PAI-1 polymorphism in cases than in controls, what may suggest the protective effect of this genotype for the risk of AAA development. None of the remaining polymorphisms tested were associated with AAA occurrence.The most important feature of abdominal aortic aneurysm (AAA) pathogenesis is an enzymatic degradation of elastic lamellae and extracellular matrix proteins particularly with participation of matrix metalloproteinases. Plasmin, which is responsible for the dissolution of fibrin in blood vessels, plays also a key role in the cascade for activation of the metalloproteinases. The purpose of this study was to evaluate the influence of selected polymorphisms in genes coding for tissue plasminogen activator (−7351 C/T polymorphism), urokinase-type plasminogen activator (1788 C/T polymorphism) and plasminogen activator inhibitor 1 (−675 4G/5G and −844 G/A polymorphism) on the susceptibility to AAA. We performed a case–control study of 153 polish patients hospitalized due to AAA and compared them with matched healthy control subjects. The polymorphisms were ascertained through genotyping by polymerase chain reaction and restriction digestion of amplified fragments or through high-resolution melting analysis. In this study we have found lower frequency of wild-type GG genotype of the −844G/A  PAI-1 may prevent the development of AAA [Fibrinolytic system is a proteolytic cascade of activation events responsible for the enzymatic dissolution of fibrin blood clots. It is composed of plasminogen, its active form—plasmin, plasminogen activators—tissue-type (t-PA) and urokinase type (u-PA), inhibitors which regulate activation of plasminogen—plasminogen activator inhibitor type 1(PAI-1) and type 2 (PAI-2) and activity of plasmin—α2-antiplasmin. Plasmin, which is formed from plasminogen by t-PA or u-PA, is an enzyme responsible for the dissolution of fibrin in blood vessels, but also plays a key role in the cascade for activation of matrix metalloproteinases (MMPs) –7. It ist of AAA .u-PA consisting in a transition of cytosine to thymine at position 1788 (rs2227564) causes a missense mutation (Pro141Leu) in the kringle domain of uPA protein, which has been shown to decrease the affinity for fibrin [Helicobacter pylori infection [Tissue type plasminogen activator is mainly involved in intravascular thrombolysis, while urokinase mediates pericellular proteolysis during cell migration, wound healing, and tissue remodeling under physiological and pathological conditions –11. −735r fibrin . This ponfection .PAI-1 gene cause the elevated PAI-1 level [PAI-1 promoter results in the disappearance of one binding site for a DNA-binding protein that acts as a transcriptional repressor [PAI-1 gene promoter includes a consensus sequence binding site for the Ets nuclear protein and can be implicated in the regulation of the PAI-1 gene expression [PAI-1 belongs to the serpin superfamily of proteins and is the main regulatory protein of the fibrinolytic system. In addition to its role as a regulator of haemostasis, PAI-1 also plays a significant role in several biological processes which include: wound healing, atherosclerosis, such metabolic disturbances as obesity and insulin resistance, tumor angiogenesis, chronic stress, bone remodeling, asthma, rheumatoid arthritis, fibrosis, glomerulonephritis, sepsis, and others , 23. Bot-1 level , 25. Theepressor , 27. Thupression , 29.The purpose of our study was to examine the occurrence of selected polymorphisms in genes coding for t-PA (−7351 C/T), u-PA (1788 C/T) and PAI-1 (−675 4G/5G insertion/deletion and −844 G/A) in patients suffering from AAA.The study population was comprised of 153 Polish patients with AAA, who were hospitalized in the M. Pirogow Regional Specialist Hospital in Lodz. Control group consisted of 152 healthy volunteers who had no history or symptoms of aneurysms and other cardiovascular diseases. The study was approved by Ethics Committee of Medical University of Lodz (consent number: RNN/19/07/KE). All subjects gave written informed consent to participate.Venous blood samples were taken from all study participants. Genomic DNA was isolated from leukocytes obtained from whole blood samples using phenol–chloroform extraction method or QIAamp DNA Blood Mini Kit .u-PA 1788 C/T, PAI-1 -844 G/A and PAI-1 -675 4G/5G polymorphisms were genotyped using polymerase chain reaction-restriction fragment length polymorphism sites (PCR–RFLP). t-PA −7351 C/T polymorphism was determined by melting curve analysis using Light Scanner™ HR I 96 . The PCR primer pairs, annealing temperature, PCR product sizes and restriction enzymes are listed in Table 2), 2 μl of 2.5 mM dNTPs, 100 pmol of each primer, 1 U of Taq DNA polymerase , 2 μl of genomic DNA and deionized water added to a final volume of 20 μl. Moreover, in the case of PAI-1 −844 G/A polymorphisms there were additional 4 μl of 25 mM MgCl2 used. Basically, the amplification reactions were carried out as follows: 94 °C for 5 min, followed by 35 cycles of 95 °C for 30 s for DNA denaturation, 56–63 °C , 72 °C for 1 min. Amplification and correct sizes of PCR products were confirmed on polyacrylamide gels in Tris/acetate/EDTA buffer in comparison with 100 bp or 1 kb DNA Ladder (Fermentas). After staining with ethidium bromide the gels were photographed under UV light using Image Master VDS system (Pharmacia Biotech).PCR reactions for amplification of polymorphic segments were performed in reaction mixture containing 2 μl of 10× PCR buffer (with 15 mM MgClu-PA and PAI polymorphisms) was then digested overnight with an excess of proper restriction enzyme  in 96-well plate coated with 20 μl of mineral oil. Each reaction mixture contained 4 μl of Light Scanner Master Mix , 4 μl of water, 0.5 μl of each primer (100 pmol/μl) and 1 μl of DNA template. The conditions of the PCR were as follows: 94 °C for 5 min, followed by 20 cycles of 94 °C for 30 s for DNA denaturation, 60 °C for 30 s for primers annealing and 72 °C for 5 s for DNA extension. Amplified products of 60 base pairs were subjected to high-resolution melting analysis (HRM) in the same plate using the light scanner instrument.In the case of To confirm the results obtained using PCR–RFLP and HRM technique, selected samples (every tenth) were subjected to DNA sequence analysis carried out by Genomed S.A. .2 square test and the 95 % confidence intervals (CI) for disease odds ratio (OR) calculated with the use of logistic regression. All the analyses were derived by means of Statistica v8.0 software. The level of significance was set at p < 0.05.The results are reported as percentages (%) or means with standard deviations (±SD). Analysis of association between genotype/allele distribution and AAA risk was performed with the use of the χp > 0.20).The study group consisted of 51 females (33.3 %) and 102 males (66.6 %) with an age range from 44 to 73 years. The mean age in the study population was 59 years, SD = 7. The control group consisted of 152 healthy volunteers: 60 females (39.5 %) and 92 males (60.5 %) with their ages ranging from 37 to 70 . The control subjects had no history or symptoms of aneurysms and other cardiovascular diseases. The cases and controls were homogenous in gender and age distribution   (Table p = 0.008). Thus, it could be suggested that the GG wild-type genotype may have protective effect for the risk of AAA development.We did not observe any statistical differences in the distribution of genotypes and alleles of The pathogenesis of AAA has not yet been completely defined. The literature data show that proteolytic activity of fibrinolytic system may be one of the factors contributing to the development of aneurysms , 7. Thust-PA −7351C/T polymorphism with ischemic stroke [p = 0.28).The TT genotype of −7351 C/T tissue-type plasminogen activator gene polymorphism has been reported to be associated with reduced t-PA release rates due to the decreased transcriptional activity , 13. It c stroke , 32. To We have also studied, for the first time, the association between the 1788 C/T urokinase-type plasminogen activator gene polymorphism and risk of AAA. Since it is documented that elevated urokinase expression may participate in AAA formation and rapture –35, and PAI-1 gene promoter (−675 4G/5G and −844 G/A), that increase the PAI-1 expression and plasma concentration. Elevated levels of PAI-1 decrease plasmin formation resulting in the inhibition of the proteolytic activity of MMPs, and thus could prevent the formation of aneurysm. Our study indicates a lower frequency of wild-type GG genotype of the −844 G/A PAI-1 polymorphism in cases than in controls, what may suggest the protective effect of this genotype for the risk of AAA development. Although the −844 G/A PAI-1 polymorphism influences the plasma PAI-1 level, there is at present a lack of studies regarding the association of this polymorphism and risk of AAA. However, there are some literature data on the relationship between this polymorphism and other vascular system diseases involving myocardial infarction [Another fibrinolytic gene which we have studied regarding to its contribution to the aneurysm disease is plasminogen activator inhibitor 1. We have chosen two polymorphisms in the farction , ischemifarction  or deep farction . But allPAI-1 promoter polymorphism and AAA occurrence in Polish population. Nevertheless, there are some data concerning the correlation of 4G/5G PAI-1 polymorphism with AAA. Jones et al. [PAI-1 5G/5G genotype presented with AAAs of largest diameter. Whereas another report found the influence of 5G/5G genotype on the aneurysm development, but only in the case of typical familial AAA [PAI-1 promoter polymorphism is also reported to be associated with other cardiovascular diseases, including thrombosis [Our study revealed also the lack of correlation between −675 4G/5G s et al.  noticed s et al.  documentlial AAA . −675 4Grombosis , myocardrombosis , 36, 42 rombosis , 44.t-PA, 1788 C/T u-PA and −675 4G/5G PAI-1 polymorphisms and the risk of AAA development. Only in the case of −844 G/A PAI-1 polymorphism the study revealed a slight difference in the wild-type genotype frequency between AAA patients and healthy controls. This result should be confirmed in a larger population.In conclusions, in this study no association was observed between −7351 C/T


Proximal humeral fractures are common lesions of the elderly, but there are no established treatment guidelines. A surgical treatment for comminuted and displaced fractures of the proximal humerus was developed and is still evolving. The aim of this study was to perform a quantitative review to evaluate the risk of avascular necrosis (AVN) in patients with proximal humeral fractures who were treated by operative fixation compared with conservative treatment.We searched the PubMed, MEDLINE, Springer, Elsevier Science Direct, Cochrane Library, Google Scholar, China National Knowledge Infrastructure (in Chinese), and Wanfang database (in Chinese) up to December 2013 to identify studies related to operative fixation and AVN in patients with proximal humeral fractures.p = 0.64]. The subgroup meta-analysis by study design (retrospective/prospective), sample size (≤40/>40), and ethnicity (European/Asian) demonstrated similar results. However, the subgroup analysis by specific operative approach (plate fixation/tension band wiring fixation/others) indicated that plate fixation was associated with a higher rate of AVN than conservative treatment .Seven studies with a total of 291 patients (142 operative fixation cases and 149 conservative treatment cases) with proximal humeral fractures were considered in the meta-analysis. The overall meta-analysis showed no significant difference in the incidence of AVN between the two groups [odds ratio (OR) 1.42, 95% confidence interval (CI) 0.33–6.11, Plate fixation was associated with a higher risk of AVN development than conservative treatment in patients with proximal humeral fractures. Although proximal humeral fractures are common lesions of the elderly, accounting for 5% of all fractures and 45% of all humeral fractures-3, thereCurrently, many surgical solutions such as operative fixations , and Wanfang (in Chinese) up to December 2013. The key words ‘avascular necrosis’, ‘operative fixation’, and ‘proximal humeral fractures’ were used for the searches. References from retrieved papers were checked for additional studies. We collected data only from fully published papers—not from meeting or conference abstracts. The publication date was not restricted in our research.Studies were included in the meta-analysis if they met the following criteria. The study had to contain (1) the occurrence rates of AVN in patients with proximal humeral fractures , (2) operative fixation versus conservative treatment data, and (3) the effect size reported as odds ratios (ORs) and 95% confidence intervals (CIs) or it could be calculated. The sample size and age range were not limited. Studies were excluded if the study (1) described only conservative treatment data in a review or report, (2) was a reduplicated study, or (3) did not compare operative fixation with conservative treatment.Articles were reviewed and filtered out independently by two investigators according to our criteria. The data were then extracted independently in duplicate using a standardized form to assess eligibility for inclusion. Data items included study details  of the study, year of the study's publication, location of participants, design of the study, follow-up time) and characteristics of the participants . Discrepancies were resolved by consensus.Evaluation of quality mainly included the sample size and recruitment of the studies. The papers were first selected by reading the document titles and abstracts. Then, we read the full text of each paper to determine whether the study conformed to our inclusion criteria. Two investigators independently completed this task. Assuming that differences would occur, agreements were to be reached by discussion.Q-statistic[I2 parameter testing[p < 0.05 or I2 > 50% was considered to indicate a heterogeneous nature. When substantial heterogeneity was detected, we calculated the summary ORs and their 95% CIs with the DerSimonian and Laird method in the random effect model[The meta-analysis was performed in fixed or random effect models as appropriate. The effect sizes of the ORs with 95% CIs were pooled to assess the AVN incidence associated with operative fixation in proximal humeral fractures compared with conservative treatment. Heterogeneity among studies was evaluated by Cochran's tatistic and I2 p testing. Also, pct model. If hetect model. To testWe further conducted subgroup analysis according to the study design (retrospective/prospective), sample size (≤40/>40), ethnicity (European/Asian), and specific operative approach (plate fixation/tension band wiring fixation/others) to investigate the impact of study characteristics on our outcomes. Publication bias was also assessed by Egger's regression asymmetry test.http://ims.cochrane.org/revman) and the STATA software package version 11.0 . All p values were two-sided. A value of p < 0.05 was considered to indicate statistical significance.Analyses were performed using the software Review Manager 5.1  among studies. Thus, the random effect model was applied. The results showed that there was a relatively higher risk of AVN in patients with proximal humeral fractures who underwent operative fixation than in those treated conservatively , but the difference was not statistically significant. The result of Egger's linear regression test .In an overall analysis of the seven selected studies, the heterogeneity test showed that there were heterogeneities (p > 0.05). In the stratified analysis by ethnicity, Asian patients  showed less risk of developing AVN than European patients  when treated with operative fixation. The result of the subgroup analysis by operative approach demonstrated that plate fixation had a significantly increased risk of AVN compared with conservative treatment , whereas tension band wiring fixation showed a risk similar to that with conservative treatment .The outcomes of the subgroup analysis stratified by study design, sample size, ethnicity, and operative approach are shown in Table p > 0.05), and the heterogeneity test showed that there were heterogeneities . Also, the estimates changed little, which implied that our results were statistically reliable.We performed the sensitivity analysis by removing one study each time and rerunning the model to determine the effect on each overall estimate. By omitting the Fjalestad et al. study, for insp value for AVN was 0.19, which indicated no statistical significance for publication bias.Egger's linear regression test was performed to assess the publication bias of the articles. In this study, the total Previous studies,11,18,21p > 0.05). This finding was consistent with results in a study by Ilchmann and coworkers, who found that the risk of developing AVN was at an OR (95% CI) of 1.50 (0.29–7.75)[p > 0.05) showed that operative fixation was associated with a nonsignificantly decreased risk of AVN, which was inconsistent with the overall estimates. Hence, study design and ethnicity might be factors that cause heterogeneity.Findings from the meta-analysis showed that in patients with proximal humeral fractures, operative fixation was associated with a nonsignificantly higher risk of AVN development than conservative treatment . AccordiAlthough the overall analysis demonstrated a nonsignificant difference in the incidence of AVN after operative fixation versus conservative treatment, the subgroup analysis by operative approach showed a markedly increased risk of AVN after plate fixation compared with conservative treatment. Tension band wiring fixation and conservative treatment had similar incidences of AVN. Multiple materials have been developed for open reduction and internal fixation including cerclage wires, rotary self-locking intramedullary nails, intramedullary interlocking nails, and six-hole dynamic compression plates-24. AmonTreatment of displaced proximal humeral fractures remains a challenge for the orthopedic surgeon, and theThe treatment and prognosis of proximal humeral fractures have improved increasingly with the development of medical technology. The study of Poeze et al. indicated that radiographic evaluation in patients with minimally displaced proximal humeral fractures is helpful for predicting functional outcome during conservative treatment. HumeralThe present study was an updated meta-analysis on the risk of AVN after operative fixation of a proximal humeral fracture. There are some limitations of this study. First, only seven published studies were included in the meta-analysis. Second, significant heterogeneities were detected in the meta-analysis for clinical neurological outcome, which might be distorting the outcomes as heterogeneity is one of the major concerns in meta-analyses regarding validity. Also, dThe findings of the present meta-analysis suggested that plate fixation in patients with proximal humeral fractures had a significantly higher risk of AVN development than conservative treatment, whereas tension band wiring fixation and conservative treatment had similar risks. Thus, further investigation with more high-quality studies should be carried out to obtain a more precise outcome.The authors declare that they have no competing interests.JX and CZ participated in the design of this study and performed the statistical analysis. JX carried out the study together with CZ, collected important background information, and drafted the manuscript. TW conceived of this study, participated in its design, and helped draft the manuscript. All authors read and approved the final manuscript.


PCSK9) E670G polymorphism and coronary artery disease (CAD) and lipid levels, but the results were controversial. Thus, we performed this meta-analysis to investigate the association between PCSK9 E670G polymorphism and lipid levels and the susceptibility to CAD.Studies had investigated the associations between proprotein convertase subtilisin/kexin type 9  with 95 % CI was used for the meta-analysis of PCSK9 E670G polymorphism and lipid levels. The publication bias was examined by using Begg’s funnel plots and Egger’s test.All relevant articles according to the inclusion criteria were retrieved and included in the present meta-analysis. Odds ratios (ORs) with 95 % confidence interval (CI) were used to analyze the strength of the association between P < 0.001), as well as in allelic genetic model . When the subgroup analysis stratified by ethnicity and HWE was performed, the positive result existed in most of the subgroups. For lipid levels association, the pooled effects indicated that the G allele carriers had higher TC and LDL-C levels than the non-carriers . There was no difference in the levels of TG and HDL-C between the G carriers and the non-carriers in the whole population . When the studies were stratified by ethnicity and type of study, the G carriers had higher TC levels than the non-carriers  in the non-Asian subgroup. The similar results existed in cohort subgroup. The association between PCSK9 E670G polymorphism and LDL-C levels was significant in all subgroups. Meanwhile, the G carriers had higher TG levels than the non-carriers  in the case–control subgroup. AG + GG genotypes had lower HDL-C levels than AA genotype in Asian subgroup  and in case–control subgroup .A total of seventeen studies met the inclusion criteria. For CAD association, the pooled effects indicated that the G allele carriers had higher risk of CAD than non-carriers in dominant genetic model  contains supplementary material, which is available to authorized users. Coronary artery disease (CAD) is the major manifestations of atherosclerotic process, which is expected to remain one of the leading causes of mortality until at least 2030 . EpidemiProprotein convertase subtilisin/kexin type 9 (PCSK9), originally named neural apoptosis regulated convertase-1 (NARC-1), is the ninth member of the proprotein convertase family . PCSK9 iThe coding gene for human PCSK9 protein is located on chromosome 1p32.3 and it encompasses 12 exons and encodes a 692 amino acid glycoprotein. It contains a signal peptide (1–30), a pro-domain (31–152) and a catalytic domain (153–451) followed by a cysteine-histidine rich domain (526–692). The pro-domain acts as an intramolecular chaperone, which ensures the catalytic domain to fold correctly in the ER.PCSK9 were identified as the cause of a third monogenetic form of autosomal-dominant hypercholesterolemia (ADH), except LDLR and apolipoprotein B (apoB) genes [PCSK9 in the serum lipids homeostasis and the pathogenesis of CAD.The mutations of B) genes . RecentlPCSK9 gene have been found, including 153 substitutions, eight insertions and two deletion mutations . These mutations are divided into gain-of-function (GOF) and loss-of-function (LOF) mutations. LOF mutations in PCSK9 gene increase the concentration of LDLR on the cell surface, which promotes the uptake of serum LDL-C and prevents the procession of CAD.GOF mutations enhance the degradation of LDLR, which reduces the uptakes of LDL-C and ultimately increases the circulating LDL-C levels. In addition, it may also regulate the production and secretion of apoB and enhance the levels of very low-density lipoprotein (VLDL) [Up to date, a total of 163 mutations in PCSK9 E670G variant (rs505151), a common variant in exon 12, is located within the cysteine-rich C-terminal domain, which participates in the regulation of auto-catalysis [atalysis , but theatalysis . For insatalysis . On the atalysis .PCSK9 gene polymorphism and lipid levels and the risk of CAD.To enhance statistical power to produce a more precise result, we performed this meta-analysis based on available data aimed to derive a more precise association of the PCSK9 E670G polymorphism and serum lipid levels and/or the risk of CAD. We also searched the Chinese electronic databases, such as Wanfang Data and China National Knowledge Infrastructure (CNKI) databases for Chinese literatures. The following keywords were used in the search: “PCSK”, “proprotein convertase subtilisin/kexin type 9”, “neural apoptosis regulated convertase-1”, “NARC-1”, “polymorphism”, “gene”, “rs505151”, “mutation”, “variant”, “lipid”, “dyslipidemia”, “coronary heart disease”,“myocardial infarction”, “coronary artery disease”, “ischemic heart disease”, “acute coronary syndrome”, “CAD” and “CHD”. The reference lists of the relevant papers that we identified were also checked.The meta-analysis followed the Perferred Reporting Items for Systematic Reviews and Meta-analysis  For the lipid levels, the data were presented as mean ± standard deviations (SD) and the study had one of the serum lipid levels, at least including TC, triglyceride (TG), LDL-C or high density lipoprotein cholesterol (HDL-C); 3) For risk of CAD, the frequencies of genotypes in controls and CAD groups were clear.The eligible study must met the following criteria: 1) Case–control study or cohort study investigating the relationships between Exclusion criteria: 1) The study was not conducted in human; 2) Family-based study. If there are several multiple publications from the same population, the most recent literature with the largest sample size was adopted.A special form was prepared for recording the available information. Two reviewers (Cai and Zhang) independently extracted the following data from each eligible study: first author, year of publication, country, ethnicity, genotype and allele distributions, geontyping method, lipid levels, type of study, Hardy-Weinberg equilibrium (HWE). The disagreements were resolved by consulting with the third author (Shi).PCSK9 E670G genotype distributions was assessed by Fisher’s exact test. The odds ratios (ORs) with 95 % confidence interval (CI) were applied to evaluate the strength of the association between the PCSK9 E670G polymorphism and the susceptibility to CAD. Because the frequency of genotype GG in most of studies was low, the pooled ORs were only performed for allelic model (G vs. A) and dominant model (AG + GG vs. AA). The pooled standardized mean difference (SMD) with 95 % CI was used for the meta-analysis of PCSK9 E670G polymorphism and lipid levels. We defined PCSK9 670G carriers as having the AG and GG genotypes. If the unit of the lipids is presented as mg/dl, we converted it to mmol/l. In several studies, the subjects were divided into different subgroups , so we treated each subpopulation as a separate comparison in our meta-analysis. Between-study heterogeneity was investigated and measured by using Cochran’s Q test. It was also detected by using the I2 statistic. If the between-study heterogeneity was significant , a random-effect model (a Dersimonian-Laird method) was used to calculate the results. Otherwise, the fixed effect model (a Mantel-Haenszel method) was adopted [The deviation from the HWE for the  adopted .Because of the significant heterogeneity among studies, we carried out sensitivity analysis while evaluating the results again by omitting one single study each time. We also performed the subgroup analysis stratified by ethnicity (“Asian” or “non-Asian”) and type of study (“case–control” or “cohort”) to explore the sources of heterogeneity. The publication bias of literature was examined by Begg’s funnel plots, which was verified by Egger’s linear regression test.The STATA version 12.0  was used for the meta-analysis.n = 2); (2) data were obviously wrong (n = 2); (3) data were not obtained (n = 2); (4) family-based study (n = 1). In the end, 17 literatures met all the inclusion criteria and are involved in the present meta-analysis [Through the initial retrieval, a total of 147 records were selected. 123 records were excluded according to abstract. Full-text articles were retrieved and seven literatures were excluded for the following reasons: (1) data were repetitive or overlapping  varied greatly from 2.3 to 48.3 %. None of the distribution of genotypes in controls deviated from HWE, except Gao et al’s study. Among them, seven studies were performed among Asians, one among Caucasians and one among Africans, respectively. The genotyping method in most studies was PCR-RFLP, except for Mo et al’s study.The characteristics of the included studies were listed in Tables PCSK9 E670G polymorphism and serum lipid levels, including 14,558 participants. Of these, 22 comparisons from 14 papers, 20 comparisons from 13 papers, 21 comparisons from 14 papers and 23 comparisons from 14 papers presented the data on TC, TG, HDL-C and LDL-C, respectively. 11 comparisons were conducted in Asians, 11 in Caucasians, and 3 in other ethnic populations. Genotype distributions in 19 populations or subpopulations were consistent with HWE. Seven papers were case–control studies and eight papers were cohort studies.For lipid levels, ten papers separately provided data for more than one subpopulation , and we treated each subpopulation as a separate comparison. Therefore, 25 comparisons from 15 papers were used to evaluate the relationship between PCSK9 E670G polymorphism with the risk of CAD. Because of absence of significant between-study heterogeneity, the fixed-effect model was applied. The pooled effects indicated that the G allele carriers had higher risk of CAD than non-carriers in dominant model , as well as in allelic model   Table .Table 3SP < 0.001; for allelic model, OR: 1.503, 95 % CI: 1.244–1.816, P < 0.001) and non-Asians . Subgroup analysis was also performed by HWE, significant association only existed in subgroup in HWE , suggesting that the results are credible.Then the subgroup analyses stratified by ethnicity and HWE were performed. In the subgroup analyses stratified by ethnicity, significant associations were found among both Asians    Fig. . There wly) Fig. .Table 4SP = 0.027). The similar results existed in cohort subgroup. The association between PCSK9 E670G polymorphism and LDL-C levels was significant in all subgroups. Meanwhile, the G carriers had higher TG levels than the non-carriers  in the case–control subgroup. AG + GG genotypes had lower HDL-C levels than AA genotype in Asian subgroup  and in case–control subgroup .The subgroup analyses stratified by ethnicity and type of study were also performed. In the non-Asian subgroup, the G carriers had higher TC levels than the non-carriers . For lipid levels, there was significant heterogeneity among the total comparisons for TC, HDL-C and LDL-C . In order to explore the possible sources of heterogeneity, subgroup analyses stratified by ethnicity (Asian or non-Asian) and type of study (cohort or case–control) were performed. But the between-study heterogeneity was not obviously reduced under the subgroup analyses.For CAD, the between-study heterogeneity was not significant  . The dominant model suggested that 670G carriers were at a 1.601-fold higher risk of CAD than non-carriers. In the whole population, there was no difference in the levels of TG and HDL-C between the 670G carriers and non-carriers.In 2003, Abifadel et al.  firstly nclusive , 26, 28.nclusive , 21. The factors . In addiP = 0.027) and in cohort studies . The association between this mutation and LDL-C levels was significant in all subgroups. Interestingly, the 670G carriers had higher TG levels in case–control subgroup . Moreover, the 670G carriers had lower HDL-C levels in Asian population  and case–control subgroup . The sensitivity analysis found that the pooled effects did not change after excluding single study each time, which indicated that the results were stable.For the lipid study, there was a significant between-study heterogeneity, which may affect the interpretation of the results. To explore the heterogeneity, the subgroup analyses stratified by ethnicity and type of study were performed. But, we still could not explain the source of heterogeneity entirely. In the subgroup analyses, we found that 670G carriers had high TC levels in non-Asian populations (SMD: 0.126, 95 % CI: 0.014-0.238, As other meta-analyses, there were several inherent limitations in this study. Firstly and mainly, the between-study heterogeneity is significant, which was a potential problem that may affect the interpretation of the results. As we all know, the between-study heterogeneity may be influenced by age, sex, sample size, type of study, and so on. Although the subgroup analyses were performed, the heterogeneity was not explained entirely. Secondly, the relationships between this SNP and CAD risk and lipid levels did not consider the confounding factors, such as age, sex, smoking, drinking and other lifestyle factors. In the study conducted by Evans et al, E670G polymorphism was associated with increased LDL-C levels in man but not in women. Thirdly, the number of studies included in the meta-analysis was small for CAD risk. Only one study was conducted in Caucasian population and one study was in African population. Fourthly, although the Egger’s tests indicated no remarkable publication bias in our meta-analysis, the inevitable publication bias may remain in the results because the papers having negative result were probably more difficult to be accepted for publication. Finally, the present meta-analysis was not able to assess gene-gene and gene-environment interactions.PCSK9 E670G polymorphism was associated with CAD risk and lipid levels.Despite the limitations, the present meta-analysis concluded that


FMO3), a microsomal antioxidant defense enzyme, has been suggested to contribute to essential hypertension (EH). The present study was designed to investigate whether common functional polymorphism E158K (rs2266782) of the FMO3 gene is associated with EH susceptibility in a Russian population. A total of 2 995 unrelated subjects from Kursk  and Belgorod (357 EH patients and 422 population controls) regions of Central Russia were recruited for this study. DNA samples from all study participants were genotyped for the FMO3 gene polymorphism through PCR followed by RFLP analysis. We found that the polymorphism E158K is associated with increased risk of essential hypertension in both discovery population from Kursk region  and replication population from Belgorod region  after adjustment for gender and age using logistic regression analysis. Further analysis showed that the increased hypertension risk in carriers of genotype 158KK gene occurred in cigarette smokers, whereas nonsmoker carriers of this genotype did not show the disease risk. This is the first study reporting the association of the FMO3 gene polymorphism and the risk of essential hypertension.Gene encoding flavin-containing monooxygenase 3 ( FMO3 gene have been found to contribute to the disease trimethylaminuria (TMAuria), an inborn error of metabolism resulting from diminished oxidation of the tertiary amine trimethylamine to trimethylamine N-oxide resulting in a severe body odour in affected individuals [Oxidative stress resulting from the enhanced production of reactive oxygen species (ROS) and decreased activity of antioxidant defense enzymes has been implicated in pathogenesis of essential hypertension (EH) , 2. The ividuals . Interesividuals , 5. FMO3 gene and hypertension in Irish population [ FMO3 gene polymorphisms was associated with hypertension risk. This negative result can be explained by a relatively low statistical power which is related with a small sample size in the study. The present study was designed to investigate whether common functional polymorphism E158K (rs2266782) of the FMO3 gene is associated with susceptibility to essential hypertension in Russia. Pursuing this aim, two independent populations of ethnic Russians from Kursk and Belgorod regions of Central Russia were included in this study to be used as discovery and replication cohorts, respectively.A single study investigated the relationship between common polymorphisms of thepulation . HoweverP > 0.05). Diagnosis of essential hypertension in both populations was verified by qualified cardiologists. Individuals were defined as hypertensive according to World Health Organization criteria or if they had a history of receiving any antihypertensive drug. Diagnosis of EH in untreated patients was defined by a seated systolic and/or diastolic blood pressure greater than 140 and/or 90 mmHg, respectively, on at least 2 separate measurements. All EH patients had no clinical signs, symptoms, and laboratory findings suggestive of secondary hypertension.The study was approved by Ethical Review Committee of Kursk State Medical University. A total of 2 995 Russian unrelated subjects from Kursk (discovery cohort) and Belgorod (replication population) regions of Central Russia were included in the study. The discovery cohort comprising 2 216 subjects  was recruited at Cardiology Clinics of Kursk Regional Clinical Hospital and Neurology Clinics of Kursk Emergency Medicine Hospital over two periods: between 2003 and 2006  and betw FMO3 gene was done using PCR followed by RFLP analysis as described elsewhere [Genomic DNA was isolated from peripheral blood samples using a standard phenol/chloroform procedure. Genotyping of polymorphism E158K of thelsewhere . The gen FMO3 genotype was assessed according to dominant, recessive, and additive genetic models, and the chi-squire  odds ratio with 95% confidence interval was calculated. Odds ratios were calculated as a measure of the association of the FMO3 genotype with hypertension risk, with the effects of the allele 158K assumed to be additive , dominant (with scores of 0 for EE genotype and 1 for EK and KK genotypes combined), or recessive (with scores of 0 for EE and EK genotypes combined and 1 for KK genotype). The statistical significance was established at P ≤ 0.05. Bonferroni correction for P values (Padj) was applied in cases when multiple tests were performed. Statistical calculations were performed with Statistica for Windows 8.0 .The association between the polymorphism and hypertension risk was estimated by odds ratio (OR) with 95% confidence interval (CI) using multiple logistic regression analysis with adjustment for confounding variables such as age and gender. EachAs can be seen from  FMO3 genotype frequencies were in agreement with Hardy-Weinberg equilibrium (HWE) in control groups from both populations (P > 0.05). However, the genotype frequencies in hypertensive groups from both Kursk and Belgorod populations showed a deviation from HWE (P < 0.05) due to a decreased heterozygosity. Data on allele and genotype frequencies in Kursk and Belgorod populations are shown in  FMO3 genotypes was observed between the case and control groups of Kursk population . A statistically significant difference in the distribution of the FMO3 genotypes between the case and control groups was also observed in Belgorod population (P = 0.02). The genotype 158KK was found to be associated with increased risk of hypertension in both Kursk and Belgorod populations after adjustment for gender and age using multiple logistic regression analysis.TheP = 0.18) and dominant  genetic models did not show a significant effect of the FMO3 gene polymorphism on hypertension risk. Meanwhile, recessive model of the genetic association between the polymorphism and hypertension risk was established to be statistically significant .Logistic regression analysis revealed that additive , whereas nonsmoker carriers of the genotype did not show the disease risk .Data on the analysis for the FMO3 gene polymorphism contributes to the development of essential hypertension. Firstly, it is known that an amino acid substitution Glu-to-Lys at a position 158  is associated with decreased activity of the enzyme in oxidation of catecholamine releasing agents such as tyramine, phenylethylamine, adrenaline, and noradrenaline possessing vasopressor effects [ FMO3 gene variant may be responsible for decreased catabolism of the vasoactive hormones, thereby contributing to hypertension risk through increased vasoconstriction and heart rate modulation. Secondly, a diminished activity of FMO3 in carriers of genotype 158KK may enhance ROS generation and induce oxidative stress, one of the proposed pathogenetic mechanisms of human hypertension [We can suggest two possible mechanisms by which the effects , 10. Thirtension , 11–13. rtension , enhanci FMO3 gene polymorphism is associated with susceptibility to essential hypertension, the result which was originally found in Kursk population and then validated in an independent population from Belgorod region. We also observed in the Kursk population that the increased risk of hypertension in carriers of heterozygous genotype 158KK of the FMO3 gene occurs only in smokers, whereas nonsmokers possessing this genotype do not have the risk of the disease. This finding is an indication of the gene-environment interaction, a situation when adverse effects of cigarette smoking on hypertension can be enhanced by the phenotypic effects of the FMO3 polymorphism. Although much remains to be learned regarding the biological basis of the relationship between the FMO3 gene and hypertension risk, it is already clear that further studies focusing on the investigation of gene-environment interactions may improve our understanding of the disease pathogenesis and define novel therapeutic and preventive options as a means of personalized medicine in cardiologic practice.The present study demonstrated for the first time that the


In stratified analyses, we found TNF-α-308A allele was significantly associated with higher risk of NHL, B-cell lymphomas (BCL), T-cell lymphomas (TCL) and diffuse large B-cell lymphomas (DLBCL) in Caucasians . Interestingly, it was associated with decreased risk of NHL, BCL and DLBCL in Asians . These findings also suggest TNF-α might play a distinct role in pathogenesis of NHL in different populations.Tumor necrosis factor-α (TNF-α) is an immunoregulatory cytokine involved in B- and T-cell function, and also plays an important role in inflammation and cancer. TNF-α-308G>A has been associated with constitutively elevated TNF-α expression. Several studies have reported the association between the TNF-α-308G>A polymorphism and non-Hodgkin lymphomas (NHL) risk, however, results are still inconsistent. To solve these conflicts, we conducted the first meta-analysis to assess the effect of TNF-α-308G>A polymorphism on the risk of NHL and various subtypes (additive model) including 10,619 cases and 12,977 controls in Caucasian and Asian populations. Our meta-analysis indicated that TNF-α-308G>A polymorphism is not associated with NHL risk when pooling all studies together (OR = 1.06, 95% CI: 0.92–1.23,  MalignaTumor necrosis factor-α (TNF-α) is one of the most important pro-inflammatory and tumor-related cytokines for its regulating immune response, inflammation, Th1/Th2 balance and lymphomagenesis . IncreasIn this study, we conducted the first comprehensive meta-analysis to test whether the TNF-α-308 polymorphism is associated with NHL overall risk or its subtypes, especially BCL, TCL, DLBCL, FL, chronic lymphocytic leukemia/small lymphocytic lymphomas (CLL/SLL), mantel cell lymphomas (MCL), mucosal-associated lymphomas (MALT), peripheral T-cell lymphomas (PTCL) and natural killer/T-cell lymphomas (NK/TCL). We also performed subgroup analysis by descent (Caucasians and Asians) to assess a possible factor that might influence the overall results. Therefore, this study might have more statistical power and increase precision to estimate association between TNF-α-308 polymorphism and its effect on NHL.2.2.1.p = 0.916). But TNF-α-308G>A in all controls in the initial report was not consistent with HWE (p = 0.0007). We analyzed the initial report composed of 8 subgroups comprehensively [et al. [In the initial screening for key words, 405 potential articles were identified in PubMed, Embase and Cochrane Library. After removing duplication, 321 articles were needed for further assessment. Among them, 293 were excluded because of inappropriate study design or control samples. Of the remaining 28 relevant articles, 8 articles were excluded for using the same patients. 2 articles were also excluded for their controls in concordance with Hardy-Weinberg equilibrium (HWE). With strict including criteria, the final pool of eligible articles consisted of 18 articles involving a total of 10,619 patients with NHL and 12,977 healthy controls. Because of the large sample size, Caucasians and Asians were considered as population stratification in this meta-analysis. ensively . Finally [et al.  conducte [et al. –38, and  [et al. –42. Seve2.2.vs. G) which stratified by ethnicity (Caucasians and Asians). Results of meta-analysis and primary data extracted from studies are listed in Based on a large pooled sample size, we analyzed TNF-α-308G>A polymorphism effects on risks of NHL, BCL, TCL and subtypes  in additive model . In subgroup analysis based on population, significant associations were detected in Caucasian and Asian populations, respectively. In Caucasians with 7893 cases and 8447 controls, participants with TNF-308A allele had an increased NHL risk . However, in Asians with 2726 cases and 4530 controls, decreased risk was observed . vs. G) stratified by ethnicity.Of the combined 10,619 patients with NHL and 12,977 controls, no risk association was observed in TNF-308G>A polymorphism and NHL with significant heterogeneity between studies . These indicated that the results of this meta-analysis were reliable and had not been overly influenced by any one of studies.pBegg’s = 0.529). Egger’s test also suggested no significant publication bias existed in this meta-analysis .We performed Begg’s funnel plot and Egger’s test to evaluate the publication bias of all included studies. 2.2.2.Thirteen studies comprising a total of 20,064 participants  and 4 studies including 11,832 participants  were analyzed for an association between TNF-α-308G>A polymorphism and BCL or TCL risk.p = 0.411; I2 = 74.4%, phet < 0.001). Increased risk was found in Caucasians , decreased risk was detected in Asians . In TCL analysis, TNF-308A associated with a higher NHL risk in Caucasians .For BCL analysis, the pooled OR across all studies was not statistically significant , but with a decreased risk in Asians . The pooled OR was 0.97 .For DLBCL, the most common NHL subtype, there were 3353 cases and 10,817 controls included in the analysis. Consistent with the results for all NHL, the TNF-α-308A allele was associated with an increased risk of DLBCL in Caucasians  by ligand passing and signal transduction. TNFR1 has a death domain that could interact with TNF-α receptor-associated death domain (TRADD), sequentially recruiting proteins to induce caspase-3 activation for apoptosis. TRADD could also bind TNF receptor-associated factor 2 (TRAF2) to recruits proteins activating IKK, GCK, and RIP, which finally leads to the NF-κB, JNK, and MAPK pathway activation for anti-apoptosis and cell survival. Although TNFR2 lacks the death domain, it could also bind TRAF2 to active an anti-apoptosis pathway ,43. AberPrevious studies suggest higher expression of TNF-α is associated with NHL risk at the time of diagnosis –17. ThesTNF-α and LT-α gene lie in the major histocompatibility complex class III, telemetric to the class II and centrometric to class I gene. Therefore, TNF-α being in linkage disequilibrium (LD) with these genes may also be linked to another region, haplotype or extended, that can influence NHL development . This meBecause of relatively low incidence of NHL, sample sizes of some studies included in this analysis are very small. The major strength of this study is the larger pooled sample size involving a total of 10,619 patients with NHL and 12,977 healthy controls for association study, which would largely minimize the possibility of chance findings. In addition, patients in our study were all Caucasians or Asians, which would exclude the biased results due to population stratification.In conclusion, we performed the first comprehensive meta-analysis involving 10,619 patients with NHL and 12,977 controls from 18 articles to evaluate the association between TNF-α-308G>A polymorphism and NHL risk. Our study showed that TNF-α-308G>A SNP in the promoter region of TNF-α gene is associated with NHL risk. In addition, TNF-α-308A increases risks of NHL, BCL, TCL and DLBCL in the Caucasian population; however, interestingly, it reduces risks of NHL, BCL and DLBCL in the Asian population. This association might be mediated by constitutive changes of TNF-α expression in individuals carrying the -308A allele, to induce inflammatory responses or the alternative pathway which be involved in NHL initiation and progression. Our meta-analysis emphasizes that genetic variation plays a crucial role in cancer; theTNF-α-308G>A polymorphism might play a role in a specific subtype of NHL and its importance varies in different populations. Further studies should focus on the function of how variants affect NHL in different populations and the elucidation of the pathway involved which may eventually lead to a better understanding of tumorigenesis and contribute to the prevention of NHL.3.3.1.We carried out a search in three electronic databases PubMed, Embase and Cochrane Library to find relevant publications up to November 2013, using key words related to the TNF-α gene polymorphism in combination with various NHL subtypes . The sea3.2.Selection of studies had to meet the following criteria: (1) case-control studies, family or sibling pairs studies were excluded; (2) published in English; (3) subjects were limited to adult and without autoimmune diseases, studies with children were also excluded; (4) DNA was extracted from peripheral blood leukocytes; (5) study described the association between TNF-α-308 polymorphism and NHL risk; (6) sufficient data for estimating odds ratio (OR) and its corresponding 95% confidence interval (95% CI); (7) control group fulfilled HWE. When the same subject group occurred in more than one study, only the complete study was chosen to be included in this meta-analysis.3.3.An initial screening of title and abstract was performed for the first step, followed by further screening based on full-text review. Information was independently extracted from all eligible publications by two investigators (K.Z. and J.D.), including the first author, publication year, ethnicity, sample size, genotyping method, the number of each genotype in cases and controls. For studies with subjects of different ethnic groups and had sufficient information, we extracted data separately for each ethnicity. Disagreements were resolved through discussion.3.4.2 statistics. In this meta-analysis, I2 > 50% was defined as heterogeneity. Fixed-effect model (Mantel-Haenszel method) was used to evaluate inter-study heterogeneity. If heterogeneity existed, random-effect model (DerSimonian-Laird method) was used. Z test was used to determine the pooled OR and 95% CI. Analyses were also conducted on the subgroups of studies based on ethnicity. The potential influence of publication bias was assessed using Begg’s funnel plot and Egger’s linear regression test [2 test. All statistical tests were carried out with SPSS 16.0  and Stata 12.0 . A 2-tailed p < 0.05 was considered as statistical significance.We assessed the association between TNF-α-308G>A polymorphism and NHL risk by crude ORs and 95% CIs in an additive model. Heterogeneity among studies was examined with Iion test ,53. To eion test . HWE in 4.We performed the first comprehensive meta-analysis involving 10,619 patients with NHL and 12,977 controls from 18 articles to evaluate the association between TNF-α-308G>A polymorphism and NHL risk. Our study showed that TNF-α-308G>A SNP in the promoter region of TNF-α gene is associated with NHL risk. In addition, TNF-α-308A increases risks of NHL, BCL, TCL and DLBCL in the Caucasian population; interestingly, this polymorhism reduces risks of NHL, BCL and DLBCL in the Asian population. This association might be mediated by constitutive changes of TNF-α expression in individuals carrying the -308A allele, to induce inflammatory responses or the alternative pathway which be involved in NHL initiation and progression. Our meta-analysis emphasizes that genetic variation plays a crucial role in cancer; the TNF-α-308G>A polymorphism might play a role in a specific subtype of NHL and its importance may vary in different populations. Further studies should be focused on how variants affect NHL in different populations and the elucidation of the pathways involved which may eventually lead to a better understanding of tumorigenesis and contribute to the prevention of NHL.


COQ2 gene led us to question whether variants in the COQ2 gene are associated with PD in Han Chinese in a case-control study. A total of 564 patients with PD were genotyped using the ligase detection rection, together with 484 gender- and age-matched healthy subjects. The M128V and R387X variants of COQ2 were not detected in patients or controls; instead, we detected only the heterozygous V393A variant (CT genotype). The frequency of the CT genotype encoding the V393A mutation was significantly higher in patients PD (4.08%) than in controls (1.86%), corresponding to an odds ratio of 2.24 . The frequency of the C allele of the V393A variant was significantly higher in patients with PD than in controls , and this was also observed in a meta-analysis of studies from mainland China, Taiwan and Japan. Subgroup analysis of our data showed that the V393A variant was significantly associated with early-onset PD  but not with late-onset disease . Gender was not significantly associated with either genotype or minor allele frequencies. In conclusion, our findings show for the first time that the V393A variant in the COQ2 gene increases risk of PD among the population of east Asia. These results, combined with research on Japanese, lend genetic support to the hypothesis that oxidative stress underlies pathogenesis of both PD and MSA.Both Parkinson’s disease (PD) and multiple system atrophy (MSA) are neurodegenerative diseases of uncertain etiology, but they show similarities in their pathology and clinical course. The fact that the gene encoding α-synuclein is associated with both diseases also suggests that they share some genetic determinants. Recent studies in Japan associating MSA with a variant in the  SNCA gene; however, the etiology of both disorders is unclear [SNCA, DJ-1, GBA, LRRK2 and other genes [SNCA transgene has been established [interleukin (IL) 21A, IL-1B, IL-8, tumor necrosis factor, α-1-antichymotrypsin, SLC1A4, SQSTM1 and EIF4EBP1 [Multiple system atrophy (MSA) and Parkinson’s disease (PD) are neurodegenerative diseases that share several common features in their pathogenesis and their clinical manifestations. In both diseases, the characteristic pathological hallmark is aggregation of α-synuclein, encoded by the  unclear . PD is rablished . MSA, orablished . Since tEIF4EBP1 –5.SNCA variants rs3822086 and rs3775444 [SNCA polymorphism.Evidence that MSA and PD may share genetic determinants came with the announcement of an association of MSA with the COQ2 gene (M128V-V393A/M128V-V393A). They also identified heterozygous COQ2 mutations encoding single substitutions (V393A) or double substitutions (R387X/V393A). The COQ2 gene product is an enzyme that participates in the synthesis of coenzyme Q10, and the polymorphism that leads to the V393A enzyme variant leads to lower enzyme expression and therefore lower production of coenzyme Q10 [COQ2 variants may be associated with risk of PD. Indirect evidence for this has come from studies showing that serum from patients with PD has significantly lower coenzyme Q10 concentrations than serum from healthy individuals, and that coenzyme Q10 supplementation shows promise for treating PD [A study of familial MSA in Japan recently identified a homozygous mutation in the In order to explore possible overlap in genetic determinants of PD and MSA and in the corresponding pathophysiological mechanisms, we conducted a case-control study in Han Chinese. We examined whether the homozygous mutation M128V-V393A/M128V-V393A, the compound heterozygous mutation R387X/V393A and the single heterozygous mutation V393A are associated with PD.Han Chinese patients with sporadic PD  were consecutively recruited from the movement disorder center of West China Hospital, Sichuan University. Their mean age was 62.64±10.83 years; patients were divided into a group with late-onset PD  if the age at onset was >50 years , and a group with early-onset PD , in which the mean age at onset was 44.98±4.66. PD was diagnosed in all patients independently by two movement disorder specialists according to criteria for idiopathic PD from the UK PD Society Brain Bank [To provide a control group to compare with the entire PD group as well as with the EOPD and LOPD subgroups separately, we recruited 484 healthy Han Chinese  who were unrelated to the PD patients  and who were age- and gender-matched with them. his study protocol was approved by the Ethics Committee of West China Hospital, Sichuan University. All study participants provided written informed consent.COQ2 polymorphisms was then performed by the Shanghai BioWing Applied Biotechnology Company using the ligase detection reaction  was verified, and genotype frequencies were compared using the chi-squared test. Associations of gender with particular alleles and genotypes were also assessed by chi-squared test, whereas age differences among the three groups were assessed using the t test. Two-tailed P < 0.05 was considered significant. Since only one variant(V393A) was involved in our research, there is no need to perform the multiple test or multiple correction. SPSS 17.0  was used to perform all statistical analysis, including chi-squared and t tests.Age was reported as mean ± SD, and gender, allele and genotype frequencies were reported as percentages. Allele and genotype frequencies were determined by direct counting of 2 = 3.167, p = 0.160) and gender   and controls . The frequency of the CT genotype for the V393A variant was significantly higher in patients (4.08%) than in controls (1.86%)  (The PD patients and controls showed similar distributions of age (χ= 0.492) . We dete (1.86%) . Effect = 0.039) .COQ2 polymorphism and risk of PD, we compared genotype and minor allele frequencies in PD subgroups based on gender or age at onset. Gender was not significantly associated with genotype or minor allele frequencies  and complex II (succinate-ubiquinone oxidoreductase) of the mitochondrial electron transport chain. It also serves as an antioxidant [COQ2 leads to lower production of coenzyme Q10 [The ioxidant . Coenzymioxidant . Oral adioxidant . The V39zyme Q10 ,13, resuzyme Q10 .COQ2 V393A variant. One study in Japan [This is the first study to confirm an association between PD and the in Japan  and two in Japan ,17 failein Japan , and a min Japan ; anotherin Japan . StudiesSubgroup analysis of our data showed significant differences in genotype and minor allele frequencies between EOPD patients and controls, but not between LOPD patients and controls. EOPD usually involves obvious lysosomal and mitochondrial dysfunction, which helps increase oxidative stress; LOPD, in contrast, is usually more associated with synaptic transmission . These fCOQ2 polymorphisms in our patients since we did not sequence the entire gene. We did not detect the M128V or R387X variants, nor did we detect the homozygous CC genotype of the V393A variant. Finally, we did not take into account possible gene-gene or gene-environment interactions that may be confounding our results. These limitations highlight the need for larger studies that consider the complete range of COQ2 polymorphisms in different ethnicities, especially Asians, as well as possible interactions of the polymorphisms with other genes and the environment.Our study has several limitations. First, although we included only patients with probable PD, diagnosis was not confirmed by pathology in any of our patients. This raises the possibility that some may not have had disease. Second, the relatively small sample of patients with PD meant that the statistical power for calculating genetic associations with PD was only 53.7%. Third, we examined only a limited array of COQ2 gene and risk of PD in Han Chinese and the followed meta-analysis extend this association to the population of east Asia. These results lend genetic support to the hypothesis that oxidative stress underlies the pathogenesis of both neurodegenerative diseases(PD and MSA).In conclusion, our study suggests an association between the V393A variant of the


PTPN11, which encodes tyrosine phosphatase Shp2, is a critical gene mediating cellular responses to hormones and cytokines. Loss of Shp2 promotes hepatocellular carcinoma (HCC), suggesting that PTPN11 functions as a tumor suppressor in HCC tumorgenesis. The aim of this study was to evaluate the effects of the short tandem repeat (STR) polymorphism (rs199618935) within 3'UTR of PTPN11 on HCC susceptibility in Chinese populations.in-silico studies were used to evaluate the possible functional significance of this polymorphism. Logistic regression analysis showed that compared with individuals carrying shorter alleles (11 and 12 repeats), those subjects who carry longer alleles (13 and 14 repeats) had a significantly decreased risk of HCC , with the risk decreased even further in those carrying allele 15 and 16 . Biochemical investigations showed that longer alleles of rs199618935 conferred higher PTPN11 expression in vivo and in vitro. The altered luciferase activities in reporter gene system suggested that STR regulation of PTPN11 expression could be a transcriptional event. Finally, in-silico prediction revealed that different alleles of rs199618935 could alter the local structure of PTPN11 mRNA.We analyzed the associations in 400 patients from Jiangsu province of China, validating the findings in an additional 305 patients from Shanghai of China. Unconditional logistic regression was used to analyze the association between rs199618935 and HCC risk. Additional biochemical investigations and PTPN11 contributes to hepatocarcinogenesis, possibly by affecting PTPN11 expression through a structure-dependent mechanism. The replication of our studies and further functional studies are needed to validate our findings.Taken together, our findings suggested that the STR polymorphism within  Hepatocellular carcinoma (HCC) is one of the most common malignancies and the third leading cause of cancer death PTPN11) encodes the non-receptor protein tyrosine phosphatase SHP2, which is critical for RAS/ERK pathway activation in most receptor tyrosine kinase, cytokine receptor, and integrin signaling pathways PTPN11 have been shown to be directly associated with the pathogenesis of Noonan syndrome and childhood leukemias Protein tyrosine phosphatase, non-receptor type 11  polymorphism (rs199618935) and conducted a two-stage case-control study to analyze the genetic effect of the polymorphism on the susceptibilities to HCC in Chinese populations. Additional experimental and in-silico studies were used to evaluate the possible functional significance of this polymorphism.Previous studies have reported genetic variation within This study was approved by the Ethical Committee of Soochow University. Written informed consent was obtained from each participant before investigation.The case-control study was performed on genomic DNA extracted from peripheral blood of 705 newly diagnosed incident HCC cases together with 723 controls after obtaining informed consent. All subjects recruited were unrelated ethnic Han Chinese. For the Jiangsu's case-control study (Panel I), the case series were comprised of 400 HCC patients diagnosed, hospitalized and treated in the affiliated hospitals of Soochow University from 2007 to 2011. For the Shanghai's case-control study (Panel II), 305 HCC patients were recruited at Nantong Tumor Hospital from 2003 to 2005. Controls were cancer-free individuals selected from a community nutritional survey that was conducted in the same regions during the same period as the recruitment of cancer patients. Controls without clinical evidence of liver disease were frequency matched for age (±5 years) and sex to each set of HCC individuals. The diagnosis of the cases, the inclusion and exclusion criteria for the cases and controls, and the definition of smokers and drinkers were described previously in details Additional 48 tumor tissues and adjacent non-HCC tissues from patients with a diagnosis of HCC were collected from Department of General Surgery, the First Affiliated Hospital of Soochow University from 2011 to 2012. All cases had histological confirmation of their tumor diagnosis and none of these patients had received preoperative chemotherapy or radiotherapy. After surgical resection, the fresh tissues were immediately stored at −80°C until the DNA/RNA extraction was processed.5′- GTGTCCCTTCTACTTCCCTCT-3′, Reverse primer: 5′- GCTGGGCTTGTGACTTGTTT-3′). The PCR products were analyzed by 7% non-denaturing polyacrylamide gel electrophoresis (PAGE) and visualized by silver staining Genomic DNA of peripheral blood samples, tissue samples and HCC cell lines were isolated using genomic DNA purification kit (Qiagen). DNA fragments containing rs199618935 were amplified with a pair of genotyping primers , and 50–100 ng of cDNA. The negative control experiments were performed with distilled H2O as template. The expression levels of target genes were normalized with GAPDH using a 2−ΔΔCT method The Hep-G2, Hep3B and Huh7 hepatoma cell lines were obtained directly from Shanghai Cell Bank of Chinese Academy of Sciences. Cells were cultured in Dulbecco's Modified Eagle's Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and 1% penicillin-streptomycin at 37°C in a humidified 5% CO2 incubator before RNA extraction. Total RNA was isolated from hepatoma cell lines and tissue samples using RNA isolation kit (Qiagen). cDNA was generated using random primers and Superscript II reverse transcriptase (Invitrogen). A SYBR Green real-time PCR was performed using Roche LightCycler 480 to quantify relative PTPN11 expression in these samples. GAPDH was chosen as the internal control. Primer sequences used for PTPN11 and GAPDH were as follows: PTPN11-F: 5′-GATCTCTAGACCCCAACTGTTAGTCAATCTGAGC-3′ and reverse primer 5′- CATGGATCCTTGTCCCAGCTACTGTAAGCAGC-3′ from three homozygous human genomic DNA samples. The PCR products were separated in agarose gel and extracted, purified, and cloned with TA cloning Kit (Promega). The repeat numbers of different alleles were confirmed by sequencing. Finally, the 3′UTR of Renilla luciferase in the vector pRL-SV40 (Promega) was replaced with the cloned 3′UTR of PTPN11 by restriction enzymes XbaI and BamHI. The resulting constructs were verified by sequencing.The partial structures (∼570 bp) of human PTPN11-3′UTR containing allele 12, 14 and 15 of rs199618935 were amplified with forward primer 5 cells per well in 24-well plates (BD Biosciences). Twenty-four hours after the plating, cells were transfected by Lipofectamine 2000 according to manufacturer's manual. In each well, 500 ng constructed pRL-SV40 vector and 50 ng pGL3 control vector were cotransfected. Six replicates were performed for each group and each experiment was repeated at least three times. After transfection for 24 hr, cells were harvested by the addition of 100 µl passive lysis buffer. Renilla luciferase activities in cell lysate were measured with the Dual Luciferase assay system (Promega) in TD-20/20 luminometer (Turner Biosystems) and were normalized with the firefly luciferase activities.The Hep-G2, Hep3B, sk-Hep-1 and Huh7 hepatoma cell lines were seeded at 1×10As certain conserved structures more likely serve important biological functions, a 60-bp region covering the polymorphism was analyzed using RNAfold to predict the putative influence of different alleles on local folding structures of PTPN11 using default parameters 2 test. Since rs199618935 is a multi-allele polymorphism, the genotypic frequencies were calculated by a specific counting method based on different alleles. The samples would be classified into specific genotypic groups provided it has one or two specific alleles. Genotypic and allelic frequencies for each allele between HCC patients and controls were compared by χ2 test. To facilitate analysis, alleles with frequencies lower than 3% were combined with the adjacent alleles . Unconditional logistic regression was used to analyze the association between rs199618935 and HCC risk, adjusted by gender, age, smoking, drinking and HBV infection status. As HBV infection was one of the major risk factors, a stratified analysis by HBV infection status for overall population was performed using binary logistic regression model. Student's t test was used to examine the differences in luciferase reporter gene expression. The normalized expression values of PTPN11 in tissue samples and hepatoma cell lines were compared with student's t test and one-way ANOVA, respectively. These statistical analyses were implemented in Statistic Analysis System software . P<0.05 was used as the criterion of statistical significance. All statistical tests were two sided.The genotype distribution was analyzed by Hardy-Weinberg equilibrium using χP values >0.05).The demographic characteristics of the 705 HCC patients and 723 controls from two independent case-control sets were summarized in Six different alleles  were detected corresponding to 11–16 repeats  and there were totally 11 and 12 different genotypes observed in overall cases and controls, respectively. Genotypic and allelic frequencies of rs199618935 as well as its associations with HCC susceptibility were presented in P<0.01). To validate our findings in tissue samples, we further examined the genotype-phenotype correlations in three common hepatoma cell lines . Compared with Hep-G2 cell lines carrying 12–15 genotype, the PTPN11 mRNA expression levels of Huh-7 (14–14 genotype) and Hep3B (14–14 genotype) were significantly increased , which validated the tumor-inhibiting effect of PTPN11 in HCC. Similarly, Ptpn11 has also been proved to be a tumor suppressor in cartilage and involved in metachondromatosis by inducing hedgehog signaling Most recent experimental data suggest that in contrast to the proto-oncogene effect of dominant-active mutants, PTPN11 may act as a tumor suppressor in hepatocarcinogenesis PTPN11 3′UTR regulates mRNA level, we transfected three different constructs containing different alleles  into Huh7, Hep3B, sk-Hep-1 and Hep-G2 cells and then assayed luciferase levels. Our data provided first evidence that the construct containing allele 14 displayed the highest luciferase activity, which was significantly higher than that of constructs containing allele 12 and 15. The altered luciferase activities we observed in the reporter gene system suggested that STR regulation of PTPN11 expression can be a transcriptional event, such as changed RNA stability in a post-transcriptional level. Early studies have shown that STR polymorphism in the 3′UTR formed structural elements (stem-loops) and contributed to mRNA regulation in-silico studies have shown different structures formed by sequences containing different alleles (To test whether the different allele in human  alleles . Based oPTPN11 mutations have been found in various types of human cancer Deregulation of PTPN11 causes hyperactivation of ERK, leading to growth abnormality. Gain-of-function PTPN11 STR polymorphism and HCC risk, the significance of this finding is limited by the relative small sample size used in this study. However, result from our genetic association analysis provides a preliminary data and evokes the need for future study with different or expanded case-control populations to confirm our observations. Furthermore, the underlined molecular mechanisms between the STR polymorphism and PTPN11 expression still need to be fully investigated both at genetic and functional levels.Although this is the first report for the possible association between In summary, we have provided initial evidence that the length variation of the “TCA” repeats within human PTPN11 3′UTR may play a functional role in regulating the expression of PTPN11 and subsequently affect development of HCC. Therefore, PTPN11 may be a promising marker for personalized diagnosis and therapy of HCC.


P = 0.001) in European subgroup. No publication bias was detected. Our results showed that the SNP49 polymorphism of CTLA4 gene was related to increased risk of GO.Many studies have established that T-lymphocyte antigen-4 (CTLA4) is a susceptible gene for Graves' disease (GD). Also many studies showed the association between the CTLA4 exon-1 49A/G polymorphism and the risk of developing Graves' ophthalmopathy (GO) in GD patients. But those results were inconsistent. In recent years many new studies were published which helped to shed light on the relationship of CTLA4 SNP49 with GO. So we performed the meta-analysis to explore the association between the SNP49 and GO susceptibility in GD patients. Studies up to February 29, 2012, were searched by using PubMed. The odds ratio was used to evaluate the strength of the association. Altogether 12 case-control studies involving 2,505 participants were included in the meta-analysis. Results showed that the G allele was related to the increased risk of GO compared with the A allele under allelic genetic model (OR = 1.14, 95% CI: 1.14–1.72,  Graves' disease is a thyroid autoimmune disorder with 25–50% individuals having ophthalmopathy which is called thyroid-associated ophthalmopathy (TAO) or GO. The clinical manifestations of GO can mostly be explained by the increased volume of extraocular muscles and orbital connective tissues . The symHuman CTLA-4 gene, located on 2q33, encodes a molecule which plays an important role in the downregulation of CD28 interaction with the ligands on the surface of antigen-presenting cells (APCs). The important inhibitory role of CTLA-4 in T-cell function has made it be one candidate gene when exploring autoimmune diseases.Many reports have established the association of CTLA-4 with GD –9. The COne meta-analysis exploring the association of CTLA-4 gene polymorphism with GO was performed in 2006 which found no definite result . Many neWe searched PubMed up to February 29, 2012, for all possible publications on the association between the CTLA-4 49A/G polymorphism and GO. Search terms were used as follows: “Cytotoxic T-lymphocyte associated antigen-4 or CTLA4,” “Graves' disease,” and “ophthalmopathy.” We also manually searched all reference lists of the relevant articles for additional papers. Language was limited to English. All eligible studies met the following criteria: (1) case-control design investigating the associations of CTLA4 49A/G polymorphism and GO; (2) providing genotype distribution information in both cases and controls or provided odds ratio (OR) with 95% confidence interval (95% CI) . When a study reported results on different subpopulations, we treated each subpopulation as a separate comparison in the meta-analysis. For overlapping data, the most complete or recent study was included.Two authors (Pengfei Du and Changjiang Wang) independently extracted data and reached consensus on all items. The following information was sought from each report: authors, journal and year of publication, country of origin, selection and characteristics of GO cases and controls, demographics, ethnic group of the study population, eligible and genotyped cases and controls, and number of cases and controls for the genotype. For studies including subjects of different ethnic groups, data were extracted separately for each ethnicity whenever possible.Z test was used to determine the significance of the pooled OR and P value <0.05 was considered significant. The between-study heterogeneity across all eligible comparisons was estimated using the chi-square based Q statistic [P < 0.10. Data were combined using both fixed-effects (Mantel-Haenszel) and random effects (DerSimonian and Laird) models [τ correlation coefficient) [P values were two-sided.The primary analysis compared GO cases with controls for the frequency of G versus A alleles. This analysis aims to detect overall differences. Pooled OR with 95% CI was used to evaluate the strength of the associations of CTLA-4 49A/G polymorphism and GO. When zero events occurred, we treated this problem by adding 0.5 to all the 2 × 2 cells of the contingency table to calculate the study-specific OR just as implemented in RevMan software. We examined the association of 49A/G polymorphism and GO for allelic contrast (G versus A). Furthermore, we conducted subgroup analyses by stratifying ethnicity into Europeans and Asians separately. A tatistic . Heterog) models . Random ) models . Inverteficient)  evaluateThe primary literature search yielded 17 papers , 28, 29.P heterogeneity = 0.01), and the random effects pooled OR was not significant: OR = 1.14, 95% CI: 0.94–1.38, P = 0.18  [P = 0.002, P heterogeneity = 0.32)  . The pooled frequency of G allele was 68.1% and 51.8% for cases and controls, respectively. Two studies suggested an at-risk effect of G allele , 24 and P = 0.18 . One stu = 0.19) . Sensiti = 0.32) . The G c = 0.01) .The current meta-analysis including 12 case-control studies was in an effort to clarify the relationship between CTLA-4 gene polymorphism and GO susceptibility. The overall results indicated that the CTLA-4 49A/G polymorphism was associated with susceptibility of GO which was more significant in European population. Although obvious heterogeneity was detected for 49A/G associations, sensitivity analyses did not materially alter the overall and subgroup results under different genetic models, indicating that the results were stable and reliable.The polymorphism of CTLA-4 had been suggested in many diseases which included Addison's disease, autoimmune hypothyroidism, and rheumatoid arthritis. The CTLA-4 49A/G SNP in exon-1 leads to the substitution of Ala with Thr in the signal peptide part which was reported to cause misprocessing of CTLA-4 in the ER resulting in less efficient glycosylation and diminished surface expression of CTLA-4 protein . StudiesThe association of CTLA-4 49A/G SNP with GO was more significant in European population but not in Asian population. The CTLA4 49A/G SNP shows the same effect on the GO development with different significance. The results are confusing but not conflicting. This can be explained by the following reasons. Firstly, both GD and GO are multigenic conditions. It is unlikely to have one major susceptibility gene contributing to the development of GO. In the studies of Xu et al., they transiently transfected a T-cell line with a CTLA-4 construct harboring either the G or the A allele of the 49A/G SNP and found no difference in CTLA-4 expression and/or function harboring the A or the G allele . It is lIn conclusion, our analysis first intensified the association between 49A/G and GO susceptibility, which showed that the G allele is a risk factor for GO. More risk factors should be considered in future association studies of the CTLA-4 SNP with GO. The gene-gene and gene-environment interaction should also be taken into consideration in future research. The identification of the candidate gene and understanding of the mechanisms by which they cause disease will identify those individuals at risk for developing GO in the future. Treatment will evolve towards more preventive strategies. The treatment will be personalized by targeting the specific pathways that are most contributory to GO development. We believe that the future will witness the engagement of novel molecules, such as altered peptide ligands  and spec


EPHX1 Tyr113His and His139Arg polymorphisms in the EPHX1 gene and the risk of head and neck cancer.To evaluate the association between the EPHX1 Tyr113His and His139Arg polymorphisms with HNC performed up until June 1st, 2014, were identified using a predefined search strategy. Summary odds ratios (ORs) and 95% confidence intervals (CIs) were used to evaluate the strength of these associations.Studies on the association of Tyr113His (1890 cases and 1894 controls) and 10 studies of His139Arg polymorphisms (1982 cases and 2024 controls), were considered eligible for inclusion. Overall, the pooled results indicated that the EPHX1 Tyr113His polymorphism was significantly associated with increased HNC risk . However, no significant association was found between the His139Arg polymorphism and HNC risk. In the subgroup analysis, a statistically significant association between the EPHX1 Tyr113His polymorphism and HNC was observed in population-based case-control studies (PCC), which involved less than 500 participants and genotype frequencies in HWE. This association showed minimal heterogeneity after excluding studies that were determined to contribute to heterogeneity. After categorizing the studies by publication time, a sensitivity analysis and cumulative meta-analysis of the two associations were conducted, and the results of the two analyses were consistent.In this meta-analysis, 10 case-control studies, which included 9 studies of EPHX1 Tyr113His polymorphism may be a risk factor for HNC, while the EPHX1 His139Arg polymorphism has no association with HNC risk.Our meta-analysis suggests that  Head and neck cancer (HNC) is the fifth most common type of human cancer , with apmEH) (EPHX1), which is one of the phase I detoxification enzymes found on the endoplasmic reticulum in many tissues, plays a role in the metabolism of potential carcinogens in tobacco smoke analysis was not performed. However, the activity of EPHX1 can be affected by single Tyr113His and His139Arg polymorphisms or a combination of these polymorphisms [SNP) analyses were considered, which prevented us from performing a pooled analysis. Finally, significant heterogeneity was found in studies examining EPHX1 polymorphisms, and in some studies, the genotype distribution showed a deviation from HWE. As a retrospective study, the quality of a meta-analysis is dependent on the methodological limitations of the original studies. To minimize bias, we first designed a detailed protocol and performed a meticulous search as a predefined search strategy.However, the meta-analysis described herein had some limitations. First, unadjusted OR estimates were adopted in this meta-analysis because we could not obtain sufficient information to calculate adjusted ORs with potential confounders, such as age and sex. Second, the study size and the sample size for some subgroup analyses were limited, which contributes to the possibility of type I and type II errors. Thus, the reliability of our results may need to be further tested. Third, similar to previous studies, this meorphisms –47. In tEPHX1 polymorphisms and HNC risk. Our meta-analysis suggests that the EPHX1 Tyr113His polymorphism may be a risk factor for HNC, while the EPHX1In conclusion, this meta-analysis evaluates the relationship between His139Arg polymorphism has no association with HNC risk. In subgroup analysis, a statistically significant association between the EPHX1 Tyr113His polymorphism and HNC was observed in population-based case-control studies, studies that enrolled more than 500 participants and studies in which the genotype frequencies were in HWE. When stratified by ethnicity, no significant associations were found in this study; thus, well-designed studies with a large sample size that examine multiple ethnicities are required. Moreover, further meta-analyses must be performed to estimate the effects of both single SNP analysis and combination (two-SNP) analysis. Gene–environment interactions should also be examined to determine the association between the EPHX1 polymorphisms and HNC risk.S1 PRISMA Checklist(PDF)Click here for additional data file.S1 FigHis/His+ Tyr/His vs. Tyr/Tyr; B: Arg/Arg+ Arg/His vs. His/His. The studies outside the range between -2 and 2 were seen as the outliers and the major source of heterogeneity.A: (TIF)Click here for additional data file.S2 FigHis/His+ Tyr/His vs. Tyr/Tyr; B. Arg/Arg+Arg/His vs. His/His.A. (TIF)Click here for additional data file.S3 FigEPHX1 Tyr113His polymorphism.(A) For (TIF)Click here for additional data file.S4 FigEPHX1 His139Arg polymorphism.(B) For (TIF)Click here for additional data file.


Background. Cisplatin and carboplatin cause nephrotoxicity by forming platinum-DNA adducts and lead to cell death. Methods. One-hundred and sixteen Taiwanese lung cancer patients who received cisplatin or carboplatin more than twice were recruited, and their genotypes were determined. The risk of renal dysfunction, injury to the kidney, failure of kidney function, loss of kidney function, and end-stage kidney disease (RIFLE) criteria were used to evaluate the occurrence of nephrotoxicity. A logistic regression, multiple regression with a classification and regression tree (CART), and the Framingham study risk score were used to analyze interactions between genetic and nongenetic factors in producing platinum-induced nephrotoxicity. Results. ERCC1 118C and TP53 72Arg polymorphisms were associated with increased risks of platinum-induced nephrotoxicity. Other risk factors found included the platinum type, baseline serum creatinine (Scr), coadministration of vinorelbine, and the number of chemotherapy cycles. The overall prediction rate of the CART was 82.7%, with a sensitivity of 0.630 and specificity of 0.896. The Framingham study risk prediction model contained 7 factors. Its prediction rate was 84.5%, with a sensitivity of 0.643 and specificity of 0.909. Conclusions. Genetic polymorphisms of ERCC1 and TP53 are risk factors for nephrotoxicity. The CART analysis may provide a clinically applicable model to predict the risk of cisplatin- and carboplatin-induced nephrotoxicity. Unfortunately, these platinum-containing chemotherapies can induce severe nephrotoxicity, which limits their usage. Nephrotoxicity occurs in approximately one-third of cisplatin-treated patients, even after aggressive hydration . Carbopltoxicity , 4. Othe ERCC1 C118T was associated with a higher risk of developing cancer and negative clinical outcomes in lung cancer [ ERCC1 C118T polymorphism and platinum-induced nephrotoxicity is still inconclusive [Platinum binds to DNA to form platinum-DNA adducts, which induce cell death. Both tumor and renal cells are able to uptake large amounts of platinum. The concentration of cisplatin in proximal tubular cells was reported to be 5 times higher than that in serum . The toxg cancer , 11. Hownclusive , 11–13.As the gatekeeper of cell cycle or death, platinum-induced apoptosis is strongly regulated by tumor suppressor protein 53 (TP53) . The impThe goals of this study were to examine the roles of ERCC1 and TP53 polymorphisms in platinum-induced nephrotoxicity and develop a prediction model which can identify patients susceptible to platinum-induced nephrotoxicity. The relationships of genetic and nongenetic factors with cisplatin- and carboplatin-induced nephrotoxicity were first assessed by a multivariable regression. Given that platinum-induced nephrotoxicity involves multiple factors and results from multivariable regressions are not easily explainable clinically, two other classification methods were also evaluated. The classification and regression tree (CART) analysis is a nonparametric statistical method, which manages multiple categorical and continuous variables at the same time to generate a tree-shaped classification model . The CARThis study recruited lung cancer patients who were admitted to Wan Fang Hospital, Taipei Medical University, between January 2005 and March 2011. Patients who had received more than two cycles of cisplatin- or carboplatin-containing chemotherapy for lung cancer and were aged 12~100 years at the time of diagnosis were eligible. Patients who were pregnant or infected by the human immunodeficiency virus (HIV) were excluded. Because of the known risk of nephrotoxicity, patients who were coadministrated ifosfamide with cisplatin or carboplatin were also excluded . PeriphePatients who met any level of risk of renal dysfunction, injury to the kidney, failure of kidney function, loss of kidney function, or end-stage kidney disease (RIFLE) criteria were defined as having cisplatin- or carboplatin-induced nephrotoxicity in this study . An Scr DNA from peripheral blood mononuclear cells was extracted by proteinase K digestion followed by the conventional phenol-chloroform method as previously described . TP53 codon 72 (rs1042522) were determined by polymerase chain reaction-restriction fragment-length polymorphism (PCR-RFLP). The PCR was performed in a total volume of 25 μL, containing 0.2 mM dNTPs , 1 mM MgCl2 (Protech), 1x Taq buffer (Mg+-free) (Protech), 0.04 U Taq polymerase (Protech), 2 μL primers (Protech), and 1 μL DNA. Primers were 5′-GAAGACCCAGGTCCAGATGA-3′ (forward) and 5′-ACTGACCGTGCAAGTCACAG-3′ (reverse). Amplification was carried out under the following conditions: 1 cycle of 94°C for 3 min, followed by 35 cycles of 94°C for 30 s, 55°C for 30 s, and 72°C for 30 s, and 1 cycle of 72°C for 10 min, followed by cooling down to 4°C. The PCR product of TP53 codon 72 was digested with BstUI (New England Biolabs), separated on a 2% agarose gel, and visualized by ethidium bromide staining.Genotypes of ERCC1 codon 118 (rs11615) were determined by a 5′ nuclease assay (TaqMan). Each PCR mixture with a volume of 20 μL contained 2x TaqMan Master Mix (Applied Biosystem), 40x TaqMan SNP Genotyping Assay Mix (Applied Biosystem), and 0.5 μL DNA. Amplification conditions were 1 cycle of 95°C for 10 min, followed by 40 cycles of 92°C for 15 s and 60°C for 1 min. The fluorescence detection and PCR were carried out in an ABI Prism 7300 (Applied Biosystem).Genotypes ofFor each SNP, 20 randomly selected samples (17%) were selected to be genotyped by direct sequencing in the ABI Prism 3100 (Applied Biosystem) again to validate the accuracies of the PCR-RFLP and 5′ nuclease assay. The accuracies of both methods were 100%.P < 0.05 was considered statistically significant in this study. The Hardy-Weinberg equilibrium of alleles at individual loci was evaluated. SPSS version 16.0  for Windows was used to analyze data. Chi-squared and Fisher's exact tests were used to analyze ordinal and categorical data, and Student's t-test was used for continuous data. A multivariable logistic regression was applied to calculate the adjusted odd ratios (ORs) and 95% confidence intervals (CIs) of specific genotypes or variables of interest after adjusting for age, gender, body weight, alcohol consumption, smoking, baseline Scr, treatment regimen, number of chemotherapy cycles, and cancer histology.A two-tailed The classification and regression tree (CART) and nephrotoxicity risk score system were performed to establish the most useful model to predict nephrotoxicity. Then the sensitivity, specificity, and total prediction rate of the CART and risk score models were compared. The Hosmer and Lemeshow method was also applied by using variables included in the CART and Framingham models to examine the goodness-of-fit with the Akaike information criterion (AIC) as an adjuvant parameter. Smaller values of the AIC indicate better models. Gini index was used as a splitting criterion to stratify the data into various risk subgroups with maximum homogeneity to build the model. Tree-building continued until the terminal nodes had no statistically significant splits or reached a minimum size of 10 subjects for each terminal node. The ORs and 95% CIs of the terminal nodes were calculated using a logistic regression.The CART based on binary recursive partitioning was used to explore gene-gene and gene-environment interactions . Data weP < 0.1) and biological relevance, such as smoking [A nephrotoxicity risk score was established according to the statistical analysis method in the Framingham study, which used a point system to predict a patient's risk . Variabl smoking . The ini smoking .P = 0.034).Demographic characteristics of the 28 (24.1%) cases and 88 (75.9%) controls are listed in P > 0.1) were coadministration of vinorelbine, the number of chemotherapy cycles, and polymorphisms of ERCC1 C118T and TP53 Arg72Pro between cases and controls.Alleles at individual loci fulfilled a Hardy-Weinberg distribution in both cases and controls . In the In an effort to transform gene-gene and gene-environment relationships to prediction systems that are convenient to use clinically, variables of characteristics and genetic polymorphisms were analyzed by two different statistical models.The first evaluated model used a CART analysis. The decision tree of the CART is shown in  TP53 Pro allele, and ERCC1 C/C and subjects in node 5 with the TP53 Arg/Arg genotype were the subgroups most susceptible to nephrotoxicity. The case ratios were 71.4% and 63.6%, respectively. In contrast, subjects who received carboplatin (node 2) and those who received cisplatin with baseline Scr values of >1 mg/dL (node 4) had a lower nephrotoxicity risk, with respective case ratios of 9.8% and 8.3%. No interaction was found in patients treated with carboplatin due to a limited number of subjects with nephrotoxicity.Subjects in node 8 who had received cisplatin with baseline Scr values of ≤1 mg/dL, then = 86/104). Youden's index, the sensitivity, and specificity were 0.499, 0.630 (n = 17/27), and 0.869 (n = 69/77), respectively. Using the four variables identified by the CART, including the platinum type, baseline Scr, TP53, and ERCC1, the AIC value was 109.714.The overall prediction rate of the CART model was 82.7% (n = 98/116). The sensitivity and specificity for each score are shown in The second prediction model evaluated in this study was the risk score. The best model of the nephrotoxicity risk score is shown in In the present study, we investigated the association of ERCC1 and TP53 gene polymorphisms with the risk of cisplatin- or carboplatin-induced nephrotoxicity by logistic regression and multifactor analytical approaches. No significant association of TP53 or ERCC1 with nephrotoxicity was detected by the multivariable regression. However, two multifactor analytical approaches generated different results.2/wk). There are several possible explanations. It is possible that the renal function of patients with a baseline Scr of ≤1 mg/dL was usually overestimated. The cisplatin dose used in these patients may have been too high. Another possible reason may be inherent, such as TP53 and ERCC1, and can be explained by sequent nodes in the CART model. The relationship between baseline Scr and cisplatin-induced nephrotoxicity needs to be explored in future studies.Cisplatin, with 30% prevalence of nephrotoxicity, is known to be more nephrotoxic than carboplatin . TherefoTP53 is an important protein that regulates cell cycle arrest and cell death, and variants with attenuated activity may therefore be less susceptible to drug-induced cell apoptosis . Many inNotably, an interesting finding of the TP53 polymorphism in our current CART model was that subjects with the TP53 mutation could be divided into two groups by the ERCC1 polymorphism. One group carrying the ERCC1 mutation had mildly increased Scr, but patients carrying the ERCC1 wild-type had obviously increased Scr. This phenomenon was also observed in an animal study reported by Wei et al. . After cAccording to the nephrotoxicity risk score, simultaneously considering both genetic (TP53 and ERCC1) and nongenetic factors  can better predict the occurrence of platinum-induced nephrotoxicity than either type of factors alone. It was reported by de Jongh et al. that smoking may be a risk factor for cisplatin-induced nephrotoxicity . HoweverBoth CART and risk score models demonstrated similar excellent predictive abilities . Although the prediction rate of the risk score model was slightly higher than the CART model, more factors were used in the risk score than in the CART model (7 versus 4). Taking clinical usefulness into consideration, the CART model, containing fewer factors and a clinically similar prediction rate, might possess better clinical convenience than the risk score model.Our CART model included only 4 factors  but still had a high overall prediction rate, sensitivity, and specificity . The CART analysis was applied to explore multifactor risks for various cancers, such as bladder cancer and oral premalignant lesion –23, 25. Our study has several limitations. First of all, the sample size of our study was small. Some risk factors might not have been picked up by the statistical analysis due to the limited number of cases. For example, there were only 28 cases in 116 subjects and only 11 females with nephrotoxicity among the cases. Thus, sexual differences in the genetic effects might not have been indicated if they existed. Additionally, the range of the distribution of patient age was very wide, with 17 patients with more than 60 years. The small sample size also resulted in a large 95% CI in terminal nodes of the CART model that was split by 4 factors. In addition, no interaction was found in patients who received carboplatin because of a limited number of subjects who developed nephrotoxicity. Second, most of the patients (more than 70%) were subjected to combined treatment, such as cisplatin with gemcitabine, which can interfere in the nephrotoxicity. However, there was no statistically significant difference in the percentages of patients who were coadministered with gemcitabine in the case and control groups . Third,  TP53 Arg72Pro and ERCC1 C118T play roles in cisplatin-induced nephrotoxicity. The study also demonstrated that classification statistical methods, such as the CART and risk score models, may be useful for evaluating risks of platinum-induced nephrotoxicity and develop dosing regimens for lung cancer patients. The results of this study need to be validated by future studies.To sum up, the present study indicated that