Integration of Meta-Multi-Omics Data Using Probabilistic Graphs and External Knowledge.

Multi-omics has the promise to provide a detailed molecular picture of biological systems. Although obtaining multi-omics data is relatively easy, methods that analyze such data have been lagging. In this paper, we present an algorithm that uses probabilistic graph representations and external knowledge to perform optimal structure learning and deduce a multifarious interaction network for multi-omics data from a bacterial community. Kefir grain, a microbial community that ferments milk and creates kefir, represents a self-renewing, stable, natural microbial community. Kefir has been shown to have a wide range of health benefits. We obtained a controlled bacterial community using the two most abundant and well-studied species in kefir grains: Lentilactobacillus kefiri and Lactobacillus kefiranofaciens. We applied growth temperatures of 30 °C and 37 °C and obtained transcriptomic, metabolomic, and proteomic data for the same 20 samples (10 samples per temperature). We obtained a multi-omics interaction network, which generated insights that would not have been possible with single-omics analysis. We identified interactions among transcripts, proteins, and metabolites, suggesting active toxin/antitoxin systems. We also observed multifarious interactions that involved the shikimate pathway. These observations helped explain bacterial adaptation to different stress conditions, co-aggregation, and increased activation of L. kefiranofaciens at 37 °C.

Serum Protein Signatures Using Aptamer-Based Proteomics for Minimal Change Disease and Membranous Nephropathy.

Introduction: Minimal change disease (MCD) and membranous nephropathy (MN) are glomerular diseases (glomerulonephritis [GN]) that present with the nephrotic syndrome. Although circulating PLA2R antibodies have been validated as a biomarker for MN, the diagnosis of MCD and PLA2R-negative MN still relies on the results of kidney biopsy or empirical corticosteroids in children. We aimed to identify serum protein biomarker signatures associated with MCD and MN pathogenesis using aptamer-based proteomics. Methods: Quantitative SOMAscan proteomics was applied to the serum of adult patients with MCD (n = 15) and MN (n = 37) and healthy controls (n = 20). Associations between the 1305 proteins detected with SOMAscan were assessed using multiple statistical tests, expression pattern analysis, and systems biology analysis. Results: A total of 208 and 244 proteins were identified that differentiated MCD and MN, respectively, with high statistical significance from the healthy controls (Benjamin-Hochberg [BH] P < 0.0001). There were 157 proteins that discriminated MN from MCD (BH P < 0.05). In MCD, 65 proteins were differentially expressed as compared with MN and healthy controls. When compared with MCD and healthy controls, 44 discriminatory proteins were specifically linked to MN. Systems biology analysis of these signatures identified cell death and inflammation as key pathways differentiating MN from MCD and healthy controls. Dysregulation of fatty acid metabolism pathways was confirmed in both MN and MCD as compared with the healthy subjects. Conclusion: SOMAscan represents a promising proteomic platform for biomarker development in GN. Validation of a greater number of discovery biomarkers in larger patient cohorts is needed before these data can be translated for clinical care.

Differentiation of memory processing stages and effect of demographic variables with alternative scoring approaches to the Rey auditory verbal learning test.

BACKGROUND: The Rey Auditory Verbal Learning Test (RAVLT) is the third most popular verbal memory test and the tenth most frequently used neuropsychological test. The original scoring system of RAVLT does not differentiate stages of memory processing, but a recently developed composite scoring system has this potential. The objectives were to compare the two systems in terms of their capacity to differentiate the stages of memory processing and to study the effect of demographic variables on the learning trials (T) of the Turkish form of RAVLT (T-RAVLT). METHOD: The sample consisted of 600 Caucasian Turkic adults, who were categorized into three levels of age, three levels of education, and two levels of gender. Individual administration of T-RAVLT was performed using the standard procedures of RAVLT. RESULTS: The components in the exploratory factor analysis (EFA) and latent variables in the confirmatory factor analysis (CFA) of the original scores were consistent with sequentially ordered T-RAVLT stages. Demographic variables (age, education, and gender) affected performances in all of the learning trials. The composite scores revealed retrieval and retention as separate components, but these scores could not be predicted from the relevant T-RAVLT scores. CONCLUSIONS: Findings recommend a combined utilization of the two scoring systems: The original system to provide scores on the performance at each stage of T-RAVLT and the combined system to provide separate scores on learning, retention, and retrieval, the three stages of memory processing. A selective effect of demographic variables on T1 was not observed, indicating a need for cross-cultural studies that are meticulously controlled for age and education.

KEGG2Net: Deducing gene interaction networks and acyclic graphs from KEGG pathways.

The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway database provides a manual curation of biological pathways that involve genes (or gene products), metabolites, chemical compounds, maps, and other entries. However, most applications and datasets involved in omics are gene or protein-centric requiring pathway representations that include direct and indirect interactions only between genes. Furthermore, special methodologies, such as Bayesian networks require acyclic representations of graphs. We developed KEGG2Net, a web resource that generates a network involving only the genes represented on a KEGG pathway with all of the direct and indirect gene-gene interactions deduced from the pathway. KEGG2Net offers four different methods to remove cycles from the resulting gene interaction network, converting them into directed acyclic graphs (DAGs). We generated synthetic gene expression data using the gene interaction networks deduced from the KEGG pathways and performed a comparative analysis of different cycle removal methods by testing the fitness of their DAGs to the data and by the number of edges they eliminate. Our results indicate that an ensemble method for cycle removal performs as the best approach to convert the gene interaction networks into DAGs. Resulting gene interaction networks and DAGs are represented in multiple user-friendly formats that can be used in other applications, and as images for quick and easy visualisation. The KEGG2Net web portal converts KEGG maps for any organism into gene-gene interaction networks and corresponding DAGS representing all of the direct and indirect interactions among the genes.

Comparative analysis of single-cell transcriptomics in human and Zebrafish oocytes.

BACKGROUND: Zebrafish is a popular model organism, which is widely used in developmental biology research. Despite its general use, the direct comparison of the zebrafish and human oocyte transcriptomes has not been well studied. It is significant to see if the similarity observed between the two organisms at the gene sequence level is also observed at the expression level in key cell types such as the oocyte. RESULTS: We performed single-cell RNA-seq of the zebrafish oocyte and compared it with two studies that have performed single-cell RNA-seq of the human oocyte. We carried out a comparative analysis of genes expressed in the oocyte and genes highly expressed in the oocyte across the three studies. Overall, we found high consistency between the human studies and high concordance in expression for the orthologous genes in the two organisms. According to the Ensembl database, about 60% of the human protein coding genes are orthologous to the zebrafish genes. Our results showed that a higher percentage of the genes that are highly expressed in both organisms show orthology compared to the lower expressed genes. Systems biology analysis of the genes highly expressed in the three studies showed significant overlap of the enriched pathways and GO terms. Moreover, orthologous genes that are commonly overexpressed in both organisms were involved in biological mechanisms that are functionally essential to the oocyte. CONCLUSIONS: Orthologous genes are concurrently highly expressed in the oocytes of the two organisms and these genes belong to similar functional categories. Our results provide evidence that zebrafish could serve as a valid model organism to study the oocyte with direct implications in human.

Construct Validity of Auditory Verbal Learning Test. / Isitsel Sözel Ögrenme Testi Yapi Geçerligi.

OBJECTIVE: Auditory Verbal Learning Test (AVLT) is frequently used in neuropsychology literature to comprehensively assess the memory. The test measures verbal learning as immediate and delayed free recall, recognition, and retroactive and proactive interference. Adaptation of AVLT to the Turkish society has been completed, whereas research and development studies are still underway. The purpose of the present study is to investigate the construct validity of the test in order to contribute to the research and development process. METHOD: In line with this purpose, the research data were obtained from 78 healthy participants aged between 20 and 69. The exclusion criteria included neurological and/or psychiatric disorders as well as untreated auditory/visual disorders. AVLT was administered to participants individually by two trained psychologists. RESULTS: Principal component analysis that is used to investigate the components represented by the AVLT scores consisted of learning, free recall and recognition, in line with the construct of the test. Distractors were also added to these two components in structural equation model. Analyses were carried out on descriptive level to establish the relatioships between age, education, gender and AVLT scores. CONCLUSION: These findings, which are consistent with the literature indicating that memory is affected by the developmental process, suggest that learning/free recall, recognition, and distractor scores of the AVLT demonstrate a component pattern consistent with theoretical knowledge. This conclusion suggests that AVLT is a valid measurement test for the Turkish society.

[A Study to Determine the Norms for the Trail Making Test For the Age Range of 20-49 in Turkey]. / Iz Sürme Testi’nin 20-49 Yas Araliginda Türkiye Için Norm Belirleme Çalismasi.

OBJECTIVE: The main goal of the study is to determine the normative values of Trail Making Test (TMT) for people between 20-49 years of age; to examine the effect of age, education and sex variables over TMT scores and identify the reliability coefficient of the test. METHOD: The sample of the study consisted of 133 women and 130 men, 261 voluntary and healthy participants in total. The data of the research was collected according to 3 x 2 x 2 factorial experimental design; and the participants were distributed to experimental conditions well balanced in terms of the levels of age, education and sex. TMT A and B form, and Beck Depression Scale (BDS) were applied for the assessment. RESULTS: Seven scores were calculated determining for Turkish normative values; 3 x 2 x 2 factorial multivariate variance analysis was applied in order to identify the effect of levels of age, education and sex over TMT scores. According to the results of the analyses, it was found that main effect of education was significant, while the main effects of age and sex variables were not (p<.05). The test-retest reliability coefficients of the TMT changed between .71 and .87. CONCLUSION: This study indicated that especially the TMT B and subtest scores were affected by the education in the range of 20-49 years of age. Additionally, normative values depending on the means of TMT scores for 20-49 age group were obtained in the study; it was shown that TMT was a reliable assessment tool.

Metagenomic analysis of the microbial community in kefir grains.

Kefir grains as a probiotic have been subject to microbial community identification using culture-dependent and independent methods that target specific strains in the community, or that are based on limited 16S rRNA analysis. We performed whole genome shotgun pyrosequencing using two Turkish Kefir grains. Sequencing generated 3,682,455 high quality reads for a total of ∼1.6 Gbp of data assembled into 6151 contigs with a total length of ∼24 Mbp. Species identification mapped 88.16% and 93.81% of the reads rendering 4 Mpb of assembly that did not show any homology to known bacterial sequences. Identified communities in the two grains showed high concordance where Lactobacillus was the most abundant genus with a mapped abundance of 99.42% and 99.79%. This genus was dominantly represented by three species Lactobacillus kefiranofaciens, Lactobacillus buchneri and Lactobacillus helveticus with a total mapped abundance of 97.63% and 98.74%. We compared and verified our findings with 16S pyrosequencing and model based 16S data analysis. Our results suggest that microbial community profiling using whole genome shotgun data is feasible, can identify novel species data, and has the potential to generate a more accurate and detailed assessment of the underlying bacterial community, especially for low abundance species.

Whole genome sequence of a Turkish individual.

Although whole human genome sequencing can be done with readily available technical and financial resources, the need for detailed analyses of genomes of certain populations still exists. Here we present, for the first time, sequencing and analysis of a Turkish human genome. We have performed 35x coverage using paired-end sequencing, where over 95% of sequencing reads are mapped to the reference genome covering more than 99% of the bases. The assembly of unmapped reads rendered 11,654 contigs, 2,168 of which did not reveal any homology to known sequences, resulting in ∼1 Mbp of unmapped sequence. Single nucleotide polymorphism (SNP) discovery resulted in 3,537,794 SNP calls with 29,184 SNPs identified in coding regions, where 106 were nonsense and 259 were categorized as having a high-impact effect. The homo/hetero zygosity (1,415,123∶2,122,671 or 1∶1.5) and transition/transversion ratios (2,383,204∶1,154,590 or 2.06∶1) were within expected limits. Of the identified SNPs, 480,396 were potentially novel with 2,925 in coding regions, including 48 nonsense and 95 high-impact SNPs. Functional analysis of novel high-impact SNPs revealed various interaction networks, notably involving hereditary and neurological disorders or diseases. Assembly results indicated 713,640 indels (1∶1.09 insertion/deletion ratio), ranging from -52 bp to 34 bp in length and causing about 180 codon insertion/deletions and 246 frame shifts. Using paired-end- and read-depth-based methods, we discovered 9,109 structural variants and compared our variant findings with other populations. Our results suggest that whole genome sequencing is a valuable tool for understanding variations in the human genome across different populations. Detailed analyses of genomes of diverse origins greatly benefits research in genetics and medicine and should be conducted on a larger scale.

[Evaluation of visual-motor integration functions in children between 6-15 years of age]. / 6-15 Yas Arasi Cocuklarda Görsel Motor Birlestirme Islevlerinin Degerlendirilmesi.

OBJECTIVE: Visual-motor integration skills are considered an essential domain of clinical and psycho-educational assessment. The goal of the present investigation is to provide the Turkish norms for the Beery-Buktenica Developmental Visual-Motor Integration Test (VMI-4th) for children and adolescents between the ages of 6-15 years as part of a comprehensive neuropsychological test battery. METHOD: A total of 1887 children from elementary and high schools in the city of Bursa were recruited for this study. From this sample 44 children were re-tested 3-4 weeks following the first administration for test-retest reliability. RESULTS: Findings showed clear developmental trajectories in visual-motor integration skills. Significant performance increments were observed in six month intervals for ages 6 and 7. Starting from age 8, norms were established for each age group separately. Girls and boys performed similarly on the VMI-4. Test- retest correlation was modest but within an acceptable range. CONCLUSION: The age-based norms established for the VMI-4 in this study can be used to assess children between the ages of 6-15 years as part of a clinical neuropsychological and a psycho-educational assessment. The mean VMI scores presented in this study represent performance of children in middle and middle-upper socio-economic status and may not represent the normal performance range of children from lower SES.

Proteomic identification of interleukin-2 therapy response in metastatic renal cell cancer.

PURPOSE: To detect a predictive protein profile that distinguishes interleukin-2 therapy responders and nonresponders among patients with metastatic renal cell carcinoma we used surface-enhanced laser desorption/ionization time-of-flight mass spectrometry. MATERIALS AND METHODS: Protein extracts from 56 patients with metastatic clear cell patients renal cell carcinoma obtained from radical nephrectomy specimens before interleukin-2 therapy were applied to protein chip arrays of different chromatographic properties and analyzed using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry. A class prediction algorithm was applied to identify a subset of protein peaks with expression values associated with interleukin-2 response status. Multivariate analysis was performed to assess the association between the proteomic profile and interleukin-2 response status, controlling for the effect of lymphadenopathy. RESULTS: From 513 protein peaks we discovered a predictor set of 11 that performed optimally for predicting interleukin-2 response status with 86% accuracy (Fisher's p <0.004, permutation p <0.01). Results were validated in an independent data set with 72% overall accuracy (p <0.05, permutation p <0.01). On multivariate analysis the proteomic profile was significantly associated with the interleukin-2 response when corrected for lymph node status (p <0.04). CONCLUSIONS: We identified and validated a proteomic pattern that is an independent predictor of the interleukin-2 response. The ability to predict the probability of the interleukin-2 response could permit targeted selection of the patients most likely to respond to interleukin-2, while avoiding unwanted toxicity in patients less likely to respond. This proteomic predictor has the potential to significantly aid clinicians in the decision making of appropriate therapy for patients with metastatic renal cell carcinoma.

Genomic expression pathways associated with brain injury after cardiopulmonary bypass.

OBJECTIVES: Neurologic injury after cardiac surgery, often manifested as neurocognitive decline, is a common postoperative complication without clear cause. We studied acute variations in gene-expression profiles of patients with neurocognitive decline (NCD group) compared with those without neurocognitive decline (NORM group) after cardiopulmonary bypass. METHODS: Forty-two patients undergoing coronary artery bypass grafting, valve procedures, or both by using cardiopulmonary bypass were administered a validated neurocognitive battery preoperatively and postoperatively at day 4. Neurocognitive decline was defined as 1 standard deviation from baseline on 25% or greater of tasks. Whole-blood mRNA was isolated preoperatively and at 6 hours after surgical intervention for fold-change calculation. Relative gene expression in the NCD versus the NORM group was assessed by using Affymetrix GeneChip U133 Plus 2.0 (>40,000 genes) from mRNA samples collected. Differential expression, clustering, gene ontology, and canonical pathway analysis were performed. Validation of microarray gene expression was performed with SYBR Green real-time polymerase chain reaction. RESULTS: Patients with neurocognitive decline (17/42 [40.5%] patients) were associated with a significantly different gene-expression response compared with that of healthy patients. Compared with preoperative samples, 6-hour samples had 531 upregulated and 670 downregulated genes uniquely in the NCD group compared with 2214 upregulated and 558 downregulated genes uniquely in the NORM group (P < .001; lower confidence bound, > or =1.2). Compared with patients in the NORM group, patients with neurocognitive decline had significantly different gene-expression pathways involving inflammation (including FAS, IL2RB, and CD59), antigen presentation (including HLA-DQ1, TAP1, and TAP2), and cellular adhesion (including ICAM2, ICAM3, and CAD7) among others. CONCLUSIONS: Patients with neurocognitive decline have inherently different genetic responses to cardiopulmonary bypass compared with those of patients without neurocognitive decline Genetic variations in inflammatory, cell adhesion, and apoptotic pathways might be important contributors to the pathophysiology of neurologic injury after cardiopulmonary bypass and could become a target for prevention and risk stratification.

Prediction of diabetic nephropathy using urine proteomic profiling 10 years prior to development of nephropathy.

OBJECTIVE: We examined whether proteomic technologies identify novel urine proteins associated with subsequent development of diabetic nephropathy in subjects with type 2 diabetes before evidence of microalbuminuria. RESEARCH DESIGN AND METHODS: In a nested case-control study of Pima Indians with type 2 diabetes, baseline (serum creatinine <1.2 mg/dl and urine albumin excretion <30 mg/g) and 10-year urine samples were examined. Case subjects (n = 31) developed diabetic nephropathy (urinary albumin-to-creatinine ratio >300 mg/g) over 10 years. Control subjects (n = 31) were matched to case subjects (1:1) according to diabetes duration, age, sex, and BMI but remained normoalbuminuric (albumin-to-creatinine ratio <30 mg/g) over the same 10 years. Surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS) was performed on baseline urine samples, and training (14 cases:14 controls) and validation (17:17) sets were tested. RESULTS: At baseline, A1C levels differed between case and control subjects. SELDI-TOF MS detected 714 unique urine protein peaks. Of these, a 12-peak proteomic signature correctly predicted 89% of cases of diabetic nephropathy (93% sensitivity, 86% specificity) in the training set. Applying this same signature to the independent validation set yielded an accuracy rate of 74% (71% sensitivity, 76% specificity). In multivariate analyses, the 12-peak signature was independently associated with subsequent diabetic nephropathy when applied to the validation set (odds ratio [OR] 7.9 [95% CI 1.5-43.5], P = 0.017) and the entire dataset (14.5 [3.7-55.6], P = 0.001), and A1C levels were no longer significant. CONCLUSIONS: Urine proteomic profiling identifies normoalbuminuric subjects with type 2 diabetes who subsequently develop diabetic nephropathy. Further studies are needed to characterize the specific proteins involved in this early prediction.

Restoration of liver mass after injury requires proliferative and not embryonic transcriptional patterns.

Normal adult liver is uniquely capable of renewal and repair after injury. Whether this response represents simple hyperplasia of various liver elements or requires recapitulation of the genetic program of the developing liver is not known. To study these possibilities, we examined transcriptional programs of adult liver after partial hepatectomy and contrasted these with developing embryonic liver. Principal component analysis demonstrated that the time series of gene expression during liver regeneration does not segregate according to developmental transcription patterns. Gene ontology analysis revealed that liver restoration after hepatectomy and liver development differ dramatically with regard to transcription factors and chromatin structure modification. In contrast, the tissues are similar with regard to proliferation-associated genes. Consistent with these findings, real-time polymerase chain reaction showed transcription factors known to be important in liver development are not induced during liver regeneration. These three lines of evidence suggest that at a transcriptional level restoration of liver mass after injury is best described as hepatocyte hyperplasia and not true regeneration. We speculate this novel pattern of gene expression may underlie the unique capacity of the liver to repair itself after injury.

The transcriptome of human oocytes.

The identification of genes and deduced pathways from the mature human oocyte can help us better understand oogenesis, folliculogenesis, fertilization, and embryonic development. Human metaphase II oocytes were used within minutes after removal from the ovary, and its transcriptome was compared with a reference sample consisting of a mixture of total RNA from 10 different normal human tissues not including the ovary. RNA amplification was performed by using a unique protocol. Affymetrix Human Genome U133 Plus 2.0 GeneChip arrays were used for hybridizations. Compared with reference samples, there were 5,331 transcripts significantly up-regulated and 7,074 transcripts significantly down-regulated in the oocyte. Of the oocyte up-regulated probe sets, 1,430 have unknown function. A core group of 66 transcripts was identified by intersecting significantly up-regulated genes of the human oocyte with those from the mouse oocyte and from human and mouse embryonic stem cells. GeneChip array results were validated using RT-PCR in a selected set of oocyte-specific genes. Within the up-regulated probe sets, the top overrepresented categories were related to RNA and protein metabolism, followed by DNA metabolism and chromatin modification. This report provides a comprehensive expression baseline of genes expressed in in vivo matured human oocytes. Further understanding of the biological role of these genes may expand our knowledge on meiotic cell cycle, fertilization, chromatin remodeling, lineage commitment, pluripotency, tissue regeneration, and morphogenesis.