Expression of ATP binding cassette-transporter ABCG1 prevents cell death by transporting cytotoxic 7beta-hydroxycholesterol.

Oxysterols result from cholesterol by enzymatic or oxidative processes. Some exert cytotoxic effects leading to necrosis or apoptosis. Detoxification of these compounds mainly occurs in the liver and requires transport from peripheral tissues towards it. Some ATP-binding cassette transporters are involved in export of cytotoxic compounds. In the current study, we investigated whether ABC transporter family member G1 (ABCG1) may be involved in oxysterol transport, since its gene expression is highly responsive to oxysterol loading. TetOff HeLa cells stably expressing ABCG1 showed decreased mass uptake of 7beta-hydroxycholesterol (7beta-HC) whereas that of other physiologically relevant oxysterols was unaffected. Application of 7beta-HC to ABCG1 expressing cells induced hyperpolarization of mitochondrial membrane potential and production of reactive oxygen species, indicating energy consumption by the ATP-binding cassette transporter when it is activated by its correct substrate. Our study points to detoxification as one of potential cellular functions of ABCG1. We assume that ABCG1 protects against 7beta-HC-induced cell death, an important role in prevention of neurodegenerative and cardiovascular disease.

Expression and functional characterization of ABCG1 splice variant ABCG1(666).

The ATP-binding cassette transporter ABCG1 mediates the transport of excess cholesterol from macrophages and other cell types to high density lipoprotein (HDL) but not to lipid-depleted apolipoprotein AI. Several splice variants which may have different functions have been identified in mammals. In the current study, we characterized the human splice variant ABCG1(666), which differs from full-length ABCG1(678) by absence of an internal segment of 12 amino acids (VKQTKRLKGLRK). Accordingly spliced ABCG1 transcripts were detected in macrophages and liver in approximately twofold higher amounts than the alternatively spliced ABCG1 form encoding full-length ABCG1. We used transient and stable expression of ABCG1(666) fusion proteins to characterize glycosylation, subcellular localization, molecular interaction and functions of this ABCG1 variant. It could be demonstrated that ABCG1(666) is located at the cell surface and has the ability to form cholesterol transport competent homodimers which affect cellular cholesterol export in a similar manner as previously characterized forms of ABCG1. Our results support that ABCG1(666) may in fact be the most prominent form of functional ABCG1 expressed in the human.

Induction of an anti-inflammatory human monocyte subtype is a unique property of glucocorticoids, but can be modified by IL-6 and IL-10.

Glucocorticoids (GC) are the most widely used immunosuppressive agents in clinical medicine. Recently we showed that GC enhance survival of human monocytes and induce a specific anti-inflammatory monocyte subtype which actively induces resolution of inflammation. We now investigated if cytokines IL-4, IL-6 and IL-10, which, like GC, have mostly anti-inflammatory effects on macrophages, would have GC-like effects also on monocytes. Human monocytes were stimulated with either cytokine, GC or combination thereof, and resulting effects on apoptosis, adherence, migration, phagocytosis, ROS production and cell surface phenotype were determined. We found that IL-4, IL-6, and IL-10 had either less or different effects on various anti-inflammatory functions of monocytes compared to GC. As such, IL-4 and IL-6 alone did not delay apoptosis while IL-10 even enhanced it. However, IL-6 or IL-10 increased GC-mediated protection from apoptosis when applied together with GC. Thus, the potential of GC to induce anti-inflammatory human monocytes is unique and not mimicked by the investigated cytokines. However, IL-6 and IL-10 amplify GC-induced anti-inflammatory and pro-resolution mechanisms by enhancing survival of GC-induced monocytes and thus sustaining their function. This combined effect of GC and cytokines could be important for the physiological switch from amplification towards resolution phase of inflammation.