AcLDLAcetylated Human Low Density Lipoprotein
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The acLDL uptake of mvECs was analyzed to evaluate EC functionality.
After 14 days in indirect co-culture with diffASCs, mvECs in CoM and ECM again orientated themselves in even monolayers, expressed CD31 and vWF throughout and were able to take up acLDL (Fig.
Bind to receptor modified low density lipoproteins (LDL) by oxidation (oxLDL) or acetylation (acLDL).
When discovered, the first function described for SR-A was to provide adhesiveness to monocytes and macrophages to glycated collagen-IV-coated surfaces, and to mediate the endocytosis of acLDL [95].
It is important that in contrast to most SRs, SR-A expression is not downregulated by chronic exposure to endogen ligands such as acLDL. On the opposite, SR-A expression can be reversibly increased by incubating macrophages with SR-A ligands [108].
In both groups, the number of clusters dissipated within 7 days of culture, giving rise to spindle-shaped cells (Figure 4A), which stained positive for Sca-1 and Flk-1, as well as for acLDL and UE-lectin (Figure 4C).
Other modified forms of LDL, such as oxLDL, acLDL, and gLDL, were used for antibody testing.
Cross-reactivity of anti-CLDL antibody to gLDL, acLDL, and oxLDL, evaluated by indirect ELISA, was extremely low (ratios, 380 for CLDL/gLDL, 810 for CLDL/acLDL, and 2630 for CLDL/ oxLDL).
The medium was changed on days 3 and 5, and on day 6 BMM were treated for 24 hr with RPMI 1640 containing 1% FBS, 1% P/S, and 25 [micro]g/mL acLDL. On day 7 cells were equilibrated for 2 hr in RPMI-1640 containing 0.2% fatty acid-free BSA, dislodged from the tissue culture flask with nonenzymatic cell dissociation buffer (Sigma), centrifuged, and suspended in MEM prior to intraperitoneal injection of 2.5 x [10.sup.6] cells/0.5 mL.
Briefly, J774 cells (~60% confluent) in 12-well plates were labeled for 48 hr with 0.2 [micro]Ci/mL [[sup.3]H]cholesterol (35-50 Ci/mmol, Amersham Biosciences) in RPMI-1640 supplemented with 10% heat-inactivated FBS, 50 [micro]g/mL gentamicin, and 50 [micro]g/mL acLDL. Cells were then washed three times with PBS containing 1 mg/mL BSA (PBS-BSA) and equilibrated overnight in RPMI-1640 containing 0.2% fatty acid-free BSA (RPMI-BSA).
In our system, we have found that about one quarter of all differentially expressed genes are of this type, i.e., they are differentially expressed from cell preparation to preparation, but not in response to the manipulation under investigation (in our case, loading of the cells with AcLDL or OxLDL).
These ECs have "spindle-shaped" morphology and were characterized by various markers, an ability to uptake acLDL and an ability to bind UEA-1.