Nor Azian Abdul Murad

DRD and GRIN2B polymorphisms and their association with the development of impulse control behaviour among Malaysian Parkinson’s disease patients

BackgroundImpulse control disorder (ICD) and behaviours (ICB) represent a group of behavioural disorders that have become increasingly recognised in Parkinson’s disease (PD) patients who previously used dopaminergic medications, particularly dopamine agonists and levodopa. It has been suggested that these medications can lead to the development of ICB through the abnormal modulation of dopaminergic transmission and signalling in the mesocorticolimbic dopaminergic system. Several studies have reported an association between polymorphisms in the dopamine receptor (DRD) and N-methyl-D-aspartate 2B (GRIN2B) genes with the development of ICB in PD (PD-ICB) patients. Thus, this study aimed to investigate the association of selected polymorphisms within the DRD and GRIN2B genes with the development of ICB among PD patients using high resolution melt (HRM) analysis.MethodWe used high resolution melt (HRM) analysis to genotype 11 polymorphisms in 5 DRD genes [DRD1 (rs4532, rs4867798 and rs265981), DRD2 (ANKK1 rs1800497, rs104894220 and rs144999500), DRD3 (rs3732783 and rs6280), DRD4 (rs1800443), and DRD5 (rs144132215)] and 1 polymorphism in GRIN2B (rs7301328) in PD patients with (cases, n = 52) and without (controls, n = 39) ICB. Cases were obtained from two tertiary movement disorder centres [UKMMC (n = 9) and UMMC (n = 43)]. At both centres, the diagnosis of ICB was made using the QUIP questionnaire. Controls were recruited from PD patients who attended UKMMC and were found to be negative for ICB using the QUIP questionnaire.ResultsThe HRM analysis showed that 7 of 11 polymorphisms [DRD1 (rs4532, rs4867798, and rs265981), DRD2 (ANKK1 rs1800497), DRD3 (rs3732783 and rs6280), and GRIN2B (rs7301328)] exhibited a clear distinction between wild-type and variant alleles. Variants of DRD2/ANKK1 rs1800497 (OR = 3.77; 95% CI, 1.38-10.30; p = 0.0044), DRD1 rs4867798 (OR = 24.53; 95% CI, 1.68-357.28; p = 0.0054), DRD1 rs4532 (OR = 21.33; 95% CI, 1.97-230.64; p = 0.0024), and GRIN2B rs7301328 (OR = 25.07; 95% CI, 1.30-483.41; p = 0.0097) were found to be associated with an increased risk of developing ICB among PD patients.ConclusionOur findings suggest that polymorphisms in dopamine [DRD1 (rs4532 and rs4867798) and DRD2/ANKK1 rs1800497] and glutamate (GRIN2B rs7301328) receptor genes confer increased risk of ICB development among PD patients.

MicroRNAs and Target Genes As Biomarkers for the Diagnosis of Early Onset of Parkinson Disease

Among the neurodegenerative disorders, Parkinsons disease (PD) ranks as the second most common disorder with a higher prevalence in individuals aged over 60 years old. Younger individuals may also be affected with PD which is known as early onset PD (EOPD). Despite similarities between the characteristics of EOPD and late onset PD (LODP), EOPD patients experience much longer disease manifestations and poorer quality of life. Although some individuals are more prone to have EOPD due to certain genetic alterations, the molecular mechanisms that differentiate between EOPD and LOPD remains unclear. Recent findings in PD patients revealed that there were differences in the genetic profiles of PD patients compared to healthy controls, as well as between EOPD and LOPD patients. There were variants identified that correlated with the decline of cognitive and motor symptoms as well as non-motor symptoms in PD. There were also specific microRNAs that correlated with PD progression, and since microRNAs have been shown to be involved in the maintenance of neuronal development, mitochondrial dysfunction and oxidative stress, there is a strong possibility that these microRNAs can be potentially used to differentiate between subsets of PD patients. PD is mainly diagnosed at the late stage, when almost majority of the dopaminergic neurons are lost. Therefore, identification of molecular biomarkers for early detection of PD is important. Given that miRNAs are crucial in controlling the gene expression, these regulatory microRNAs and their target genes could be used as biomarkers for early diagnosis of PD. In this article, we discussed the genes involved and their regulatory miRNAs, regarding their roles in PD progression, based on the findings of significantly altered microRNAs in EOPD studies. We also discussed the potential of these miRNAs as molecular biomarkers for early diagnosis.

Quantification of Her-2/Neu Gene in Breast Cancer Patients using Real Time-Polymerase Chain Reaction (Q-PCR) and Correlation with Immunohistochemistry Findings

BACKGROUND HER-2/neu is a proto-oncogene that encodes a transmembrane tyrosine kinase growth factor which is crucial for stimulating growth and cellular motility. Overexpression of HER-2/neu is observed in 10-35% of human breast cancers and is associated with pathogenesis, prognosis as well as response to therapy. Given the imperative role of HER-2/neu overexpression in breast cancer, it is important to determine the magnitude of amplification which may facilitate a better prognosis as well as personalized therapy in affected patients. In this study, we determined HER-2/neu protein expression by immunohistochemistry (IHC) concurrently with HER-2/neu DNA amplification by quantitative real time-polymerase chain reaction (Q-PCR). MATERIALS AND METHODS A total of 53 paired tissue samples from breast cancer patients were frozen-sectioned to characterize the tumour and normal tissues. Only tissues with 80% tumour cells were used in this study. For confirmation, Q-PCR was used to determine the HER-2/neu DNA amplification. RESULTS We found 20/53 (37.7%) of the tumour tissues to be positive for HER-2/neu protein overexpression using IHC. Out of these twenty, only 9/53 (17%) cases were in agreement with the Q-PCR results. The concordance rate between IHC and Q-PCR was 79.3%. Approximately 20.7% of positive IHC cases showed no HER-2/neu gene amplification using Q-PCR. CONCLUSION In conclusion, IHC can be used as an initial screening method for detection of the HER-2/neu protein overexpression. Techniques such as Q-PCR should be employed to verify the IHC results for uncertain cases as well as determination of HER-2/neu gene amplification.

NFIX as a master regulator for lung cancer progression

About 40% of lung cancer cases globally are diagnosed at the advanced stage. Lung cancer has a high mortality and overall survival in stage I disease is only 70%. This study was aimed at finding a candidate of transcription regulator that initiates the mechanism for metastasis by integrating computational and functional studies. The genes involved in lung cancer were retrieved using in silico software. 10 kb promoter sequences upstream were scanned for the master regulator. Transient transfection of shRNA NFIXs were conducted against A549 and NCI-H1299 cell lines. qRT-PCR and functional assays for cell proliferation, migration and invasion were carried out to validate the involvement of NFIX in metastasis. Genome-wide gene expression microarray using a HumanHT-12v4.0 Expression BeadChip Kit was performed to identify differentially expressed genes and construct a new regulatory network. The in silico analysis identified NFIX as a master regulator and is strongly associated with 17 genes involved in the migration and invasion pathways including IL6ST, TIMP1 and ITGB1. Silencing of NFIX showed reduced expression of IL6ST, TIMP1 and ITGB1 as well as the cellular proliferation, migration and invasion processes. The data was integrated with the in silico analyses to find the differentially expressed genes. Microarray analysis showed that 18 genes were expressed differentially in both cell lines after statistical analyses integration between t-test, LIMMA and ANOVA with Benjamini-Hochberg adjustment at p-value < 0.05. A transcriptional regulatory network was created using all 18 genes, the existing regulated genes including the new genes PTCH1, NFAT5 and GGCX that were found highly associated with NFIX, the master regulator of metastasis. This study suggests that NFIX is a promising target for therapeutic intervention that is expected to inhibit metastatic recurrence and improve survival rate.

Apolipoprotein Eε4: A Biomarker for Executive Dysfunction among Parkinson's Disease Patients with Mild Cognitive Impairment

Background: Cognitive impairment is prevalent in Parkinsons disease (PD), affecting 15–20% of patients at diagnosis. α-synuclein expression and genetic polymorphisms of Apolipoprotein E (ApoE) have been associated with the presence of cognitive impairment in PD although data have been inconsistent. Objectives: To determine the prevalence of cognitive impairment in patients with PD using Montreal Cognitive Assessment (MoCA), Comprehensive Trail Making Test (CTMT) and Parkinsons disease-cognitive rating scale (PDCRS), and its association with plasma α-synuclein and ApoE genetic polymorphisms. Methods: This was across-sectional study involving 46 PD patients. Patients were evaluated using Montreal cognitive assessment test (MoCA), and detailed neuropsychological tests. The Parkinsons disease cognitive rating scale (PDCRS) was used for cognitive function and comprehensive trail making test (CTMT) for executive function. Blood was drawn for plasma α-synuclein measurements and ApoE genetic analysis. ApoE polymorphism was detected using MutaGELAPoE from ImmunDiagnostik. Plasma α-synuclein was detected using the ELISA Technique (USCN Life Science Inc.) according to the standard protocol. Results: Based on MoCA, 26 (56.5%) patients had mild cognitive impairment (PD-MCI) and 20 (43.5%) had normal cognition (PD-NC). Based on the PDCRS, 18 (39.1%) had normal cognition (PDCRS-NC), 17 (37%) had mild cognitive impairment (PDCRS-MCI), and 11 (23.9%) had dementia (PDCRS-PDD). In the PDCRS-MCI group, 5 (25%) patients were from PD-NC group and all PDCRS-PDD patients were from PD-MCI group. CTMT scores were significantly different between patients with MCI and normal cognition on MoCA (p = 0.003). Twenty one patients (72.4%) with executive dysfunction were from the PD-MCI group; 17 (77.3%) with severe executive dysfunction and 4 (57.1%) had mild to moderate executive dysfunction. There were no differences in the plasma α-synuclein concentration between the presence or types of cognitive impairment based on MoCA, PDCRS, and CTMT. TheApoEe4 allele carrier frequency was significantly higher in patients with executive dysfunction (p = 0.014). Conclusion: MCI was prevalent in our PD population. PDCRS appeared to be more discriminatory in detecting MCI and PDD than MoCA. Plasma α-synuclein level was not associated with presence nor type of cognitive impairment, but the ApoEe4 allele carrier status was significantly associated with executive dysfunction in PD.

Prospective Advances in Circular RNA Investigation

circRNAs have emerged as one of the key regulators in many cellular mechanisms and pathogenesis of diseases. However, with the limited knowledge and current technologies for circRNA investigations, there are several challenges that need to be addressed for. These include challenges in understanding the regulation of circRNA biogenesis, experimental designs, and sample preparations to characterize the circRNAs in diseases as well as the bioinformatics pipelines and algorithms. In this chapter, we discussed the above challenges and possible strategies to overcome those limitations. We also addressed the differences between the existing applications and technologies to study the circRNAs in diseases. By addressing these challenges, further understanding of circRNAs roles and regulations as well as the discovery of novel circRNAs could be achieved.

Association of job sectors with type 2 diabetes mellitus, hypercholesterolemia and obesity: a cross-sectional study from the Malaysian Cohort (TMC) project

Abstract Background The investigation of risk factors of cardiovascular disease (e.g., major endocrine, nutritional and metabolic diseases) across job sectors is useful for targeted public health intervention. This study examined the occurrence of type 2 diabetes mellitus (T2DM), hypercholesterolemia and obesity in 21 job sectors in the general population. Methods A baseline cross-sectional analysis of the Malaysian Cohort was conducted, which included 105 391 adults. Multiple logistic regression analyses were conducted for these three diseases across 20 job sectors compared with the unemployed/homemaker sector. Results The prevalence of T2DM, hypercholesterolemia and obesity was 16.7%, 38.8% and 33.3%, respectively. The Accommodation & Food Service Activities and Transportation & Storage sectors had significantly higher odds for T2DM (adjusted [adj.] prevalence odds ratio [POR] 1.18, p=0.007 and adj. POR 1.15, p=0.008, respectively). No job sector had significantly higher odds for hypercholesterolemia compared with the unemployed/homemaker sector. Only the Accommodation & Food Service Activities sector had significantly higher odds for obesity (adj. POR 1.17, p≤0.001). Conclusions Many job sectors were significantly associated with lower odds of having these three diseases when compared with the unemployed/homemaker sector. These differing associations between diverse job sectors and these diseases are important for public health intervention initiatives and prioritization.

Renin–Angiotensin–Aldosterone System Gene Polymorphisms and Type 2 Diabetic Nephropathy in Asian Populations: An Updated Meta–analysis.

BACKGROUND The association of polymorphisms in the renin-angiotensin-aldosterone system candidate genes, namely Angiotensin-Converting Enzyme (ACE) Insertion/Deletion (I/D), Angiotensinogen (AGT) M235T and Angiotensin II Receptor Type 1 (AGTR1) A1166C with Diabetic Nephropathy (DN) has been studied for decades. OBJECTIVE This meta-analysis aimed to assess the updated pooled effects of these polymorphisms with DN among Asian populations with type 2 diabetes mellitus. METHODS The PubMed electronic database was searched without duration filter until August 2017 and the reference list of eligible studies was screened. The association of each polymorphism with DN was examined using odds ratio and its 95% confidence interval based on dominant, recessive and allele models. Subgroup analyses were conducted based on region, DN definition and DM duration. RESULTS In the main analysis, the ACE I/D (all models) and AGTR1 A1166C (dominant model) showed a significant association with DN. The main analysis of the AGT M235T polymorphism did not yield significant findings. There were significant subgroup differences and indication of significantly higher odds for DN in terms of DM duration (≥10 years) for ACE I/D (all models), AGT M235T (recessive and allele models) and AGTR1 A1166C (recessive model). Significant subgroup differences were also observed for DN definition (advanced DN group) and region (South Asia) for AGTR1 A1166C (recessive model). CONCLUSION In the Asian populations, ACE I/D and AGTR1 A1166C may contribute to DN susceptibility in patients with T2DM by different genetic models. However, the role of AGT M235T needs to be further evaluated.

Genotype-phenotype correlation among Malaysian patients with osteogenesis imperfecta

BACKGROUND Osteogenesis imperfecta (OI) is a rare genetic bone disease characterized by bone fragility and low bone mass. OI was mainly caused by genetic mutations in collagen genes, COL1A1 and COL1A2. Nevertheless, new genes have been identified to be causally linked to OI. The clinical features between each OI groups share great similarities and it is sometimes difficult for clinicians to diagnose the disease accurately. Here, we identify the genetic mutations of OI patients from Malaysia and correlate the genetic mutations with the clinical features. METHOD Targeted sequencing of fourteen genes panel was performed to identify the mutations in 29 OI patients with type I, III, IV and V disease. The mutations were determined using Ion Torrent Suite software version 5 and variant annotation was conducted using ANNOVAR. The identified mutations were confirmed using Sanger sequencing and in silico analysis was performed to evaluate the effects of the candidate mutations at protein level. RESULTS Majority of patients had mutations in collagen genes, 48% (n = 14) in COL1A1 and 14% (n = 4) in COL1A2. Type I OI was caused by quantitative mutations in COL1A1 whereas most of type III and IV were due to qualitative mutations in both of the collagen genes. Those with quantitative mutations had milder clinical severity compared to qualitative mutations in terms of dentinogenesis imperfecta (DI), bone deformity and the ability to walk with aid. Furthermore, a few patients (28%, n = 8) had mutations in IFITM5, BMP1, P3H1 and SERPINF1. CONCLUSION Majority of our OI patients have mutations in collagen genes, similar to other OI populations worldwide. Genotype-phenotype analysis revealed that qualitative mutations had more severe clinical characteristics compared to quantitative mutations. It is crucial to identify the causative mutations and the clinical severity of OI patients may be predicted based on the types of mutations.

Exome sequencing identifies SLC26A4 , GJB2 , SCARB2 and DUOX2 mutations in 2 siblings with Pendred syndrome in a Malaysian family

BackgroundPendred syndrome (PDS, MIM #274600) is an autosomal recessive disorder characterized by congenital sensorineural hearing loss and goiter. In this study, we describing the possible PDS causal mutations in a Malaysian family with 2 daughters diagnosed with bilateral hearing loss and hypothyroidism.Methods and ResultsWhole exome sequencing was performed on 2 sisters with PDS and their unaffected parents. Our results showed that both sisters inherited monoallelic mutations in the 2 known PDS genes, SLC26A4 (ENST00000265715:c.1343C > T, p.Ser448Leu) and GJB2 (ENST00000382844:c.368C > A, p.Thr123Asn) from their father, as well as another deafness-related gene, SCARB2 (ENST00000264896:c.914C > T, p.Thr305Met) from their mother. We postulated that these three heterozygous mutations in combination may be causative to deafness, and warrants further investigation. Furthermore, we also identified a compound heterozygosity involving the DUOX2 gene (ENST00000603300:c.1588A > T:p.Lys530* and c.3329G > A:p.Arg1110Gln) in both sisters which are inherited from both parents and may be correlated with early onset of goiter. All the candidate mutations were predicted deleterious by in silico tools.ConclusionsIn summary, we proposed that PDS in this family could be a polygenic disorder which possibly arises from a combination of heterozygous mutations in SLC26A4, GJB2 and SCARB2 which associated with deafness, as well as compound heterozygous DUOX2 mutations which associated with thyroid dysfunction.