Background: Periodontitis is an inflammatory condition rendering in degradation of tooth supporting tissue. In the inflammatory process cytokines, amongst others TNF-a, IL-1b and IL-6 play an important role in regulating the immune response. Periodontal Ehlers Danlos syndrome (pEDS) is a rare connective tissue disorder characterized by distinct oral manifestations such as periodontitis. Studies has shown that individuals with pEDS exhibit a mutation in the complement factors C1r and C1s. Aim: The aim of this study was to analyze the gene expression of C1r and C1s in human gingival fibroblasts and to investigate if the expression of these genes’ changes during inflammatory conditions. Methods: Human gingival fibroblasts (hGFBs) were cultured with TNF-a or IL-1b for 24h. RNA isolation, extraction and complementary DNA synthesis was made at the timepoint of 6h and 24h. An qRT-PCR analyses of the genes C1r, C1s and IL-6 were performed and the relative gene expression of C1r, C1s and IL-6 were calculated. Results: The expression of C1r and C1s increased after 24h in TNF-a and IL-1b compared to control by measuring the relative fold change of the genes by culturing fibroblast over time. The result was taken from three parallel samples and the result was significant. Conclusion: The findings point to a link between the inflammatory classical complement pathway and the connective tissue homeostasis. Hopefully, our findings will be one piece of the puzzle regarding the molecular events rendering in pEDS and thereby a potential treatment target.

<p>Haptoglobin (Hp) is a serum protein known for its ability to form a tight complex with hemoglobin (Hb) and thereby inhibiting the oxidative activity of Hb. </p><p>Mammalian Hp is synthesized as a precursor (proHp) that undergoes proteolytic cleavage by a previously unidentified enzyme in the endoplasmic reticulum (ER). In this study, a proHp-cleaving enzyme was isolated from human serum and identified as complement C1r-like protein (C1rLP). Co-expression of C1rLP with proHp in mammalian cells resulted in cleavage of the latter protein in the ER. Mutation of either the active site serine residue in C1rLP or the arginine residue in the cleavage site of Hp abolished the cleavage of proHp by C1rLP. RNAi studies in mammalian cells identified the proHp-cleaving enzyme as C1rLP.</p><p>Hp has been found in all mammals studied to date but its presence in non-mammalian species has not been unambiguously shown. By searching currently available genomic DNA and cDNA sequence databases, a gene orthologous to mammalian <i>Hp</i> was found in bony fish. Hp-like protein expressed from this gene was demonstrated to be a major Hb protein in fish serum. Surprisingly, no Hp-like gene was found in the genomes of either frog or chicken. In chicken, a protein previously described as Hp was identified as PIT54, a member of a scavenger receptor cysteine-rich family of proteins. Interestingly, ostrich serum seemed to contain two Hb-binding proteins; one similar to PIT54 and one to mammalian Hp. We are not aware of any other case where the function of one gene has been taken over by another, completely unrelated gene</p><p>Fish Hp (fHp) is composed of a serine proteinase-related domain preceded by an extension consisting of several aminoa acids and a signal peptide. The extension contains a consensus motif for cleavage by subtilisin-like proprotein convertases (SPCs). fHp was found to be cleaved by SPCs in the Golgi complex.</p><p>Collectively, this thesis presents evidence that Hp has undergone significant changes during evolution with respect to its molecular organization and to the mechanism of its proteolytic cleavage.</p>

Haptoglobin (Hp) is a serum protein known for its ability to form a tight complex with hemoglobin (Hb) and thereby inhibiting the oxidative activity of Hb. Mammalian Hp is synthesized as a precursor (proHp) that undergoes proteolytic cleavage by a previously unidentified enzyme in the endoplasmic reticulum (ER). In this study, a proHp-cleaving enzyme was isolated from human serum and identified as complement C1r-like protein (C1rLP). Co-expression of C1rLP with proHp in mammalian cells resulted in cleavage of the latter protein in the ER. Mutation of either the active site serine residue in C1rLP or the arginine residue in the cleavage site of Hp abolished the cleavage of proHp by C1rLP. RNAi studies in mammalian cells identified the proHp-cleaving enzyme as C1rLP. Hp has been found in all mammals studied to date but its presence in non-mammalian species has not been unambiguously shown. By searching currently available genomic DNA and cDNA sequence databases, a gene orthologous to mammalian Hp was found in bony fish. Hp-like protein expressed from this gene was demonstrated to be a major Hb protein in fish serum. Surprisingly, no Hp-like gene was found in the genomes of either frog or chicken. In chicken, a protein previously described as Hp was identified as PIT54, a member of a scavenger receptor cysteine-rich family of proteins. Interestingly, ostrich serum seemed to contain two Hb-binding proteins; one similar to PIT54 and one to mammalian Hp. We are not aware of any other case where the function of one gene has been taken over by another, completely unrelated gene Fish Hp (fHp) is composed of a serine proteinase-related domain preceded by an extension consisting of several aminoa acids and a signal peptide. The extension contains a consensus motif for cleavage by subtilisin-like proprotein convertases (SPCs). fHp was found to be cleaved by SPCs in the Golgi complex. Collectively, this thesis presents evidence that Hp has undergone significant changes during evolution with respect to its molecular organization and to the mechanism of its proteolytic cleavage.