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Reference limits are estimators for ‘extreme’ percentiles of the distribution of a quantitative diagnostic marker in the healthy population. In most cases, interest will be in the 90% or 95% reference intervals. The standard parametric method of determining reference limits consists of computing quantities of the form X ̅± c · S . The proportion of covered values in the underlying population coincides with the specificity obtained when a measurement value falling outside the corresponding reference region is classified as diagnostically suspect. Nonparametrically, reference limits are estimated by means of so-called order statistics. In both approaches, the precision of the estimate depends on the sample size. We present computational procedures for calculating minimally required numbers of subjects to be enrolled in a reference study. The much more sophisticated concept of reference bands replacing statistical reference intervals in case of age-dependent diagnostic markers is also discussed.

Research into lipoprotein metabolism has developed because understanding lipoprotein metabolism has important clinical indications. Lipoproteins are risk factors for cardiovascular disease. Recent advances include the identification of factors in the synthesis and secretion of triglyceride rich lipoproteins, chylomicrons (CM) and very low density lipoproteins (VLDL). These included the identification of microsomal transfer protein, the cotranslational targeting of apoproteinB (apoB) for degradation regulated by the availability of lipids, and the characterization of transport vesicles transporting primordial apoB containing particles to the Golgi. The lipase maturation factor 1, glycosylphosphatidylinositol-anchored high density lipoprotein binding protein 1 and an angiopoietin-like protein play a role in lipoprotein lipase (LPL)-mediated hydrolysis of secreted CMs and VLDL so that the right amount of fatty acid is delivered to the right tissue at the right time. Expression of the low density lipoprotein (LDL) receptor is regulated at both transcriptional and post-transcriptional level. Proprotein convertase subtilisin/kexin type 9 (PCSK9) has a pivotal role in the degradation of LDL receptor. Plasma remnant lipoproteins bind to specific receptors in the liver, the LDL receptor, VLDL receptor and LDL receptor-like proteins prior to removal from the plasma. Reverse cholesterol transport occurs when lipid free apoAI recruits cholesterol and phospholipid to assemble high density lipoprotein (HDL) particles. The discovery of ABC transporters (ABCA1 and ABCG1) and scavenger receptor class B type I (SR-BI) provided further information on the biogenesis of HDL. In humans HDL-cholesterol can be returned to the liver either by direct uptake by SR-BI or through cholesteryl ester transfer protein exchange of cholesteryl ester for triglycerides in apoB lipoproteins, followed by hepatic uptake of apoB containing particles. Cholesterol content in cells is regulated by several transcription factors, including the liver X receptor and sterol regulatory element binding protein. This review summarizes recent advances in knowledge of the molecular mechanisms regulating lipoprotein metabolism.

Rheumatoid arthritis, the most prominent of systemic autoimmune rheumatic diseases, represents an important social health problem. Recent insights into the immunopathogenic mechanism of this complex and multiform illness might open new perspectives for a more appropriate laboratory approach. In this review we focus on the most relevant pathogenetic mechanism; indicating the laboratory biomarkers specifically linked to early diagnosis, prognosis, evolutive aspects of the disease, and therapeutic efficacy. Evidence based on laboratory medicine could provide the best outcome for patients.

Background: Early notification of critical values by the clinical laboratory to the treating physician is a requirement for accreditation and is essential for effective patient management. Many laboratories automatically repeat a critical value before reporting it to prevent possible misdiagnosis. Given today’s advanced instrumentation and quality assurance practices, we questioned the validity of this approach. We performed an audit of repeat-testing in our laboratory to assess for significant differences between initial and repeated test results, estimate the delay caused by repeat-testing and to quantify the cost of repeating these assays. Methods: A retrospective audit of repeat-tests for sodium, potassium, calcium and magnesium in the first quarter of 2013 at Tygerberg Academic Laboratory was conducted. Data on the initial and repeat-test values and the time that they were performed was extracted from our laboratory information system. The Clinical Laboratory Improvement Amendment criteria for allowable error were employed to assess for significant difference between results. Results: A total of 2308 repeated tests were studied. There was no significant difference in 2291 (99.3%) of the samples. The average delay ranged from 35 min for magnesium to 42 min for sodium and calcium. At least 2.9% of laboratory running costs for the analytes was spent on repeating them. Conclusions: The practice of repeating a critical test result appears unnecessary as it yields similar results, delays notification to the treating clinician and increases laboratory running costs.

Background: The Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) equation seems to correct the overdiagnosis of chronic kidney disease (CKD) provided by Modification of Diet in Renal Disease (MDRD) equation. However, this point has not been tested in some ethnic groups. This study investigated the performance of MDRD and CKD-EPI equations in South Brazilian individuals. Methods: This cross-sectional study included 354 individuals including healthy volunteers, diabetic and non-diabetic individuals with or without CKD. Glomerular filtration rate (GFR) was measured by the 51 Cr-EDTA single-injection method ( 51 Cr-GFR). Accuracy (P30), bias, and Bland-Altman agreement plots were evaluated. Results: In the group as a whole, 51 Cr-GFR was 87±37 (6-187), CKD-EPI eGFR, 82±30 (6-152), and MDRD eGFR, 77±28 (6-156) mL/min/1.73 m 2 (p<0.001 for all comparisons). Analyzing the subset of individuals with 51 Cr-GFR <60 mL/min/1.73 m 2 , P30 values were, respectively, 76% and 84% for MDRD and for CKD-EPI (p<0.001) while for 51 Cr-GFR ≥60 mL/min/1.73 m 2 , P30 values were 57.5% for both equations (p=1.000). For MDRD and CKD-EPI, mean bias were negative for GFRs <60 (–11 vs. –12, p=0.221) and positive for values >60 (16 vs. 9, p<0.001). In multivariate analysis, absolute bias was unfavorably influenced by measured GFR >60 (for MDRD) and being diabetic or younger (for CKD-EPI). Conclusions: CKD-EPI reduces GFR underestimation in individuals with GFRs >60, but still presents a quite low accuracy at this GFR range. Moreover, it tends to overestimate GFR in subjects with GFRs <60 mL/min/1.73 m 2 . CKD stages 1 and 2, diabetes and young age had a negative influence on the performance of the equations.

Background: In chronic kidney diseases of various etiologies, the urinary excretion of uromodulin is usually decreased in parallel with the glomerular filtration rate. This study aimed to investigate whether serum uromodulin is associated with kidney function. Methods: Within the framework of the Seniorlabor study, a subset of subjectively healthy individuals 60 years of age and older were included in the study. Serum uromodulin was measured with ELISA. The relationship between serum uromodulin and different stages of kidney function (i.e., cystatin C-based 2012-CKD-EPI eGFR CysC >90 mL/min/1.73 m 2 , 60–89 mL/min/1.73 m 2 , 45–59 mL/min/1.73 m 2 , and <45 mL/min/1.73 m 2 ) was investigated. Furthermore, the relationship between serum uromodulin and other markers of kidney function (i.e., creatinine, cystatin C, and urea) was assessed. Results: In total, 289 participants (140 males/149 females; mean age 71±7 years) were included in the study. There were significant differences in serum uromodulin among the four groups according to different kidney function stages (p<0.001). Serum uromodulin displayed inverse relationships with creatinine (r=–0.39), cystatin C (r=–0.42), and urea (r=–0.30) and, correspondingly, a positive relationship with eGFR CysC (r=0.38, p<0.001 for all). These associations remained intact when fitting a regression model that incorporated age, gender, body mass index, and current smoking status as covariates. Conclusions: Serum uromodulin behaves in a manner opposite that of the different conventional renal retention markers by displaying lower concentrations with decreasing kidney function. As uromodulin is produced by the cells of the thick ascending limb of the loop of Henle, lower uromodulin serum levels may reflect a reduction in number or function of these cells in chronic kidney disease.

Background: Peripheral smear review is a critical, but labor intensive adjunct for evaluation of lymphocytosis. Standard practice based on consensus guidelines is to review cases with absolute lymphocyte count (ALC) >5×10 9 /L. We hypothesize that identifying cases for review by applying appropriately adjusted ALC and age discriminators will decrease laboratory workload without compromising patient care. Methods: 1170 complete blood counts with ALCs >5×10 9 /L analyzed in the core laboratory during a 2-year period were included. Patients were categorized into diagnostic groups based on follow-up criteria. A total of 402 patients with new onset lymphocytosis who met criteria for reactive lymphocytosis (82%) or lymphoproliferative disorder (18%) were used to establish optimal ALC and age thresholds from receiver operating characteristic (ROC) curve analysis. Results: ALC as a discriminator for neoplastic lymphocytosis had an ROC area under the curve (AUC) of 0.732. Selecting cases with ALC >10×10 9 /L enriched the proportion of neoplastic cases in the review pool (90% specificity); however, many cases with ALC below this threshold were also neoplastic (52% sensitivity). For cases with ALC between 5 and 10×10 9 /L, age as a discriminator had an ROC AUC of 0.886. Selecting patients >50 years old in this group for review captured the neoplastic cases while excluding the reactive cases (93% sensitivity, 62% specificity). When applied to a validation cohort, the predictive performance of the thresholds was maintained while reducing smears reviewed by 50%. Conclusions: We show that modifying the standard 5×10 9 /L ALC smear review threshold through retrospective analysis of institutional data can reduce laboratory workload without compromising quality.

Background: The Sysmex XN (XN) modular system (Sysmex, Kobe, Japan) is a new automated hematology analyzer equipped with different principles from its previous version, Sysmex XE-2100. We compared the performances of Sysmex XN and XE-2100 in umbilical cord blood (CB) specimens. Methods: In 160 CB specimens, complete blood count (CBC) parameters and white blood cells (WBC) differentials were compared between the two analyzers. Their flagging performances for blasts, abnormal/atypical lymphocytes, immature granulocytes and/or left-shift (IG), and nucleated red blood cells (NRBC) counts were compared with manual counts. For the blast flagging, Q values by Sysmex XN were further compared with manual slide review. Results: Sysmex XN and XE-2100 showed high or very high correlations for most CBC parameters but variable correlations for WBC differentials. Compared with XE-2100, XN showed significantly different flagging performances for blasts, abnormal/atypical lymphocytes, and IG. The flagging efficiency for blasts was significantly better on Sysmex XN than on XE-2100 (85.0% vs. 38.8%): Sysmex XN showed a remarkably increased specificity of blast flag, compromising its sensitivity of blast flag. Among the 24 specimens with blasts (range, 0.5%–1.5%), only one (4.2%) showed a positive Q value. Conclusions: This study highlighted the remarkable differences of flagging performances between Sysmex XN and XE-2100 in CB specimens. The Sysmex XN modular system seems to be a suitable and practical option for the CB specimens used for hematopoietic stem cell transplantation as well as for the specimens from neonates.

Background : We evaluated the new body fluid mode on the UF-1000 i urinalysis analyzer for counting total white blood cells (WBC) and red blood cells (RBC) in continuous ambulatory peritoneal dialysis (CAPD), ascites and pleural fluids. Methods : We collected 154 body fluid samples, and compared the results of the UF-1000 i BF mode with the Fuchs-Rosenthal counting chamber and the XN-1000 BF mode. Linearity, carry over and precision were also assessed. Results : Method comparison results showed acceptable WBC agreement between UF-1000 i and chamber (y=1.27x+3.13, n=135, r=0.99) and between UF-1000 i and XN (y=1.15x+0.31, n=135, r=1.00). Comparison between the UF-1000 i and both comparison methods showed good agreement for RBC counts. Overall results were better when UF-1000 i was compared with the XN-1000 than with the Fuchs-Rosenthal chamber. The lower limit of quantitation was defined at 9×10 6 WBC/L and at 25×10 6 RBC/L. Linearity for both WBC (r=1.00) and RBC (r=0.99) was good. Carry over was negligible, and it never exceeded 0.01%. In one sample, a high discrepancy was observed between WBC results for both automated analyzers and the counting chamber. This discrepancy was due to interfering factors, such as bacteria and yeast cells, and it led to a false increased WBC count on both automated systems. Conclusions : The UF-1000 i BF mode offers rapid and reliable total WBC and RBC counts for initial screening of CAPD, ascites and pleural fluid, and can improve the workflow in a routine laboratory; however, when using automated analyzers, the inspection of scattergrams is required to ensure the most accurate results are obtained.

Background: Sickle cell disease is a major health burden in India. The aim of the study was to compare the diagnostic utility of two different approaches on automated high performance liquid chromatography (HPLC) for newborn screening for sickle cell disorders and other haemoglobinopathies in India. Methods: Newborn babies of sickle heterozygous mothers were tested by HPLC using two different kits, the β-thal short kit, which is routinely used for screening for haemoglobinopathies in most laboratories, and the sickle cell short kit which is specific only for neonatal samples. Confirmation of the sickle and α genotypes was done by molecular analysis. Results: Of the 601 babies tested, 276 were normal, 284 were sickle heterozygous and 41 were sickle homozygous using the β-thal short kit. Using the sickle cell short kit, a discrepancy was seen in one newborn sample where a normal baby was identified as a sickle heterozygous baby. α-Genotyping was done in 42 babies and 16 of them had α gene deletions. The presence of α thalassaemia could be suspected in 15 of these 16 babies based on a spike at the start of the chromatogram using the β-thal short kit. In comparison, using the sickle cell short kit the diagnosis of α thalassaemia was difficult based on the percentage of the FAST peak. Further, other rare α chain Hb variants were also missed. Conclusions: The β-thal short kit was more versatile than the sickle cell short kit for screening for haemoglobinopathies in newborns in our population.

Background: Antithrombin (AT) is a slow-acting progressive inhibitor of activated clotting factors, particularly thrombin and activated factor X (FXa). However, the presence of heparin or heparan sulfate accelerates its effect by several magnitudes. AT deficiency, a severe thrombophilia, is classified as type I (quantitative) and type II (qualitative) deficiency. In the latter case mutations may influence the reactive site, the heparin binding-site (HBS) and exert pleiotropic effect. Heterozygous type II-HBS deficiency is a less severe thrombophilia than other heterozygous subtypes. However, as opposed to other subtypes, it also exists in homozygous form which represents a very high risk of venous thromboembolism. Methods: A modified anti-FXa chromogenic AT assay was developed which determines both the progressive (p) and the heparin cofactor (hc) activities, in parallel. The method was evaluated and reference intervals were established. The usefulness of the assay in detecting type II-HBS AT deficiency was tested on 78 AT deficient patients including 51 type II-HBS heterozygotes and 18 homozygotes. Results: Both p-anti-FXa and hc-anti-FXa assays showed excellent reproducibility and were not influenced by high concentrations of triglyceride, bilirubin and hemoglobin. Reference intervals for p-anti-FXa and hc-anti-FXa AT activities were 84%–117% and 81%–117%, respectively. Type II-HBS deficient patients demonstrated low (heterozygotes) or very low (homozygotes) hc-anti-FXa activity with normal or slightly decreased p-anti-FXa activity. The p/hc ratio clearly distinguished wild type controls, type II-HBS heterozygotes and homozygotes. Conclusions: Concomitant determination of p-anti-FXa and hc-anti-FXa activities provides a reliable, clinically important diagnosis of type II-HBS AT deficiency and distinguishes between homozygotes and heterozygotes.

Background: Previous studies with mainly selected populations have proposed contradicting reference ranges for thyroid-stimulating hormone (TSH) and have disagreed on how screening, age and gender affect them. This study aimed to determine a TSH reference range on the Abbott Architect ci8200 integrated system in a large, nationwide, stratified random sample. To our knowledge this is the only study apart from the NHANES III that has addressed this issue in a similar nationwide setting. The effects of age, gender, thyroid peroxidase antibody (TPOAb)-positivity and medications on TSH reference range were also assessed. Methods: TSH was measured from 6247 participants randomly drawn from the population register to represent the Finnish adult population. TSH reference ranges were established of a thyroid-healthy population and its subpopulations with increasing and cumulative rigour of screening: screening for overt thyroid disease (thyroid-healthy population, n=5709); screening for TPOAb-positivity (risk factor-free subpopulation, n=4586); and screening for use of any medications (reference subpopulation, n=1849). Results: The TSH reference ranges of the thyroid-healthy population, and the risk factor-free and reference subpopulations were 0.4–4.4, 0.4–3.7 and 0.4–3.4 mU/L (2.5th–97.5th percentiles), respectively. Although the differences in TSH between subgroups for age (p=0.002) and gender (p=0.005) reached statistical significance, the TSH distribution curves of the subgroups were practically superimposed. Conclusions: We propose 0.4–3.4 mU/L as a TSH reference range for adults for this platform, which is lower than those presently used in most laboratories. Our findings suggest that intensive screening for thyroid risk factors, especially for TPOAb-positivity, decreases the TSH upper reference limit.

Background: β-Trace protein (BTP) has been proposed as an alternative endogenous marker of glomerular filtration rate. Data on BTP reference ranges in young children are scarce. We therefore aim to establish reference ranges and examine the developmental course of serum BTP in basically healthy children younger than 1 year of age. Methods: Single blood samples were taken from healthy children (born at gestational age ≥37 weeks) <12 months of age. Serum BTP was measured using the N latex B-trace protein assay (Siemens Diagnostics, Deerfield, IL, USA) on an Immage ® 800 Rate Nephelometer (Beckman Coulter Inc. Brea, CA, USA). Serum creatinine and cystatin C were additionally determined and compared to reference values to confirm a normal renal function. Results: From June 2010 to January 2014, 95 blood samples were collected from 95 children {67.4% male; median age 120 days [inter quartile range 57–166]}. BTP was normally distributed (mean concentration 0.84±standard deviation 0.35 mg/L). Considering all children, the 50th centile BTP reference concentration was 0.82 mg/L (5th–95th centiles; 0.27–1.38). BTP concentrations were the highest in neonates and steadily declined with increasing age (Spearman’s rank correlation was –0.415, p=0.002). No gender differences were found. Conclusions: Our data provide a BTP reference range for the first year of life. Seeing the biological pattern of BTP, with only a limited postnatal decline, this marker might offer a promising alternative to serum creatinine-based methods for estimating glomerular filtration rate in newborns.

Background: A nationwide multicenter study was organized to establish reference intervals (RIs) in the Turkish population for 25 commonly tested biochemical analytes and to explore sources of variation in reference values, including regionality. Methods: Blood samples were collected nationwide in 28 laboratories from the seven regions (≥400 samples/region, 3066 in all). The sera were collectively analyzed in Uludag University in Bursa using Abbott reagents and analyzer. Reference materials were used for standardization of test results. After secondary exclusion using the latent abnormal values exclusion method, RIs were derived by a parametric method employing the modified Box-Cox formula and compared with the RIs by the non-parametric method. Three-level nested ANOVA was used to evaluate variations among sexes, ages and regions. Associations between test results and age, body mass index (BMI) and region were determined by multiple regression analysis (MRA). Results: By ANOVA, differences of reference values among seven regions were significant in none of the 25 analytes. Significant sex-related and age-related differences were observed for 10 and seven analytes, respectively. MRA revealed BMI-related changes in results for uric acid, glucose, triglycerides, high-density lipoprotein (HDL)-cholesterol, alanine aminotransferase, and γ-glutamyltransferase. Their RIs were thus derived by applying stricter criteria excluding individuals with BMI >28 kg/m 2 . Ranges of RIs by non-parametric method were wider than those by parametric method especially for those analytes affected by BMI. Conclusions: With the lack of regional differences and the well-standardized status of test results, the RIs derived from this nationwide study can be used for the entire Turkish population.

Background: Low circulating homoarginine as well as high levels of asymmetric dimethylarginine (ADMA) and symmetric dimethylarginine (SDMA) have been associated with impaired cardiovascular (CV) outcome and mortality in patients at risk and in the general population. The present study aimed to formulate reference intervals for serum homoarginine, ADMA, and SDMA to improve risk stratification between healthy individuals and individuals at risk. Methods: We determined age- and sex-specific reference intervals for homoarginine, ADMA, and SDMA in a subgroup of 1359 healthy participants (no diabetes mellitus, CV disease, increased blood pressure, elevated blood lipids, chronic kidney disease stadium III or IV, or a body mass index >30) of the Study of Health in Pomerania (SHIP) using quantile regression analyses. Homoarginine, ADMA, and SDMA serum concentrations were measured using liquid chromatography-tandem mass spectrometry. Results: Median age of the investigated cohort was 36 (25th; 75th percentile 28; 47) years, with 62% women. Median serum concentrations of homoarginine, ADMA, and SDMA were 2.63 (2.08; 3.32) μmol/L, 0.64 (0.57; 0.73) μmol/L, and 0.43 (0.37; 0.49) μmol/L, respectively. Serum levels of homoarginine, ADMA, and SDMA showed material age- and sex-related differences (p<0.05 for all). Overall reference ranges were 1.41–5.00 and 1.20–5.53 μmol/L (2.5th; 97.5th percentile; for men and women, respectively) for homoarginine, 0.41–0.95 and 0.43–0.96 μmol/L for ADMA, and 0.30–0.67 and 0.27–0.63 μmol/L for SDMA. Conclusions: We formulated for the first time homoarginine, ADMA, and SDMA reference intervals in serum. These reference intervals might be useful for individual CV risk stratification.

Background : Aberrant expression of SRY-box containing gene 17 ( SOX17 ) has been observed in several solid tumors. However, little is known about SOX17 expression in acute myeloid leukemia (AML). The purpose of this study was to investigate the alteration of SOX17 expression and to explore its clinical significance in AML. Methods : Real-time quantitative PCR (RQ-PCR) was performed to analyze the status of SO1X17 expression in 103 patients with de novo AML and 26 normal controls. The clinical relevance of SOX17 expression was analyzed in AML. Results : SOX17 level in AML was significantly down-regulated compared to controls (p<0.001). Receiver operating characteristic curve (ROC) curve analysis revealed that an area under the ROC curve (AUC) of 0.834 (95% CI 0.765–0.903; p<0.0001) or 0.789 (95% CI 0.690–0.888, p<0.001) in discriminating all patients or cytogenetically normal patients from controls, respectively. The cohort of AML patients was divided into two groups according to the cut-off value of 0.017 (60% sensitivity and 100% specificity, respectively). Cytogenetically normal patients with low SOX17 expression had significantly shorter OS than those with high SOX17 expression (median 4 vs. 25 months, respectively, p=0.035). Multivariate analysis confirmed low SOX17 expression as an independent risk factor. Conclusions: Our findings indicated that low SOX17 level may define an important risk factor in AML with normal cyotgenetics.

Background: Laboratory testing for KRAS (v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog) mutations in metastatic colorectal cancer (mCRC) is performed by various methods in China, but there is no standardized system for proficiency testing or assay performance evaluations. The aim of this study was to evaluate assay and laboratory performance with artificial samples derived from formalin-fixed, paraffin-embedded (FFPE) cell lines. Methods: Artificial FFPE samples were prepared from cultured cell lines to construct a proficiency panel of 10 samples covering eight KRAS mutations and two wild-type samples. The samples were validated by Sanger sequencing and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). The panel was distributed to participating laboratories and their reported results were compared to the reference sequences. Results: The percentages of mutant KRAS alleles in each mutant sample were more than 50% by MALDI-TOF-MS. Sixty-three laboratories reported results, including 41 hospital laboratories and 22 commercial laboratories and reagent manufacturers. Only 55.6% (35/63) of the laboratories correctly identified the mutations in all samples and 33.3% (21/63) reported at least one false-positive result. The false-positive ratio was 7.1% (45/630) and the false-negative ratio was 3.0% (15/504). Conclusions: KRAS mutations can be missed even by the most sensitive methods if the procedures are not performed correctly. False-positive results are a substantial problem in KRAS testing; laboratories must use sufficient negative controls to identify cross-contamination from PCR-amplified products or between samples during handling and DNA extraction.

Background: CTNNA1 gene is a putative tumor suppressor for its roles in inhibiting proliferation and promoting apoptosis. Aberrant expression of CTNNA1 is regulated by epigenetic mechanisms including both promoter methylation and histone deacetylation in hematopoietic malignancies. However, the clinical relevance of CTNNA1 methylation remains rarely known in myelodysplastic syndrome (MDS). Methods : We investigated the methylation status of CTNNA1 promoter using methylation-specific PCR (MSP) and analyzed its clinical significance in Chinese MDS patients. Results: Aberrant hypermethylation of CTNNA1 gene was identified in 22% (18/83) of the patients. CTNNA1 expression was significantly correlated with promoter methylation status (p<0.05). No significant differences were observed in the age, sex, and blood parameters between patients with and without CTNNA1 hypermethylation (p>0.05). The frequency of CTNNA1 hypermethylation was significantly higher in patients with isolated del(5q) (3/4, 75%) than those with other abnormal karyotypes (4/23, 17%) and also than those with normal karyotypes (11/54, 20%) (p=0.042 and 0.040, respectively). The patients with higher IPSS risks (Int-2/High) had significantly increased incidence of CTNNA1 methylation than those with lower risks (Low/Int-1) (36% vs. 15%, p=0.049). Although the estimated 50% survival time of the CTNNA1 -methylated group [median 13 months, 95% confidence interval (CI) 3–22 months] was shorter than that of CTNNA1 -unmethylated group (median 24 months, 95% CI 7–41 months), the difference was not statistically significant (p=0.330). Conclusions: Our data confirm that aberrant CTNNA1 methylation is a common event and is associated with higher IPSS risk in MDS.

Background: The small conductance calcium-activated potassium, subfamily N, member 3 ( KCNN3 ) gene rs13376333 and rs1131820 have been shown to be strongly associated with lone atrial fibrillation (AF), while replication association studies between rs13376333 in KCNN3 gene and risk of AF showed conflicting results. The current study tried to validate the impact of SNP rs13376333 and rs1131820 of KCNN3 gene on the risk of AF in the Chinese Han population. Methods: A total of 889 AF patients and 1015 controls were enrolled. Two hundred and seventy-eight cases of AF were lone AF. KCNN3 gene SNP rs13376333 and rs1131820 were genotyped by allele-specific MALDI-TOF mass spectrometry. Results: The genotype distribution and allele frequency of rs13376333 polymorphism were not different between total AF patients and controls. However, the genotype distribution of rs13376333 polymorphism was significantly different between lone AF and control group (p<0.001); and T allele frequency was significantly higher in lone AF group than that in controls (7.6% vs 3.6%, p<0.001). Multivariable logistic regression analysis showed that T allele carriers of rs13376333 was significantly associated with lone AF (OR=2.31, 95% CI 1.41–3.78, p=0.001). No relationship between rs1131820 polymorphism and total AF or lone AF was found in this study. Conclusions: KCNN3 rs13376333 polymorphism was associated with lone AF in the Chinese Han population and the T allele carriers may be an independent predictive factor for lone AF.