aUniversity Hospitals of Cleveland and Case Western Reserve University Center for AIDS Research, Cleveland, OH, USA; and bHuman Genome Sciences, Inc., Rockville, MD, USA.
Sponsorship: This work was partly supported by grants AI 38858 and AI 36219 from the National Institutes of Health.
This work was presented in part at the 9th Conference on Retroviruses and Opportunistic Infections. Seattle, WA, 24–28 February 2002 [Abstract 267-T].
W.F. and T.B. are employees of Human Genome Sciences, Inc., owner of the BLyS protein trademark.
Received: 20 September 2002; revised: 13 March 2003; accepted: 19 March 2003.
We measured the plasma levels of B-lymphocyte stimulator (BLyS) in 101 HIV-1-infected patients and 18 controls. BLyS levels were higher among HIV-positive patients [median 5.70 (3.90) versus 4.62 (1.04) ng/ml, P = 0.002], who had significantly higher BLyS and total serum globulin levels with decreasing CD4 cell counts. Moreover, BLyS levels increased exponentially below 100 CD4 cells/μl. BLyS and globulin levels increase as HIV disease progresses, suggesting a role for BLyS in the hypergammaglobulinemia of HIV infection.
HIV infection is often accompanied by polyclonal hypergammaglobulinemia [1–5], resulting from a state of generalized, non-specific B-cell activation [6–8]. The mechanism underlying this phenomenon has not been conclusively established. B-lymphocyte stimulator protein (BLyS) is a member of the tumor necrosis factor ligand superfamily that regulates the survival, proliferation and differentiation of B lymphocytes [9–13]. In vitro, BLyS induces B-cell activation and expansion [12,14]; in vivo, exogenous administration of BLyS to mice leads to the expansion of B-cell populations in lymphoid tissue and increased serum immunoglobulin levels [11,12]. In humans, BLyS levels are elevated in autoimmune disorders associated with hypergammaglobulinemia [15–18] and in follicular non-Hodgkin's lymphoma . Given the similarities between the biological actions of BLyS and the humoral immune derangements seen in HIV infection, we hypothesized that BLyS levels may be abnormal in HIV disease.
A total of 101 adult HIV-infected patients and 18 healthy, HIV-uninfected volunteers were included in the study. Demographic data, nadir CD4 T-cell counts, highest plasma HIV-RNA levels, history of antiretroviral treatment, and the CD4 T-cell count, plasma HIV-RNA level and serum globulin level values closest in time to the measurement of BLyS were recorded for HIV-infected patients. HIV-infected patients were subdivided into three groups according to their most recent CD4 T-cell count: group 1, greater than 500 cells/μl (n = 27); group 2, 201–500 cells/μl (n = 42) and group 3, 200 cells/μl or less (n = 32). BLyS levels were measured by enzyme-linked immunosorbent assay on stored plasma samples as previously described .
BLyS levels were significantly higher among HIV-infected patients than among controls [median (interquartile range; IQR), 5.70 (3.90) versus 4.62 (1.04) ng/ml, P = 0.002]. Ninety per cent of the controls had BLyS levels between 3.4 and 5.6 ng/ml. None of the controls had a BLyS level above 5.7 ng/ml, in agreement with previous reports, which have generally found BLyS levels below 10 ng/ml in normal individuals [15–17]. By comparison, 51 of the HIV-infected patients (50.5%) had levels above 5.7 ng/ml and 22 of them (21.8%) had levels above 10 ng/ml. BLyS levels in the subgroups of HIV-1-infected patients and the controls are depicted in Fig. 1a. There was a graded trend towards increasing BLyS levels with more advanced stages of HIV disease. Median (IQR) BLyS levels among uninfected controls and HIV-infected patients with over 500, 201–500 and fewer than 200 CD4 T cells/μl were 4.62 (1.04), 4.85 (2.97), 5.51 (2.20) and 8.28 (7.43) ng/ml, respectively (P < 0.001). The corresponding serum globulin levels among HIV-infected patients were 3.3 (0.4), 3.6 (1) and 3.8 (6.8) g/dl (P = 0.026), paralleling the trend observed in BLyS levels (Fig. 1b); however, the linear correlation between the two did not reach statistical significance. There was a significant but non-linear correlation between BLyS levels and CD4 T-cell counts, with BLyS levels remaining relatively stable at higher CD4 T-cell counts and increasing exponentially at the lower extreme of the CD4 T-cell count spectrum. There was a weak, but statistically significant direct linear correlation between plasma HIV-RNA and BLyS levels.
The elevation of BLyS levels in our HIV-infected patients is consistent with findings in a previous report on the production of autoantibodies at different stages of HIV infection . Although both plasma BLyS and serum globulin levels proved to be strongly associated with the stage of HIV disease in our cohort, we were unable to confirm a direct correlation between them. This does not exclude the possibility that an underlying correlation between BLyS and immunoglobulin levels might have been obscured in this analysis by other serum globulins or by the effect of regulatory signals other than BLyS on immunoglobulin production. In a previous report on a similar cohort of HIV-infected patients , no significant association could be demonstrated between BLyS and IgG levels regardless of the CD4 T-cell count. In patients with autoimmune disorders, a weak association between BLyS and immunoglobulin levels has been found by some investigators , but not others .
This study does not allow us to determine the mechanism of BLyS level elevation in HIV infection. The cells that primarily express BLyS, including dendritic cells, monocytes and macrophages [9,10,12,20], are all targets for HIV, suggesting that HIV might directly drive the overexpression of BLyS by these cells. Alternatively, the expression of BLyS may be upregulated as a homeostatic response to lymphopenia or by soluble factors that are, in turn, elevated in HIV infection. Both IFN-γ and IL-10 upregulate BLyS expression and secretion [12,21]. Because its production is impaired in the more advanced stages of HIV disease , IFN-γ is an unlikely mediator of the increased levels of BLyS seen in HIV-infected patients. IL-10 levels, on the other hand, are often elevated in HIV infection [23–25], and are inversely correlated with CD4 T-cell counts . Moreover, HIV Nef protein directly induces IL-10 expression and production in vitro , and effective antiretroviral treatment leads to a rapid decline in IL-10 levels . One could thus speculate that in HIV infection, increased IL-10 production might result in the upregulation of BLyS with subsequent B-cell activation and hypergammaglobulinemia.
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