Inhibition of HIV-1 binding with antisense oligonucleotides complementary to HLE mRNA. HIV-1 permissive clone 10 was transfected with antisense oligonucleotides complementary to the HLE start codon (▧), 3′ splice site of HLE exon 4 (□), or β-globin (▪). Homogeneous receptor density at equilibrium was achieved by exposing cells to α1PI by culturing in medium containing 10% FBS throughout transfection. (A) Cell-surface expression of HLE, CD4, and CXCR4 was determined by flow cytometric analysis 48 hours after transfection (Table 1). MFI of cells transfected with β-globin antisense was used to estimate baseline expression. Results are expressed as percent baseline expression and were calculated as 100 × (MFI after HLE antisense/MFI after β-globin antisense). Cell viability was determined prior to staining. (B) PCR amplification of HIV-1 minus-strand, strong-stop DNA using R/U5 primer pairs resulted in a minimally detected product in lysates from cells transfected with antisense complementary for the 3′ splice site of HLE exon 4 (lane 1), and slightly less product in lysates from cells transfected with antisense complementary to the HLE start codon (lane 2), and strongly detected product in lysates from cells transfected with negative control antisense oligomer complementary for β-globin (lane 3). Transfection and PCR amplification were repeated twice and representative data are presented.
