Primary Isolated Human Brain Microvascular Endothelial Cells Express Diverse HIV/SIV-Associated Chemokine Coreceptors and DC-SIGN and L-SIGN

Abstract

Chemokines have received increasing attention due to their inhibitory activities on human immunodeficiency virus type-1 (HIV-1) and simian immunodeficiency virus (SIV) replication and the potential for chemokine receptors to assist in HIV-1/SIV entry into permissive cells. Besides CD4, which is the major receptor for HIV-1 and SIV, a number of chemokine receptors including but not limited to APJ, CCR3, CXCR4, and CCR5 may be coreceptors for HIV-1/SIV, not only in peripheral blood and lymphoid tissues but also in the central nervous system (CNS). The present studies reveal the lack of CD4, but the significant expression of various chemokine receptors, APJ, CCR3, CXCR4, and CCR5, plus C-type lectins DC-SIGN and L-SIGN on isolated primary human brain microvascular endothelial cells (MVECs). As these MVECs do not express CD4, this suggests a CD4-independent HIV/SIV entry/infection of these cells, which are the major cells constituting the human blood-brain barrier. We also found that chemokines for cognate chemokine receptors individually were unable to block binding of HIV-1 to brain MVECs. These results reveal that in primary isolated brain MVECs viral attachment is mediated by a possible previously unknown receptor(s) or by cooperative activity of various receptors. Moreover, mRNA transcripts for DC-SIGN/L-SIGN, as well as DC-SIGN protein expression, suggest the capability of MVECs to attach viral particles on cell surfaces, even though polyclonal antisera for DC-SIGN did not affect viral binding to these cells. These data will assist in further understanding lentiviral entry into the CNS.

Contribution of Proteoglycans to Human Immunodeficiency Virus Type 1 Brain Invasion

“…We found that BMECs express high levels of HSPGs likely contributed by syndecan-2 and -4. As expected, we did not detect CD4 or GalCer and found that BMECs express low levels of CXCR4 and CCR5, as previously described (47,57). Importantly, we found that like HSPGs, CSPGs are also abundantly expressed on the surface of BMECs.…”

“…CXCR4 and CCR5 inhibitors did not block pNL-ADA or pNL-HXB attachment to BMECs (Fig. 6A), in agreement with previous work by Liu et al and Mukhtar et al, who showed that SDF-1␣, RANTES, and MIP-1␣ and ␤ do not prevent HIV-1 binding to BMECs (47,57). Together, these results indicate that HIV-1 uses receptors other than CXCR4 and CCR5 to initially attach to the surface of BMECs.…”

“…The microvessels were harvested, explanted into collagen type I-coated T-25 flasks, and propagated as monolayers. As a second source of BMECs, primary human BMECs were obtained from Cell System Corp. (ACBRI; Kirkland, Wash.) and cultured in human endothelial growth medium (Cell System Corp.) as described previously (8,56,57). Importantly, primary BMECs (mainly from brain cortex) obtained from Cell System Corp. were not passaged more than four times prior to use, to maintain the original features of primary BMECs.…”

Inhibition of HIV-1 fusion with small interfering RNAs targeting the chemokine coreceptor CXCR4

“…This unique coreceptor has been shown to bind and allow infection of cells by a number of HIV-1 strains. [61][62][63][64] Its presence in the central nervous system siRNA and CXCR4 N Zhou et al (CNS) is of potential importance, [64][65][66] and investigation of siRNAs' effects on this moiety is ongoing in our laboratories. CXCR4, as well, has been demonstrated on CNS-based cells.…”

“…CXCR4, as well, has been demonstrated on CNS-based cells. 66,67 It has been recently demonstrated that RNAi also functions in mammalian neurons in vitro. 68 As such, targeting CXCR4 and/or APJ in human CNS cells may be an important first step in targeting RNAi toward HIV-1-induced encephalopathy.…”