Publications by authors named "Krishnan Viswanathan"

21 Publications

  • Page 1 of 1

Sclerostin and Its Role as a Bone Modifying Agent in Periodontal Disease.

J Oral Biosci 2021 Apr 17. Epub 2021 Apr 17.

Department of Periodontology, Rajah Muthiah Dental College & Hospital, Annamalai University, Annamalai Nagar, Chidambaram - 608002, Tamil Nadu, India. Electronic address:

Background: Periodontitis is a highly prevalent inflammatory disease affecting the periodontium that results from an imbalance between periodontopathogens and host mechanisms. Continuous progression of the disease may lead to tissue and bone destruction, eventually resulting in tooth loss. The extent of bone loss depends on the dysregulated host immune response. Various host-elicited molecules play a major role in disease progression. The discovery of the glycoprotein sclerostin and its role as a regulator of bone mass has led to a better understanding of bone metabolism.

Highlight: Sclerostin, which is primarily expressed by osteocytes, is a negative regulator of bone formation. It is a potent antagonist of the canonical Wingless-related integration site (Wnt) pathway, which is actively involved in bone homeostasis. Sclerostin is known to stimulate bone resorption by altering the osteoprotegerin (OPG)/receptor activator of nuclear factor kappa- β ligand (RANKL) balance. Additionally, in periodontitis, activation of the inflammatory cascade also increases the synthesis of sclerostin.

Conclusion: The recently discovered sclerostin antibody has emerged as a positive therapeutic tool for the treatment of metabolic bone diseases. It has been reported to improve bone strength, bone formation, osseointegration around implants and lower the risk of bone fractures in various animal and human models. This review describes the properties and action of sclerostin, its role in periodontal diseases, and the advent and efficacy of sclerostin antibodies.
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http://dx.doi.org/10.1016/j.job.2021.04.002DOI Listing
April 2021

NMR spectroscopy analysis reveals differential metabolic responses in arabidopsis roots and leaves treated with a cytokinesis inhibitor.

PLoS One 2020 6;15(11):e0241627. Epub 2020 Nov 6.

Department of Plant Sciences, University of California, Davis, CA, United States of America.

In plant cytokinesis, de novo formation of a cell plate evolving into the new cell wall partitions the cytoplasm of the dividing cell. In our earlier chemical genomics studies, we identified and characterized the small molecule endosidin-7, that specifically inhibits callose deposition at the cell plate, arresting late-stage cytokinesis in arabidopsis. Endosidin-7 has emerged as a very valuable tool for dissecting this essential plant process. To gain insights regarding its mode of action and the effects of cytokinesis inhibition on the overall plant response, we investigated the effect of endosidin-7 through a nuclear magnetic resonance spectroscopy (NMR) metabolomics approach. In this case study, metabolomics profiles of arabidopsis leaf and root tissues were analyzed at different growth stages and endosidin-7 exposure levels. The results show leaf and root-specific metabolic profile changes and the effects of endosidin-7 treatment on these metabolomes. Statistical analyses indicated that the effect of endosidin-7 treatment was more significant than the developmental impact. The endosidin-7 induced metabolic profiles suggest compensations for cytokinesis inhibition in central metabolism pathways. This study further shows that long-term treatment of endosidin-7 profoundly changes, likely via alteration of hormonal regulation, the primary metabolism of arabidopsis seedlings. Hormonal pathway-changes are likely reflecting the plant's responses, compensating for the arrested cell division, which in turn are leading to global metabolite modulation. The presented NMR spectral data are made available through the Metabolomics Workbench, providing a reference resource for the scientific community.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0241627PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7647083PMC
December 2020

Validation of Enthalpy-Entropy Compensation Mechanism in Partial Amide Bond Rotation.

ACS Omega 2020 Apr 13;5(16):9348-9355. Epub 2020 Apr 13.

Department of Chemistry, California State University, Fresno, California 93740, United States.

The concept of enthalpy-entropy compensation (EEC) is one of the highly debated areas of thermodynamics. The conformational change due to restricted double-bond rotation shows a classic two-site chemical exchange phenomenon and has been extensively studied. Fifty-four analogs of ,-diethyl--toluamide (DEET) as a model system were synthesized to study the thermodynamics of the partial amide bond character using nuclear magnetic resonance (NMR) spectroscopy. Line-shape analysis as a function of temperature is used to estimate the chemical exchange. Eyring analysis was then used to convert the chemical exchange rates to determine the transition state enthalpy and entropy of the molecules. The experimental design follows selective variations that perturb one aspect of the molecular system and its influence on the observed thermodynamic effect. The results of the study demonstrate that amide bond resonance in analogs of DEET follows an EEC mechanism. Simple modifications made to DEET's structural motif alter both the enthalpy and entropy of the system and were limited overall to a temperature compensation factor, = 292.20 K, 95% CI [290.66, 293.73]. We suggest EEC as a model to describe the kinetic compensation seen in chemical exchange phenomena in analogs of DEET.
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http://dx.doi.org/10.1021/acsomega.0c00332DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7191833PMC
April 2020

Role of glycosylation on the ensemble of conformations in the MUC1 immunodominant epitope.

J Pept Sci 2020 Jan 14;26(1):e3229. Epub 2019 Nov 14.

Department of Chemistry, California State University, 2555 E San Ramon Avenue, Fresno, CA, 93740, USA.

MUC1 is a membrane glycoprotein, which in adenocarninomas is overexpressed and exhibits truncated O-glycosylation. Overexpression and altered glycosylation make MUC1 into a candidate for immunotherapy. Monoclonal antibodies directed against MUC1 frequently bind an immunodominant epitope that contains a single site for O-glycosylation. Glycosylation with tumor carbohydrate antigens such as the Tn-antigen (GalNAc-O-Ser/Thr) results in antibodies binding with higher affinity. One proposed model to explain the enhanced affinity of antibodies for the glycosylated antigen is that the addition of a carbohydrate alters the conformational properties, favoring a binding-competent state. The conformational effects associated with Tn glycosylation of the MUC1 antigen was investigated using solution-state NMR and molecular dynamics. NMR experiments revealed distinct substructures of the glycosylated MUC1 peptides compared with the unglycosylated peptide. Molecular dynamics simulations of the MUC1 glycopeptide and peptide revealed distinguishing differences in their conformational preferences. Furthermore, the glycopeptide displayed a smaller conformational sampling compared with the peptide, suggesting that the glycopeptide sampled a narrower conformational space and is less dynamic. A comparison of the computed ensemble of conformations assuming random distribution, NMR models, and molecular dynamics simulations indicated that the MUC1 glycopeptide and aglycosylated peptide sampled structurally distinctly ensembles and that these ensembles were different from that of the random coil. Together, these data support the hypothesis that that conformational pre-selection could be an essential feature of these peptides that dictates the binding affinities to MUC1 specific antibodies.
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http://dx.doi.org/10.1002/psc.3229DOI Listing
January 2020

The Ensemble of Conformations of Antifreeze Glycoproteins (AFGP8): A Study Using Nuclear Magnetic Resonance Spectroscopy.

Biomolecules 2019 06 17;9(6). Epub 2019 Jun 17.

Department of Chemistry, California State University, Fresno, CA 93740, USA.

The primary sequence of antifreeze glycoproteins (AFGPs) is highly degenerate, consisting of multiple repeats of the same tripeptide, Ala-Ala-Thr*, in which Thr* is a glycosylated threonine with the disaccharide AFGPs seem to function as intrinsically disordered proteins, presenting challenges in determining their native structure. In this work, a different approach was used to elucidate the three-dimensional structure of AFGP8 from the Arctic cod and the Antarctic notothenioid . Dimethyl sulfoxide (DMSO), a non-native solvent, was used to make AFGP8 less dynamic in solution. Interestingly, DMSO induced a non-native structure, which could be determined via nuclear magnetic resonance (NMR) spectroscopy. The overall three-dimensional structures of the two AFGP8s from two different natural sources were different from a random coil ensemble, but their "compactness" was very similar, as deduced from NMR measurements. In addition to their similar compactness, the conserved motifs, Ala-Thr*-Pro-Ala and Ala-Thr*-Ala-Ala, present in both AFGP8s, seemed to have very similar three-dimensional structures, leading to a refined definition of local structural motifs. These local structural motifs allowed AFGPs to be considered functioning as effectors, making a transition from disordered to ordered upon binding to the ice surface. In addition, AFGPs could act as dynamic linkers, whereby a short segment folds into a structural motif, while the rest of the AFGPs could still be disordered, thus simultaneously interacting with bulk water molecules and the ice surface, preventing ice crystal growth.
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http://dx.doi.org/10.3390/biom9060235DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6628104PMC
June 2019

Molecular Thermodynamics Using Nuclear Magnetic Resonance (NMR) Spectroscopy.

Inventions (Basel) 2019 Mar 21;4(1). Epub 2019 Feb 21.

Department of Chemistry, California State University, Fresno, CA 93740, USA.

Nuclear magnetic resonance (NMR) spectroscopy is perhaps the most widely used technology from the undergraduate teaching labs in organic chemistry to advanced research for the determination of three-dimensional structure as well as dynamics of biomolecular systems... The NMR spectrum of a molecule under a given experimental condition is unique, providing both quantitative and structural information. In particular, the quantitative nature of NMR spectroscopy offers the ability to follow a reaction pathway of the given molecule in a dynamic process under well-defined experimental conditions. To highlight the use of NMR when determining the molecular thermodynamic parameters, a review of three distinct applications developed from our laboratory is presented. These applications include the thermodynamic parameters of (a) molecular oxidation from time-dependent kinetics, (b) intramolecular rotation, and (c) intermolecular exchange. An experimental overview and the method of data analysis are provided so that these applications can be adopted in a range of molecular systems.
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http://dx.doi.org/10.3390/inventions4010013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6528671PMC
March 2019

Proteomic profiles by multiplex microsphere suspension array.

J Immunol Methods 2018 10 10;461:1-14. Epub 2018 Jul 10.

Omni Array Biotechnology, Rockville, MD 20855, United States. Electronic address:

Advances in high-throughput proteomic approaches have provided substantial momentum to novel disease-biomarker discovery research and have augmented the quality of clinical studies. Applications based on multiplexed microsphere suspension array technology are making strong in-roads into the clinical diagnostic/prognostic practice. Conventional proteomic approaches are designed to discover a broad set of proteins that are associated with a specific medical condition. In comparison, multiplex microsphere immunoassays use quantitative measurements of selected set(s) of specific/particular molecular markers such as cytokines, chemokines, pathway signaling or disease-specific markers for detection, metabolic disorders, cancer, and infectious agents causing human, plant and animal diseases. This article provides a foundation to the multiplexed microsphere suspension array technology, with an emphasis on the improvements in the technology, data analysis approaches, and applications to translational and clinical research with implications for personalized and precision medicine.
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http://dx.doi.org/10.1016/j.jim.2018.07.002DOI Listing
October 2018

Western Diet-Induced Dysbiosis in Farnesoid X Receptor Knockout Mice Causes Persistent Hepatic Inflammation after Antibiotic Treatment.

Am J Pathol 2017 Aug 12;187(8):1800-1813. Epub 2017 Jul 12.

Department of Medical Pathology and Laboratory Medicine, University of California, Davis, Sacramento, California. Electronic address:

Patients who have liver cirrhosis and liver cancer also have reduced farnesoid X receptor (FXR). The current study analyzes the effect of diet through microbiota that affect hepatic inflammation in FXR knockout (KO) mice. Wild-type and FXR KO mice were on a control (CD) or Western diet (WD) for 10 months. In addition, both CD- and WD-fed FXR KO male mice, which had hepatic lymphocyte and neutrophil infiltration, were treated by vancomycin, polymyxin B, and Abx (ampicillin, neomycin, metronidazole, and vancomycin). Mice were subjected to morphological analysis as well as gut microbiota and bile acid profiling. Male WD-fed FXR KO mice had the most severe steatohepatitis. FXR KO also had reduced Firmicutes and increased Proteobacteria, which could be reversed by Abx. In addition, Abx eliminated hepatic neutrophils and lymphocytes in CD-fed, but not WD-fed, FXR KO mice. Proteobacteria and Bacteroidetes persisted in WD-fed FXR KO mice even after Abx treatment. Only polymyxin B could reduce hepatic lymphocytes in WD-fed FXR KO mice. The reduced hepatic inflammation by antibiotics was accompanied by decreased free and conjugated secondary bile acids as well as changes in gut microbiota. Our data revealed that Lactococcus, Lactobacillus, and Coprococcus protect the liver from inflammation.
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http://dx.doi.org/10.1016/j.ajpath.2017.04.019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5530909PMC
August 2017

Gender Differences in Bile Acids and Microbiota in Relationship with Gender Dissimilarity in Steatosis Induced by Diet and FXR Inactivation.

Sci Rep 2017 05 11;7(1):1748. Epub 2017 May 11.

Department of Medical Pathology and Laboratory Medicine, University of California, Davis, Sacramento, CA, USA.

This study aims to uncover how specific bacteria and bile acids (BAs) contribute to steatosis induced by diet and farnesoid X receptor (FXR) deficiency in both genders. A control diet (CD) and Western diet (WD), which contains high fat and carbohydrate, were used to feed wild type (WT) and FXR knockout (KO) mice followed by phenotyping characterization as well as BA and microbiota profiling. Our data revealed that male WD-fed FXR KO mice had the most severe steatosis and highest hepatic and serum lipids as well as insulin resistance among the eight studied groups. Gender differences in WD-induced steatosis, insulin sensitivity, and predicted microbiota functions were all FXR-dependent. FXR deficiency enriched Desulfovibrionaceae, Deferribacteraceae, and Helicobacteraceae, which were accompanied by increased hepatic taurine-conjugated cholic acid and β-muricholic acid as well as hepatic and serum lipids. Additionally, distinct microbiota profiles were found in WD-fed WT mice harboring simple steatosis and CD-fed FXR KO mice, in which the steatosis had a potential to develop into liver cancer. Together, the presented data revealed FXR-dependent concomitant relationships between gut microbiota, BAs, and metabolic diseases in both genders. Gender differences in BAs and microbiota may account for gender dissimilarity in metabolism and metabolic diseases.
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http://dx.doi.org/10.1038/s41598-017-01576-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5431816PMC
May 2017

Comprehensive Laboratory Evaluation of a Highly Specific Lateral Flow Assay for the Presumptive Identification of Bacillus anthracis Spores in Suspicious White Powders and Environmental Samples.

Health Secur 2016 Sep-Oct;14(5):351-65

We conducted a comprehensive, multiphase laboratory evaluation of the Anthrax BioThreat Alert(®) test strip, a lateral flow immunoassay (LFA) for the rapid detection of Bacillus anthracis spores. The study, conducted at 2 sites, evaluated this assay for the detection of spores from the Ames and Sterne strains of B. anthracis, as well as those from an additional 22 strains. Phylogenetic near neighbors, environmental background organisms, white powders, and environmental samples were also tested. The Anthrax LFA demonstrated a limit of detection of about 10(6) spores/mL (ca. 1.5 × 10(5) spores/assay). In this study, overall sensitivity of the LFA was 99.3%, and the specificity was 98.6%. The results indicated that the specificity, sensitivity, limit of detection, dynamic range, and repeatability of the assay support its use in the field for the purpose of qualitatively evaluating suspicious white powders and environmental samples for the presumptive presence of B. anthracis spores.
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http://dx.doi.org/10.1089/hs.2016.0041DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5041547PMC
May 2017

Equilibrium Dynamics of β-N-Methylamino-L-Alanine (BMAA) and Its Carbamate Adducts at Physiological Conditions.

PLoS One 2016 11;11(8):e0160491. Epub 2016 Aug 11.

Department of Chemistry, College of Science and Mathematics, California State University, Fresno, California, 93740, United States of America.

Elevated incidences of Amyotrophic Lateral Sclerosis/Parkinsonism Dementia complex (ALS/PDC) is associated with β-methylamino-L-alanine (BMAA), a non-protein amino acid. In particular, the native Chamorro people living in the island of Guam were exposed to BMAA by consuming a diet based on the cycad seeds. Carbamylated forms of BMAA are glutamate analogues. The mechanism of neurotoxicity of the BMAA is not completely understood, and BMAA acting as a glutamate receptor agonist may lead to excitotoxicity that interferes with glutamate transport systems. Though the interaction of BMAA with bicarbonate is known to produce carbamate adducts, here we demonstrate that BMAA and its primary and secondary adducts coexist in solution and undergoes a chemical exchange among them. Furthermore, we determined the rates of formation/cleavage of the carbamate adducts under equilibrium conditions using two-dimensional proton exchange NMR spectroscopy (EXSY). The coexistence of the multiple forms of BMAA at physiological conditions adds to the complexity of the mechanisms by which BMAA functions as a neurotoxin.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0160491PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4981398PMC
July 2017

Upregulation of cystathionine β-synthase and p70S6K/S6 in neonatal hypoxic ischemic brain injury.

Brain Pathol 2017 07 28;27(4):449-458. Epub 2016 Aug 28.

Departments of Pathology and Neurosurgery, Division of Neuropathology, Microvascular and Molecular Neuro-Oncology Laboratory, Perlmutter Cancer Center, New York University Langone Medical Center, New York, NY.

Encephalopathy of prematurity (EOP) is a complex form of cerebral injury that occurs in the setting of hypoxia-ischemia (HI) in premature infants. Using a rat model of EOP, we investigated whether neonatal HI of the brain may alter the expression of cystathionine β-synthase (CBS) and the components of the mammalian target of rapamycin (mTOR) signaling. We performed unilateral carotid ligation and induced HI (UCL/HI) in Long-Evans rats at P6 and found increased CBS expression in white matter (i.e. corpus callosum, cingulum bundle and external capsule) as early as 24 h (P7) postprocedure. CBS remained elevated through P21, and, to a lesser extent, at P40. The mTOR downstream target 70 kDa ribosomal protein S6 kinase (p70S6K and phospho-p70S6K) and 40S ribosomal protein S6 (S6 and phospho-S6) were also overexpressed at the same time points in the UCL/HI rats compared to healthy controls. Overexpression of mTOR components was not observed in rats treated with the mTOR inhibitor everolimus. Behavioral assays performed on young rats (postnatal day 35-37) following UCL/HI at P6 indicated impaired preference for social novelty, a behavior relevant to autism spectrum disorder, and hyperactivity. Everolimus restored behavioral patterns to those observed in healthy controls. A gait analysis has shown that motor deficits in the hind paws of UCL/HI rats were also significantly reduced by everolimus. Our results suggest that neonatal HI brain injury may inflict long-term damage by upregulation of CBS and mTOR signaling. We propose this cascade as a possible new molecular target for EOP-a still untreatable cause of autism, hyperactivity and cerebral palsy.
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http://dx.doi.org/10.1111/bpa.12421DOI Listing
July 2017

Data mining strategies to improve multiplex microbead immunoassay tolerance in a mouse model of infectious diseases.

PLoS One 2015 23;10(1):e0116262. Epub 2015 Jan 23.

Center for Comparative Medicine, University of California Davis, Davis, California, United States of America; Department of Chemistry, California State University, Fresno, California, United States of America; Department of Pathology and Laboratory Medicine, University of California School of Medicine, Davis, California, United States of America.

Multiplex methodologies, especially those with high-throughput capabilities generate large volumes of data. Accumulation of such data (e.g., genomics, proteomics, metabolomics etc.) is fast becoming more common and thus requires the development and implementation of effective data mining strategies designed for biological and clinical applications. Multiplex microbead immunoassay (MMIA), on xMAP or MagPix platform (Luminex), which is amenable to automation, offers a major advantage over conventional methods such as Western blot or ELISA, for increasing the efficiencies in serodiagnosis of infectious diseases. MMIA allows detection of antibodies and/or antigens efficiently for a wide range of infectious agents simultaneously in host blood samples, in one reaction vessel. In the process, MMIA generates large volumes of data. In this report we demonstrate the application of data mining tools on how the inherent large volume data can improve the assay tolerance (measured in terms of sensitivity and specificity) by analysis of experimental data accumulated over a span of two years. The combination of prior knowledge with machine learning tools provides an efficient approach to improve the diagnostic power of the assay in a continuous basis. Furthermore, this study provides an in-depth knowledge base to study pathological trends of infectious agents in mouse colonies on a multivariate scale. Data mining techniques using serodetection of infections in mice, developed in this study, can be used as a general model for more complex applications in epidemiology and clinical translational research.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0116262PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4304816PMC
May 2016

Plasma antibody profiles in non-human primate tuberculosis.

J Med Primatol 2014 Apr 22;43(2):59-71. Epub 2014 Jan 22.

Center for Comparative Medicine, University of California, Davis, CA, USA.

Background: Tuberculosis (TB) in non-human primates (NHPs) is highly contagious, requiring efficient identification of animals infected with Mycobacterium tuberculosis. Tuberculin skin test is usually used but lacks desirable sensitivity/specificity and efficiency.

Methods: We aimed to develop an immunoassay for plasma antibodies against M. tuberculosis. A key challenge is that not all infected animals contain antibodies against the same M. tuberculosis antigen. Therefore, a multiplex panel of 28 antigens (Luminex(®) -Platform) was developed.

Results: Data revealed antibodies against eight antigens (Rv3875, Rv3875-Rv3874 fusion, Rv3874, Rv0934, Rv3881, Rv1886c, Rv2031, Rv3841) in experimentally infected (M. tuberculosis strains: Erdman and H37Rv) NHPs (rhesus and cynomolgus macaques). In a naturally acquired M. tuberculosis infection, rhesus macaques (n = 15) with lung TB pathology (n = 10) contained antibodies to five additional antigens (Rv0831, Rv2220, Rv0054, Rv1099, and Rv0129c).

Conclusions: Results suggest that this user-friendly and easily implementable multiplex panel, containing 13 M. tuberculosis antigens, may provide a high-throughput alternative for NHP TB screening.
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http://dx.doi.org/10.1111/jmp.12097DOI Listing
April 2014

Exploratory study on plasma immunomodulator and antibody profiles in tuberculosis patients.

Clin Vaccine Immunol 2013 Aug 12;20(8):1283-90. Epub 2013 Jun 12.

Center for Comparative Medicine, University of California, Davis, Davis, CA, USA.

Host immune responses to Mycobacterium tuberculosis are generally able to contain infection and maintain a delicate balance between protection and immunopathology. A shift in this balance appears to underlie active disease observed in about 10% of infected individuals. Effects of local inflammation, combined with anti-M. tuberculosis systemic immune responses, are directly detectable in peripheral circulation, without ex vivo stimulation of blood cells or biopsy of the affected organs. We studied plasma immunomodulator and antibody biomarkers in patients with active pulmonary tuberculosis (TB) by a combination of multiplex microbead immunoassays and computational tools for data analysis. Plasma profiles of 10 immunomodulators and antibodies against eight M. tuberculosis antigens (previously reported by us) were examined in active pulmonary TB patients in a country where TB is endemic, Pakistan. Multiplex analyses were performed on samples from apparently healthy individuals without active TB from the same community as the TB patients to establish the assay baselines for all analytes. Over 3,000 data points were collected from patients (n = 135) and controls (n = 37). The data were analyzed by multivariate and computer-assisted cluster analyses to reveal patterns of plasma immunomodulators and antibodies. This study shows plasma profiles that in most patients represented either strong antibody or strong immunomodulator biomarkers. Profiling of a combination of both immunomodulators and antibodies described here may be valuable for the analysis of host immune responses in active TB in countries where the disease is endemic.
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http://dx.doi.org/10.1128/CVI.00213-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3754529PMC
August 2013

Role of receptors of advanced glycation end-products (RAGE) in type 2 diabetic and non-diabetic individuals with chronic periodontal disease: an immunohistochemical study.

J Investig Clin Dent 2011 Nov 5;2(4):287-92. Epub 2011 Aug 5.

Faculty of Dentistry, Annamalai University, Annamalai Nagar, India.

Aim:   The relationship between diabetes and periodontal disease is well established. It has been shown that advanced glycation end-products might exert noxious effects on several tissues of the body through its receptor. Evidence for the role of receptors of advanced glycation end-products in periodontal disease for diabetes is limited, and their presence in human gingival tissues has been demonstrated in few studies. In this study, we demonstrate the presence of receptors of advanced glycation end-products in patients with chronic periodontitis, with and without type 2 diabetes.

Methods:   Gingival biopsies from 19 patients with both type 2 diabetes and chronic periodontitis, and 18 healthy controls with chronic periodontitis, were immunohistochemically stained for receptors of advanced glycation end-products.

Results:   On immunohistochemical analysis, positive staining for receptors of advanced glycation end-products was seen in the endothelium and the basal and spinous layers of the inflamed gingival epithelium in both type 2 diabetes and non-diabetes tissue, with a statistically-significant difference between both groups (P <0 .05).

Conclusions:   There was a significant difference in receptors of advanced glycation end-product immune reactivity between both groups. Receptors of advanced glycation end-product increase in type 2 diabetes gingival tissue might indicate possible involvement of this receptor in periodontal destruction in individuals with type 2 diabetes.
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http://dx.doi.org/10.1111/j.2041-1626.2011.00079.xDOI Listing
November 2011

Plasma antibody profiles as diagnostic biomarkers for tuberculosis.

Clin Vaccine Immunol 2011 Dec 5;18(12):2148-53. Epub 2011 Oct 5.

Center for Comparative Medicine, University of California at Davis, Hutchison Rd. and County Rd. 98, Davis, CA 95616, USA.

Two billion people are infected with Mycobacterium tuberculosis, the etiological agent of tuberculosis (TB), worldwide. Ten million to 20 million of the infected individuals develop disease per year. TB is a treatable disease, provided that it is diagnosed in a timely manner. The current TB diagnostic methods are subjective, inefficient, or not cost-effective. Antibody-based blood tests can be used efficiently and cost-effectively for TB diagnosis. A major challenge is that different TB patients generate antibodies against different antigens. Therefore, a multiplex immunoassay approach is needed. We have developed a multiplex panel of 28 M. tuberculosis antigen-coated microbeads. Plasma samples were obtained from over 300 pulmonary TB patients and healthy controls in a country where TB is endemic, Pakistan. Multiplex data were analyzed using computational tools by multivariate statistics, classification algorithms, and cluster analysis. The results of antibody profile-based detection, using 16 selected antigens, closely correlated with those of the sputum-based diagnostic methods (smear microscopy and culture) practiced in countries where TB is endemic. Multiplex microbead immunoassay had a sensitivity and specificity of approximately 90% and 80%, respectively. These antibody profiles could potentially be useful for the diagnosis of nonpulmonary TB, which accounts for approximately 20% of cases of disease. Since an automated, high-throughput version of this multiplex microbead immunoassay could analyze thousands of samples per day, it may be useful for the diagnosis of TB in millions of patients worldwide.
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http://dx.doi.org/10.1128/CVI.05304-11DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3232686PMC
December 2011

Validation of multiplex microbead immunoassay for simultaneous serodetection of multiple infectious agents in laboratory mouse.

J Immunol Methods 2010 Dec 20;363(1):51-9. Epub 2010 Oct 20.

Center for Comparative Medicine, University of California, Davis, CA 95616, United States.

Multiplex methodologies enable simultaneous detection of antibodies against several infectious agents allowing sample conservation, cost effectiveness, and amenability to high-throughput/automation. We have previously described a multiplex microbead immunoassay for serodetection of ten, high-priority mouse infectious pathogens. Here, we present a validation of this multiplex diagnostic system using approximately four hundred serum samples from different groups of mice. Computer assisted multivariate analysis of the resulting high volume data (8000 data points) was performed. This computational approach enabled presentation of data in a variety of easily interpretable formats (e.g., correlation tables and heat maps). Importantly, this computer aided approach was instrumental for the evaluation of assay accuracy, sensitivity, specificity, and robustness during the study. Crucial pieces of information were obtained to make timely adjustments for assay refinement. This progressive approach to developing an implementation-ready clinical assay, facilitated by computational analysis, produced a highly efficient, accurate and dependable serodiagnostics system. This system has effectively replaced the current state-of-the-art methodology (ELISA) used in mouse colony health management at the University of California and the Jackson Laboratory. A pathway to develop multiplex serology tests for infectious disease diagnosis described here serves as a model for multiplex immunoassay design, clinical validation, refinement and implementation.
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http://dx.doi.org/10.1016/j.jim.2010.10.003DOI Listing
December 2010

A bimodal distribution of two distinct categories of intrinsically disordered structures with separate functions in FG nucleoporins.

Mol Cell Proteomics 2010 Oct 5;9(10):2205-24. Epub 2010 Apr 5.

Department of Molecular, Cell, and Developmental Biology, University of California, Santa Cruz, California 95064, USA.

Nuclear pore complexes (NPCs) gate the only conduits for nucleocytoplasmic transport in eukaryotes. Their gate is formed by nucleoporins containing large intrinsically disordered domains with multiple phenylalanine-glycine repeats (FG domains). In combination, these are hypothesized to form a structurally and chemically homogeneous network of random coils at the NPC center, which sorts macromolecules by size and hydrophobicity. Instead, we found that FG domains are structurally and chemically heterogeneous. They adopt distinct categories of intrinsically disordered structures in non-random distributions. Some adopt globular, collapsed coil configurations and are characterized by a low charge content. Others are highly charged and adopt more dynamic, extended coil conformations. Interestingly, several FG nucleoporins feature both types of structures in a bimodal distribution along their polypeptide chain. This distribution functionally correlates with the attractive or repulsive character of their interactions with collapsed coil FG domains displaying cohesion toward one another and extended coil FG domains displaying repulsion. Topologically, these bipartite FG domains may resemble sticky molten globules connected to the tip of relaxed or extended coils. Within the NPC, the crowding of FG nucleoporins and the segregation of their disordered structures based on their topology, dimensions, and cohesive character could force the FG domains to form a tubular gate structure or transporter at the NPC center featuring two separate zones of traffic with distinct physicochemical properties.
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http://dx.doi.org/10.1074/mcp.M000035-MCP201DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2953916PMC
October 2010

Application of NMR methods to identify detection reagents for use in development of robust nanosensors.

Methods Mol Biol 2005 ;300:141-63

Biology and Biotechnology Research Program, Lawrence Livermore National Laboratory, Livermore, CA, USA.

Nuclear magnetic resonance (NMR) spectroscopy is a powerful technique for studying bimolecular interactions at the atomic scale. Our NMR laboratory is involved in the identification of small molecules, or ligands, that bind to target protein receptors such as tetanus neurotoxin (TeNT) and botulinum neurotoxin, anthrax proteins, and HLA-DR10 receptors on non-Hodgkin lymphoma cancer cells. Once low-affinity binders are identified, they can be linked together to produce multidentate synthetic high-affinity ligands (SHALs) that have very high specificity for their target protein receptors. An important nanotechnology application for SHALs is their use in the development of robust chemical sensors or biochips for the detection of pathogen proteins in environmental samples or body fluids. Here we describe a recently developed NMR competition assay based on transferred nuclear Overhauser effect spectroscopy that enables the identification of sets of ligands that bind to the same site, or a different site, on the surface of TeNT fragment C (TetC) than a known "marker" ligand, doxorubicin. Using this assay, one can identify the optimal pairs of ligands to be linked together for creating detection reagents, as well as estimate the relative binding constants for ligands competing for the same site.
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http://dx.doi.org/10.1385/1-59259-858-7:141DOI Listing
April 2005

Dynamics of antifreeze glycoproteins in the presence of ice.

Biophys J 2002 Jan;82(1 Pt 1):464-73

Section of Molecular and Cellular Biology, University of California, Davis, California 95616, USA.

Antifreeze glycoproteins from the Greenland cod Boreogadus saida were dimethylated at the N-terminus (m*AFGP) and their dynamics and conformational properties were studied in the presence of ice using (13)C-NMR and FTIR spectroscopy. (13)C-NMR experiments of m*AFGP in D(2)O, in H(2)O, and of freeze-dried m*AFGP were performed as a function of temperature. Dynamic parameters ((1)H T(1 rho) and T(CH)) obtained by varying the contact time revealed notable differences in the motional properties of AFGP between the different states. AFGP/ice dynamics was dominated by fast-scale motions (nanosecond to picosecond time scale), suggesting that the relaxation is markedly affected by the protein hydration. The data suggest that AFGP adopts a similar type of three-dimensional fold both in the presence of ice and in the freeze-dried state. FTIR studies of the amide I band did not show a single prevailing secondary structure in the frozen state. The high number of conformers suggests a high flexibility, and possibly reflects the necessity to expose more ice-binding groups. The data suggest that the effect of hydration on the local mobility of AFGP and the lack of significant change in the backbone conformation in the frozen state may play a role in inhibiting the ice crystal growth.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1302486PMC
http://dx.doi.org/10.1016/S0006-3495(02)75411-8DOI Listing
January 2002