadruplicate for each sample and the 2-ΔΔCT method was used for analysis [DNA copy number was validated using the Applied Biosystems 7900 HT Fast Real-Time PCR System . Primers and probes were designed to a region spanning exon 2 in analysis .OXTR to determine maternal or paternal contributions. Five markers on chromosome 3  were fluorescently labeled using PCR . Primers were designed to bisulfite-converted regions of CpG islands in the promoter region of the ividuals  , according to a standard ΔΔCt Syber Green protocol [PPIA (MIM accession no.: 123840) cDNA was measured in parallel and used as a standard for normalization. Due to relatively low neocortical OXTR gene expression levels compared to PPIA, PCR reactions were performed using 1:5 and 1:25 cDNA dilutions, respectively. OXTR primers were targeted to exon 2, which is shared by all four alternative mRNA transcripts, and designed as follows: F2, 5'-CTGAACATCCCGAGGAACTG-3', and R2, 5'-CTCTGAGCCACTGCAAATGA-3'. To ensure specificity, PCR products were initially sequenced using a CEQ8000 DNA sequencer , and afterwards checked by running a melting curve at the end of each experiment. Analysis was performed using the 2-ΔΔCT method [Total RNA was extracted from four case-control pairs matched for age, sex and PMI  in a 0.6 Mb deletion on chromosome 8q22.2, has been previously implicated in autism spectrum disorders (ASDs) via an observation in Rett syndrome [OXTR deletion described below, by microsatellite and quantitative real-time PCR analysis. Genomic rearrangements of note included a 12.5 Mb deletion within 2q24.1-2q24.3, a region previously linked to autism [LMCD1 (MIM accession no.: 604859); C3orf32; Caveolin 3, CAV3 (MIM accession no.: 601253); oxytocin receptor, OXTR (MIM accession no.: 167055); and RAD18 (MIM accession no.: 605256)  were generated by sex-mismatch array CGH experiments using controls from our normal-normal analysis. A total of 113 loci within 111 autistic individuals contained at least 1 genomic deletion or duplication, with a range of 1-6 CNVs per individual . We choo autism ,46 and co autism -49 and go autism ; and a 0ffspring .OXTR and the previous data implicating OXTR in the development of autism [OXTR was not a de novo event  of all four family members . Kusui et al. had shown that the methylation status of a specific 405 base-pair fragment of this CpG island  is important for tissue-specific expression of OXTR . This island has previously been experimentally defined as an EcoRI-KpnI fragment that was less methylated in myometrial DNA than in the DNA from peripheral blood leukocytes [OXTR differed in methylation level between family members. Specifically, CpG dinucleotides at positions -959, -934, -924, -901 and -860 (relative to translation start site) showed the most dramatic differences in methylation status, with the proband's affected sibling being heavily methylated at sites -934, -924 and -901. These specific residues fall within the critical region (MT2) in the CpG island of OXTR shown by Kusui et al. to regulate the expression of OXTR in liver [OXTR deletion. The 100% methylation of a specific CpG could indicate maintenance of an aberrant methylation mark, making this a more dramatic change.Based on the CNV data within the ometrium ,57. We cs Figure . The firukocytes . All CpGin liver . ImportaOXTR deletion family, we hypothesized that DNA methylation of residues found within the critical region of OXTR regulation may extend beyond this family and represent a more general epigenetic contribution to autism spectrum disorders. We extended our BSS analysis of the 5 differentiallly methylated CpG dinucleotides to PBMCs from 20 phenotypically normal control individuals  and 20 individuals with autism . BSS data showed significantly increased methylation in the PBMCs of CpG -860 (P = 0.0009), -934 (P = 0.0081) and -959 (P = 0.0026) in individuals with autism compared to controls , CpG -901 (P = 0.0149), CpG -924 (P = 0.0448) and CpG -934 (P = 0.0233) between autism cortex tissue compared to the normal cortex controls  that has previously been implicated in autism spectrum disorders [OXTR) that is contained within the deletion to the etiology of autism [ has had copy number data deposited from a syndromic patient who has developmental delay and cognitive impairment presumably emanating from a novel 1.1 Mb deletion that contains OXTR, amongst other genes. Detailed DNA methylation analysis of the promoter region of OXTR, prompted by the observation that the proband's autistic brother did not carry the novel deletion, identified a single CpG dinucleotide within a known OXTR control region [in utero during the first few weeks of pregnancy that lead to shifts in methylation status of OXTR. Similar influences have been seen in IGF2 methylation based on in utero exposures such as smoking and folic acid intake in humans [For several years the characterization of CNVs in autism patients has identified genes that potentially contribute to the etiology of the disorder ,58-60. Iisorders . CNVs obisorders ,46, two isorders -23,47-49isorders ,62, assof autism -53. Morel region  that, inn humans . Since eOXTR expression was supported by analysis of the corresponding mRNA from the temporal cortex of autism cases matched for age and sex and controls. As one would hypothesize given their increased methylation of OXTR, the autism cases also showed statistically lower transcript levels of OXTR expression compared to the normal controls. Because deletions encompassing OXTR have not been observed in other studies characterizing structural variation in autism [OXTR in autism, resulting in a decrease in OXTR expression, may more commonly contribute to the disorder. While the mechanism for the development of autism from a germ line deletion of OXTR versus epigenetic silencing of the gene in the temporal cortex may clearly be different, the effect on oxytocin signaling could be the same. Together, these data provide strong evidence for the role of OXTR and oxytocin signaling pathway in the etiology of autism and, for the first time, show that the epigenetic regulation of OXTR may be an important factor in the development of the disorder.The impact of this epigenetic mark on n autism ,64,65 suMECP2, (MIM accession no.: 300005)) is the causative agent of Rett syndrome [et al have shown that 79% of autism cases have a decrease in MECP2 expression in the frontal cortex and that an increase in aberrant DNA methylation correlates with this decrease in MECP2 expression [OXTR identified in this study.Epigenetic control of autism susceptibility is a recent concept and most certainly a topic of great interest in the field. Over the past decade, researchers have uncovered suggestive links between epigenetics and autism, for example, autism is associated with duplications of 15q11-13 , an imprinted region in the genome where DNA methylation status has been linked to Prader-Willi syndrome (PWS) and Angelman syndrome (AS) ; mutatiosyndrome ; and mutsyndrome . Nagarajpression . These dOXTR, is spatially and temporally regulated by a complex combination of sex hormones, inflammatory cytokines, oxytocin (OXT) feedback and epigenetic mechanisms [Oxt [Oxtr [Oxtr exhibited social impairment by failure to recognize individuals of their own species. It has been suggested that the normal social behavior of the female knockout mice may be attributed to the compensatory function of vasopressin or developmental compensation [OXTR deletion in the mother who does not present with autism. Sex-specific effects of OXTR function from animal models, together with statistical data that suggest dominantly acting de novo mutations may have reduced penetrance in females [OXTR in the mother from our study does not result in autism while the male proband displays the disorder.The oxytocin receptor, chanisms ,57. It ichanisms , includichanisms  who showchanisms -73. Subssms [Oxt  and Oxtrxt [Oxtr  knockoutxt [Oxtr  showed tensation ,77. In aensation  that des females , may expOXTR.We identified a single CpG dinucleotide within a predicted CpG island that showed statistically significant hypermethylation in PBMCs and the temporal cortex of individuals with autism compared to controls. Although the dominant paradigm for regulation of transcription by DNA methylation suggests that clusters of CpGs are necessary for silencing, recent evidence has indicated that methylation of single CpG dinucleotides may also lead to transcriptional changes via interference of transcription factor binding or recruitment of silencing factors . This isOXTR deletion, the data we collected from the affected sibling shows hypermethylation of the CpG dinucleotide (-934) in the predicted CpG island upstream of OXTR. Given our results suggesting that DNA methylation of OXTR in PBMCs may be a marker of methylation status in the temporal cortex, it follows that the affected sibling may also exhibit a decrease in OXTR expression due to methylation instead of deletion; thus implicating two separate mechanisms of transcriptional regulation with the same outcome. These results suggest that because the methylation status of site -934 is significantly different in the temporal cortex and PBMCs between autism cases and controls, measuring the methylation status of OXTR from the blood for autism cases could act as a surrogate for methylation status in the temporal cortex and be diagnostic of affection status, together with traditional diagnostic criteria.In this study, we focused on the temporal cortex (Brodmann areas 41/42 and 22) because studies have shown that localized temporal dysfunction is linked with clinical symptoms in autism, such as perceptive, emotional and cognitive deficits -35. AlthOXTR may contribute to the development of autism. We acknowledge that screening for these genomic changes in a large, well controlled autism sample is necessary to determine the prevalence of these changes and their relationship to the disease. However, regardless of the sample size, we provide strong evidence that the epigenetic regulation of OXTR may be an important factor in the development of the disorder and suggest that further studies begin to focus on this regulatory event and its relationship to disease.In summary, the data presented suggest that genomic deletion or aberrant methylation of The authors declare that they have no competing interests.SGG conceived the experimental design and wrote the paper; JJC assisted with experimental design, carried out experiments and wrote the paper; AT, JJ, DB, DB, CM and CL carried out the experiments; RKA, HHW, GW ascertained autism families for the paper; PE and CFL assisted with experimental design, SKM assisted with the writing of the paper; MLC helped ascertain autism families and assisted with clinical assessment of the study population; AP and MAP-V assisted with experimental design and in the writing of the paper.The pre-publication history for this paper can be accessed here:Cloned bisulfite sequencing results for each peripheral blood mononuclear cell (PBMC) sample.Click here for filePrimer pairs used for this study.Click here for fileGenomic duplication and deletion events within 54 control and 119 autistic individuals identified by whole genome tilepath array comparative genomic hybridization (CGH).Click here for fileOXTR promoter CpG methylation status in peripheral blood mononuclear cells (PBMCs) of 10 autistic females and 10 age-matched controls.Click here for fileOXTR promoter CpG methylation status in the cortex of eight autism cases and controls matched for age and sex.Click here for file


Gestational diabetes mellitus (GDM) is defined as the glucose intolerance that is not present or recognized prior to pregnancy. Several risk factors of GDM depend on environmental factors that are thought to regulate the genome through epigenetic mechanisms. Thus, epigenetic regulation could be involved in the development of GDM. In addition, the adverse intrauterine environment in patients with GDM could also have a negative impact on the establishment of the epigenomes of the offspring. Late pregnancy is characterized by moderate peripheral insulin resistance and hyperinsulinemia, both of which are necessary to ensure an appropriate supply of nutrient to the fetus. It is counteracted by the adaptation of the islets of Langerhans to the higher insulin demand. This adaptation is characterized by an increase of insulin biosynthesis, enhanced glucose-stimulated insulin secretion, and an increase of b-cell mass. It is not completely understood why, in some individuals, the b-cell mass and function fail to adapt to the metabolic demands of pregnancy and why blood glucose concentration rises to pathological levels , 2. GestThe incidence of GDM differs among ethnic populations, with higher rates in African American, Hispanic, American Indian, and Asian women than in white women; values range from 1.4% to 14% but overall the condition commonly affects between 2% and 5% of pregnant women [The frequency of GDM varies in direct proportion to the prevalence of type II diabetes in populations, and women who develop GDM during pregnancy have a higher risk of developing type 2 diabetes (T2D) later in their lives. These observations are important for relating the two pathological situations and they probably arise from common physiopathological mechanisms . There are two different methods for classifying diabetes in pregnancy. The most recent is the American Diabetes Association (ADA) classification  but the Several risk factors are associated with the development of GDM. The most common is obesity (body mass index over 30) diagnosed before pregnancy . Being aThere is enough evidence to assert that T2D has a strong genetic component. The concordance of T2D in monozygotic twins is approximately 70%, compared with 20%–30% in dizygotic twins , 14.Family studies have prompted the emergence of new perspectives regarding the identification of GDM susceptibility genes. Familial clustering of GDM and T2D have been studied, linking T2D and impaired glucose tolerance in families with GDM  and therThe latest genome-wide studies have identified a number of genetic variants that can explain some of the interindividual discrepancies in diabetes susceptibility , 18 and Some strands of scientific opinion suggest a possible role for epigenetic factors in the complex interplay between genes and the environment that are related to insulin resistance, T2D, and GDM mellitus . These pThe pathogenesis of the most common type of GDM have been described similar to that of type 2 diabetes (T2D), with both pancreatic b-cell dysfunction and chronic insulin resistance playing crucial roles . Pregnanα [Moreover, women with GDM after delivery often have an increased risk for metabolic syndrome, and they have been shown to express early markers of vascular dysfunction such as increased intimamedia thickness of carotid arteries . Thus, pα , leptin α , human pα .There are outstanding medical complications that have been related to GDM for mother and also offspring. For mothers, GDM development have been related to overweight and obesity prior pregnancy  and alsoCongenital anomalies are not more common in patients with GDM but there is an increased incidence of stillbirth when glucose control is poor . OrganogThis situation is also important because periconceptual hyperglycemia is known to be teratogenic and undiagnosed T2D might have a role in pregnancy outcome , 43. ConRecent literature defines macrosomia as identical to large for gestational age (LGA), which is size >90th percentile for gestational age  but thisMoreover, postpartum neonatal hypoglycemia may occur when neonates are no longer exposed to high levels of maternal glucose. At delivery, neonates continue producing elevated rates of insulin, but without high levels of maternal glucose that leads to a virtual neonatal hyperinsulinemia and subsequent hypoglycemias, requiring glucose infusion after delivery . Finally, other important complications have been described in offspring of diabetic gestations with repercussions in future. Children who have been diagnosed LGA at birth and were exposed to an intrauterine environment of either diabetes or maternal obesity developed an increased risk of metabolic syndrome (MS) during childhood. This increased rate of obesity prevalence among children and adults may have implications for perpetuating the cycle of obesity, insulin resistance, and their consequences  in subsequent generations –54.Epigenetics is currently known as a science that studies changes in gene activity that take place without a change in the nucleotide sequence. Epigenetic modifications are transmitted from one cell generation to the next (mitotic inheritance) and can also be transmitted down organismal generations (meiotic inheritance) . EpigeneEpigenetic modifications include DNA methylation, histone modification, and those of the microRNA machinery. These mechanisms explain how cells with identical genomic information can differentiate into distinct cell types with different phenotypes . Cytosinhistone modifying enzymes can modify their NH2-terminal tails by acetylation, methylation, phosphorylation, sumoylation, or ubiquitination [The enzymes that lead to DNA methylation are methyltransferases (DNMTs), of which there are types: DNMT1, which copies the pattern of DNA methylation during replication (methylation maintenance), and DNMT3a and DNMT3b, which are responsible for de novo DNA methylation . In conttination . These mtination . Althougtination . Native lysine residues on histone tails bear a positive charge that can bind negatively charged DNA to form a condensed structure with low transcriptional activity. An early suggestion was that histone acetylation removes these positive charges, thereby relaxing chromatin structure and facilitating access to the DNA for the transcriptional machinery to initiate transcription . HoweverAs we previously commented, pregnancy is a pathological condition that may indicate a predisposition to develop T2D in near future. Moreover, several studies point towards GDM and T2D as different faces of the same disease at diverse stages of life . High maIn T2D, b-cell damage usually arises as a result of a combination of genetic susceptibility and acquired damage. Numerous genes have been associated with islet cell dysfunction in T2D, including some that encode for transcription factors, glucose metabolism proteins, and molecules of the insulin signaling pathways . Pdx-1 is a pancreatic homeobox transcription factor that regulates pancreas development and b-cell differentiation. A reduction in Pdx-1 expression in humans and animal models has been shown to cause T2D, while mutations in this gene also cause the monogenic form of diabetes (MODY 4) . Also, PGlucose is an important physiological fuel for the b-cell, initializing ATP production, insulin secretion, and proliferation. However, chronic hyperglycemia has severe adverse effects on b-cell function, including glucose desensitization, b-cell exhaustion and finally a lack of insulin secretion and insulin storage .Increased oxidative stress found in obese and T2D patients, lead to decreased insulin gene transcription by decreasing Pdx1 . The mecGlucose transporters are involved in the transport of glucose in most cells. GLUT4 is a member of the facilitative glucose transporter (GLUT) family, characterized by preferential expression in muscle and adipose tissues, where it is responsible for insulin-stimulated glucose uptake. GLUT4 is essential for the maintenance of normal glucose homeostasis . In patients with T2D, insulin resistance is a product of decreased insulin-stimulated skeletal muscle glycogen synthesis, which can mostly be attributed to decreased insulin-stimulated glucose transport (Glut 4) activity  and simiPPARGC1A expression is correlated with impaired glucose-stimulated insulin secretion, which is known to require ATP, and suggests that epigenetic mechanisms may regulate insulin secretion in human islets.Insulin secretion in pancreatic islets is dependent upon mitochondrial function and production of ATP. The transcriptional coactivator peroxisome proliferator-activated receptor gamma coactivator-1 alpha  is a master regulator of mitochondrial genes, and its expression is weak and related to impaired oxidative phosphorylation in the muscle of patients with T2D . DNA metPPARGC1A is a coactivator of PPARG and PPARA and regulates genes involved in energy metabolism . An alteThe epigenome is most vulnerable to dysregulation during those phases of life associated with higher rates of change for tissue development and growth, like embryogenesis, fetal, and neonatal life and puberty , 90. Classical theories of disease development assumed that genetic susceptibility and unhealthy adult lifestyle give rise to insulin resistance and T2D, but observational studies showed a link between intrauterine growth retardation and subsequent development of T2D . More reIn the case of GDM, glucose travels freely from the mother to the fetus, but maternal insulin does not. Thus, maternal GDM exposes the fetus to higher concentrations of glucose than normal, forcing the fetus to increase its own insulin production . GDM hasAn adverse intrauterine milieu affects the development of the fetus by modifying expression in both pluripotential cells and terminally differentiated, poorly replicated cells and is associated with epigenetically induced downregulation of key genes controlling b-cell development, differentiation, and function –99.Gestational diabetes is a growing health concern, especially in certain predisposed populations. Although traditionally deemed not as dangerous for the developing fetus as pregestational diabetes, recent findings indicate that GDM may have serious long-term consequences for both child and mother. A conceptual developmental disease framework for GDM emerges from the findings reviewed here. Epigenetic mechanisms may control insulin resistance in mothers during pregnancy diagnosed with GDM but could also predispose offspring to developing T2D in later stages of life. An increased need for insulin by the fetus to deal with the high levels of glucose caused by GDM is an environmental circumstance that probably triggers epigenetic changes in that early stage of life, involving genes critical to pancreatic development and b-cell function, peripheral glucose uptake and insulin resistance. Understanding the role of developmental programming genes is crucial to our understanding of GDM and its consequences for the mother and child and it might stimulate the development of further epigenetic therapeutic agents as modern tools for treating this disease.


Mycobacterium leprae that affects the peripheral nerves, skin, and potentially other organs Leprosy is a chronic infection caused by M. leprae mounted by the host will determine the different potential clinical states. The Ridley-Joplin system uses clinical and histopathological features and the bacteriologic index and includes the polar categories (lepromatous [LL] and tuberculoid [TT]) and the borderline states  The immunological response to A 31-year-old Brazilian male living in the United States for the previous four years presented with progressive crops of new nontender nodules on all four extremities over a 16-month period  and 2. HWe recommended that he continue his MDT until his BI  decreased below 2 (dead bacteria may be present in the skin up to 10 years). Despite the WHO recommendation of 12 months of MDT for multibacillary cases such as our patient, many experts still advocate 24 months of MDT for patients with a BI >4 at diagnosis Six months after the initiation of MDT, patient A experienced the abrupt onset of multiple new erythematous and ulcerated skin nodules (1–2 cm) distributed in the upper limbs, trunk, and right leg . The oriBased on the clinical history and signs, our patient was diagnosed with a type 2 leprosy reaction Reactional states represent a change in the immunological state of a patient with leprosy. These reactions are characterized by inflammatory episodes of the skin, nerves, and other organs that can occur anytime within the course of leprosy, but are often observed after starting treatment, and can result in irreversible nerve damage M. lepraeM. leprae as compared to lepromatous states Type 1 reactions are considered to be an upgrading in the cell-mediated immune response to Type 2 reactions are characterized by an inflammatory infiltrate of neutrophils with associated vasculitis and/or panniculitis Type 1 reactions affect patients within the borderline spectrum , while type 2 reactions occur among patients with BL and LL states In the MDT era, reactions occur in approximately 15%–39% of patients with leprosy The diagnosis of leprosy reactions is based primarily on clinical history and examination Type 2 reactions often follow a protracted clinical course MDT should be continued concomitantly to anti-reaction drugs in type 1 and type 2 reactions. In those who have completed MDT, management of reactions does not require restarting MDT Thalidomide is sometimes used in combination with corticosteroids for controlling severe type 2 reactions M. leprae (Clofazimine is considered an anti-reaction agent in addition to its role in MDT against . leprae . ManagemThe use of tumor necrosis factor inhibitors (TNF-α inhibitors) has been considered in patients with type 2 reactions. However, the TNF-α inhibitor infliximab has been associated with the clinical development of leprosy and, upon its discontinuation, the occurrence of type 1 reactions A 28-year-old Indian male, living in the United States for 2 years, presented with an 8-month history of multiple skin plaques that were well defined with their edges slightly raised, and scaly, and localized mostly in the face, upper and lower limbs, and trunk . Some ofAlmost 2 months after he began receiving MDT, he developed a sudden onset of bilateral lower limb edema, with inflammation of previously existent skin plaques distributed in his four limbs and in the trunk. No new skin lesions were identified . He alsoType 1 reactions are characterized by increasing swelling, erythema, and tenderness of previously existing skin lesions accompanied by increased neuritis manifested as pain or tenderness in one or more nerves associated with pain and nerve function impairment The foremost priority in the management of type 1 reactions is the treatment of neuritis followed by decreasing acute inflammation, easing pain, and reversing nerve and eye damage If the reversal reaction becomes chronic, requiring high doses of corticosteroids for extended periods, adjunctive clofazimine, started at 100 mg three times daily and subsequently tapered over many months, may be used  [5]. CycThe clinical management of leprosy reactions is challenging. Leprosy reactions occur among patients with borderline and lepromatous leprosy and may occur prior to the initiation of MDT, during MDT, or even years after its completion. Corticosteroids are considered the standard of care for managing type 1 and type 2 reactions, sometimes in combination with clofazimine or thalidomide. Long-term follow-up of patients diagnosed with leprosy and who have suffered leprosy reactions is needed, even after completion of MDT. Further research regarding their pathogenesis and optimal medical management is warranted.Leprosy reactions contribute significantly to neurological sequelae and functional and anatomical impairment.Leprosy reactions occur among patients with borderline and lepromatous leprosy and may occur prior to the initiation of MDT, during MDT, or even years after its completion.Early detection and effective treatment of reactions is critical to prevent nerve damage.The treatment of choice for both types of reactions (and neuritis) is corticosteroids.Alternative Language Abstract S1Translation of the abstract into Spanish by Carlos Franco-Paredes(0.03 MB DOC)Click here for additional data file.