Nelson DeMille."The mind can cook up very subtle syndromes to throw at our bodies".  Astrid Alauda.The varying clinical presentations of not so well recognized diseases, which continue to haunt patients is well described in the above quotes. Clinicians may not recognize associated subtle early presentations of the disease, till the disease has fully manifested itself. Of all the disorders of human body, autonomic syndromes are still the least understood. Presentation and association of autonomic diseases with other disorders may be so varied and subtle that clinicians may fail to recognize autonomic disturbance as an individual clinical entity within the disease and therefore appropriate management still remains inadequate. Similar association between joint hypermobility disorder, in which postural orthostatic tachycardia syndrome and dysautonomia is increasingly being recognized as an associated clinical entity, has been further elaborated.Postural orthostatic tachycardia syndrome (POTS) is a form of dysautonomia affecting predominantly young females, consisting of a constellation of symptoms with symptomatic orthostatic intolerance and by definition, a change from supine to an upright position causes an increase in heart rate of more than 30 beats per minute or to a heart rate greater than 120 beats per minute within 10 minutes of upright position . POTS isPrimary POTS is related to partial dysautonomia and hyperadrenergic state . The cauSecondary POTS is due to peripheral autonomic denervation. Commonly associated conditions are diabetes mellitus, amyloidosis, sarcoidosis, alcoholism, lupus, heavy metal intoxication, Sjogren's syndrome and post-chemotherapy. Pure dysautonomic syndromes due to sympathetic under-activity may have POTS as a presenting feature. POTS has also been associated with musculoskeletal disorders like Ehlers-Danlos syndrome, Marfans syndrome, fibromyalgia and joint hypermobility syndrome.Joint Hypermobility syndrome (JHS) was first described by Kirk et al in 1967 to describe a disorder of generalized joint hypermobility and musculoskeletal pain . The terIn the current issue of this journal Kanjwal et al  have desThere still remain some unanswered, but important, questions. How exactly are JHS and POTS associated? Why do both POTS and JHS have female preponderance? Is it related to genetic, hormonal or environmental modification of the nervous system along with differences in musculoskeletal structure in patients that predispose to the above or is there a single cause of all these contiguous syndromes which we are yet to discover? Based on what was remarkably said by Charles Dickens "Oh the nerves, the nerves; the mysteries of this machine called man!  Oh the little that unhinges it, poor creatures that we are!" we possibly would continue to recognize new syndromes of dysautonomia and its associations with various clinical entities out of this machine called man.


Both soluble and nuclear oestrogen receptors have been measured in at least two separate sections from 72 endometrial cancers and 12 normal endometria. Concentration of oestrogen receptor is shown to be, in our hands, more meaningful when expressed per unit DNA than per unit protein, whether for soluble or nuclear receptor. Endometrial cancer cells from the central part of the tumour are shown to be receptor negative more frequently than those from peripheral tumour. Thus, in large cancers, biopsies from different areas are required before a tumour can be correctly designated as receptor positive, heterogeneous or receptor negative. The intratumoral variation of receptor status may relate to poor prognosis, since patients with homogeneous receptor-positive disease survive significantly longer than those with tumours showing either heterogeneous distribution of receptor or homogeneous absence of receptor. Intratumoral variation in receptor status is found to be more common in the group of patients who are within 7 years of their menopause, than in older patients.


To test the hypothesis that steroid hormone receptor expression is particularly pronounced in breast tumours when compared with non-neoplastic tissue, mastectomy samples were divided into 16 sectors. Multifocal tumours, of varying receptor phenotype were found in 4 patients and in addition different regions of large tumours also showed varying receptor contents. Remaining samples were found to consist of normal tissue, with fat, connective tissue and some benign breast disease. In the 9 patients with oestrogen receptor positive tumours (ER), ER content was invariably much greater in the tumours than in the remainder of the breast. Progesterone receptor (PR) content was not closely related to ER, and was lowest in the poorly differentiated tumours. This relation to differentiation was not seen in ER. The data support the view that ER concentration in ER positive tumours may reflect the transformed nature of neoplastic tissue.


Twenty-one multifocal urinary tract transitional cell carcinomas, mostly bladder tumours, from a total of six patients were processed for cytogenetic analysis after short-term culturing of the tumour cells. Karyotypically related, often identical, cytogenetically complex clones were found in all informative tumours from each case, including the recurrent tumours. Rearrangement of chromosome 9, leading to loss of material from the short and/or the long arm, was seen in all cases, indicating that this is an early, pathogenetically important event in transitional cell carcinogenesis. The presence of related clones with great karyotypic similarity in anatomically distinct tumours from the same bladder indicates that multifocal uroepithelial tumours have a monoclonal origin and arise via intraluminal seeding of viable cancer cells shed from the original tumour. Later lesions may develop also from cells shed from the so called second primary tumours. The relatively complex karyotypes seen in all lesions from most cases argue that the seeding of tumour cells is a late event that succeeds the acquisition by them of multiple secondary genetic abnormalities. © 1999 Cancer Research Campaign


Different imaging modalities as MRI, SPECT, PET and optic imaging are used together with molecular probes specific for the biological process of interest. Molecular imaging of transcription factor activity is done in animal models and mostly in transgenic reporter mice, where the transgene essentially consists of a promoter that regulates a reporter gene. During inflammation, the transcription factor NF-κB is widely involved in orchestration and regulation of the immune system and almost all imaging studies in this field has revolved around the role and regulation of NF-κB. We here present a brief introduction to experimental use and design of transgenic reporter mice and a more extensive review of the various studies where molecular imaging of transcriptional regulation has been applied during inflammation.Molecular imaging enables non-invasive visualization of the dynamics of molecular processes within living organisms  The imaging modalities in molecular imaging are SPECT and PET that detect γ- and β- radiation; MRI that detect differences in relaxation time and optical imaging that mainly record luminescent and fluorescent light [Historically conventional imaging techniques as radiography, computed tomography, ultrasonography and magnetic resonance imaging (MRI) were developed to visualize anatomical properties and changes for diagnostic purposes. Molecular imaging, which has emerged as a new discipline during the last decade, attempts to visualize functional properties. European Society for Molecular Imaging defines molecular imaging as the characterization of the dynamics of the molecular processes in the living organisms nt light . EssentiInflammation involves changes in hemodynamics, recruitment of leucocytes and platelets, and release of numerous signaling and effector molecules. All of this is adapted to type of tissue and stimuli ; additionally it is timely regulated and adjusted to severity of insult. Ideally the inflammatory response is initiated on insult and terminated after homeostasis is reestablished. However, inflammation can become chronic, which is the case for diseases like rheumatoid arthritis and inflammatory bowel disease. The complexity of the inflammatory response requires that its many functional elements are controlled coordinately in some situations and independently in others. This regulation occurs through the specificity of recruited immune cells and their differentiation, signaling pathways and gene expression. Cellular protein composition is crucial for regulation at all levels, which gives transcriptional regulation a central role in orchestrating the inflammatory process. It is suggested that various sets of genes encode the different functional elements and that these genes are coordinately regulated by dedicated transcription factors . For insThere are several hundred transcription factors involved in inflammation. In spite of this, imaging studies of NF-κB has dominated research in this field. NF-κB is attractive for inflammation studies due to the early activation, and the involvement in the large numbers of signaling pathways and the many genes related to immune functions that it controls ,5. The NAs imaging of transcription factor activity is basically only examined in transgenic reporter mice, we here briefly describe this technology. Furthermore, we review different studies related to transcription and inflammation, foremost connected to NF-κB.Transgenic reporter mice have genomically inserted an engineered DNA construct  essentially composed of a promoter and a reporter gene. Studies using reporter mice can determine activity of specific promoters and transcription factors that regulate them, which further can reflect physiological processes, disease progression and experimental manipulations. Reporter mice make possible non-invasive dynamic studies in living animals; meaning that a particular biological process can be monitored both over time and in all organs in the same animal.Promoters used in transgenes are in principle designed by two approaches: natural promoters taken from a gene of interest and artificial promoters where a set of selected cis-elements are combined. Often combinations of the two strategies are used. A promoter is composed of a core promoter containing DNA elements necessary for binding the polymerase and initiations of transcription, and a proximal promoter, placed upstream of the core promoter, containing regulatory cis-elements bound by transcription factors Fig.. In addiReporter genes are genetic markers that encode easily detectable proteins. The half-life of reporter proteins should be within hours when studying dynamic patterns to obtain close relationship with the biological process. Additionally, a short half-life will prevent accumulation of the reporter caused by potential background activity of the promoter. Extensive accumulation of the reporter can mask an induced expression. In basically all imaging studies of transgenic reporter mice, genes that encode bioluminescent or fluorescent proteins are used . The bioCo-regulated expression of different types of reporter genes is required to examine the same biological process with different imaging modalities. While bioluminescence is more sensitive in live animal imaging, fluorescent reporters are nearly always necessary for identification of single cells in tissue samples. In clinical imaging, expression of a therapeutic gene can be followed by co-regulated expression of a PET reporter gene . To obtain vivo, which have the potential for cell specific transfection [Transgenic mice are most frequently generated by pronuclear injection of fertilized eggs . With thsfection . To obtasfection ,30. A liin vivo either used the HIV long terminal repeat as promoter [ex vivo images was equal to the luciferase activity recorded in tissue homogenates. This confirms that the luminescent signal recorded in tissue reflects the actual level of reporter protein. These reporter mice have been used to examine the role of NF-κB during inflammation, inflammatory mechanisms in general and to evaluate therapeutic strategies. A selection of these studies is reviewed in this chapter with a summary listed in table The two first transgenic mice that convincingly showed that NF-κB activity could be imaged promoter  or a synpromoter . Both miPrevious observations have shown that following acute brain injury, leukocytes are recruited particularly from the liver to the damaged brain. To test the hypothesis that NF-κB has a critical role in this process, the dynamics of NF-κB activity was imaged after induction of brain injury by intracerebral injection of IL-1β. This led to an exceptionally rapid NF-κB activation in the liver, suggestive of a signal transfer that involves the neural system. To determine the role of hepatic NF-κB, it was selectively inhibited by intravenous adenoviral-mediated delivery of an IκBα super-repressor. This treatment significantly reduced the number of neutrophils recruited to the brain .in vivo imaging of the gut region after blocking signaling from the vagus nerve by transection showed increased NF-κB activity. Furthermore, in mice with experimentally induced colitis cutting the vagus nerve, and thus removing the cholinergic mediated anti-inflammatory signal, exaggerated the NF-κB activity. This elevated NF-κB activity coincided with disease severity and reduction in regulatory T-cells [The vagus nerve is shown to stimulate anti-inflammatory processes in the gut through cholinergic modulation of macrophages , and in  T-cells .In vivo imaging was used to confirm up-regulation of NF-κB activity in the lung, which also correlated well with disease parameters as cytokines, chemokines and recruitment of neutrophils. Imaging was also used to investigate to what extent duration of NF-κB activation correlated with outcome of lung inflammation [Various constitutively active forms of IκB kinases have been virally introduced in airway epithelium to test whether activation of NF-κB pathways are sufficient to generate lung inflammation . In vivoammation . Models ammation ,39.Mastitis is defined as inflammation of the mammary gland mainly caused by microbial pathogens, such as bacteria. Mastitis is quite common in breast feeding women, and in the dairy industry intra-mammary infections are of great economical importance due to loss of milk production. The dynamics of NF-κB activity was investigated in a mouse model of mastitis where E.coli was inoculated in the mammary glands of lactating mice . NF-κB wex vivo imaging of the small intestine demonstrated autoimmune disease, which had clinical parameters in agreement with celiaki [in vivo imaging of NF-κB activation is a good marker for autoimmune disease in experimental mouse models.Despite intense research efforts, the etiology of most autoimmune diseases remains obscure. It is previously shown that B-cells can present fragments of the variable region of their immunoglobulins, called idiotype (Id), on their MHC class II, which further can be recognized by T-helper cells. Such T-cells with Id-specific receptors have been described in a number of autoimmune diseases in humans -45. In a celiaki . Additio celiaki . These rNumerous molecular processes are involved in inflammation and probes that can detect some of these events have been developed. Imaging of NF-κB activation together with such probes has been performed to examine correlation of different but still connected processes. A near infrared fluorescent probe that emits light when cleaved by the protease activity of cathepsin B and K was utilized in a model of rheumatoid arthritis . The intThe discovery that diet affects gene regulation has been crucial for the understanding of diet's role in health and disease. It has been demonstrated that dietary components can be both pro-inflammatory and anti-inflammatory. Intake of high fat diet can for instance lead to low grade inflammation, which again is linked to metabolic syndrome and type 2 diabetes . In two Besides NF-κB reporter mice, very few other transgenic reporter models exists for reporting specific transcription factors related to inflammation. One exception is the luciferase reporter mice for the transcription factors Smad 2 and 3 . ActivatIn vivo imaging following induction of inflammation by various stimuli showed robust activation in all the reporter mice. In addition they showed, by exploiting specific inhibitors and activators, that NF-κB is central to the regulation of all the promoters. However, the influence of NF-κB differed between the four. The SAA and GADD45β promoter were primarily regulated by NF-κB, whereas the IκB promoter was in addition influenced by the p38 signaling pathway and the iNOS promoter also needed interferon regulated factor for maximal activation [In addition to imaging inflammatory regulation of single transcription factors, several reporter mice have been developed for studies of natural promoters involved in inflammation. Such reporter mice are valuable both for assessing the transcriptional regulation during inflammation and to evaluate the relative contribution of distinct inflammatory genes. Zhang et al., have developed four different transgenic luciferase reporter mice with promoters from important response genes in inflammation: Serum Amyloid A (SAA), inducible Nitric Oxide Synthetase (iNOS), IκB and Growth arrest and DNA-damage-inducible β (GADD45β). tivation -60.in vivo. In this study, luciferase activity could be quantified and followed over several weeks after LPS stimulation [While basically all studies of molecular imaging of transcriptional regulation in inflammations is done in mice produced by pronuclear injection there are some studies with other methods. For instance, transfection and expression of reporter genes can specifically be obtained in liver following intravenous injection of naked DNA. Such an approach has been used to image luciferase activity under the control of NF-κB . In thesmulation .in vivo. We have basically only reviewed studies on NF-κB regulation in inflammation, which have dominated this research area. The various studies illustrate the importance and advantage both to track activity in individual mice over time, to quantify the relative changes in activity and to visualize the spatial patterns of activation.Molecular imaging of transcription factors is a young research field, but has demonstrated the usefulness of visualizing regulation of transcription factors The present reporter models of NF-κB have provided valuable information in a variety of experiments, but still have potential to elucidate a number of unresolved questions related to inflammation. However, NF-κB, whose binding sites are represented in more than 200 genes, is a rather ubiquitous marker of inflammation and it is also present in genes not directly involved in inflammation. Therefore, development of promoters that are more specifically regulated is attractive in order to investigate distinct functional elements of inflammation. Such promoters must be under regulation of defined sets of transcription factors. An important improvement in this regard will be the ability to more precisely image gene regulation in individual organs, and functional events such as activation of specific cell types and productions of distinct cytokines.Due to the rapid evolution of imaging technology combined with advances in molecular biology more quantitative and localized characterization of the reporter signal will represent an important and valuable improvement. The studies presented in this review have utilized conventional two dimensional imaging of reporter gene activity, which indeed has created valuable information from distinct organs such as liver, lungs, brain and intestine; however, due to light scattering and poor tissues penetration, it is difficult to localize and quantify signals from deeper lying tissues. Information on inflammation in specific organs is vital for the understanding of biological mechanisms and to validate more precisely the effect of a treatment regime. New developments of brighter versions of optical reporter proteins as well as more red shifted fluorescent proteins is therefore crucial for obtaining tissue specific imaging. The combination of anatomical images (Computer tomography and MRI) with molecular imaging is forthcoming  and willHC owns stocks in the company Cgene AS, who holds the commercial rights of certain NF-κB-luciferase reporter mice. AK has been partly employed by Cgene AS.AK and HC authored the manuscript. Both authors read and approved the final manuscript.This work was funded by grants from the EU consortium DiMI (LSHB-CT-2005-512146) and the Norwegian Research Council.


Sir,We recently came across a case of concurrent sternal and pulmonary langerhans cell histiocytosis (PLCH), who was successfully diagnosed by a careful review of high-resolution computed tomography (HRCT) and using ultrasound (US) guidance for tissue diagnosis. PLCH is part of the spectrum of the disorder manifesting as an interstitial lung disease with an Although sternal involvement of LCH is rare and manifests with nonspecific physical and radiologic signs, PLCH presents with specific HRCT signs. Therefore, chest radiograph of a patient with a painful sternal mass should be scrutinized for any interstitial infiltrates that may lead to a specific imaging diagnosis, which was PLCH in this case.To date, sternal LCH has been diagnosed using various surgical techniques. However, tissue diagnosis can be done percutaneously using US guidance as in this case, reducing the morbidity and the length of hospital stay.


Asymmetric strand segregation has been proposed as a mechanism to minimize effective mutation rates in epithelial tissues. Under asymmetric strand segregation, the double-stranded molecule that contains the oldest DNA strand is preferentially targeted to the somatic stem cell after each round of DNA replication. This oldest DNA strand is expected to have fewer errors than younger strands because some of the errors that arise on daughter strands during their synthesis fail to be repaired. Empirical findings suggest the possibility of asymmetric strand segregation in a subset of mammalian cell lineages, indicating that it may indeed function to increase genetic fidelity. However, the implications of asymmetric strand segregation for the fidelity of epigenetic information remain unexplored. Here, I explore the impact of strand-segregation dynamics on epigenetic fidelity using a mathematical-modelling approach that draws on the known molecular mechanisms of DNA methylation and existing rate estimates from empirical methylation data. I find that, for a wide range of starting methylation densities, asymmetric—but not symmetric—strand segregation leads to systematic increases in methylation levels if parent strands are subject to de novo methylation events. I found that epigenetic fidelity can be compromised when enhanced genetic fidelity is achieved through asymmetric strand segregation. Strand segregation dynamics could thus explain the increased DNA methylation densities that are observed in structured cellular populations during aging and in disease. Through my investigations of the fidelity of epigenetic inheritance, I became intrigued by the interplay of genetic and epigenetic fidelities. Cairns proposed in 1975 that the lifetime risk of epithelial cancers would be reduced if chromosomes containing the oldest DNA strands were selectively segregated to somatic stem cells. I wondered about the implications of such asymmetric strand segregation for the fidelity of epigenetic information. To address this issue, I modelled the partitioning of DNA molecules after replication, with special attention to the molecule that contained the oldest strand. I found that the enhanced genetic fidelity that may be achieved through asymmetric strand segregation could, under some scenarios, compromise epigenetic fidelity. I am excited to pursue these studies as they apply to epigenetic changes observed to occur during aging and in human diseases, including several cancers. Cairns proposed A few reports have discussed possible epigenetic causes and consequences of asymmetric strand segregation (i) when de novo methylation events were permitted to occur on both parent and daughter strands, asymmetric strand segregation resulted in population-mean and oldest-strand methylation densities that increased monotonically  when de novo methylation events occured on both parent and daughter strands, symmetric segregation yielded population-mean and oldest-strand methylation densities that, although dynamic, remained very near the predicted equilibrium  when de novo methylation events were limited to the daughter strand, population-mean and oldest-strand methylation densities under both asymmetric and symmetric segregation remained very close to starting values (dotted lines, I compared the dynamics of mean and oldest-strand methylation densities under asymmetric and symmetric strand segregation . Three kThe population-epigenetic model I develop here reveals that asymmetric strand segregation in somatic stem cells could lead to monotonic increases in DNA methylation densities in structured cellular populations. These increases are predicted to occur when de novo methylation occurs on parent as well as daughter strands, but not when de novo methylation events are limited to the daughter strand.The predictions of my model are made using empirical estimates of methylation rates in differentiated cells, for which substantial amounts of data are available. Further work will be necessary directly to ascertain methylation rates in somatic stem cells. Nevertheless, the essential findings of my study are consistent across a broad range of parameter values (see, for instance, The accumulation of aberrant methylation predicted by my model may have different time courses depending on the biological properties of a given lineage of somatic stem cells, and on the initial methylation density of a given locus. When somatic stem cell division always gives rise to one stem cell and one differentiated cell, as I model here, substantial increases in DNA methylation densities can occur over just a few cell divisions . When soWhat might be the functional implications of the increased DNA methylation densities predicted under asymmetric strand segregation? The accumulation of methyl groups on a long-lived DNA strand could serve as a signal to guide asymmetric strand segregation itself My results may also have implications for the etiology of cancer in humans. Several epithelial cancers are associated with reductions in epigenetic fidelity, including the accumulation of aberrant methylation and abnormal gene silencing Thus, it is possible that directional change in epigenetic information may be a cost of the increased genetic fidelity achieved through asymmetric strand segregation, with implications for human disease.I developed a simplified model of an epithelial crypt with which to track methylation dynamics . Each crI modelled replication and methylation dynamics for a single, methylated locus such as one of those on the hemizygous X chromosome in a human male, or on the inactive X chromosome in a human female. Prior to cell division, the locus undergoes semi-conservative replication, producing two double-stranded DNA molecules. Each molecule is composed of a parent strand from the original double-stranded molecule, and a newly-synthesized daughter strand. The model to be described in detail below compares methylation dynamics under asymmetric  and symmMethyl groups are added to CpG cytosines in DNA by two different processes: maintenance methylation and de novo methylation. Maintenance methylation is performed by maintenance methyltranferases, which exhibit a preference for hemimethylated CpG/CpG dyads, are thought to localize to the replication fork, and operate principally during DNA replication Here, the maintenance methylation rate, in vivo on the parent strand was addressed experimentally during the 1970s, yielding conflicting data The question of whether or not de novo methylation events can occur I developed vector l crypts . The iniThe first element of I developed matrix The first row of matrix asymmetric strand segregation, I multiplied matrix Both asymmetric and symmetric strand segregation are modelled using matrix symmetric strand segregation, I used a similar approach, multiplying matrix To model a single round of DNA replication and cell division under Several authors have investigated the rate of maintenance methylation, bona fide biological variation among CpG cytosine sites, and perhaps among loci.Comparatively few studies have investigated the rate of de novo methylation. The estimates that do exist exhibit substantial variation. Pfeifer et al. For de novo methylation rates, I first chose parameter values that yield an expected equilibrium methylation density of 0.8 under the model described previously


The literature on hallucinations is reviewed, including history; theoretical background from physiological, biochemical and psychological points of view; classification; causation; presentation in different psychiatric and neurological disorders and in normal persons. The available evidence suggests that hallucinations result from a failure of the metacognitive skills involved in discriminating between self-generated and external sources of information. Management of hallucinations is briefly discussed. Hallucinations are intriguing psychological phenomena that have a number of important clinical, theoretical and empirical implications; they are also among the most severe and puzzling forms of psychopathology. Usually regarded as characteristic of psychoses, they are found in a wide range of medical and psychiatric conditions. Moreover, a substantial minority of normal individuals report hallucinatory experiences. Phenomenologically they are the commonest and the most important disorder of perception. Although the exact cause and pathogenesis of hallucinations are not known, evidence points toward multiple etiological factors for the hallucinatory phenomenon.Freud (1953) felt that hallucinations are very similar to dreams and that both conditions represent a psychotic state in which there is a complete lack of time sense. In this process, thoughts are transformed into visual images, mainly of a visual sort, that is, word presentations are taken back to corresponding “thing” presentations. According to Kolb and Brodie (1982), hallucinations represent a breakthrough of preconscious or unconscious material into consciousness in response to certain psychological situations and needs, e.g., wish fulfillment, enhancement of self-esteem, guilt feelings. The contents of hallucinations are thought to reflect their psychodynamic significance. Hallucinations often occurring with delusions during psychotic states may represent the concrete symbolic expression of delusional ideas that are seeking other routes of expression.et al. (1950) demonstrated that electrical stimulation of certain cortical or subcortical structures induced different types of hallucinations. He proposed the theory of abnormal brain excitation as a mechanism of production of hallucinations. The neurophysiologic dissociation theory  proposes that hallucinations result from a dissociation between primary sensory cortex and cortical association areas which exert a regulatory influence on the former. The perceptual release theory  postulated the presence of a censorship mechanism in the brain which actively excluded from the consciousness the majority of sensory information that is received continually by the brain. But the censorship mechanism can operate only when there is constant flow of sensory inputs. If by any chance there is stoppage or impairment of sensory input , then earlier perception or memory traces emerge into the conscious, and the individual experiences hallucinations. This explains the occurrence of hallucinations following specific sensory modality deprivation.Hughlings Jackson (1932) suggested that hallucinations occur when the usual inhibitory influences of the uppermost level are impeded, thus leading to release of middle-level activity, which takes the form of hallucinations. This model is known as disinhibition model. Penfield In schizophrenia (SCZ), there is evidence that very high levels of dopamine in the limbic system play a major role in emergence of hallucinations and delusions. Antipsychotic medications, which block central dopamine activity, alleviate the hallucinations of psychosis. Drugs with strong dopaminergic effect, such as L-dopa, methylphenidate, bromocriptine, pramipexole and piribedil, may induce hallucinations. D-amphetamine, a direct dopamine agonist, may also induce psychosis and hallucinations. The fact that hallucinations were also described in Parkinson’s disease before the introduction of L-dopa indicates that not only hyperdopaminergic states but also hypodopaminergic states, presumably due to progressive loss of dopamine projections to the cortex, can induce hallucinations.A deranged cholinergic neurotransmission has also been involved in the pathophysiology of hallucinations. For example, alteration of consciousness and hallucinations have been described widely since ancient times for members of the Solanaceae family of plants (belladonna and dhatura), which contain scopolamine, atropine and other antimuscarinic agents. Hallucinations occur in about 30% of patients with Alzheimer’s disease and 60% of patients with Lewy body dementia, which are characterized by reduction in acetylcholine and abnormalities in nicotinic and muscarinic receptor expression.2A receptor agonist or partial agonists. In addition, hallucinations have been reported as side effects of Selective serotonin reuptake inhibitors (SSRIs), which increase the availability of serotonin in the synaptic cleft.Serotonin has also been implicated in the causation of hallucinations, based on the fact that a number of hallucinogenic drugs, like lysergic acid diethylamide (LSD), mescaline, psilocybin and ecstasy, appear to act, at least in part, as serotonin 5 HTA possible role of glutamate in hallucinations is suggested by the finding that glutamate antagonists like phencyclidine and ketamine can induce hallucinations. This has led to the hypothesis that psychotic symptoms may in part be attributed to hypofunction of NMDA receptors.PET and SPECT studies using GABA-A receptor ligands showed that the intensity of hallucinations was strongly associated with diminished GABA-A binding, specifically in the left medial temporal region.et al. (2007) suggested that in healthy subjects, EEG synchrony preceding speech reflects the action of a forward model system, which dampens auditory responsiveness to self-generated speech. However, the dampening was deficient in SCZ patients with hallucinations. Hubl et al. (2007) investigated the responsiveness of the auditory cortex during hallucinations in SCZ patients using the N100 evoked potential in response to auditory stimulation. During hallucinations, the N100 amplitude was smaller, presumably because of a reduced left temporal responsivity. They concluded that the findings indicate competition between auditory stimuli and hallucinations for physiological resources in the primary auditory cortex. Abnormal activation of the primary auditory cortex may thus be a constituent of auditory hallucinations.Using cognitive tasks involving speech production and perception, Heinks-Maldonado et al., 2009). Volume reductions in the prefrontal and cerebellar cortices have also been reported and may be associated with impairment in the monitoring or awareness and volition of internal speech. MRI brain scans of 66 patients of type I bipolar disorder and 66 controls indicated that hallucinations and positive symptoms were associated with gray matter reduction in the left middle temporal gyrus .In a systematic review of 46 studies (including 1444 patients with SCZ and 1327 controls) on superior temporal gyrus (STG), volumetry MRI studies in SCZ revealed that left STG or subregions appear to be more involved in the generation of hallucinations and thought disorder than right sides. The majority of 5 follow-up studies found evidence of progressive changes in volumes  found increased rCBF in the left temporal region in hallucinating patients. McGuire et al. (1993) reported that hallucinatory state was associated with greater blood flow in language-related areas, significantly so in a region corresponding to Broca’s area and anterior cingulate and left temporal cortices. Nonpsychotic visual hallucinations in Parkinson’s disease (PD) were associated with hypoperfusion in the right fusiform gyrus and hyperperfusion in the right superior and middle temporal gyri. These temporal regions are important for visual object recognition, and these rCBF changes are associated with inappropriate visual processing and are responsible for nonpsychotic visual hallucinations in PD . Using PET during auditory verbal hallucinations (AVH), patients demonstrated a significant activation of the supplementary motor area, anterior cingulum, medial superior frontal area and cerebellum. Activation was also observed in the left superior frontal area, right superior temporal pole and right orbitofrontal region . Stephane et al. (2006) suggest that the abnormal laterality of the supplementary motor area activity accounts for the failure to attribute speech generated by one’s own brain to one’s self and that the activation of Wernicke’s area accounts for the perceptual nature (hearing) of the patient’s experience. Lahti et al. (2006) using PET reported a positive correlation between hallucinations and activation of anterior cortex and a negative correlation with hippocampus-parahippocampus. Functional MRI revealed that AVHs were associated with activation in the right STG and posterior insula, the right middle temporal gyrus, and hippocampus/parahippocampal gyrus. Somatic hallucinations were associated with activation in the thalamus, insula and posterior cingulate gyrus bilaterally. Further activation was evident in the left middle frontal gyrus, the precentral gyrus, twin foci in the postcentral gyrus, inferior parietal lobule, the middle and superior temporal gyri with right-sided inferior frontal gyrus, and precuneus . Functional imaging studies have consistently shown the involvement of associative sensory areas. Moreover, the hallucinating brain may be characterized by stronger inputs from subcortical centers ; reduced control by the dorsolateral prefrontal cortex; and aberrant activation of dorsal anterior cingulate, which is involved in source monitoring.Stephane et al. (2008) proposed that auditory hallucinations are internally generated speech perceptions that are localized in the left temporal lobe, in the left peri-Sylvian region. If hallucinations are misidentified perceptions originating in the speech perception area of the left temporal lobe, then hallucinating patients should have problems identifying a simultaneously presented external speech sound, especially when the sound is presented lateralized to the left hemisphere. Using a dichotic listening paradigm with pair-wise presentations of consonant-vowel syllables, they showed that patients with schizophrenia who experience frequent and/or intense auditory hallucinations fail to show an expected right ear advantage (REA), which may be indicative of a functional deficit in this brain region. Patients who experience frequent hallucinations are impaired in their ability to use top-down cognitive strategies to overcome a ‘bottom-up stimulus’-driven right ear effect when instructed to focus attention on the left or right ear stimulus. The behavioral data are supported by MR brain imaging data looking for pathology of gray matter density in patients with schizophrenia, particularly in the left hemisphere. The reviewed studies revealed significant reductions in gray matter density in the left peri-Sylvian region, including thinning of the cortical mantle, also supported by altered neuronal activation as seen in the fMRI results.Hallucinations cause sensory modality-specific activation in cerebral areas involved in normal sensation. A disturbance in perception of speech seems to have a central role in occurrence of auditory hallucinations. Anatomically, auditory hallucinations appear to involve primary and association cortices; Broca’s and Wernicke’s areas; subcortical, paralimbic, limbic regions; ventral striatum; and thalamus. Furthermore, they are suggested to be associated with the dysmodulation of the information flow from ventral striatum to thalamus and cortex caused by increased dopaminergic activity in mesolimbic pathway. Hugdahl et al. (1994) using psycholinguistic approach postulated that auditory hallucinations stem from disruption in language production caused by parasitic memory. David (1994) linked verbal hallucinations to inner speech. He argued that inner speech is produced and controlled by an inner-voice-inner-ear system, and the disruption of the system may lead to verbal hallucinations. Hamsley postulated that normal perception is dependant on an interaction between current sensory input and regularities that have been detected in previous sensory input. Memory of previous regularities and the current context result in ‘expectancies’ or ‘response biases’ towards the current sensory input. The central defect in hallucinations is the reduction of the influence of regularities of past experiences on current perception. This weakened influence would result in ambiguous and unstructured sensory input. Thus the unstructured sensory input would fail to inhibit the emergence of material from long-term memory into awareness; its emergence would be experienced as hallucinations .Hallucinations can be seen as erroneous perception or “sensory deception.” Hence researchers have been investigating the integrity of perceptual functions in persons with hallucinations. Basically, 2 approaches can be distinguished. The first is concerned with bottom-up, or data-driven, perceptual processing, such as deficits in the processing of incoming visual stimuli in primary visual cortex; hearing impairment and auditory hallucinations associated with sensory impairment in older people; or the emergence of hallucinations in sensory deprivation. The second approach is concerned with top-down, or conceptual, processing in perception. This refers to processes that contribute to perception that do not originate in the external world but in the brain of the perceiver. Examples of such top-down factors are prior knowledge, perceptual expectations, attentional modulation and mental imagery. Bentall (1990) proposed that discrimination between internal and external sources of information is a metacognitive skill called source monitoring. Hallucinations result from failure of this skill. Hoffman et al. (2005) proposed a PAD model of complex visual hallucination, in which the hallucinatory images generally occur in the focus of the visual field and are seen against the background of the existing visual scene. According to the PAD model, both sensory impairment and attentional abnormalities are needed for hallucinations to arise. More specifically, a combination of impaired attentional binding and poor sensory activation of a correct proto-object, in conjunction with a relatively intact scene representation, biases perception to allow the intrusion of a hallucinatory proto-object into a scene perception. Proto-objects refer to holistic or part-based abstracted object representations that are segmented from visual information and act as candidates for further processing but are at such an early processing stage that they have not yet entered conscious awareness. Such proto-objects are in multiple competitions for further processing. The interplay of top-down and bottom-up biasing information will eventually determine which proto-object will “win” and enter conscious awareness. Cholinergic dysfunction may result in a failure to properly integrate sensory information (bottom-up) and prior expectation (top-down). The PAD model proposes that impaired attentional binding is due to abnormal lateral frontal activity; poor sensory activation of a correct proto-object is due to abnormal ventral visual stream activity; and intrusion of a hallucinatory proto-object is mediated by increased temporal versus frontal activity. Diederich et al. (2005) suggested that visual hallucinations should be considered a dysregulation of the gating and filtering of external perception and internal image production. Contributive factors for their model include poor primary vision, reduced activation of primary visual cortex, aberrant activation of associative visual and frontal cortex, lack of suppression or spontaneous emergence of internally generated imagery through the ponto-geniculo-occipital system, intrusion of rapid eye movement dreaming imagery into wakefulness, erratic changes of the brainstem filtering capacities through fluctuating vigilance and medication-related overactivation of mesolimbic system. Not all of these have to be present, and different combinations will lead to differences in phenomenology. Llinas and Pare (1991) suggested that conscious perception is subserved by intrinsic activity in thalamocortical circuits that is constrained or modulated by sensory input. They considered the primary difference between conscious perception and dream imagery to be found in the weight given to sensory input, which is a large weight in conscious perception, and hence sensory factors largely determine the final percept, whereas this weight is negligible in dream imagery. Integrating the sensory impairment approach with the top-down approach, Behrendt and Young (2004) viewed hallucinations as “underconstrained perceptions” that arise when the impact of sensory input on thalamocortical circuits is reduced. If sensory constraints  are weak, e.g., due to sensory impairment, attentional mechanisms (i.e. top-down influences) may become the dominant modulatory influence on thala mocortical oscillatory activity that gives rise to conscious percepts — in this case, hallucinations.Collerton To test different hypotheses regarding neurocognitive basis of hallucination, Hoffman and McGlasher (2006) developed a neural network simulation. Their model specifically concerns auditory hallucinations as typically reported in people with schizophrenia and is based on 2 popular hypotheses regarding neurobiological basis of schizophrenia: (a) overzealous pruning of synapses that are an extension of normal developmental pruning during adolescence and (b) alterations in the dopaminergic neuromodulatory system in schizophrenia. To assess and compare these 2 hypotheses, a computer simulation of some aspects of speech perception was developed. This system was found to produce spontaneous percepts simulating hallucinated speech when the working memory component was excessively pruned or when neuronal responses were modulated to simulate a hyperdopaminergic system. Comparing the performance of the network with that of actual hallucinating patients and normal controls while tracking (repeating while simultaneously listening to) speech that was phonetically degraded, revealed that the neural network simulation producing the best match to speech-tracking performance of human hallucinators was an overpruned system with compensatory hypodopaminergic adjustments.Metacognition refers to “thinking about thinking”or to beliefs and attitudes held about cognition. Most recent models of hallucinations assume that hallucinations are related to misattribution of private events. These private events may include a number of types and modalities, such as inner images, inner speech, voices, intrusive thoughts, vivid daydreams and bodily sensations. Different approaches have attempted to explain this misattribution, with the most prominent of these relating hallucinations to a misattribution of inner speech, problems in source monitoring, and relations between hallucinations and beliefs. Recent cognitive models of hallucination have attributed these misattributions to cognitive bias or cognitive deficit. Cognitive biases are present when some forms of information are processed preferentially in comparison to others. Examples of cognitive bias include the preferential recall of negative information in patients with depression. Cognitive deficits occur when there are disruptions of specific cognitive functions, including, for example, problems with working memory, inhibition or attention. It is probable that disturbances in cognitive processes as a result of cognitive deficits  and cognitive bias factors (top-down factors) are both responsible for hallucinations.et al., 2004) on source-monitoring tasks.Some studies provide evidence for Bentall’s notion that hallucinators are impaired in their ability to discriminate between real and imagined events and reveal a specific bias towards attributing their thoughts to an external source. This externalizing bias seems related, at least in part, to the effects of both an inadequate use of cognitive effort cues and the emotional salience of stimuli . An important aspect of this theory concerns research showing similarities between intrusive thoughts and hallucinations, because the genesis of hallucinations is seen as a reaction to intrusive experiences. Like hallucinations, intrusions are also often experienced as ego dystonic and uncontrollable; share similarities in terms of form, content and triggers ; and are usually accompanied by subjective discomfort. The theory also states that the need to attribute intrusive thoughts to an external source is due to motivational factors. The presence of certain intrusive thoughts may lead to negative affect in the subject in the form of anxiety or cognitive dissonance. According to cognitive dissonance theory, dissonance occurs when 2 cognitions  contradict each other, resulting in an uncomfortable state from which an individual is motivated to escape. Morrison et al. (1995) argued that to reduce the levels of negative affect, the subject chooses to externalize the intrusive thought, resulting in hallucinations. Morrison also posited the important role that metacognitive beliefs may play in this misattribution process. Metacognitive beliefs are beliefs concerning one’s own mental process. This may include beliefs that mental events should be controllable, that thoughts are dangerous or harmful or that intrusive thoughts are acceptable and beneficial. When the occurrence of intrusive thoughts does not comply with the subject’s metacognitive beliefs, an aversive state of arousal results (cognitive dissonance), which the subject tries to escape by externalizing the intrusive thoughts , thus maintaining consistency in his belief system. A number of studies have found evidence for an association between metacognitive beliefs and the presence of hallucinations.Morrison is in agreement with the theory that hallucinations are internal cognitive events that are misattributed to an external source. His explanation for this misattribution is based on the fact that hallucinations are in some way linked to normal intrusive thoughts and that because of motivational factors these intrusive thoughts become externalized by the individual (Morrison Alemon and Laroi (2008) presumed that an integrated network of cognitive processes may contribute to perceptual experience, and proposed that 4 different routes may result in a hallucination: (a) release phenomena due to lesions in sensory pathways/the arousal system ; (b) irritative processes acting on cortical centers that integrate perceptual information; (c) unconstrained activation of the attentional spotlight; and (d) a cognitive route with key roles for affective state, top-down factors and source-/self-monitoring.Release phenomena: Disturbed visual input may cause hallucinations by means of an abnormal release of central processing, though the level at which the release occurs is unclear. The implication of primary and sensory cortices seems straightforward, but changes have also been observed in subcortical brain areas  in patients with optic nerve lesions. The capacity for generating complex visual hallucinations may be located in the association cortex, which is released by restricted lesions, a loss of cortico-cortical inputs, and alteration of activity via the reticular activating system. In this model, release hallucinations would arise from damage to the component sensory input or its connecting pathway to sensory experience.Irritative processes: Hallucinations caused by spontaneous activation of sensory cortical areas have been primarily described in epilepsy. The fact that focal cortical resection can result in complete remission of epileptic hallucinations indicates that these hallucinations cannot be attributed to a release phenomenon. According to this model, irritative processes would be confined to aberrant activation of the module sensory experience.Attentional spotligh: The thalamic reticular nucleus (TRN) acts as a searchlight to guide attention to certain features of the environment and thereby intensifies processing of these features. The thalamus is acknowledged as one of the three important nodes in a network underlying selective attention that comprises the prefrontal cortex, parietal cortex and the thalamus. The TRN facilitates sensory processing by facilitating detection and discrimination through the initiation of burst-firing (when a population of neurons fires with a high-frequency burst in contrast to a continuous low-frequency baseline firing) in the lateral geniculate nucleus (LGN). Although the encoding of an externally presented stimulus becomes distorted during burst-firing, the background noise decreases significantly, thereby enhancing signal detectability. This model proposes that activation of the TRN in the absence of sensory input could explain the involuntary character of hallucinations. Such activation is possible via connections from the prefrontal cortex to the TRN, via feedback from sensory association cortex and via neurotransmitters such as dopamine and acetylcholine.Cognitive route: The model assumes that hallucinations may arise from a cognitive route, in which emotion, top-down perceptual mechanisms and metacognitive functions  play a decisive role. The idea here is that an affective state (stress and anxiety) negatively influences monitoring systems, resulting in external misattribution of internally generated events, but can also influence top-down factors such as perceptual expectations. There is evidence that emotional factors can modulate verbal self-monitoring and source-monitoring. Neuro-imaging studies show that amygdala activation in response to emotional stimuli can boost activation of perceptual areas, which indicates that the affective state can influence perception and imagery.The top-down factors’ subsystem in the model refers to the influence of prior knowledge, perceptual expectations and attention on perception. Excessive activation of such top-down factors can “push” images to the right within the sensory experience box . As a reThe monitoring subsystem in the model  refers tHallucination is a fundamental symptom in psychiatry. Two hundred years of research into this phenomenon has not yet answered the following questions: (1) Are hallucinations pathognomic of psychosis? (2) Can the presence of hallucinations as such or in different modalities and forms include or exclude certain diagnoses? (3) What is the neural substrate of hallucination? These questions are very basic to the understanding of mental diseases, and more research in both phenomenological and theoretical areas is necessary to fathom the secret. Conventionally, hallucinations are treated as psychotic features. However, there is ample evidence to support the fact that hallucination could be present in nonpsychotic conditions. Mechanism and nosological status of these conditions are yet not clear. Assessing cultural background in the evaluation of hallucination is important as the concept of reality varies across cultures and there is a possibility of culturally sanctioned hallucination. A part from effective pharmacological treatment, more awareness is needed regarding psychological treatment of hallucination, which can help us deal with refractory hallucinations.


Glucocorticoids are the mainstay of asthma therapy. They are primarily used to suppress airway inflammation, which is the central pathological change in asthmatic patients’ airways. This is achieved by many different mechanisms. The classical mechanism is by suppression of the genetic transcription of many inflammatory cytokines that are key in asthma pathophysiology (transrepression). On the other hand, the transcription of certain inhibitory cytokines is activated by glucocorticoids (transactivation), a mechanism that also mediates many of the adverse effects of glucocorticoids. The onset of action through these mechanisms is often delayed (4-24 hours). Other mechanisms mediated through non-genomic pathways are increasingly appreciated. These are delivered in part by binding of glucocorticoids to nonclassical membrane-bound glucocorticoid receptors or by potentiating the α1-adrenergic action on the bronchial arterial smooth muscles, in addition to other mechanisms. These effects are characterized by their rapid onset and short duration of action. Understanding these different mechanisms will help in the development of new and better drugs to treat this common disease and to develop new improved strategies in our approach to its management. Here, the genomic and non-genomic mechanisms of actions of glucocorticoids in asthma are briefly reviewed, with special emphasis on the current updates of the non-genomic mechanisms. Asthma is one of the most common chronic illnesses worldwide that has a huge burden on every society. It is the most common chronic illness in childhood and is one of the most common causes of admission to the hospital among children and adults.2 Its preRecent data from the United States indicate improved outcomes of asthma morbidity and mortality, with fewer annual hospitalizations for asthmatic attacks and fewer asthma-related deaths. This couIn this short review I will discuss, focusing on recent updates, the different genomic and non-genomic mechanisms by which glucocorticoids act.genomic action of glucocorticoids implicates the activation or repression of multiple genes. Glucocorticoids significantly suppress airway inflammation, mainly through genomic mechanisms. This action involves many steps and therefore takes effect with a time lag of about 4 to 24 hours. L22 Ltranstivation  However,tivation 25transrepression  Cla Clatranspression  Targetedpression  In additpression  NO contrpression 32 Gene tpression  These cypression 33 Other pression  Activatipression The development of glucocorticoids has passed through different stages since the introduction of cortisone, which has glucocorticoid and mineralocorticoid activity, to the development of prednisone, which has less mineralocorticoid activity, and then the development of glucocorticoids with almost no mineralocorticoid activity, like dexamethasone. AdvancedGlucocorticoids can act through regulation of mRNA stability. They have been shown to block the production of several pro-inflammatory cytokines by increasing their mRNA degradation rate. In additnon-genomic action. This mode of action entails mechanisms that do not directly and initially influence gene expression, and their effects are not blunted by inhibitors of gene transcription such as actinomycin D.[A more recently recognized and increasingly understood mode of action of glucocorticoids is the omycin D. Unlike tomycin D.Several basic mechanisms of the non-genomic action of glucocorticoids have been identified so far. Different attempts were made to classify these mechanisms. First, the Mannheim classification includedangiogenesis.[Bronchial airway vasculature plays an important role in asthma pathogenesis by contributing to the inflammatory process. Several changes are noted in the vasculature of asthmatic patients’ airways, including vasodilation, hyperperfusion and increased microvascular permeability. These changes are essential to edema formation and inflammatory cell recruitment.46 The aiogenesis.481 -adrenergic agonists has been shown to be potentiated in the airways of asthmatic patients.[The significant increase in airway mucosal blood flow in asthmatic patients with stable asthma or acute exacerbation has been shown in many studies in comparison with healthy subjects.50 Calcul5150patients.541-adrenoceptors or to be re-uptaken by neurons and released again. Pre-treatment with 5 mg terazosin, a selective α1-adrenoceptor antagonist, inhibited the effect of fluticasone on bronchial blood flow, which supports this hypothesis.[How can glucocorticoids cause vasoconstriction? Evidence suggests that glucocorticoids decrease airway blood flow by modulating sympathetic control of vascular tone. Glucocorticoids inhibit the extraneuronal monoamine transporter–mediated uptake of norepinephrine by bronchial arterial smooth muscle cells56 Figur. Therefopothesis. This effpothesis.Several lines of evidence from clinical research suggested an important role for this mode of action in the treatment of asthma. A protective effect was demonstrated 2 hours after administration of a single dose of 100 mcg of fluticasone proprionate inhaler using bronchial provocation with cyclic adenosine monophosphate (cAMP). Higher d5962Inhaled glucocorticoids were found to have additional benefit to Systemic glucocorticoids in the management of patients with acute asthma exacerbation in the ER. A recent meta-analysis comparing inhaled to systemic glucocorticoids; or a combination of inhaled and systemic glucocorticoid to a combination of placebo and systemic glucocorticoid in the ER, included 17 different randomized double-blind placebo-controlled trials with data from 1133 subjects (470 adults and 663 children). It concl2-agonist and ipratropium bromide to children with acute asthma exacerbation, in addition to giving systemic steroids.Taken all together, inhaled glucocorticoids may need to be administered simultaneously with bronchodilators in high and repeated or sequential doses as a way to obtain an early effect and maintain it throughout the time. However,Other vascular effects of glucocorticoids have been reported, but their relevance to asthma is not clear.–70++ influx upon induction of allergic inflammation.[Glucocorticoids can be incorporated into cell membranes, leading to changes in the physicochemical properties of the membrane . Subsequ71ammation.Glucocorticoids were shown to have a rapid T-cell immunosuppressive action that is mediated through membrane-bound GRs . These r74762 and cysteinyl leukotrienes, which are known to be potent inducers of bronchoconstriction, excessive mucus secretion, and edema of the bronchial mucosa. When glucocorticoids bind their own receptors in the cytosol, some GR protein complex components, like src, dissociate and block the EGF-mediated cPLA2 activation and subsequently arachidonic acid production.[in vitro studies and animal studies have demonstrated decrease in baseline and post-challenge production of cysteinyl leukotrienes after treatment with glucocorticoids, in vivo studies generally failed to show a similar effect.[Arachidonic acid is produced by the action of cytosolic phospholipase A2 (cPLA2) on membrane phospholipids. By binding to its receptor on the cell membrane, epidermal growth factor (EGF) can activate cPLA2 and therefore arachidonic acid production. Arachidonic acid is a precursor to prostaglandin Doduction.79 Althour effect. This sugIn conclusion, glucocorticoids, as the gold standard asthma therapy, deliver their effects by many ways but most importantly through transrepression by binding key pro-inflammatory transcription factors. Non-genomic mechanisms are being increasingly recognized, especially those affecting bronchial vascular blood flow. Understanding the different mechanisms by which glucocorticoids act should enable the development of new agents that have more desired effects while avoiding many adverse effects by being more specific and selective. It should also open the door for new indications and strategies in the treatment of asthma, things that we may observe in the near future.


The many millions of humans who have life-long virus infections represent a major health issue for the 21st century but also a unique opportunity for investigative virologists. For persistent virus infections to endure, two ingredients are essential. The first is a unique strategy of viral replication; that is, instead of killing its host cell, the pathogen causes little to no damage so it can continue to reside in those cells. The second requirement for persistent virus infection is an immune response that does not react to or remove virus-infected cells. Overall, our knowledge of how viral genes and cellular factors interact to allow persistence to occur is incomplete. Although our libraries contain volumes of facts on this subject, many physiologic functions and interrelationships of viral genes with host genes that establish persistence remain, in large part, unknown. We do know that acutely infected cells express viral peptides, which, when attached to host major histocompatibility complex (MHC) molecules on their surfaces, signal the immune system to kill such cells. However, viruses apply numerous avoidance strategies to persist. One is direct selective pressure to suppress the infected host's innate and/or adoptive immune system that would otherwise destroy them reviewed  [3]. ForViruses can cause persistent infection early in life directly from mother to child in utero or in newborns whose immune system is immature, and even later in adults after the immune system has matured. Infection in early life was initially attributed to immunologic tolerance, that is, deletion or removal of cell clones that generate an antiviral immune response However, results from our work Specific antiviral CTLs are rarely found in adult mice or in humans persistently infected early in life. Reconstitution of virus-specific T cells first shown by adoptive transfer of anti-LCMV-specific CD4 and CD8 T cells readily purged viruses and cleared infection from blood and tissues of LCMV-carrier mice Therefore, we conclude that viral infection initiated in neonates or in utero does not lead to deletion of T or B cell clones specific for the virus Immune responses reflect the sum of positive versus negative regulators of that response. Negative regulators function to prevent an excessive immunologic response leading to immunopathologic disease. Of those so far identified Viruses that initiate persistent infections in juveniles or adults take advantage of this negative regulatory system by actively causing hosts to make regulators that turn off or exhaust the expected antiviral immune response. The discovery that viruses can escape immunologic attack and persist by this means again came from the study of LCMV infection in its natural murine host In the LCMV persistent infection induced in adults, IL-10 is produced primarily by virus-infected cDCs and perhaps by B cells PD-1 expression increases on T cells during persistent LCMV infection and, as stated above, these T cells become hyporesponsive or exhausted and cannot clear infection The exploration and understanding of negative immune regulators, including IL-10 and PD-1, are still at an early stage; nevertheless, the implications are important and profound. First, exhausted or hyporesponsive T cells found in persistent infections can be resurrected to functional capacity. This is true not only for the LCMV model in which the phenomenon was uncovered Clearance of a persistent LCMV infection requires virus-specific CD4 T cell help to assist virus-specific CD8 T cells Currently our laboratory and others are engaged in the discovery of additional negative immune regulators and their signaling pathway(s) using gene chip and forward genetics technology. These projects have a multitude of applications. Some examples are the development of pharmacologic small molecules as effective antagonists of negative immune regulators, the use of transient negative regulator blockers as an adjuvant approach to enhance both prophylactic and therapeutic vaccination, and the determination of how long during the course of persistent virus infection exhausted T cells can be rescued to become antiviral effector T cells. As always, the goal is to understand basic principles in viral pathogenesis and to extend results in the murine model to resolve persistent infections of humans.


With the help of large disease databases, medical scientists have begun to discern how such patterns occur over the course of a lifetime. A new review focused on developmental immunotoxicology explores how this integrative perspective might inspire novel strategies for lowering the risk and prevalence of immune-based diseases influenced by environmental stimuli Many chronic diseases share three common features: 1) early-life exposures to chemical agents or pathogens, 2) evidence of immune insult or dysfunction, and 3) the appearance of disease biomarkers in exposed children although disease itself may not manifest until later in life. One example of interlinked disease conditions highlighted by the authors is metabolic syndrome, defined as the co-occurrence of at least three of five conditions: insulin resistance, obesity, high blood pressure, elevated triglycerides, and reduced HDL cholesterol.Immune dysfunction is central to the underlying physiology of metabolic syndrome, and the authors posit that the seeds of such dysfunction may be planted in childhood. They describe pre- and postnatal exposures to environmental risk factors that produce postnatal lipid dysregulation and immune dysfunction. However, it is not yet known whether immune dysfunction is an underlying cause of metabolic syndrome or simply an associated or disease-facilitating characteristic.The practical key to preventing metabolic syndrome may lie in treatments that address overall patterns and their progression, not just the initial presenting condition. “For those patterns of disease with immune involvement,” the authors write, “preventing the underlying immune dysfunction is the single most effective option to minimize the risk of one or more chronic diseases later in life.” This will require more information about risk factors for immune dysfunction that are encountered during development or childhood. Therefore, the authors also recommend that chemicals and pharmaceuticals be tested for developmental immunotoxicity end points; currently, safety assessments are based solely on adult exposures.The authors say patterns of disease can be used to better predict, prevent, and treat diseases associated with an immune-related pattern of diseases, and may also serve as the basis for environmental protection and testing to prevent exposure to developmental immunotoxicants that may contribute to multiple interconnected diseases. But pattern-based evaluation, prevention, and treatment will require a shift from the prevailing single-organ approach to disease classification and management.


SirBritish Journal of Cancer. The study compares findings in primary colorectal cancer and paired metastases in 38 patients, showing differences with respect to EGFR pathway deregulation, which may imply a different response to anti-EGFR monoclonal antibodies. As a consequence, the authors suggest that the analysis of metastatic lesions should be considered both for patient management and when planning clinical trials with anti-EGFR drugs.We read with great interest the paper by Statistical analysis is lacking in terms of study question definition and sample size calculation and therefore all the considerations made are not supported by a formal hypothesis verification;KRAS mutational status, BRAF mutational status, PTEN protein expression, EGFR protein expression and EGFR gene status, respectively, showing that only for EGFR gene status the concordance did not reach a ‘good’ value. As no intra-sample concordance results are available, the question whether these differences are because of a true discordance between primary tumour and metastases or rather due to an intra-sample variability  between different areas within the same sample because of biological heterogeneity  that post hoc analyses of randomised studies, which led to the regulatory restriction for anti-EGFR MoAbs to KRAS wild-type patients were conducted almost exclusively on primary tumours  that the only universally accepted determinant of resistance to anti-EGFR MoAbs are


We have found that although wild-type numbers of chromatophore precursors are generated in the first day of development and migrate normally in enz mutants, the numbers of all three chromatophore cell types that ultimately develop are reduced. Further, differentiated melanophores and xanthophores subsequently lose dendricity, and iridiphores are reduced in size. We demonstrate that enz function is required cell autonomously by melanophores and that the enz locus is located on chromosome 7. In addition, zebrafish enz appears to selectively regulate chromatophore development within the neural crest lineage since all other major derivatives develop normally. Our results suggest that enz is required relatively late in the development of all three embryonic chromatophore types and is normally necessary for terminal differentiation and the maintenance of cell size and morphology. Thus, although developmental regulation of different chromatophore sublineages in zebrafish is in part genetically distinct, enz provides an example of a common regulator of neural crest-derived chromatophore differentiation and morphology.The development of neural crest-derived pigment cells has been studied extensively as a model for cellular differentiation, disease and environmental adaptation. Neural crest-derived chromatophores in the zebrafish (Danio rerio) consist of three types: melanophores, xanthophores and iridiphores. We have identified the zebrafish mutant  The neural crest is a transient vertebrate embryonic cell population that gives rise to a wide variety of cell types, including chromatophores, craniofacial cartilage, and neurons and glia of the peripheral nervous system Vertebrate chromatophore populations are readily observed, as they produce their own visible intrinsic markers. In addition, chromatophores are not strictly required for viability microphthalmia-associated transcription factor (mitf), which is both necessary and sufficient for melanophore development, and dopachrome tautomerase (dct), an enzyme in the melanin synthesis pathway c-kit is also expressed by melanoblasts, and appears to be necessary for their migration and survival kit ortholog fms, which has no known role in mammalian melanocyte development, is required for the migration of embryonic xanthophores and the specification of a subset of adult melanophores in zebrafish fms is expressed by embryonic xanthoblasts and macrophages, which can be distinguished from one another based on location and cellular morphology xanthine dehydrogenase (xdh), which is correspondingly expressed by xanthoblasts fms and xdh are co-expressed in a subset +mitf cells in the premigratory neural crest, which may represent uncommitted precursors of melanophores or xanthophores c-kit, however, and both have been used as specific diagnostic markers of xanthoblasts at migratory and post-migratory stages of chromatophore development gch expression is observed in both melanoblasts and xanthoblasts endothelin receptor (ednr) b, is also expressed by neural crest-derived chromatophore precursors ednrb mutant mice are almost completely devoid of melanocytes ednrb1/rose mutants display defects in subsets of adult melanophores and iridiphores but lack an embryonic chromatophore phenotype ednrb1 is initially expressed by all chromatophore sublineages, but by late embryonic/early larval stages, is restricted to iridiblasts and iridiphores Studies from several vertebrates, including zebrafish, have led to the extensive characterization of melanophore development, and to a lesser extent, xanthophore and iridiphore development mitf expression in mammals Morphologically, differentiated melanophores and xanthophores are large and dendritic with many processes, while iridiphores are rounded in shape ednrb is necessary for both melanocyte and enteric neuron development ednrb1 appears to be dispensable for their embryonic development Despite these and other data that have been amassed regarding chromatophore development in a variety of vertebrate systems, many questions remain. For example, relatively few genes have been identified that are required for the development of all embryonic chromatophores, and yet are specific to chromatophore sublineages within the neural crest. For example, Sox10 is necessary for development of chromatophore sublineages, but is also required for some crest-derived neurons and glia, and mammalian b431endzone , which was isolated based on the abnormal appearance of embryonic chromatophores. Our results indicate that all three chromatophore types are similarly reduced in cell size and number in enz mutant embryos. We show that enz is required specifically for chromatophore sublineages within the neural crest, and that this requirement is cell autonomous. Further, we report the identification of multiple enz alleles and progress toward the molecular identification of the enz locus. We suggest that enz is a relatively late cue required by pigment cell sublineages of the neural crest during embryogenesis, and is indicative of common requirements of chromatophores that are distinct from other neural crest derivatives.We report here characterization of the zebrafish mutant enz was identified in a chemical  mutagenesis screen for mutations affecting neural crest derivatives as previously described enz mutant was identified based on its altered pigment pattern. At 54 hours post-fertilization (hpf), the morphology of melanophores is dramatically altered being smaller and less stellate in b431enz mutant embryos compared to wild-type siblings , although they remain smaller than wild-type siblings. embryos . In addil stages . enz mutt 30 dpf . As adulenz mutant embryos, we investigated whether the development of other cell types derived from the neural crest were affected by the enz mutation. Molecular markers indicated that neural crest-derived peripheral neuron and cranial glial populations develop normally in enz homozygotes  at 3 dpf, as well as fms expression between 48 and 53 hpf, revealed that while many xanthophores are present in enz homozygotes, their numbers are reduced in a manner qualitatively similar to that of melanophores and iridiphores . At 27 hpf, enz melanophores are stellate, but pale compared to those of wild-type siblings . Together, these data indicate that mutations in enz similarly affect all three neural crest-derived chromatophore types with respect to cell size and also result in loss of the the typical stellate morphology of melanophores and xanthophores.In wild-type zebrafish embryos raised at 28.5°C, neural crest-derived melanophores normally begin to differentiate at approximately 25 hpf, while xanthophores and iridiphores begin to overtly differentiate at about 42 hpf and 72 hpf, respectively siblings . After 2nophores  [11], [1at 3 dpf . Similar hpf see . While tenz mutant, and b1golden embryos. To facilitate the identification of donor-derived melanophores, particularly from enz donors, we utilized hosts homozygous for the b1golden mutation, which results in pale melanophores that are distinct from wild-type melanophores enz hosts , there are several factors that prevent conclusions to be drawn from the experiments. First, TUNEL, as well as most other reliable methods to detect cell death in vivo are only useful in the instantaneous detection of apoptotic cells. Thus, if cell death were to occur at a low rate over an extended period, differences in cell death rate would be very difficult to detect. This would be especially true if, as is the case in enz mutants, the cell types of interest are only slightly reduced in number compared to control embryos. Nevertheless, it is potentially significant to note that multiple studies have shown that chromatophores differentiate and undergo apoptosis in response to factors that control chromatophore motile responses, that is, the redistribution of pigment organelles within chromatophores . Further, these factors are likely to utilize shared intracellular signaling pathways in different pigment cell types. Thus, the chromatophore phenotypic consequences and cell autonomous action of enz are at least consistent with a role for enz in such a pathway and the regulation of differentiation and apoptosis via such a pathway. Finally, in addition to the molecular identification of enz it will also be critical to determine whether any of the enz alleles we have identified result in loss of function. While the majority of chromatophores develop in enz mutants, the same situation is observed in most mutants and morphants affecting chromatophore development enz alleles exhibit altered morphology. Thus, it is possible that one or more of the enz alleles we have identified result in loss of function, but this will only be possible to ascertain by cloning the enz locus.The mechanism by which disruption of enz larvae die after 2–3 weeks of development, a minority of individuals does survive to adulthood but were found to be significantly smaller than wild-type siblings. Because neural crest-derived chromatophores are not required for viability and are not thought to regulate growth enz must function in other cells required for growth and viability. Interestingly, there is abundant evidence demonstrating roles for pituitary hormone signaling systems in chromatophore development and growth control enz acts cell autonomously, enz would be predicted to be a downstream effector if it acts in such a hormonal signaling pathway.While the vast majority of homozygous enz that disrupt genes involved in organellar biosynthesis and transport ATPase 6 subunit v0c (atp6v0c) and vacuolar protein sorting protein 18 (vps18) result in punctate melanophore appearance, apparent loss of xanthophores and reduced iridiphore numbers ATP synthase and vacuolar protein sorting genes generated in a viral insertion-based mutagenesis screen vps35 ortholog assigned to chromosome 7 (http://www.sanger.ac.uk/), however, sequencing of the four enz alleles reported here did not reveal any molecular lesions in the vps35 coding sequence. This does not rule out the possibility that enz encodes vps35, as lesions may be present in non-coding regulatory regions of this gene in enz mutants. Rawls and colleagues have identified a mutation linked to chromosome 7 and SSLP marker z10441 that phenotypically resembles enzenz mutants complement this mutation. Further, the genomic region in which the enz gene is located contains several other genes involved in organellar function that may be good candidates for the enz locus (http://zfin.org). Thus, it is also possible that the enz locus encodes a protein required for normal formation or activity of cellular organelles. Specifically in the case of enz, such a gene would have to regulate chromatophore cell morphology instead of or in addition to organelle synthesis and distribution. The characterization of enz mutant embryos and larvae presented here may provide clues to the mechanisms behind phenotypes described for organellar biosynthesis mutants and morphants described previously. In any case, the potential relevance of the speculation discussed here will be resolved with the molecular identification of the enz locus.It is also potentially informative to note that recent reports have described mutants and morphants with live phenotypes similar to enz homozygotes. Together with our finding that the enz mutation acts cell autonomously with respect to melanophore development, this suggests that the enz locus does not encode a gene generally required by neural crest-derived cells, but rather a molecule intrinsic to chromatophores themselves. Our results strongly suggest that the differentiation and maintenance of cell morphology of all three zebrafish chromatophore cell types depends on enz and are thus at least partially coordinately regulated during terminal differentiation. However, since the development of all three chromatophore lineages is unaffected in enz mutants until late in their embryonic development and many mutants and identified genes selectively affect individual or subsets of chromatophore types enz, will provide important contributions to ultimately determining the genetic regulation of chromatophore development and leading to potential insights into clinically relevant conditions in humans involving pigment cells. Further, many of these mutants are likely to provide valuable insights into the genetic regulation of color changes and light adaptation in fish and other organisms.Although many loci have been identified that are required for crest-derived chromatophore development, relatively few of them affect all three chromatophore cell types found in zebrafish Adult zebrafish and embryos were maintained in the Ohio State University zebrafish facility. Adults and embryos were reared at 28.5°C and embryos were staged based on morphological criteria, according to Kimmel et al., 1995. Mutant lines were maintained in the AB* and WIK backgrounds. Homozygous mutant embryos and wild-type siblings were obtained by crossing heterozygous carriers.enz embryos were imaged on a Zeiss Axioplan compound microscope. Individual cells were outlined and areas calculated using Zeiss Axioplan software with appropriate scalings. Iridiphores in 6 dpf larvae were imaged on a Leica dissecting microscope. Individual cells were outlined and areas calculated using Spot Advanced software with appropriate scalings. A minimum of 3 melanophores or iridiphores in at least 3 wild-type and 3 enz mutant individuals were utilized for each set of measurements. Standard errors of mean were calculated for wild-type and enz cell areas, and mean numbers were also subjected to a one-tailed t-test to determine whether differences observed between wild-type and enz chromatophore areas were statistically significant.Melanophores in 2 days post-fertilization (dpf) wild-type and enz mutant larvae were placed in epinephrine (10 mg/ml) at 4 dpf for ∼10 minutes, that results in redistribution of melanosomes to the center of the cell body in wild-type melanophores . Larvae were fixed in 4% paraformaldehyde at 4°C overnight, rinsed and stored in 1∶1 PBS∶glycerol. Larvae were then deyolked, mounted on single bridge cover slips, and viewed on a Zeiss Axioplan microscope. Melanophores in the dorsal stripes were counted from somite 5 to somite 14. Iridiphores in 6 dpf larvae were counted in the dorsal and ventral stripes. Live fish were viewed under incident light using a Zeiss dissecting microscope. Iridiphores were counted posterior of the hindbrain in the dorsal stripe, and caudally from the posterior edge of the yolk ball in the ventral stripe. Standard errors of mean were calculated for wild-type and enz cell counts. The mean numbers of cells in wild-type and enz embryos were subjected to a one-tailed T-test to determine whether the decrease in cell numbers in enz mutant embryos compared to wild-type siblings was statistically significant.Melanophores in 4 dpf larvae were counted in dorsal stripes. To better visualize distinct melanophores, wild-type and mitfa and c-ret cDNAs were provided by D. Raible c-kit and fms were obtained from D. Parichy dct and ednrb1 cDNA clones were provided by R. Kelsh In situ hybridizations were performed as described by Thisse, et al. (1993) with minor modifications. A detailed protocol will be provided upon request. + cells within whole-mount wild-type and endzone larvae.Antibody labeling was performed as previously described +/b431enz) crosses were manually dechorionated and injected at the one to two cell stage with 2–5% lysinated rhodamine dextran  in 0.2 M KCl. To facilitate the identification of donor-derived melanophores, we utilized hosts homozygous for the b1golden mutation, which results in pale melanophores that are distinct from wild-type melanophores Genetic mosaics were produced using cell transplantation techniques enz alleles were maintained in the AB* background. For mapping purposes, enz carriers in this background were crossed to a wild-type WIK line, which is polymorphic to AB* enz carriers in the WIK background were then used to generate parthenogenetic diploid progeny by suppressing the second meiotic division with early pressure enz embryos and wild-type siblings were identified by live phenotype and used to obtain DNA. enz was initially placed on the zebrafish genomic map based on PCR amplification of simple sequence length polymorphisms (SSLPs) from diploid genomes b431enz was initially demonstrated to be linked to SSLP markers near the centromere of chromosome 7 based on the MGH mapping panel (http://zebrafish.mgh.harvard.edu/zebrafish/index.htm). enz was shown to cosegregate with z10441 (map position 36.7 cM), z11625 (map position 51.1 cM) and z6819 (map position 45.0 cM). Further linkage analysis was performed on enz using haploid genomes, or meioses enz locus distal to z6819 and z55887 and proximal to z8252 and z10441.