Chemical fragmentation for massively parallel sequencing library preparation.

Fragmentation is essential in most library preparation protocols for use with massively parallel sequencing systems. Complexes that generate hydroxyl radicals, such as iron-EDTA, can be used to introduce random DNA cleavage. Here we describe a chemical fragmentation method that can be incorporated into library preparation protocols for next-generation sequencing workflows. This protocol has been validated by whole genome, amplicon and exome sequencing. Chemical fragmentation is a cost-effective alternative to current fragmentation methods that has no observable sequence bias and requires no instrumentation.

Environment and the serotonergic system.

In summary, genetics, as well as foetal and early life environmental factors shape the size or capacity of our monoamine systems, of which the serotonergic one might play a leading role. Those constitutional properties then form the biological basis for personality traits, such as impulsiveness and "sensation seeking", which interact with psychosocial settings and life events to form a pattern of reactivity to a current life event or psychosocial situation, shown as a high or low order of magnitude of gene-environment interaction. In the present paper emphasis is put on the role of genotypes of the serotonin transporter, of monoamine oxidases A and B, and of platelet monoamine oxidase B activity, which all have been shown to be of importance for behaviour and with obvious effects of interactions with environment. Under unfortunate circumstances constitutional properties might be strong enough to result in vulnerability for suicide, even with a modest influence of environment.

Monoamine oxidases: activities, genotypes and the shaping of behaviour.

The importance of an interaction between environment and biological factors for the expression for a particular behaviour is illustrated by results from a series of adolescents in which effects of platelet MAO activity and psychosocial environment on criminality was investigated. In a favourable environment platelet MAO-B activity was not associated with criminality, while a very strong association was found in adolescents from a bad psychosocial environment. Essentially similar findings were obtained when a MAO-A promoter polymorphism was analysed instead of platelet MAO-B activity. In boys, presence of the low functioning allele seemed to be protective against criminal activity in combination with a good environment, while it predisposed for criminality in a bad psycho-social environment. In girls, instead, homozygosity for the high activity MAO-A allele interacted with environment to predict criminality. Possible mechanisms underlying the role of monoamine oxidases for behaviour are discussed.

Platelet MAO and personality--function and dysfunction.

Research on the association between platelet monoamine oxidase (MAO) activity and personality traits, such as sensation seeking and impulsiveness, is reviewed with an emphasis on early history and current situation. The effects of MAO-inhibiting compounds in cigarette smoke for the interpretation of this association are discussed and recent results confirming a true association between platelet MAO activity and personality and vulnerability, for e.g. type 2 alcoholism are presented. From a clinical point of view, the link between platelet MAO activity, which is highly genetically regulated and is stable in the individual, and personality traits, has had its greatest impact on the understanding of the nature of constitutional factors making individuals vulnerable, for e.g. substance abuse and other forms of sociopathic behaviour. The molecular mechanisms underlying the association between platelet MAO and behaviour are discussed and evidence that common transcriptional factors, e.g. within the AP-2 family, regulating both the expression of platelet MAO and components of the central monoaminergic systems, such as synthesising enzymes, receptors and transporters, are presented. A hypothesis is put forward, that such common transcription factors may not directly regulate platelet MAO expression, but rather mitochondrial density, or outer mitochondrial membrane surface.

Smoking only explains part of the associations between platelet monoamine oxidase activity and personality.

Platelet monoamine oxidase (MAO) activity has been shown to be inversely associated with personality traits such as sensation seeking, impulsiveness and extraversion. Those personality traits have also been linked to vulnerability for substance abuse, e.g. tobacco smoking and early onset or "type 2" alcoholism. Compounds in cigarette smoke have been shown to be inhibitors of MAO, which has led several authors to claim that there is no association between alcoholism, which is the most studied psychiatric condition, and platelet MAO if the effect of smoking is removed. With regard to the association between personality and platelet MAO, authors have in general been cautious. In the present paper we describe a number of results which show that there is such an association, both in clinical series if the effect of smoking is removed and in series where smoking have never taken place. A cornerstone in this regard is the significant association between platelet MAO activity and both behaviour/personality, voluntary alcohol intake and biochemical measures of CNS serotonergic activity in non-human primates. Strong evidence that the regulation of platelet MAO activity takes place on a transcriptional level with an involvement of transcription factors, likely to also regulate central monoaminergic activity, are presented.

Female alcoholism: differences between female alcoholics with and without a history of additional substance misuse.

The purpose of the present study was to investigate personality traits and platelet monoamine oxidase (MAO) activity, as well as occurrence of personality disorder criteria, family characteristics, and criminal activity as related to misuse pattern in a sample of female in-patients with severe alcohol dependence. In accordance with theoretical assumptions and empirical findings of underlying neurobiological correlates of disinhibition in individuals with double or multiple misuse patterns, the female alcoholics were divided into subgroups with: (1) alcohol as their single drug of misuse (ALC; n = 16); (2) a history of additional substance misuse (ALC-DRUG; n = 11). Platelet MAO activity of the female subgroups was studied in comparison to a control group of non-clinical female subjects. Marked differences were obtained between the two subgroups. The ALC-DRUG females scored significantly higher with regard to Karolinska Scales of Personality (KSP) impulsiveness, anxiety-related traits, and non-conformity, displayed higher frequency of depression in first degree relatives, were more frequently involved in criminal activity, and more frequently diagnosed for any personality disorder according to DSM-IV, axis II. Finally, the ALC-DRUG females displayed significantly lower platelet MAO activity than controls. In conclusion, the results support the usefulness of a classification of severe alcoholic female subjects according to having/not having a history of additional substance misuse. The present female alcoholics with a history of additional misuse formed a group of individuals fulfilling the criteria similar for male type 2 alcoholism.

Genetic mechanisms of behavior--don't forget about the transcription factors.

Major changes in psychiatric phenotypes due to genetic factors are seldom the result of single gene polymorphisms, but more often the result of several genetic mechanisms. In this millennium article we discuss the notion that the expression of numerous candidate genes could be regulated by the same transcription factors, and that polymorphisms in transcription factor genes might explain some phenotypes. We describe recent results of studies on the biological marker thrombocyte monoamine oxidase (trbc MAO) and the transcription factor AP-2beta. Low levels of trbc MAO is associated with temperamental characteristics such as sensation seeking and impulsiveness, and the enzyme is genetically regulated by specific transcriptional mechanisms. Transcription factor AP-2beta is important for the development of midbrain structures and AP-2beta has several binding sites in the regulatory regions of genes encoding key proteins in the monoamine transmitter systems. We have recently shown AP-2beta to be linked to personality, binge-eating disorder, treatment with antidepressant drugs, and also to trbc MAO. Regardless of whether transcriptions factors, such as AP-2beta, regulate the expression of eg, the number of monoamine neurons or a variety of candidate genes within the monoamine systems, or both, we would like to emphasize the role of transcription factors, besides polymorphisms in monoaminergic candidate genes, when explaining inter-individual differences in temperament and psychiatric vulnerability.

Expression of microtubule-associated protein 2 in benign and malignant melanocytes: implications for differentiation and progression of cutaneous melanoma.

Cutaneous melanocytic neoplasms are known to acquire variable characteristics of neural crest differentiation. Melanocytic nevus cells in the dermis and desmoplastic melanomas often display characteristics of nerve sheath differentiation. The extent and nature of neuronal differentiation characteristics displayed by primary and metastatic melanoma cells are not well understood. Here, we describe induction of a juvenile isoform of microtubule-associated protein 2 (MAP-2c) in cultured metastatic melanoma cells by the differentiation inducer hexamethylene bisacetamide. Up-regulation of this MAP-2 isoform, a marker for immature neurons, is accompanied by extended dendritic morphology and down-regulation of tyrosinase-related protein 1 (TYRP1/gp75), a melanocyte differentiation marker. In a panel of cell lines that represent melanoma tumor progression, MAP-2c mRNA and the corresponding approximately 70-kd protein could be detected predominantly in primary melanomas. Immunohistochemical analysis of 61 benign and malignant melanocytic lesions showed abundant expression of MAP-2 protein in melanocytic nevi and in the in situ and invasive components of primary melanoma, but only focal heterogeneous expression in a few metastatic melanomas. In contrast, MAP-2-positive dermal nevus cells and the invasive cells of primary melanomas were TYRP1-negative. This reciprocal staining pattern in vivo is similar to the in vitro observation that induction of the neuronal marker MAP-2 in metastatic melanoma cells is accompanied by selective extinction of the melanocytic marker TYRP1. Our data show that neoplastic melanocytes, particularly at early stages, retain the plasticity to express the neuron-specific marker MAP-2. These observations are consistent with the premise that both benign and malignant melanocytes in the dermis can express markers of neuronal differentiation.

The genotype of human transcription factor AP-2beta is associated with platelet monoamine oxidase B activity.

Platelet monoamine oxidase B (MAO; EC 1.4.3.4.) activity is stable in the individual and is mainly genetically regulated. Levels of MAO-B in platelets have repeatedly been shown to be associated with personality traits. We have recently also demonstrated an association between the genotype of AP-2beta to a variety of personality traits as well as binge-eating disorder. In the present study we have analysed blood samples from 158 males and 64 females with regard to platelet MAO activity and genotype of transcription factor AP-2beta. In both sexes homozygotes for the long allele [CAAA](5) were significantly associated with low platelet MAO activity P<0.0001 (males) and P=0.0158 (females). This study represents a novel approach to increase the understanding about the molecular mechanisms for how the MAOB gene is regulated in blood cells and how this regulation is linked to personality traits.

Fusiform incisional biopsy for pigmented skin lesions.

Current recommendations for biopsy of a pigmented skin lesion suspected of being melanoma include complete excision when possible. Because benign pigmented lesions do not warrant excisional biopsy, especially when they are large and/or on cosmetically sensitive sites, incisional biopsy is desirable. For such lesions we recommend a fusiform incisional biopsy. This tissue sampling technique not only yields an excellent cosmetic outcome, but also provides the pathologist with an optimal biopsy specimen in which he/she can assess the symmetry, circumscription, breadth, and depth of the lesion.

Cytomorphologic features of Merkel cell carcinoma in fine needle aspiration biopsies. A study of two atypical cases.

OBJECTIVE: To report atypical cytomorphologic features in fine needle aspiration biopsies (FNABs) from two cases of Merkel cell carcinoma (MCC), a primary neuroendocrine neoplasm of skin. STUDY DESIGN: Retrospective review of FNABs with histologic correlation from six patients with MCC and a report of findings from two whose smears showed atypical features. RESULTS: Typically the aspirates produce highly cellular smears of loosely clustered and individual, relatively monomorphic, small tumor cells with round to oval, regularly contoured nuclei. In two of our cases, the tumor cell nuclei exhibited a spectrum of pleomorphism ranging from moderately complex nuclear membranes with cleaves, indentations and protrusions in one case to large, markedly bizarre, convoluted nuclei and multinucleate tumor cells in the extreme case. Both cases were primary neoplasms, and the diagnosis was based on clinical, histologic and immunohistochemical data. Additionally, electron microscopy was performed on the tumor with bizarre nuclei and demonstrated rare, dense core neurosecretory granules and paranuclear bundles of intermediate filaments.

Erythrocyte and brain methionine adenosyltransferase activities in patients with schizophrenia.

The activity of methionine adenosyltransferase (MAT) was investigated in erythrocytes and postmortem brain specimens (cortex gyrus frontalis, hippocampus and thalamus) of patients with schizophrenia treated with neuroleptics. In comparison with the control group, abnormally low values of MAT Vmax and an increased MAT affinity towards methionine (lower Km values) were found in erythrocytes. In the brain, a regionally selective decrease of MAT Km was found in cortex gyrus frontalis but the Vmax values were however, unchanged. In the regions of cortex gyrus frontalis and hippocampus, but not in thalamus, the values of Vmax and Km were inversely correlated with the duration of schizophrenia. In rats treated for 28 days with the typical neuroleptic haloperidol and the atypical clozapine, a significant increase of MAT activity was found in the corpus striatum. There is the possibility that the changes observed in MAT activity in patients with schizophrenia are attributed to the neuroleptic medication.

Platelet monoamine oxidase activity in relation to alleles of dopamine D4 receptor and tyrosine hydroxylase genes.

Human personality characteristics and vulnerability to psychopathology are to a large extent dependent upon genetic factors which have yet to be fully defined. The allele distribution of the dopamine D4 receptor (D4DR) and thrombocyte monoamine oxidase (trbc MAO) activity have both been associated with personality traits which are supposedly related, namely 'sensation seeking' according to Zuckerman and 'novelty seeking' according to Cloninger, respectively. In this report, the D4DR allele distribution and trbc MAO activity were studied in 31 psychiatric patients and 21 control subjects. Trbc MAO activity is a biochemical marker of personality that has been shown to be under strong genetic influence. However, no association between the D4DR alleles and trbc MAO could be observed in this material. To our knowledge, this is the first report comparing these two markers, and based upon the results obtained, we speculate that they may be connected with different types of overlapping personality characteristics. The allele distribution of the tyrosine hydroxylase (TH) gene was also determined. TH is the rate-limiting enzyme in the biosynthesis of catecholamines, and it is believed to be involved in different kinds of psychopathology. No covariation between TH gene alleles and trbc MAO activity or D4DR alleles was observed in this material.

Is severity of premenstrual symptoms related to illness in the climacteric.

The purpose of this study was to investigate whether women in the climacteric who have previously had severe premenstrual problems have a different pattern of reporting sick than other women. The material in the study comprised women with different degrees of severity of premenstrual symptoms who participated in an epidemiological study in 1982 on patterns of reporting sick and those women who at this time were of premenopausal or menopausal age were included. Women who previously had had such serious premenstrual problems that they had to be put on the sick-list were compared with women who had had moderate or no premenstrual problems. Information about to which extent these women have reported themselves sick has been obtained from the Regional Social Insurance Office in Uppsala County. The results show that women who have previously had severe premenstrual problems and who are now of menopausal age have a significantly greater number of diagnoses and occasions and have more long-term absences from work than women in the two control groups. They also have a greater number of sick days per year compared with statistics from the National Social Insurance Board on all women in the country.

Leiomyosarcoma of the esophagus: radiographic findings in 10 patients.

OBJECTIVE: Leiomyosarcomas of the esophagus are rare malignant smooth-muscle tumors that have been described only anecdotally in the radiology literature. The objective of this study was to evaluate the clinical and radiographic findings of this unusual lesion. MATERIALS AND METHODS: A search of the radiology archives of the Armed Forces Institute of Pathology revealed 10 cases of esophageal leiomyosarcomas. Clinical and radiographic findings were reviewed retrospectively. RESULTS: All but one patient presented with dysphagia. The average duration of the dysphagia was 6.7 months, but five patients had dysphagia for 3 or fewer months. Frontal chest radiographs revealed a mediastinal mass in five patients. Barium studies revealed intramural lesions in six patients, intraluminal lesions in two, and infiltrative lesions in two. The intramural Lesions all had large exophytic components, and three contained ulceration or tracking. One of the intraluminal lesions appeared as a polypoid expansile mass and the other, as a smooth expansile sausage-shaped mass mimicking a fibrovascular polyp. CT revealed a mass involving the esophagus in five patients; three of these patients had heterogeneous lesions containing large exophytic components, central areas of low density, and extraluminal gas or contrast material within the tumor. In two patients, MR imaging revealed large masses that were isointense with skeletal muscle on T1-weighted images and hyperintense on T2-weighted images. CONCLUSION: Our experience suggests that esophageal leiomyosarcomas have radiographic findings similar to those of leiomyosarcomas elsewhere in the gastrointestinal tract. Esophageal leiomyosarcomas have a better prognosis than squamous cell carcinomas and are often amenable to surgical cure.

Fibrovascular polyps of the esophagus: clinical, radiographic, and pathologic findings in 16 patients.

OBJECTIVE: Fibrovascular polyps of the esophagus are rare benign nonneoplastic intraluminal masses. Most published reports of patients with these polyps have been anecdotal. The purpose of this study was to reassess the clinical, radiographic, and pathologic findings in a relatively large series of patients with this unusual tumorlike lesion. MATERIALS AND METHODS: A search of the radiologic archives of the Armed Forces Institute of Pathology revealed 16 cases of fibrovascular polyps of the esophagus. We reviewed the clinical, radiographic, and pathologic findings in these 16 cases. RESULTS: All 16 patients were symptomatic. Fourteen (87%) had dysphagia and four (25%) had respiratory symptoms. The average duration of symptoms was 17 months, but seven patients (44%) had symptoms for 6 or fewer months. Two patients (12%) had a history of regurgitating the tumor into the pharynx or mouth, but none had the known complication of asphyxiation due to occlusion of the larynx. Chest radiographs revealed a right-sided superior mediastinal mass and/or anterior tracheal bowing in seven patients (44%). Barium studies revealed smooth but variably lobulated intraluminal masses that originated in the lower cervical esophagus and had variable sizes and distal extents, with an average length of 15 cm. Depending on the amount of fat and fibrovascular tissue in the lesion, CT revealed a heterogeneous appearance in four patients, lesions of predominantly fat density in two, and lesions of predominantly soft-tissue density in two. CONCLUSION: Knowledge of the clinical and radiographic features of fibrovascular polyps of the esophagus is important because surgical removal of these lesions is warranted in most patients.