Relationship between in vitro human immunodeficiency virus type 1 replication rate and virus load in plasma

Abstract

Although plasma human immunodeficiency virus type 1 (HIV-1) RNA concentration is a major determinant of the rate of HIV-1 disease progression, the reasons for variability in plasma virus loads among infected individuals are not fully understood. We conducted investigations with 15 HIV-1-infected individuals who were not receiving antiretroviral therapy to evaluate the hypothesis that HIV-1 replication rate in vitro is a significant determinant of plasma virus load. Virus could not be isolated from one subject. Two subjects were excluded because they had features previously associated with distinct plasma virus loads and altered rates of disease progression; one harbored a syncytium-inducing virus and the second was heterozygous for a 32-bp deletion from the CCR5 gene. HIV-1 replication rates were determined by culturing autologous virus isolates in phytohemagglutinin-treated peripheral blood mononuclear cells (PBMC) and determining the rate of p24 antigen production during the logarithmic phase of viral replication. The contribution of HIV-1 reverse transcriptase (RT) and protease (PR) alleles to replication capacity was assessed using recombinant viruses in a single-cycle infection assay. HIV-1 replication rates ranged from 0.15 to 0.76 log(10) pg/ml/day and were reproducible within the same donor PBMC (coefficient of variation +/- 4%). RT-PR replication capacity ranged from 14 to 95% of that of control virus and was linearly related to replication rate (r(2) = 0.53; P = 0.007). Plasma HIV-1 RNA concentration was linearly related to replication rate (r(2) = 0.71; P < 0.001) and RT-PR replication capacity (r(2) = 0.44; P = 0.019). These data suggest that different RT-PR alleles are important determinants of HIV-1 replication rates and that HIV-1 replication rate explains much of the variability in plasma virus load in chronic HIV-1 infection.

FIG. 1.

Growth curves for an HIV-1 isolate in PHA lymphoblasts inoculated on day 3 (closed circles) and day 7 (open circles). Closed circles show the typical triphasic growth curve for an HIV-1 isolate after inoculation of lymphoblasts on day 3, with a lag phase (days 0 to 1), exponential phase (days 1 to 6) and plateau phase (days 6 to 10). Open circles show growth of the same HIV-1 isolate on PHA lymphoblasts inoculated on day 7. All data points are the mean ± range of two parallel cultures.

FIG. 2.

Measurement of HIV-1 replication rate. PHA lymphoblasts from a single donor were infected with NSI HIV-1 isolates from 12 different subjects on day 0. Data during the phase of exponential increase of supernatant p24 were fitted by linear regression. The coefficient of determination (r²) and slope (m) for each regression are shown. The slope (m) is the viral replication rate. All data points are the mean ± range of two parallel cultures. (A) Subject 1. (B) Subject 3. (C) Subject 4. (D) Subject 5. (E) Subject 10. (F) Subject 11. (G) Subject 13. (H) Subject 17. (I) Subject 20. (J) Subject 23. (K) Subject 25. (L) Subject 26.

FIG. 3.

HIV-1 replication rate in PHA lymphoblasts was linearly related to HIV-1 RT and PR replication capacity. HIV-1 replication rate is the slope (m) of the regressions in Fig. 2. RT and PR replication capacity were determined in a single cycle-based assay using recombinant virus that contained the RT and PR genes of each HIV-1 isolate. The replication capacity is the percentage of virus replication relative to the reference virus strain, NL4-3. Solid line indicates fit of data by linear regression (r² = 0.53; P = 0.007). Dashed lines indicate the 95% confidence interval for the regression.

FIG. 4.

HIV-1 replication rate in PHA lymphoblasts was linearly related to plasma HIV-1 RNA concentration. Plasma HIV-1 RNA values are from Table 1 and virus replication rate is the slope of the regressions in Fig. 2. Solid line indicates fit of data by linear regression (r² = 0.71; P < 0.001). Dashed lines indicate the 95% confidence interval for the regression.

FIG. 5.

HIV-1 RT and PR replication capacity was linearly related to plasma HIV-1 RNA concentration. RT and PR replication capacity was determined in a single cycle-based assay using recombinant virus as described in the legend to Fig. 3. Solid line indicates fit of data by linear regression (r² = 0.44; P = 0.019). Dashed lines indicate the 95% confidence interval for the regression.