Sensen Shen

Recent advances in lipidomics for disease research

Lipidomics is an important branch of metabolomics, which aims at the detailed analysis of lipid species and their multiple roles in the living system. In recent years, the development of various analytical methods for effective identification and characterization of lipids has greatly promoted the process of lipidomics. Meanwhile, as many diseases demonstrate a remarkable alteration in lipid profiles compared with that of healthy people, lipidomics has been extensively introduced to disease research. The comprehensive lipid profiling provides a chance to discover novel biomarkers for specific disease. In addition, it plays a crucial role in the study of lipid metabolism, which could illuminate the pathogenesis of diseases. In this review, after brief discussion of analytical methods for lipidomics in clinical research, we focus on the recent advances of lipidomics related to four types of diseases, including cancer, atherosclerosis, diabetes mellitus, and Alzheimers disease.

Metabolomics Approach Reveals Integrated Metabolic Network Associated with Serotonin Deficiency.

Serotonin is an important neurotransmitter that broadly participates in various biological processes. While serotonin deficiency has been associated with multiple pathological conditions such as depression, schizophrenia, Alzheimer’s disease and Parkinson’s disease, the serotonin-dependent mechanisms remain poorly understood. This study therefore aimed to identify novel biomarkers and metabolic pathways perturbed by serotonin deficiency using metabolomics approach in order to gain new metabolic insights into the serotonin deficiency-related molecular mechanisms. Serotonin deficiency was achieved through pharmacological inhibition of tryptophan hydroxylase (Tph) using p-chlorophenylalanine (pCPA) or genetic knockout of the neuronal specific Tph2 isoform. This dual approach improved specificity for the serotonin deficiency-associated biomarkers while minimizing nonspecific effects of pCPA treatment or Tph2 knockout (Tph2-/-). Non-targeted metabolic profiling and a targeted pCPA dose-response study identified 21 biomarkers in the pCPA-treated mice while 17 metabolites in the Tph2-/- mice were found to be significantly altered compared with the control mice. These newly identified biomarkers were associated with amino acid, energy, purine, lipid and gut microflora metabolisms. Oxidative stress was also found to be significantly increased in the serotonin deficient mice. These new biomarkers and the overall metabolic pathways may provide new understanding for the serotonin deficiency-associated mechanisms under multiple pathological states.

Cysteine-Functionalized Metal–Organic Framework: Facile Synthesis and High Efficient Enrichment of N-Linked Glycopeptides in Cell Lysate

Cysteine-functionalized metal-organic framework (MOF) was synthesized via a common and facile two-step method of in situ loading of Au nanoparticles on amino-derived MOF followed by l-cysteine (Cys) immobilization. Owing to the large specific surface area and ultrahigh hydrophilicity of this nanocomposite, excellent performance was observed in the enrichment of N-linked glycopeptides in both model glycoprotein and HeLa cell lysate. By using this nanocomposite, 16 and 31 glycopeptides were efficiently extracted from digest of horseradish peroxidase (HRP) and human serum immunoglobulin G (IgG), respectively. The short incubation time (5 min), large binding capacity (150 mg/g, IgG digest to material), good selectivity (1:50, molar ratio of IgG and bovine serum albumin (BSA) digest), high recovery (over 80%), and low detection limit (1 fmol) ensure the effectiveness and robustness of MIL-101(NH2)@Au-Cys in complex HeLa cell lysate. As a result, 1123 N-glycosylation sites corresponding to 1069 N-glycopeptides and 614 N-glycoproteins were identified from the lysate. Compared with those of previously reported hydrophilic methods, to our knowledge, it was the best result. This work paves a new way for fast functionalization of MOF and also provides a novel idea for material design in sample preparation, especially in glycoproteome and related analysis.

Lipidomic profiling of tryptophan hydroxylase 2 knockout mice reveals novel lipid biomarkers associated with serotonin deficiency

Serotonin is an important neurotransmitter that regulates a wide range of physiological, neuropsychological, and behavioral processes. Consequently, serotonin deficiency is involved in a wide variety of neurodegenerative diseases, including Alzheimer’s disease, Parkinson’s disease, schizophrenia, and depression. The pathophysiological mechanisms underlying serotonin deficiency, particularly from a lipidomics perspective, remain poorly understood. This study therefore aimed to identify novel lipid biomarkers associated with serotonin deficiency by lipidomic profiling of tryptophan hydroxylase 2 knockout (Tph2−/−) mice. Using a high-throughput normal-/reversed-phase two-dimensional liquid chromatography–quadrupole time-of-flight mass spectrometry (NP/RP 2D LC–QToF-MS) method, 59 lipid biomarkers encompassing glycerophospholipids (glycerophosphocholines, lysoglycerophosphocholines, glycerophosphoethanolamines, lysoglycerophosphoethanolamines glycerophosphoinositols, and lysoglycerophosphoinositols), sphingolipids (sphingomyelins, ceramides, galactosylceramides, glucosylceramides, and lactosylceramides) and free fatty acids were identified. Systemic oxidative stress in the Tph2−/− mice was significantly elevated, and a corresponding mechanism that relates the lipidomic findings has been proposed. In summary, this work provides preliminary findings that lipid metabolism is implicated in serotonin deficiency.

Lipidomic analysis of p-chlorophenylalanine-treated mice using continuous-flow two-dimensional liquid chromatography/quadrupole time-of-flight mass spectrometry.

RATIONALE Although serotonin deficiency is involved with various physiological disorders such as Alzheimers disease, Parkinsons disease, schizophrenia and depression, the serotonin-dependent pathomechanisms remain poorly understood, particularly from a lipidomics perspective. METHODS This study therefore aimed to identify novel lipid biomarkers associated with serotonin deficiency by lipid profiling of p-chlorophenylalanine (pCPA)-treated, serotonin-deficient mice using continuous-flow normal-phase/reversed-phase two-dimensional liquid chromatography/quadrupole time-of-flight mass spectrometry (NP/RP 2D LC/QTOFMS). Principal component analysis (PCA) was performed to distinguish significantly altered lipids between the pCPA-treated mice and control mice. RESULTS Eighteen lipid biomarkers were associated with pCPA-induced serotonin deficiency. Specifically, lipid species of lysophosphatidylethanolamine (LPE), phosphatidylethanolamine (PE), sphingomyelin (SM), galactosylceramide (GalCer), glucotosylceramide (GluCer), lactosylceramide (LacCer) and triacylglycerol (TG) were down-regulated whereas glycerophosphocholine (PC) and phosphatidylinositol (PI) were up-regulated in the pCPA-treated mice compared with control mice. CONCLUSIONS This work demonstrates the significant effects of serotonin deficiency on lipid metabolisms and will facilitate improved understanding of pathomechanisms in serotonin deficiency, particularly from a lipidomics perspective.

Study on the interaction of uranyl with sulfated beta-cyclodextrin by affinity capillary electrophoresis and molecular dynamics simulation.

The study on sulfated beta‐cyclodextrin binding to uranyl ion helps to get a better understanding of uranyl compounds’ intermolecular interaction mechanism and facilitates the structure‐based design of uranyl binding molecules. Here we investigated the electromigration of the inclusion complex by using affinity capillary electrophoresis in acidic solution. The binding constant was determined to be logK = 2.96 ± 0.02 (R2 = 0.996) through nonlinear regression approach. The possible configurations and structural features of the inclusion complex were further studied by molecular dynamics simulation. The results suggest the distinctions of coordination environment and hydration compared with bare uranyl ion in aqueous solution. Thus, two water oxygen atoms coordinated with uranyl in the first hydration shell at 2.55 angstrom instead of five in the same distance range. The binding free energy was calculated as –12.10 ± 1.46 kcal/mol by means of thermodynamic perturbation method. The negative value indicates that the process of S‐β‐CD capture uranyl ion in the aqueous media is spontaneous.

Lipidomic analysis of plasma in patients with lacunar infarction using normal-phase/reversed-phase two-dimensional liquid chromatography-quadrupole time-of-flight mass spectrometry

AbstractStroke is a major cause of mortality and long-term disability worldwide. The study of biomarkers and pathogenesis is vital for early diagnosis and treatment of stroke. In the present study, a continuous-flow normal-phase/reversed-phase two-dimensional liquid chromatography–quadrupole time-of-flight mass spectrometry (NP/RP 2D LC-QToF/MS) method was employed to measure lipid species in human plasma, including healthy controls and lacunar infarction (LI) patients. As a result, 13 lipid species were demonstrated with significant difference between the two groups, and a “plasma biomarker model” including glucosylceramide (38:2), phosphatidylethanolamine (35:2), free fatty acid (16:1), and triacylglycerol (56:5) was finally established. This model was evaluated as an effective tool in that area under the receiver operating characteristic curve reached 1.000 in the discovery set and 0.947 in the validation set for diagnosing LI patients from healthy controls. Besides, the sensitivity and specificity of disease diagnosis in validation set were 93.3% and 96.6% at the best cutoff value, respectively. This study demonstrates the promising potential of NP/RP 2D LC-QToF/MS-based lipidomics approach in finding bio-markers for disease diagnosis and providing special insights into the metabolism of stroke induced by small vessel disease. Graphical abstractFlow-chart of the plasma biomarker model establishment through biomarker screening and validation

A plasma lipidomics strategy reveals perturbed lipid metabolic pathways and potential lipid biomarkers of human colorectal cancer

To explore underlying molecular mechanisms and identify novel lipid biomarkers promising for colorectal cancer (CRC) diagnosis, a continuous-flow two dimensional liquid chromatography-quadrupole time-of-flight mass spectrometry (2D LC-QToF/MS) method was employed to comprehensively measure lipid species in human plasma of CRC patients and healthy controls. With a total of 427 annotated lipid species, we identified 64 lipid species with corrected p value less than 0.05 and fold change more than 1.5. These significantly altered lipid species were mainly involved in glycerolipids and glycerophospholipids metabolism and sphingolipids metabolism. After the diagnosis ability evaluation based on the receiver operating characteristic (ROC) curve, phosphatidylglycerol (34:0), sphingomyelin (42:2), ceramide (44:5), lysophosphatidylcholine (18:3), lysophosphatidylcholine (18:2), phosphatidylethanolamine (O-36:3), phosphatidylethanolamine (O-38:3) and sphingomyelin (38:8) were finally proposed as the potential biomarkers with the area under the curve (AUC) more than 0.900. These results suggest that this 2D LC-QToF/MS-based lipidomics profiling has great potential as a noninvasive diagnostic method in detecting CRC and hopefully provide new clues to understand its underlying mechanism.

Metabolomic Analysis of Mouse Embryonic Fibroblast Cells in Response to Autophagy Induced by Acute Starvation

Autophagy-related protein 7 (Atg7) is essential in the formation of the autophagophore and is indispensable for autophagy induction. Autophagy will exist in lower level or even be blocked in cells without Atg7. Even though the possible signaling pathways of Atg7 have been proposed, the metabolomic responses under acute starvation in cells with and without Atg7 have not been elucidated. This study therefore was designed and aimed to reveal the metabolomics of Atg7-dependent autophagy through metabolomic analysis of Atg7−/− mouse embryonic fibroblast cells (MEFs) and wild-type MEFs along with the starvation time. 30 significantly altered metabolites were identified in response to nutrient stress, which were mainly associated with amino acid, energy, carbohydrate, and lipid metabolism. For the wild-type MEFs, the induction of autophagy protected cell survival with some up-regulated lipids during the first two hours’ starvation, while the subsequent apoptosis resulted in the decrease of cell viability after four hours’ starvation. For the Atg7−/− MEFs, apoptosis perhaps led to the deactivation of tricarboxylic acid (TCA) cycle due to the lack of autophagy, which resulted in the immediate drop of cellular viability under starvation. These results contributed to the metabolomic study and provided new insights into the mechanism associated with Atg7-dependent autophagy.

Lipid profiling of cyanobacteria Synechococcus sp. PCC 7002 using two‐dimensional liquid chromatography with quadrupole time‐of‐flight mass spectrometry

Glycerolipid is a main component of membranes in oxygenic photosynthetic organisms. Up to now, the majority of publication in this area has focused on the physiological functions of glycerolipids and lipoprotein complexes in photosynthesis, but the study on the separation and identification of glycerolipids in thylakoid membrane in cyanobacteria is relatively rare. Here we report a new method to separate and identify five photosynthetic glycerolipid classes, including monoglucosyl diacylglycerol, monogalactosyl diacylglycerol, digalactosyl diacylglycerol, sulfoquinovosyl diacylglycerol, and phosphatidylglycerol, in cyanobacteria Synechococcus sp. PCC 7002 by two-dimensional (normal- and reversed-phase) liquid chromatography online coupled to quadrupole time-of-flight mass spectrometry. Over twice as many lipid species were detected by our method compared to the previously reported methods. Ten new odd-chain fatty acid glycerolipids were discovered for the first time. Moreover, complete separation of isomers of monogalactosyl diacylglycerol and monoglucosyl diacylglycerol was achieved. According to the tandem mass spectrometry results, we found that the head group of monoglucosyl diacylglycerols was not as stable as that of monogalactosyl diacylglycerols, which might explain why the organism chose monogalactosyl diacylglycerols and digalactosyl diacylglycerols instead of monoglucosyl diacylglycerols as the main content of the photosynthetic membranes in the history of evolution. This work will benefit further research on the physiological function of glycerolipids.