'n Ondersoek na die hiperlipidemie-veroorsakende mutasies en hul geassosieerde haplotipes in Suid-Afrikaanse hiperlipidemiese kleurlinge

Simple item page
dc.contributor.advisorOdendaal, H. J.en_ZA
dc.contributor.advisorKotze, Maritha J.en_ZA
dc.contributor.authorLoubser, Odellen_ZA
dc.contributor.otherStellenbosch University. Faculty of Medicine and Health Sciences. Department of Biomedical Sciences.
dc.date.accessioned2012-08-27T11:39:00Z
dc.date.available2012-08-27T11:39:00Z
dc.date.issued1994
dc.descriptionThesis (MSc)--Stellenbosch University, 1994.
dc.description.abstractENGLISH ABSTRACT: Coronary heart disease (CHD), common among South African whites, is increasingly manifesting itself in the coloured population of South Africa. More than 70% of coloureds participating in the CRISIC I study, a study undertaken to investigate CHD risk factors in the coloureds of the Cape Peninsula, had serum cholesterol levels imparting CHD risk. Increased levels of plasma low-density lipoprotein cholesterol (LDL-C) are the hallmark of the disease familial hypercholesterolemia (FH). The underlying molecular defect of FH consists of mutations in the low-density lipoprotein (LDL) receptor gene. The coloured study population was screened for the presence of the three founder related point mutations in the LDL receptor gene that account for more than 90% of FH cases in the Afrikaner population of South Africa. The prevalence of the FH Afrikaner mutations was determined in 31 FH heterozygotes and 39 hyperlipidemic coloureds. The 39 hyperlipidemic coloureds did not meet the criteria for heterozygous FH. The FH Afrikaner-1 (Asp₂₀₆→Glu , FH-1) en FH-Afrikaner-2 (Val₄₀₈→Met FH-2) mutations had the same prevalence, 12.9% (4 individuals), in the FH group. The FH Afrikaner-3 (Asp₁₅₄→Asn, FH-3) mutation was absent in the FH group. The three FH Afrikaner mutations were not detected in the hyperlipidemic group of 39 individuals. Haplotype analysis in the families with the FH-1 mutation showed that the chromosomal background of the mutation was compatible with that described in the Afrikaner patients. In the FH-2 heterozygous patients the mutation was found on a haplotype that differed from the Afrikaner haplotype at polymorphic sites at both sides of the defect in exon 9. These results indicate that identical LDL-receptor gene mutations originated in two different South African population groups due to independent events at a potential CpG dinucleotide "hotspot". The clinical effects of the FH-1 and FH-2 mutations were the same as described in the Afrikaners. Heteroduplex analysis detected a 3 bp deletion that causes deletion of amino acid Gly₁₉₇ (FH Lithuania) in one of 23 FH heterozygous coloured patients (4.3 %) and in one indiviaual of 90 hyperlipidemic coloureds (1.1%) already screened for the FH Afrikaner mutations , by means of heteroduplex analysis. A hyperlipidemic group of 66 coloureds, screened for the first time for mutations in exon 4 of the LDL-receptor gene by means of heteroduplex analysis, showed one individual (1.5%) with the FH Lithuania mutation. In all cases, the haplotype of the mutant allele was compatible with that described in Ashkenazi Jews. The heteroduplex screening method was sensitive enough to detect the FH-1 mutation, as 5 individuals of the group of 66 hyperlipidemic coloureds were found to have the FH-1 mutation. A total of 22 FH heterozygous coloured patients, negative for the above mentioned mutations, were screened for new mutations in exon 4 of the LDL-receptor gene by means of the single-strand-conformation-polymorphism (SSCP) technique and the heteroduplex method. A new mutation, hitherto not described, was detected in one patient by means of the SSCP technique. A point mutation (A to C) at nucleotide 671 in exon 4 of the LDL receptor cDNA results in an amino acid change from aspartic acid to alanine at residue 203 in the cysteine rich ligand binding domain of the LDL receptor. The mutation gives rise to an additional Hae Ill restriction site in the DNA of affected subjects. The accurate diagnosis of this so-called FH Tygerberg mutation in subjects with FH is possible now by means of the amplification of genomic DNA (using the polymerase chain reaction) and restriction enzyme analysis. The allele frequencies of four restriction-fragment-length-polymorphisms (RFLP's) in the LDL-receptor gene were determined for 17 FH heterozygous patients and 21 control individuals. Statistical analysis indicated a significant rise in the frequency of the least frequent allele of the Ava 11 polymorphism (x2= 7.14, P<0.01). This indicates population association of the allele with a gene that causes FH. The heterozygosities and PIC values of each RFLP site were determined, and these values disclosed that the Sma I site would be the most informative in genetic linkage studies, followed by the Ava II and Nco I sites. These three sites were all moderately polymorphic. The Stu I site was the least polymorphic and therefore would be the least informative of the four RFLP sites in genetic studies. A 478 bp deletion in exon 18 of the LDL-receptor gene was detected in 2 FH heterozygous coloured patients. One of the two patients was heterozygous for the FH-1 mutation. This deletion is most likely a polymorphism as it was detected in the 3'-nontranslated region of the LDL-receptor gene and therefore it may be used in genetic marker analysis. This study suggests that the moderately informative RFLP sites (Sma I, Ava II and Nco I) must be combined with the TA dinucleotide repeat locus, also moderately informative, to indirectly diagnose FH in families where the direct molecular diagnosis of FH, by means of the detection of the FH associated mutations, is not possible. A better molecular diagnostic service is now possible for the coloureds with a high risk for CHD by combining the screening for FH associated mutations with RFLP and dinucleotide repeat polymorphism studies.en_ZA
dc.description.abstractAFRIKAANSE OPSOMMING: Koronêre hartsiekte (KHS), wat algemeen is onder Suid-Afrikaanse blankes, is toenemend besig om te manifesteer in die Kleurlingpopulasie van Suid-Afrika. Meer as 70% van Kleurlinge in die CRISIC I studie, wat onderneem is om die risikofaktore vir die ontwikkeling van KHS in die Kleurlinge van die Kaapse Skiereiland te bepaal, het serum cholesterolwaardes gehad wat 'n hoë risiko vir die ontwikkeling van KHS aandui. Verhoogde plasma lae-digtheid lipoproteïen cholesterol- (LDL-C) vlakke is die algemeenste kenmerk van die siekte familiële hipercholesterolemie (FH), wat veroorsaak word deur mutasies in die lae-digtheid lipoproteïen (LDL) reseptor geen. Die kleurling studie-populasie is gesif vir die drie stigterverwante puntmutasies in die LDL reseptor geen, wat verantwoordelik is vir meer as 90% van FH-gevalle in die Afrikanerpopulasie van Suid-Afrika. Die voorkoms van die FH-Afrikaner mutasies is ondersoek in 31 FH heterosigote, asook 39 hiperlipidemiese Kleurlinge wat nie aan die kriteria voldoen het vir die insluiting in die FH heterosirotiese groep nie. Die FH-Afrikaner-1 (Asp₂₀₆→Glu , FH-1) en FH-Afrikaner-2 (Val₄₀₈→Met FH-2) mutasies se voorkoms in die FH-groep was dieselfde, nl. 12.9% (4 individue), terwyl die FH-Afrikaner-3 (Asp₁₅₄→Asn, FH-3) mutasie afwesig was. Die drie FH-Afrikaner mutasies is nie opgespoor in die hiperlipidemiese groep van 39 individue nie. Haplotipe-analise in families met die FH-1 mutasie het getoon dat die mutasie op dieselfde chromosomale agtergrond voorkom as wat voorheen in die Afrikaners beskryf is. In die FH-2 heterosigote was die mutasie geassosieerd met 'n haplotipe wat by polimorfiese setels aan beide kante van die defek in ekson 9 verskil van die Afrikanerhaplotipe. Die resultate dui dus aan dat identiese LDL reseptor geen mutasies in die twee verskillende Suid-Afrikaanse populasiegroepe as gevolg van onafhanklike gebeure by 'n potensiële CpG mutasie-geneigde setel ontstaan het. Die kliniese effek van die FH-1 en FH-2 mutasies in die Kleurlingpasiënte stem ooreen met wat reeds beskryf is in die Afrikaners. 'n Bekende 3 bp delesie van die aminosuur Gly₁₉₇ (FH-Lithuania) is waargeneem in een individu (4.3%) van die 23 Kleurling FH heterosigote en in een individu van 90 hiperlipidemiese Kleurlinge (1 .1%), by wie die FH-Afrikaner mutasies reeds uitgeskakel is, deur gebruik te maak van heterodupleks-analise. Uit 'n hiperlipidemiese groep van 66 Kleurlinge, waarvan PKR-produkte van ekson 4 van die LDL reseptor geen vir die eerste keer aan mutasie-analise onderwerp is. het een individu (1 .5%) die FH-Lithuania mutasie getoon. In alle gevalle het die mutasie op dieselfde haplotipe voorgekom as wat beskryf is in Askenazi Jode. Die heterodupleksmetode was sensitief genoeg om die FH-1 mutasie te identifiseer, aangesien 5 individue met die mutasie opgespoor is in die 66 hiperlipidemiese Kleurlinge wat nog nie voorheen aan mutasie-analise onderwerp is nie. 'n Totaal van 22 FH heterosigote, negatief vir bg. mutasies, is gesif vir nuwe mutasies in ekson 4 van die LDL reseptor geen d.m.v. heterodupleks-analise en die enkelstring-konformasie-polimorfisme (ESKP) opsporingstegniek. In een pasiënt is 'n nuwe mutasie d.m.v. die ESKP tegniek geïdentifiseer. Die A na G puntmutasie by nukleotiedposisie 671 in ekson 4 van die LDL reseptor cDNA, lei tot die aminosuurverandering van aspartiensuur na alanien by kodon 203 in die sisteïenryke ligandbindende gebied van die LDL reseptor. Aangesien die nuwe mutasie 'n Hae Ill restriksie-endonuklease setel skep, kan restriksie-endonuklease analise van PKR-geamplifiseerde genomiese DNA die akkurate diagnose van hierdie sogenaamde FH-Tygerberg mutasie toelaat. Die alleelfrekwensies van vier restriksie-fragment-lengte-polimorfismes (RFLP's) van die LDL reseptor geen is ondersoek in 17 FH heterosigote en 21 kontrole individue. 'n Statisties beduidende verhoging in die skaars aileel van die Ava II polimorfisme (x2 = 7.14, P<0.01) is waargeneem. Dit dui op populasie assosiasie van die skaars alleel van die Ava II polimorfisme met 'n geen wat FH veroorsaak. Die heterosigositeit- en Pll waardes is vir elke RFLP setel bereken en daaruit het dit geblyk dat die Sma I setel die informatiefste sal wees in genetiese koppelingstudies, gevolg deur die Ava II en Nco I setels. Laasgenoemde drie setels is almal redelik informatief, terwyl die Stu I setel die minste informatief is van die vier RFLP setels. In 2 FH heterosigote is 'n 478 bp delesie in ekson 18 van die LDL reseptor geen waargeneem. Een van hierdie pasiënte was heterosigoties vir die FH-1 mutasie. Die delesie word as 'n polimorfisme beskou, aangesien dit in die 3'ongetransleerde gedeelte van die LDL reseptor geen voorkom. Dit mag moontlik in genetiese merkerstudies gebruik word. Uit hierdie studie kan afgelei word dat die redelik informatiewe RFLP setels (Sma I, Ava II en Nco I), en die TA-herhalende-dinukleotiedlokus, ook redelik informatief, gekombineer moet word om FH indirek te diagnoseer in families waar die direkte molekulêre diagnose van FH, deur die aantoon van FH-verwante mutasies, nie moontlik is nie. 'n Beter molekulêre diagnostiese diens is nou moontlik vir die Kleurlingpopulasie, met 'n hoë risiko vir die ontwikkeling van koronêre hartsiekte, deur die kombinering van die sifting vir FH-verwante mutasies met RFLP en dinukleotied-herhalende-polimorfisme studies.af_ZA
dc.format.extent151 pages
dc.identifier.urihttp://hdl.handle.net/10019.1/58483
dc.language.isoaf_ZA
dc.publisherStellenbosch : Stellenbosch University
dc.rights.holderStellenbosch University
dc.subjectHyperlipidemiaen_ZA
dc.subjectDissertations -- Medicineen_ZA
dc.subjectUCTDen_ZA
dc.title'n Ondersoek na die hiperlipidemie-veroorsakende mutasies en hul geassosieerde haplotipes in Suid-Afrikaanse hiperlipidemiese kleurlingeaf_ZA
dc.typeThesis
Files
Original bundle
Now showing 1 - 1 of 1
Thumbnail Image
Name:
loubser_ondersoek_1994.pdf
Size:
16.17 MB
Format:
Adobe Portable Document Format
Description:
Download Thesis
Download
Collections
Masters Degrees (Molecular Biology and Human Genetics)