Post-transcriptional effects of extracellular pH on tumour necrosis factor-α production in RAW 246.7 and J774 A.1 cells

Abstract

The present studies determined the effects of extracellular pH (pH(o)) on the production of tumour necrosis factor-alpha (TNF-alpha) in the macrophage-like cell lines RAW 246.7 and J774 A.1. The cells were activated with lipopolysaccharide (LPS) at pH(o) 5.5, 6.5 or 7.4. TNF-alpha gene transcription was monitored by Northern blot analysis. Synthesis of the cytokine was monitored by ELISA measurements of the TNF-alpha content of cell-conditioned media (extracellularly released TNF-alpha) and cell lysates (cytosolic TNF-alpha). The magnitude of the TNF-alpha response differed markedly between the two cell lines. RAW cells were more responsive to LPS than were J774 cells. However, the effects of pH(o) on TNF-alpha production were similar in the two cell lines. TNF-alpha gene transcription was insensitive to experimental pH(o). The pH(o) had no effect on the abundance of TNF-alpha mRNA at 2, 4 or 18 h. Nonetheless, synthesis of TNF-alpha was affected significantly by pH(o). The TNF-alpha contents of cell-conditioned medium and cell lysate at 18 h were reduced progressively at lower pH(o) values. The data indicate that pH(o) alters TNF-alpha production in RAW and J774 cells at a post-transcriptional level. These findings suggest that pH(o) influences the phenotypic responses of macrophages to activating stimuli and modifies the role that macrophages play in inflammatory and immune actions.

Carbon dioxide differentially affects the cytokine release of macrophage subpopulations exclusively via alteration of extracellular pH

“…Similar to West et al [20,21], Heming et al [8], and Kopernik et al [10], we observed a decreased release of IL-6 of NR 8383 cells incubated in a CO 2 atmosphere. These in vitro data fit well with the known reduction of circulating IL-6 levels in patients following laparoscopic surgery with CO 2 pneumoperitoneum compared to open surgery or laparoscopy with laparolifting techniques and room air contamination [16], as well as results of other reports that show a suppression of the inflammatory response of peritoneal macrophages following laparoscopy.…”

“…Moreover, they demonstrated that the effects of CO 2 could be mimicked via pharmacological modulation of the intracellular pH and thus concluded that the decrease in intracellular pH is essential for the effects of CO 2 . In contrast to the modulation of intracellular pH, Heming et al [7,8] investigated the impact of extracellular pH (pH 0 ) on the production and secretion of TNF-a by peritoneal and alveolar macrophage cultures. They demonstrated that the reduction of extracellular pH could significantly depress the release of TNF-a and simultaneously increase the accumulation of intracellular TNF-a in stimulated alveolar macrophages.…”

Effects of extracellular pH on tumour necrosis factor-α production by resident alveolar macrophages

“…No obvious trend with time was observed in the cytosolic TNF-α content of LPS-activated alveolar mφ at pH o 7.4 (see Figure 6). In contrast, previous studies by us [22] and others [34] found significant increases with time in the cytosolic TNF-α content of mφ-like cell lines ( J774 and RAW cells) following cell activation with LPS. The reason for this discrepancy is unclear, but cellspecific differences in the LPS-induced production of TNF-α have been documented previously [35].…”

“…In contrast, West and co-workers [20,21] reported that acidification of LPS-activated peritoneal mφ with CO # suppressed TNF-α secretion without altering TNF-α mRNA abundance. We recently documented similar effects (specifically, a decrement in LPS-induced TNF-α secretion without a change in transcript abundance) in mφ-like cell lines ( J774 A.1 and RAW 246.7 cells) incubated in acidic culture media [22]. In still other studies, incubation of peritoneal mφ in acidic dialysis fluids has been shown to decrease both LPS-induced TNF-α transcript abundance and TNF-α secretion [23].…”

Simvastatin inhibits acidic extracellular pH-activated, outward rectifying chloride currents in RAW264.7 monocytic-macrophage and human peripheral monocytes

“…Recently, it has been suggested that acidic microenvironments have a considerable effect on immune cells and their function [4,5]. Acidosis has been associated with proinflammatory effects on nuclear factor-κB (NF-κB) DNA binding levels and tumor necrosis factor (TNF) synthesis in macrophages [6,7]. Kos et al [8] found that a decrease in extracellular pH affected IL-6 release in macrophage subpopulations.…”