Fig 7. Gammaherpesvirus accelerates reactivation from latency and increased virus replication in physiological oxygen tensions.

MHV68 latently infected splenocytes from C57bl/6J mice (n = 3) were collected and processed on 16 dpi. (A) Limiting dilution assay was performed on 21% O₂ or 1% O₂ to determine frequency of viral reactivation in low oxygen levels. At 63.2% the frequency reactivating splenocytes (dotted line) normoxia reactivation was 1 in 42,743 and from physioxic conditions 1 in 29,498. Symbols represent the mean percentage of wells positive for virus detection +/- standard error of the mean. Curve fit line were derived from nonlinear regression analysis. Data represents results of one experiment, performed in triplicates, out of more than three independent experiments. (B-C) Different ratios of splenocytes to 10⁵ MEFs, (10⁴ Left, 10⁵ Center, 10⁶ Right) were plated and incubated at 21% O₂ or 1% O₂. Graphs represent one out of two experiments. (B) Supernatants were collected, and viral titers were quantified by plaque assay on days 4, 5 and 6 post infection. *, p<0.05. **, p<0.01. ****, p<0.001. Statistical significance was determined by Multiple student’s t-test (C) At 3 dpi, RNA was isolated from cell layer of co-cultures and RTA levels were measured by qRT-PCR. *, p<0.05. Statistical significance determined by Student’s t-test.