Ex vivo culture of human colorectal tissue for the evaluation of candidate microbicides.
Fletcher, Patricia S a; Elliott, Julie b; Grivel, Jean-Charles c; Margolis, Leonid c; Anton, Peter b; McGowan, Ian b; Shattock, Robin J a
20(9):1237-1245, June 12, 2006.
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Objectives: Establishment of an in vitro model to evaluate rectal safety and the efficacy of microbicide candidates.
Design: An investigation and characterization of human colorectal explant culture for screening candidate microbicides to prevent rectal transmission of HIV-1 infection.
Methods: Human colorectal explants were cultured at the liquid-air interface on gelfoam rafts. Phenotypic characterization of HIV-1 target cells was performed by fluorescence-activated cell sorter analysis. HIV-1 infection was determined by the measurement of p24 antigen release, viral RNA, and proviral DNA accumulation.
Results: Colorectal explant CD4 T cells expressed higher CCR5 and CXCR4 levels compared with blood. Minor differences between the rectal and sigmoid colon were observed with a trend for slightly higher CCR5 and HLA-DR expression in cells from the sigmoid colon. Favourable culture conditions were established for colorectal tissue. Although tissue structure degenerated with time, CD4: CD8 cell ratios remained constant, and tissue supported productive HIV-1 infection. The ability of candidate microbicides to inhibit R5 HIV-1 infection was evaluated. Polyanion candidates, PRO2000 and dextrin sulphate, provided 99% protection at 1 [mu]g/ml and 1 mg/ml, respectively, equivalent to 1/5000 and 1/40 of the vaginal formulations. The nucleotide reverse transcriptase inhibitor (NRTI) 9-[2-(phosphonomethoxy)propyl]adenine (PMPA) provided protection at concentrations 1000-fold lower (10 [mu]g/ml) than the proposed vaginal formulation (1%). Furthermore, non-NRTI UC-781 and TMC-120 provided greater than 99% inhibition at 3.3 or 0.33 [mu]g/ml, respectively. No products demonstrated toxicity to rectal mucosa at inhibitory concentrations.
Conclusion: Colorectal explant culture was shown to be a useful tool for the preclinical evaluation of potential microbicides. The data suggest that rectally applied microbicides might provide protection from HIV-1 transmission.
(C) 2006 Lippincott Williams & Wilkins, Inc.