Acidic pH Inhibits Non-MHC-Restricted Killer Cell Functions

Immunotherapeutic strategies in advanced stages of solid tumors have generally met with little success. Various mechanisms have been discussed permitting the escape of tumor cells from an effective antitumoral immune response. Solid tumors are known to develop regions with acidic interstitial pH. In a recent study performed in the human system, we were able to demonstrate that non-MHC-restricted cytotoxicity is inhibited by an acidic microenvironment. To get more insight into the mechanisms leading to this reduced cytotoxic activity, we have now investigated the influence of an acidic extracellular pH (pHe) on the killing process in detail. Unstimulated PBMC and LAK cells were used as effector cells. Both populations are able to kill tumor cells in a MHC-independent manner via perforin/granzymes or TNFα, whereas only IL-2-activated cells can use the killing pathway via Fas/FasL. We studied the influence of a declining pHe on the different killing pathways against TNFα-sensitive and -resistant, as well as Fas-positive and -negative, target cells. Experiments in the absence of extracellular Ca2+ were used to discriminate the Ca2+-dependent perforin-mediated killing. Here we show that the release of perforin/granzyme-containing granules, the secretion of TNFα, and also the cytotoxic action of Fas/FasL interaction or of membrane-bound TNFα were considerably inhibited by declining pHe. Furthermore, the secretion of the activating cytokine IFNγ, as well as the release of the down-regulating cytokines IL-10 and TGF-β1, was strictly influenced by surrounding pH. As a pHe of 5.8 resulted in a nearly complete loss of cytotoxic effector cell functions without affecting their viability, we investigated the influence of pHe on basic cellular functions, e.g., mitochondrial activity and regulation of intracellular pH. We found an increasing inhibition of both functions with declining pHe. Therefore, an acidic pHe obviously impairs fundamental cellular regulation, which finally prevents the killing process. In summary, our data show a strict pHe dependence of various killer cell functions. Thus, an acidic microenvironment within solid tumors may contribute to the observed immunosuppression in vivo, compromising antitumoral defense and immunotherapy in general, respectively.