康柏西普联合激光光凝治疗糖尿病黄斑水肿43 例临床分析

目的:探讨康柏西普玻璃体腔注射联合激光光凝治疗糖尿病黄斑水肿的临床疗效及安全性。方法:收取2013年6月至2015年8月间我院收治的糖尿病黄斑水肿患者87例(87眼)作为研究对象,采用随机数字表法将其分为观察组及对照组,观察组43例(43眼)给予康柏西普玻璃体腔注射联合激光光凝治疗,对照组44例(44眼)给予单纯激光光凝治疗。对两组患者治疗前后视力(best corrected visual acuilty,BCVA)、视网膜厚度(central macular thickness,CMT)、临床疗效、并发症以及生活质量进行观察与比较。结果:治疗前两组患者BCVA、CMT均无显著差异,治疗后两组BCVA、CMT均有所提高,同时间点观察组明显高于对照组,差异有统计学意义(P0.05)。观察组治疗后6个月视力提高率为81.40%,对照组为13.64%,观察组远高于对照组,差异有统计学意义(P0.05)。观察组患者共有3例出现一过性眼压升高,对照组共有2例患者出现一过性眼压增高,两组相较无统计学差异(P0.05)。观察组生活质量显著高于对照组,差异有统计学意义(P0.05)。结论:康柏西普玻璃体腔注射联合激光光凝治疗糖尿病黄斑水肿具有良好的疗效及安全性,有利于患者生活质量提高,值得临床推广应用。     

Development of a Semi-Automatic Segmentation Method for Retinal OCT Images Tested in Patients with Diabetic Macular Edema

Purpose

To develop EdgeSelect, a semi-automatic method for the segmentation of retinal layers in spectral domain optical coherence tomography images, and to compare the segmentation results with a manual method.

Methods

SD-OCT (Heidelberg Spectralis) scans of 28 eyes (24 patients with diabetic macular edema and 4 normal subjects) were imported into a customized MATLAB application, and were manually segmented by three graders at the layers corresponding to the inner limiting membrane (ILM), the inner segment/ellipsoid interface (ISe), the retinal/retinal pigment epithelium interface (RPE), and the Bruch''s membrane (BM). The scans were then segmented independently by the same graders using EdgeSelect, a semi-automated method allowing the graders to guide/correct the layer segmentation interactively. The inter-grader reproducibility and agreement in locating the layer positions between the manual and EdgeSelect methods were assessed and compared using the Wilcoxon signed rank test.

Results

The inter-grader reproducibility using the EdgeSelect method for retinal layers varied from 0.15 to 1.21 µm, smaller than those using the manual method (3.36–6.43 µm). The Wilcoxon test indicated the EdgeSelect method had significantly better reproducibility than the manual method. The agreement between the manual and EdgeSelect methods in locating retinal layers ranged from 0.08 to 1.32 µm. There were small differences between the two methods in locating the ILM (p = 0.012) and BM layers (p<0.001), but these were statistically indistinguishable in locating the ISe (p = 0.896) and RPE layers (p = 0.771).

Conclusions

The EdgeSelect method resulted in better reproducibility and good agreement with a manual method in a set of eyes of normal subjects and with retinal disease, suggesting that this approach is feasible for OCT image analysis in clinical trials.     

Factors Affecting Reading Speed in Patients with Diabetic Macular Edema Treated with Laser Photocoagulation

Purpose

To study the factors that may affect reading speed in patients with diabetic macular edema previously treated with laser photocoagulation.

Methods

Consecutive patients with type II diabetes treated with laser photocoagulation for diabetic macular edema (DME) at least twelve months previously, with best corrected visual acuity of better than 65 letters (approximately 20/40) measured with Early Treatment Diabetic Retinopathy Study (ETDRS) charts were included in this study. Patients previously treated with pan-retinal photocoagulation, vitrectomy, intravitreal steroid or anti-VEGF therapy were excluded. Any other ocular co-morbidities that may influence reading ability such as cataract, glaucoma or macular degeneration were also excluded. All patients were refracted by a certified examiner, the following measurements were collected: best corrected visual acuity (BCVA), contrast sensitivity with Pelli-Robson chart, reading speed with MNREAD chart, microperimetry with Nidek MP1, and central subfield thickness with Zeiss spectral domain optical coherent topography.

Results

The slow reading group had poorer contrast sensitivity (p = 0.001), reduced retinal sensitivity (p = 0.027) and less stable fixation (p = 0.013). Most interestingly the reduced retinal sensitivity findings were driven by the microperimetry value on the right subfield (p = 0.033), (nasal to the fovea in the right eye and temporal to the fovea in the left eye). Multiple linear regression analysis showed that contrast sensitivity is probably the most important factor that affects reading speed (p = 0.001).

Conclusion

Reduced retinal sensitivity after laser treatment is associated with reduced reading speed in patients with diabetic macular edema.     

Analysis of Rare Variants in the C3 Gene in Patients with Age-Related Macular Degeneration

Age-related macular degeneration (AMD) is a progressive retinal disorder affecting over 33 million people worldwide. Genome-wide association studies (GWASs) for AMD identified common variants at 19 loci accounting for 15–65% of the heritability and it has been hypothesized that the missing heritability may be attributed to rare variants with large effect sizes. Common variants in the complement component 3 (C3) gene have been associated with AMD and recently a rare C3 variant (Lys155Gln) was identified which exerts a large effect on AMD susceptibility independent of the common variants. To explore whether additional rare variants in the C3 gene are associated with AMD, we sequenced all coding exons in 84 unrelated AMD cases. Subsequently, we genotyped all identified variants in 1474 AMD cases and 2258 controls. Additionally, because of the known genetic overlap between AMD and atypical hemolytic uremic syndrome (aHUS), we genotyped two recurrent aHUS-associated C3 mutations in the entire cohort. Overall, we identified three rare variants (Lys65Gln (P = 0.04), Arg735Trp (OR = 17.4, 95% CI = 2.2–136; P = 0.0003), and Ser1619Arg (OR = 5.2, 95% CI = 1.0–25; P = 0.05) at the C3 locus that are associated with AMD in our EUGENDA cohort. However, the Arg735Trp and Ser1619Arg variants were not found to be associated with AMD in the Rotterdam Study. The Lys65Gln variant was only identified in patients from Nijmegen, the Netherlands, and thus may represent a region-specific AMD risk variant.     

变应性鼻炎与IRF5的单核苷酸多态性关系

目的:探讨变应性鼻炎与干扰素调节因子5(IRFS)的单核苷酸多态性(SNP)的相关性.方法:采用聚合酶链(PCR)和限制性片段长度多态性(RFL P)方法在新加坡华人中检测110例变应性鼻炎患者及101健康对照组IRF5位点上的3个单核苷酸多态性(rs4728142,rs11770589,rs2280714)计算基因型和等位基因频率.结果:IRF5基因的3个位点的基因型和等位基因频率在变应性鼻组和对照组间差异无统计学意义.结论:新加坡华人IRF5 rs4728142,rsl1770589,rs2280714多态性与变应性鼻炎无明显相关性.     

Study of 27 Aqueous Humor Cytokines in Type 2 Diabetic Patients with or without Macular Edema

The aim of the present study was to compare the changes in the levels of 27 aqueous humor cytokines between diabetic patients with macular edema (ME) and diabetic patients without ME. Undiluted aqueous humor samples were obtained from 68 consecutive type 2 diabetic patients without ME and 56 consecutive type 2 diabetic patients with ME. The concentrations of 27 cytokines in the aqueous humor samples were measured using a multiplex bead immunoassay. Compared with diabetic patients without ME, diabetic patients with ME had significantly higher concentrations of IL-1β, IL-6, IL-8, IP-10, MCP-1, and VEGF in the aqueous humor. However, the concentrations of IL-10 and IL-12 were significantly lower in the diabetic patients with ME. The aqueous humor levels of IL-1β, IL-6, IL-8, MCP-1, IP-10, and VEGF were closely correlated with retinal macular thickness, retinal macular volume and the severity of ME. In addition, the aqueous humor levels of IL-10 and IL-12 decreased with increasing the severity of ME. A variety of cytokines associated with inflammation and angiogenesis may contribute to the pathogenesis of diabetic macular edema, and both anti-inflammatory and antiangiogenic agents should be included in the treatment of ME simultaneously.     

糖尿病性黄斑水肿3D-OCT与mERG检查结果的相关性研究

目的:探讨糖尿病性黄斑水肿的三维光学相干断层扫描技术(3D-OCT)与多焦视网膜电图(m ERG)检查结果的相关性。方法:随机选择糖尿病性黄斑水肿患者40人60只眼,经荧光造影检查示黄斑部有持续性的荧光渗漏、囊样水肿,3D-OCT示黄斑中心凹厚度大于170um,对黄斑中心凹旁上、下、鼻、颞约750um处视网膜神经上皮层处厚度进行OCT自带软件的分析测量。随机选择18人35只年龄相配的正常眼为对照组以同样方法行3D-OCT黄斑旁四个方位的神经上皮厚度测量。两组用同种方法行常规黄斑部m ERG检查。所有数据应用SPSS13.0统计软件进行处理,分析3D-OCT与m ERG在影像学方面的相关性。结果:两组被检查者检查结果经统计学处理后对比发现:被检查者黄斑中央5°、10°区域N1、P1波反应密度绝对值与黄斑中心凹神经上皮层厚度具有显著的相关性(P0.05)。结论:糖尿病性黄斑水肿的神经上皮厚度的改变与黄斑区m ERG的改变具有显著相关性,主要表现为黄斑中央5°和10°区域m ERG各波反应密度的改变,且有神经上皮厚度增加的越大,m ERG各波反应密度的降低的幅度也随之变大的趋势。     

糖尿病白内障术后黄斑水肿与房水细胞因子的关系

为了探讨2型糖尿病白内障术后黄斑水肿与房水中细胞因子的关系,本研究选取2016年2月至2017年7月在我院治疗的2型糖尿病白内障患者117例117眼,均行超声乳化白内障摘除术,观察术后4周黄斑水肿发生情况以及房水中多种细胞因子水平。研究结果显示:术后4周,有37眼发生黄斑水肿(观察组),未发生黄斑水肿80眼(对照组);观察组术后4周黄斑区视网膜厚度为(247.28±18.37)μm,明显高于对照组(p<0.05),且明显高于术前水平(p<0.05);观察组术后4周干扰素诱导蛋白-10 (IP-10)、巨噬细胞趋化蛋白(MCP-1)、血管内皮生长因子(VEGF)和细胞因子转化生长因子-β2 (TGF-β2)水平分别为(6.01±0.89) pg/mL、(340.03±45.39) pg/mL、(812.28±40.06) pg/mL和(40.05±10.03) pg/mL,明显高于对照组(p<0.05),且高于术前水平(p<0.05);观察组和对照组手术前后粒细胞-巨噬细胞集落刺激因子(GM-CSF)和碱性成纤维细胞生长因子(b-FGF)比较差异无统计学意义(p>0.05);术后4周IP-10、MCP-1、VEGF和TGF-β2水平与黄斑区视网膜厚度呈正相关(r=0.452, 0.501, 0.466和0.470, p<0.05)。本研究的结果初步说明,房水中IP-10、MCP-1、VEGF和TGF-2水平与2型糖尿病白内障术后黄斑水肿有一定关系,有可能成为黄斑水肿的特异性评价指标,值得进一步研究。     

曲安奈德玻璃体注射治疗黄斑水肿的安全性长期随访分析

目的:观察2 mg、4 mg曲安奈德玻璃体注射治疗黄斑水肿的长期安全性。方法:将54例黄斑水肿患者分别采用2 mg和4mg不同剂量的曲安奈德玻璃体内注射,随访2年,观察眼压、晶状体、眼内炎症、视网膜、玻璃体等并发症。结果:两种剂量曲安奈德玻璃体内注射均可产生高眼压、白内障、非感染性眼内炎症;4 mg组白内障发生率明显高于2 mg组,其余并发症未见明显差异;两组患者均未出现严重、不可逆并发症。结论:2 mg和4 mg曲安奈德玻璃体注射治疗黄斑水肿均较安全,4 mg剂量更容易发生白内障,两种计量均需长期随访。     

Trends in the Care of Diabetic Macular Edema: Analysis of a National Cohort

Purpose

To evaluate how the monitoring and treatment for diabetic macular edema (DME) has changed in a national sample.

Design

Retrospective cohort study.

Methods

Setting: Administrative medical claims data from a large, national U.S. insurer. Study population: Beneficiaries of a U.S. insurance company. Observation procedures: All incident cases of DME were found. Those in years 2002/3, 2006 and 2010 were followed for a 2-year observation period and those from 2009, 2010 and 2011 for a 1-year observation period. Main Outcome Measures: Types and frequencies of treatment were tallied and compared over each of the cohorts.

Results

Two-year cohorts had 233, 251 and 756 patients in 2002/3, 2006 and 2010 respectively. One-year cohorts had 1002, 1119 and 1382 patients in 2009, 2010 and 2011, respectively. Both percentage of patients receiving therapy and number of treatments given increased across the 2-year cohorts for both focal laser and anti-vascular endothelial growth factor (anti-VEGF) (p<0.001). The highest use of anti-VEGF agents in any of the cohorts was in the 2011 1-year group that only averaged 3.78 injections. Focal laser was used 2.5x as frequently as anti-VEGF injections in the most recent cohorts with only a high of 14.0% of DME patients receiving anti-VEGF therapy in any of the cohorts.

Conclusion

Regardless of treatment modality (laser or injection) DME patients received vastly fewer treatments than patients in randomized control trials. Despite the proven superior visual outcomes of anti-VEGF agents over focal laser in DME, focal laser was still used more frequently.     

The Expression of Two Splice Variants of Metabotropic Glutamate Receptor Subtype 5 in the Rat Brain and Neuronal Cells During Development

Abstract: We previously reported that a variant with extra amino acid residues exists in the metabotropic glutamate receptor subtype 5 (mGluR5). Either of the two isoforms, named mGluR5b and mGluR5a for the isoforms with and without the inserted sequence, respectively, generated Ca²⁺-activated Cl⁻ current when expressed in Xenopus oocytes. We herein report that these two isoforms are produced by the alternative splicing of the exon skipping type. When examined during the course of postnatal development, the major mGluR5 isotype mRNA was observed to switch from mGluR5a to mGluR5b in the rat hippocampus and the cerebral cortex. We also investigated two cell lines that could be differentiated into neuron-like cells in vitro. Whereas the mGluR5b mRNA was hardly detectable in either undifferentiated or differentiated NG108-15 cells, the relative amounts of the two variant mRNAs changed after the induction of differentiation in the P19 cells. An extracellular application of trans - d,l -1-amino-1,3-cyclopentanedicarboxylate on the neuron-like P19 cells induced intracellular Ca²⁺ mobilization, thus suggesting that the cells could express functional mGluR(s) coupled to phospholipase C and other components that could mediate the signal transduction pathway. This cell line may thus provide a model system for studying both mGluR5 expression and other mGluR-induced phenomena at the molecular level.     

Morphology and Function over a One-Year Follow Up Period after Intravitreal Dexamethasone Implant (Ozurdex) in Patients with Diabetic Macular Edema

Background

To investigate changes in macular morphology and function after an intravitreal dexamethasone implant for diabetic macular edema (DME).

Methods

Twenty-seven eyes in 27 treatment-naive patients affected by DME were treated with intravitreal Ozurdex® injections (IVOI) and followed up 12 months to evaluate morphological and functional changes by means of best-corrected visual acuity (BCVA), microperimetry (MP1), multifocal electroretinography (mfERG), pattern electroretinography (PERG) and spectral domain optical coherence tomography (SD-OCT).

Results

Both BCVA and retinal sensitivity improved significantly at one month after the IVOI (p = 0.031 and p<0.0001, respectively). After five months, the improvement of BCVA remained statistically significant compared with baseline values (p = 0.022); retinal sensitivity improvement was statistically significant for up to four months after the IVOI (p = 0.059). Moreover, central macular thickness significantly decreased for up to four months. Interestingly, PERG and mfERG values did not change significantly for up to four months post-IVOI, but then began to worsen.

Conclusions

In eyes with DME, intravitreal dexamethasone implant determined morphological and functional improvement as soon as one month and for up to four months after the treatment.     

Copy Number Variants in German Patients with Schizophrenia

Large rare copy number variants (CNVs) have been recognized as significant genetic risk factors for the development of schizophrenia (SCZ). However, due to their low frequency (1∶150 to 1∶1000) among patients, large sample sizes are needed to detect an association between specific CNVs and SCZ. So far, the majority of genome-wide CNV analyses have focused on reporting only CNVs that reached a significant P-value within the study cohort and merely confirmed the frequency of already-established risk-carrying CNVs. As a result, CNVs with a very low frequency that might be relevant for SCZ susceptibility are lost for secondary analyses. In this study, we provide a concise collection of high-quality CNVs in a large German sample consisting of 1,637 patients with SCZ or schizoaffective disorder and 1,627 controls. All individuals were genotyped on Illumina''s BeadChips and putative CNVs were identified using QuantiSNP and PennCNV. Only those CNVs that were detected by both programs and spanned ≥30 consecutive SNPs were included in the data collection and downstream analyses (2,366 CNVs, 0.73 CNVs per individual). The genome-wide analysis did not reveal a specific association between a previously unknown CNV and SCZ. However, the group of CNVs previously reported to be associated with SCZ was more frequent in our patients than in the controls. The publication of our dataset will serve as a unique, easily accessible, high-quality CNV data collection for other research groups. The dataset could be useful for the identification of new disease-relevant CNVs that are currently overlooked due to their very low frequency and lack of power for their detection in individual studies.     

Single-Step Purification of Two Functional Human Apolipoprotein E Variants Hyperexpressed inEscherichia coli

We have cloned, from total human liver RNA, the cDNA encoding apolipoprotein E3 (apoE3). Site-directed mutagenesis was used to obtain the cDNA encoding the apoE4 isoform, a major variant of this apolipoprotein in man. These two cDNAs were subcloned into the procaryotic expression vector pAHRS. A polyhistidine tag was added at the NH₂-termini of the recombinant proteins (apoE3 and apoE4) to enable rapid purification. The resulting plasmids (pAHRS-apoE3 and pAHRS-apoE4) were introduced into theEscherichia colistrain BL21(DE3). Recombinant strains were grown at 37°C in a Luria and Bertani medium and the addition of isopropyl β-thiogalactoside resulted in the expression of large amounts of apoE protein (40.5 kDa), representing at least 15% of cellular proteins. The recombinant apoE isoforms were purified, under denaturating conditions, in one step by affinity chromatography on a Ni-chelated agarose column, yielding to about 20 mg of 96% pure protein per liter of culture. Compared to plasma apoE3 purified from human very low density lipoproteins, the two renatured recombinant apoE isoforms have the same secondary structure content, as revealed by circular dichroism measurement. Moreover, the recombinant apoE3 isoform shares similar properties for the association with lipids, compared to the human protein, indicating that the addition of the amino-terminal polyhistidine peptide does not influence the structure and the lipid binding properties of this recombinant apoE isoform. No differences in the secondary structure of recombinant apoE4 were detected, whereas this isoform presents specific reactivity with lipids. This simple and rapid procedure for the expression and the purification of functional recombinant apoE should therefore enable structural and physiological studies requiring large amounts of these apolipoproteins.     

Functional Analysis of Two PLA2G2A Variants Associated with Secretory Phospholipase A2-IIA Levels

Background

Secretory phospholipase A2 group IIA (sPLA2-IIA) has been identified as a biomarker of atherosclerosis in observational and animal studies. The protein is encoded by the PLA2G2A gene and the aim of this study was to test the functionality of two PLA2G2A non-coding SNPs, rs11573156 C>G and rs3767221 T>G where the rare alleles have been previously associated with higher and lower sPLA2-IIA levels respectively.

Methodology/Principal Findings

Luciferase assays, electrophoretic mobility shift assays (EMSA), and RNA expression by RT-PCR were used to examine allelic differences. For rs3767221 the G allele showed ∼55% lower luciferase activity compared to the T allele (T = 62.1 (95% CI 59.1 to 65.1) G = 27.8 (95% CI 25.0 to 30.6), p = 1.22×10⁻³⁵, and stronger EMSA binding of a nuclear protein compared to the T-allele. For rs11573156 C >G there were no luciferase or EMSA allelic differences seen. In lymphocyte cell RNA, from individuals of known rs11573156 genotype, there was no allelic RNA expression difference for exons 5 and 6, but G allele carriers (n = 7) showed a trend to lower exon 1–2 expression compared to CC individuals. To take this further, in the ASAP study (n = 223), an rs11573156 proxy (r² = 0.91) showed ∼25% higher liver expression of PLA2G2A (1.67×10⁻¹⁷) associated with the G allele. However, considering exon specific expression, the association was greatly reduced for exon 2 (4.5×10⁻⁵) compared to exons 3–6 (10⁻¹⁰ to 10⁻²⁰), suggesting rs11573156 G allele-specific exon 2 skipping.

Conclusion

Both SNPs are functional and provide useful tools for Mendelian Randomisation to determine whether the relationship between sPLA2-IIA and coronary heart disease is causal.     

Interobserver Agreement in Detecting Spectral-Domain Optical Coherence Tomography Features of Diabetic Macular Edema

PurposeTo evaluate interobserver agreement for the detection of spectral-domain optical coherence tomography (SDOCT) features of diabetic macular edema (DME).MethodCross-sectional study in which 2 retinal specialists evaluated SDOCT scans from eyes receiving treatment for DME. Scans from 50 eyes with DME of 39 patients were graded for features of DME including intra-retinal fluid (IRF), diffuse retinal oedema (DRE), hyper-reflective foci (HRF), subretinal fluid (SRF), macular fluid and vitreomacular traction (VMT). Features were graded as present or absent at zones involving the fovea, 1mm from the fovea and the whole scan of 49 line scans. Analysis was performed using cross-tabulations for percentage concordance and kappa values (κ).ResultsIn the 2950 line scans analysed, there was an increase in percentage concordance for DRE and HRF when moving from a foveal line scan, 1mm zone and then to a whole scan analysis (88% vs 94% vs 96%) and (88% vs 94% vs 94%) respectively with κ ranging from substantial to almost perfect. Percentage concordance for SRF was 96% at all 3 regions analysed, whilst IRF was 96% at fovea and 98% at higher number of line-scans analysed. Concordance for MF was 100% at fovea and 98% at 1mm zone and whole scan with almost perfect and substantial κ respectively. κ agreement was substantial for VMT at all regions analysed.ConclusionWe report a high level of interobserver agreement in the detection of SDOCT features of DME. This finding is important as detection of macular fluid is used to guide retreatment with anti-angiogenic agents.     

Influence of Uncomplicated Phacoemulsification on Central Macular Thickness in Diabetic Patients: A Meta-Analysis

ObjectiveTo evaluate the effect of uncomplicated phacoemulsification on central macular thickness (CMT) and best corrected visual acuity (BCVA) in both diabetic patients without diabetic retinopathy (DR) and diabetic patients with mild to moderate non-proliferative diabetic retinopathy (NPDR).MethodsPotential prospective observational studies were searched through PubMed and EMBASE. Standardized mean difference (SMD) and 95% confidence interval (CI) for changes in CMT and BCVA were evaluated at postoperative 1, 3 and 6 months. The pooled effect estimates were calculated in the use of a random-effects model.ResultsA total of 10 studies involving 190 eyes of diabetic patients without diabetic retinopathy and 143 eyes of diabetic patients with NPDR were identified. CMT values demonstrated a statistically significant increase after uncomplicated phacoemulsification at 1 month (SMD, -0.814; 95%CI, -1.230 to -0.399), 3 months (SMD, -0.565; 95%CI, -0.927 to -0.202) and 6 months (SMD, -0.458; 95%CI, -0.739 to -0.177) in diabetic patients with NPDR. There was no statistical difference in CMT values at postoperative 1 month (SMD, -1.206; 95%CI, -2.433 to 0.021)and no statistically significant increase in CMT values at postoperative3 months (SMD, -0.535; 95%CI, -1.252 to 0.182) and 6 months (SMD, -1.181; 95%CI, -2.625 to 0.263) in diabetic patients without DR.BCVA was significantly increased at postoperative 1 month (SMD, 1.149; 95%CI, 0.251 to 2.047; and SMD,1.349; 95%CI, 0.264 to 2.434, respectively) and 6 months (SMD, 1.295; 95%CI, 0.494 to 2.096; and SMD, 2.146; 95%CI, 0.172 to 4.120, respectively) in both diabetic patients without DR and diabetic patients with NPDR. Sensitivity analysis showed that the results were relatively stable and reliable.ConclusionUncomplicated phacoemulsification in diabetic patients with mild to moderate NPDR seemed to influence significantly the subclinical thickening of the macular zones at postoperative 1, 3 and 6 months compared with diabetic patients without DR. BCVA was significantly improved in both diabetic patients without DR and diabetic patients with mild to moderate NPDR.     

Unbiased Functional Clustering of Gene Variants with a Phenotypic-Linkage Network

Groupwise functional analysis of gene variants is becoming standard in next-generation sequencing studies. As the function of many genes is unknown and their classification to pathways is scant, functional associations between genes are often inferred from large-scale omics data. Such data types—including protein–protein interactions and gene co-expression networks—are used to examine the interrelations of the implicated genes. Statistical significance is assessed by comparing the interconnectedness of the mutated genes with that of random gene sets. However, interconnectedness can be affected by confounding bias, potentially resulting in false positive findings. We show that genes implicated through de novo sequence variants are biased in their coding-sequence length and longer genes tend to cluster together, which leads to exaggerated p-values in functional studies; we present here an integrative method that addresses these bias. To discern molecular pathways relevant to complex disease, we have inferred functional associations between human genes from diverse data types and assessed them with a novel phenotype-based method. Examining the functional association between de novo gene variants, we control for the heretofore unexplored confounding bias in coding-sequence length. We test different data types and networks and find that the disease-associated genes cluster more significantly in an integrated phenotypic-linkage network than in other gene networks. We present a tool of superior power to identify functional associations among genes mutated in the same disease even after accounting for significant sequencing study bias and demonstrate the suitability of this method to functionally cluster variant genes underlying polygenic disorders.     

Analysis of Ancestral and Functionally Relevant CD5 Variants in Systemic Lupus Erythematosus Patients

Objective

CD5 plays a crucial role in autoimmunity and is a well-established genetic risk factor of developing RA. Recently, evidence of positive selection has been provided for the CD5 Pro224-Val471 haplotype in East Asian populations. The aim of the present work was to further analyze the functional relevance of non-synonymous CD5 polymorphisms conforming the ancestral and the newly derived haplotypes (Pro224-Ala471 and Pro224-Val471, respectively) as well as to investigate the potential role of CD5 on the development of SLE and/or SLE nephritis.

Methods

The CD5 SNPs rs2241002 (C/T; Pro224Leu) and rs2229177 (C/T; Ala471Val) were genotyped using TaqMan allelic discrimination assays in a total of 1,324 controls and 681 SLE patients of Spanish origin. In vitro analysis of CD3-mediated T cell proliferative and cytokine response profiles of healthy volunteers homozygous for the above mentioned CD5 haplotypes were also analyzed.

Results

T-cell proliferation and cytokine release were significantly increased showing a bias towards to a Th2 profile after CD3 cross-linking of peripheral mononuclear cells from healthy individuals homozygous for the ancestral Pro224-Ala471 (CC) haplotype, compared to the more recently derived Pro224-Val471 (CT). The same allelic combination was statistically associated with Lupus nephritis.

Conclusion

The ancestral Ala471 CD5 allele confers lymphocyte hyper-responsiveness to TCR/CD3 cross-linking and is associated with nephritis in SLE patients.     

Wider Retinal Artery Trajectories in Eyes with Macular Hole Than in Fellow Eyes of Patients with Unilateral Idiopathic Macular Hole

Purpose

To determine whether the width of the retinal artery (RA) trajectory was associated with the presence of a macular hole (MH).

Methods

A retrospective cross sectional case-control study was performed. The fundus photographs were rotated 90 degrees, and the coordinates of the best fit curve of the RA trajectory were determined automatically based on these plots using the ImageJ program. The converted coordinates were fit to a second degree polynomial (ax²/100 + bx + c) equation. The width and steepness of the RA trajectory, “a”, of the eyes with a MH eye were compared to that of the fellow eyes.

Results

One hundred and ten eyes of 55 consecutive patients (30 women) with a unilateral MH and healthy fellow eyes were analyzed. The mean age was 64.9 years (range 47-81 years). The constant ‘a’ was significantly smaller in eyes with a MH than that of the fellow eyes (0.379 ± 0.094 vs 0.416 ± 0.121, P = 0.001, paired t test), indicating that the RA trajectory was wider in the MH eyes than in the fellow eyes. There was a significant correlation between the axial length and ‘a’ of the RA trajectory in the MH eyes (R = 0.273, P = 0.044) and in the fellow eyes (R = 0.356, P = 0.008; Spearman’s rank correlation coefficient).

Conclusions

Because eyes with a MH have a significantly wider and flatter RA trajectory, there may be greater traction on the fovea which is located between the RA arches. The causative role of this finding is still unclear.