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Ubrzani razvoj analitičkih tehnika u posljednjih nekoliko desetljeća omogućio je temeljito istraživanje procesa N-glikozilacije u brojnim patofioziološkim stanjima, a uočene promjene u mnogima od njih dovele su do strelovitog rasta interesa za glikobiologiju. Promjene u N-glikozilaciji danas se smatraju vrijednim biljezima raznih bolesti, te se njihova specifičnost pokušava iskoristiti u diferencijalne, dijagnostičke i prognostičke svrhe. Prethodne studije pokazale su da je N-glikanski profil plazmatskih proteina promijenjen u šećernoj bolesti tipa 1 (ŠBT1). Jedna od izraženije uočenih promjena bila je ona u zastupljenosti visoko-manoznih glikanskih struktura u plazmatskom N-glikomu. Ove strukture dominantno bi mogle potjecati s komponente komplementa C3, budući da je to glikoprotein s isključivo visoko-manoznim glikanima vezanima na proteinsku okosnicu za kojeg je poznato da doprinosi razvoju ŠBT1 pojačavanjem autoimunih upalnih procesa. Iz tog je razloga u ovom radu razvijena visokoprotočna metoda za analizu N-glikozilacije pojedinih glikozilacijskih mjesta ljudskog C3 proteina upotrebom tekućinske kromatografije spregnute sa spektrometrom masa (LC-MS) temeljena na njegovu obogaćivanju iz plazme pomoću lektinskog afinitetnog medija visokog afiniteta za manozu. Novorazvijenom metodom zatim je analizirana krvna plazma ispitanika novodijagnosticiranih šećernom bolesti tipa 1 i njihove zdrave braće i sestara te su uočene značajne promjene C3 N-glikoma. ŠBT1 povezana je s porastom manje procesuiranih struktura s više manoznih podjedinica na oba N-glikozilacijska mjesta. Nadalje, C3 N-glikozilacija je analizirana u odrasloj populaciji ispitanika s različitim stupnjem najčešćih komplikacija ŠBT1 – retinopatije i albuminurije. Pokazano je da se C3 N-glikom značajno mijenja u teškoj albuminuriji u ŠBT1, no da je neovisan o trajanju bolesti. Retinopatija je pak dovela do promjene samo jedne glikoforme, dok su svi osim jednog C3 glikopeptida značajno povezani s razinama hemoglobina A1c (HbA1c). Predstavljene spoznaje ukazuju na uključenost N-glikozilacije i C3 komponente komplementa u patofiziologiju šećerne bolesti tipa 1 te upućuju na značaj C3 N-glikoma kao potencijalnog dijagnostičkog i prognostičkog biljega ove bolesti i njoj pridruženih komplikacija. Rapid development of analytical techniques in the last few decades has enabled a thorough investigation of the N-glycosylation process in numerous pathophysiological conditions, and the observed changes in many of them have led to a rapid growth of interest in the field of glycobiology. Changes in N-glycosylation are now considered valuable biomarkers of various diseases, and their specificity is being exploited for differential, diagnostic and prognostic purposes. Previous studies have shown that the N-glycan profile of plasma proteins is altered in type 1 diabetes (T1D). One of the more pronounced changes observed was in he abundance of high-mannose glycans in the plasma N-glycome. These structures could predominantly originate from the complement component C3, a glycoprotein with exclusively high-mannose glycans attached to its protein backbone, which is known to contribute to the development of T1D by enhancing autoimmune inflammatory processes. For this reason, a high-throughput method for the site-specific N-glycosylation analysis of human C3 protein was developed in this study using liquid chromatography coupled to a mass spectrometer (LC-MS), based on its enrichment from plasma using lectin affinity matrix. The newly developed method was then used to analyze blood plasma of subjects newly diagnosed with type 1 diabetes and their healthy siblings, and significant changes in C3 N-glycome were observed. T1D was associated with an increase in less processed structures with more mannose subunits at both C3 N-glycosylation sites. Furthermore, C3 N-glycosylation was analyzed in the adult population of T1D subjects with different degrees of the most common complications of T1D – retinopathy and albuminuria. It was shown that C3 N-glycome significantly changes in severe albuminuria in T1D, but is independent of disease duration. Retinopathy led to a change in only one glycoform, while all but one C3 glycopeptide was significantly associated with levels of hemoglobin A1c (HbA1c). The findings presented indicate the involvement of N-glycosylation and the C3 complement component in the pathophysiology of type 1 diabetes and indicate the importance of C3 N-glycome as a potential diagnostic and prognostic biomarker of this disease and its associated complications.

The aim of the present study was to compare the frequencies of the F allele of C3 complement component and the Leiden mutation of coagulation factor V in patients with severe coronary heart disease (CHD) who survived myocardial infarction (MI; group A), and those who had no MI in their case history (group B). We have determined the C3 allele frequencies by electrophoresis, and Leiden mutation by PCR in 338 patients with severe CHD and in 490 and 523 healthy controls, respectively. The C3*F allele frequency was significantly (p = 0.006) higher in group A (0.213) that in group B (0.132). A significant (p = 0.045) difference was found between ≤60-year group A (0.077) and group B (0.029) patients in the frequency of Leiden mutation. These findings indicate that the C3*F allele and the Leiden mutation may be associated with an increased risk of developing myocardial infarction in CHD patients.

Several phenolic constituents of propolis and their synthetic analogs were derivatized with ʟ-lysine. The ability of these complexes to alter complement activity was estimated in vitro in human serum. The influence of selected complexes on C3 hemolytic activity via classical pathway (CP) and alternative pathway (AP) and on zymosan-induced AP activation was determined. The results suppose that the anticomplement effect of the complexes might be related to the interaction with C3 complement component.

In order to determine the allele and genotype frequencies for the polymorphism of the gene coding the C3 component of the complement system, 432 sheep from nine breeds of different productive types were studied. Tests were carried out with Mouton Charollais, Ile de France, Trakia Merino breeds, milk crosses [Stara Zagora×East Friesian and (Stara Zagora × East Friesian) × Pleven Blackhead], White Maritsa, Patch-faced Maritsa, Karnobat, Pleven Blackhead and Stara Zagora breeds, with 48 animals studied from each breed. The animals were reared at the Agricultural Institute – Stara Zagora, the Agricultural Institute – Karnobat and private farms. Seven alleles were discovered (S, S1, S5, S7, S10, F, and F1), forming nine genotypes (SS, SS1, FS, FS5, FS7, FS10, FF, FF1, F1F1). The S allele frequency was the highest in sheep from the Stara Zagora and Trakia Merino breeds. In all other breeds, most common was the F allele, with this tendency being most evident in the Mouton Charollais and Pleven Blackhead breeds. The very rare S1 allele was found in three breeds with a frequency of 0.01 to 0.05. Another rare allele discovered over the course of his study was F1, with frequency varying between 0.2 and 0.13. The frequencies of the nine established genotypes (SS, SS1, FS, FS5, FS7, FS10, FF, FF1, F1F1) varied in each breed. The SS genotype was most commonly encountered in the Stara Zagora breed (0.42), the heterozygous FS genotype was most common in the Mouton Charollais breed and milk crosses (0.23), whereas the animals homozygous for the F allele were prevalent among the Pleven Blackhead breed (0.54). The heterozygous FS5 genotype was also notable, having a frequency of 0.33 in the Karnobat breed.

Sensitivity to cow's whole milk investigated in six patients with the idiopathic nephrotic and an elemental diet was given. After proteinuria had decreased to less than or equal to 500 mg/24 hours on consecutive days patients were challenged with cow's milk. This resulted in the return of significant proteinuria, oedema, and decreased urine volume together with a decrease in serum-IgG concentrations in four patients. Acute alteration of plasma C3 complement component accompanied milk challenge in all 6 patients. There were no consistent alterations in other immunoglobulin concentrations. Intradermal skin testing with cow's milk extract was positive in all patients. These results suggest that in some individuals clinical and biochemical manifestations of the nephrotic syndrome of childhood may be related to hypersensitivity of food products.

The serum concentrations of circulating immune complexes, immunoglobulins G and M, and the C3 complement component of thirty patients with one to three large periapical lesions were measured and compared with those of patients with no periapical lesions. The results indicate that there is no statistical difference between the two groups, and it appears that chronic periapical lesions cannot act as a focus to cause systemic diseases via immune complexes.

Population and family data collected at the Reference Laboratory for the polymorphism of the C3 complement component of man support the view that there is very little, if any, gene frequency variation among Caucasian populations. The observed segregation ratios are in close agreement with the expected ratios. Three ‘new’ variants are reported in 1973/74: ‘F 0.9’, ‘S 1.0’ and ‘S 1.6’. Since rare variants cannot be designated in terms of absolute mobilities, the recommendation of the First International Symposium and Workshop at Bonn in 1972 is renewed to compare ‘new’ rare variants with reference standard sera in the Reference Laboratory. The common C3 polymorphism is also detected in the C3c conversion product as revealed by the Pt typing technique. Rare variants, however, are not reliably determined in this method.

Serum C3 and C4 levels have been determined in patients with Addisonian pernicious anaemia (PA) and megaloblastic anaemia due to vitamin B12 deficiency from other causes, before and after treatment, in order to study the interaction between vitamin B12 deficiency and complement and the role of complement in the pathogenesis of the gastric lesion of PA. C3 levels are significantly reduced in vitamin B12 deficiency and return to normal on treatment; C3 levels correlate with the degree of anaemia but not with serum vitamin B12 levels at diagnosis. C4 levels are normal. These observations suggest that the observed C3 hypocomplementaemia is not a consequence of immune mechanisms, but may be due to altered synthesis of C3 complement component.

Immunopathologic cutaneous lesions resembling human systemic lupus erythematosus (SLE) can be induced in mice sensitized to ultraviolet (UV)-irradiated DNA following whole body irradiation with UV light. The lesions are characterized by the formation of immune complexes at different skin sites. The role played by cellular and humoral mediators in the pathogenesis of this experimental model was investigated. The results obtained suggest that inflammation that follows UV radiation is the major factor responsible for this pathology. Accordingly mice that were rendered neutrophil (PMN) deficient did not manifest skin lesions, and depletion of C3 complement component left them unchanged. In addition time course studies showed that PMN depletion did not prevent a delayed skin involvement. Thus multiple factors seem to mediate the onset of the immunopathologic changes previously described.

The serum concentrations of circulating immune complexes, immunoglobulins G, M, and E, and the C3 complement component of 35 patients with acute apical abscesses were measured. These levels were compared with those of people without acute apical abscesses. Statistically significant differences between the levels of immune complexes, immunoglobulins G and M, and C3 complement component were found between the two groups. In addition, significant differences were also noted in eight patients in the mean levels of concentration of immune complexes, immunoglobulins G, M, and E, and C3 complement component before and after root canal therapy or extraction of involved teeth. It appeared that acute periapical lesions may lead to measurable systemic immunological reactions, but the clinical significance of these changes remains unclear.