Drug-induced liver injury (DILI) has become a leading cause of severe liver disease in Western countries and therefore poses a major clinical and regulatory challenge. Whereas previously drug-specific pathways leading to initial injury of liver cells were the main focus of mechanistic research and classifications, current concepts see these as initial upstream events and appreciate that subsequent common downstream pathways and their attenuation by drugs and other environmental and genetic factors also have a profound impact on the risk of an individual patient to develop overt liver disease. This review summarizes current mechanistic concepts of DILI in a 3-step model that limits its principle mechanisms to three main ways of initial injury, i.e. direct cell stress, direct mitochondrial impairment, and specific immune reactions. Subsequently, initial injury initiates further downstream events, i.e. direct and death receptor-mediated pathways leading to mitochondrial permeability transition, which then results in apoptotic or necrotic cell death. For all mechanisms, mitochondria play a central role in events leading to apoptotic vs. necrotic cell death. New treatment targets consequently focus on interference with downstream pathways that mediate injury and therefore determine the ultimate outcome of DILI. Genome wide and targeted pharmacogenetic as well as metabonomic approaches are now used in order to reach the key goals of a better understanding of mechanisms in hepatotoxicity, and to develop new strategies for its prediction and treatment. However, the complexity of interactions between genetic and environmental risk factors is considerable, and DILI therefore currently remains unpredictable for most hepatotoxins. The liver may be considered as the most important organ in drug toxicity for two reasons: on the one hand it is functionally interposed between the site of absorption and the systemic circulation and is a major site of metabolism and elimination of foreign substances; but on the other hand these features also render it a preferred target for drug toxicity. Drug-induced liver injury (DILI) therefore poses a major clinical problem. DILI has become the leading cause of acute liver failure and transplantation in Western countries , 2. IntrOne of its most intriguing features is the fact that DILI can mimic all forms of acute or chronic liver disease. Another aspect of particular interest is the unpredictable idiosyncratic occurrence of DILI and the chemical heterogeneity of hepatotoxins causing idiosyncratic DILI. These aspects already indicate the wide range of targeted structures and cell types, the diversity of the involved mechanisms, and the importance of individual risk factors. During recent years the increasing insight into the complex processes and sequence of events involved in liver cell injury in general and DILI in particular had a substantial impact on current approaches to mechanistic hepatotoxicity and the subsequent definition of future research targets. Whereas previously drug-specific pathways leading to the initial injury of liver cells were the main focus of mechanistic research and classifications, current concepts see these as so-called initial “upstream” events and appreciate that subsequent common “downstream” pathways and their activation or inhibition by drugs and other environmental and genetic factors also have a profound impact on the risk of an individual patient to develop overt liver disease. In particular, these concepts emphasize the central role of mitochondria, of events leading to apoptotic vs. necrotic cell death, and of factors that balance injurious vs. protective and regenerative responses towards initial cell stress.This review will therefore focus on these new concepts and their implication for our understanding of mechanisms and risk factors including their complex interactions, as well as for the development of new treatments in DILI.or idiosyncratic hepatotoxins, and that dose or direct cell injury play no role in idiosyncratic hepatotoxicity. However, unpredictable rare severe DILI often develops on a background of frequent and dose-dependent mild increases in transaminases. Isoniazid is an example for such a hepatotoxin that is associated with mild intrinsic as well as severe idiosyncratic occurrence of DILI. On the other hand APAP, considered as a classic intrinsic hepatotoxin in doses above 10 to 15 g per day, may cause severe liver disease in therapeutic doses below 4 g per day in some patients [DILI is commonly classified into intrinsic vs. idiosyncratic hepatotoxicity, and the latter further into allergic vs. non-allergic. Intrinsic hepatotoxicity is regarded as dose-dependent and predictable above an approximate threshold dose, whereas idiosyncratic hepatotoxicity occurs without obvious dose-dependency and in an unpredictable fashion. Allergic idiosyncratic hepatotoxicity is characterized by the presence of typical symptoms and signs of adaptive immune reactions, including fever, skin reactions, eosinophilia, formation of autoantibodies, and a short latency time particularly after re-exposure. Other clinical classifications differentiate e.g. between hepatocellular, cholestatic or mixed liver enzyme patterns, histological criteria, acute vs. chronic onset, or severity. These classifications are useful in clinical practice because they describe typical clinical signatures of DILI for specific drugs, and they can also give useful hints regarding the involved mechanisms. Nevertheless, one must realize that these classifications are descriptive and based on clinical or histopathological criteria. They can therefore be misleading if mixed with mechanistic concepts and may indeed be the foundation of some classic paradigms that are now challenged by recent advances in mechanistic hepatotoxicity. For example, common misconceptions are that specific substances can be clearly classified as either intrinsic patients , 10, whepatients . And evepatients , 13, andpatients .A major challenge for mechanistic classifications is that DILI is not sufficiently characterized by the initial injury, but always involves several mechanisms, regulatory systems and risk factors with complex interactions; this also explains why for most hepatotoxins there are no experimental models available, a drug’s hepatotoxic potential is often not recognized before marketing, the exact contribution of different processes leading to DILI in humans remain largely unknown, and targeted treatments of DILI are not available except for APAP-induced hepatotoxicity. A possible approach to this dilemma is a general working model that integrates the principle initial specific mechanisms of toxic liver cell injury with recently gained knowledge of the unspecific complex regulation of injurious vs. protective processes involved in liver cell injury.1). According to this model DILI involves three subsequent main steps. It also includes details on the intrinsic and extrinsic pathways emphasizing the central role of mitochondria for the mechanisms leading to apoptosis vs. necrosis. Fig. (2) additionally demonstrates the important role of risk factors in DILI.Such a general mechanistic model is presented in Fig.  family that mediates oxidative phase-I drug metabolism. However, conjugative phase-II metabolism may also result in hepatotoxic metabolites, e.g. acyl glucuronides are well known to cause DILI , 16.ABCB11 gene), in which case the subsequent intracellular accumulation of its substrates may cause secondary toxic hepatocyte damage [Reactive metabolites can exert initial cell stress through a wide range of mechanisms including depletion of glutathione (GSH), or binding to enzymes, lipids, nucleic acids and other cell structures. Furthermore reactive metabolites or parent drugs may specifically inhibit other hepatocellular functions such as the apical  bile salt efflux pump , inhibit β-oxidation leading to steatosis , damage mitochondrial DNA or interfere with its replication, or directly cause mitochondrial permeability transition (MPT), i.e. opening of the “MPT pore” located in their inner membrane -21. Therful here -24.Specific immune responses involving cytotoxic T-cells with concomitant release of inflammatory cytokines can be evoked by reactive metabolites that covalently bind to proteins and are subsequently recognized as neo-antigens (hapten formation). Their subsequent major histocompatibility complex (MHC)-dependent presentation on antigen presenting cells may then activate formation of antibodies against haptens or autoantibodies against cell structures such as CYP450 enzymes , 26.In some instances initial injury also targets nonparenchymal liver cells. Examples include toxicity against biliary epithelial cells by reactive flucloxacillin metabolites , direct Different hepatotoxins are typically associated with specific patterns regarding their initial mechanism of injury. However, one should be aware of the fact that a single drug may concomitantly act through several of these initial mechanisms, and that for many drugs at least some of the involved mechanisms currently remain unknown.These initial specific injurious mechanisms can also be referred to as “upstream events”. In the next step they lead to subsequent rather unspecific “downstream events” that involve the innate immune system, which balances pro- and anti-inflammatory responses and therefore determines the further progress to severe injury or recovery.2.via a direct pathway initiated by severe cell stress (intrinsic pathway), or via an indirect death receptor-amplified pathway that is triggered by mild cell stress and/or specific immune reactions (extrinsic pathway) [Second, initial cell stress and/or initial specific immune reactions lead to MPT. If the initial mechanism does not directly target and impair mitochondrial function, this occurs in two principle ways, i.e. either pathway) .In the intrinsic pathway severe intracellular stress activates the endoplasmic reticulum pathway, lysosomal permeabilization, or c-jun N-terminal kinase (JNK) -34. SubsIn the extrinsic pathway an initial mild injury may be amplified if inflammatory responses due to mild stress and/or additional factors have modulated the innate immune system, where signaling cytokines that promote e.g. IL-12) or prevent  injury are usually well balanced. As a consequence, sensitized liver cells become more susceptible to lethal effects of tumor necrosis factor alpha (TNFα), Fas ligand (FasL) and interferon gamma (IFγ). This is particularly important if one considers that the liver as the central organ of detoxification is constantly exposed to cell stress that can activate TNFα and FasL. If the initial event is a specific immune reaction, MHC-dependent antigen presentation will activate the release of TNFα and FasL from Kupffer cells (hepatic macrophages) and cytotoxic T-cells. According to the danger hypothesis for autoimmune diseases, haptenization alone may be insufficient to trigger the development of frank allergic hepatotoxicity, which requires an additional stimulation, a so-called danger-signal  or preve. Therefo3.Third, impaired mitochondrial function and energy production leads to apoptotic or necrotic cell death. MPT allows massive influx of protons through the inner mitochondrial membrane, which stops mitochondrial ATP synthesis. Mitochondrial ATP depletion resulting from MPT  causes matrix expansion and mitochondrial outer membrane permeabilization and rupture with release of cytochrome c and other pro-apoptotic mitochondrial proteins from the intermembrane space into the cytosol , 37-39.In the case of apoptosis, cytochrome C then binds to a cytoplasmic scaffold (apaf-1) and pro-caspase 9, forming a complex called apoptosome, which activates signaling procaspase 9. This, however, is an active process that requires ATP and can therefore only start if MPT did not rapidly and simultaneously occur in all mitochondria. Only if some mitochondria are left intact and continue to synthesize ATP, activated pro-caspase 9 and possibly also other pro-apoptotic mitochondrial proteins subsequently activate executioner caspase 3. Caspase 3 will then cleave specific cell proteins and further activate pro-caspases 6,7 and 2, which have their own target proteins . These pNecrosis, in contrast, develops if the initial injury is so severe that MPT quickly occurs in all mitochondria, or if other mechanisms cause rapid severe mitochondrial ATP depletion, preventing the apoptotic pathway. This is typical for hepatotoxins that directly cause profound initial cell stress. However, in the absence of ATP also activation of the extrinsic pathway may lead to necrotic cell death. Cell swelling and lysis that follow severe disturbance of cell functions characterize necrosis. Necrotic cell lysis induces inflammatory responses including release of cytokines, which is important because these may amplify initial injury through a sensitization of surrounding hepatocytes and therefore cause further collateral damage.Finally it should be mentioned that the distinction between apoptosis and necrosis is not always clear-cut. Mixed phenomena have been described, and the same hepatotoxin may cause one or the other, or even the concomitant occurrence of both, depending on the circumstances including dose and preexisting vulnerability of hepatocytes , 42. ApoIn conclusion one can state that mitochondria stand in the center of life and death in hepatotoxicity: they can be targets of initial direct toxicity, MPT plays a key role in the further signaling of extrinsic and intrinsic pathways, and because mitochondria generate most of the cell’s ATP supply and are also the main intracellular source of oxygen and nitrogen free radicals, the extent of mitochondrial impairment finally determines whether hepatocytes die by apoptosis or necrosis .Additional factors and pathways modulate intrinsic and extrinsic pathways and the subsequent induction of apoptosis and necrosis. In addition to its DISC-mediated injurious effects, TNFα on the other hand also induces nuclear transcription factor kappa B (NF-κB)-dependent survival gene expression. NF-κB responsive genes inhibit JNK and promote upregulation of antioxidant genes . SimilarBecause of the liver’s extraordinary capacities to regenerate, full recovery is even possible if severe DILI has already caused extensive hepatocyte death . Differe2)). In particular it emphasizes the importance of several amplification pathways and provides a plausible explanation for the rare idiosyncratic occurrence of severe DILI that develops on a background of frequent mild liver injury. In the absence of cofactors such mild injury may be necessary though not sufficient to cause severe DILI. But in the presence of cofactors mild injury can be amplified and then tip the balance of injurious vs. protective processes toward death pathways. This triggering role of cofactors is also able to plausibly explain the very long latency time that has been reported in some cases of DILI [The 3-step model represents an attractive approach to hepatotoxicity for several reasons. An important feature is that it truly focuses on mechanisms and does not blend them with descriptive classifications, which eliminates some apparent contradictions that exist from other viewpoints. The model limits the principle mechanisms of DILI to three main ways of initial injury and on a vertical axis to three sequential main steps that lead to cell death. But at the same time it also integrates the crucial role of risk factors by appreciating the complex interactions of a large number of modulating factors and the fragile balance between injurious and protective factors that occur on all levels of the vertical axis .Several clinical risk factors for DILI such as high age, female gender and concomitant diseases or drugs have been described. Many of those may actually be associated with altered pharmacokinetics, which are an obvious risk factor whenever toxicity is dose-dependent and altered pharmacokinetics result in elevated concentrations of parent drugs and/or their toxic metabolites. However, from a current mechanistic point of view, it is most interesting to ask where risk factors exert their effect based on a multi-step model of hepatotoxicity that differentiates specific upstream vs. non-specific downstream mechanisms. In addition, risk factors can also be classified as either genetic or environmental. Such a view on risk factors superimposed on the 3-step model is shown in Fig.  may indeed be in accordance with this theoretical concept: several cases of acute liver failure during kava intake showed symptoms and signs of hypersensitivity indicating a specific immune reaction , 66; at via CYP450 2E1, leading to its reactive metabolite NAPQI that causes oxidative cell stress unless it is detoxified by GSH. Once GSH stores are depleted covalent binding to proteins occurs. This was long considered as the principle direct and sufficient hepatotoxic mechanism. Several risk factors that relate to APAP metabolism and NAPQI-induced oxidative stress were consequently identified: ethanol can induce but also competitively inhibit the formation of NAPQI via CYP450 2E1 [The complex role of different mechanisms and risk factors in hepatotoxicity can also be demonstrated by the example of APAP. APAP is a “classic” hepatotoxin for which an animal model is available, and also clinical APAP-induced DILI has been studied extensively. Toxification of APAP involves metabolism P450 2E1 , ethanolP450 2E1 , 70, inhP450 2E1 , and actP450 2E1 , 73. HowP450 2E1 , 74-78. P450 2E1 , 70, 79.P450 2E1 . FurtherP450 2E1 , and aftP450 2E1 .Studies of APAP-induced hepatotoxicity clearly demonstrate the role of many environmental risk factors and the role of toxic metabolites, oxidative stress and defense against it with a crucial role for GSH, and the contribution of both, initial toxic cell stress, as well as subsequent downstream events, for DILI. The relevance of other risk factors, particularly genetic factors, has been shown with hepatotoxins that typically cause idiosyncratic hepatotoxicity. Like environmental factors, genetic variations can principally affect all mechanisms involved in DILI, from drug metabolism over initial injury and mitochondrial impairment, to cell death and regeneration.Examples of genetic factors that affect drug metabolism or detoxification and have been associated with an increased risk of DILI are genetic variations in the GSH S-transferases GSTT1 and GSTM1, CYP450 2E1, N-acetyltransferase, as well as manganese superoxide dismutase (SOD2) -86. Intevia an impairment of mitochondrial function could be obesity and diabetes associated with steatosis and steatohepatitis [Cytokine polymorphisms associated with diclofenac-induced hepatotoxicity are another genetic risk factor of particular interest, because they relate to a downstream mechanism in DILI and could therefore also be relevant for other idiosyncratic hepatotoxins . The samepatitis -103.ABCB11) gene has been associated with an increased risk of DILI [Other genetic risk factors that recently gained much attention are determinants of hepatic transport proteins. Genetic  factors that affect basolateral  or apical  transport systems will determine exposure of hepatocytes not only to toxic drugs but also to toxic bile constituents and therefore represent both, risk factors as well as mechanisms for cholestatic DILI , 104-108 of DILI , 111, wh of DILI .Considering the important role of inflammatory mediators that can sensitize hepatocytes to injurious effects to cytokines (vide supra), it is not surprising that inflammation has not only been shown to enhance hepatotoxicity in an animal model , but alsvia its action on upstream events, such as the prevention of NAPQI-induced initial injury, but also via its effect on downstream processes that are not specific to APAP toxicity may justify trials that evaluate possible beneficial effects in the treatment of other forms of DILI including idiosyncratic hepatotoxicity.A better understanding of mechanisms of hepatotoxicity is a major future challenge in order to develop new solutions for the prevention and treatment of DILI. Recent important advances in the understanding of general hepatotoxic mechanisms are the foundation for further studies on mechanisms for specific hepatotoxic drugs. The increased appreciation of the crucial role and complex regulation of downstream events that modulate the critical switch from recovery to unopposed injury also implies that they represent promising new therapeutic targets. For example, specific JNK inhibitors are in clinical development for the treatment of APAP-induced hepatotoxicity, and they could also be protective against other hepatotoxins. Other potential downstream targets are the inhibition of any amplifying pathways, e.g. through an inhibition of injurious or a stimulation of protective cytokines and Bcl-2 proteins. Also specific caspase inhibitors that may prevent apoptosis are a possible approach. Given that apoptosis activates stellate cells leading to liver fibrosis, attenuation of apoptosis may even lead to the resolution of fibrosis. In fact, according to some reports, the beneficial effects of ursodeoxycholic acid are partly mediated through an inhibition of apoptosis by modulating MPT , 119. NeKnowledge of genetic and environmental contributors that determine actual idiosyncratic DILI in humans is necessary in order to focus mechanistic in-vitro and animal toxicology studies. Likewise, also prevention of DILI in clinical practice is inevitably linked to the identification of risk factors that predict severe DILI with sufficient sensitivity and specificity. Pharmacogenetic approaches such as case-control studies that consider interactions between genetic but also environmental factors may be able to reach that goal. However, the complexity of interactions being sufficient to cause severe DILI, the difficulties in the reliable identification of environmental risk factors, and also in the reliable causality assessment of clinical cases of DILI must not be underestimated. Therefore it is currently unknown whether this is a realistic scenario for clinically important hepatotoxins.Considering the importance of hepatotoxic metabolites, strategies aimed at identifying early markers of DILI that may precede ALT elevations are focusing on the use of metabonomics, e.g. high resolution NMR of urine samples. Patients that go to develop elevated ALT levels may have a different metabolome, and the spectra obtained before ALT values exceed the upper limit of normal might indicate a predictive metabolome signature. In fact, trials conducted with known idiosyncratic hepatotoxins such as isoniazid could be used to identify “Hy’s law” cases in serial collections , 121, alIdentification of risk factors for rare idiosyncratic hepatotoxicity requires special networks that contribute to data collection and subsequent identification of environmental as well as genetic risk factors for clinical cases of idiosyncratic DILI. With that in mind the DILI network project links several university medical centers that collect detailed information on cases of DILI including standardized causality assessments and samples for genetic analyses. The data collection focuses on idiosyncratic hepatotoxicity and particularly aims to identify upstream as well as downstream genetic determinants of hepatotoxicity , 123. FuFrom a clinical standpoint the important role of triggers and risk factors that may be required in idiosyncratic hepatotoxicity should stimulate clinicians to search for such factors when collecting, analyzing and reporting data of individual cases and case series of DILI. Furthermore, the increasing recognition of the hepatotoxic potential of herbal products should alert physicians as well as consumers. In fact it is well possible that a substantial proportion of cases of so-called “idiopathic” acute liver failure are indeed due to herbal hepatotoxicity. Every clinician should therefore include herbal products of any kind in the differential diagnosis and actively search for them when taking the history of a patient with liver disease of unclear origin. From a regulatory point of view the issue of herbal hepatotoxicity also raises the question whether not only herbal drugs but also herbal dietary supplements may generally require a stricter regulation of their distribution and use -6. For d


Several thousand human genome epidemiology association studies are published every year investigating the relationship between common genetic variants and diverse phenotypes. Transparent reporting of study methods and results allows readers to better assess the validity of study findings. Here, we document reporting practices of human genome epidemiology studies.Articles were randomly selected from a continuously updated database of human genome epidemiology association studies to be representative of genetic epidemiology literature. The main analysis evaluated 315 articles published in 2001–2003. For a comparative update, we evaluated 28 more recent articles published in 2006, focusing on issues that were poorly reported in 2001–2003.During both time periods, most studies comprised relatively small study populations and examined one or more genetic variants within a single gene. Articles were inconsistent in reporting the data needed to assess selection bias and the methods used to minimize misclassification  or to identify population stratification. Statistical power, the use of unrelated study participants, and the use of replicate samples were reported more often in articles published during 2006 when compared with the earlier sample.We conclude that many items needed to assess error and bias in human genome epidemiology association studies are not consistently reported. Although some improvements were seen over time, reporting guidelines and online supplemental material may help enhance the transparency of this literature. Human genome epidemiology (HuGE) is a rapidly emerging scientific field that examines the influence of genomic variation on human health -4. AlthoIn 2001, the Human Genome Epidemiology Network (HuGENet) established the HuGE Published Literature database (HuGE Pub Lit), a continually updated, searchable, online database of population-based, genetic epidemiology articles . RelevanOur evaluation was designed in 2004 and data collection and analyses were conducted in 2004–2007. For the main analysis, we drew a five percent simple random sample (SRS) of articles that were returned by the query described above, published from 2001 to 2003, and curated in HuGE Pub Lit before May 30th, 2004  to yield a dataset of 406 articles. To provide an updated description of reporting practices and to assess improvements in reporting, we randomly selected (SRS) 40 articles that were published during 2006 from articles that were returned by our database query, added to PubMed in 2006, and curated in HuGE Pub Lit before May 18, 2007 . After each article was read, 91 from 2001–2003 and 12 from 2006 were excluded from the analysis for the following reasons: not written in English , population screening studies , clinical trials or pharmacogenomic studies , not full-length articles  , failed to fulfill the inclusion criteria for HuGE Pub Lit [Data were abstracted from each original publication in duplicate by two independent data extractors. All discrepancies between the independent extractors were discussed and a consensus was reached.For the 2001–2003 articles, a standardized abstraction form was developed and piloted for 10 articles; the form was revised according to the results of this pilot study to ensure that the definitions for the collected items were clear and unambiguous. Items on this final form were designed to collect information on the reporting of study design, genotyping method, population stratification, analytical methods , and study inferences. In addition, the final form accommodated different observational study designs, multiple groups of study participants, and the consideration of more than one postulated genetic risk factor as well as additional environmental factors. Articles were coded as potentially misclassifying the disease or environmental exposure status of study participants when the article did not explicitly state that these factors were directly measured for all participants in the study population. When multiple groups of study participants were reported in a study, we recorded the sample size of the largest group for cohort and cross-sectional studies and the largest case and control groups for case-control studies. Items were collected separately for case and control groups for case-control studies and for all study participants regardless of disease status for cohort and cross-sectional studies. For the purpose of this analysis, data collected for case and control groups were combined so that statistics could be calculated for all study participants. Information  was considered as given for all study participants only if it was provided for all case and control groups. Additionally, for case-control studies, we recorded whether cases and controls were described as drawn from the same population according to one or more of the following definitions: 1) geographic region, 2) clinical population, 3) general population , or whether information on the choice of suitable controls was missing or incomplete.Fourteen items were assessed in the HuGE articles published in 2006. These included the number of study participants, genes, polymorphisms, and environmental factors assessed in gene-environment interactions. In addition, we selected ten items that were applicable to all study designs and that had been reported in fewer than 50% of the articles published in 2001–2003.The data analysis was conducted using SAS 9.13 . Counts and percents were calculated for the items abstracted from the articles. Comparisons of articles published in 2006 vs. those published in 2001–2003 used the Mann-Whitney U test for continuous variables and Fisher's exact test for binary variables.The 315 articles selected for analysis were published in 194 journals and reported on the findings of 227 case-control, 32 cohort, and 56 cross-sectional studies. In addition to population-based studies, three articles also described family-based analyses. Data pertaining exclusively to these family-based analyses were not included in this report. As shown in Table Seven percent of studies reported that the genotyping results were validated with the use of replicate samples, and an additional 9.8 percent reported that a different method of validation was used Table . A blindAs shown in Table Approximately one-half of the articles stated that they examined whether the study populations were in Hardy-Weinberg equilibrium; of these, 6.6 percent reported that the genotype frequencies deviated from those expected under Hardy-Weinberg equilibrium Table . SummaryOverall, less than 40 percent of the articles discussed the public health, medical, or clinical implications of their findings. Less than one in six articles claimed to be the first to analyze a particular association. For the articles that did not make this claim, 8.6 percent clearly made reference to the first study on the issue. Six percent of articles clearly referenced a systematic review, and 1.9 percent referenced a non-systematic review.Nearly two-thirds of the articles (N = 201) investigated multiple genetic variants, often in more than one gene Table . These sGene-environment interactions were discussed in 15.2 percent (n = 48) of the articles Table . Among tThe number of study participants, genes and polymorphisms analyzed, and environmental factors examined in gene-environment interactions were similar for the two time periods Table . ArticleThree of the ten items that were reported in fewer than 50 percent of the articles from 2001–2003 were reported significantly more often in the 2006 articles Table . StudiesMany published claims of gene-disease association have not been replicated when studied in independent samples ,6. SuspeRepresentative of the literature in this field, most of the studies in our samples were small: only about 10 percent of the studies reported sample sizes that exceeded 1,000. Several meta-analyses have found significant differences between the results of small and large genetic association studies ,19. GrowOur ability to assess the potential for selection biases was severely limited in many of the studies we examined. Although most studies provided some qualitative descriptions of the study participants (such as origin and enrollment criteria), reporting was sporadic for even simple descriptors, such as age and sex. Potentially important details, such as the number of exclusions or the number of samples that could not be genotyped, were often omitted.Misclassification can severely limit study power and bias the results ,23,25-27Population stratification may occur when study participants are selected from subpopulations with a different prevalence of the phenotypes and genotypes ,30,32,33As a result of the selective reporting of significant results from multiple analyses and publication bias, the extent of type I error in the published literature may be great ,30,40,41The use of common reporting standards could increase the transparency of research methodology, thus helping to identify selective reporting and sources of bias and confounding while allowing for a more complete synthesis of data across consortia or in meta-analyses ,10,42. TBy publishing supplementary information online, journals increasingly provide authors the opportunity to present their study methods and results in greater detail than is permitted in print ,46. ReceIn summary, our results provide evidence that many details needed to assess the validity of study findings are not consistently reported in human genome epidemiology studies, though some improvement has been seen recently. The use of standard reporting guidelines and online supplements could help readers to better judge the scientific evidence. As large-scale genotyping platforms are rapidly introduced in human genome epidemiology, the importance of transparent reporting of the background, epidemiological methods, and population characteristics cannot be understated given the challenge of assessing, interpreting, and discussing ever-greater amounts of data.The authors declare that they have no competing interests.AY assisted in data collection, performed the analysis, and drafted the manuscript. EE, FKK, and NAP assisted in data collection and the revision of the manuscript. MC identified and indexed relevant articles for the HuGE Published Literature database. MCW and BKL created the standardized abstraction form and assisted in the sampling process and data collection. WY curates the HuGE Published Literature and assisted in the sampling process. MG, JPAI, and MJK designed the study, oversaw the project, and revised the manuscript. All authors read and approved the final manuscript.The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention.The pre-publication history for this paper can be accessed here:


Biological evidence suggests that inflammation might induce type 2 diabetes (T2D), and epidemiological studies have shown an association between higher white blood cell count (WBC) and T2D. However, the association has not been systematically investigated.Studies were identified through computer-based and manual searches. Previously unreported studies were sought through correspondence. 20 studies were identified . Estimates of the association of WBC with T2D were combined using random effects meta-analysis; sources of heterogeneity as well as presence of publication bias were explored.−18). Substantial heterogeneity was present (I2 = 83%). For granulocytes the RR was 1.38 , for lymphocytes 1.26 , and for monocytes 0.93  comparing top to bottom tertile. In cross-sectional studies, RR was 1.74 , while in cohort studies it was 1.48 . We assessed the impact of confounding in EPIC-Norfolk study and found that the age and sex adjusted HR of 2.19  was attenuated to 1.82  after further accounting for smoking, T2D family history, physical activity, education, BMI and waist circumference.The combined relative risk (RR) comparing the top to bottom tertile of the WBC count was 1.61 (95% CI: 1.45; 1.79, p = 1.5*10A raised WBC is associated with higher risk of T2D. The presence of publication bias and failure to control for all potential confounders in all studies means the observed association is likely an overestimate. Chronic inflammation, characterized by the increased production of cytokines and acute-phase reactants and activation of inflammatory signalling networks Evidence from epidemiological studies suggests an association between total peripheral white blood cell (WBC) or leukocyte count, a non-specific marker of inflammation, and diabetes riskIn the present study we systematically review and meta-analyse existing studies of the association between differential WBC count and T2D, including previously unpublished data from 5,021 cases and 43,508 non-cases  obtained through correspondence with investigators. We also explore the potential roles of reverse causality, publication bias and confounding.SM version 4.7 (©Thomson Reuters 2009) and Embase (© 2009 Elsevier B.V.), initially without limits with regard to publication date or language. Last searches were conducted in April 2010. Two authors  independently reviewed all identified titles , and subsequently abstracts (n = 136) and full articles  count and T2D. The search terms included (“leukocyte” OR “leucocyte “OR “white blood”) combined (AND) with diabetes (diabetes” OR “glucose” OR “metabolic syndrome” OR “hyperglycaemia” OR “hyperglycemia”). We searched Pubmed 2.0  entering each search term as a MeSH, ISI Web of Knowledgearticles . We inclarticles . DiscrepWe identified studies that appeared during the literature review and did not report on the association between WBC and T2D but had potentially collected pertinent data, as evident from their study description. Corresponding authors of these studies (n = 19) were contacted by electronic and regular mail (two electronic reminders) and invited to submit data using a standardized data extraction sheet and uniform analysis plan. We requested odds ratios, relative risks or hazard ratios for the association between WBC and its sub-fractions where available, comparing top to bottom tertile of each measure, adjusting for age, sex, smoking, BMI and waist circumference and using the WHO definition of diabetesWe additionally included unpublished results from the EPIC Norfolk study, described in more detail below, according to the protocol used for obtaining unpublished evidence from other investigators.2 statistic, which represents the proportion of variation in the effect sizes that is attributable to genuine differences across studies rather than to random error For the purposes of the meta-analysis, we considered all of odds ratios, risk ratios and hazard ratios as estimates of the relative risk. These relative risks were combined across studies using random effects meta-analysis. Heterogeneity was assessed using the IIn addition to including results from the European Prospective Investigation of Cancer in Norfolk, UK (EPIC-Norfolk) as part of the overall meta-analysis according to the standard protocol described above, we used prospective data from this cohort to investigate the effect of adjusting for a range of potential confounding factors not consistently available across other studies.EPIC-Norfolk is a population-based cohort study, which has previously been described in detail We excluded participants with a history of stroke, myocardial infarction, or cancer  or prevalent or unconfirmed diabetes at baseline (n = 688).A detailed self-completed health and lifestyle questionnaire was completed at baseline, including questions on family history of diabetes, prescribed medications, occupational social class, smoking status, educational level, and physical activity assessed by a four point indexBiochemical assays were carried out on samples drawn with participants in the non-fasted state. Blood samples for WBC measurement were stored overnight at room temperature and were collected each morning and transported to the EPIC-Norfolk laboratory in Attleborough (UK). The samples were analysed in a random order using impedance counting technique with an MD18 haematology analyser . Quality controls were carried out daily. In addition, the Haematology Department of Addenbrooke's Hospital included the EPIC Laboratory in a monthly quality control scheme. The WBC count coefficient of variation for the period of study was ≤3.0%. The standard deviation values for the differential granulocyte, lymphocyte and monocyte percentages were less than or equal to 1.5, 1.5, and 3.0, respectively Ascertainment of incident cases of type 2 diabetes involved review of multiple sources of evidence including self-report  during follow-up, linkage to primary and secondary care registers, hospital admissions and mortality dataCox proportional hazards regression was used to estimate hazard ratios for the incidence of diabetes by tertiles of WBC, granulocyte, lymphocyte, and monocyte count, using the lowest tertile as the reference category. Results from age and sex adjusted models were compared to those additionally adjusting for smoking status , waist circumference (continuous), BMI (continuous), educational level (below ‘A’ level vs. ‘A’ level and above where ‘A’ level is school education to age 18 years), a positive family history of diabetes, and physical activity level (4 categories ranging from sedentary to active).Analyses were restricted to participants with full information on total WBC, granulocyte, lymphocyte or monocyte count . We further excluded 158 participants without complete information on covariates or exclusion criteria.15,550 participants (499 incident diabetes cases) remained in the analyses. We calculated a Health Behaviours Score (HBS), including information on physical activity, alcohol intake, plasma vitamin C (a biomarker of fruit and vegetable intake) and smoking, as proposed by Khaw et al p values were based on 2-sided tests.All statistical analyses were performed using Stata/SE 10.0 . All Initial searches identified 12,705 articles and abstracts . After e−18). (−4) for granulocytes, 1.26  for lymphocytes, and 0.93  for monocytes, comparing the top to bottom tertile of the distribution of each measure (The combined relative risk (RR) comparing the top to bottom tertile of the total WBC count distribution was 1.61 . . The com measure .2 = 83%, p<0.001). I2 was slightly smaller but still statistically significant among prospective cohort studies . Sensitivity analyses were used to identify potential sources of heterogeneity between studies. Stratification by study design showed a combined RR for WBC count of 1.73  for prevalent  and 1.48 .for incident  T2D. Subgroup analyses (−25). and no significant difference comparing published to unpublished evidence included in this report  versus RR 1.72, 95% CI: 1.48; 2.00, p = 1.1*10−12). The combined RR was 1.66  versus 1.52  when comparing studies including more than 70% versus less than 70% European descent participants respectively.For total WBC, there was a high degree of heterogeneity between the 20 studies , number of cases, number of participants, source of data (investigator provided versus published data), and type of study  in a meta-regression model resulted in a decrease in the value of IA funnel plot indicated the presence of publication bias in these studies . SignifiSince limited information was available from the published studies, we were unable to assess the impact of confounding based on results from published studies. Instead, we assessed the impact of confounding in the EPIC-Norfolk cohort as described below.We assessed the impact of confounding in EPIC-Norfolk study participants with detailed covariate information . The following were associated with lower WBC counts: female sex, lower BMI, lower waist circumference, lower age, never smoking .a priori to include T2D family history, physical activity and educational level in our models since they have been previously associated with peripheral WBC Baseline HbA1c data were available for only a subset of EPIC-Norfolk (n = 9558). 9392 individuals had a baseline HbA1c of less than 6.5%, 166 of whom developed incident diabetes. The age and sex adjusted HR for WBC count of 2.07 (95% CI: 1.41 3.04) was reduced to 1.87  after further accounting for smoking status, T2D family history, physical activity, education, BMI and waist circumference, comparing the top and bottom tertiles of the total WBC distribution.9/L) results were similar to when including participants irrespective of the normal range .The present meta-analysis includes evidence about the association between WBC count and T2D from 20 cross-sectional and prospective cohort observational studies, comprising a total of 8,647 T2D cases and 85,040 non cases. Total WBC count was significantly associated with T2D, after adjustment for age, sex, smoking, BMI, waist circumference. Total granulocyte (and subset neutrophil) as well as lymphocyte but not monocyte count were also significantly associated with T2D, after adjustment for age, sex, smoking, BMI, waist circumference The findings were similar for incident T2D in the EPIC-Norfolk cohort analysis, where further adjustment for measured confounders showed the potential for residual confounding.The available biological data have strongly suggested that T2D is an inflammatory disease We did not find an increased risk of incident T2D for participants belonging to the higher monocyte tertile. Our findings agree with the previously reported results of the ARIC study Possible mechanisms that could link inflammation and diabetes include interruptions of the insulin signalling in the liver by inflammatory molecules like IL-6 Measurement error could affect our assessment of exposures, outcomes and confounding factors. Differences in the methods used to measure WBC counts as well as different performance of the same assays at different time periods might have contributed to error in longitudinal studies. In general, such measurement error would attenuate the measure of association if non-differential with regard to the case status. Some cases of type 1 diabetes (T1D) might have been inadvertently included; however, such misclassification is likely to have a minimal impact on effect sizes, given that T1D constitutes only 5–10% of all diabetes cases. Also, given the ascertainment methods used, misclassification is unlikely in prospective cohorts; all except three  included incident cases occurring after age 35 years. The majority of cross-sectional studies asked about the type of diabetes when assessing prevalent diabetes or excluded participants using insulin within the first year of diagnosis.It is possible that hyperglycaemia itself has an impact on WBC levels. In people with diabetes, WBC levels are lowered by treatment with rosiglitazoneResults of this study are based on a systematic and comprehensive literature review, including data from both prospective cohorts and cross-sectional studies, and previously unpublished data for a large number of participants. Incomplete retrieval of available results and studies is possible, since the studies included were all published in the English language. However, additional searches showed that no article in a language other that English that fitted the inclusion criteria could be identified. To address variations in design and diabetes ascertainment we asked corresponding authors to report their cases following the same diabetes definition (WHO 1999) and standardize their effect estimates for the same set of covariates . Despite these attempts of standardisation, heterogeneity between investigator sought studies was not totally eliminated. The funnel plot does indicate the potential for publication bias despite our efforts to obtain data that were not published. This could have been due to the fact that our search strategy, although exhaustive, was more likely to identify studies with reported results on WBC and T2D associations. Sensitivity analyses revealed a significantly lower RR in larger (≥500 cases) compared to smaller studies but no significant difference comparing published to unpublished evidence.In summary, these results suggest that WBC is positively associated with the risk of T2D. However, the presence of publication bias and failure to control for all potential confounders in all studies suggests that the observed association may be an overestimate of the truth. We cannot exclude the possibility of reverse causality or residual confounding. Approaches such as the use of genetic determinants of WBC as instrumental variables may be useful to deal with these as yet unresolved issues.Table S1Summary of the studies of association between WBC and T2D included in the meta-analysis(0.09 MB RTF)Click here for additional data file.PRISMA Checklist S1PRISMA Checklist of items to include when reporting a systematic review or meta-analysis (diagnostic review consisting of cohort studies)(0.07 MB DOC)Click here for additional data file.


GIPR SNPs  in German families with at least one obese index patient, two case-control studies and two cross-sectional population-based studies.Gastric inhibitory polypeptide (GIP) is postulated to be involved in type 2 diabetes mellitus and obesity. It exerts its function through its receptor, GIPR. We genotyped three Genotyping was performed by MALDI-TOF, ARMS-PCR and RFLP. The family-study: 761 German families with at least one extremely obese child or adolescent  and both parents . Case-control study: (a) German obese children (n = 333) and (b) obese adults (n = 987) in comparison to 588 adult lean controls. The two cross-sectional population-based studies: KORA  and SHIP .TDT-Test = 0.0089). In the combined case-control sample, we estimated an odd ratio of 1.54  for homozygotes of the rs2302382 A-allele compared to individuals with no A-allele. A similar trend was found in KORA where the rs2302382 A-allele led to an increase of 0.12 BMI units (p = 0.136). In SHIP, however, the A-allele of rs2302382 was estimated to contribute an average decrease of 0.27 BMI units .We detected over-transmission of the A-allele of rs2302382 in the German families (pGIPR variants for obesity. However, additional studies are warranted in light of the conflicting results obtained in one of the two population-based studies.Our data suggest a potential relevance of  Obesity is a serious health problem in both developed and in developing countries. It results from an interplay of environmental and genetic factors that mediate energy intake and expenditure. From twin studies it is estimated that up to 80% of the variance in body mass index (BMI) might be genetically determined -4. Theseob/ob) rodents . -48. GIPRGIPR variants potentially involved in obesity. Most likely variations in the putative regulatory region of the gene (e.g. rs2302382) are the most promising candidates for independent validations in case-control samples or in selected family samples as well as ultimately in functional studies. If our findings indeed are truly positive, this study also supports the observation [In conclusion, our data provide a first step towards identification of ervation ,49 that The authors declare that they have no competing interests.CIGV participated of the study design, perform the genotyping and drafted the manuscript; AS participated of the study design, performed statistical analysis and drafted the manuscript. TTN performed statistical analysis. AB and HG performed genotyping. GB, HJW, DR, HV, TR, WR, TI, and HEW participated of the study design; JH and AH conceived the study, participated in its design and coordination and drafted the manuscript. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:


The objective of this study is to conduct a systematic review of studies reporting the frequency of neurocysticercosis (NCC) worldwide.PubMed, Commonwealth Agricultural Bureau (CAB) abstracts and 23 international databases were systematically searched for articles published from January 1, 1990 to June 1, 2008. Articles were evaluated for inclusion by at least two researchers focusing on study design and methods. Data were extracted independently using standardized forms. A random-effects binomial model was used to estimate the proportion of NCC among people with epilepsy (PWE). Overall, 565 articles were retrieved and 290 (51%) selected for further analysis. After a second analytic phase, only 4.5% of articles, all of which used neuroimaging for the diagnosis of NCC, were reviewed. Only two studies, both from the US, estimated an incidence rate of NCC using hospital discharge data. The prevalence of NCC in a random sample of village residents was reported from one study where 9.1% of the population harboured brain lesions of NCC. The proportion of NCC among different study populations varied widely. However, the proportion of NCC in PWE was a lot more consistent. The pooled estimate for this population was 29.0% (95%CI: 22.9%–35.5%). These results were not sensitive to the inclusion or exclusion of any particular study.Only one study has estimated the prevalence of NCC in a random sample of all residents. Hence, the prevalence of NCC worldwide remains unknown. However, the pooled estimate for the proportion of NCC among PWE was very robust and could be used, in conjunction with estimates of the prevalence and incidence of epilepsy, to estimate this component of the burden of NCC in endemic areas. The previously recommended guidelines for the diagnostic process and for declaring NCC an international reportable disease would improve the knowledge on the global frequency of NCC. Taenia solium, which infects humans and pigs. There have been increasing case reports and epidemiological studies on this disease, but its global frequency has never been determined, partly due to the fact that blood tests are not very good for the diagnosis of NCC. We present here a systematic review of the literature on the frequency of NCC diagnosed with neuroimaging worldwide. Overall, 565 articles were retrieved and 290 (51%) selected for further review. Of those, only 26 had information valid enough to estimate the frequency of NCC in various populations. Only one study estimated the prevalence of NCC in the general population. The most striking finding was that the proportion of NCC among persons with epilepsy was very consistent and estimated at 29.6% (95%CI: 23.5%–36.1%) from 12 studies conducted in Latin America, Sub-Saharan Africa and Southeast Asia. A reinforcement of the suggested universal guidelines for the diagnostic process, declaring NCC an international reportable disease and standardizing procedures for data collection could improve our understanding of the frequency of NCC worldwide and hence its global burden.Neurocysticercosis (NCC) is a parasitic infection of the brain caused by the tapeworm  Scientific evidence is instrumental to improving global public health, as health policies should be based on accurate and meaningful data. In early 1990, the first Global Burden of Diseases (GBD) study, commissioned by the World Bank, was launched to develop a method to estimate and compare the burden of 107 diseases and injuries around the world. A standardized indicator, the “Disability Adjusted Life Years” (DALY) method, was developed for this purpose Taenia solium cysticercosis, has never been estimated.Neglected tropical diseases are a public health issue worldwide and especially in developing countries, where risk factors for their transmission are common T. solium is illustrated in T. solium migrate to the brain, has been reported as the most frequent helminthic infection of the central nervous system (CNS) T. solium remains largely unknown The lifecycle of The primary objective of this study was to conduct a systematic review of the literature to gather data on estimates of NCC frequency between 1990 and 2008 by age group and region. A secondary aim was to estimate the proportion of people living with epilepsy (PWE) who have NCC. This study was commissioned by the World Health Organization's Foodborne Disease Burden Epidemiology Reference Group (FERG). The FERG is the World Health Organization's advisory body to estimate the global burden of foodborne diseases.A systematic search of the literature was conducted to gather valid information on the frequency  of NCC between 1990 and 2008.Taenia solium”, “taeniasis”, “cysticercosis”, and “neurocysticercosis”. Only one copy of duplicated documents was kept for analysis. Studies were selected that included original epidemiological data on NCC frequency. Books and conference abstracts were excluded because they were unlikely to present original data or to have sufficient details on methods to judge the validity of the study. Dissertations, theses, and memoirs were included. Moreover, due to the under-representation of publications from Sub-Saharan Africa, three unpublished studies (at that time) were reviewed in addition to one paper published online in November 2008 The search strategy was conducted in three phases. In phase I, PubMed, Commonwealth Agricultural Bureau (CAB) Abstracts, and 23 international databases  were scrAnother component of this project was to assess the proportion of sequelae associated with NCC (details reported elsewhere). This search led to the finding of four additional studies which described the proportion of NCC among people with epilepsy and seizures a priori. In phase I, all documents retrieved were screened based on title and abstract. The exclusion criteria for phase I were: 1) wrong agent ; 2) animal data only; 3) no original data on the frequency of NCC; 4) case series with less than 20 participants; 5) review article without original data; and 6) editorials or letters to the editors without original data. Documents without abstracts were included in the next phase. All eligible documents after phase I were obtained in full. Each full document was read and reviewed by at least two investigators and subjected to the two subsequent phases of review. Phase II and III corresponded to a qualitative and quantitative appraisal of the information, respectively. The exclusion criteria for phase II included all criteria used in phase I in addition to: 1) no neuro-imaging (CT-scans or MRI) or autopsies used for the diagnosis of NCC; 2) high potential for selection bias ; or 3) all available data from before 1990 or after 2008 (except for sub-Saharan Africa where two studies published in 2009 were included). The data from documents included after phase II were extracted in phase III.The inclusion and exclusion criteria were defined During phase I, the full reference of each article, the country where the study was conducted, the decision on inclusion for phase II, the reason for exclusion (if applicable) and the language of the document were entered into an Excel spreadsheet . Data were extracted independently by two authors and a third author checked a random sample of 10% of all the entries. Any differences were resolved by discussion until agreement was reached.The quality assessment (phase II) and data extraction (phase III) for each document were carried out by two reviewers (except for Chinese articles) one of whom was a senior researcher (HC or CB). All documents published in French, Portuguese or Spanish were reviewed by those who could read those languages (PN and HC). All documents published in Chinese were reviewed by a Chinese collaborator (Y-JQ). No articles in German, Romanian or Italian were identified. In addition, a random sample of 10% of all English documents was reviewed by all reviewers. Disagreements were resolved through discussion in a meeting with all the reviewers. Data were entered into standardized electronic forms of the data extraction tool which was developed in Access  specifically for this review (available from the authors on request). Data were collected on the study characteristics , participant selection, case definitions, and ascertainment of outcome. For documents that could potentially be included in the review but had incomplete or missing data, the authors were contacted at least twice for clarification and/or additional information.The number of documents included in each phase of the systematic review was first plotted geographically using ArcView GIS software . Results from studies reporting separately the overall prevalence or incidence rate of NCC in the population are reported for each study.Measures of frequency: We report the proportion of patients with NCC in studies where a specific group of patients seeking care in a hospital or clinic were included. This is obtained by dividing the number of people with lesions of NCC by the number of people included in the study . We report the proportion of NCC among PWE using the same approach. The term prevalence is applied to the proportion of people with lesions of NCC at the CT-scan in studies where the general population or community residents without epilepsy were randomly sampled. The 95% Confidence Intervals (95% CI) of those proportions was estimating using the Clopper-Pearson exact interval provided in the Stata  software. The annual incidence rate of hospitalized cases of NCC was calculated by dividing the number of cases discharged with an International Classification of Disease (ICD) code for cysticercosis by the person-years living in the U.S. state where the study was conducted (based on census data). The prevalence of NCC-associated epilepsy was estimated by multiplying the prevalence of epilepsy by the proportion of NCC conditional on having epilepsy in community-based studies where both these estimates were available. This prevalence was estimated using WinBugs 1.4.3© and represents the proportion of people in a community estimated to have both epilepsy and NCC.www.R-project.org). We used the inverse variance method 2 index was used to summarize the total variability in proportion estimates due to between-study variation A random-effects model was used to summarize the data on the proportion of NCC among PWE using R META package  and the METFOR package  from R statistical software  We found only one study from a developed country (United States) reporting the percentage of NCC among people with seizures attending emergency rooms Four studies reported the proportion of NCC among people who were autopsied The estimated prevalence of people with NCC-associated epilepsy in community-based studies ranged from 0.1% in India 2 statistic indicated that 92.5% (95% CI: 88.1%–94.6%) of the total variability in the percentage values was due to between-study variation. No study had a significant impact on the result and the between-study variability was not explained by a single study.The proportion of NCC among PWE was remarkably homogeneous across studies conducted in children and adults . The lowWe ran the model excluding the study from Peru where some single seizures cases were included We also ran random-effect models stratified by the target population . The estimates were 31.7% (95% CI: 25.6%–38.2%) for community-based and 25.4% (95%CI: 16.3%–35.7%) for clinical-based studies. A mixed-effects regression model was used to determine whether the study setting  significantly influenced the percentage value. The estimated percentage among clinic-based studies is expected to be 5.9% lower (absolute difference) than that for community-based studies (95% CI: 17.2% lower to 5.5% higher), which is not statistically significant (p = 0.31).2 statistic suggested that 43.1% (95%: CI 0%–76.1%) of the total variability in the values was due to between-study variation. In the analysis of adults aged 20 to 54 years, the estimate of the percentage of NCC among PWE was more variable, ranging from 14.2% (95% CI: 8.6%–21.5%) in an outpatient clinic in Tanzania 2, the percentage of total variation across studies due to between-study heterogeneity, was 74.8% (95% CI: 46.6%–88.2%).Only five of the 12 studies had sufficient information to obtain estimates stratified by two broad and consistent age groups. In patients less than 20 years of age, the percentage of NCC among PWE ranged from 11.1% (95% CI: 2.4%–29.2%) in a community-based study in Burkina Faso  to 45.2% (95% CI: 27.3%–64.0%) in the Eastern Cape Province of South Africa This is the first study to systematically collect data on the frequency of NCC worldwide Our results demonstrate that there is not sufficient evidence at this time to estimate the prevalence of NCC globally. However, our study is the first meta-analysis to summarize the proportion of NCC among PWE, and suggest that nearly one-third of PWE living in endemic communities show lesions of NCC in their brain. Our study is particularly comprehensive because it used 25 different databases to identify published information and included documents published in four different languages.After a systematic review, only studies that are likely to be valid and are similar enough in their methods and definitions should be reported T. solium infections The second most common reason for exclusion of documents, after an absence of measurement of NCC frequency, was the lack of neuro-imaging for the diagnosis of NCC. Underdeveloped countries where sanitation and proper pig management methods are lacking are often endemic for The third most common reason for exclusion of a study was the use of flawed methodology. Studies were excluded that did not mention when and where they were conducted as were primary studies with a high potential of selection bias, since the strength of a systematic review depends on the quality of the primary studies that are included Target and study populations were very different across the documents we analyzed. For example, studies were conducted on patients with Japanese encephalitis (JE) The only cross-sectional study among a random sample of the population found a prevalence of NCC lesions, all calcified, of 9.1% T. solium in their community. Given that the incubation of NCC is unknown, those participants testing negative to the EITB may have been exposed a long time ago but have seen their immunity wane with time. Those cases may also have never developed antibodies to the brain infection. Such studies are unlikely to be repeated in the future. Due to potential adverse effects of the contrast materials used for CT or MRI, it is usually considered unethical to perform neuro-imaging in apparently healthy individuals. This limits our ability to truly measure the burden of NCC as some people may be asymptomatic for extended periods of time In another study, people who were sero-positive to the EITB and an age-sex-village matched sample of sero-negative people underwent a CT-scan of the brain We identified four studies conducted in autopsied patients from large hospitals in Brazil and Mexico The prevalence of people with NCC-associated epilepsy in community-based studies varied considerably. This prevalence is the product of the prevalence of PWE in the community and the proportion of NCC among PWE. Since the prevalence of NCC among PWE tends to be similar across studies (about one-third), the prevalence of epilepsy in communities is the parameter that contributes the most to the observed variability across communities. There are diverse, competing etiologies for epilepsy across countries and in addition to NCC include malaria, paragonomiasis, toxocariasis, and other parasites of the brain The results from our meta-analysis show that epilepsy is consistently associated with NCC in over one quarter of patients residing in endemic regions. This result was very robust, regardless of the type of epilepsy, if single epileptic seizures were included or not, and where and among whom the study was conducted. In an older study of 100 consecutive patients with epilepsy, Medina et al. found a prevalence of 50% for NCC It is difficult to determine if our finding of the proportion of NCC lesions among PWE is an over or underestimate of the truth. First, epidemiological studies are generally conducted in areas where the infection is expected to be common. Second, as describe in the Mexican study In three of the studies, single calcifications and/or single enhancing lesions were not considered as lesions compatible with NCC. This goes against the lesions described in Del Brutto et al. It is generally believed that NCC is a cause of late-onset epilepsy. Our meta-analysis contradicts this belief by obtaining very similar pooled estimates of the proportion of NCC among PWE in adults (aged 20 years old or more) and children (aged less than 20 years old). The proportion of NCC among children with partial seizures varied considerably. However, it does support the fact that children are affected by NCC.Our study has some limitations with regards to missing data, potential biases, and misclassification. Although a very broad search in seven different languages was conducted, relevant papers may have been overlooked. This situation may have the consequence of introducing a bias in the synthesis we were aiming to produce. Another potential bias may be publication bias, as we mainly considered published papers Another important limitation is that the ascertainment of NCC cases remains a problem. Although CT and MRI are considered the best tools to diagnose NCC, they can miss early stages of the larvae infestation in the brain A further limitation of this systematic review is that most published studies have been based on small sample sizes. Gender and age-specific data were often not available and, when reported, the age groups were not consistent. Most of the authors we contacted for additional information did not answer our correspondence. Hence, we could only report the prevalence estimates from a sub-sample of the studies and in two very broad age groups. Interestingly, we noticed that the prevalence of NCC among PWE in children aged less than 20 was much higher in South Africa. This supports prior reports suggesting that NCC may be more common in children in South Africa than elsewhere Finally, all the available literature is based on cross-sectional studies in communities or clinics that selected PWE and offered them a CT-scan of the brain. It is impossible to determine the temporality of the link between NCC and epilepsy in such study design. Unfortunately, a cohort study of people developing brain lesions of NCC which follows them to see if they develop epilepsy is not ethically feasible. Among the literature reviewed, one was a prevalence case-control study and reported a prevalence odds ratio of 6.9 between NCC and epilepsy This systematic review has shown several challenges for the assessment of NCC's global burden. One way to improve the assessment of the global burden of NCC would be to encourage and enforce the use of a standard diagnosis for NCC, such as that developed by Del Brutto et al. T. solium were associated with NCC. While this estimate may be biased due to measurement error, its robustness across populations and studies suggest that it is likely to be accurate. The number of DALYs lost due to epilepsy worldwide was estimated to be 6,223,000, with slightly higher values for males  than for females  Despite these challenges, this study found that approximately one-third of people with epilepsy living in regions endemic for Given the very robust estimate of the proportion of people with NCC lesions among PWE and, in some cases people with epileptic seizures, it may be possible to use the range of the percentage of NCC among PWE to estimate the number of NCC-associated epilepsy cases in endemic areas. To achieve this goal, we will need to obtain information on the prevalence of epilepsy in areas that are endemic for cysticercosis, that is, those countries were sanitation is poor, pork consumption occurs, and pigs have access to human feces. In the literature that we have reviewed, we have not come across publications suggesting that NCC is not endemic in areas where these three conditions are met. By multiplying the range of prevalence of epilepsy by the range of the proportion of NCC among PWE in each endemic area, we would obtain a range of values for the prevalence of NCC-associated epilepsy. Since epilepsy has been reported as the most common manifestation of NCC, such estimates will probably capture the majority of NCC-associated burden. This information will ultimately lead to the estimation of prevalence DALYs associated with NCC.Checklist S1PRISMA checklist.(0.07 MB DOC)Click here for additional data file.Flowchart S1PRISMA flow diagram.(0.06 MB DOC)Click here for additional data file.


MDR1/ABCB1) gene encodes the transport protein P-glycoprotein (a phase III xenobiotic transporter). P-glycoprotein is present in the intestinal mucosal lining and restricts absorption of certain carcinogens, among these polycyclic aromatic hydrocarbons. Moreover, P-glycoprotein transports various endogenous substrates such as cytokines and chemokines involved in inflammation, and may thereby affect the risk of malignity. Hence, genetic variations that modify the function of P-glycoprotein may be associated with the risk of colorectal cancer (CRC). We have previously found an association between the MDR1 intron 3 G-rs3789243-A polymorphism and the risk of CRC in a Danish study population. The aim of this study was to investigate if this MDR1 polymorphism was associated with risk of colorectal adenoma (CA) and CRC in the Norwegian population.Smoking, dietary factors, and alcohol consumption are known life style factors contributing to gastrointestinal carcinogenesis. Genetic variations in carcinogen handling may affect cancer risk. The multidrug resistance 1 and 95 confidence interval (95% CI) were estimated by binary logistic regression.Using a case-control design, the association between the MDR1 polymorphism (G-rs3789243-A) and colorectal adenomas or cancer. Carriers of the variant allele of MDR1 intron 3 had odds ratios (95% CI) of 0.97 (0.72–1.29) for developing adenomas, and 0.70 (0.41–1.21) for colorectal cancer, respectively, compared to homozygous wild type carriers.No association was found between the MDR1 intron 3 (G-rs3789243-A) polymorphism was not associated with a risk of colorectal adenomas or carcinomas in the present Norwegian study group. Thus, this MDR1 polymorphism does not seem to play an important role in colorectal carcinogenesis in this population.The  Colorectal cancer (CRC) is one of the leading causes of cancer-related mortality in the Western World, with great impact on the life quality of the affected persons . Both geMDR1/ABCB1) gene codes for P-glycoprotein, a membrane-bound transporter genotyping was performed on an Mx3000 machine , using allelic discrimination with TaqMan chemistry, as earlier described [Genomic DNA was isolated from blood samples according to standard procedures  with minescribed . In shorThe genotype frequencies were compared using a logistic regression model, measured as odds ratio (OR) with 95% confidence interval (CI). Two separate ORs were calculated; one crude (OR^a) and one adjusted for age and gender (OR^b). In addition we analyzed for carriers of the polymorphism separately. MiniTab Statistical Software, Release 13.1 Xtra (Minitab Inc.) for Windows was used for the statistical calculations Given the allele frequency of 0.5, we had 80% chance of detecting an OR of approximately 1.7 in carcinomas and 1.5 in adenomas at a 5% significance level .MDR1 polymorphism (G-rs3789243-A) and the risk of colorectal adenomas or cancer (Table MDR1 intron 3 had ORs (95% CI) of 0.96 (0.72–1.29) for developing adenomas, and 0.70 (0.41–1.21) for developing colorectal cancer, respectively, compared to homozygous wild type carriers.Characteristics of the study population and risk factors for CRC are shown in Table  G-rs378943-A and MDR1 polymorphism and the risk of colorectal adenomas or carcinomas in the present Norwegian study group. The risk of colorectal adenomas and carcinomas were not different for homozygous or heterozygous carriers of the variant genotype of MDR1 G-rs3789243-A compared with the homozygous wild type carriers. A power calculation showed that we had 80% chance of detecting an OR of approximately 1.7 in carcinomas and 1.5 in adenomas. The statistical power to look into interactions was therefore limited, and interaction studies were therefore not done.We found no associations between the studied MDR1 genotypes have previously been studied in relation to colorectal cancer susceptibility [MDR1 G-rs3789243-A polymorphism have previously been associated with a risk of colorectal cancer in the Danish population  and of ulcerative colitis in the Scottish population [tibility ,41,42. Ipulation . In the MDR1 C3435T polymorphism was studied in a Polish study population [The pulation . They fopulation .Several factors may contribute to the differences seen between the studies. First of all, the first report of genetic effects is likely to represent an overestimated odds ratios due to reporting bias . TherefoMDR1 shows a high degree of heterogeneity between different population [CARD15 polymorphisms in European populations [CARD15 variants in the Northern countries compared to the rest of Europe has been demonstrated.Population heterogeneity may also contribute. However, although pulation ,21,44, iulations -47, wherFurthermore, differences in carcinogenic exposure may modify the inherent risk associated with genetic susceptibility. For instance, the average monthly intake of alcohol was 11 units, corresponding to 88 g alcohol per month among the controls in the present Norwegian  cohort, MDR1 G-rs3789243-A polymorphism was not associated with risk of colorectal adenomas or carcinomas in the present Norwegian study group, indicating that this polymorphism does not play an important role in the colorectal carcinogenesis in the Norwegian population.The MDR1: Multidrug Resistance 1; OR: odds ratio; CI: confidence interval; PAH: polycyclic aromatic hydrocarbons; QPCR: real-time quantitative PCR; SNP: single nucleotide polymorphism; IRR: incidence rate ratio.CA: Colorectal Adenoma; CRC: Colorectal Cancer; HCA: heterocyclic amines; The authors declare that they have no competing interests.LA, DJ, MØ carried out the genotyping. VA and LA drafted the manuscript. EK established the KAM study. EK, MS and JH participated in sample preparation and collection, MS performed the statistical analyses and JH administered the KAM database. VA and UV conceived the genotyping study, and participated in its design and coordination and helped to draft the manuscript. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:


Myeloperoxidase (MPO) is an enzyme stored in azurophilic granules of polymorphonuclear neutrophils and macrophages and released into extracellular fluid in the setting of inflammatory process. The observation that myeloperoxidase is involved in oxidative stress and inflammation has been a leading factor to study myeloperoxidase as a possible marker of plaque instability and a useful clinical tool in the evaluation of patients with coronary heart disease. The purpose of this review is to provide an overview of the pathophysiological, analytical, and clinical characteristics of MPO and to summarize the state of art about the possible clinical use of MPO as a marker for diagnosis and risk stratification of patients with acute coronary syndrome (ACS). Myeloperoxidase (MPO) isa well-known enzyme, mainly released by activated neutrophils, characterised bypowerful pro-oxidative and proinflammatory properties. Recently, myeloperoxidasehas been proposed as a useful risk marker and diagnostic tool in acute coronarysyndromes and in patients admitted to emergency room for chest pain.Oxidative stress and inflammationplay important roles in the pathogenesis of destabilization of coronary arterydisease (CAD) leading to acute coronary syndromes (ACS). Infiltratingmacrophages and neutrophils participate in the transformation of stablecoronary artery plaques to unstable lesions , 2.ReceMajor evidence for MPO as enzymatic catalyst foroxidative modification of lipoproteins in the artery wall has been suggested ina number of studies performed with low-density lipoprotein .In contMPO activity can be measuredin blood and tissues by spectrophotometric assays using hydrogen peroxide and o-dianisidinedihydrochloride as substrates. In addition, MPO content can be measured inneutrophils as an index of degranulation with the Coulter counter and flowcytometry and circulating MPO by ELISA. Very recently, commercial methodsallowing low-cost and high-volume measurements have been proposed. Theintroduction of these methods of measurement might make MPO a new and usefulcardiac biomarker.n = 1138) were apparently healthy men and womenwho developed CAD during 8 years of follow-up; control subjects (n = 2237)matched for age, gender, and enrollment time, remained free of CAD. TheMPO levels were significantly higher in case subjects than in control subjectsand correlated with C-reactive protein (CRP) and white blood cell count. Riskof future CAD increased in consecutive quartiles of MPO concentration, with anodds ratio (OR) of 1.49 in the top versus bottom quartile. Afteradjustment for traditional risk factors, the OR in the top quartileremained significant at 1.36 (95% CI 1.07 to 1.73). Of interest in this study, serum MPO levels were associatedwith the risk of future development of CAD in apparently healthy individuals,but the association was weaker than that of traditional risk factors and CRP.However MPO, at variance from CRP, was largely independent from classical riskfactors.There have been a few but important clinicalstudies examining the role of MPO as a marker of risk for CAD. Using an enzymeassay, Zhang et al. showed tIn ACS, MPO has been consistently found to beassociated with the presence of instability and risk of future events in the studiesthat have explored these topics. Biasucci et al. first obUsing the same method, Buffon et al. studied μg/L wasassociated with an adjusted hazard ratio was 2.25 . Theeffects were particularly impressive in patients with undetectable cardiactroponin T (cTnT < 0.01 μg/L), in whom the hazard ratio was 7.48 . Ofinterest, the increase in risk was already evident after 72 hours, increasingonly slightly thereafter (P < .001) and in those with low C-reactiveprotein serum levels , suggesting that recruitmentand degranulation of neutrophils is a primary event and is followed by releaseof other systemic mediators and acute-phase proteins such as C-reactiveprotein. At variance from CRP levels, levels of MPO were not influenced bytroponin, suggesting a prognostic role of MPO independent from troponin andconfirming that inflammation is a primary phenomenon in ACS.The potential usefulness for risk stratification of blood concentrations of MPO wasexamined in 2 recent studies. In the CAPTURE trial ,MPO masereafter . This obμg/L. Furthermore,while troponin T takes three to six hours to rise to measurablecirculating levels after myocardial injury, MPO levels were significantlyelevated at baseline (even within two hours after the onset of symptoms) in patients who were initially negative for troponin T. These findings suggestthat measurement of MPO levels may be useful in triage in the emergencydepartment and that elevated plasma MPO levels may be a marker of unstableangina preceding myocardial necrosis and therefore, a predictor of vulnerableplaque. Interestingly, C-reactiveprotein levels predicted the risk of myocardial infarction at presentation forthe entire cohort but were not predictive of major adverse cardiac events inthe group that was negative for troponin T. In the cohort that was consistentlynegative for troponin T, the areas under the curve for MPO were significantlyhigher than those for troponin T, CK-MB, and C-reactive protein.In a study of 604 consecutive patients presenting to the emergency department withchest pain, Brennan et al. demonstrTaken together, these datasuggest that CRP and MPO may be complementary and explore different fields: CRP is a marker of diseaseactivity and vascular inflammation, and is useful for long-term riskstratification while MPO is a marker of plaque instability and neutrophilactivation and may be associated with short-term stratification, in particularin patients with troponin negative levels.P < .01.(2) Gensini score in the fourth quartile group of MPO level was significantly higherthan that in the first quartile group (P < .01). WBC in the fourthquartile group was also significantly higher than that in the first quartile group, P < .05. In addition, Kaplan-Meierevent rate curve showed that there was a significant difference in outcome  between the groups with MPO ≥ 62.9 AUU/L and ≤62.9 AUU/L of MPO serum level at 6-month follow-up visit (chi(2) = 13.5, P = .01).Furthermore, Cavusoglu et al. [More recently, several studies have alsoinvestigated the value of MPO in predicting long-term outcomes. Recently, S.-H. Liet al. have stuu et al. have invMocatta et al. have investigated the relationshipbetween plasma MPO and clinical outcome after AMI , 16.TheMPO is a marker of inflammation and oxidative stress that has been consistently demonstrated to beelevated in patients with ACS. However, the data so far available arerelatively few; therefore, more studies are requested to precisely define therole of MPO. In particular all studies involving MPO assessment have useddifferent methods, thus a standardization effort is needed. Furthermore, increasedMPO is not likely to be specific for cardiac diseases, as activation ofneutrophils and macrophages can occur in any infectious, inflammatory, orinfiltrative process, therefore, more studies should address and clarify thesepoints. So far, as reported in