Paediatrics (RCH) - Research Publications
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ItemNeonatal DNA methylation profile in human twins is specified by a complex interplay between intrauterine environmental and genetic factors, subject to tissue-specific influence(COLD SPRING HARBOR LAB PRESS, PUBLICATIONS DEPT, 2012-08)Comparison between groups of monozygotic (MZ) and dizygotic (DZ) twins enables an estimation of the relative contribution of genetic and shared and nonshared environmental factors to phenotypic variability. Using DNA methylation profiling of ∼20,000 CpG sites as a phenotype, we have examined discordance levels in three neonatal tissues from 22 MZ and 12 DZ twin pairs. MZ twins exhibit a wide range of within-pair differences at birth, but show discordance levels generally lower than DZ pairs. Within-pair methylation discordance was lowest in CpG islands in all twins and increased as a function of distance from islands. Variance component decomposition analysis of DNA methylation in MZ and DZ pairs revealed a low mean heritability across all tissues, although a wide range of heritabilities was detected for specific genomic CpG sites. The largest component of variation was attributed to the combined effects of nonshared intrauterine environment and stochastic factors. Regression analysis of methylation on birth weight revealed a general association between methylation of genes involved in metabolism and biosynthesis, providing further support for epigenetic change in the previously described link between low birth weight and increasing risk for cardiovascular, metabolic, and other complex diseases. Finally, comparison of our data with that of several older twins revealed little evidence for genome-wide epigenetic drift with increasing age. This is the first study to analyze DNA methylation on a genome scale in twins at birth, further highlighting the importance of the intrauterine environment on shaping the neonatal epigenome.
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ItemNew forearm elements discovered of holotype specimen Australovenator wintonensis from Winton, Queensland, Australia.(Public Library of Science (PLoS), 2012)New skeletal elements are reported of the holotype specimen Australovenator wintonensis, from the type locality, near Winton, central western Queensland. New elements include left and right humeri, right radius, right radiale, right distal carpal 1, near complete right metacarpal I, left manual phalanx II-1, left manual phalanx II-2, near complete left manual phalanx II-3 and a left manual phalanx III-3. These new elements combined with those previously described are compared against other neovenatorids.
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ItemVariable lengths of stay among ischemic stroke subtypes in Chinese general teaching hospitals.(Public Library of Science (PLoS), 2012)BACKGROUND: Length of stay (LOS) is one of the most important quantitative indexes that measures health service utilization within a hospital. Many studies have examined the association of three major stroke categories with LOS. Our aim is to investigate the differences of LOS among ischemic stroke subtypes, results from which are helpful to healthcare providers and government agencies to improve health care delivery efficiency. METHODOLOGY/PRINCIPAL FINDINGS: Using the Beijing Municipal Health Bureau's hospitalization summary reports, we performed a retrospective study among first-ever in-hospital patients with ischemic stroke (ICD-10 I63) in three general teaching hospitals in Beijing, China, from 2006 to 2010 with generalized linear model. In our study, 5,559 patients (female, 36.0%; age, 64.4 ± 12.9 years) were included. The estimated mean LOS of ischemic stroke was 17.4 ± 1.8 days. After adjusting for confounders, LOS of lacunar infarction (14.7 days; p<0.001) and LOS of small cerebral infarction (17.0 days; p=0.393) were shorter than that of single cerebral infarction (17.9 days, p<0.001). LOS of multi-infarct (19.0 days; p=0.028), brainstem infarction (19.3 days; p=0.045), basal ganglia infarction (18.5 days; p=0.452) and other subtypes of ischemic stroke (18.9 days; p=0.327) were longer than that of single cerebral infarction. CONCLUSIONS: LOS of ischemic stroke patient differes across single cerebral infarction, lacunar infarction, multi-infarct and brainstem infarction patients. The ascending order of LOS was lacunar infarction, small cerebral infarction, single cerebral infarction, basal ganglia infarction, other subtypes of ischemic stroke, multi-infarct and brainstem infarction.
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Item[Effects of tea polyphenols on the expression of NF-κB, COX-2 and survivin in Lewis lung carcinoma xenografts in C57BL/6 mice].( 2012-05)BACKGROUND AND OBJECTIVE: Lung cancer is the leading cause of cancer-related deaths worldwide. The anti-angiogenic effect of tea polyphenols (TPs) on lung carcinoma was confirmed in our previous study. The effect of TPs on the expression of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), cyclooxygenase-2 (COX-2), and Survivin in Lewis lung cancer xenograft in mice was observed to explore the anti-angiogenic mechanism of TPs. METHODS: Lewis lung carcinoma was successfully implanted in 32 C57BL/6 mice, which were then randomly divided into the model control group, the Thalidomide group, the TPs group and the TPs combined with Thalidomide group. The tumor inhibition rates in these groups were determined, whereas the expressions of NF-κB, COX-2 and Survivin were detected using immunohistochemical techniques to investigate the molecular mechanism of the anti-tumor effect of TPs. RESULTS: (1) The tumor inhibition rates in the Thalidomide control group, the TPs group and the TPs plus Thalidomide group were 17.26%, 20.81%, 44.32% respectively. Compared with the model control group, the tumor inhibition rate was significantly higher in the TPs plus Thalidomide group (P<0.05); (2) Compared with the model control group, the expression of NF-κB was significantly decreased in the TPs group and the TPs plus Thalidomide group, and it was significantly downregulated in the latter (P<0.05); (3) The expression of COX-2 decreased in all treatment groups, and was significantly downregulated in the TPs plus Thalidomide group compared with the model control group (P<0.05); (4) Compared with the model control group, Survivin expression was significantly downregulated in all treatment groups (P<0.05), and was most obviously reduced in the TPs plus the Thalidomide group (P<0.01). CONCLUSION: TPs plus Thalidomide significantly inhibits Lewis lung cancer growth. TPs possibly downregulates the expression of NF-κB, COX-2 and Survivin in tumor tissue, thereby playing important roles in anti-angiogenesis.
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ItemCongenital Hydrocephalus and Abnormal Subcommissural Organ Development in Sox3 Transgenic Mice(PUBLIC LIBRARY SCIENCE, 2012-01-26)Congenital hydrocephalus (CH) is a life-threatening medical condition in which excessive accumulation of CSF leads to ventricular expansion and increased intracranial pressure. Stenosis (blockage) of the Sylvian aqueduct (Aq; the narrow passageway that connects the third and fourth ventricles) is a common form of CH in humans, although the genetic basis of this condition is unknown. Mouse models of CH indicate that Aq stenosis is associated with abnormal development of the subcommmissural organ (SCO) a small secretory organ located at the dorsal midline of the caudal diencephalon. Glycoproteins secreted by the SCO generate Reissner's fibre (RF), a thread-like structure that descends into the Aq and is thought to maintain its patency. However, despite the importance of SCO function in CSF homeostasis, the genetic program that controls SCO development is poorly understood. Here, we show that the X-linked transcription factor SOX3 is expressed in the murine SCO throughout its development and in the mature organ. Importantly, overexpression of Sox3 in the dorsal diencephalic midline of transgenic mice induces CH via a dose-dependent mechanism. Histological, gene expression and cellular proliferation studies indicate that Sox3 overexpression disrupts the development of the SCO primordium through inhibition of diencephalic roof plate identity without inducing programmed cell death. This study provides further evidence that SCO function is essential for the prevention of hydrocephalus and indicates that overexpression of Sox3 in the dorsal midline alters progenitor cell differentiation in a dose-dependent manner.
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ItemClinical findings and treatment in cattle with caecal dilatation.(Springer Science and Business Media LLC, 2012-07-09)BACKGROUND: This retrospective study describes the clinical and laboratory findings, treatment and outcome of 461 cattle with caecal dilatation. RESULTS: The general condition and demeanor were abnormal in 93.1% of cases, and 32.1% of the patients had colic. Ruminal motility was reduced or absent in 78.3% of cattle. In 82.6% of cases, swinging and/or percussion auscultation were positive on the right side, and 82.4% had little or no faeces in the rectum. Caecal dilatation could be diagnosed via rectal palpation in 405 (88.0%) cattle. There was caudal displacement of the dilated caecum in 291 patients, torsion around the longitudinal axis in 20 and retroflexion in 94. The most important laboratory finding was hypocalcaemia, which occurred in 85.1% of cases. Of the 461 cattle, 122 (26.5%) initially received conservative therapy (intravenous fluids, neostigmine, calcium borogluconate) and 329 (71.4%) underwent surgical treatment. Ten patients were slaughtered or euthanased after the initial physical examination. Of the 122 cattle that received conservative treatment, 42 did not respond after one to two days of therapy and required surgical treatment. The final number of cattle that were operated was 371 (80.5%). Because of a grave prognosis, 24 cases were euthanased or slaughtered intraoperatively. Another 24 cattle did not respond to one or more operations and were euthanased or slaughtered. Of the 461 patients, 403 (87.4%) responded to either conservative or surgical treatment and were cured, and 58 were euthanased or slaughtered. CONCLUSIONS: Caecal dilatation can usually be diagnosed based on clinical findings and treated conservatively or surgically. Swinging and percussion auscultation as well as rectal examination are important diagnostic tools. Conservative treatment is not rewarding in cattle considered surgical candidates with suspected caecal torsion or retroflexion and surgery should not be delayed in these patients.
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ItemClinical Features and Correlates of Outcomes for High-Risk, Marginalized Mothers and Newborn Infants Engaged with a Specialist Perinatal and Family Drug Health Service(HINDAWI LTD, 2012)Background. There is a paucity of research in Australia on the characteristics of women in treatment for illicit substance use in pregnancy and the health outcomes of their neonates. Aims. To determine the clinical features and outcomes of high-risk, marginalized women seeking treatment for illicit substance use in pregnancy and their neonates. Methods. 139 women with a history of substance abuse/dependence engaged with a perinatal drug health service in Sydney, Australia. Maternal (demographic, drug use, psychological, physical, obstetric, and antenatal care) and neonatal characteristics (delivery, early health outcomes) were examined. Results. Compared to national figures, pregnant women attending a specialist perinatal and family drug health service were more likely to report being Australian born, Aboriginal or Torres Strait Islander, younger, unemployed, and multiparous. Opiates were the primary drug of concern (81.3%). Pregnancy complications were common (61.9%). Neonates were more likely to be preterm, have low birth weight, and be admitted to special care nursery. NAS was the most prevalent birth complication (69.8%) and almost half required pharmacotherapy. Conclusion. Mother-infant dyads affected by substance use in pregnancy are at significant risk. There is a need to review clinical models of care and examine the longer-term impacts on infant development.
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ItemRegulation of immune responses by histone deacetylase inhibitors.(Hindawi Limited, 2012)Both genetic and epigenetic factors are important regulators of the immune system. There is an increasing body of evidence attesting to epigenetic modifications that influence the development of distinct innate and adaptive immune response cells. Chromatin remodelling via acetylation, methylation, phosphorylation, and ubiquitination of histone proteins as well as DNA, methylation is epigenetic mechanisms by which immune gene expression can be controlled. In this paper, we will discuss the role of epigenetics in the regulation of host immunity, with particular emphasis on histone deacetylase inhibitors. In particular, the role of HDAC inhibitors as a new class of immunomodulatory therapeutics will also be reviewed.
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ItemBone to pick: the importance of evaluating reference genes for RT-qPCR quantification of gene expression in craniosynostosis and bone-related tissues and cells.(Springer Science and Business Media LLC, 2012-05-08)BACKGROUND: RT-qPCR is a common tool for quantification of gene expression, but its accuracy is dependent on the choice and stability (steady state expression levels) of the reference gene/s used for normalization. To date, in the bone field, there have been few studies to determine the most stable reference genes and, usually, RT-qPCR data is normalised to non-validated reference genes, most commonly GAPDH, ACTB and 18 S rRNA. Here we draw attention to the potential deleterious impact of using classical reference genes to normalise expression data for bone studies without prior validation of their stability. RESULTS: Using the geNorm and Normfinder programs, panels of mouse and human genes were assessed for their stability under three different experimental conditions: 1) disease progression of Crouzon syndrome (craniosynostosis) in a mouse model, 2) proliferative culture of cranial suture cells isolated from craniosynostosis patients and 3) osteogenesis of a mouse bone marrow stromal cell line. We demonstrate that classical reference genes are not always the most 'stable' genes and that gene 'stability' is highly dependent on experimental conditions. Selected stable genes, individually or in combination, were then used to normalise osteocalcin and alkaline phosphatase gene expression data during cranial suture fusion in the craniosynostosis mouse model and strategies compared. Strikingly, the expression trends of alkaline phosphatase and osteocalcin varied significantly when normalised to the least stable, the most stable or the three most stable genes. CONCLUSION: To minimise errors in evaluating gene expression levels, analysis of a reference panel and subsequent normalization to several stable genes is strongly recommended over normalization to a single gene. In particular, we conclude that use of single, non-validated "housekeeping" genes such as GAPDH, ACTB and 18 S rRNA, currently a widespread practice by researchers in the bone field, is likely to produce data of questionable reliability when changes are 2 fold or less, and such data should be interpreted with due caution.
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ItemDevelopment of an efficient, non-viral transfection method for studying gene function and bone growth in human primary cranial suture mesenchymal cells reveals that the cells respond to BMP2 and BMP3.(Springer Science and Business Media LLC, 2012-08-03)BACKGROUND: Achieving efficient introduction of plasmid DNA into primary cultures of mammalian cells is a common problem in biomedical research. Human primary cranial suture cells are derived from the connective mesenchymal tissue between the bone forming regions at the edges of the calvarial plates of the skull. Typically they are referred to as suture mesenchymal cells and are a heterogeneous population responsible for driving the rapid skull growth that occurs in utero and postnatally. To better understand the molecular mechanisms involved in skull growth, and in abnormal growth conditions, such as craniosynostosis, caused by premature bony fusion, it is essential to be able to easily introduce genes into primary bone forming cells to study their function. RESULTS: A comparison of several lipid-based techniques with two electroporation-based techniques demonstrated that the electroporation method known as nucleofection produced the best transfection efficiency. The parameters of nucleofection, including cell number, amount of DNA and nucleofection program, were optimized for transfection efficiency and cell survival. Two different genes and two promoter reporter vectors were used to validate the nucleofection method and the responses of human primary suture mesenchymal cells by fluorescence microscopy, RT-PCR and the dual luciferase assay. Quantification of bone morphogenetic protein (BMP) signalling using luciferase reporters demonstrated robust responses of the cells to both osteogenic BMP2 and to the anti-osteogenic BMP3. CONCLUSIONS: A nucleofection protocol has been developed that provides a simple and efficient, non-viral alternative method for in vitro studies of gene and protein function in human skull growth. Human primary suture mesenchymal cells exhibit robust responses to BMP2 and BMP3, and thus nucleofection can be a valuable method for studying the potential competing action of these two bone growth factors in a model system of cranial bone growth.