Mindi Zhao

Acute kidney injury (AKI) is a frequent complication following on-pump coronary artery bypass graft surgery (CABG). To better investigate the effect of CABG on kidney function and the pathological process of AKI during the operation, the urine metabolic and protein signatures of four time points in 55 postoperative AKI and 104 non-AKI patients after CABG were analyzed by using high-resolution tandem mass spectrometry combined with pattern recognition and functional annotation. In the longitudinal urinary metabolomics study, the protein catabolism pathway is prominent in AKI patients throughout the surgery and recovery process. The integration of metabolomic and proteomic data revealed that alterations in purine metabolism exhibited significant differences preoperatively between the AKI and the non-AKI cohorts. A panel of two proteins (ARFIP1 and SOD2) and two metabolites (tyrosyl-γ-glutamate and l-methionine) was discovered to have a good predicting performance (area under the curve, 0.92) in the preoperative urine. Our research suggests that the panel could play a significant role in distinguishing AKIs from non-AKIs prior to surgery. We propose targeting purine metabolism pathways that may help to identify potential renal protective approaches in acute kidney injury. This study helps to reveal changes in urinary metabolomics and proteomics after CABG and may provide diagnostic clues for patients with postoperative renal injury.

Multiple sclerosis is a chronic autoimmune demyelinating disease of the central nervous system and is difficult to diagnose in early stages. Without homeostatic control, urine was reported to have the ability to accumulate early changes in the body. We expect that urinary proteome can reflect early changes in the nervous system. The early urinary proteome changes in a most employed multiple sclerosis rat model (experimental autoimmune encephalomyelitis) were analysed to explore early urinary candidate biomarkers, and early treatment of methylprednisolone was used to evaluate the therapeutic effect. Twenty-five urinary proteins were altered at day 7 when there were no clinical symptoms and obvious histological changes. Fourteen were reported to be differently expressed in the serum/cerebrospinal fluid/brain tissues of multiple sclerosis patients or animals such as angiotensinogen and matrix metallopeptidase 8. Functional analysis showed that the dysregulated proteins were associated with asparagine degradation, neuroinflammation and lipid metabolism. After the early treatment of methylprednisolone, the incidence of encephalomyelitis in the intervention group was only 1/13. This study demonstrates that urine may be a good source of biomarkers for the early detection of multiple sclerosis. These findings may provide important information for early diagnosis and intervention of multiple sclerosis in the future.

The chemical modification of proteins refers to the covalent group reaction involved in their amino acid residues or chain ends which, in turn, change the molecular structure and function of the proteins. There are many types of molecular modifications in the human plasma proteome, such as phosphorylation, methylation, and acetylation. In this study, two groups of human plasma proteome at different age groups (old and young) were used to perform a comparison of global chemical modifications, as determined by tandem mass spectrometry (MS/MS) combined with non-limiting modification identification algorithms. The sulfhydryl in the cysteine A total of 4 molecular modifications were found to have significant differences: the succinylation and phosphorylation modification of cysteine (Cys, C) and the modification of lysine (Lys, K) with threonine (Thr, T) were significantly higher in the old group than in the young group, while the carbamylation of lysine was lower in the young group. Cysteine residue is an important group for forming disulphide bonds and maintaining the structure of the protein. Differential cysteine-related sulfydryl modifications may cause structural and functional changes. Lysine is a basic amino acid, and the modification of its amino group will change the charge state of the protein, which may affect the structure and function of the protein. In summary, four types of protein chemical modifications and substitutes were found to be significantly different in the plasma proteome in different age groups and their probabilities of random generation are lower by passing random grouping test. We speculate that there is an increase in certain modified proteins in the blood of the old people which, in turn, changes the function of those proteins. This change may be one of the reasons why the old people are more likely than the young people to be at risk for age-related diseases, such as metabolic diseases, cerebral and cardiovascular diseases, and cancer.

Purpose: As an important part of the data‐independent acquisition‐mass spectrometry (DIA‐MS) analysis approach, spectral library generation is closely related to the final protein identification and quantitation.Experimental design: In this study, an attempt is made to evaluate the influence of different sample separation methods (reversed‐phase liquid chromatography (RPLC) linear gradient elution and spin column step gradient fractionation) and data acquisition modes (data‐dependent acquisition (DDA) and DIA) on library generation of the human urine proteome.Results: The RPLC separation method provides more proteins for generating the library than the spin column; however, the spin column library leads to identification of more proteins when used for the urine proteome analysis. The DDA mode can provide a larger spectral library than the DIA mode and the DDA‐library leads to more protein identifications in the study. Use of a combined urinary protein library provides the most complete protein and peptide information, and the DIA approach can provide accurate protein quantitative information even for low‐abundance proteins.Conclusions and clinical relevance: When the combined library is used to analyze the normal individual urine proteome (18 males and 18 females), gender‐related and age‐related proteins are discovered and functionally analyzed. This strategy may benefit urine proteome DIA analysis.

Purpose: Body fluid is considered a rich source of disease biomarkers. Proteins in many body fluids have potential clinical applications for disease diagnostic and prognostic predictions.Experimental design: To determine differences in the protein components and functional features of body fluids, a proteomic comparison of five body fluids (plasma, urine, cerebrospinal fluid, saliva, and amniotic fluid) was conducted by high‐resolution mass spectrometry.Results: A total of 4717 nonredundant proteins were identified, and the concentrations of 3433 proteins were estimated by an intensity‐based algorithm quantitation method. Among them, 564 proteins were shared among the five body fluids, with common functions in the coagulation/prothrombin system and inflammatory response. A total of 36.7% of the proteins were detected in only one body fluid and were closely related to their adjacent tissues by function. The functional analysis of the remaining 2986 proteins showed that similar functions might be shared among different body fluids, which highlighted intimate connection in the body.Conclusions and clinical relevance: The quantitative comparative functional analysis indicated that body fluids might reflect the diverse functions of the whole body rather than the characteristics of their adjacent tissues. The above data might indicate the potential application of body fluids for biomarker discovery.

Scope: Pulmonary fibrosis (PF) is a progressive and devastating lung disease. With limited effective treatments available in the late stage, PF has a very poor prognosis. Molecular biomarkers are highly desired for PF, especially for its early phase.Materials and methods: Urine is a good biomarker source, and accumulates systemic changes in the body especially in the early‐stage of diseases. In this study, a bleomycin (BLM)‐induced rat model is used to mimic PF. Using labeled proteome quantitation, some urinary proteins are identified as candidate biomarkers of PF for early detection and disease monitoring. Then, prednisone treatment is administered at different phases of fibrosis.Results: Our results suggested that urine proteins could enable early detection and monitoring of both disease progression and treatment efficacy in the BLM‐induced PF model. Early prednisone treatment effectively inhibited pulmonary fibrosis, whereas the same treatment at a later phase had very limited effects. Meanwhile, five proteins showed the potential for monitoring therapeutic response.Conclusion: Urinary proteomics has been underutilized in respiratory diseases. These findings will improve our understanding of the pathogenesis of PF and accelerate biomarker discovery in respiratory diseases.

Multiple sclerosis is a chronic autoimmune demyelinating disease of the central nervous system and is difficult to diagnose in early stages. Without homeostatic control, urine was reported to have the ability to accumulate early changes in the body. We expect that urinary proteome can reflect early changes in the nervous system. In this study, the early urinary proteome changes in a most employed multiple sclerosis rat model (experimental autoimmune encephalomyelitis (EAE)) were analyzed to explore early urinary candidate biomarkers, and early treatment of methylprednisolone were used to evaluate the therapeutic effect. Compare with controls, twenty-five urinary proteins were altered at day 7 when there were no clinical symptoms and no obvious histological changes. Among them, twenty-three have human homologs and fourteen were reported to be differently expressed in the serum/cerebrospinal fluid/brain tissues of multiple sclerosis patients or animal models. Functional analysis showed that the dysregulated proteins were associated with asparagine degradation, neuroinflammation and lipid metabolism. After the early treatment of methylprednisolone, the incidence of encephalomyelitis in the intervention group was only 1/13. This study demonstrates that urine may be a good source of biomarkers for the early detection of multiple sclerosis and early treatment can significantly delay disease progression. These findings may provide important information for early diagnosis and intervention of multiple sclerosis in the future.

Urine is a better source than plasma for biomarker studies, as it can accumulate all changes in the body. Various candidate urinary biomarkers of physiological conditions, kidney disease and even brain dysfunction, have been detected in urine; however, urine has rarely been used to reflect cardiac diseases. As the clinical presentations of myocarditis are heterogeneous, reliable and sensitive diagnostic biomarkers of myocarditis are very important. In this study, candidate urinary biomarkers in the myosin-induced autoimmune myocarditis rat models were characterized using the isobaric tandem mass tag labeling approach coupled with high-resolution mass spectrometry. Compared with controls, forty-six urinary proteins were significantly changed in the myocarditis rats; among them, ten had previously been associated with myocarditis, twelve corresponding gene products have been annotated as mainly cardiovascular network genes by the Ingenuity Pathway Analysis, and four urinary proteins were validated by western blot.