Emerging role of metabolomics in protein conformational disorders.

Introduction: Metabolomics focuses on interactions among different metabolites associated with various cellular functions in cells, tissues, and organs. In recent years, metabolomics has emerged as a powerful tool to identify perturbed metabolites, pathways influenced by the environment, for protein conformational diseases (PCDs) and also offers wide clinical application.Area Covered: This review provides a brief overview of recent advances in metabolomics as applied to identify metabolic variations in PCDs, such as Alzheimer's disease, Parkinson's disease, Huntington's disease, prion disease, and cardiac amyloidosis. The 'PubMed' and 'Google Scholar' database search methods have been used to screen the published reports with key search terms: metabolomics, biomarkers, and protein conformational disorders.Expert opinion: Metabolomics is the large-scale study of metabolites and is deemed to overwhelm other omics. It plays a crucial role in finding variations in diseases due to protein conformational changes. However, many PCDs are yet to be identified. Metabolomics is still an emerging field; there is a need for new high-resolution analytical techniques and more studies need to be carried out to generate new information.

Secretome Profiling and Computational Biology of Human Leiomyoma Samples Unravel Molecular Signatures with Potential for Diagnostic and Therapeutic Interventions.

In recent years, significant advancements have been made in the way the complex proteome samples are compared but the ultimate goal of routine biomarker discovery has yet to be achieved. Based on reverse genetic strategy, our study involved the spotting of genes showing expressional variability in uterine leiomyoma females. Serum samples were taken from uterine leiomyomas subjects (n=6) and healthy control subjects (n=6) for proteomic studies. Additionally, leiomyoma tissue samples (n=25) and normal myometrium samples (n=25) were taken for validation studies. In this study, we profiled the proteomes of uterine leiomyoma patient's serum and healthy control, along with relative quantification using Nano LC-MS/MS analysis. A total of 146 proteins were reported to be significantly differentially expressed (P value less than 0.05) in case and control sample. Statistical analysis identified a number of molecular signatures distinguishing healthy from diseased serum. Among these, five proteins lumican, ficolin, MASP2, EMSY, and kallistatin were further chosen according to their function for validation. Kallistatin was downregulated while ficolin, MASP2, lumican, and EMSY were found to be upregulated in the diseased sample. The expression modulations in the identified genes were further validated in twenty-five cases. Interactions among the differentially expressed proteins were identified followed with network analysis. Network analysis emphasized important pathways that are highly deregulated in myoma, and functional significance of these pathways in the pathology of the disease was discussed. Comparative expression analysis reveals distinct molecular signatures and their probable role in diagnosis of the disease.

Differential levels of CHMP2B, LLPH, and SLC25A51 proteins in secondary renal amyloidosis.

OBJECTIVES: Renal amyloidosis (RA) is a rare disease, typically manifested with proteinuria, nephrotic syndrome, and ultimately leads to renal failure. The present study aims to profile the proteomes of renal amyloidosis patient's serum and healthy controls, along with relative quantification to find out robust markers for RA. METHODS: In this study, 12 RA patients and their corresponding age and gender-matched healthy controls were recruited from the Nephrology department of Max Super Specialty Hospital, New Delhi. We employed gel-based proteomic approach coupled with MALDI-TOF MS to compare protein expression patterns in RA patients and controls. Furthermore, validation of differential proteins (selected) was done using bio-layer interferometry. RESULTS: Eleven proteins showed remarkably altered expression levels. Moreover, expression modulation of three proteins (LLPH, SLC25A51, and CHMP2B) was validated which corroborated with two-dimensional gel electrophoresis (2-DE) results showing significant upregulation (p < 0.05) in RA patients followed by ROC analysis which demonstrated the diagnostic potential of these proteins. A protein-protein master network was generated implicating the above identified proteins along with their interactors, fishing out the routes leading to amyloidosis. CONCLUSION: This study indicates that the identified serum proteomic signatures could improve early diagnosis and lead to possible therapeutic targets in RA.

Differential expression of MAP3K7 and TROPONIN C proteins and related perturbations in renal amyloidosis.

OBJECTIVES: Renal amyloidosis (RA) is a rare protein misfolding disorder that prompts progressive renal insufficiency. This study aimed to decipher proteomic changes in human sera to understand the pathophysiology and molecular mechanisms underlying the disease development, hence assisting in the diagnosis of RA. METHODS: Serum proteomic analysis was performed using a gel-based approach followed by MALDI-TOF MS. RA patients with age and sex matched healthy volunteers were recruited from Max Super Speciality Hospital, New Delhi, India. RESULTS: Proteome profiles of serum revealed eight differentially expressed proteins namely, Zinc finger protein 624, Protein FAM183A, Calcium-binding mitochondrial carrier protein Scamc-3, V-type proton ATPase 116 kDa subunit A isoforms 2, Protein TXNRD3NB, ATP - dependent RNA helicase, Troponin C and Mitogen-activated protein kinase kinase kinase 7. These proteins were reported first time in RA. The increased levels of MAP3K7 and TROPONIN C were validated by bio-layer interferometry and their diagnostic accuracy was evaluated by ROC curve analysis. The differentially expressed proteins were predominantly associated with vesicular trafficking, transcriptional regulation, metabolic processes, apoptotic process and mitochondrial metabolism. CONCLUSION: The results indicate that these proteomic signatures may be considered as potential molecular targets for RA diagnostics and therapeutics subject to validation on large sample size. Abbreviations: AßP= Amyloid-beta protein, Aß=Amyloid-beta, AL= Light chain amyloidosis, AA= Amyloid A, ALECT2= LECT2 amyloidosis, APS= Ammonium persulfate CKD= Chronic Kidney Diseases, EBRT= external beam radiation therapy, ESRD= End-Stage Kidney Disease, Glis2= Gli-similar 2, JNK= c-Jun NH 2-terminal kinase, MAPK= Mitogen-Activated Protein Kinase, MM=Multiple Myeloma, PHD= Prolyl hydroxylase, RA = Renal Amyloidosis, SAA= Serum Amyloid A, SD= Standard Deviation, Sepp= Selenoprotein, SCC= Squamous cell carcinoma, SDS= Sodium dodecyl sulfate, TEMED = tetramethyl ethylenediamine, TGF-Beta-1=Transforming growth factor- Beta-1, Trx = Thioredoxin, TrxR= Thioredoxin reductase.

Renal amyloidosis: an update on diagnosis and pathogenesis.

Amyloidosis is a diverse group of protein conformational disorder which is caused by accumulation and deposition of insoluble protein fibrils in vital tissues or organs, instigating organ dysfunction. Renal amyloidosis is characterized by the acellular Congo red-positive pathologic deposition of amyloid fibrils within glomeruli and/or the interstitium. It is generally composed of serum amyloid A-related protein or an immunoglobulin light chain; other rare forms lysozyme, gelsolin, fibrinogen alpha chain, transthyretin, apolipoproteins AI/AII/AIV/CII/CIII; and the recently identified form ALECT2. This disease typically manifests with heavy proteinuria, nephrotic syndrome, and finally progression to end-stage renal failure. Early diagnosis of renal amyloidosis is arduous as its symptoms appear in later stages with prominent amyloid deposition. The identification of the correct type of amyloidosis is quite troublesome as it can be confused with another related form. Therefore, the exact typing of amyloid is essential for prognosis, treatment, and correct management of renal amyloidosis. The emanation of new techniques of proteomic analysis, for instance, mass spectroscopy/laser microdissection, has provided greater accuracy in amyloid typing. This in-depth review emphasizes on the clinical features, renal pathological findings, and diagnosis of the AL and non-AL forms of renal amyloidosis.

Differential Expression of MARK4 Protein and Related Perturbations in Females with Ovulatory PCOS.

BACKGROUND: Ovulatory PCOS (OPCOS) is the mildest form of the polycystic ovarian syndrome among all four determined phenotypes. Though the females with OPCOS are ovulating, hyperandrogenism and polycystic ovarian morphology increase the susceptibility of cardiovascular diseases, insulin resistance, hyperlipidemia and metabolic syndrome in these females. OBJECTIVES: The aim of the study was to identify the significance associated with OPCOS phenotype through serum proteomic profiling of OPCOS females and normal age-matched healthy ovulating females. METHODS: One and two-dimensional gel-based proteomic approaches were adopted to fractionate the complex serum proteome. Differential protein profiles generated were analyzed with PD-QUEST Software. Protein spots differing in intensity by >2-fold were selected and identified further by MALDI-TOF MS. Validation of identified protein was carried out by Biolayer Interferometry. RESULTS: One and two-dimensional gel profiles revealed a differential expression pattern of proteins. 10 selected spots were identified as GMP synthase [glutamine hydrolyzing], zinc finger protein 518A, pericentriolar material 1 protein, BCLAF1 and THRAP3 family member 3, MAP/microtubule affinityregulating kinase 4, H/ACA ribonucleoprotein complex subunit 1, Melanoma-associated antigen B3 and Zinc finger protein 658B. Expression of MAP/microtubule affinity-regulating kinase 4 (MARK4) was found to be downregulated in OPCOS females as compared to controls on validation. CONCLUSION: Reduced expression of MARK4 protein in OPCOS increases the associated risk of hyperlipidemia, hyperandrogenism and metabolic syndrome, thus the protein holds strong candidature as a drug target for the syndrome.

Effect of Saliva on the Tensile Bond Strength of Different Generation Adhesive Systems: An In-Vitro Study.

BACKGROUND: Newer development of bonding agents have gained a better understanding of factors affecting adhesion of interface between composite and dentin surface to improve longevity of restorations. OBJECTIVE: The present study evaluated the influence of salivary contamination on the tensile bond strength of different generation adhesive systems (two-step etch-and-rinse, two-step self-etch and one-step self-etch) during different bonding stages to dentin where isolation is not maintained. MATERIALS AND METHODS: Superficial dentin surfaces of 90 extracted human molars were randomly divided into three study Groups (Group A: Two-step etch-and-rinse adhesive system; Group B: Two-step self-etch adhesive system and Group C: One-step self-etch adhesive system) according to the different generation of adhesives used. According to treatment conditions in different bonding steps, each Group was further divided into three Subgroups containing ten teeth in each. After adhesive application, resin composite blocks were built on dentin and light cured subsequently. The teeth were then stored in water for 24 hours before sending for testing of tensile bond strength by Universal Testing Machine. The collected data were then statistically analysed using one-way ANOVA and Tukey HSD test. RESULTS: One-step self-etch adhesive system revealed maximum mean tensile bond strength followed in descending order by Two-step self-etch adhesive system and Two-step etch-and-rinse adhesive system both in uncontaminated and saliva contaminated conditions respectively. CONCLUSION: Unlike One-step self-etch adhesive system, saliva contamination could reduce tensile bond strength of the two-step self-etch and two-step etch-and-rinse adhesive system. Furthermore, the step of bonding procedures and the type of adhesive seems to be effective on the bond strength of adhesives contaminated with saliva.

Reference values for three channels of amplitude-integrated EEG using the Brainz BRM3 cerebral function monitor in normal term neonates: a pilot study.

BACKGROUND: Amplitude-integrated electroencephalography (EEG) is a form of continuous EEG using a select number of electrodes (2-4), which can be used for bedside monitoring of brain functions in critically ill neonates. There is a paucity of normative amplitude-integrated EEG data for term healthy neonates especially for unilateral channels that are available for newer cerebral function monitors. OBJECTIVE: To define absolute amplitudes for all three available channels and also to determine if route of delivery or presence of a caput succedaneum would affect amplitude-integrated EEG amplitude voltages. METHODS: This is a prospective observational study of 80 healthy term neonates (gestational age ≥ 38 weeks) who had three-channel amplitude-integrated EEG recorded for 90 minutes within 12 hours of birth using the Brainz BRM3 cerebral function monitor. RESULTS: Median maximum and median minimum voltages obtained were 16.96 µV and 8.13 µV for the cross-cerebral (CC), 14.42 µV and 7.13 µV for the right unilateral, and 13.16 µV and 6.51 µV for the left unilateral aEEG channels, respectively. There were no statistically significant difference amplitude voltages for any channel based on route of delivery. The presence of a caput succedaneum was associated with a decrease in the median and mean of the maximum and minimum amplitude voltages for CC channel. Median maximum and median minimum voltages for the CC channel among the caput and normal scalp examination groups were 14.62 µV vs 17.27 µV (P = 0.022) and 7.21 µV vs 8.24 µV (P = 0.004), respectively. Similarly, mean maximum and mean minimum voltages for the CC channel were 15.42 µV vs 17.59 µV (P = 0.038) and 7.27 µV vs 8.25 µV (P = 0.005) in the caput and normal scalp examination groups, respectively.

Assessment of the effect of macromolecular crowding on aggregation behaviour of a model amyloidogenic Peptide.

Accumulation of ordered protein aggregates (or amyloids) represents a hallmark of many diseases (e.g., Alzheimer's disease, type II diabetes, Parkinson's diseases etc.), results from intermolecular association of partially unfolded proteins/ peptides. Such associations usually take place in highly crowded conditions. The aggregates, which are formed under in vitro and in vivo conditions exhibit substantial variations in their structure and function. Such heterogeneities in amyloids might arise due to macromolecular crowding that is usually omitted under in vitro conditions. The current study is an attempt to assess the effects of macromolecular crowding on amyloid formation using a model amyloidogenic peptide. The sequence of the peptide was derived from C-terminal region (RATQIPSYKKLIMY) of PAP(248-286), which naturally occurs in human semen as amyloid aggregates and is known for boosting HIV infectivity. This model peptide forms sedimentable and fibrillar aggregates in aqueous buffer and shows the characteristic features of amyloids. In the presence of macromolecular crowders the morphological features of the amyloids are significantly altered and resulted in the formation of shorter amyloid aggregates. The current study assumes the hypothesis that macromolecular crowding in the biological system favours formation of heterogeneous classes of aggregates and each of them might differ in their biophysical and biological properties.

Epidemiologic findings of cancer detected in a pediatric emergency department.

This study describes epidemiologic findings of pediatric cancer diagnosed in the emergency department (ED) setting. Medical records are retrospectively reviewed on all patients in the hospital's cancer database between 2000 and 2004 who were diagnosed as having cancer or whose oncologic diagnosis was missed during their ED presentation. Of 427 patients identified in the cancer database, 18% (77 of 427) are analyzed. Oncologic diagnosis was missed in 5% (4 of 77) of the eligible patients initially presenting to the ED. The incidence of cancer in the ED is 22.8 cases per 100,000 ED visits. The most prevalent cancer is related to the hematologic system (37.7%), followed by the central nervous system (31.2%) and the abdomen (22.1%). Hematologic, central nervous system, and abdominal cancers constitute approximately 90% of all childhood cancers. Cancer is diagnosed frequently in our ED patient population. Based on the prevalence of certain tumors, the diagnostic approach to children with hematologic, neurologic, or abdominal complaints should include evaluation for any underlying cancer.