Diss. (sammanfattning) Umeå : Umeå universitet, 2005. / Härtill 4 uppsatser.

Lai Ching Ha. / Thesis (Ph.D.)--Chinese University of Hong Kong, 1995. / Includes bibliographical references (leaves 225-243). / Title --- p.1 / Abstract --- p.2 / Acknowledgments --- p.4 / Contents --- p.5 / Abbreviations --- p.10 / List of Figures --- p.12 / List of Tables --- p.15 / Chapter Chapter 1 --- Review on Inherited Metabolic Diseases --- p.18 / Chapter 1.1 --- Development of the concept of inherited metabolic diseases (IMD) --- p.18 / Chapter 1.2 --- Frequency of inherited metabolic diseases --- p.20 / Chapter 1.3 --- Molecular basis of mutations in inherited metabolic diseases --- p.22 / Chapter 1.3.1 --- Point mutations --- p.22 / Chapter 1.3.2 --- Small deletions and insertions --- p.25 / Chapter 1.3.3 --- large deletions or duplications --- p.26 / Chapter 1.4 --- Pathological consequences of protein defect resultingin IMD --- p.27 / Chapter 1.4.1 --- End product --- p.28 / Chapter 1.4.2 --- Precursor accumulation --- p.28 / Chapter 1.4.3 --- Unusual metabolites --- p.29 / Chapter 1.5 --- Heterogeneity of inherited metabolic diseases --- p.29 / Chapter 1.5.1 --- Genetic heterogeneity --- p.29 / Chapter 1.5.2 --- Variations of expression in different cells --- p.31 / Chapter 1.6 --- Diagnosis of inherited metabolic diseases --- p.32 / Chapter 1.6.1. --- Biochemical investigations --- p.32 / Chapter 1.6.2 --- Identification of accumulated or missing metabolites --- p.33 / Chapter 1.6.3 --- Direct analysis of enzymes and proteins --- p.34 / Chapter 1.6.4 --- Molecular investigations --- p.34 / Chapter 1.7 --- Treatment of inherited metabolic diseases --- p.40 / Chapter 1.7.1 --- Treatment at the clinical phenotype level --- p.41 / Chapter 1.7.2 --- Treatment at the metabolite level --- p.41 / Chapter 1.7.3 --- Treatment at the dysfunctional protein level --- p.43 / Chapter 1.7.4 --- Transplantation --- p.44 / Chapter 1.7.5 --- Gene therapy --- p.45 / Chapter 1.8 --- Inherited metabolic diseases in Hong Kong --- p.47 / Chapter 1.9 --- General Aim --- p.48 / Chapter Chapter 2 --- Study of Inherited Metabolic Diseases in Mentally Retarded Patients --- p.49 / Chapter 2.1 --- Introduction --- p.49 / Chapter 2.2 --- Aim --- p.52 / Chapter 2.3 --- Materials --- p.53 / Chapter 2.3.1 --- Standards --- p.53 / Chapter 2.3.2 --- Chemical reagents --- p.53 / Chapter 2.3.3 --- Derivatization reagents --- p.54 / Chapter 2.3.4 --- Major equipment --- p.54 / Chapter 2.4 --- Clinical materials --- p.56 / Chapter 2.4.1 --- Subjects --- p.55 / Chapter 2.4.2 --- Blood and urine samples --- p.56 / Chapter 2.5 --- Methods --- p.57 / Chapter 2.5.1 --- General biochemistry tests --- p.57 / Chapter 2.5.2 --- Metabolic screening tests --- p.57 / Chapter 2.5.3 --- Two-dimensional thin layer chromatography --- p.53 / Chapter 2.5.4 --- Identification of urinary organic acids by gas chromatography mass spectroscopy --- p.59 / Chapter 2.5.5 --- Amino acid analysis by high performance liquid chromatography --- p.66 / Chapter 2.6 --- Results --- p.71 / Chapter (A) --- Methodological Aspects / Chapter 2.6.1 --- Identification of urinary organic acids by gas chromatography-mass spectroscopy (GC-MS) --- p.71 / Chapter 2.6.2 --- Amino acid analysis by high performance liquid chromatography (HPLC) --- p.86 / Chapter (B) --- Patient Investigations / Chapter 2.6.3 --- General biochemistry tests --- p.107 / Chapter 2.6.4 --- Serum amino acid profiles --- p.113 / Chapter 2.6.5 --- Urinary organic acid analysis --- p.115 / Chapter 2.6.6 --- Case reports --- p.119 / Chapter 2.7 --- Discussion --- p.123 / Chapter 2.7.1 --- Identification of urinary organic acids by gas chromatography-mass spectroscopy (GC-MS) --- p.123 / Chapter 2.7.2. --- Amino acid analysis by high performance liquid chromatography (HPLC) --- p.130 / Chapter 2.7.3 --- Identification of inherited metabolic diseases (IMD)in an institutionalized mentally retarded patients --- p.136 / Chapter Chapter 3 --- Molecular Investigation of Maple Syrup Urine Disease --- p.140 / Chapter 3.1 --- Introduction --- p.140 / Chapter 3.1.1 --- Branched chain amino acids (BCAA) --- p.140 / Chapter 3.1.2 --- Metabolism of branched chain amino acids --- p.142 / Chapter 3.1.3 --- Maple syrup urine disease (MSUD) --- p.144 / Chapter 3.1.4 --- Classification of maple syrup urine disease --- p.146 / Chapter 3.1.5 --- Screening and diagnosis of maple syrup urine disease --- p.148 / Chapter 3.1.6 --- Treatment of maple syrup urine disease --- p.150 / Chapter 3.1.7. --- Branched chain a-ketoacid dehydrogenase complex (BCKDH) --- p.151 / Chapter 3.1.8 --- "Gene features of human E1α,E1β and E2 subunitsin branched chain α-ketoacid dehydrogenase complex" --- p.153 / Chapter 3.1.9 --- Molecular defects of the BCKDH gene complex --- p.156 / Chapter 3.1.10 --- MSUD in Hong Kong --- p.161 / Chapter 3.2 --- Aim --- p.163 / Chapter 3.3 --- Materials --- p.164 / Chapter 3.3.1 --- Source of skin fibroblasts --- p.164 / Chapter 3.3.2 --- Enzymes --- p.164 / Chapter 3.3.3 --- DNA markers --- p.164 / Chapter 3.3.4 --- Reagent Kits --- p.165 / Chapter 3.3.5 --- Primers --- p.165 / Chapter 3.3.6 --- Chemical reagents --- p.165 / Chapter 3.3.7 --- Nitrocellulose membrane --- p.166 / Chapter 3.3.8 --- Antiserum for Western blotting --- p.166 / Chapter 3.3.9 --- Radioisotopes --- p.166 / Chapter 3.4 --- Methods --- p.168 / Chapter 3.4.1 --- Preparation of buffers and solutions --- p.168 / Chapter 3.4.2 --- Agarose gel electrophoresis --- p.170 / Chapter 3.4.3 --- Preparation of native polyacrylamide gel --- p.171 / Chapter 3.4.4 --- Preparation of sodium dodecyl sulfate (SDS) polyacrylamide gel --- p.172 / Chapter 3.4.5 --- Preparation of denaturing polyacrylamide gel --- p.173 / Chapter 3.4.6 --- Branched chain α-ketoacid dehydrogenase complex enzyme assay --- p.173 / Chapter 3.4.7. --- Identification of the affected subunits in BCKDH complex of MSUD patient and her family members --- p.176 / Chapter 3.4.8 --- Screening of mutation in the BCKDH subunits by RT-PCR-SSCP --- p.178 / Chapter 3.4.9 --- Mutation analysis of whole cDNA fragments of Elα， Elβ and E2 subunits by ds DNA cycle sequencing --- p.184 / Chapter 3.5 --- Results --- p.188 / Chapter 3.5.1 --- Branched chain α-ketoacid dehydrogenase complex enzyme assay --- p.188 / Chapter 3.5.2 --- Identification of the affected subunits in BCKDH complex ofMSUD patient and her family members --- p.188 / Chapter 3.5.3 --- Screening of mutation in the BCKDH subunits by RT-PCR-SSCP --- p.192 / Chapter 3.5.4 --- "Mutation analysis of whole cDNA fragments of Ela, Elβ and E2 subunits by ds DNA cycle sequencing" --- p.204 / Chapter 3.6 --- Discussion --- p.210 / Chapter 3.6.1 --- BCKDH activity in the MSUD patient and her family members --- p.210 / Chapter 3.6.2 --- Investigation of the mutation sites --- p.212 / General Conclusion --- p.222 / Appendix --- p.224 / References --- p.225

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Thesis (doctoral)--University of Wisconsin-Madison, 2000. / Includes bibliographical references.

Research Project (M.B.A.) - Simon Fraser University, 2007. / Theses (Faculty of Business Administration) / Simon Fraser University. Senior supervisor: Dr. Aidan Vining -- Faculty of Business Administration. MBA-MOT Program. Also issued in digital format and available on the World Wide Web.

Inborn errors of metabolism (IEM) are under international spotlight because of the recent tremendous development in expanded newborn screening (NBS) and molecular genetics. IEM is a difficult subject involving more than 1,000 different disorders with protean clinical presentations and complicated diagnostic pathways. Cumulative incidence of IEM was reported up to 1 in 800. Patients can be affected in any ages. High clinical suspicion alone is not sufficient to reduce morbidities and mortalities. Notably, some IEM are amenable to treatment with promising outcome. Local data regarding the disease spectrum and incidences is largely lacking. Public awareness and readiness for expanded NBS is unknown. This renders difficulties in the consideration of expanded NBS in Hong Kong. In this study, laboratory data of classical IEM from 2005 to 2009 were retrospectively analyzed (Chapter 2). Local incidence was 1 in 4,122 and that of hyperphenylalaninemias was 1 in 29,542, similar to worldwide figures. Majority (69%) was amino acid disorders, 12% was organic acidemias and 19% was fatty acid oxidation defects. Most of these diseases are effectively amenable to treatment. Local cases including hyperphenylalaninemia, tyrosinemia type I, arginase deficiency, ornithine transcarbamylase deficiency, very long-chain acyl-CoA dehydrogenase deficiency, tyrosine hydroxylase deficiency, thermolabile carnitine palmitoyltransferase II variants and adults IEM with X-linked adrenoleukodystrophy, cerebrotendinous xanthomatosis, familial transthyretin amyloidosis, Wilson disease and PANK2-associated young-onset Parkinsonism were described (Chapters 1.1.3 and 3). Electronic chemical pathology consultation service and dried blood spot metabolic screening were implemented (Chapter 4). There were 279 consultations and 158 screening in a 12-month period. Major referral reasons were developmental delay, neurological defects and unexplained biochemical abnormalities. The incidence in high risk screening was 1 in 158. A non-derivatized tandem mass spectrometry assay for amino acids and acylcarnitines was evaluated for its precision, accuracy and reference intervals (Chapter 5). The concordance rate was 100% in inter-laboratory comparison and external quality assurance programs. The method was proven to be accurate, rapid and affordable. It is suitable for large volume testing and emergency diagnostic needs. A feasibility study of a hospital-based expanded NBS service model was conducted on 360 newborns (Chapter 6). More than 90% of babies were older than 48 hours before discharge and were fit for blood collection. The service model consisted of parent education, consent, postnatal sample collection, technical analysis, clinical interpretation, reporting and follow-up actions. Questionnaire on the knowledge and attitude towards IEM and expanded NBS was surveyed on 172 parents to investigate the psychological, social and ethical aspects (Chapter 7). Here, 99.4% demanded more education on expanded NBS; 97.6% supported to implement the program; 97.7% supported population screening even though some diseases are incurable. Availability of treatment is not the most important pre-requisite for NBS; 93.9% accepted the possibility of false positive and false negative results. Acceptance towards expanded NBS among parents was high. Our data indicate that IEM is not uncommon in Hong Kong and it is indisputable for the introduction of a local expanded NBS program. Our data serve as groundwork for policy decision and further discussion on expanded NBS. / published_or_final_version / Pathology / Master / Doctor of Medicine

Human skin fibroblasts derived from patients with all 7 known inborn errors of vitamin B$ sb{12}$ metabolism have been studied for functional integrity of methylmalonyl CoA mutase and methionine synthase. Cocultivation of cblC and cblF fibroblasts in the absence of polyethylene glycol resulted in a twofold increase over the expected in both ($ sp{14}$C) propionate and ($ sp{14}$C) methyltetrahydrofolate incorporation into acid-precipitable material, suggesting that metabolic cooperation between cells occurs. CblD fibroblasts, which are biochemically similar to cblC cells (Goodman et al, 1970; Willard et al, 1977), do not cooperate metabolically when mixed with cblF cells. Partial correction in phenotype was seen in mixtures of cblD and cblG cells, but not cblC and cblG cells. These observations lend further support for the division of cblC and cblD disease into two discrete complementation classes. Cocultivation of cblF fibroblasts with both cblE and cblG cells also resulted in partial correction in phenotype. / ($ sp{14}$C) Propionate incorporation in both cblC and cblF cells exposed to conditioned medium from control cells was increased more than twofold. ($ sp{14}$C) methyltetrahydrofolate incorporation in cblC cells exposed to conditioned medium from cblF cells was increased twofold. This suggests the presence of a diffusible factor correcting the defect in the mutant cell lines.

Prolidase deficiency (McKusick 26413) is a rare autosomal recessive disorder characterized by iminodipeptiduria, skin lesions and mental retardation. The enzyme prolidase hydrolyzes dipeptides containing C-terminal proline or hydroxyproline. / The results presented here indicate that prolidase plays a major role in the recycling of dipeptide bound proline. Control fibroblasts were able to use iminodipeptides in lieu of proline to sustain normal growth and protein synthesis whereas prolidase deficient cells were not. / Iminodipeptides added to the media of control and mutant cells showed no adverse effects on protein synthesis or cell growth. These results are consistent with a mechanism of biochemical pathology in which proline deprivation caused by the enzyme deficit is the cause of damage to skin cells. / Prolidase regulation by product and substrate was studied. A two fold decrease of prolidase activity was observed in fibroblasts grown in excess proline. However, cells grown in medium in which iminodipeptides replaced proline showed no significant difference in prolidase activity.