Abstract Body: Background: Lipoproteins are spherical particles that circulate in the blood and transport insoluble lipids and other cargos, including proteins, in the body. Proteomic studies have shown that some of the proteins associated with lipoproteins are involved in blood clotting. To understand the role of lipoproteins in blood clotting, we examined the association of lipoproteins with coagulation factors, starting with fibrinogen. Fibrinogen is of interest because it is produced exclusively by hepatocytes, where most lipoproteins are assembled, and plays a critical role in blood clotting.

Objective: To investigate the interactions of lipoproteins with the coagulation factor I, fibrinogen.

Methods: We used size-exclusion Fast Protein Liquid Chromatography (FPLC) to fractionate human or mouse plasma based on particle size. Total fibrinogen in all fractions were measured by ELISA. Human plasma was pooled from 10 healthy donors (adults aged under 40). Mouse plasma was pooled from C57BL/6 mice (7 mice, adults on normal chow diet).

Results: In pooled human plasma, fibrinogen, had a single major peak at elution volumes 24-27 mL, which are in the range of LDL fractions. Fibrinogen in pooled C57BL/6 mouse plasma fractions had a consistent peak in the range of LDL fractions (elution volume 24-28 mL). No fibrinogen was detected in other lipoprotein fractions nor free protein fractions. The highest fibrinogen concentration eluted at 26 mL in both human and mouse pooled plasma, 0.255 mg/mL and 0.360 mg/ml, respectively. Pooled human plasma was delipidated by either density gradient ultracentrifugation or a lipid removal agent, cleanascite, fibrinogen levels were negligible in de-lipidated plasma fractions.

Intravital imaging of laser-induced clot formation in mouse cremaster arterioles showed that LDL goes to the site of injury first and then fibrin accumulates subsequently and colocalizes with LDL. Immunofluorescence experiments showed abundant apoB intracellularly in mouse platelets, however, apoB did not colocalize with markers for the α- and lysosomal granules. Ongoing research continues to further investigate the localization and functional consequences in clotting.

Conclusions: These experiments indicate that fibrinogen is mostly found in the LDL fractions in both human and mouse plasma. These data fill gaps in our knowledge of the association of LDL fractions with fibrinogen and our understanding of how hemostasis and thrombosis is regulated in dyslipidemic conditions.