Ex vivo culture of human colorectal tissue for the evaluation of candidate microbicides.
OBJECTIVE
Establishment of an in vitro model to evaluate rectal safety and the efficacy of microbicide candidates.


METHODS
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 microg/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 microg/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 microg/ml, respectively. No products demonstrated toxicity to rectal mucosa at inhibitory concentrations.


CONCLUSIONS
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.
Fletcher
Patricia S
PS
St George's, University of London, Cranmer Terrace, Tooting, London SW17 0RE, UK.
Elliott
Julie
J
Grivel
Jean-Charles
JC
Margolis
Leonid
L
Anton
Peter
P
McGowan
Ian
I
Shattock
Robin J
RJ
D000287:Administration, Topical; D000998:Antiviral Agents / Q000008:administration & dosage / Q000627:therapeutic use*; D002469:Cell Separation; D003106:Colon / Q000821:virology*; D004341:Drug Evaluation / Q000379:methods*; D005434:Flow Cytometry; D005782:Gels; D015658:HIV Infections / Q000188:drug therapy* / Q000276:immunology; D015497:HIV-1; D006801:Humans; D007962:Leukocytes / Q000276:immunology; D010641:Phenotype; D012007:Rectum / Q000821:virology*; D046509:Tissue Culture Techniques; D014779:Virus Replication
D000998:Antiviral Agents; D005782:Gels