Regulatory Non-coding RNAs Network in Non-alcoholic Fatty Liver Disease

Non-alcoholic fatty liver disease (NAFLD) spectrum comprises simple steatosis and non-alcoholic steatohepatitis (NASH) that can lead to fibrosis and cirrhosis. The patients usually have no history of excessive alcohol consumption and other etiologies that can cause fatty liver. Understanding of the pathophysiology of NAFLD has revealed that non-coding RNAs (ncRNAs) play significant roles in modulating the disease susceptibility, pathogenesis and progression. Currently, the ncRNAs are grouped according to their sizes and their regulatory or housekeeping functions. Each of these ncRNAs has a wide range of involvement in the regulation of the genes and biological pathways. Here, we briefly review the current literature the regulatory ncRNAs in NAFLD pathogenesis and progression, mainly the microRNAs, long non-coding RNAs and circular RNAs. We also discuss the co-regulatory functions and interactions between these ncRNAs in modulating the disease pathogenesis. Elucidation of ncRNAs in NAFLD may facilitate the identification of early diagnostic biomarkers and development of therapeutic strategies for NAFLD

MUTATIONAL PROFILES OF F8 AND F9 IN A COHORT OF HAEMOPHILIA A AND HAEMOPHILIA B PATIENTS IN THE MULTI-ETHNIC MALAYSIAN POPULATION

Background: Haemophilia A (HA) and Haemophilia B (HB) are X-linked blood disorders that are caused by various mutations in the factor VIII (F8) and factor IX (F9) genes respectively. Identification of mutations is essential as some of the mutations are associated with the development of inhibitors. This study is the first comprehensive study of the F8 mutational profile in Malaysia. Materials and methods: We analysed 100 unrelated HA and 15 unrelated HB patients for genetic alterations in the F8 and F9 genes by using the long-range PCR, DNA sequencing, and the multiplex-ligation-dependent probe amplification assays. The prediction software was used to confirm the effects of these mutations on factor VIII and IX proteins. Results: 44 (53%) of the severe HA patients were positive for F8 intron 22 inversion, and three (3.6%) were positive for intron 1 inversion. There were 22 novel mutations in F8, including missense (8), frameshift (9), splice site (3), large deletion (1) and nonsense (1) mutations. In HB patients, four novel mutations were identified including the splice site (1), small deletion (1), large deletion (1) and missense (1) mutation. Discussion: The mutational spectrum of F8 in Malaysian patients is heterogeneous, with a slightly higher frequency of intron 22 inversion in these severe HA patients when compared to other Asian populations. Identification of these mutational profiles in F8 and F9 genes among Malaysian patients will provide a useful reference for the early detection and diagnosis of HA and HB in the Malaysian population

Characterisation of Cholinesterase Activity from Asian Swamp Eel; Monopterus albus, and Testing its Capabilities as a Biosensor on Metal Ion Contamination

The presence of high concentrations of heavy metals in the river impedes daily activities and damages the surrounding aquatic ecosystem. Continuous monitoring should be implemented to prevent continuous discharge resulting in increased levels of pollution over time. This study aims to determine the sensitivity of Monopterus albus muscle ChE to metal ions. Priorly, ChE was purified from M. albus muscle tissue using the ion exchange matrix, DEAE, with the yield percentage of 42.16% with the purification fold of 2.17. PTC was selected as a specific synthetic substrate with the highest concentration and lowest biomolecular constant at 145838 ± 7533 μM.min.-1mg.-1 and 0.26 mM, while optimal assay parameters were obtained at pH 7.5 at 20 ° C. Inhibition studies of metal ions involving Cr, Cd, Ag, As, Hg, Pb, Ni and Cu at the concentration of 10 mg/L. The activity of M. albus ChE was 56.61% inhibited by Hg2+ and the highest recorded compared to the other metals ion arranged in declining order; Cr r <Cd = Ni <Cu <Pb <As <Ag. Overall, purified ChE from M. albus muscle has proven its ability to be applied as a bio sensor that can be used for environmental monitoring programs

Autoimmune haemolytic anaemia : a cross sectional study in a Tertiary Haematological Centre

Autoimmune haemolytic anaemia (AIHA) is a group of disorders wherein autoantibody causes decompensated acquired haemolysis. There has been no epidemiological study of autoimmune haemolytic anaemia (AIHA) in Malaysia. This study retrospectively analysed the epidemiology of AIHA including Evan’s Syndrome in a Tertiary Haematology Centre in Malaysia. Patients diagnosed with AIHA and Evan’s Syndrome at 18 years old and above between 1 January 1994 to 1 October 2020 at the out-patient Haematology Clinic of Hospital Raja Permaisuri Bainun, Ipoh were selected. Patients’ information was retrieved from the outpatient clinic records. A total of 71 patients were included of which predominantly female. The mean age for both genders were comparable. Ethnic stratification revealed AIHA was higher in Malays followed by Chinese and Indian. Warm AIHA was most prevalent at 40.8%, compared to cold AIHA and Evan’s Syndrome (both 23.9%), and mixed AIHA (11.3%). Primary was more common than secondary AIHA followed by Evan’s Syndrome. Approximately half of the secondary AIHA and secondary Evan’s Syndrome were due to SLE. Overall, 67.6% of patients received corticosteroid only and 28.2% combined with immunosuppressant. Individuals at higher age and females have higher risk of developing AIHA and Evan’s Syndrome. The highest prevalence was seen among the Malay ethnic. Primary warm AIHA is the most common type and majority of Evan’s syndrome are secondary to autoimmune diseases

Water Warriors Living Lab: Towards an integrated “Heartware - Hardware – Software” Approach to Water Management

This paper presents the experience of the Water Warriors Living Lab, a bottom-up sustainable campus action research initiative in applying the integrated ‘Heartware - Hardware – Software’ approach for water management at the University of Malaya since the year 2013. It advances the proposition that a heartware approach is needed to enhance campus sustainability efforts, in addition to the more strategic hardware and software approaches. The paper describes how the Water Warrior’s experience in applying the heartware approach has been a strengthening factor in resolving water conservation issues at the university (specifically in lake management), and discusses important lessons learned to date. Our theoretical and empirical insights can hopefully provide renewed appreciation on how local, organic and humanistic factors can be essential ingredients for the long term ‘sustainability’ of campus sustainability efforts.Keyword: Campus sustainability, living lab, water management, heartware, bottom-u

Mutations in KIF27, GNAS and IFT140 genes in a patient with VACTERL association: a case report

VACTERL association is a rare genetic disorder involving at least three of the following congenital malformations: vertebral defects (V), anal atresia (A), cardiac defects (C), trachea-oesophageal fistula with or without oesophageal atresia (TE), renal anomalies (R) and limb abnormalities (L). Until now, the aetiology of VACTERL association is unknown, particularly at the molecular level. Here, we performed whole exome sequencing (WES) of an infant with VACTERL association. The patient was delivered prematurely at 30 weeks and had 4/6 of the VACTERL malformations. Trio-WES analysis was performed using Torrent Suite and ANNOVAR. Polymorphisms with an allele frequency of >0.01 were excluded, and the remaining variants were filtered based on de novo mutations, autosomal recessive, X-linked and di-genic inheritance traits. In this patient, no homozygous, compound heterozygous or X-linked mutations was associated with VACTERL. However, we identified two heterozygous mutations; KIF27 (ENST00000297814: c.3004A> C:p.N1002H) and GNAS (ENST00000371098: c.205C>A:p.H69N) genes that were inherited from her father and mother respectively. A de novo, IFT140 gene mutation (ENST00000426508: c.683C>G:p.S228C) was also identified in this patient. The VACTERL phenotype in this patient may due to heterozygous mutations affecting KIF27 and GNAS genes, inherited via autosomal recessive trait. In addition, the IFT140 gene mutation may also be involved. These genes are known to be directly or non-directly involved in the sonic hedgehog signalling that is known to be implicated in VACTERL. This is the first report of these genetic mutations in association with VACTERL

Early life determinants of Beta-cell function in the sheep

Low birth weight or intrauterine growth restriction (IUGR) consistently predict increased risk of Type 2 diabetes (T2D) through impairment of glucose tolerance, insulin resistance and inadequate insulin secretion in humans (1, 2), as well as in many experimental studies in other species (3, 4). IUGR due to insufficient supply of fetal nutrients, decreased oxygen supply and elevated exposure to stress hormones are thought to ‘program’ the impairment of β-cell mass, function and plasticity which then contributes to development of diabetes later in life, as observed in humans (5-7) and animals (4, 8). Interestingly, administration of the glucagon-like-peptide 1 (GLP1) analogue exendin-4 to neonatal IUGR rats normalised subsequent β-cell mass and insulin secretion and prevented later development of T2D (9), thus providing a possible intervention strategy to prevent T2D following IUGR. However, there are differences in the timing of pancreatic and β-cell development between species and therefore in the developmental stages during exposure to IUGR and neonatal interventions. In humans and sheep, most pancreatic and β-cell development occurs before birth (10-17). In contrast, rodents undergo later development of β-cells than sheep or humans, with the majority of pancreatic remodelling occurred at postnatal ages (18-20). It is therefore necessary to test the efficacy of neonatal exendin-4 treatment in animal models such as sheep that share similar profile of pancreatic development and growth with humans (9, 21). Therefore, this thesis will address the effects of IUGR on β-cell mass and function, expression of their molecular determinants, as well as epigenetic modifications, and the possible involvement of altered circulating adiponectin abundance and expression in adipose tissue in the young lamb from birth to 16 d of age. The efficacy of neonatal exendin-4 treatment as a postnatal intervention to prevent these adverse effects of IUGR on metabolic outcomes will also be assessed. Here, natural twin pregnancies were used as a model of IUGR in progeny and unrestricted singleton lambs as the controls. In each twin set, sibling twin lambs with high and low birth weights were alternately allocated to either vehicle or exendin-4 treatment. Effects of IUGR due to twinning and of neonatal exendin-4 treatment of the twin lambs on neonatal growth, pancreatic β-cell in vivo and in vitro insulin secretory function, β-cell mass and islet expression of key regulatory genes including microRNAs, epigenetic regulators, and adiponectin, and on adiponectin abundance were analysed. IUGR due to twinning reduced size at birth and increased neonatal growth, without altering insulin sensitivity, in vivo insulin action, β-cell mass or islet mRNA expression of β-cell mass molecular determinants when compared to CON lambs. However, in vitro glucose-stimulated insulin secretion was increased in the IUGR twin lamb relative to controls (+420%, P = 0.081), consistent with up-regulation of islet mRNA expression of GCK in this group (+80%, P = 0.017), thus suggesting up-regulated β-cell function at this age. Interestingly, IUGR twin lambs also had increased islet mRNA expression of DNMT3B relative to CON lambs (+96%, P = 0.027), which is responsible for de novo DNA methylation (22, 23). Islet mRNA expression of GCK was positively correlated with that of DNMT3B in the IUGR twin group, suggesting that altered islet GCK mRNA expression and β-cell function after IUGR may occur in part via epigenetic changes that may persist throughout life. In conjunction with enhanced β-cell function, up-regulation of adiponectin mRNA expression in omental fat (+72%, P = 0.008) and increased circulating adiponectin levels (P = 0.012) were also observed in the IUGR twin lamb group. Omental adiponectin mRNA expression and circulating adiponectin correlated positively with insulin secretion and β-cell mass in combined control and IUGR twin lamb groups, suggesting that this adipokine may play a role in regulating neonatal insulin secretion. Daily exendin-4 treatment of IUGR twin lambs during neonatal life prevented accelerated neonatal growth or catch up growth (CUG) and fat accumulation (-57%, P < 0.001), and normalised in vitro insulin secretion and GCK and DNMT3B mRNA expression in their islets, relative to vehicle-treated IUGR twins. This may retain adaptive capacity of β-cell function for later life. Glucose tolerance of twin IUGR lambs was impaired during exendin-4 treatment (+156%, P = 0.003) reflecting decreased insulin sensitivity (-46%, P = 0.002) in this group, despite having normal in vivo insulin secretion. This may be due to central actions of exendin-4 to inhibit food intake and insulin sensitivity (24-26). β-cell mass in IUGR twin lambs treated with exendin-4 tended to be higher than in their IUGR counterparts (+28%, P = 0.083), and consistent with this, islet mRNA expression of IGF1 and IGF2R was increased in this group (+62%, P = 0.058 and +63%, P = 0.005 respectively) when compared to controls. Moreover, in the IUGR+Ex-4 lambs, islet mRNA expression of PDX1 correlated positively with that of IGF1R, while IGF1 mRNA expression correlated positively with β-cell volume density, which may suggest hyperplastic effects of the IGF axis on β-cell mass during exendin-4 treatment. Despite the profound reduction in visceral fat mass induced by neonatal exendin-4 treatment, circulating adiponectin concentrations were not reduced in exendin-4-treated lambs, possibly due to upregulation of adiponectin expression in subcutaneous fat in these animals (+91%, P = 0.007). Nevertheless, the reduction in fat accumulation and normalised in vitro β-cell action of IUGR lambs during neonatal exendin-4 treatment suggest that neonatal exendin-4 might have beneficial effects on insulin-regulated glucose homeostasis in later life. These outcomes also demonstrate the biological activity of exendin-4 for the first time in the sheep, at least in the context of individuals who had undergone growth-restriction before birth. In conclusion, IUGR due to twinning induced CUG, early life up-regulation of in vitro β-cell insulin secretion and islet expression of its determinant, GCK, but did not alter in vivo insulin action, glucose tolerance or β-cell mass in young lambs at 16 d of age. These metabolic and molecular changes may be partly mediated by increases in circulating adiponectin and its expression in omental fat, as part of an adipose tissue response during neonatal fat deposition. Consistent with our hypothesis, neonatal exendin-4 treatment prevented this IUGR-induced CUG and decreased visceral fat deposition, increased 2ⁿᵈ phase insulin secretion in vivo, normalised in vitro insulin secretion and islet expression of its determinant, GCK, at the end of treatment in the IUGR twin lambs. Although exendin-4 treatment only tended to increase β-cell mass in young IUGR lamb, the up-regulation of islet expression of β-cell mass determinants after 16 days of exendin-4 treatment may suggest beneficial effects of exendin-4 to subsequently expand β-cell mass. This may protect the exendin-4-treated IUGR individual from a need to increase β-cell function, and preserve the capacity of β-cells for later plasticity of insulin secretion in response to the development of insulin resistance with ageing. Hence, a long term investigation is required to address how these changes following IUGR and neonatal exendin-4 treatment at 16 d of age will affect β-cell function and mass and insulin action and their regulation in the IUGR sheep to adulthood.Thesis (Ph.D.) (Research by Publication) -- University of Adelaide, School of Paediatrics and Reproductive Health, 2015