Interaction of Chlamydia trachomatis with Human Genital Epithelium in Culture

Abstract

Primary cultures of human endometrial and ectocervical epithelial cells were examined as a new model system to study genital infection by Chlamydia trachomatis. Initial studies demonstrated that these cells were indeed susceptible to chlamydial infection. Inocula, adjusted to produce inclusions in 50 to 80% of equivalent numbers of standard McCoy cells, resulted in infection rates of approximately 15 to 30% for the columnar cells of the endometrium and 5 to 10% for the squamous cells of the ectocervix. Exposure of cultures to DEAE-dextran and centrifugationassisted inoculation, manipulations reported to enhance infection of HeLa and McCoy cells, did not alter the number of inclusion-positive genital cells. Addition of cycloheximide to the postinoculation culture medium slightly increased numbers of inclusion-bearing cells while growth of genital cells in hormone-supplemented medium resulted in a variable effect on inclusion development and a significant reduction in the association of radiolabelled organisms with these cells. The basis for the different levels of infection in McCoy versus genital cell cultures was revealed by immunofluorescence analysis of chlamydia1 association with host cells immediately after inoculation. Chlamydiae failed to adhere to many cells in the genital cell cultures while adherence to McCoy cells was uniform. In addition, the association of radiolabelled C. trachomatis was significantly lower with genital cells than with McCoy cells. Finally, culture conditions were defined which markedly inhibited inclusion development without an immediate loss of chlamydial growth potential. This investigation indicates that primary genital cell cultures are susceptible to chlamydial infection and will be valuable for studies on the nature of C. trachomatis interactions with natural human target cells.

An in vitro Model of Chlamydia trachomatis Infection in the Regenerative Phase of the Human Endometrial Cycle

“…When outgrowths of glandular explants were either incubated with 25 pl inocula of 2 x lo5 McIFU for 2 h in a humidified chamber, or continuously incubated for 2 d with the same inoculum in 1 ml of medium, no inclusions developed. This result is at variance with that obtained by Moorman et al (1986), where centrifugation did not affect the number of inclusions that developed. However, their culture system involved the use of a different substratum (fibronectin) and a higher inoculum of Chlamydia1 injection of' human endometrium 208 1 organisms.…”

“…There was a striking dissimilarity between the two types of culture. As previously observed by Moorman et al (1986), chlamydial particles were evenly distributed over the McCoy cell monolayers, whereas in the endometrial cultures very few particles were located either on areas of confluent epithelial sheets or on the explants themselves, despite a uniform distribution of particles on the collagen-coated glass in cell-free areas. This raised the possibility that centrifugation presented chlamydiae to the cell by depositing them on the surrounding substratum, from which they were subsequently 'scavenged' during the course of radial migration.…”

Comparison of Chlamydia trachomatis serovar L2 growth in polarized genital epithelial cells grown in three-dimensional culture with non-polarized cells

“…It is interesting to note that, firstly, serovar L2 attachment to polarized epithelial cell monolayers on beads appeared evenly distributed (Fig. 1A and D) while serovar E EB attachment/entry was rather patchy and clustered throughout both HEC-1B and HeLa cell monolayers [13], a pattern reminiscent of chlamydial attachment to apical surfaces of polarized primary human and pig genital epithelial cells cultured ex vivo [15,18]. This intriguing difference between strains may possibly be due to the recognition of and binding to different receptor molecules on host cell surfaces that, for serovar E, may be enriched or clustered in some regions or microdomains of polarized epithelial cell apical membranes, such as lipid rafts or caveolae, proposed to be involved in serovar E but not in serovar L2 entry [19,20].…”

“…Crude chlamydial stocks were collected at 48 h post-infection (hpi) using a standard protocol [15], and were then resuspended in 2-SPG (0.2 M sucrose, 0.02 M phosphate buffer, and 5 mM L-glutamine), aliquoted and stored at −80 °C. Subsequently, the infectious titer of each chlamydial stock was determined in HEC-1B and HeLa cells grown in 24-well tissue culture plates, and expressed as a percent infectivity.…”

Quantification of Chlamydia trachomatis Elementary Bodies in Urine by Ligase Chain Reaction

“…Most assays rely on an indirect estimation of an infected individual's organism burden by a count of the number of chlamydial inclusion-forming units in a monolayer of tissue culture cells. These studies have been limited by the relative insensitivity of culture (9), variability of infectivity of host cells under different culture conditions (7,11), and an inability to adequately quantify a broad range of organism concentration.…”