Development of Multimarker Panel for Early Detection of Diabetic Retinopathy Using LC/MRM-MS and Antibody-Based Verification
Abstract
Diabetic retinopathy (DR) is a complication of diabetes and 80 % of diabetes mellitus (DM) patients whose DM duration is over 10 years can be expected to suffer with DR. Diagnosis of DR have been depends on ophthalmological examination and there are no molecular methods to screen the DR status yet. Nonproliferative diabetic retinopathy (NPDR) is the early DR which is hard to be noticed in early NPDR, showing significant cause of adult blindness in type 2 diabetes patients. Protein biomarkers have been valuable in the diagnosis of disease and the use of multiple biomarkers has been suggested to overcome the low specificity of single ones. For multiple biomarker development, multiple reaction monitoring (MRM) has been spotlighted as an alternative method to quantify target proteins. Then, we designed, constructed, and evaluated a systematic statistical pipeline for establishing a multimarker panel using candidate proteins from MRM and the antibody-based verification. F-test and stepwise multiple analysis of variance (S-MANOVA) were performed to select multimarker proteins that increased the discriminatory power between no DR and moderate NPDR. All prediction models, including linear discriminant analysis (LDA), support vector machine (SVM), and logistic regression (LR), were crossvalidated by leave-one-out crossvalidation (LOOCV). As results, in the comparison between no DR and moderate NPDR, the 4-marker panel had improved discriminatory performance compared, with greater sensitivity (0.95 in LDA, 0.85 in SVM and 1 in LR) and specificity (0.95, 0.9, and 0.9, respectively) and a lower error rate (5%, 12.5%, and 5%, respectively).
Role of sphingosine 1-phosphate in fatty liver and obesity
Abstract
The endoplasmic reticulum is the principal organelle in the cell for protein folding and trafficking, lipid synthesis and cellular calcium homeostasis. Perturbation of ER function results in activation of the unfolded protein response (UPR). Chronic ER stress is reported to have an important role in abnormal lipid biosynthesis and development of insulin resistance. The present study reports that transcription of sphingosine kinase 2 (Sphk2) is differentially regulated by ER stress-mediated UPR pathways. Expression of Sphk2, a major isotype of sphingosine kinase in the liver, was upregulated by tunicamycin and sphingosine 1-phosphate (S1P) was elevated in primary mouse hepatocytes. In contrast, chronic ER stress by high fat diet suppressed Sphk2 expression. Overexpression of the activating transcription factor 4 (ATF4) upregulated Sphk2 expression, whereas the spliced form of X-box binding protein 1 (sXBP1) downregulated Sphk2 as demonstrated by Sphk2 promoter assays and western blot analyses. Adenoviral Sphk2 overexpression activated pAKT with no alteration of IRS1 phosphorylation. In addition, cellular S1P levels were elevated by Sphk2 overexpression while ceramide and sphingomyelin were not altered. These results demonstrated that Sphk2 is differentially regulated by ER stress-induced UPR pathways would contribute to the amelioration of hepatic insulin resistance and steatosis via S1P production.
Lipidomic Profiling of Liver Tissue from Obesity-Prone and Obesity-Resistant Mice Fed a High Fat Diet
Abstract
Obesity is a multifactorial health problem resulting from genetic, environmental, and behavioral factors. A particularly interesting aspect of obesity is the differences observed in response to the same high-fat diet (HFD). In this study, we performed lipidomic profiling of liver from HFD-fed C57BL/6J mice using ultra-performance liquid chromatography–quadrupole time-of-flight mass spectrometry. Mice were divided into three groups: normal diet (ND), HFD-obesity prone (HFD-OP), and HFD-obesity resistant (HFD-OR). Principal components analyses showed a slight separation between the HFD-OP and HFD-OR groups. Individuals of HFD-OR group were closer to those of the ND group compared to those of the HFD-OP group. In particular, phosphocholine (PC) and triacylglyceride levels differed significantly depending on the length of the acyl chain and degree of unsaturation, respectively. PC species were either positively or negatively correlated with concentrations of glucose, insulin, leptin, and hepatic cholesterol according to the length of the acyl chain. Decreased expression of the scavenger receptor B1 and ATP-binding cassette A1 in HFD-OP mice indicated that the acyl chain length of PC species may be related to high-density lipoprotein cholesterol metabolism. This study demonstrates that the lipidomic profiling is an effective approach of analyzing global lipid alterations as they pertain to obesity.
Code / Date
SYM 1-4 / March 28(Mon) 11:08-11:25
Speaker
SooYoung Cho
Affiliation
Seoul National University, Korea
Title
Transcriptome Profiles on Weight Cycling Model Mice in White Adipose Tissue
Abstract
A Weight cycle (WC) is the cyclic loss and gain of body weight, and estimates range from 25 to 40% of the population in westernized countries. To identify functional difference between body weight fluctuation and simple weight gain event, we established WC model mice and constructed genome wide transcriptome profile using by RNA-sequencing technology in white adipose tissue (WAT). C57BL/6N mice fed on high fat diet (HFD) for 8 weeks, normal diet (ND) was challenged for 4 weeks and second HFD was challenged for 4 weeks. Cell cycle and immune response were highly enriched in WC and weight gain group. To identify different immune response between WC and weight gain group, co-expression network was constructed. We found WC association module which was following body weight fluctuation and showed increasing expression pattern in second HFD challenge. The module was highly enriched with major histocompatibility complex class II (MHC class II)-mediated antigen presentation and T cell activation function. We validated activation of adaptive immune response with qPCR. These results suggested that obesity will be memorized by adaptive immune system in MHC class II-dependent manner.
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