Overview of literature and information on "khat-related" mortality: a call for recognition of the issue and further research

During the past 20 years or so, more has become known about the properties of khat, its pharmacology, physiological and psychological effects on humans. However, at the same time its reputation of social and recreational use in traditional contexts has hindered the dissemination of knowledge about its detrimental effects in terms of mortality. This paper focuses on this particular deficit and adds to the knowledge-base by reviewing the scant literature that does exist on mortality associated with the trade and use of khat. We sought all peer-reviewed papers relating to deaths associated with khat. From an initial list of 111, we identified 15 items meeting our selection criteria. Examination of these revealed 61 further relevant items. These were supplemented with published reports, newspaper and other media reports. A conceptual framework was then developed for classifying mortality associated with each stage of the plant's journey from its cultivation, transportation, consumption, to its effects on the human body. The model is demonstrated with concrete examples drawn from the above sources. These highlight a number of issues for which more substantive statistical data are needed, including population-based studies of the physiological and psychological determinants of khat-related fatalities. Khat-consuming communities, and health professionals charged with their care should be more aware of the physiological and psychological effects of khat, together with the risks for morbidity and mortality associated with its use. There is also a need for information to be collected at international and national levels on other causes of death associated with khat cultivation, transportation, and trade. Both these dimensions need to be understood.

Dopaminergic modulation of the human reward system: a placebo-controlled dopamine depletion fMRI study

Reward related behaviour is linked to dopaminergic neurotransmission. Our aim was to gain insight into dopaminergic involvement in the human reward system. Combining functional magnetic resonance imaging with dopaminergic depletion by α-methylparatyrosine we measured dopamine-related brain activity in 10 healthy volunteers. In addition to blood-oxygen-level-dependent (BOLD) contrast we assessed the effect of dopaminergic depletion on prolactin response, peripheral markers for dopamine and norepinephrine. In the placebo condition we found increased activation in the left caudate and left cingulate gyrus during anticipation of reward. In the α-methylparatyrosine condition there was no significant brain activation during anticipation of reward or loss. In α-methylparatyrosine, anticipation of reward vs. loss increased activation in the right insula, left frontal, right parietal cortices and right cingulate gyrus. Comparing placebo versus α-methylparatyrosine showed increased activation in the left cingulate gyrus during anticipation of reward and the left medial frontal gyrus during anticipation of loss. α-methylparatyrosine reduced levels of dopamine in urine and homovanillic acid in plasma and increased prolactin. No significant effect of α-methylparatyrosine was found on norepinephrine markers. Our findings implicate distinct patterns of BOLD underlying reward processing following dopamine depletion, suggesting a role of dopaminergic neurotransmission for anticipation of monetary reward.

The therapeutic potential of the humoral pattern recognition molecule PTX3 in chronic lung infection caused by Pseudomonas aeruginosa

Chronic lung infections by Pseudomonas aeruginosa strains are a major cause of morbidity and mortality in cystic fibrosis (CF) patients. Although there is no clear evidence for a primary defect in the immune system of CF patients, the host is generally unable to clear P. aeruginosa from the airways. PTX3 is a soluble pattern recognition receptor that plays nonredundant roles in the innate immune response to fungi, bacteria, and viruses. In particular, PTX3 deficiency is associated with increased susceptibility to P. aeruginosa lung infection. To address the potential therapeutic effect of PTX3 in P. aeruginosa lung infection, we established persistent and progressive infections in mice with the RP73 clinical strain RP73 isolated from a CF patient and treated them with recombinant human PTX3. The results indicated that PTX3 has a potential therapeutic effect in P. aeruginosa chronic lung infection by reducing lung colonization, proinflammatory cytokine levels (CXCL1, CXCL2, CCL2, and IL-1β), and leukocyte recruitment in the airways. In models of acute infections and in in vitro assays, the prophagocytic effect of PTX3 was maintained in C1q-deficient mice and was lost in C3- and Fc common γ-chain-deficient mice, suggesting that facilitated recognition and phagocytosis of pathogens through the interplay between complement and FcγRs are involved in the therapeutic effect mediated by PTX3. These data suggested that PTX3 is a potential therapeutic tool in chronic P. aeruginosa lung infections, such as those seen in CF patients.

Natural variations at position 93 of the invariant Vα24-Jα18 α chain of human iNKT-cell TCRs strongly impact on CD1d binding

Human invariant natural killer T (NKT) cell TCRs bind to CD1d via an "invariant" Vα24-Jα18 chain (iNKTα) paired to semi-invariant Vβ11 chains (iNKTβ). Single-amino acid variations at position 93 (p93) of iNKTα, immediately upstream of the "invariant" CDR3α region, have been reported in a substantial proportion of human iNKT-cell clones (4-30%). Although p93, a serine in most human iNKT-cell TCRs, makes no contact with CD1d, it could affect CD1d binding by altering the conformation of the crucial CDR3α loop. By generating recombinant refolded iNKT-cell TCRs, we show that natural single-nucleotide variations in iNKTα, translating to serine, threonine, asparagine or isoleucine at p93, exert a powerful effect on CD1d binding, with up to 28-fold differences in affinity between these variants. This effect was observed with CD1d loaded with either the artificial α-galactosylceramide antigens KRN7000 or OCH, or the endogenous glycolipid β-galactosylceramide, and its importance for autoreactive recognition of endogenous lipids was demonstrated by the binding of variant iNKT-cell TCR tetramers to cell surface expressed CD1d. The serine-containing variant showed the strongest CD1d binding, offering an explanation for its predominance in vivo. Complementary molecular dynamics modeling studies were consistent with an impact of p93 on the conformation of the CDR3α loop.

Time-lapse microscopy and classification of 2D human mesenchymal stem cells based on cell shape picks up myogenic from osteogenic and adipogenic differentiation

Current methods to characterize mesenchymal stem cells (MSCs) are limited to CD marker expression, plastic adherence and their ability to differentiate into adipogenic, osteogenic and chondrogenic precursors. It seems evident that stem cells undergoing differentiation should differ in many aspects, such as morphology and possibly also behaviour; however, such a correlation has not yet been exploited for fate prediction of MSCs. Primary human MSCs from bone marrow were expanded and pelleted to form high-density cultures and were then randomly divided into four groups to differentiate into adipogenic, osteogenic chondrogenic and myogenic progenitor cells. The cells were expanded as heterogeneous and tracked with time-lapse microscopy to record cell shape, using phase-contrast microscopy. The cells were segmented using a custom-made image-processing pipeline. Seven morphological features were extracted for each of the segmented cells. Statistical analysis was performed on the seven-dimensional feature vectors, using a tree-like classification method. Differentiation of cells was monitored with key marker genes and histology. Cells in differentiation media were expressing the key genes for each of the three pathways after 21 days, i.e. adipogenic, osteogenic and chondrogenic, which was also confirmed by histological staining. Time-lapse microscopy data were obtained and contained new evidence that two cell shape features, eccentricity and filopodia (= 'fingers') are highly informative to classify myogenic differentiation from all others. However, no robust classifiers could be identified for the other cell differentiation paths. The results suggest that non-invasive automated time-lapse microscopy could potentially be used to predict the stem cell fate of hMSCs for clinical application, based on morphology for earlier time-points. The classification is challenged by cell density, proliferation and possible unknown donor-specific factors, which affect the performance of morphology-based approaches. Copyright © 2012 John Wiley & Sons, Ltd.

LUPA: a European initiative taking advantage of the canine genome architecture for unravelling complex disorders in both human and dogs

The domestic dog offers a unique opportunity to explore the genetic basis of disease, morphology and behaviour. Humans share many diseases with our canine companions, making dogs an ideal model organism for comparative disease genetics. Using newly developed resources, genome-wide association studies in dog breeds are proving to be exceptionally powerful. Towards this aim, veterinarians and geneticists from 12 European countries are collaborating to collect and analyse the DNA from large cohorts of dogs suffering from a range of carefully defined diseases of relevance to human health. This project, named LUPA, has already delivered considerable results. The consortium has collaborated to develop a new high density single nucleotide polymorphism (SNP) array. Mutations for four monogenic diseases have been identified and the information has been utilised to find mutations in human patients. Several complex diseases have been mapped and fine mapping is underway. These findings should ultimately lead to a better understanding of the molecular mechanisms underlying complex diseases in both humans and their best friend.

Accuracy of Recognition and Diagnosis of Comorbid Depression in the Nursing Home

A variety of research has documented high levels of depression among older adults in the health care setting. Additional research has shown that care providers in health care settings are not very effective at diagnosing comorbid depression.This is a troublesome finding since comorbid depression has been linked to a number of negative outcomes in older adults. Early results have indicated that comorbid depression may be associated with a number of unfavorable consequences ranging from impairments in physical functioning to increased mortality.The health care setting with arguably the highest rate of physical impairment is the nursing home and it is the nursing home where the effects of comorbid depression may be most costly. Therefore, the current analysis uses data from the Institutional Population Component of the NationalMedical Expenditure Survey (US Department of Health and Human Services, 1990) to explore rates of both recognized and unrecognized comorbid depression in the nursing home setting. Using a constructed proxy variable representative of the DSM-III-R diagnosis of depression, results indicate that approximately 8.1% of nursing home residents have an unrecognized potential comorbid depression.

Aggressive Behaviour of Different Social and Demographic Groups in Post-Totalitarian Society

The project looked at aggressiveness in different age and social groups of modern post-totalitarian society, beginning with the hypotheses that the greatest risk groups are teenagers and the unemployed, and that there is a link between aggression and the level of meaningfulness of life. The groups studied comprised about 200 persons from urban areas of eastern Ukraine, including schoolchildren, students, white collar workers, self-employed persons, the unemployed and pensioners. Workers in industry were not included as this group has virtually disappeared in Ukraine at present since most enterprises have ceased to work and most workers have moved into the groups of the unemployed or self-employed. Participants were divided into age groups of 13-14, 16-17, 18-22, 24-45, 46-60 and over 60, with each group including approximately equal number of men and women. Research methods included Buss-Darky techniques, the "hand test" (E. Wagner), the "non-existent animal" technique, a Rozenzweig picture frustration study, purpose-in-life tests and an interview. The Buss-Darky test showed that schoolchildren have the highest level of aggression, followed by students. These groups have high indexes in virtually all types of aggression, including its open form. The self-employed have relatively lower indexes, although they are more likely to manifest it openly, while such open manifestations are less likely among white-collar workers, pensioners and the unemployed. The least aggressive were the unemployed and pensioners, although the latter had a relatively high level of hostility. In terms of age, aggression was shown to decrease with age, which Ms. Ivanova attributes to the still imperfect control mechanisms of teenagers and their less mature personalities. Among the younger groups girls showed a slightly higher level of aggression, although this situation was reversed among older people. The risk groups inclined to manifest open forms of aggressiveness can therefore be seen to be teenagers and students. Other tests used show aggressiveness as a feature of the current state, rather than as an inherent feature and the results obtained were somewhat different. In the interviews, all adults referred to the increased aggressiveness in society and most stated that they themselves had become more aggressive and bad-tempered. The ability of individuals to adapt to their social environment was also investigated and schoolchildren turned out to have the lowest adaptation index and the unemployed the highest. MS. Ivanova attributes that latter, rather surprising, result to the fact that the constant frustrations facing the unemployed force them to actively seek ways and means of adapting in order to survive. The final aspect considered was the possible connection between human aggressiveness and the meaningfulness of life. Here the groups with the most meaningful lives were the self-employed and pensioners. The latter result, again rather surprising, was attributed to the desire of people who have already lived the greater part of their lives to place more weight on what they have already done, in order to prove to themselves that they have not lived in vain. The hypothesis that aggressiveness is conversely related to the meaningfulness of life was only partially confirmed. In the two extreme cases (schoolchildren and pensioners) this was indeed the case, but the remaining groups did not show any such connection. From the data obtained, Ms. Ivanova concluded that life in modern post-totalitarian society does indeed foster a rise in people's aggressiveness, and this was supported by the fact that indexes of aggressiveness proved to be higher than the norm. Her original hypothesis as to the groups in society most at risk from open aggression confirmed in the case of teenagers but not of the unemployed, who had relatively low aggressiveness indexes and the highest degree of adaptation.

Tolerance and Critical Thinking

Post-soviet countries are in the process of transformation from a totalitarian order to a democratic one, a transformation which is impossible without a profound shift in people's way of thinking. The group set themselves the task of determining the essence of this shift. Using a multidisciplinary approach, they looked at concrete ways of overcoming the totalitarian mentality and forming that necessary for an open democratic society. They studied the contemporary conceptions of tolerance and critical thinking and looked for new foundations of criticism, especially in hermeneutics. They then sought to substantiate the complementary relation between tolerance and criticism in the democratic way of thinking and to prepare a a syllabus for teaching on the subject in Ukrainian higher education. In a philosophical exploration of tolerance they began with relgious tolerance as its first and most important form. Political and social interests often lay at the foundations of religious intolerance and this implicitly comprised the transition to religious tolerance when conditions changed. Early polytheism was more or less indifferent to dogmatic deviations but monotheism is intolerant of heresies. The damage wrought by the religious wars of the Reformations transformed tolerance into a value. They did not create religious tolerance but forced its recognition as a positive phenomenon. With the weakening of religious institutions in the modern era, the purely political nature of many conflicts became evident and this stimulated the extrapolation of tolerance into secular life. Each historical era has certain acts and operations which may be interpreted as tolerant and these can be classified as to whether or not they are based on the conscious following of the principle of tolerance. This criterion requires the separation of the phenomenon of tolerance from its concept and from tolerance as a value. Only the conjunction of a concept of tolerance with a recognition of its value can transform it into a principle dictating a norm of conscious behaviour. The analysis of the contemporary conception of tolerance focused on the diversity of the concept and concluded that the notions used cannot be combined in the framework of a single more or less simple classification, as the distinctions between them are stimulated by the complexity of the realty considered and the variety of its manifestations. Notions considered in relation to tolerance included pluralism, respect and particular-universal. The rationale of tolerance was also investigated and the group felt that any substantiation of the principle of tolerance must take into account human beings' desire for knowledge. Before respecting or being tolerant of another person different from myself, I should first know where the difference lies, so knowledge is a necessary condition of tolerance.The traditional division of truth into scientific (objective and unique) and religious, moral, political (subjective and so multiple) intensifies the problem of the relationship between truth and tolerance. Science was long seen as a field of "natural" intolerance whereas the validity of tolerance was accepted in other intellectual fields. As tolerance eemrges when there is difference and opposition, it is essentially linked with rivaly and there is a a growing recognition today that unlimited rivalry is neither able to direct the process of development nor to act as creative matter. Social and economic reality has led to rivalry being regulated by the state and a natural requirement of this is to associate tolerance with a special "purified" form of rivalry, an acceptance of the actiivity of different subjects and a specification of the norms of their competition. Tolerance and rivalry should therefore be subordinate to a degree of discipline and the group point out that discipline, including self-discipline, is a regulator of the balance between them. Two problematic aspects of tolerance were identified: why something traditionally supposed to have no positive content has become a human activity today, and whether tolerance has full-scale cultural significance. The resolution of these questions requires a revision of the phenomenon and conception of tolerance to clarify its immanent positive content. This involved an investigation of the contemporary concept of tolerance and of the epistemological foundations of a negative solution of tolerance in Greek thought. An original soution to the problem of the extrapolation of tolerance to scientific knowledge was proposed based on the Duhem-Quine theses and conceptiion of background knowledge. In this way tolerance as a principle of mutual relations between different scientific positions gains an essential epistemological rationale and so an important argument for its own universal status. The group then went on to consider the ontological foundations for a positive solution of this problem, beginning with the work of Poincare and Reichenbach. The next aspect considered was the conceptual foundations of critical thinking, looking at the ideas of Karl Popper and St. Augustine and at the problem of the demarcation line between reasonable criticism and apologetic reasoning. Dogmatic and critical thinking in a political context were also considered, before an investigation of critical thinking's foundations. As logic is essential to critical thinking, the state of this discipline in Ukrainian and Russian higher education was assessed, together with the limits of formal-logical grounds for criticism, the role of informal logical as a basis for critical thinking today, dialectical logic as a foundation for critical thinking and the universality of the contemporary demand for criticism. The search for new foundations of critical thinking covered deconstructivism and critical hermeneutics, including the problem of the author. The relationship between tolerance and criticism was traced from the ancient world, both eastern and Greek, through the transitional community of the Renaissance to the industrial community (Locke and Mill) and the evolution of this relationship today when these are viewed not as moral virtues but as ordinary norms. Tolerance and criticism were discussed as complementary manifestations of human freedom. If the completeness of freedom were accepted it would be impossible to avoid recognition of the natural and legal nature of these manifestations and the group argue that critical tolerance is able to avoid dismissing such negative phenomena as the degradation of taste and manner, pornography, etc. On the basis of their work, the group drew up the syllabus of a course in "Logic with Elements of Critical Thinking, and of a special course on the "Problem of Tolerance".

Human monocytoid cells as a model to study Toll-like receptor-mediated activation

THP-1 2A9, a subclone of the monocytoid cell line THP-1 and known to be exquisitely sensitive to LPS, was tested for TNF production following triggering by excess doses of TLR ligands. TLR2, TLR4 and TLR5 agonists, but neither TLR3 nor TLR9 agonists, induced TNF production. When used at lower concentrations, priming by calcitriol strongly influenced the sensitivity of cells to LPS and different TLR2 triggers (lipoteichoic acid (LTA), trispalmitoyl-cysteyl-seryl-lysyl-lysyl-lysyl-lysine (Pam3Cys) and peptidoglycan (PGN)). Priming by calcitriol failed to modulate TLR2 and TLR4 mRNA and cell surface expression of these receptors. TNF signals elicited by TLR2 agonists were blocked by the TLR-specific antibody 2392. CD14-specific antibodies showed variable effects. CD14-specific antibodies inhibited TNF induction by LTA. High concentrations partially inhibited TNF induction by Pam3Cys. The same antibodies failed to inhibit TNF induction by PGN. Thus, THP-1 2A9 cells respond by TNF production to some, but not all TLR agonists, and the wide variety of putative TLR2 agonists interact to variable degrees also with other cell-surface-expressed binding sites such as CD14. THP-1 2A9 cells might provide a model by which to investigate in more detail the interaction of pathogen-associated molecular patterns and monocytoid cell-surface-expressed pattern recognition receptors.

Structure and binding kinetics of three different human CD1d-alpha-galactosylceramide-specific T cell receptors

Invariant human TCR Valpha24-Jalpha18+/Vbeta11+ NKT cells (iNKT) are restricted by CD1d-alpha-glycosylceramides. We analyzed crystal structures and binding characteristics for an iNKT TCR plus two CD1d-alpha-GalCer-specific Vbeta11+ TCRs that use different TCR Valpha chains. The results were similar to those previously reported for MHC-peptide-specific TCRs, illustrating the versatility of the TCR platform. Docking TCR and CD1d-alpha-GalCer structures provided plausible insights into their interaction. The model supports a diagonal orientation of TCR on CD1d and suggests that complementarity determining region (CDR)3alpha, CDR3beta, and CDR1beta interact with ligands presented by CD1d, whereas CDR2beta binds to the CD1d alpha1 helix. This docking provides an explanation for the dominant usage of Vbeta11 and Vbeta8.2 chains by human and mouse iNKT cells, respectively, for recognition of CD1d-alpha-GalCer.

Isolated mediotegmental lesion causing narcolepsy and rapid eye movement sleep behaviour disorder: a case evidencing a common pathway in narcolepsy and rapid eye movement sleep behaviour disorder

Narcolepsy is usually an idiopathic disorder, often with a genetic predisposition. Symptomatic cases have been described repeatedly, often as a consequence of hypothalamic lesions. Conversely, REM (rapid eye movement) sleep behaviour disorder (RBD) is usually a secondary disorder, often due to degenerative brain stem disorders or narcolepsy. The case of a hitherto healthy man is presented, who simultaneously developed narcolepsy and RBD as the result of an acute focal inflammatory lesion in the dorsomedial pontine tegmentum in the presence of normal cerebrospinal fluid hypocretin-1 levels and in the absence of human lymphocyte antigen haplotypes typically associated with narcolepsy and RBD (DQB1*0602, DQB1*05). This first observation of symptomatic narcolepsy with RBD underlines the importance of the mediotegmental pontine area in the pathophysiology of both disorders, even in the absence of a detectable hypocretin deficiency and a genetic predisposition.

Lipopolysaccharides from atherosclerosis-associated bacteria antagonize TLR4, induce formation of TLR2/1/CD36 complexes in lipid rafts and trigger TLR2-induced inflammatory responses in human vascular endothelial cells

Infection with bacteria such as Chlamydia pneumonia, Helicobacter pylori or Porphyromonas gingivalis may be triggering the secretion of inflammatory cytokines that leads to atherogenesis. The mechanisms by which the innate immune recognition of these pathogens could lead to atherosclerosis remain unclear. In this study, using human vascular endothelial cells or HEK-293 cells engineered to express pattern-recognition receptors (PRRs), we set out to determine Toll-like receptors (TLRs) and functionally associated PRRs involved in the innate recognition of and response to lipopolysaccharide (LPS) from H. pylori or P. gingivalis. Using siRNA interference or recombinant expression of cooperating PRRs, we show that H. pylori and P. gingivalis LPS-induced cell activation is mediated through TLR2. Human vascular endothelial cell activation was found to be lipid raft-dependent and to require the formation of heterotypic receptor complexes comprising of TLR2, TLR1, CD36 and CD11b/CD18. In addition, we report that LPS from these bacterial strains are able to antagonize TLR4. This antagonistic activity of H. pylori or P. gingivalis LPS, as well as their TLR2 activation capability may be associated with their ability to contribute to atherosclerosis.

In vitro characterization of immune-related properties of human fetal bone cells for potential tissue engineering applications

We describe herein some immunological properties of human fetal bone cells recently tested for bone tissue-engineering applications. Adult mesenchymal stem cells (MSCs) and osteoblasts were included in the study for comparison. Surface markers involved in bone metabolism and immune recognition were analyzed using flow cytometry before and after differentiation or treatment with cytokines. Immunomodulatory properties were studied on activated peripheral blood mononuclear cells (PBMCs). The immuno-profile of fetal bone cells was further investigated at the gene expression level. Fetal bone cells and adult MSCs were positive for Stro-1, alkaline phosphatase, CD10, CD44, CD54, and beta2-microglobulin, but human leukocyte antigen (HLA)-I and CD80 were less present than on adult osteoblasts. All cells were negative for HLA-II. Treatment with recombinant human interferon gamma increased the presence of HLA-I in adult cells much more than in fetal cells. In the presence of activated PBMCs, fetal cells had antiproliferative effects, although with patterns not always comparable with those of adult MSCs and osteoblasts. Because of the immunological profile, and with their more-differentiated phenotype than of stem cells, fetal bone cells present an interesting potential for allogeneic cell source in tissue-engineering applications.

Mass spectrometry-based analytical tools for the molecular protein characterization of human plasma lipoproteins

Lipoproteins are a heterogeneous population of blood plasma particles composed of apolipoproteins and lipids. Lipoproteins transport exogenous and endogenous triglycerides and cholesterol from sites of absorption and formation to sites of storage and usage. Three major classes of lipoproteins are distinguished according to their density: high-density (HDL), low-density (LDL) and very low-density lipoproteins (VLDL). While HDLs contain mainly apolipoproteins of lower molecular weight, the two other classes contain apolipoprotein B and apolipoprotein (a) together with triglycerides and cholesterol. HDL concentrations were found to be inversely related to coronary heart disease and LDL/VLDL concentrations directly related. Although many studies have been published in this area, few have concentrated on the exact protein composition of lipoprotein particles. Lipoproteins were separated by density gradient ultracentrifugation into different subclasses. Native gel electrophoresis revealed different gel migration behaviour of the particles, with less dense particles having higher apparent hydrodynamic radii than denser particles. Apolipoprotein composition profiles were measured by matrix-assisted laser desorption/ionization-mass spectrometry on a macromizer instrument, equipped with the recently introduced cryodetector technology, and revealed differences in apolipoprotein composition between HDL subclasses. By combining these profiles with protein identifications from native and denaturing polyacrylamide gels by liquid chromatography-tandem mass spectrometry, we characterized comprehensively the exact protein composition of different lipoprotein particles. We concluded that the differential display of protein weight information acquired by macromizer mass spectrometry is an excellent tool for revealing structural variations of different lipoprotein particles, and hence the foundation is laid for the screening of cardiovascular disease risk factors associated with lipoproteins.