Papers by Chamindie Punyadeera
Journal of clinical pathology, Jan 23, 2016
Heart failure (HF) affects millions of older individuals in both developed and low/middle-income ... more Heart failure (HF) affects millions of older individuals in both developed and low/middle-income countries. Serum galectin-3 levels have been shown to have prognostic value. However, its use as a diagnostic biomarker has not been explored. The aim was to establish a saliva galectin-3 reference range and to demonstrate the potential diagnostic utility of salivary and serum galectin-3 levels in assessing HF. Blood and saliva samples were collected from age-matched healthy controls (n=51) and patients with HF (n=63). Customised immunoassays were developed to quantify salivary galectin-3 levels. The diagnostic performances of these assays were evaluated by receiver operator characteristic (ROC) curves analysis. The galectin-3 concentrations were significantly elevated in saliva and serum samples of patients with HF compared with controls (p<0.001 and p<0.0001, respectively). Using ROC curve analysis, both serum and salivary galectin-3 gave area under the curve (AUC)=0.86 and AUC=0...
Biomolecules, 2019
Screening for systolic heart failure (SHF) has been problematic. Heart failure management guideli... more Screening for systolic heart failure (SHF) has been problematic. Heart failure management guidelines suggest screening for structural heart disease and SHF prevention strategies should be a top priority. We developed a multi-protein biomarker panel using saliva as a diagnostic medium to discriminate SHF patients and healthy controls. We collected saliva samples from healthy controls (n = 88) and from SHF patients (n = 100). We developed enzyme linked immunosorbent assays to quantify three specific proteins/peptide (Kallikrein-1, Protein S100-A7, and Cathelicidin antimicrobial peptide) in saliva samples. The analytical and clinical performances and predictive value of the proteins were evaluated. The analytical performances of the immunoassays were all within acceptable analytical ranges. The multi-protein panel was able to significantly (p < 0.001) discriminate saliva samples collected from patients with SHF from controls. The multi-protein panel demonstrated good performance wit...
Theranostics, 2018
Saliva as a sample matrix is rapidly gaining interest for disease diagnosis and point-of-care ass... more Saliva as a sample matrix is rapidly gaining interest for disease diagnosis and point-of-care assays because it is easy to collect (non-invasive) and contains many health-related biomarkers. However, saliva poses particular problems relative to more common urine and blood matrices, which includes low analyte concentrations, lack of understanding of biomolecule transportation and inherent viscosity variability in human samples. While several studies have sought to improve assay sensitivity, few have addressed sample viscosity specifically. The goal of this study is to minimize the effect of sample viscosity on paper-based analytical devices (PADs) for the measurement of pH and nitrite in human saliva. PADs were used to measure salivary pH from 5.0 to 10.0 with a universal indicator consisting of chlorophenol red, phenol red and phenolphthalein. Nitrite determination was performed using the Griess reaction. Artificial saliva with viscosity values between 1.54 and 5.10 mPa∙s was tested...
International Journal of Cardiology, 2015
We have previously demonstrated that circulating NT-proBNP is truncated at the N and C termini. A... more We have previously demonstrated that circulating NT-proBNP is truncated at the N and C termini. Aims of this study are three-fold: firstly to determine whether the NT-proBNP levels correlate with NYHA functional classes when measuring with different antibody pairs; secondly to evaluate the diagnostic potential of ProBNP and; thirdly to investigate whether combining NT-proBNP assays with or without ProBNP would lead to better diagnostic accuracies. Plasma samples were collected from healthy controls (n=52) and HF patients (n=46). Customized AlphaLISA® immunoassays were developed and validated to measure the concentrations of proBNP and NT-proBNP (with antibodies targeting 13-45, 13-76, 28-76). The diagnostic performance and predictive value of proBNP and NT-proBNP assays and their combinations were evaluated. Plasma proBNP assay showed acceptable diagnostic performance. NT-proBNP13-76 assay is useful in diagnosing and stratifying HF patients. The diagnostic performance of NT-proBNP13-76 demonstrated improvement over commercial NT-proBNP tests. The combination of NT-proBNP13-76 with NT-proBNP28-76 assays gave the best diagnostic assay performance. Our results demonstrate that while there is major heterogeneity in circulating NT-proBNP, specific epitopes of the peptides are extraordinarily stable, providing ideal targets for clinically useful diagnostic assays. Future new clinical diagnostic clinical trials should include a multimarker approach rather than using a single marker to diagnose HF.
Biomarker insights, 2014
Head and neck cancers (HNCs) represent a significant and ever-growing burden to the modern societ... more Head and neck cancers (HNCs) represent a significant and ever-growing burden to the modern society, mainly due to the lack of early diagnostic methods. A significant number of HNCs is often associated with drinking, smoking, chewing beetle nut, and human papilloma virus (HPV) infections. We have analyzed DNA methylation patterns in tumor and normal tissue samples collected from head and neck squamous cell carcinoma (HNSCC) patients who were smokers. We have identified novel methylation sites in the promoter of the mediator complex subunit 15 (MED15/PCQAP) gene (encoing a co-factor important for regulation of transcription initiation for promoters of many genes), hypermethylated specifically in tumor cells. Two clusters of CpG dinucleotides methylated in tumors, but not in normal tissue from the same patients, were identified. These CpG methylation events in saliva samples were further validated in a separate cohort of HNSCC patients (who developed cancer due to smoking or HPV infect...
Rapid communications in mass spectrometry : RCM, Jan 15, 2014
Diseases including cancer and congenital disorders of glycosylation have been associated with cha... more Diseases including cancer and congenital disorders of glycosylation have been associated with changes in the site-specific extent of protein glycosylation. Saliva can be non-invasively sampled and is rich in glycoproteins, giving it the potential to be a useful biofluid for the discovery and detection of disease biomarkers associated with changes in glycosylation. Saliva was collected from healthy individuals and glycoproteins were enriched using phenylboronic acid based glycoprotein enrichment resin. Proteins were deglycosylated with peptide-N-glycosidase F and digested with AspN or trypsin. Desalted peptides and deglycosylated peptides were separated by reversed-phase liquid chromatography and detected with on-line electrospray ionization quadrupole-time-of-flight mass spectrometry using a 5600 TripleTof instrument. Site-specific glycosylation occupancy was semi-quantitatively determined from the abundance of deglycosylated and nonglycosylated versions of each given peptide. Glyco...
We have previously shown that the concentrations of D-dimer are signi cantly elevated in saliva c... more We have previously shown that the concentrations of D-dimer are signi cantly elevated in saliva compared with plasma. Saliva offers several advantages compared with blood analysis. We hypothesised that human saliva contains plasminogen activator inhibitor-1 (PAI-1) and that the concentrations are not a ected by the time of saliva collection. The aim was to adopt and validate an immunoassay to quantify PAI-1 concentrations in saliva and to determine whether saliva collection time has an in uence in the measurement. Materials and methods: Two saliva samples (morning and afternoon) from the same day were collected from healthy subjects (N = 40) who have had no underlying heart conditions. A customized AlphaLISA® immunoassay (PerkinElmer®, MA, USA) was adopted and used to quantify PAI-1 concentrations. We validated the analytical performance of the customized immunoassay by calculating recovery of known amount of analyte spiked in saliva. Results: The recovery (95.03%), intra-(8.59%) and inter-assay (7.52%) variations were within the acceptable ranges. The median salivary PAI-1 concentrations were 394 pg/mL (interquartile ranges (IQR) 243.4-833.1 pg/mL) in the morning and 376 (129.1-615.4) pg/mL in the afternoon and the plasma concentration was 59,000 (24,000-110,000) pg/mL. Salivary PAI-1 did not correlate with plasma (P = 0.812). Conclusions: The adopted immunoassay produced acceptable assay sensitivity and speci city. The data demonstrated that saliva contains PAI-1 and that the its concentration is not a ected by the time of saliva collection. There is no correlation between salivary and plasma PAI-1 concentrations. Further studies are required to demonstrate the utility of salivary PAI-1 in CVD risk factor studies.
International Journal of Cardiology, 2015
We have previously demonstrated that circulating NT-proBNP is truncated at the N and C termini. A... more We have previously demonstrated that circulating NT-proBNP is truncated at the N and C termini. Aims of this study are three-fold: firstly to determine whether the NT-proBNP levels correlate with NYHA functional classes when measuring with different antibody pairs; secondly to evaluate the diagnostic potential of ProBNP and; thirdly to investigate whether combining NT-proBNP assays with or without ProBNP would lead to better diagnostic accuracies. Plasma samples were collected from healthy controls (n=52) and HF patients (n=46). Customized AlphaLISA® immunoassays were developed and validated to measure the concentrations of proBNP and NT-proBNP (with antibodies targeting 13-45, 13-76, 28-76). The diagnostic performance and predictive value of proBNP and NT-proBNP assays and their combinations were evaluated. Plasma proBNP assay showed acceptable diagnostic performance. NT-proBNP13-76 assay is useful in diagnosing and stratifying HF patients. The diagnostic performance of NT-proBNP13-76 demonstrated improvement over commercial NT-proBNP tests. The combination of NT-proBNP13-76 with NT-proBNP28-76 assays gave the best diagnostic assay performance. Our results demonstrate that while there is major heterogeneity in circulating NT-proBNP, specific epitopes of the peptides are extraordinarily stable, providing ideal targets for clinically useful diagnostic assays. Future new clinical diagnostic clinical trials should include a multimarker approach rather than using a single marker to diagnose HF.
Clinical and Translational Medicine, 2012
Background: Human saliva mirrors the body's health and can be collected non-invasively, does not ... more Background: Human saliva mirrors the body's health and can be collected non-invasively, does not require specialized skills and is suitable for large population based screening programs. The aims were twofold: to evaluate the suitability of commercially available saliva collection devices for quantifying proteins present in saliva and to provide levels for C-reactive protein (CRP), myoglobin, and immunoglobin E (IgE) in saliva of healthy individuals as a baseline for future studies. Methods: Saliva was collected from healthy volunteers (n = 17, ages 18-33 years). The following collection methods were evaluated: drool; Salimetrics® Oral Swab (SOS); Salivette® Cotton and Synthetic (Sarstedt) and Greiner Bio-One Saliva Collection System (GBO SCS®). We used AlphaLISA® assays to measure CRP, IgE and myoglobin levels in human saliva. Results: Significant (p b 0.05) differences in the salivary flow rates were observed based on the method of collection, i.e. salivary flow rates were significantly lower (p b 0.05) in unstimulated saliva (i.e. drool and SOS), when compared with mechanically stimulated methods (p b 0.05) (Salivette® Cotton and Synthetic) and acid stimulated method (p b 0.05) (SCS®). Saliva collected using SOS yielded significantly (p b 0.05) lower concentrations of myoglobin and CRP, whilst, saliva collected using the Salivette® Cotton and Synthetic swab yielded significantly (p b 0.05) lower myoglobin and IgE concentrations respectively. Conclusions: The results demonstrated significantly relevant differences in analyte levels based on the collection method. Significant differences in the salivary flow rates were also observed depending on the saliva collection method. The data provide preliminary baseline values for salivary CRP, myoglobin, and IgE levels in healthy participants and based on the collection method.
Cellular Oncology, 2014
MicroRNAs (miRNAs) are known to play an important role in cancer development by post-transcriptio... more MicroRNAs (miRNAs) are known to play an important role in cancer development by post-transcriptionally affecting the expression of critical genes. The aims of this study were two-fold: (i) to develop a robust method to isolate miRNAs from small volumes of saliva and (ii) to develop a panel of saliva-based diagnostic biomarkers for the detection of head and neck squamous cell carcinoma (HNSCC). Five differentially expressed miRNAs were selected from miScript™ miRNA microarray data generated using saliva from five HNSCC patients and five healthy controls. Their differential expression was subsequently confirmed by RT-qPCR using saliva samples from healthy controls (n = 56) and HNSCC patients (n = 56). These samples were divided into two different cohorts, i.e., a first confirmatory cohort (n = 21) and a second independent validation cohort (n = 35), to narrow down the miRNA diagnostic panel to three miRNAs: miR-9, miR-134 and miR-191. This diagnostic panel was independently validated using HNSCC miRNA expression data from The Cancer Genome Atlas (TCGA), encompassing 334 tumours and 39 adjacent normal tissues. Receiver operating characteristic (ROC) curve analysis was performed to assess the diagnostic capacity of the panel. On average 60 ng/μL miRNA was isolated from 200 μL of saliva. Overall a good correlation was observed between the microarray data and the RT-qPCR data. We found that miR-9 (P &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.0001), miR-134 (P &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.0001) and miR-191 (P &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.001) were differentially expressed between saliva from HNSCC patients and healthy controls, and that these miRNAs provided a good discriminative capacity with area under the curve (AUC) values of 0.85 (P &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.0001), 0.74 (P &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001) and 0.98 (P &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.0001), respectively. In addition, we found that the salivary miRNA data showed a good correlation with the TCGA miRNA data, thereby providing an independent validation. We show that we have developed a reliable method to isolate miRNAs from small volumes of saliva, and that the saliva-derived miRNAs miR-9, miR-134 and miR-191 may serve as novel biomarkers to reliably detect HNSCC.
Nanobiomaterials in Clinical Dentistry, 2013
European journal of endocrinology / European Federation of Endocrine Societies, 2005
It has been suggested that adiponectin regulates plasma free fatty acid (FFA) clearance by stimul... more It has been suggested that adiponectin regulates plasma free fatty acid (FFA) clearance by stimulating FFA uptake and/or oxidation in muscle. We aimed to determine changes in plasma adiponectin concentration and adiponectin receptor 1 and 2 mRNA expression in skeletal muscle during and after prolonged exercise under normal, fasting conditions (high FFA trial; HFA) and following pharmacological inhibition of adipose tissue lipolysis (low FFA trial; LFA). Furthermore, we aimed to detect and locate adiponectin in skeletal muscle tissue. Ten subjects performed two exercise trials (120 min at 50% VO(2max)). Indirect calorimetry was used to determine total fat oxidation rate. Plasma samples were collected at rest, during exercise and during post-exercise recovery to determine adiponectin, FFA and glycerol concentrations. Muscle biopsies were taken to determine adiponectin protein and adiponectin receptor 1 and 2 mRNA expression and to localise intramyocellular adiponectin. Basal plasma ad...
X‐Ray Spectrometry, 2000
Archaeology has been called 'the science of the artefact' and nothing demonstrates this point bet... more Archaeology has been called 'the science of the artefact' and nothing demonstrates this point better than the current interest displayed in provenance studies of archaeological objects. In theory, every vessel carries a chemical compositional pattern or 'fingerprint' identical with the clay from which it was made and this relationship is basic to provenance studies. The reasoning behind provenance or sourcing studies is to probe into the past and attempt to re-create prehistory by obtaining information on exchange and social interaction. This paper discusses the use of XRF spectrometry for the analysis of ancient pottery and ceramics to examine whether it is possible to predict prehistoric cultural exchanges.
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Papers by Chamindie Punyadeera