2010jassalmPhD.pdf (7.15 MB)
Establishment of a C1sA targeted mouse line and the differential expression of the complement serine protease C1sA and the related gene C1sB in mouse tissues
thesisposted on 2010-05-24, 12:59 authored by Meenu Jassal
The complement system bridges the innate and adaptive immune responses. Complement can be activated via three distinct pathways, the classical, lectin and alternative pathways, all of which play a crucial role in the identification and elimination of pathogenic microorganisms. The classical pathway (CP) is activated mainly by immune complexes (complexes of antigen and immunoglobulin). The lectin pathway (LP) is activated when a carbohydrate recognition complex and associated serine proteases bind to specific carbohydrate patterns on the surface of pathogens. In addition to providing protection from microbial infections, the LP has been shown to be involved in inducing post-ischemic inflammatory tissue loss (ischemia/ reperfusion injury), which significantly contributes to the pathology of ischemic diseases such as, myocardial, renal and GI ischemia. Separate mouse models of CP and LP deficiency are available, mice deficient in C1q (the CP recognition complex) and MASP-2 (a serine protease essential for LP activation), respectively. However, a model of combined CP and LP deficiency is not available. It has not been possible to generate mice deficient of both C1q and MASP-2, because the genes are located on the same chromosome in close proximity, and mice deficient in C4 (a constituent of both pathways) are not genuinely LP deficient, because the LP can activate C3 in the absence of C4. To establish a mouse strain deficient of both pathways, the gene for murine C1s was targeted the essential serine protease of the CP, with the aim of producing a C1s/MASP-2 double deficient mouse. A C1s deficient line would also be a valuable model to study C1q functional activity in the absence of CP activation. The mouse has two genes for C1s, C1sA and C1sB. The expression profiles for C1sA and C1sB in normal mouse tissue were assessed using qRT-PCR and in situ hybridisation, and confirmed that C1sA is widely expressed whereas C1sB expression is restricted to the testis. Thus C1sB is very unlikely to compensate for C1sA in a C1sA deficient mouse line. C1sA was targeted and the germ line transmission of the disrupted allele was confirmed. However, intercrossing of heterozygote mice did not result in the expected generation of homozygous C1sA deficient mice. Analysis of the offspring indicated that homozygous deficiency for C1sA appears to be lethal.
Supervisor(s)Schwaeble, Hans Wilhelm; Freestone, P.
Date of award2010-03-24
Awarding institutionUniversity of Leicester