Thesis (PhD)--University of Stellenbosch, 2003. / ENGLISH ABSTRACT: The solute carrier family 11 member 1 gene (SLC11A 1) is a divalent metal ion
transporter with various pleiotropic effects on macrophage function. This gene that
regulates iron, and is also regulated by cellular iron levels, has previously been linked
to many infectious and autoimmune diseases. In this analysis, in vitro studies using
the luciferase reporter system as well as case-control association studies were
applied to investigate the significance of SLC11 A1 allelic variation in patients with
diverse disease phenotypes.
For in vitro studies, five different SLC11A 1 promoter constructs were generated,
followed by transfection into U937 and THP-1 cells. The inserted fragments included
two previously described alleles (alleles 2 and 3), two novel alleles identified in this
study (alleles 8 and 9) and a C to T point mutation at nucleotide position -237 in the
presence of allele 3. The most striking finding was the opposite effect observed for
allele 3 in the presence of the -237C~ T polymorphism, similar to that of allele 2.
Although the SLC11A 1 gene has previously been implicated in iron transport, we
have demonstrated, for the first time, that the various alleles investigated cause
differential expression of the gene upon iron loading.
Association studies were performed by investigating diseases including oesophageal
cancer (DC), inflammatory bowel disease (lBO) and hereditary haemochromatosis
(HH) (or primary iron overload). Significant associations (P<O.05) were observed with
allele 3 for all three conditions investigated only after stratification according to the
presence of the -237C~ T polymorphism. Re-assessment of the promoter alleles
according to expression profiles determined by the in vitro studies, showed
statistically significant associations for allele 3 with DC and primary iron overload,
compared with the respective population-matched control groups. Additionally,
several novel variants were identified in exon 2 (112G~A, 148deIGACCAGCCC,
157insGACCAGCCCAG) and intron 1 (IVS1-28C~T), with variant IVS1-28C~T
occurring at a significantly increased frequency in patients with DC compared with
population-matched controls (P<O.05). Investigation of the SLC11A 1 gene in
individuals presenting with iron overload in the absence of homozygosity for the HFE
C282Y mutation, provided further support for the importance of sequence variation in the promoter region of the SLC11A 1 gene in modified risk of iron-related disorders.
Genes related to iron homeostasis, including HFE, SLC11A3, HAMP and DCYTB,
were investigated in individuals with similar criteria and potential disease-causing
mutations were identified in 11% White and 45% Black South African patients. The
possible significance of the SLC11A3 and DCYTB genes in iron overload in the Black
South African population, and the possible involvement of the DCYTB gene in iron
overload in general, are demonstrated for the first time.
This study contributed to a better understanding of the function of the SLC11A 1 gene
in relation to iron metabolism. The involvement of SLC11A 1 in a range of disease
phenotypes including cancer and inflammatory conditions that may involve iron
dysregulation, can probably be explained by interaction with external factors such as
infectious agents that may affect cellular iron status. Our findings provide both in vivo
and in vitro evidence that iron dysregulation mediated by allelic effects of SLC11A 1
may underlie disease susceptibility to infectious and autoimmune conditions. / AFRIKAANSE OPSOMMING: Die opgeloste stof draer familie 11 deel 1 geen (SLC11 A 1) is 'n divalente metaal ioon
vervoerder met verskeie pleiotropiese effekte op makrofaagfunksie. Die geen, wat
yster reguleer en ook deur sellulêre ystervlakke gereguleer word, is voorheen verbind
met verskeie infektiewe en outo-immune siektes. In hierdie studie is in vitro analises,
deur middel van die lusiferase verklikker sisteem, asook gevalle-kontrole assosiasie
studies gebruik om die rol van SLC11A 1 alleel variasie in pasiënte met diverse
siektefenotipes te ondersoek.
Vyf verskillende SLC11A 1 promotor variante is geskep vir in vitro studies en gevolg
deur transfeksie in U937 en THP-1 sellyne. Die ingevoegde fragmente het twee
voorheen beskryfde allele (allele 2 en 3), twee nuwe allele wat in hierdie studie
geïdentifiseer is (allele 8 en 9) en In C na T puntmutasie by nukleotied posisie -237
in die teenwoordigheid van alleel 3 ingesluit. Die opvallendste bevinding was die
teenoorgestelde effek wat waargeneem is wanneer alleel 3 in die teenwoordigheid
van die -237C~ T polimorfisme voorkom, soortgelyk aan alleel 2 uitdrukking.
Alhoewel die SLC11A1 geen voorheen geïmpliseer is in yster vervoer, is daar vir die
eerste keer aangetoon dat na yster lading, die verskillende allele differensiële
uitdrukking van die geen veroorsaak.
Verskeie siektes, insluitend slukderm kanker (OC), inflammatoriese dermsiekte (lBO)
en oorerflike hemochromatose (HH) (of primêre ysteroorlading), is ondersoek deur
middel van assosiasie studies. Betekenisvolle verskille (P<O.05) is waargeneem vir
alleel 3 tussen die kontrole- en pasiëntgroepe in al drie siektes wat ondersoek is,
maar slegs na stratifikasie volgens die teenwoordigheid van die -237C~ T
polimorfisme. Na hersiening van die promotor allele volgens ekspressie profiele
verkry met in vitro studies is statisties betekenisvolle assosiasie ook verkry vir alleel 3
met OC en primêre ysteroorlading in vergelyking met die onderskeie populasie
kontrolegroepe. Verder is verskeie nuwe variante ook geïdentifiseer in ekson 2
(112G~A, 148deIGACCAGCCC, 157insGACCAGCCCAG) en intron 1 (IVS1-
28C~ T) en 'n statisties betekenisvolle verhoogde frekwensie van variant IVS1-
28C~ T is waargeneem in pasiënte met OC in vergelyking met die populasie
kontrolegroep (P<O.05). Die belangrikheid van variasie in die promotor area van die SLC11A 1 geen as 'n modifiserende faktor in ysterverwante siektes, is verder
ondersteun deur die SLC11A 1 geen in individue met ysteroorlading in die
afwesigheid van homosigositeit vir die HFE C282Y mutasie te ondersoek. Ander
gene geassosieerd met yster homeostase, insluitend HFE, SLC11A3, HAMP and
DCYTB, is ondersoek in individue met soortgelyke seleksie kriteria en potensiële
siekte-verwante mutasies is geïdentifiseer in 11% Wit en 45% Swart Suid-Afrikaanse
pasiënte. Die moontlike belang van die SLC11A3 en DCYTB gene in ysteroorlading
in die Swart Suid-Afrikaanse populasie en die moontlike betrokkenheid van die
DCYTB geen in yster oorlading oor die algemeen, is vir die eerste keer aangetoon.
Hierdie studie dra by tot 'n beter insig in die funksie van die SLC11A 1 geen ten
opsigte van ystermetabolisme. Die betrokkenheid van SLC11A 1 in 'n reeks siekte
fenotipes, wat insluit kanker en inflammatoriese toestande wat verband kan hou met
'n yster wanbalans, kan moontlik verklaar word deur interaksie met eksterne faktore
soos infektiewe agente wat die sellulêre yster status kan beïnvloed. Ons bevindinge
verskaf beide in vivo en in vitro getuienis dat yster wanbalans, wat bemiddel word
deur alleliese effekte van SLC11A1, verantwoordelik mag wees vir vatbaarheid vir
infektiewe en outoimmune siekte toestande.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/53510 |
| Date | 12 1900 |
| Creators | Zaahl, Monique G. (Monique Glenda) |
| Contributors | Kotze, M. J., Warnich, L., Winter, T. A., Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences. |
| Publisher | Stellenbosch : Stellenbosch University |
| Source Sets | South African National ETD Portal |
| Language | en_ZA |
| Detected Language | Unknown |
| Type | Thesis |
| Format | 215 p. : ill. |
| Rights | Stellenbosch University |
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