Research title: Dietary intervention among patients with type 2 diabetes mellitus: An e-approach (2012)
Dr Chai Hwa Chia
Research title: Molecular pathology of systemic lupus erythematosus in Asians
Research title: Candidate early predictors for diabetic nephropathy (2010)
Dr Chung Pooi Yin
Research title: Elucidation of the mechanism of antimicrobial activity of pentacyclic triterpenoids on Staphylococcus aureus (2012)
Research title: Developing quantitative tools for asthma forecast in London using weather and air quality
Dr Lim Fei Tieng
Research title: Mechanism of neurogenesis and neuro-regeneration in the adult teleost brain (2011)
Dr Muhammad Kamruzzaman
Research title: Cognitive determinants of racial prejudice
Dr Phang Yen Li
Research title: Gonadotropin-releasing hormone regulates atp-binding cassette g1 and g4 to transport cholesterol in the brain (2011)
Dr Sandun Dalpatadu
Research title: Role of neurosteroid (allopregnanolone) in the modulation of kisspeptin-gonadotropin releasing hormone pathway and mood disorders (2011)
Author: Dr Amutha Ramadas
Title: Dietary intervention among patients with type 2 diabetes mellitus: an e-approach (2012)
myDIDeA was a 12-months two-armed randomised controlled trial conducted in three tertiary public hospitals in Klang Valley, Malaysia. The primary outcome was the Dietary Knowledge, Attitude and Behaviour (DKAB) score, while the secondary outcomes included measures of food intake, anthropometry measurements, blood pressure and resting heart rate, some blood biomarkers and the Dietary Stages of Change (DSOC) score. The study was designed according to the recommendations of the CONSORT statement for randomised trials of non-pharmacologic treatment. The study commenced in November 2009 after obtaining the ethical approvals from relevant authorities, and the trial has been registered with Clinicaltrials.gov (NCT01246687). After being screened for eligibility, 128 patients with Type 2 Diabetes Mellitus (T2DM) from the outpatient clinics at these hospitals were recruited with informed consent, and then randomised into the e-intervention (n=66) or the control (n=62) group. The e-intervention group received an intensive six-month dietary intervention through the study website, which was developed based on various established guidelines and recommendations but personalised according to the participants’ DSOC, in addition to the usual standard treatment at the outpatient clinics. In contrast, the control group continued their usual standard treatment for patients with T2DM in the hospitals. Data were collected at baseline, six months post-intervention and at 12 months follow-up. A pre-tested and validated questionnaire was used for this purpose. The anthropometry measurements, blood pressure and resting heart rate were measured at data collection, while details on the blood biomarkers were obtained from the hospital medical records. All data were analysed with IBM® PASW® Statistics 17.0. On average, each participant logged into the website once a week and spent 11 minutes at the website per visit. myDIDeA was found to be a successful intervention programme to improve the overall DKAB score, due to the improvement in the knowledge and attitude sub-domains. Additionally, the intervention programme also successfully improved the DSOC score of the intervention group, which suggests participants were also making small but significant progress in changing their dietary behaviour. Some additional improvements in diet quality (reduction in carbohydrate and protein intake), glycaemic control and total cholesterol were also detected. myDIDeA is one of few web-delivered dietary interventions for patients with chronic disease. The reach, flexibility, accessibility and conversion of established guidelines into a more user-friendly format have contributed to the success of this intervention programme. Future related studies are suggested to emphasise on the interactivity, familiarise the participants with the system prior to the intervention, encourage self-monitoring and built the intervention on a strong theoretical background.
Author: Dr Chai Hwa Chia
Title: Molecular pathology of systemic lupus erythematosus in Asians
Systemic lupus erythematosus (SLE) is a prototypic autoimmune disease affecting various parts of the body. Polymorphisms in genes involved in toll-like receptor (TLR)/interferon (IFN) signalling pathways have been reported previously to be associated with SLE in many populations. This study aimed to investigate the role of seven single nucleotide polymorphisms (SNPs) within TNFAIP3 (rs2230936 and rs3757173), STAT4 (rs7574865, rs10168266, and rs7601754), and IRF5 (rs4728142 and rs729302), that are involved in upstream and downstream pathway of type I IFN production, in the Malaysian SLE. Genotyping of 360 Malaysian SLE patients and 430 normal healthy individuals revealed that STAT4 rs7574865 and rs10168266 with their minor T alleles [p=0.001, odds ratio (OR)=1.40 and p=5.75x10-4, OR=1.43, respectively] and TNFAIP3 rs3757173 with its C allele (p=0.017, OR=1.66) were associated with elevated risk of SLE in the Malaysian patients, as well as in the Malays and Chinese. The minor G allele of TNFAIP3 rs2230926 was found to reduce the SLE risk (p=0.021, OR=0.53) in the Malaysian patients, particularly in the Malays. No association was observed for STAT4 rs7601754 and SNPs in IRF5 gene. Besides having haplotype TT (p=1 x 10-4, OR=1.53) and CG (p=0.02, OR=0.76) being significantly associated with SLE susceptibility, STAT4 rs7574865 and rs10168266 also formed the best model for high-risk group in multifactor dimensionality reduction (MDR) test. In conclusion, polymorphisms in STAT4 and TNFAIP3 genes could be potential genetic risk factors for SLE development in the Malaysian individuals. → Read full abstract
Author: Dr Christina Gertrude Yap
Title: Candidate early predictors for diabetic nephropathy (2010)
Since diabetic nephropathy (DN) is the most common cause of end-stage renal disease (ESRD), the search for an early predictor for DN is important because interventions may be tailored to prevent the onset of pathogenesis leading to DN instead of reversing an already altered physiological condition. Microalbuminuria is presently the preferred marker of nephropathy, but by the time the detection of microalbuminuria is initiated, advanced renal structural changes have already occurred as evidenced from renal biopsies. Recent reports have showed that the expression and levels of CYP2E1 protein is elevated in the peripheral blood lymphocytes among patients with prolonged diabetes and in those with chronic renal failure. However, there are no published reports on the correlation between elevated CYP2E1 and progression of nephropathy. Therefore, the aims of this study were to evaluate lymphocyte CYP2E1 as a candidate early predictor for diabetic nephropathy using high performance liquid chromatography (HPLC), validate the HPLC observations using real-time PCR, to construct a candidate DN gene signature using the microarray technology, as well as to determine other biomarkers besides CYP2E1 which may be suitable candidate early predictors for DN. A cross-sectional cohort study was carried out among Malaysians, consisting of control (n=28), diabetes (n=50), diabetic nephropathy (n=34) and non diabetic nephropathy (n=15) cohorts. Whole blood samples were collected, lymphocytes were isolated and aliquots were made. Microsomes were prepared for CYP2E1 analysis using HPLC. Total RNA was extracted from the lymphocytes for real time PCR and microarray studies. Microarray analysis was done using Illumina Human Ref-8 Sentrix bead chips and the raw data were analyzed using GeneSpring GX 11.0 software. A gene expression signature consisting of 59 differentially expressed genes (P<0.001) has been constructed and validated using real time PCR. Statistical significance tests were done by employing the one-way ANOVA test at P value cut-off of 0.001 using the Sigma Plot software, version 11.0. The prototype candidate gene expression pattern displayed properties of possible early predictors for DN. Genes involved in glucose metabolism and associated pathways (PGK1, ENO1, TpI1, ANGPTL4, AKR1B1, and SORD) were good observational entities for early prediction of DN. Products of these candidate signature genes may be easily measured in clinical laboratories from plasma samples. The results also showed lymphocyte CYP2E1 was detected in 38.7% of the diabetes cohort population (n=50) with normal serum creatinine, urine microalbumin, glomerular filtration rate and HbA1c levels. Lymphocyte CYP2E1 was not detectable in the control and pre-diabetic cohorts. In conclusion, this study lends early evidence that lymphocyte CYP2E1 is a more specific candidate predictor for diabetic nephropathy compared to microalbuminuria and a prototype candidate molecular signature may be tested as a possible early predictor for DN.
Author: Dr Chung Pooi Yin
Title: Elucidation of the mechanism of antimicrobial activity of pentacyclic triterpenoids on Staphylococcus aureus (2012)
Staphylococcus aureus strains with multiple resistances to antibiotics are increasingly widespread in hospitals and community settings, thus the development of new agents are urgently necessary. The availability of complete whole-genome nucleotide sequence data of various strains of S. aureus, presents an opportunity to explore novel compounds and their targets to address the challenges presented by antimicrobial drug resistance in this organism. Study compounds α-amyrin [3β-hydroxy-urs-12-en-3-ol (AM)], betulinic acid [3β-hydroxy-20(29)-lupaene-28-oic acid (BA)] and betulinaldehyde [3β-hydroxy-20(29)-lupen-28-al (BE)] belong to pentacyclic triterpenoids that are based on a 30-carbon skeleton comprising five six-membered rings or four six-membered rings and one five-membered rings. These compounds were reported to exhibit antimicrobial activities against bacteria and fungi, including S. aureus. The MIC values of these compounds against reference strains of methicillin-resistant S. aureus (MRSA ATCC 43300) and methicillin-sensitive S. aureus (MSSA ATCC 29213) ranged from 64 to 512 μg/ml while most combinations of the compounds and combinations with standard antibiotics methicillin and vancomycin showed synergistic effects. However, the response mechanisms of S. aureus to these compounds are still poorly understood. The genome-wide transcription of reference strains MRSA and MSSA treated with sub-inhibitory concentrations of the three compounds was determined using Affymetrix GeneChips. Treatment of the MRSA strain with AM resulted in the down-regulation of genes involved in the peptidoglycan biosynthesis, aminoacyl tRNA synthase and fatty acid biosynthesis pathways. When treated with BA, there was up-regulation of genes involved in β-lactam resistance, ABC transporters and DNA replication. Down-regulation of genes involved in three main pathways; i.e. fatty acid biosynthesis, peptidoglycan biosynthesis and ribosome were observed when the reference strain of MRSA was treated with BE. In addition, two genes in the ABC transporter pathway were at least 3-fold up-regulated. Similar to the effect of BE against MRSA, AM inhibits MSSA primarily via the ribosomes. The gene expression pattern of MSSA after treatment with BA exhibited minimal changes, while the effect of BE on MSSA is also primarily on the ribosome. In general, there were positive correlation between microarray data and real-time PCR data for all the genes validated. The findings showed that AM, BA and BE affected novel and previously targeted pathways which could be further explored in the development of therapeutic compounds for the treatment of S. aureus infections.
Author: Dr Ireneous Ngmenlanaa Soyiri
Research title: Developing quantitative tools for asthma forecast in London using weather and air quality
The thesis examines approaches to the forecasting of respiratory events, generally hospital admissions for asthma, but also mortality. The focus of the thesis is forecasting accuracy rather than model specification per se. The thesis is a compilation of eight papers (seven published, one "under review"), broken in to four sections, with a brief narrative drawing the themes together. The topic is introduced with a review of asthma - the main condition examined in the thesis – and the known relationships between environmental conditions and asthma events. An empirical study is then presented that examines the factors affecting length of stay (LOS) in a hospital following an asthma admission. The paper relies on National Health Service (NHS), England data for London from 2001 to 2006. The idea was to demonstrate a burden of disease, as measured in this case by LOS, as a motivation for forecasting asthma events. If there is no consequence for the health system of asthma events, then there may be no point proceeding to the forecasting. Negative binomial regression was used to model the effect(s) of demographic, temporal and diagnostic factors on the LOS, taking into account the cluster effect of each patient's hospital attendance in London. The median and mean asthma LOS over the period of study were 2 and 3 days respectively. Admissions increased over the years from 8,308 (2001) to 10,554 (2006), but LOS consistently declined within the same period. Younger individuals were more likely to be admitted than the elderly, but the latter significantly had higher LOS (p<0.001). Respiratory related secondary diagnoses, age, and gender of the patient as well as day of the week and year of admission were important predictors of LOS. Having established the burden of asthma on the health system, health forecasting as an approach is introduced in a series of three closely related, published papers. In the first paper a general overview of health forecasting is provided (Soyiri and Reidpath, 2012a). In the second paper, there is a greater emphasis on the specific modelling approaches used in forecasting, and the measures of forecasting accuracy (Soyiri and Reidpath, 2012b). The final published paper in this series introduces in a general sense a "semi structured black-box approach" to forecasting. Two modelling techniques are described Negative Binomial Models for modelling the conditional mean, and Quantile Regression Models for modelling more extreme quantiles; and these are illustrated using London data from 2005-2006 (Soyiri and Reidpath, 2012c). In the next section of the thesis, four empirical studies are presented, each looking at an approach to health forecasting in greater detail. → Read full abstract
Author: Dr Lim Fei Tieng
Title: Mechanism of neurogenesis and neuro-regeneration in the adult teleost brain (2011)
Fish retain a remarkable potential of neuro-regeneration throughout life, whereas injury to neuronal system in the mammalian central nervous system (CNS) results in degeneration and loss of function. Hypothetically, the brain of teleost will regenerate after injury, however, the molecular mechanism underlying this phenomena as well as the reason for the lack of such regenerative capability in the mammalian CNS is still unknown. To address this issue, three main study objectives were placed, (i) to understand the mechanism involved in neuro-regeneration in teleosts brain (Chapter 2); (ii) to identify novel molecular factors involved in neuro-regeneration in the zebrafish brain (Chapter 3); and (iii) to investigate the potential of identified novel molecular factors in associating adult neurogenesis in the zebrafish brain (Chapter 4). In chapter 2, the study aimed to characterize the neuro-regenerative process by observing changes in apoptotic and proliferative activities in the brain of teleost, zebrafish (Danio rerio) and tilapia (Oreochromis niloticus). Morphological observations showed complete neuro-regeneration of the habenula region by 40 days and 60 days post-damage for zebrafish and tilapia respectively. Recovery of neuronal projections and axonal re-integration further proved complete neuro-regeneration in adult teleost. In chapter 3, the study aimed to identify the molecular factor(s) that play role in the early stage of regeneration, before first batch of newborn cells were significantly detected at the injury site, protein profiling through two-dimensional difference gel electrophoresis was done in the following chapter 3. Ten proteins were identified, which included cytoskeleton protein, transcription factors and binding proteins. Among these proteins identified, sprouty-related EVH1 domain-containing protein 2 (Spred-2) was chosen for further study as there is no study between Spred-2 and neurogenesis has been reported. Finally in chapter 4, the expression of spred-2 mRNA in the adult zebrafish brain was analyzed by in situ hybridization and their histological changes were examined during the brain injury. The zebrafish spred-2 mRNA containing cells were observed in most of the cell proliferative zones, which suggests the potential role of Spred-2 in the regulation of neurogenesis. On the other hand, histological study showed a decrease of spred-2 expression with a concurrent increase in apoptotic cells at the lesion site on the first two days after the injury. Damaged and dead neural cells release growth factors and neurotrophins which can down-regulate Spred-2 expression to unlock the activation of the ERK pathway. Proliferative and phosphorylated-ERK-immunoreactive cells significantly increased on day-4 post-lesion and co-expression of spred-2 mRNA in phosphorylated-ERK-immunoreactive cells were observed in neurogenic zones near the lesion site. These observations suggest that a decrease in spred-2 after injury activates the ERK pathway to stimulate cell proliferation in the adult zebrafish brain. Collectively, these studies showed the teleost CNS possesses excellent neuro-regenerative capability. The apoptotic neural cells could be the earliest source to induce cell proliferation. Differential protein profiling revealed that Spred-2 could be a novel molecule which can regulate neurogenesis in adult teleost CNS. High conservation of Spred-2 protein sequence in vertebrates suggests the potential role for Spred-2 in neurogenesis-regulation in vertebrates including mammals. Taken together, these results provide better cellular and molecular processes underlying adult neuro-regeneration of injured CNS.
Author: Dr Muhammad Kamruzzaman
Title: Cognitive determinants of racial prejudice
Two studies, first qualitative and then quantitative, were conducted to investigate cognitive correlates of racial prejudice in Chittagong Hill Tracts (CHT) in Bangladesh - home to the decades-long interracial conflicts between indigenous Chakmas and settler Bengalis. The studies were guided by a theoretical framework comprised of cognitive behavioral perspective, indigenous psychology and grounded theory methodology. The first study employed a grounded theory approach to examine cognitive factors, especially thinking patterns and perceptions, likely to be associated with racial prejudice. In-depth interviews (IDI) were conducted with 26 respondents (12 Chakmas, 14 Bengalis), of which 16 had high and 10 had low level of prejudice. Participants were recruited from two districts of CHT through a theoretical sampling strategy. Four key-informants were also interviewed to triangulate the IDI findings. The interview data, analyzed using the qualitative software NVivo, revealed 31 types of race-related thoughts and perceptions of which 24 were associated with racial prejudice (e.g., dehumanization, disapproving contact, apprehension of negative, and victim thinking). The qualitative findings were used to generate empirical hypotheses that were tested in the quantitative study. Thirty-one cognitive factors derived from the qualitative study were reduced to 28 by a number of mergers and divisions. Three contact-related and one emotional factor were later added, making a total of 32 constructs. Thirty-three brief instruments, specific to the CHT, were developed to assess racial prejudice and all those constructs. All instruments demonstrated adequate face validity and internal consistency reliability (Cronbach’s alpha = .503 - .919; inter-item correlation r = .353 - .633) except for three (i.e., perception that opposite race is ethnocentric, apprehension of negative, and anchoring). Particularly notable was the 12-item racial prejudice scale that had high concurrent validity (r = -.791 with feeling thermometer), internal consistency (Cronbach’s alpha .916; corrected item-total r = .471 - .789), and test-retest reliability (r = .979, two weeks gap). The quantitative study was conducted on 393 respondents, conveniently recruited from a number of suburban and rural locations of the Khagrachari district in CHT, almost equally represented by the two races (50.6% Chakmas, 49.4% Bengalis). With an age range of 18-87 years (average 37), the participants were mostly male (68.2%). Stepwise multiple linear regression revealed ten significant predictors explaining 86% of the variance in racial prejudice scores (F11,381 = 203.86, p < .01). Contact disapproval appeared to be the strongest predictor followed by dehumanization, progressive orientation, perspective taking, infrahumanization, overgeneralization, maximization-minimization, emotion towards other race, rumor susceptibility, and perceiving administration as biased. Of these ten factors, three (progressive orientation, rumor susceptibility, and perceiving administration as biased) were found to be quite novel as they were never studied before. Contrary to our general expectation, contact factors (direct-, extended-, and negative contact) failed to predict racial prejudice in the CHT context. This research provides an in-depth examination of race-related attitudes and thoughts within the context of CHT. The four-tiered indigenization model used here should be considered as a methodological approach for future research, as should the large set of contextualized instruments. The results suggest practical implications for prejudice reduction strategies appropriate to the CHT.
Author: Dr Phang Yen Li
Title: Gonadotropin-releasing hormone regulates atp-binding cassette g1 and g4 to transport cholesterol in the brain (2011)
The ATP-binding cassette (ABC) transporter family G1 and G4 play an important role in the transport of lipids, especially cholesterol across cell membranes in the brain. Gonadotropin-releasing hormone (GnRH), which is widely known to be the driving force in reproduction, has been implied to play other roles apart from its direct involvement in the reproductive cascade. One of these roles is in the regulation of metabolism, where a link between GnRH and cholesterol has long been implied but was never fully investigated. Advance teleost fishes such as tilapia (Oreochromis niloticus) in particular, express all three known subtypes of GnRH, namely GnRH1, GnRH2 and GnRH3. Moreover, the three types of receptors (GnRHR1, GnRHR2 and GnRHR3) for these GnRHs are also widely distributed in the brain. Thus, the studies documented in this thesis aimed to investigate the role of GnRH in regulating cholesterol homeostasis in the brain, with particular attention on ABCG1 and ABCG4 transporters. Two novel genes (ABCG1 and ABCG4) from the brain tissue of the tilapia fish were cloned and were used in the subsequent investigations of the role of GnRH in the control of cholesterol transport. Both ABCG1 and ABCG4 mRNAs were found to be highly expressed in the central nervous system, pituitary and gonads of tilapia. Additionally, ABCG1 mRNA was found to be highly expressed in the pituitary while ABCG4 mRNA displayed wide distribution pattern of expression across different brain regions. Both ABCG1 and ABCG4 mRNAs were also detected in all three types of GnRH neurons in the tilapia. Three forms of GnRH were applied exogenously to tilapia brain cell cultures and the effect of these GnRHs in mediating the expression of ABCG1 and ABCG4 were recorded. Both ABCG1 and ABCG4 mRNA expressions were affected (differentially up-regulated or down-regulated depending on brain region investigated) in the cells from the olfactory bulb, forebrain and pituitary. The effect of cholesterol on ABCG1 and ABCG4 mRNA expressions were also investigated through loading of exogenous cholesterol into brain cell cultures. Only ABCG1 expression was observed to up-regulated in the hindbrain region. Taken together, the results from this thesis show that ABCG1 transporter gene is regulated directly by changes in cellular cholesterol level in the brain while ABCG4 transporter gene is modulated directly by GnRH1 (in the OB) and GnRH2 (in the pituitary).
Author: Dr Sandun Dalpatadu
Title: Role of neurosteroid (allopregnanolone) in the modulation of kisspeptin-gonadotropin releasing hormone pathway and mood disorders (2011)
Kisspeptin, a neuropeptide encoded by the Kiss1 gene, is a regulator of reproduction. Neurosteroids play an important role in reproduction. As a gatekeeper of reproductive activity, kisspeptin may be regulated by neurosteroids. Neurosteroid allopregnanolone (3α-hydroxy-5α-pregnan-20-one, THP) has been well characterized for its role in anxiety-like behaviour. However, the mechanisms through which THP regulate reproduction and reproductive behaviour needs further investigation. On the other hand, stress during young life is a critical factor that affects reproduction, which causes various reproductive dysfunctions. Whether neonatal stress affects THP mediated reproductive regulations remains unknown. Chapter 2 of this thesis hypothesised that THP regulates kisspeptin, the upstream regulator of gonadotropin-releasing hormone neurons in young adult (postnatal day [PN] 45-55) female mice. Indeed, THP regulated kisspeptin mRNA expression in the anteroventral periventricular area (AVPV) and not in the arcuate (Arc) nucleus. THP action target; GABA(A) receptor subunit expression in kisspeptin neurons in the AVPV was down-regulated by THP. The down-regulation of GABA(A) expression enhanced kisspeptin expression in the AVPV. Chapter 3 focuses on the effect of neonatal stress on reproductive maturation. Synthetic glucocorticoid (dexamethasone, DEX) exposure during neonatal stage reduced kisspeptin mRNA expression in the AVPV and the Arc nucleus, causing delayed reproductive maturation and irregular oestrous cycles. More salient finding of this study was, kisspeptin neurons did not express glucocorticoid receptor but corticotrophin releasing hormone (CRH) receptor type-1 and CRH neuronal projections were seen in close proximity to kisspeptin neurons. Therefore, kisspeptin neurons receive stress signals from CRH neurons. Data in Chapter 2 showed GABA(A) receptors regulate Kiss1 mRNA. GABA(A) subunit expression in kisspeptin neurons increased in the AVPV of neonatal DEX treated mice (Chapter 2). Increased GABA(A) function could have decreased kisspeptin function in the AVPV and caused reproductive irregularities. In chapter 4, the regulation of sexual discrimination behaviour by THP was investigated. Since, THP levels and function depends on the reproductive stage of rodents, two age groups were selected, young adult (PN 45-55) and adult (PN 77-84). Because neonatal DEX cause deregulation of reproductive behaviour at the adult stage, effect of neonatal DEX on THP mediated sexual discrimination was investigated. Interestingly, neither DEX nor THP treatment had any effect on sexual discrimination behaviour of mice. Therefore, regulation of sexual discrimination could occur independently of THP mediated regulation of kisspeptin function. Collectively the data in this thesis suggest that THP plays an important role in the regulation of reproduction at a molecular level through modulations of kisspeptin neurons in the AVPV. Neonatal exposure to glucocorticoids affects reproductive regulation, which shows the long-term effects of neonatal stress on reproductive physiology. However, neither DEX nor THP plays a significant role in regulating sexual discrimination behaviour in female mice. Understanding the regulation of kisspeptin by THP and neonatal stress provide insight into novel pathways regulating kisspeptin neurons. These results enhance our overall understanding of kisspeptin physiology in mice.