These studies clearly demonstrate that there is an association between CXCL13 expression and AIDS-NHL. First, in the serum studies, serum CXCL13 levels were significantly elevated prior to lymphoma diagnosis in the AIDS-NHL group, compared to the AIDS control group (). CXCL13 levels in the AIDS-NHL group were even more highly elevated when compared to the HIV(+)/non-AIDS and HIV-seronegative groups. These results suggest that assessment of CXCL13 levels could potentially be useful in the detection and diagnosis of AIDS-NHL. Elevated serum CXCL13 levels have recently been reported in other cancers, including B-cell chronic lymphocytic leukemia [46
]. Second, many primary AIDS-NHL specimens, of both the Burkitt and DLBCL subtypes, showed expression of CXCL13 ().Third, as noted in (Section 3
), the AIDS-Burkitt cell line, 2F7, expressed low levels of CXCL13 after it had been growing in culture for several months (not shown). Thus, these combined data indicate that CXCL13 is frequently expressed in AIDS-NHL. These results are not dissimilar to results obtained in studies on several types of B cell lymphoma not associated with HIV infection, including non-AIDS-related primary central nervous system lymphoma (PCNSL) [15
]. The frequent expression of CXCL13 in the AIDS-NHL tumor specimens (Section 3
, ) raises the possibility that the developing tumors themselves are a source of the elevated serum CXCL13 levels seen preceding diagnosis in the AIDS-NHL group (). If so, CXCL13 could potentially prove to be a useful biomarker for early detection of AIDS-NHL, for determining burden of disease, and for determining prognosis following chemotherapy.
The receptor for CXCL13, CXCR5, appears to be commonly expressed in AIDS-NHL, as well. In the immunohistochemistry studies, all AIDS-NHL specimens showed expression of CXCR5 (), as did both AIDS-NHL cell lines (). Additionally, the CXCR5 on the AIDS-NHL cell lines appeared to be functional, as both cell lines showed chemotaxis towards CXCL13 (). Again, these results are not dissimilar to other previously reported results for several types of B cell lymphoma not associated with HIV infection [13
]. The fact that the optimal amount of CXCL13 in chemotaxis studies on the R cell line (50
ng/mL; Section 3
) was lower than the optimal amount for the 2F7 cell line (100
ng/mL; Section 3
) is consistent with the results in , which indicate that the R cell line expresses higher levels of CXCR5 than the 2F7 cell line.
Although our studies show that both CXCL13 and CXCR5 are commonly expressed in AIDS-NHL, future studies will be needed to determine more exactly what roles these molecules may be playing in these cancers. Several mechanisms, which are not necessarily mutually exclusive, appear possible: first, it seems plausible that CXCR5 and CXCL13 could be playing a role in the initial development of AIDS-NHL. As noted in Section 1
, serum CXCL13 levels appear to be elevated during HIV infection, and at least some circulating B cells appear to (abnormally) express CXCL13, and to lose expression of CXCR5 [25
]. As CXCL13/CXCR5 are important in guiding recirculating B cells into B cell zones of secondary lymphoid tissues [26
], if expression of these molecules is disturbed, it is possible that B cells will not home normally to these tissues. This could lead to inappropriate homing to other tissues and/or inappropriate activation, promoting lymphomagenesis. In support of this possibility is the fact that a large proportion of AIDS-associated lymphomas are extranodal, in contrast to B cell lymphomas not associated with HIV infection [47
]. It could be that the continued expression of CXCL13/CXCR5 in developed AIDS-NHLs, as shown in the current studies, is a reflection of processes that occurred during lymphomagenesis. While these molecules might continue to have an effect on AIDS-NHL biology, it is also possible that their expression is merely an epiphenomenon that is reflective of the earlier role they played in the initial development of these tumors.
A second possibility is that the CXCL13 and/or CXCR5 could be directly promoting AIDS-NHL tumor growth, or performing some other tumor-promoting functions, such as inhibiting apoptosis or enhancing angiogenesis. Other chemokines, such as SDF-1, have been shown to directly induce tumor cell growth in non-AIDS-related cancers [11
], and it is possible that CXCL13 could be doing so, also. This promotion of cell growth could be coming from high levels of circulating CXCL13 produced elsewhere in the body, or from CXCL13 produced locally to the tumor, either by tumor cells themselves or by local nontumor cells. Our studies do suggest that autocrine interactions could be occurring in AIDS-NHL, as some tumor cells appeared to express both CXCR5 and CXCL13 in both the immunohistochemistry and AIDS-NHL cell line studies (Section 3
Third, it seems possible that CXCL13 could be directing movement and/or metastasis of AIDS-NHL cells in the body. As noted earlier (), both AIDS-NHL cell lines used in our studies demonstrated chemotaxis towards CXCL13. Although CXCL13 has often described as being produced in secondary lymphoid tissues, it is now clear that it can be produced in other tissues as well, including brain, liver, and lung, particularly in the context of inflammation [48
]. Perhaps CXL13 produced in such organs chemoattracts AIDS-NHL tumor cells to metastasize to these locations.
It should be noted that at least aspects of these potential mechanisms involving CXCR5 and CXCL13 could be operating in some non-AIDS-related B cell lymphomas, as well, given the similarities noted above between the expression/function of these molecules in AIDS-NHL in the current study, and their previously reported expression/function in several types of non-AIDS-related B cell lymphoma [13
]. However, it also seems possible that some of these mechanisms could prove to be unique to AIDS-NHL, given our previous finding that HIV infection is associated with an overproduction of CXCL13, even in subjects who do not develop lymphoma [25
]. Such an overproduction of CXCL13 may not necessarily be occurring prior to the initiation of tumor development in otherwise healthy subjects who develop B cell lymphoma outside of the context of HIV infection. Thus, further study of CXCR5 and CXCL13 expression and function in AIDS-NHL, and in non-AIDS-associated NHL, could ultimately reveal some unique differences between the pathogenesis of AIDS-related and non-AIDS-related B cell lymphomas.
In summary, we have demonstrated an association between expression of the chemokine, CXCL13, and its receptor, CXCR5, and AIDS-NHL. CXCL13 has potential as a biomarker for AIDS-NHL. Future studies will be needed to more fully determine the feasibility of using CXCL13 as a biomarker for AIDS-NHL, as well as to define the function of CXCL13 and CXCR5 in the pathogenesis of AIDS-NHL.