Xian-Long Cheng

Urinary metabonomics study on biochemical changes in an experimental model of chronic renal failure by adenine based on UPLC Q-TOF/MS

BACKGROUND Chronic renal failure (CRF) is a serious clinical symptom, occurring as the end result of all kinds of chronic kidney disease and its pathophysiological mechanism is not yet well understood. We investigated the metabolic profiling of urine samples from CRF model rats to find potential disease biomarkers and research pathology of CRF. METHODS An animal model of CRF was produced by adenine. Metabolic profiling of the urine was performed by using ultra performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC Q-TOF/MS). Acquired data were subjected to principal component analysis (PCA) for differentiating the CRF and the normal control groups. Potential biomarkers were screened by using S-plot and were identified by the accurate mass, isotopic pattern and MS(E) fragments information obtained from UPLC Q-TOF/MS analysis. RESULTS 12 metabolites in urine were identified as potential biomarkers. Adenine-induced CRF rats were characterized by the increase of phytosphingosine, adrenosterone, tryptophan, 2,8-dihydroxyadenine, creatinine, and dihydrosphingosine together with the decrease of N-acetylleucine, 3-O-methyldopa, ethyl-N2-acetyl-L-argininate, dopamine, phenylalanine and kynurenic acid in urine. The altered metabolites demonstrated perturbations of amino acids metabolism, phospholipids metabolism and creatinine metabolism in CRF rats. CONCLUSION This work shows that metabonomics method is a valuable tool in CRF mechanism study and assists in clinical diagnosis of CRF.

Intrarenal Metabolomic Investigation of Chronic Kidney Disease and its TGF-β1 Mechanism in Induced-adenine Rats using UPLC Q-TOF/HSMS/MSE

Chronic kidney disease (CKD) is becoming a worldwide public health problem. In this study, a kidney metabonomics method based on the ultra performance liquid chromatography/high-sensitivity mass spectrometry with MS(E) data collection technique was undertaken to explore the excretion pattern of low molecular mass metabolites in rat model of adenine-induced chronic renal failure (CRF). Coupled with blood biochemistry and kidney histopathology results, the significant difference in metabolic profiling between the adenine-induced CRF group and the control group by using pattern recognition analysis indicated that changes in global tissue metabolites were occurred. Some significantly changed metabolites like fatty acids, p-cresol sulfate, and indoxyl sulfate have been identified. The results showed that the most important CRF-related metabolites were polyunsaturated fatty acids, indoxyl sulfate, and p-cresyl sulfate. Indoxyl sulfate and p-cresyl sulfate (uremic toxins) were significantly increased in CRF rats. Indoxyl sulfate and p-cresyl sulfate stimulate progressive tubulointerstitial fibrosis by increasing the expression of transforming growth factor-β1 (TGF-β1). These biochemical changes in tissue metabolites are related to the perturbations of fatty acid metabolism and amino metabolism, which may be helpful to further understand the TGF-β1 mechanisms of tubulointerstitial fibrosis. This work shows that the metabonomics method is a valuable tool for studying the essence of CKD.

Effect of ergosta-4,6,8(14),22-tetraen-3-one (ergone) on adenine-induced chronic renal failure rat: a serum metabonomic study based on ultra performance liquid chromatography/high-sensitivity mass spectrometry coupled with MassLynx i-FIT algorithm.

BACKGROUND Ergosta-4,6,8(14),22-tetraen-3-one (ergone) has been proven to prevent the progression of renal injury and the subsequent renal fibrosis. We investigated the therapeutic effects and mechanism of ergone on a chronic renal failure model of rats induced by adenine. METHODS A serum metabonomic method based on the UPLC Q-TOF/MS was undertaken to explore the excretion pattern of low molecular mass metabolites. RESULTS Coupled with blood biochemistry and kidney histopathology results, the significant difference in metabolic profiling between adenine-induced chronic renal failure group and the ergone treated group by using pattern recognition analysis indicated that changes in global serum metabolites occurred. Some significantly changed metabolites like lysophosphatidylcholines, adenine, dopamine, creatinine, aspartic acid and phenylalanine have been found and identified. These biochemical changes in serum metabolites are related to the perturbations of amino acid metabolism and lecithin metabolism, which may be helpful to further understand the chronic renal failure and therapeutic mechanisms of ergone. CONCLUSION The work shows that the metabonomic method is a valuable tool for studying the essence of chronic kidney disease and therapeutic effect mechanism of preclinical or clinical drug.

Serum metabonomics study of adenine-induced chronic renal failure in rats by ultra performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry

An ultra performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC Q-TOF MS) metabonomics approach was employed to study the serum metabolic profiling of adenine-induced chronic renal failure (CRF) rats. Acquired data were subjected to principal component analysis (PCA) for differentiating the CRF and the normal control groups. Potential biomarkers were screened by using S-plot and were identified by the accurate mass, isotopic pattern and MS/MS fragments information obtained from UPLC Q-TOF MS analysis. Significant differences in the serum level of creatinine, amino acids and LysoPCs were observed, indicating the perturbations of amino acid metabolism and phospholipid metabolism in adenine-induced CRF rats. This research proved that metabonomics is a promising tool for disease research.

Application of faecal metabonomics on an experimental model of tubulointerstitial fibrosis by ultra performance liquid chromatography/high-sensitivity mass spectrometry with MSE data collection technique

Chronic renal failure (CRF) is a major challenge for the public healthcare problem. A novel UPLC Q-TOF/MS method with MSE data collection mode was developed as a very effective biochemical analytical tool for precise identification of important biomarkers in the adenine-induced CRF rats. Nine endogenous metabolites were identified by using metabonomic method combined with multivariate data analysis, the accurate mass, isotopic pattern, MSE fragments information and MassLynx i-FIT algorithm. The identified metabolites indicated the perturbations of bile acid and phospholipid metabolism are related to CRF rats. This work shows that metabonomics method is a valuable tool in CRF mechanism study.

UPLC-Q-TOF/HSMS/MSE-based metabonomics for adenine-induced changes in metabolic profiles of rat faeces and intervention effects of ergosta-4,6,8(14),22-tetraen-3-one

Ergosta-4,6,8(14),22-tetraen-3-one (ergone), isolated from the medicinal fungus Polyporus umbellatus, has been proven to prevent the progression of renal injury and the subsequent renal fibrosis. Ultra performance liquid chromatography coupled with quadrupole time-of-flight high-sensitivity mass spectrometry and a novel mass spectrometry(Elevated Energy) (MS(E)) data collection technique was employed to investigate metabonomic characters of chronic renal failure (CRF) induced adenine and the protective effects of ergosta-4,6,8(14),22-tetraen-3-one (ergone). Coupled with blood biochemistry and kidney histopathology results, the significant difference in metabolic profiling between adenine-induced CRF group and ergone-treated CRF group by using pattern recognition analysis indicated that changes in global faecal metabolites were occurred. Seven endogenous metabolites were identified by using metabonomic method combined with multivariate data analysis, the accurate mass, isotopic pattern, MS(E) fragments information and MassLynx i-FIT algorithm. These biochemical changes in faecal metabolites are related to the perturbations of bile acid metabolism and phospholipid metabolism, which may be helpful to further understand the CRF and therapeutic mechanisms of ergone. This research proved that MS(E) can simultaneous acquire precursor ion information and fragment ion data at high and low collision energy in one analytical run, which facilitated the fast structural characterization of metabolites.

Metabolomics analysis reveals the association between lipid abnormalities and oxidative stress, inflammation, fibrosis, and Nrf2 dysfunction in aristolochic acid-induced nephropathy

Alternative medicines are commonly used for the disease prevention and treatment worldwide. Aristolochic acid (AAI) nephropathy (AAN) is a common and rapidly progressive interstitial nephropathy caused by ingestion of Aristolochia herbal medications. Available data on pathophysiology and molecular mechanisms of AAN are limited and were explored here. SD rats were randomized to AAN and control groups. AAN group was treated with AAI by oral gavage for 12 weeks and observed for additional 12 weeks. Kidneys were processed for histological evaluation, Western blotting, and metabolomics analyses using UPLC-QTOF/HDMS. The concentrations of two phosphatidylcholines, two diglycerides and two acyl-carnitines were significantly altered in AAI treated rats at week 4 when renal function and histology were unchanged. Data obtained on weeks 8 to 24 revealed progressive tubulointerstitial fibrosis, inflammation, renal dysfunction, activation of NF-κB, TGF-β, and oxidative pathways, impaired Nrf2 system, and profound changes in lipid metabolites including numerous PC, lysoPC, PE, lysoPE, ceramides and triglycerides. In conclusion, exposure to AAI results in dynamic changes in kidney tissue fatty acid, phospholipid, and glycerolipid metabolisms prior to and after the onset of detectable changes in renal function or histology. These findings point to participation of altered tissue lipid metabolism in the pathogenesis of AAN.

UPLC-based metabonomic applications for discovering biomarkers of diseases in clinical chemistry

OBJECTIVES Metabonomics is a powerful and promising analytic tool that allows assessment of global low-molecular-weight metabolites in biological systems. It has a great potential for identifying useful biomarkers for early diagnosis, prognosis and assessment of therapeutic interventions in clinical practice. The aim of this review is to provide a brief summary of the recent advances in UPLC-based metabonomic approach for biomarker discovery in a variety of diseases, and to discuss their significance in clinical chemistry. DESIGN AND METHODS All the available information on UPLC-based metabonomic applications for discovering biomarkers of diseases were collected via a library and electronic search (using Web of Science, Pubmed, ScienceDirect, Springer, Google Scholar, etc.). RESULTS Metabonomics has been used in clinical chemistry to identify and evaluate potential biomarkers and therapeutic targets in various diseases affecting the liver (hepatocarcinoma and liver cirrhosis), lung (lung cancer and pneumonia), gastrointestinal tract (colorectal cancer) and urogenital tract (prostate cancer, ovarian cancer and chronic kidney disease), as well as metabolic diseases (diabetes) and neuropsychiatric disorders (Alzheimers disease and schizophrenia), etc. CONCLUSIONS The information provided highlights the potential value of determination of endogenous low-molecular-weight metabolites and the advantages and potential drawbacks of the application of UPLC-based metabonomics in clinical setting.

Metabolomics insights into chronic kidney disease and modulatory effect of rhubarb against tubulointerstitial fibrosis

Chronic kidney disease (CKD) is a major public health problem worldwide. Rhubarb has been shown to have nephroprotective and anti-fibrotic activities in patients with CKD. However, bioactive fractions and biochemical mechanism of anti-fibrotic properties of rhubarb remain unclear. Here we applied ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry together with univariate and multivariate statistical analyses to investigate the urinary metabolite profile in rats with adenine-induced CKD treated with the petroleum ether (PE)-, ethyl acetate (EA)- and n-butanol (BU)- extracts of rhubarb. Significant differences in renal function, kidney histopathology as well as metabolic profiles were observed between CKD and control rats. Changes in these parameters reflected characteristic phenotypes of CKD rats. We further identified a series of differential urinary metabolites for CKD rats, suggesting metabolic dysfunction in pathway of amino acid, purine, taurine, and choline metabolisms. Treatment with EA, BU and PE extracts of rhubarb improved renal function and histopathological abnormalities including interstitial fibrosis and inflammation, and either fully or partially reversed the abnormalities of the urinary metabolites. Among them, the nephroprotective effect of EA extract was stronger than BU and PE extracts. This work provides important mechanistic insights into the CKD and nephroprotective effects of different rhubarb extract against tubulo-interstitial fibrosis.

Ergosta-4,6,8(14),22-tetraen-3-one induces G2/M cell cycle arrest and apoptosis in human hepatocellular carcinoma HepG2 cells.

BACKGROUND Mushrooms have been used in Asia as traditional foods and medicines for a long time. Ergosta-4,6,8(14),22-tetraen-3-one (ergone) is one of the well-known bioactive steroids, which exists widely in various medicinal fungi such as Polyporus umbellatus, Russula cyanoxantha, and Cordyceps sinensis. Ergone has been demonstrated to possess cytotoxic activity. However, the molecular mechanisms by which ergone exerts its cytotoxic activity are currently unknown. METHODS In the present study, ergone possessed a remarkable anti-proliferative activity toward human hepatocellular carcinoma HepG2 cells. We assayed the cell cycle by flow cytometry using PI staining; investigated the exposure of phosphatidylserine at the outer layer of the cytoplasmic membrane by the FITC-annexin V/PI staining; observed the nuclear fragmentation by Hoechst 33258 staining and studied the protein expression of Bax, Bcl-2, p-53, procaspase-3, -8, -9, PARP and cleaved PARP by Western blotting analysis. RESULTS Cells treated with ergone showed typical markers of apoptosis: G2/M cell cycle arrest, chromatin condensation, nuclear fragmentation, and phosphatidylserine exposure. Furthermore, PARP-cleavage; activation of caspase-3, -8, -9; up-regulation of Bax and down-regulation of Bcl-2 were observed in HepG2 cells treated with ergone, which show that both the intrinsic and extrinsic apoptotic pathways are involved in ergone-induced apoptosis in HepG2 cells. Ergosta-4,6,8(14),22-tetraen-3-one induces G2/M cell cycle arrest and apoptosis in HepG2 cells in a caspase-dependent manner. GENERAL SIGNIFICANCE In this study, we reported for the first time that ergone-induced apoptosis through activating the caspase. These results would be useful for the further utilization of many medicinal fungi in cancer treatment.