A hospital-based surveillance of rotavirus gastroenteritis in children <5 years of age in Singapore.

BACKGROUND: In Singapore, 2 rotavirus vaccines were licensed in October 2005 and July 2007, respectively, for vaccinating infants aged ≥ 6 weeks against rotavirus gastroenteritis. These vaccines are optional and are not included in the National Childhood Immunization Program. This study aimed to determine the incidence of rotavirus gastroenteritis-associated hospitalizations among children <5 years of age. METHODS: Children <5 years, who were hospitalized for acute gastro enteritis, were enrolled between September 2005 and April 2008. Stool samples were tested for the presence and serotyping of rotavirus. Incidence and proportion of gastroenteritis and rotavirus gastroenteritis cases were calculated with 95% confidence intervals. RESULTS: Among 1976 children included in the according-to-protocol cohort, 781 were rotavirus positive with a median age of 24 months (range: 0-59 months). The overall incidence of rotavirus gastroenteritis hospitalizations during the entire study period in children <5 years of age was 4.6 (95% confidence interval: 4.3-4.9) per 1000 person-years with the highest number of cases observed in children 13-24 months of age (26.5%). G1P[8] (18.3%) and G9P[8] (9.9%) were the most common rotavirus types. Rotavirus gastroenteritis hospitalizations peaked between January and March. CONCLUSION: Rotavirus infection was the primary cause of acute gastro enteritis hospitalizations among children <5 years of age, constituting nearly one-third of gastroenteritis hospitalizations in Singapore. The predominant strain observed in Singapore was G1P[8]. Results of this study suggest the need for implementation of rotavirus vaccination into National Childhood Immunization Program in Singapore.

Paediatric inflammatory bowel disease in a multiracial Asian country.

INTRODUCTION: This study examined the characteristics and trends in the incidence of inflammatory bowel disease (IBD) among children in Singapore. METHODS: We retrospectively reviewed all patients under 18 years diagnosed with IBD over a 14-year period. Information on demographics, disease presentation, laboratory findings, radiological investigations, and endoscopic and histological findings were obtained from the patients' medical records. RESULTS: 32 patients were diagnosed with IBD, 30 of whom had Crohn's disease and 2 had ulcerative colitis. The incidence of IBD rose from an initial rate of 2.2 per 100,000 patients in the year 2000 to a peak of 11.4 patients per 100,000 patients by 2008. Median age of onset of symptoms was 10.5 years. There were more boys (63%) than girls in the group and a higher representation of Indians (34.4%). The most common presenting symptoms were abdominal pain (87.5%), diarrhoea (75.0%) and weight loss (71.9%). Extraintestinal manifestations such as fever and arthralgia were found in over 50% of patients. The most common physical findings were perianal abnormalities (56.3%), mouth ulcers (37.5%) and growth failure (15.6%). Abnormal laboratory findings such as low albumin, raised erythrocyte sedimentation rate, anaemia, thrombocytosis and high C-reactive protein were found in nearly half of the patients. Endoscopic and histological findings showed that a majority of patients (90.6%) also had evidence of inflammation in the upper gastrointestinal tract. CONCLUSION: Paediatric IBD is on the rise. The higher occurrence in Indians, earlier onset and more florid presentation may suggest different genetic and environmental influences specific to Asian children.

Cell-delivery therapeutics for liver regeneration.

For acute, chronic, or hereditary diseases of the liver, cell transplantation therapies can stimulate liver regeneration or serve as a bridge until liver transplantation can be performed. Recently, fetal hepatocytes, stem cells, liver progenitor cells, or other primitive and proliferative cell types have been employed for cell transplantation therapies, in an effort to improve the survival, proliferation, and engraftment of the transplanted cells. Reviewing earlier studies, which achieved success by transplanting mature hepatocytes, we propose that there is a switch-like regulation of liver regeneration that changes state according to a stimulus threshold of extracellular influences such as cytokines, matrices and neighboring cells. Important determinants of a successful clinical outcome include sufficient quantities and functional levels of the transplanted cells (even for short periods to alter the environment), rather than just engraftment levels or survival durations of the exogenously transplanted cells. The relative importance of these determining factors will impact future choices of cell sources, delivery vehicles, and sites of cell transplantation to stimulate liver regeneration for patients with severe liver diseases.

Fibro-C-Index: comprehensive, morphology-based quantification of liver fibrosis using second harmonic generation and two-photon microscopy.

We develop a standardized, fully automated, quantification system for liver fibrosis assessment using second harmonic generation microscopy and a morphology-based quantification algorithm. Liver fibrosis is associated with an abnormal increase in collagen as a result of chronic liver diseases. Histopathological scoring is the most commonly used method for liver fibrosis assessment, where a liver biopsy is stained and scored by experienced pathologists. Due to the intrinsic limited sensitivity and operator-dependent variations, there exist high inter- and intraobserver discrepancies. We validate our quantification system, Fibro-C-Index, with a comprehensive animal study and demonstrate its potential application in clinical diagnosis to reduce inter- and intraobserver discrepancies.

An update of paediatric intussusception incidence in Singapore: 1997-2007, 11 years of intussusception surveillance.

INTRODUCTION: Understanding baseline epidemiology of intussusception (IS) in different geographical settings is important for the safety assessment of rotavirus vaccines. This paper presents IS surveillance data from Singapore between 1997 and 2007, including the period between November 2005 and December 2007 when rotavirus vaccines (primarily Rotarix) were available to newborns in Singapore. MATERIALS AND METHODS: Case ascertainment, collection, analyses and presentation of IS data was done as per recommendations of the Brighton Collaboration Working Group. For estimating the IS incidence rate in infants, live births for the years of the study were used as denominators, while for incidence in children age <2 years, the expected numbers of infant deaths occurring between 1 and 2 years of age was deducted from the combined live births for the 2 years, to obtain the denominator. RESULTS: The incidence of IS among children aged <1 year throughout this 10-year period was higher than the incidence of IS in children between 1 and 2 years of age. In 2005, 2006 and 2007, the incidence of IS per 100,000 was 39.9, 26.4 and 35.6 in children aged <1 year and 26.2, 23.8 and 28.7 in children <2 years. CONCLUSION: This IS surveillance study provides reassuring preliminary evidence that there is no increase in the incidence of IS in Singapore after the introduction of rotavirus vaccines (including Rotarix) in Singapore.

Nonlinear optical microscopy: use of second harmonic generation and two-photon microscopy for automated quantitative liver fibrosis studies.

Liver fibrosis is associated with an abnormal increase in an extracellular matrix in chronic liver diseases. Quantitative characterization of fibrillar collagen in intact tissue is essential for both fibrosis studies and clinical applications. Commonly used methods, histological staining followed by either semiquantitative or computerized image analysis, have limited sensitivity, accuracy, and operator-dependent variations. The fibrillar collagen in sinusoids of normal livers could be observed through second-harmonic generation (SHG) microscopy. The two-photon excited fluorescence (TPEF) images, recorded simultaneously with SHG, clearly revealed the hepatocyte morphology. We have systematically optimized the parameters for the quantitative SHG/TPEF imaging of liver tissue and developed fully automated image analysis algorithms to extract the information of collagen changes and cell necrosis. Subtle changes in the distribution and amount of collagen and cell morphology are quantitatively characterized in SHG/TPEF images. By comparing to traditional staining, such as Masson's trichrome and Sirius red, SHG/TPEF is a sensitive quantitative tool for automated collagen characterization in liver tissue. Our system allows for enhanced detection and quantification of sinusoidal collagen fibers in fibrosis research and clinical diagnostics.

Crigler-Najjar syndrome type 2.

Crigler-Najjar syndrome is a rare disorder of bilirubin metabolism with two distinct forms: type 1 and type 2. We report three patients with Crigler-Najjar syndrome type 2 (CN-2). All patients had serum bilirubin values higher than 171 micromol/L and deep yellow skin color. The results of other liver function tests, glucose-6-phosphate dehydrogenase activity and hematology tests were normal, and immunologic tests for hepatitis A, B and C were negative, although one patient had slightly elevated alanine aminotransferase level (45 IU/L). Polymerase chain reaction and sequence analysis of the UDP-glucuronosyltransferase 1A1 (UGT1A1) gene revealed a novel homozygous T>A mutation at nucleotide 479 in exon 1 (Val160Glu) of patient 1, a novel homozygous A>G mutation at nucleotide 610 in exon 1 (Met204Val) of patient 2, and a homozygous T>G variation at nucleotide 1456 in exon 5 (Tyr486Asp) plus a heterozygous G>A variation at nucleotide 211 in exon 1 (Gly71Arg/normal) of patient 3. Two of these mutations were novel and variations identified within the coding region of the UGT1A1 gene were considered the cause of CN-2 in all three patients.

Significance of low or normal serum gamma glutamyl transferase level in infants with idiopathic neonatal hepatitis.

INTRODUCTION: We evaluated the significance of low/normal serum gamma glutamyl transferase (GGT) level in infants with idiopathic neonatal hepatitis (INH). MATERIALS AND METHODS: A retrospective review of the hospital records of 103 infants less than 3 months of age who were diagnosed with INH between August 1991 and November 2000 was performed. Variables including age at which jaundice was noticed, age at presentation, perinatal risk factors, family history of liver disease, parental consanguinity, initial ultrasound scan, liver biopsy, laboratory values at the first visit, the peak levels of total bilirubin, aspartate aminotransferase (AST), GGT and alkaline phosphatase (ALP) in the first 3 months of follow-up and interval for normalisation of serum bilirubin and AST were compared between infants presenting with low/normal GGT (100 U/L). RESULTS AND DISCUSSION: Infants with low/normal GGT levels presented earlier (median 36.5 days versus 44 days; p=0.016) and had significantly higher bilirubin and AST levels at presentation (bilirubin 167.5 micromol/L versus 133 micromol/L; p<0.005 and AST 187.5 U/L versus 106 U/L; p<0.001) and at peak levels (bilirubin 170 micromol/L versus 146 micromol/L; p=0.024 and AST 210.5 U/L versus 129 U/L; p=0.001). A significant correlation was also found between GGT levels and serum albumin levels (p=0.004). Patients with low/normal GGT levels were more likely to have giant cell hepatitis on histology (p=0.015). There was no difference in time taken to recovery. CONCLUSION: Low/normal levels of GGT in INH infants may be a predictor of more severe but recoverable disease.

UCH-L1 aggresome formation in response to proteasome impairment indicates a role in inclusion formation in Parkinson's disease.

Aggresomes are associated with many neurodegenerative disorders, including Parkinson's disease, and polyglutamine disorders such as Huntington's disease. These inclusions commonly contain ubiquitylated proteins. The stage at which these proteins are ubiquitylated remains unclear. A malfunction of the ubiquitin/proteasome system (UPS) may be associated with their formation. Conversely, it may reflect an unsuccessful attempt by the cell to remove them. Previously, we demonstrated that overexpression of Parkin, a ubiquitin-protein ligase associated with autosomal recessive juvenile Parkinsonism, generates aggresome-like inclusions in UPS compromised cells. Mutations in the de-ubiquitylating enzyme, UCH-L1, cause a rare form of Parkinsonism. We now demonstrate that overexpression of UCH-L1 also forms ribbon-like aggresomes in response to proteasomal inhibition. Disease-associated mutations, which affect enzymatic activities, significantly increased the number of inclusions. UCH-L1 aggresomes co-localized with ubiquitylated proteins, HSP70, gamma-tubulin and, to a lesser extent, the 20S proteasome and the chaperone BiP. Similar to Parkin inclusions, we found UCH-L1 aggresomes to be surrounded by a tubulin rather than a vimentin cage-like structure. Furthermore, UCH-L1 aggregates with Parkin and alpha-synuclein in some, but not all inclusions, suggesting the heterogeneous nature of these inclusion bodies. This study provides additional evidence that aggregation-prone proteins are likely to recruit UPS components in an attempt to clear proteins from failing proteasomes. Furthermore, UCH-L1 accumulation is likely to play a pathological role in inclusion formation in Parkinson's disease.

Human homologue of ariadne promotes the ubiquitylation of translation initiation factor 4E homologous protein, 4EHP.

Human homologue of Drosophila ariadne (HHARI) is a RING-IBR-RING domain protein identified through its ability to bind the human ubiquitin-conjugating enzyme, UbcH7. We now demonstrate that HHARI also interacts with the eukaryotic mRNA cap binding protein, translation initiation factor 4E homologous protein (4EHP), via the N-terminal RING1 finger of HHARI. HHARI, 4EHP and UbcH7 do not form a stable heterotrimeric complex as 4EHP cannot immunoprecipitate UbcH7 even in the presence of HHARI. Overexpression of 4EHP and HHARI in mammalian cells leads to polyubiquitylation of 4EHP. By contrast, HHARI does not promote its own autoubiquitylation. Thus, by promoting the ubiquitin-mediated degradation of 4EHP, HHARI may have a role in both protein degradation and protein translation.

Inhibition of proteasomal activity causes inclusion formation in neuronal and non-neuronal cells overexpressing Parkin.

Association between protein inclusions and neurodegenerative diseases, including Parkinson's and Alzheimer's diseases, and polyglutamine disorders, has been widely documented. Although ubiquitin is conjugated to many of these aggregated proteins, the 26S proteasome does not efficiently degrade them. Mutations in the ubiquitin-protein ligase Parkin are associated with autosomal recessive juvenile Parkinsonism. Although Parkin-positive inclusions are not detected in brains of autosomal recessive juvenile Parkinsonism patients, Parkin is found in Lewy bodies in sporadic disease. This suggests that loss of Parkin ligase activity via mutation, or sequestration to Lewy bodies, is a contributory factor to sporadic disease onset. We now demonstrate that decreased proteasomal activity causes formation of large, noncytotoxic inclusions within the cytoplasm of both neuronal and nonneuronal cells overexpressing Parkin. This is not a general phenomenon as there is an absence of similar inclusions when HHARI, a structural homolog of Parkin, is overexpressed. The inclusions colocalize with ubiquitin and with proteasomes. Furthermore, Parkin inclusions colocalize with gamma-tubulin, acetylated alpha-tubulin, and cause redistribution of vimentin, suggesting aggresome-like properties. Our data imply that lower proteasomal activity, previously observed in brain tissue of Parkinson's disease patients, leads to Parkin accumulation and a concomitant reduction in ligase activity, thereby promoting Lewy body formation.