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Oral epigallocatechin gallate reduces intestinal nadolol absorption via modulation of Oatp1a5 and Oct1 transcriptional levels in spontaneously hypertensive rats

Tan HJ, Ling WC, Chua AL, Lee SK

Phytomedicine, 2021 Sep;90:153623.
PMID: 34303263 DOI: 10.1016/j.phymed.2021.153623

BACKGROUND: Concurrent use of epigallocatechin-3-gallate (EGCG) and medication may lead to botanical-drug interactions, subsequently therapeutic failure or drug toxicity. It has been reported that EGCG reduces plasma nadolol bioavailability in normotensive models. Nevertheless, evidence on the effects of EGCG on hypertensive model, and the possible underlying mechanism have not been elucidated.
OBJECTIVES: This study aims (i) to investigate the effects of EGCG on nadolol pharmacokinetics (maximum plasma concentration, time to achieve maximum concentration, area under the time-plasma concentration curve, plasma half-life and total clearance) and subsequently its impact on blood pressure control; and (ii) to identify transcriptional regulatory roles of EGCG on the nadolol intestinal and hepatic drug-transporters in SHR.
METHODS: Male SHR were pre-treated with a daily dose of EGCG (10 mg/kg body weight, i.g.) for 13 days. On day-14, a single dose of nadolol (10 mg/kg body weight) was given to the rats 30 min after the last dose of EGCG administration. Systolic blood pressure (SBP) was measured at 6-h and 22-h post-nadolol administration. Plasma and urinary nadolol concentrations were quantified using high-performance liquid chromatography, and pharmacokinetic parameters were analyzed by using non-compartmental analysis. Hepatic and ileal Oatp1a5, P-gp, and Oct1 mRNA expressions were determined by real-time PCR.
RESULTS: SBP of SHR pre-treated with EGCG and received nadolol was significantly higher than those which were not pre-treated with EGCG but received nadolol. Pre-treatment of EGCG resulted in a marked reduction of plasma nadolol maximum concentration (Cmax) and area under the time-plasma concentration curve (AUC) by 53% and 51% compared to its control. The 14-day treatment with oral EGCG led to a significant downregulation of mRNA levels of ileal Oatp1a5, P-gp, and Oct1 genes by 4.03-, 8.01- and 4.03-fold; and hepatic P-gp, and Oct1 genes by 2.61- and 2.66-fold.
CONCLUSION: These data concluded that exposure to EGCG could lead to reduced nadolol bioavailability and therefore, uncontrolled raised blood pressure and higher risks of cardiovascular events. Our data suggest that the reduced nadolol bioavailability is associated with the downregulation of ileal Oatp1a5 and Oct1 mRNA levels that subsequently lead to poor absorption of nadolol to the systemic circulation.
Development of a dry reagent-based triplex PCR for the detection of toxigenic and non-toxigenic Vibrio cholerae

Chua AL, Elina HT, Lim BH, Yean CY, Ravichandran M, Lalitha P

J Med Microbiol, 2011 Apr;60(Pt 4):481-485.
PMID: 21183596 DOI: 10.1099/jmm.0.027433-0

Vibrio cholerae has caused severe outbreaks of cholera worldwide with thousands of recorded deaths annually. Molecular diagnosis for cholera has become increasingly important for rapid detection of cholera as the conventional methods are time-consuming and labour intensive. However, traditional PCR tests still require cold-chain transportation and storage as well as trained personnel to perform, which makes them user-unfriendly. The aim of this study was to develop a thermostabilized triplex PCR test for cholera which is in a ready-to-use form and requires no cold chain. The PCR test specifically detects both toxigenic and non-toxigenic strains of V. cholerae based on the cholera toxin A (ctxA) and outer-membrane lipoprotein (lolB) genes. The thermostabilized triplex PCR also incorporates an internal amplification control that helps to check for PCR inhibitors in samples. PCR reagents and the specific primers were lyophilized into a pellet form in the presence of trehalose, which acts as an enzyme stabilizer. The triplex PCR was validated with 174 bacteria-spiked stool specimens and was found to be 100 % sensitive and specific. The stability of the thermostabilized PCR was evaluated using the Q10 method and it was found to be stable for approximately 7 months at 24 °C. The limit of detection of the thermostabilized triplex PCR assay was 2×10(4) c.f.u. at the bacterial cell level and 100 pg DNA at the genomic DNA level, comparable to conventional PCR methods. In conclusion, a rapid thermostabilized triplex PCR assay was developed for detecting toxigenic and non-toxigenic V. cholerae which requires minimal pipetting steps and is cold chain-free.
Development of an Optimized Tetra-Amplification Refractory Mutation System PCR for Detection of 12 Pathogenic Familial Hypercholesterolemia Variants in the Asian Population

Rosman N, Nawawi HM, Al-Khateeb A, Chua YA, Chua AL

J Mol Diagn, 2022 02;24(2):120-130.
PMID: 35074074 DOI: 10.1016/j.jmoldx.2021.10.006

Early detection of genetic diseases such as familial hypercholesterolemia (FH), and the confirmation of related pathogenic variants, are crucial in reducing the risk for premature coronary artery disease. Currently, next-generation sequencing is used for detecting FH-related candidate genes but is expensive and time-consuming. There is a lack of kits suitable for the detection of the common FH-related variants in the Asia-Pacific region. Thus, this study addressed that need with the development of an optimized tetra-amplification mutation system (T-ARMS) PCR-based assay for the detection of 12 pathogenic variants of FH in the Asian population. The two important parameters for T-ARMS PCR assay performance-annealing temperature and the ratio of outer/inner primer concentrations-were optimized in this study. The optimal annealing temperature of all 12 T-ARMS PCR reactions was 64.6°C. The ideal ratios of outer/inner primer concentrations with each pathogenic variant were: A1, 1:2; A2, 1:4; L1, 1:10; L2, 1:1; L3, 1:2; L4, 1:8; L5, 1:1; L6, 1:2; L7, 1:8; L8, 1:8; L9, 1:2; and L10, 1:8. The lowest limit of detection using DNA extracted from patients was 0.1 ng. The present article highlights the beneficial findings on T-ARMS PCR as part of the development of a PCR-based detection kit for use in detecting FH in economically developing countries in Asia with a greater prevalence of FH.
Identification of carriers among individuals recruited in the typhoid registry in Malaysia using stool culture, polymerase chain reaction, and dot enzyme immunoassay as detection tools

Chua AL, Aziah I, Balaram P, Bhuvanendran S, Anthony AA, Mohmad SN, et al.

Asia Pac J Public Health, 2015 Mar;27(2):NP2740-8.
PMID: 23000800 DOI: 10.1177/1010539512458521

Chronic carriers of Salmonella Typhi act as reservoirs for the organism and become the agents of typhoid outbreaks in a community. In this study, chronic carriers in Kelantan, Malaysia were first identified using the culture and polymerase chain reaction method. Then, a novel serological tool, designated Typhidot-C, was evaluated in retrospect using the detected individuals as control positives. Chronic carriage positive by the culture and polymerase chain reaction method was recorded at 3.6% (4 out of 110) among individuals who previously had acute typhoid fever and a 9.4% (10 out of 106) carriage rate was observed among food handlers screened during outbreaks. The Typhidot-C assay was able to detect all these positive carriers showing its potential as a viable carrier screening tool and can be used for efficient detection of typhoid carriers in an endemic area. These findings were used to establish the first carrier registry for S Typhi carriers in Malaysia.
Dry-reagent gold nanoparticle-based lateral flow biosensor for the simultaneous detection of Vibrio cholerae serogroups O1 and O139

Yu CY, Ang GY, Chua AL, Tan EH, Lee SY, Falero-Diaz G, et al.

J Microbiol Methods, 2011 Sep;86(3):277-82.
PMID: 21571011 DOI: 10.1016/j.mimet.2011.04.020

Cholera is a communicable disease caused by consumption of contaminated food and water. This potentially fatal intestinal infection is characterised by profuse secretion of rice watery stool that can rapidly lead to severe dehydration and shock, thus requiring treatment to be given immediately. Epidemic and pandemic cholera are exclusively associated with Vibrio cholerae serogroups O1 and O139. In light of the need for rapid diagnosis of cholera and to prevent spread of outbreaks, we have developed and evaluated a direct one-step lateral flow biosensor for the simultaneous detection of both V. cholerae O1 and O139 serogroups using alkaline peptone water culture. Serogroup specific monoclonal antibodies raised against lipopolysaccharides (LPS) were used to functionalize the colloidal gold nanoparticles for dual detection in the biosensor. The assay is based on immunochromatographic principle where antigen-antibody reaction would result in the accumulation of gold nanoparticles and thus, the appearance of a red line on the strip. The dry-reagent dipstick format of the biosensor ensure user-friendly application, rapid result that can be read with the naked eyes and cold-chain free storage that is well-suited to be performed at resource-limited settings.