EFFECT OF BICARBONATE-BASED DIALYSIS SOLUTIONS ON INTRACELLULAR PH (PH(I)) AND TNF-ALPHA PRODUCTION BY PERITONEAL-MACROPHAGES

Objective: To compare the effect of Dianeal and two newly-formulated b icarbonate-based peritoneal solutions on intracellular pH (pH(i)), tum or necrosis factor-alpha (TNF alpha) mRNA level, and TNF alpha secreti on by peritoneal macrophages (PM Phi). Design and Measurements: Perito neal macrophages were isolated from dialysates collected after overnig ht dwells in peritonitis-free continuous ambulatory peritoneal dialysi s patients. Dialysis solutions contained 1.5% or 4.25% dextrose. HCO3 concentrations of bicarbonate(TB) and bicarbonate/lactate-buffered (TB L) solution were 38 mM and 25 mM, respectively. TBL also contained lac tate at a concentration of 15 mM. pCO(2) levels were 78 mmHg and 51 mm Hg, respectively. In all experiments pCO(3) was carefully maintained a t a stable level. The pH(i) was measured by spectrofluorometry in BCEC F-loaded PM Phi exposed to different dialysis solutions or Rank's bala nced salt solution. TNF alpha levels were measured by ELISA in the sup ernatant of lipopolysaccharide-(LPS) stimulated PM Phi after their inc ubation in different solutions for 15 and 30 minutes. TNF alpha mRNA w as measured by reverse transcriptase polymerase chain reaction (RT-PCR ) of total RNA extracted from LPS-stimulated PM Phi after their incuba tion in different solutions for 30 minutes. beta-actin mRNA was used a s the control. Results: Dianeal caused a profound drop in pH(i) to bel ow 6.2. Following an initial drop, pH(i) stabilized after 4 minutes at levels of 6.96 and 6.8 after incubation in TB and TBL, respectively. In comparison to the control solution, a fall of 11% and 21% in TNF al pha secretion was seen after incubation in TB for 15 and 30 minutes, r espectively, and 15% and 26% after incubation in TBL. Under identical conditions, Dianeal (Baxter, McGaw Park, IL, U.S.A.) caused 59% and >9 5% suppression of TNF alpha secretion. Accordingly, TNF alpha mRNA lev el in PM Phi was severely depressed by Dianeal but no detectable inhib ition was observed following incubation for 30 minutes in TB and TBL. When dextrose concentration in TB and TBL was increased from 1.5% to 4 .25%, TNF alpha secretion by PM Phi was not suppressed by more than 49 %, even after 30 minutes incubation. Moreover, suppression of TNF alph a mRNA levels could not be detected with TB or TBL even at high dextro se concentrations. Conclusions: In contrast to Dianeal, both bicarbona te-based solutions caused only a mild drop in pH(i) of PM Phi. We post ulate this effect to be responsible for the improved capacity of PM Ph i to secrete TNF alpha when incubated in bicarbonate-based solutions c ompared to Dianeal. Reflecting its known cytotoxicity, dextrose in hig h concentrations diminishes the protective effect of TB and TBL on imm une function of PM Phi. TBL is as effective as TB in preventing the de leterious effect of Dianeal on PM Phi function.