Expression of Heat Shock Protein 72 in Peritoneal Leukocytes is Induced by Peritoneal Dialysis

Abstract

Background One of the main limitations of peritoneal dialysis (PD) is deterioration of functional and morphological characteristics of the peritoneum. This complication appears to be related to the low biocompatibility profile of PD fluids. Recently, induction of the heat shock protein (HSP) stress response was demonstrated in cultured human mesothelial cells exposed to PD fluid in vitro. We investigated whether expression of heat shock protein 72 (HSP-72) in peritoneal macrophages is induced upon exposure to PD fluid during continuous ambulatory PD. Methods Peritoneal leukocytes were isolated from 4-hour dwell dialysate; peripheral blood mononuclear cells (PBMC) and peripheral blood monocytes isolated from the same patients were used as a control. In separate experiments, PBMC from healthy individuals were exposed in vitro to different PD fluids or to culture media. Expression of HSP-72 was assessed by Western immunoblotting, flow cytometry, and reverse-transcription polymerase chain reaction analysis. Results Macrophages and leukocytes isolated from dialysis effluent expressed significantly increased HSP-72 and mRNA levels compared to blood monocytes and PBMC of the same patients. In vitro exposure of PBMC to fresh PD fluids resulted in significantly higher expression of HSP-72 compared to those incubated in culture medium. PBMC exposed in vitro to standard lactate-buffered dialysis fluids also expressed significantly more HSP-72 compared to cells exposed to bicarbonate/lactate-buffered fluids. Conclusion Our results indicate that exposure to PD fluids during dialysis triggers a shock response in peritoneal cells, which is manifested by significantly increased HSP-72 expression at both protein and mRNA levels. Analysis of this protein expression in peritoneal macrophages could be a new, convenient, and relevant way to assess the biocompatibility of PD fluids ex vivo.

The HSP72 stress response of monocytes from patients on haemodialysis is impaired

“…This is in agreement with our data, showing the same baseline amounts of HSP72 in Mphi of healthy rats and those with increased serum BUN (Figure 2), in urea-treated Mphi (Figure 6), as well as in patients and controls (Figures 4 and 5). Consistent with Marzec et al [45], monocytes from rats with renal insufficiency and from patients on IHD responded to HS but, in the case of HSP72, to a significantly lower extent than the ones from controls (Figures 2, 4, and 5). In contrast, skeletal muscle of chronic dialysis patients in the steady state contains increased amounts of protective HS proteins [51].…”

“…As diabetes-related stress and patient's metabolism considerably differ between diabetic and non-diabetic patients and the measurements were performed on EDTA-treated lymphocytes (leading via calcium efflux to pre-and de-activation of cellular pathways), the results may vary. Further, monocytes of patients on peritoneal dialysis exhibit significant HS response after contact with peritoneal dialysis fluid meaning that they are not overstimulated [45]. Unfortunately, this study lacks the healthy controls.…”

Addition of Alanyl-Glutamine to Dialysis Fluid Restores Peritoneal Cellular Stress Responses – A First-In-Man Trial

“…This calculation included an expected 10% drop-out rate and an assumed standard deviation of within-subject period differences of 50 percentage points. Predicted HSP expression levels were estimated from previous results in the ex-vivo setting [ 25 ] and from the literature [ 32 ]. For this reason, an interim analysis was planned to assess actual standard deviation of HSP expression after clinical evaluation for the first 20 patients, in order to re-calculate and to adjust final sample size.…”

“…The planned interims analysis revealed HSP expression to be quantifiable by 1D immunoblot under both PET conditions in only 4 of the 20 patients, secondary to peritoneal cell pellet lysate protein yields far lower than predicted in literature [ 32 ]. This outcome led to premature termination of patient recruitment and to the decision to analyze remaining portions of effluent cell protein samples by combining robust and highly sensitive saturation-labeling two-dimensional difference gel electrophoresis (2D-DIGE) with the specificity of 2-D Western blot, using saturation labeling (Saturn-2D Labeling Kit, NH DyeAGNOSTICS, Halle, Germany).…”

Heat shock proteins in chronic kidney disease

“…The investigations of HSP in peritoneal dialysis have mainly focused on the impact of the peritoneal fluid, as a stressor, on the function of the peritoneum. The induction of the Hsp72 expression has been confirmed in both mesothelial cells incubated with the dialysis fluid [ 31 ] and in macrophages from the dialysis effluent collected after a 4-h dwell [ 32 ]. In the meantime, the susceptibility of this protein to the fluid content has been proven, thus pointing to Hsp72 as a possible marker of the peritoneal fluid biocompatibility [ 32 , 33 ].…”