Publications by authors named "Grant Morahan"

107 Publications

Mapping of Diabetes Susceptibility Loci in a Domestic Cat Breed with an Unusually High Incidence of Diabetes Mellitus.

Genes (Basel) 2020 Nov 19;11(11). Epub 2020 Nov 19.

Centre for Diabetes Research, Harry Perkins Institute for Medical Research, University of Western Australia, Nedlands 6009, Australia.

Genetic variants that are associated with susceptibility to type 2 diabetes (T2D) are important for identification of individuals at risk and can provide insights into the molecular basis of disease. Analysis of T2D in domestic animals provides both the opportunity to improve veterinary management and breeding programs as well as to identify novel T2D risk genes. Australian-bred Burmese (ABB) cats have a 4-fold increased incidence of type 2 diabetes (T2D) compared to Burmese cats bred in the United States. This is likely attributable to a genetic founder effect. We investigated this by performing a genome-wide association scan on ABB cats. Four SNPs were associated with the ABB T2D phenotype with values <0.005. All exons and splice junctions of candidate genes near significant single-nucleotide polymorphisms (SNPs) were sequenced, including the genes and . Six candidate polymorphisms were followed up in a larger cohort of ABB cats with or without T2D and also in Burmese cats bred in America, which exhibit low T2D incidence. The original SNPs were confirmed in this cohort as associated with the T2D phenotype, although no novel coding SNPs in any of the seven candidate genes showed association with T2D. The identification of genetic markers associated with T2D susceptibility in ABB cats will enable preventative health strategies and guide breeding programs to reduce the prevalence of T2D in these cats.
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http://dx.doi.org/10.3390/genes11111369DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7699364PMC
November 2020

Micro-CT scan with virtual dissection of left ventricle is a non-destructive, reproducible alternative to dissection and weighing for left ventricular size.

Sci Rep 2020 08 17;10(1):13853. Epub 2020 Aug 17.

Australian National University Medical School, Canberra, ACT, Australia.

Micro-CT scan images enhanced by iodine staining provide high-resolution visualisation of soft tissues in laboratory mice. We have compared Micro-CT scan-derived left ventricular (LV) mass with dissection and weighing. Ex-vivo micro-CT scan images of the mouse hearts were obtained following staining by iodine. The LV was segmented and its volume was assessed using a semi-automated method by Drishti software. The left ventricle was then dissected in the laboratory and its actual weight was measured and compared against the estimated results. LV mass was calculated multiplying its estimated volume and myocardial specific gravity. Thirty-five iodine-stained post-natal mouse hearts were studied. Mice were of either sex and 68 to 352 days old (median age 202 days with interquartile range 103 to 245 days) at the time of sacrifice. Samples were from 20 genetically diverse strains. Median mouse body weight was 29 g with interquartile range 24 to 34 g. Left Ventricular weights ranged from 40.0 to 116.7 mg. The segmented LV mass estimated from micro-CT scan and directly measured dissected LV mass were strongly correlated (R = 0. 97). Segmented LV mass derived from Micro-CT images was very similar to the physically dissected LV mass (mean difference = 0.09 mg; 95% confidence interval - 3.29 mg to 3.1 mg). Micro-CT scanning provides a non-destructive, efficient and accurate visualisation tool for anatomical analysis of animal heart models of human cardiovascular conditions. Iodine-stained soft tissue imaging empowers researchers to perform qualitative and quantitative assessment of the cardiac structures with preservation of the samples for future histological analysis.
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http://dx.doi.org/10.1038/s41598-020-70734-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7431593PMC
August 2020

Changes in pancreatic exocrine function in young at-risk children followed to islet autoimmunity and type 1 diabetes in the ENDIA study.

Pediatr Diabetes 2020 09 9;21(6):945-949. Epub 2020 Jun 9.

Robinson Research Institute, Adelaide Medical School, University of Adelaide, Adelaide, South Australia, Australia.

Backgrounds: We aimed to monitor pancreatic exocrine function longitudinally in relation to the development of islet autoimmunity (IA) and type 1 diabetes (T1D) in at-risk children with a first-degree relative with T1D, who were followed prospectively in the Environmental Determinants of Islet Autoimmunity (ENDIA) study.

Methods: Fecal elastase-1 (FE-1) concentration was measured longitudinally in 85 ENDIA children from median age 1.0 (IQR 0.7,1.3) year. Twenty-eight of 85 children (progressors) developed persistent islet autoantibodies at median age of 1.5 (IQR 1.1,2.5) years, of whom 11 went on to develop clinical diabetes. The other 57 islet autoantibody-negative children (non-progressors) followed similarly were age and gender-matched with the progressors. An adjusted linear mixed model compared FE-1 concentrations in progressors and non-progressors.

Results: Baseline FE-1 did not differ between progressors and non-progressors, or by HLA DR type or proband status. FE-1 decreased over time in progressors in comparison to non-progressors (Wald statistic 5.46, P = .02); in some progressors the fall in FE-1 preceded the onset of IA.

Conclusions: Pancreatic exocrine function decreases in the majority of young at-risk children who progress to IA and T1D.
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http://dx.doi.org/10.1111/pedi.13056DOI Listing
September 2020

E2f8 and Dlg2 genes have independent effects on impaired insulin secretion associated with hyperglycaemia.

Diabetologia 2020 07 30;63(7):1333-1348. Epub 2020 Apr 30.

Department of Medicine (Austin Health), Austin Hospital, University of Melbourne, Level 7, Lance Townsend Building, Studley Road, Heidelberg, VIC, 3084, Australia.

Aims/hypothesis: Reduced insulin secretion results in hyperglycaemia and diabetes involving a complex aetiology that is yet to be fully elucidated. Genetic susceptibility is a key factor in beta cell dysfunction and hyperglycaemia but the responsible genes have not been defined. The Collaborative Cross (CC) is a recombinant inbred mouse panel with diverse genetic backgrounds allowing the identification of complex trait genes that are relevant to human diseases. The aim of this study was to identify and characterise genes associated with hyperglycaemia.

Methods: Using an unbiased genome-wide association study, we examined random blood glucose and insulin sensitivity in 53 genetically unique mouse strains from the CC population. The influences of hyperglycaemia susceptibility quantitative trait loci (QTLs) were investigated by examining glucose tolerance, insulin secretion, pancreatic histology and gene expression in the susceptible mice. Expression of candidate genes and their association with insulin secretion were examined in human islets. Mechanisms underlying reduced insulin secretion were studied in MIN6 cells using RNA interference.

Results: Wide variations in blood glucose levels and the related metabolic traits (insulin sensitivity and body weight) were observed in the CC population. We showed that elevated blood glucose in the CC strains was not due to insulin resistance nor obesity but resulted from reduced insulin secretion. This insulin secretory defect was demonstrated to be independent of abnormalities in islet morphology, beta cell mass and pancreatic insulin content. Gene mapping identified the E2f8 (p = 2.19 × 10) and Dlg2 loci (p = 3.83 × 10) on chromosome 7 to be significantly associated with hyperglycaemia susceptibility. Fine mapping the implicated regions using congenic mice demonstrated that these two loci have independent effects on insulin secretion in vivo. Significantly, our results revealed that increased E2F8 and DLG2 gene expression are correlated with enhanced insulin secretory function in human islets. Furthermore, loss-of-function studies in MIN6 cells demonstrated that E2f8 is involved in insulin secretion through an ATP-sensitive K channel-dependent pathway, which leads to a 30% reduction in Abcc8 expression. Similarly, knockdown of Dlg2 gene expression resulted in impaired insulin secretion in response to glucose and non-glucose stimuli.

Conclusions/interpretation: Collectively, these findings suggest that E2F transcription factor 8 (E2F8) and discs large homologue 2 (DLG2) regulate insulin secretion. The CC resource enables the identification of E2f8 and Dlg2 as novel genes associated with hyperglycaemia due to reduced insulin secretion in pancreatic beta cells. Taken together, our results provide better understanding of the molecular control of insulin secretion and further support the use of the CC resource to identify novel genes relevant to human diseases.
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http://dx.doi.org/10.1007/s00125-020-05137-0DOI Listing
July 2020

Variants of innate CD8 T cells are associated with Grip2 and Klf15 genes.

Cell Mol Immunol 2020 Sep 9;17(9):1007-1009. Epub 2020 Jan 9.

Department of Anatomy, Pusan National University School of Medicine, Yangsan, 50612, South Korea.

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http://dx.doi.org/10.1038/s41423-019-0357-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7608311PMC
September 2020

Higher frequency of vertebrate-infecting viruses in the gut of infants born to mothers with type 1 diabetes.

Pediatr Diabetes 2020 03 7;21(2):271-279. Epub 2020 Jan 7.

School of Women's and Children's Health, University of New South Wales, Sydney, New South Wales, Australia.

Background: Microbial exposures in utero and early life shape the infant microbiome, which can profoundly impact on health. Compared to the bacterial microbiome, very little is known about the virome. We set out to characterize longitudinal changes in the gut virome of healthy infants born to mothers with or without type 1 diabetes using comprehensive virome capture sequencing.

Methods: Healthy infants were selected from Environmental Determinants of Islet Autoimmunity (ENDIA), a prospective cohort of Australian children with a first-degree relative with type 1 diabetes, followed from pregnancy. Fecal specimens were collected three-monthly in the first year of life.

Results: Among 25 infants (44% born to mothers with type 1 diabetes) at least one virus was detected in 65% (65/100) of samples and 96% (24/25) of infants during the first year of life. In total, 26 genera of viruses were identified and >150 viruses were differentially abundant between the gut of infants with a mother with type 1 diabetes vs without. Positivity for any virus was associated with maternal type 1 diabetes and older infant age. Enterovirus was associated with older infant age and maternal smoking.

Conclusions: We demonstrate a distinct gut virome profile in infants of mothers with type 1 diabetes, which may influence health outcomes later in life. Higher prevalence and greater number of viruses observed compared to previous studies suggests significant underrepresentation in existing virome datasets, arising most likely from less sensitive techniques used in data acquisition.
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http://dx.doi.org/10.1111/pedi.12952DOI Listing
March 2020

The impact of angiogenesis inhibitors on survival of patients with small cell lung cancer.

Cancer Med 2019 10 21;8(13):5930-5938. Epub 2019 Aug 21.

Department of Thoracic Surgery, Nanfang Hospital, Southern Medical University, Guangzhou, P. R. China.

Background: Small cell lung cancer (SCLC) is a highly invasive and lethal neuroendocrine tumor. Antiangiogenic drugs have been reported in the treatment of SCLC. We aimed to provide a comprehensive evaluation of the impact of angiogenic inhibitors on SCLC survival using network meta-analysis.

Methods: The impact of five angiogenesis inhibitors, that is, vandetanib (Van), bevacizumab (Bev), Rh-endostatin (End), sunitinib (Sun), and thalidomide (Tha), on progression-free survival (PFS) and overall survival (OS) was evaluated by conducting a network meta-analysis. RNA sequencing data were downloaded from publicly available databases.

Results: Nine phase II and III randomized controlled trials (RCTs), that involved 1599 participants, that investigated angiogenesis inhibitors in the treatment of SCLC were included in this meta-analysis. Sun and Bev achieved better PFS than Tha (Bev VS. Tha, HR = 0.88, 95% CI: 0.79-0.98, Sun VS. Tha, HR = 0.80, 95% CI: 0.65-1.00). Moreover, Sun and Bev were superior to placebo in terms of PFS (Bev VS. Placebo, HR = 0.89, 95%CI: 0.81-0.97, Sun VS. Placebo, HR = 0.81, 95% CI: 0.66-1.00). Based on this study, we found no significant difference of OS of SCLC. The angiogenesis pathway and expression of target genes were globally deactivated in SCLC tissue.

Conclusion: Results of this network meta-analysis indicate that the PFS outcome of SCLC with Sun or Bev drugs is superior to that of Tha. The improved therapeutic impact of angiogenesis inhibitors on SCLC needs more evidence, such as long-term observation in clinical trials, to be validated.
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http://dx.doi.org/10.1002/cam4.2462DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6792507PMC
October 2019

Genetic characterization of early renal changes in a novel mouse model of diabetic kidney disease.

Kidney Int 2019 10 30;96(4):918-926. Epub 2019 May 30.

Centre for Diabetes Research, Harry Perkins Institute of Medical Research, the University of Western Australia, Perth, Western Australia, Australia. Electronic address:

Genetic factors influence susceptibility to diabetic kidney disease. Here we mapped genes mediating renal hypertrophic changes in response to diabetes. A survey of 15 mouse strains identified variation in diabetic kidney hypertrophy. Strains with greater (FVB/N(FVB)) and lesser (C57BL/6 (B6)) responses were crossed and diabetic F2 progeny were characterized. Kidney weights of diabetic F2 mice were broadly distributed. Quantitative trait locus analyses revealed diabetic mice with kidney weights in the upper quartile shared alleles on chromosomes (chr) 6 and 12; these loci were designated as Diabetic kidney hypertrophy (Dkh)-1 and -2. To confirm these loci, reciprocal congenic mice were generated with defined FVB chromosome segments on the B6 strain background (B6.Dkh1/2f) or vice versa (FVB.Dkh1/2b). Diabetic mice of the B6.Dkh1/2f congenic strain developed diabetic kidney hypertrophy, while the reciprocal FVB.Dkh1/2b congenic strain was protected. The chr6 locus contained the candidate gene; Ark1b3, coding aldose reductase; the FVB allele has a missense mutation in this gene. Microarray analysis identified differentially expressed genes between diabetic B6 and FVB mice. Thus, since the two loci identified by quantitative trait locus mapping are syntenic with regions identified for human diabetic kidney disease, the congenic strains we describe provide a valuable new resource to study diabetic kidney disease and test agents that may prevent it.
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http://dx.doi.org/10.1016/j.kint.2019.04.031DOI Listing
October 2019

Gut microbiome dysbiosis and increased intestinal permeability in children with islet autoimmunity and type 1 diabetes: A prospective cohort study.

Pediatr Diabetes 2019 08 20;20(5):574-583. Epub 2019 May 20.

Department of Diabetes and Endocrinology, Women's and Children's Hospital, North Adelaide, South Australia, Australia.

Aims/hypothesis: To investigate the longitudinal relationship between the gut microbiome, circulating short chain fatty acids (SCFAs) and intestinal permeability in children with islet autoimmunity or type 1 diabetes and controls.

Methods: We analyzed the gut bacterial microbiome, plasma SCFAs, small intestinal permeability and dietary intake in 47 children with islet autoimmunity or recent-onset type 1 diabetes and in 41 unrelated or sibling controls over a median (range) of 13 (2-34) months follow-up.

Results: Children with multiple islet autoantibodies (≥2 IA) or type 1 diabetes had gut microbiome dysbiosis. Anti-inflammatory Prevotella and Butyricimonas genera were less abundant and these changes were not explained by differences in diet. Small intestinal permeability measured by blood lactulose:rhamnose ratio was higher in type 1 diabetes. Children with ≥2 IA who progressed to type 1 diabetes (progressors), compared to those who did not progress, had higher intestinal permeability (mean [SE] difference +5.14 [2.0], 95% confidence interval [CI] 1.21, 9.07, P = .006), lower within-sample (alpha) microbial diversity (31.3 [11.2], 95% CI 9.3, 53.3, P = .005), and lower abundance of SCFA-producing bacteria. Alpha diversity (observed richness) correlated with plasma acetate levels in all groups combined (regression coefficient [SE] 0.57 [0.21], 95% CI 0.15, 0.99 P = .008).

Conclusions/interpretation: Children with ≥2 IA who progress to diabetes, like those with recent-onset diabetes, have gut microbiome dysbiosis associated with increased intestinal permeability. Interventions that expand gut microbial diversity, in particular SCFA-producing bacteria, may have a role to decrease progression to diabetes in children at-risk.
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http://dx.doi.org/10.1111/pedi.12865DOI Listing
August 2019

Advanced Genetic Approaches in Discovery and Characterization of Genes Involved With Osteoporosis in Mouse and Human.

Front Genet 2019 2;10:288. Epub 2019 Apr 2.

School of Biomedical Sciences, The University of Western Australia, Perth, WA, Australia.

Osteoporosis is a complex condition with contributions from, and interactions between, multiple genetic loci and environmental factors. This review summarizes key advances in the application of genetic approaches for the identification of osteoporosis susceptibility genes. Genome-wide linkage analysis (GWLA) is the classical approach for identification of genes that cause monogenic diseases; however, it has shown limited success for complex diseases like osteoporosis. In contrast, genome-wide association studies (GWAS) have successfully identified over 200 osteoporosis susceptibility loci with genome-wide significance, and have provided most of the candidate genes identified to date. Phenome-wide association studies (PheWAS) apply a phenotype-to-genotype approach which can be used to complement GWAS. PheWAS is capable of characterizing the association between osteoporosis and uncommon and rare genetic variants. Another alternative approach, whole genome sequencing (WGS), will enable the discovery of uncommon and rare genetic variants in osteoporosis. Meta-analysis with increasing statistical power can offer greater confidence in gene searching through the analysis of combined results across genetic studies. Recently, new approaches to gene discovery include animal phenotype based models such as the Collaborative Cross and ENU mutagenesis. Site-directed mutagenesis and genome editing tools such as CRISPR/Cas9, TALENs and ZNFs have been used in functional analysis of candidate genes and . These resources are revolutionizing the identification of osteoporosis susceptibility genes through the use of genetically defined inbred mouse libraries, which are screened for bone phenotypes that are then correlated with known genetic variation. Identification of osteoporosis-related susceptibility genes by genetic approaches enables further characterization of gene function in animal models, with the ultimate aim being the identification of novel therapeutic targets for osteoporosis.
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http://dx.doi.org/10.3389/fgene.2019.00288DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6455049PMC
April 2019

The development and application of Laboratory Animal Science in China.

Animal Model Exp Med 2018 Dec 21;1(4):247-249. Epub 2018 Dec 21.

Centre for Diabetes Research Harry Perkins Institute of Medical Research Perth Western Australia Australia.

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http://dx.doi.org/10.1002/ame2.12047DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6388048PMC
December 2018

Variable outcomes of human heart attack recapitulated in genetically diverse mice.

NPJ Regen Med 2019 4;4. Epub 2019 Mar 4.

1Australian Regenerative Medicine Institute, Monash University, Clayton, VIC Australia.

Clinical variation in patient responses to myocardial infarction (MI) has been difficult to model in laboratory animals. To assess the genetic basis of variation in outcomes after heart attack, we characterized responses to acute MI in the Collaborative Cross (CC), a multi-parental panel of genetically diverse mouse strains. Striking differences in post-MI functional, morphological, and myocardial scar features were detected across 32 CC founder and recombinant inbred strains. Transcriptomic analyses revealed a plausible link between increased intrinsic cardiac oxidative phosphorylation levels and MI-induced heart failure. The emergence of significant quantitative trait loci for several post-MI traits indicates that utilizing CC strains is a valid approach for gene network discovery in cardiovascular disease, enabling more accurate clinical risk assessment and prediction.
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http://dx.doi.org/10.1038/s41536-019-0067-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6399323PMC
March 2019

Synthesis of new arylsulfonylspiroimidazolidine-2',4'-diones and study of their effect on stimulation of insulin release from MIN6 cell line, inhibition of human aldose reductase, sorbitol accumulations in various tissues and oxidative stress.

Eur J Med Chem 2019 Apr 15;168:154-175. Epub 2019 Feb 15.

Department of Chemistry, Quaid-i-Azam University, Islamabad, 45320, Pakistan. Electronic address:

A novel class of spiroimidazolidine-2',4'-diones substituted with aryl sulfonyl group at different positions was designed and synthesized. The target compounds were evaluated for their potential to release insulin from MIN6 cell line derived from in-vivo immortalized insulin-secreting pancreatic cells. The MIN6 cells represent an important model of beta cells, which as passage numbers increases, lose the first phase but retain partial second phase glucose stimulated insulin secretion (GSIS), similar to patients in early type 2 diabetes onset. Some of the compounds exhibited high potency. Compound 2d and 3f exhibited excellent insulin release activity from MIN6 cells when compared with standard drug, tolbutamide. Some of these compounds had a potent inhibitory activity for human recombinant aldose reductase (ALR2), an enzyme which converts glucose into sorbitol and plays a key role in development of complications arising from diabetes, such as retinopathy, nephropathy, neuropathy and cataract formation. Against human recombinant ALR2, compounds 2a, 3a-d, and 3f-h displayed effective inhibition activities. The results were augmented by the ability of the compounds to prevent sorbitol accumulation in the isolated rat lenses, sciatic nerves and erythrocytes. Some of the compounds were found to possess excellent dual activity, hence they may be promising candidates to modify and evaluate their dual action, i.e., insulin release to combat diabetes and ALR2 inhibition to prevent/treat diabetic complications. The compounds were also found to possess good antioxidant efficacy. Furthermore, most of the compounds lack toxicity as determined on human embryonic kidney cell lines 293 (HEK293).
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http://dx.doi.org/10.1016/j.ejmech.2019.02.036DOI Listing
April 2019

Distinct Gut Virome Profile of Pregnant Women With Type 1 Diabetes in the ENDIA Study.

Open Forum Infect Dis 2019 Feb 16;6(2):ofz025. Epub 2019 Jan 16.

School of Women's and Children's Health, University of New South Wales, Sydney, Australia.

Background: The importance of gut bacteria in human physiology, immune regulation, and disease pathogenesis is well established. In contrast, the composition and dynamics of the gut virome are largely unknown; particularly lacking are studies in pregnancy. We used comprehensive virome capture sequencing to characterize the gut virome of pregnant women with and without type 1 diabetes (T1D), longitudinally followed in the Environmental Determinants of Islet Autoimmunity study.

Methods: In total, 61 pregnant women (35 with T1D and 26 without) from Australia were examined. Nucleic acid was extracted from serial fecal specimens obtained at prenatal visits, and viral genomes were sequenced by virome capture enrichment. The frequency, richness, and abundance of viruses were compared between women with and without T1D.

Results: Two viruses were more prevalent in pregnant women with T1D: picobirnaviruses (odds ratio [OR], 4.2; 95% confidence interval [CI], 1.0-17.1; = .046) and tobamoviruses (OR, 3.2; 95% CI, 1.1-9.3; = .037). The abundance of 77 viruses significantly differed between the 2 maternal groups (≥2-fold difference; < .02), including 8 types present at a higher abundance in women with T1D.

Conclusions: These findings provide novel insight into the composition of the gut virome during pregnancy and demonstrate a distinct profile of viruses in women with T1D.
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http://dx.doi.org/10.1093/ofid/ofz025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6386807PMC
February 2019

Different genetic mechanisms mediate spontaneous versus UVR-induced malignant melanoma.

Elife 2019 01 25;8. Epub 2019 Jan 25.

Drug Discovery Group, QIMR Berghofer Medical Research Institute, Herston, Australia.

Genetic variation conferring resistance and susceptibility to carcinogen-induced tumorigenesis is frequently studied in mice. We have now turned this idea to melanoma using the collaborative cross (CC), a resource of mouse strains designed to discover genes for complex diseases. We studied melanoma-prone transgenic progeny across seventy CC genetic backgrounds. We mapped a strong quantitative trait locus for rapid onset spontaneous melanoma onset to , a gene involved in detection and repair of DNA damage. In contrast, rapid onset UVR-induced melanoma was linked to the ribosomal subunit gene . Ribosome biogenesis was upregulated in skin shortly after UVR exposure. Mechanistically, variation in the 'usual suspects' by which UVR may exacerbate melanoma, defective DNA repair, melanocyte proliferation, or inflammatory cell infiltration, did not explain melanoma susceptibility or resistance across the CC. Instead, events occurring soon after exposure, such as dysregulation of ribosome function, which alters many aspects of cellular metabolism, may be important.
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http://dx.doi.org/10.7554/eLife.42424DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6428585PMC
January 2019

Common Heritable Immunological Variations Revealed in Genetically Diverse Inbred Mouse Strains of the Collaborative Cross.

J Immunol 2019 02 26;202(3):777-786. Epub 2018 Dec 26.

Department of Immunology-Oncology, Maisonneuve-Rosemont Hospital, Montreal, Quebec H1T 2M4, Canada;

Variations in the proportion and number of specific immune cell types among healthy individuals are influenced by both heritable and nonheritable factors. Mouse models, subjected to fewer nonheritable factors than humans, allow the identification of genetic factors that shape the immune system. We characterized immunological trait variability in the Collaborative Cross (CC), a powerful genetic resource of recombinant inbred mouse strains derived from eight diverse founder strains. Of the 18 immunological traits studied in more than 60 CC strains, eight showed genome-wide significant linkage, revealing new genetic loci linked to specific immune traits. We also found that these traits were highly subject to heritable influences. As for humans, mouse immunological traits varied as a continuum rather than as discrete immunophenotypes. The CC thus represents a useful resource to identify factors that determine immunological variations, as well as defining other immune traits likely to be heritable in humans.
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http://dx.doi.org/10.4049/jimmunol.1801247DOI Listing
February 2019

Genetic variation in the mitogen-activated protein kinase/extracellular signal-regulated kinase pathway affects contact hypersensitivity responses.

J Allergy Clin Immunol 2018 09 12;142(3):981-984.e7. Epub 2018 Jun 12.

UQ Diamantina Institute, Translational Research Institute, University of Queensland, Brisbane, Australia. Electronic address:

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http://dx.doi.org/10.1016/j.jaci.2018.04.028DOI Listing
September 2018

Profiling and genetic control of the murine immunoglobulin G glycome.

Nat Chem Biol 2018 05 9;14(5):516-524. Epub 2018 Apr 9.

Glycoscience Research Laboratory, Genos Ltd., Zagreb, Croatia.

Immunoglobulin G (IgG) glycosylation is essential for function of the immune system, but the genetic and environmental factors that underlie its inter-individual variability are not well defined. The Collaborative Cross (CC) genetic resource harnesses over 90% of the common genetic variation of the mouse. By analyzing the IgG glycome composition of 95 CC strains, we made several important observations: (i) glycome variation between mouse strains was higher than between individual humans, despite all mice having the same environmental influences; (ii) five genetic loci were found to be associated with murine IgG glycosylation; (iii) variants outside traditional glycosylation site motifs affected glycome variation; (iv) bisecting N-acetylglucosamine (GlcNAc) was produced by several strains although most previous studies have reported the absence of glycans containing the bisecting GlcNAc on murine IgGs; and (v) common laboratory mouse strains are not optimal animal models for studying effects of glycosylation on IgG function.
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http://dx.doi.org/10.1038/s41589-018-0034-3DOI Listing
May 2018

Keratinocyte Sonic Hedgehog Upregulation Drives the Development of Giant Congenital Nevi via Paracrine Endothelin-1 Secretion.

J Invest Dermatol 2018 04 11;138(4):893-902. Epub 2017 Nov 11.

Drug Discovery Group, QIMR Berghofer Medical Research Institute, Herston, Queensland, Australia. Electronic address:

Giant congenital nevi are associated with clinical complications such as neurocutaneous melanosis and melanoma. Virtually nothing is known about why some individuals develop these lesions. We previously identified the sonic hedgehog (Shh) pathway regulator Cdon as a candidate nevus modifier gene. Here we validate this by studying Cdon knockout mice, and go on to establishing the mechanism by which Shh exacerbates nevogenesis. Cdon knockout mice develop blue nevi without the need for somatic melanocyte oncogenic mutation. In a mouse model carrying melanocyte NRAS, we found that strain backgrounds that carry genetic variants that cause increased keratinocyte Shh pathway activity, as measured by Gli1 and Gli2 expression, develop giant congenital nevi. Shh components are also active adjacent to human congenital nevi. Mechanistically, this exacerbation of nevogenesis is driven via the release of the melanocyte mitogen endothelin-1 from keratinocytes. We then suppressed nevus development in mice using Shh and endothelin antagonists. Our work suggests an aspect of nevus development whereby keratinocyte cytokines such as endothelin-1 can exacerbate nevogenesis, and provides potential therapeutic approaches for giant congenital nevi. Furthermore, it highlights the notion that germline genetic variation, in addition to somatic melanocyte mutation, can strongly influence the histopathological features of melanocytic nevi.
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http://dx.doi.org/10.1016/j.jid.2017.10.032DOI Listing
April 2018

Influence of Biotechnological Processes, Speed of Formulation Flow and Cellular Concurrent Stream-Integration on Insulin Production from β-cells as a Result of Co-Encapsulation with a Highly Lipophilic Bile Acid.

Cell Mol Bioeng 2018 Feb 3;11(1):65-75. Epub 2017 Oct 3.

Biotechnology and Drug Development Research Laboratory, School of Pharmacy, Curtin Health Innovation Research Institute, Curtin University, Perth, WA Australia.

Introduction: We have shown that incorporation of the hydrophilic bile acid, ursodeoxycholic acid, into β-cell microcapsules exerted positive effects on microcapsules' morphology and size, but these effects were excipient and method dependent. Cell viability remained low which suggests low octane-water solubility and formation of highly hydrophilic dispersion, which resulted in low lipophilicity dispersion and compromised cellular permeation of the incorporated bile acid. Thus, this study aimed at investigating various microencapsulating methodologies using highly lipophilic bile acid (LPBA), in order to optimise viability and functions of microencapsulated β-cells.

Methods: Four different types of microcapsules were produced with (test) and without (control) LPBA, totalling eight different microcapsules. Microencapsulating methodologies were screened for best microcapsule-cell functions and microencapsulating processes were examined in terms of frequency, formulation flow, total bath-gelation time and cellular concurrent stream-integration rate, cell-viability, insulin production and inflammatory profile.

Results: Optimum biotechnological processes include formation frequency (Hz) of 2350, formulation flow (ml/min) of 1.2, total gelation time (min) of 18 and cellular concurrent stream-integration rate (ml/min) of 0.7. In all formulations, LPBA consistently improved cell viability, insulin production, mitochondrial activities and ameliorated inflammation.

Conclusion: The deployed biotechnological processes and LPBA optimised formation and functions of β-cell microcapsules, which suggests potential applications in diabetes mellitus via the creation of more stable β-cell microcapsules capable of delivering adequate levels of insulin to control glycaemia and potentially curing diabetes.
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http://dx.doi.org/10.1007/s12195-017-0510-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6816685PMC
February 2018

Alginate-combined cholic acid increased insulin secretion of microencapsulated mouse cloned pancreatic β cells.

Ther Deliv 2017 Oct;8(10):833-842

Biotechnology & Drug Development Research Laboratory, School of Pharmacy, Curtin Health Innovation Research Institute, Curtin University, Perth, Western Australia, Australia.

Aim: A semisynthetic primary bile acid (PBA) has exerted hypoglycemic effects in Type 1 diabetic animals, which were hypothesized to be due to its anti-inflammatory and cellular glucose-regulatory effects. Thus, the research purpose aimed to examine antidiabetic effects of a PBA, in terms of cellular inflammation and survival and insulin release, in the context of supporting β-cell delivery and Type 1 diabetic treatment.

Materials & Methods: 10 formulations were prepared, five without PBA (control) and five with PBA (test). Formulations were used to microencapsulate pancreatic β cells and the microcapsules were examined for morphology, cell viability, insulin release and inflammation.

Results & Conclusion: PBA improved cell viability, insulin release and reduced inflammation in a formulation-dependent manner, which suggests potential use in cell delivery and diabetes treatment. [Formula: see text].
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http://dx.doi.org/10.4155/tde-2017-0042DOI Listing
October 2017

Variable cardiac α-actin (Actc1) expression in early adult skeletal muscle correlates with promoter methylation.

Biochim Biophys Acta Gene Regul Mech 2017 Oct 26;1860(10):1025-1036. Epub 2017 Aug 26.

Harry Perkins Institute of Medical Research, QEII Medical Centre, Nedlands, Western Australia, Australia; School of Biomedical Sciences, Faculty of Health and Medical Sciences, University of Western Australia, QEII Medical Centre, Nedlands, Western Australia, Australia. Electronic address:

Different genes encode the α-actin isoforms that are predominantly expressed in heart and skeletal muscle. Mutations in the skeletal muscle α-actin gene (ACTA1) cause muscle diseases that are mostly lethal in the early postnatal period. We previously demonstrated that the disease phenotype of ACTA1 mouse models could be rescued by transgenic over-expression of cardiac α-actin (ACTC1). ACTC1 is the predominant striated α-actin isoform in the heart but is also expressed in developing skeletal muscle. To develop a translatable therapy, we investigated the genetic regulation of Actc1 expression. Using strains from The Collaborative Cross (CC) genetic resource, we found that Actc1 varies in expression by up to 24-fold in skeletal muscle. We defined significant expression quantitative trait loci (eQTL) associated with early adult Actc1 expression in soleus and heart. eQTL in both heart and soleus mapped to the Actc1 locus and replicate an eQTL mapped for Actc1 in BXD heart and quadriceps. We built on this previous work by analysing genes within the eQTL peak regions to prioritise likely candidates for modifying Actc1 expression. Additionally we interrogated the CC founder haplotype contributions to enable prioritisation of genetic variants for functional analyses. Methylation around the Actc1 transcriptional start site in early adult skeletal muscle negatively correlated with Actc1 expression in a strain-dependent manner, while other marks of regulatory potential (histone modification and chromatin accessibility) were unaltered. This study provides novel insights into the complex genetic regulation of Actc1 expression in early adult skeletal muscles.
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http://dx.doi.org/10.1016/j.bbagrm.2017.08.004DOI Listing
October 2017

Electrokinetic potential-stabilization by bile acid-microencapsulating formulation of pancreatic β-cells cultured in high ratio poly-L-ornithine-gel hydrogel colloidal dispersion: applications in cell-biomaterials, tissue engineering and biotechnological applications.

Artif Cells Nanomed Biotechnol 2018 Sep 4;46(6):1156-1162. Epub 2017 Aug 4.

a Biotechnology and Drug Development Research Laboratory, School of Pharmacy , Curtin Health Innovation Research Institute, Curtin University , Perth , WA , Australia.

Introduction: Current trials for β-cell transplantation have been hindered by poor cell viability and function post-transplantation. Recently, electric charges of the microencapsulating formulation carrying β-cells have shown significant effects on cell survival and function. Thus, this study aimed at investigating the effects of electric charge, of novel colloidal formulation containing β-cells, on cell viability, biological activity and insulin release.

Methods: A new formulation, containing high ratios of poly-L-ornithine, suspending electrical-stimulation hydrogel and polystyrene sulphone (1:1:0.1 ratio), was used to form microcapsules utilizing 800 V and 2000 Hz encapsulating conditions. The bile acid, ursodeoxycholic acid, was added into the microcapsules to measure its effects on electric charges.

Results: The electric charge of the microencapsulating formulation was enhanced by bile acid addition, and resulted in better cell viability and function.

Conclusion: Ursodeoxycholic acid microencapsulated with poly-L-ornithine, suspending electrical-stimulation hydrogel and polystyrene sulphone at 1:1:0.1 ratio, using 800 V and 2000 Hz microencapsulating conditions, produced enhanced electrokinetic parameters of microcapsules with optimized cell functions. This suggests that electric charge of formulations containing pancreatic β-cell may have significant effects on cell mass and functions, post-transplantation.
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http://dx.doi.org/10.1080/21691401.2017.1362416DOI Listing
September 2018

Gene Expression Networks in the Murine Pulmonary Myocardium Provide Insight into the Pathobiology of Atrial Fibrillation.

G3 (Bethesda) 2017 09 7;7(9):2999-3017. Epub 2017 Sep 7.

Harry Perkins Institute for Medical Research.

The pulmonary myocardium is a muscular coat surrounding the pulmonary and caval veins. Although its definitive physiological function is unknown, it may have a pathological role as the source of ectopic beats initiating atrial fibrillation. How the pulmonary myocardium gains pacemaker function is not clearly defined, although recent evidence indicates that changed transcriptional gene expression networks are at fault. The gene expression profile of this distinct cell type was examined to investigate underlying molecular events that might contribute to atrial fibrillation. Via systems genetics, a whole-lung transcriptome data set from the BXD recombinant inbred mouse resource was analyzed, uncovering a pulmonary cardiomyocyte gene network of 24 transcripts, coordinately regulated by chromosome 1 and 2 loci. Promoter enrichment analysis and interrogation of publicly available ChIP-seq data suggested that transcription of this gene network may be regulated by the concerted activity of NKX2-5, serum response factor, myocyte enhancer factor 2, and also, at a post-transcriptional level, by RNA binding protein motif 20. Gene ontology terms indicate that this gene network overlaps with molecular markers of the stressed heart. Therefore, we propose that perturbed regulation of this gene network might lead to altered calcium handling, myocyte growth, and contractile force contributing to the aberrant electrophysiological properties observed in atrial fibrillation. We reveal novel molecular interactions and pathways representing possible therapeutic targets for atrial fibrillation. In addition, we highlight the utility of recombinant inbred mouse resources in detecting and characterizing gene expression networks of relatively small populations of cells that have a pathological significance.
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http://dx.doi.org/10.1534/g3.117.044651DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5592927PMC
September 2017

New Biotechnological Microencapsulating Methodology Utilizing Individualized Gradient-Screened Jet Laminar Flow Techniques for Pancreatic β-Cell Delivery: Bile Acids Support Cell Energy-Generating Mechanisms.

Mol Pharm 2017 08 17;14(8):2711-2718. Epub 2017 Jul 17.

Biotechnology and Drug Development Research Laboratory, School of Pharmacy, Curtin Health Innovation Research Institute, Curtin University , Perth, Western Australia 6102, Australia.

In previous studies, we developed a new technique (ionic gelation vibrational jet flow; IGVJF) in order to encapsulate pancreatic β-cells, for insulin in vivo delivery, and diabetes treatment. The fabricated microcapsules showed good morphology but limited cell functions. Thus, this study aimed to optimize the IGVJF technique, by utilizing integrated electrode tension, coupled with high internal vibration, jet-flow polymer stream rate, ionic bath-gelation concentrations, and gelation time stay. The study also utilized double inner/outer nozzle segmented-ingredient flow of microencapsulating dispersion, in order to form β-cell microcapsules. Furthermore, a microcapsule-stabilizing bile acid was added, and microcapsule's stability and cell functions measured. Buchi-based built-in system utilizing IGVJF technology was screened to produce best microcapsule-containing β-cells with or without a stabilizing-enhancing bile acid. Formed microcapsules were examined, for physical characteristics, and encapsulated cells were examined for survival, insulin release, and inflammatory profiles. Optimized microencapsulating parameters, using IGJVF, were: 1000 V voltage, 2500 Hz frequency, 1 mL/min flow rate, 3% w/v ionic-bath gelation concentration, and 20 min gelation time. Microcapsules showed good morphology and stability, and the encapsulated cells showed good survival, and insulin secretion, which was optimized by the bile acid. Deployed IGVJF-based microencapsulating parameters utilizing stability-enhancing bile acid produced best microcapsules with best pancreatic β-cells functions and survival rate, which, suggests potential application in cell transplantation.
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http://dx.doi.org/10.1021/acs.molpharmaceut.7b00220DOI Listing
August 2017

Effects of Type 1 Diabetes Risk Alleles on Immune Cell Gene Expression.

Genes (Basel) 2017 Jun 21;8(6). Epub 2017 Jun 21.

Centre for Diabetes Research, Harry Perkins Institute of Medical Research, Nedlands, WA 6009, Australia.

Genetic studies have identified 61 variants associated with the risk of developing Type 1 Diabetes (T1D). The functions of most of the non-HLA (Human Leukocyte Antigen) genetic variants remain unknown. We found that only 16 of these risk variants could potentially be linked to a protein-coding change. Therefore, we investigated whether these variants affected susceptibility by regulating changes in gene expression. To do so, we examined whole transcriptome profiles of 600 samples from the Type 1 Diabetes Genetics Consortium (T1DGC). These comprised four different immune cell types (Epstein-Barr virus (EBV)-transformed B cells, either basal or after stimulation; and cluster of differentiation (CD)4+ and CD8+ T cells). Many of the T1D-associated risk variants regulated expression of either neighboring (-) or distant (-) genes. In brief, 24 of the non-HLA T1D variants affected the expression of 31 nearby genes () while 25 affected 38 distant genes (). The effects were highly significant (False Discovery Rate < 0.001). In addition, we searched in public databases for expression effects of T1D single nucleotide polymorphisms (SNPs) in other immune cell types such as CD14+ monocytes, lipopolysaccharide (LPS) stimulated monocytes, and CD19+ B cells. In this paper, we review the (expression quantitative trait loci (eQTLs) associated with each of the 60 T1D variants and provide a summary of the genes impacted by T1D risk alleles in various immune cells. We then review the methodological steps involved in analyzing the function of genome wide association studies (GWAS)-identified variants, with emphasis on those affecting gene expression. We also discuss recent advancements in the methodologies and their advantages. We conclude by suggesting future study designs that will aid in the study of T1D risk variants.
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http://dx.doi.org/10.3390/genes8060167DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5485531PMC
June 2017

Analysis of early mesothelial cell responses to Staphylococcus epidermidis isolated from patients with peritoneal dialysis-associated peritonitis.

PLoS One 2017 24;12(5):e0178151. Epub 2017 May 24.

Translational Renal Research Group, Harry Perkins Institute of Medical Research, Nedlands, Western Australia, Australia.

The major complication of peritoneal dialysis (PD) is the development of peritonitis, an infection within the abdominal cavity, primarily caused by bacteria. PD peritonitis is associated with significant morbidity, mortality and health care costs. Staphylococcus epidermidis is the most frequently isolated cause of PD-associated peritonitis. Mesothelial cells are integral to the host response to peritonitis, and subsequent clinical outcomes, yet the effects of infection on mesothelial cells are not well characterised. We systematically investigated the early mesothelial cell response to clinical and reference isolates of S. epidermidis using primary mesothelial cells and the mesothelial cell line Met-5A. Using an unbiased whole genome microarray, followed by a targeted panel of genes known to be involved in the human antibacterial response, we identified 38 differentially regulated genes (adj. p-value < 0.05) representing 35 canonical pathways after 1 hour exposure to S. epidermidis. The top 3 canonical pathways were TNFR2 signaling, IL-17A signaling, and TNFR1 signaling (adj. p-values of 0.0012, 0.0012 and 0.0019, respectively). Subsequent qPCR validation confirmed significant differences in gene expression in a number of genes not previously described in mesothelial cell responses to infection, with heterogeneity observed between clinical isolates of S. epidermidis, and between Met-5A and primary mesothelial cells. Heterogeneity between different S. epidermidis isolates suggests that specific virulence factors may play critical roles in influencing outcomes from peritonitis. This study provides new insights into early mesothelial cell responses to infection with S. epidermidis, and confirms the importance of validating findings in primary mesothelial cells.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0178151PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5443531PMC
September 2017

The effect of molecular weights of microencapsulating polymers on viability of mouse-cloned pancreatic β-cells: biomaterials, osmotic forces and potential applications in diabetes treatment.

Pharm Dev Technol 2018 Feb 9;23(2):145-150. Epub 2017 May 9.

a Biotechnology and Drug Development Research Laboratory, School of Pharmacy , Curtin Health Innovation Research Institute, Curtin University , Perth , Western Australia , Australia.

Introduction: Ideal cell-containing microcapsules should be capable of maintaining cell viability and exhibit significant structural stability to support cellular functionality. To date, such microcapsules remain unavailable; thus, this study used our well-established microencapsulating methods to examine a total of 32 different microencapsulating formulations and correlate polymers' molecular weights (M) and UDCA addition, with cell viability and microcapsules' stability, postmicroencapsulation.

Methods: MIN6 mouse-cloned pancreatic β-cells were microencapsulated using control (n = 16; without UDCA) and test (n = 16; with UDCA) different polymers. Confocal microscopic imaging, cell viability, and microcapsules' stability were assessed.

Results: Best cell viability (>50%) was obtained at average M of 50,000 g/mol (poly-l-ornithine), followed by 110,000 g/mol (poly-l-lysine). There was no linear correlation between M and viability. Confocal imagining showed similar microcapsules' shape and cell distribution among all different polymers' molecular weights, which suggests that the microencapsulating method was efficient and maintained microcapsules' uniformity. UDCA addition resulted in enhanced osmotic stability of the microcapsules and improved cell viability, when the formulation contained 1% polylornithine, 1% polyethylene glycol, 20% Eudragit NM30D, 1% polytetrafluoroethylene, or 5% pentamethylcyclopentasiloxane.

Conclusions: UDCA addition improved microenvironmental conditions within the microcapsules but this effect was largely dependent on the polymer systems used.
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http://dx.doi.org/10.1080/10837450.2017.1321664DOI Listing
February 2018