Low Glucose Degradation Product Content Research Articles

#6062 EFFECT OF KIDNEY TRANSPLANTATION ON ARTERIOLAR MOLECULAR PATHWAYS IN CHILDREN WITH PRECEDING PERITONEAL DIALYSIS

Abstract Background and Aims Patients on peritoneal dialysis suffer from progressive peritoneal membrane transformation and accelerated vascular disease, underlying molecular machineries have recently been described. Molecular effects of kidney transplantation (KTx) on vascular and peritoneal pathophysiology following PD are hardly understood. Method Fat surrounded omental arterioles (i.e. not directly exposed to PD fluids) from age-matched (median 6,5 years) children with chronic kidney disease (CKD5, n = 8), 5 children on PD [neutral pH, low glucose degradation product content, median PD duration 22 (5,50) months] and 6 children, who underwent successful KTx 4-5 weeks after a median 22 (12, 36) months of PD were analyzed by digital histomorphometry. Following microdissection from fat tissue, arterioles underwent transcriptome and proteome analyses, followed by Gene Set Enrichment Analysis (GSEA), weighted gene co-expression network analysis (WGCNA), gene ontology (GO) and protein interaction analysis (string-db.org). Key pathways were validated in independent, matched cohorts by quantitative immunohistochemistry (n = 15/group) and in vitro. Results 5000 most variable arteriolar transcripts (cut off p-value<0.05 and |r| >0.5) were grouped into 23 modules by WGCNA, of which 8 significantly differed between CKD5, PD and KTx. Seven were specifically related to KTx and one module to PD. Four out of seven KTx specific modules resulted in significant GO terms summarized as fatty acid biosynthesis, negative regulation of RNA metabolism, cell cycle arrest, and apoptosis. The PD specific module was associated with muscle cell proliferation, detoxification and complement activation (with thrombospondin as most interconnected gene). Multi-omics analysis of KTx vs. PD demonstrated concordant upregulation of lipid and fatty acid biosynthesis with the hub gene fatty acid synthase (FASN) and downregulation of positive cell cycle regulation. In independent, matched cohorts, arteriolar abundance of thrombospondin and terminal complement complex were higher in children on PD, arterioles from children after KTx had levels comparable to arterioles from children with CKD5. Key drivers of fibrotic process, arteriolar TGF-ß and pSMAD2/3, were also higher in PD arterioles compared to CKD5 and persistently high after KTx. Arteriolar cell cycle arrest marker p16 and apoptosis marker cleaved Casp3 were higher after KTx as compared to PD and CKD5. FASN was abundant in intima and media layers of all arterioles, and three-fold higher after KTx then in children with PD and CKD5, respectively. Neutral lipids (oil red staining) were present in all three groups, more abundant in the media than in the intima, and two-fold increased in the media of children after KTx compared to children on PD (p = 0.018) and with CKD5 (p = 0.002). In vitro, methylprednisolone and tacrolimus, but not mycophenolate mofetil increased FASN abundance and activity in human umbilical arterial endothelial and vascular smooth muscle cells compared to media controls. Conclusion We for the first time comprehensively describe molecular pathways activated after KTx in children following a standard immunosuppressive regime. After KTx, arteriolar fatty acid biosynthesis, apoptosis and cell cycle arrest markers are increased as compared to arterioles from children with PD and CKD5; profibrotic pathways induced by PD are persistently activated. Arteriolar hubgene FASN is three-fold more abundant after KTx and neutral lipid deposition is increased, presumably due to the induction of FASN activity by methylprednisolone and tacrolimus.

FC 102PD INDUCED ARTERIOLAR AND PERITONEAL PATHOMECHANISMS ARE PARTIALLY REVERSED AFTER KIDNEY TRANSPLANTATION

Abstract Background and Aims Due to the unphysiological composition of PD fluids, chronic peritoneal dialysis (PD) induces progressive peritoneal fibrosis, hypervascularization, and vasculopathy. The evolution of the PD membrane and vasculopathy following kidney transplantation (KTx) is largely unknown. Method Arteriolar and peritoneal tissues were obtained from 107 children with chronic kidney disease (CKD5), 72 children on PD (treated with neutral pH PD fluids, with low glucose degradation product content, GDP) and 21 children, who underwent KTx 4-5 weeks after a median 21 months of PD. Specimen underwent standardized digital quantitative histomorphometry. Molecular mechanisms were studied in omental arterioles microdissected from surrounding fat by multi-omics followed by Gene Set Enrichment Analysis (GSEA); key findings were validated in parietal tissues of independent, matched cohorts by quantitative immunohistochemistry (n=15/group). Results Arteriolar transcriptome and proteome GSEA revealed suppression of leucocyte migration and T-cell activation / secretory pathways regulation, of sprouting angiogenesis biological processes and of epithelial proliferation and cell cycle after KTx as compared to PD. Lipid / fatty acid metabolism, autophagy and ATP synthesis pathways were activated. Transcriptome analysis including KTx, PD and CKD5 specifically attributed regulation of arteriolar lipid and fatty acid metabolism to transplantation and comprised 140 transcripts; their regulation was confirmed on the proteome level. Hub gene fatty acid synthase was identified by protein interaction analysis (string-db.org). 15 arteriolar genes activated by PD were inactivated after KTx and included glucose metabolisms and cytoskeleton related transcripts. 24 transcripts and 10 corresponding proteins induced by PD were still active after KTx and associated with biological processes related to TGF-ß signaling, fibrosis and mineral absorption. In line with arteriolar multi-omics findings, peritoneal hypervascularization induced by chronic PD was reversed after Tx to CKD5 level. CD45 positive tissue infiltrating leucocytes count was reduced by 40% and was independently associated with microvessel density in multivariable analysis including PD vintage, daily GDP exposure and recent KTx. Peritoneal lymphatic vessel density, submesothelial thickness, activated fibroblast, fibrin deposit, macrophage and EMT cell counts remained unchanged after KTx compared to PD. Arteriolar lumen to vessel ratios (a marker of vasculopathy) were similar in both groups. Vessel-homeostasis-related proteins in independent, matched cohorts demonstrated increased caspase-3 abundance in peritoneal arterioles after KTx. Arteriolar VEGF-A, thrombospondin, angiopoietin1/2, and hypoxia-inducible factor-1 (HIF-1a) were unchanged, while submesothelial HIF-1a and angiopoietin1/2 were decreased after Tx, favoring vessel maturation. The abundance of the key driver of fibrosis, TGF-ß-effector pSMAD2/3, was unchanged in the peritoneum and arterioles after Tx. Conclusion Our multi-omics analyses of fat covered omental arterioles, not directly exposed to PD fluids, demonstrate inhibition of PD induced immune response and angiogenesis pathways, of glucose metabolism and cytoskeleton regulation to levels similar as seen in children with CKD5. Arteriolar lipid and fatty acid metabolism is selectively altered after KTx. Reversal of low GDP PD induced hypervascularization and inflammation of the parietal peritoneum after KTx, mirror molecular changes in omental arterioles, while profibrotic activity persists after KTx in omental arterioles and in the parietal peritoneum.

New peritoneal dialysates, for which benefits ?

The ideal peritoneal dialysis solution should allow efficient withdrawal of waste products of the metabolism and water and solutes equilibrium with minimal side effects for the patient and the peritoneal membrane. Glucose degradation products (GDP) resulting from the manufacturing process play a major toxic role and new biocompatible PD solutions with low GDP content and a more physiological bicarbonate or bicarbonate/lactate buffer have brought a clear benefit in experimental studies; however, in clinical cohorts and meta-analysis, the benefits of these solutions appear limited to better preservation of residual renal function and of diuresis. Glucose is the principal osmotic agent although hypertonic glucose solutions have a deleterious effect on PD, and their use should be restrained. Dialysate concentrations of sodium, calcium and magnesium are close to plasma; their diffusive transport is thus limited and their net peritoneal transport mainly depends on the ultrafiltration volume. A dialysate calcium concentration above 1.25 mmol/l generates a calcium load which may contribute to the high prevalence of adynamic bone disease in PD patients; this should be avoided. Low sodium dialysis solutions experimentally improve sodium diffusive transport and extraction in PD patients; the clinical benefit of this approach has to be confirmed.

Bicarbonate buffered peritoneal dialysis fluid upregulates angiopoietin-1 and promotes vessel maturation.

BackgroundUltrafiltration decline is a progressive issue for patients on chronic peritoneal dialysis (PD) and can be caused by peritoneal angiogenesis induced by PD fluids. A recent pediatric trial suggests better preservation of ultrafiltration with bicarbonate versus lactate buffered fluid; underlying molecular mechanisms are unknown.MethodsAngiogenic cytokine profile, tube formation capacity and Receptor Tyrosine Kinase translocation were assessed in primary human umbilical vein endothelial cells following incubation with bicarbonate (BPDF) and lactate buffered (LPDF), pH neutral PD fluid with low glucose degradation product content and lactate buffered, acidic PD fluid with high glucose degradation product content (CPDF). Peritoneal biopsies from age-, PD-vintage- and dialytic glucose exposure matched, peritonitis-free children on chronic PD underwent automated histomorphometry and immunohistochemistry.ResultsIn endothelial cells angiopoietin-1 mRNA and protein abundance increased 200% upon incubation with BPDF, but decreased by 70% with LPDF as compared to medium control; angiopoietin-2 remained unchanged. Angiopoietin-1/Angiopoietin-2 protein ratio was 15 and 3-fold increased with BPDF compared to LPDF and medium. Time-lapse microscopy with automated network analysis demonstrated less endothelial cell tube formation with BPDF compared to LPDF and CPDF incubation. Receptor Tyrosine Kinase translocated to the cell membrane in BPDF but not in LPDF or CPDF incubated endothelial cells. In children dialyzed with BPDF peritoneal vessels were larger and angiopoietin-1 abundance in CD31 positive endothelium higher compared to children treated with LPDF.ConclusionBicarbonate buffered PD fluid promotes vessel maturation via upregulation of angiopoietin-1 in vitro and in children on dialysis. Our findings suggest a molecular mechanism for the observed superior preservation of ultrafiltration capacity with bicarbonate buffered PD fluid with low glucose degradation product content.

Update on Peritoneal Dialysis: Core Curriculum 2016

Update on Peritoneal Dialysis: Core Curriculum 2016

Biocompatible peritoneal dialysis fluids: clinical outcomes.

Peritoneal dialysis (PD) is a preferred home dialysis modality and has a number of added advantages including improved initial patient survival and cost effectiveness over haemodialysis. Despite these benefits, uptake of PD remains relatively low, especially in developed countries. Wider implementation of PD is compromised by higher technique failure from infections (e.g., PD peritonitis) and ultrafiltration failure. These are inevitable consequences of peritoneal injury, which is thought to result primarily from continuous exposure to PD fluids that are characterised by their “unphysiologic” composition. In order to overcome these barriers, a number of more biocompatible PD fluids, with neutral pH, low glucose degradation product content, and bicarbonate buffer have been manufactured over the past two decades. Several preclinical studies have demonstrated their benefit in terms of improvement in host cell defence, peritoneal membrane integrity, and cytokine profile. This paper aims to review randomised controlled trials assessing the use of biocompatible PD fluids and their effect on clinical outcomes.

Peritoneal Dialysis Tailored to Pediatric Needs

Consideration of specific pediatric aspects is essential to achieve adequate peritoneal dialysis (PD) treatment in children. These are first of all the rapid growth, in particular during infancy and puberty, which must be accompanied by a positive calcium balance, and the age dependent changes in body composition. The high total body water content and the high ultrafiltration rates required in anuric infants for adequate nutrition predispose to overshooting convective sodium losses and severe hypotension. Tissue fragility and rapid increases in intraabdominal fat mass predispose to hernia and dialysate leaks. Peritoneal equilibration tests should repeatedly been performed to optimize individual dwell time. Intraperitoneal pressure measurements give an objective measure of intraperitoneal filling, which allow for an optimized dwell volume, that is, increased dialysis efficiency without increasing the risk of hernias, leaks, and retrofiltration. We present the concept of adapted PD, that is, the combination of short dwells with low fill volume to promote ultrafiltration and long dwells with a high fill volume to improve purification within one PD session. The use of PD solutions with low glucose degradation product content is recommended in children, but unfortunately still not feasible in many countries.

3,4-Dideoxyglucosone-3-ene as a mediator of peritoneal demesothelization

The mesothelium contributes significantly to the functional, structural and homeostatic properties of the peritoneum. Bioincompatible peritoneal dialysis solutions contribute to mesothelial cell loss during chronic peritoneal dialysis. Cell death has been implicated in mesothelial cell loss, but the molecular mechanisms have not been adequately characterized. We now report the modulation of mesothelial cell death by the glucose degradation product 3,4-dideoxyglucosone-3-ene (3,4-DGE). Human mesothelial cells were cultured from the effluents of stable dialysis patients. Apoptosis was quantified in cultured mesothelial cells and in peritoneal effluents. Confocal microscopy and inhibitors were used to assess molecular mechanisms. Peritoneal dialysis solutions with a high content of both glucose and glucose degradation products, but not those with low glucose degradation product content, induced mesothelial cell apoptosis and loss of cell viability in culture and in vivo. 3,4-DGE also induced mesothelial cell apoptosis. Apoptosis induced by peritoneal dialysis solutions and 3,4-DGE was associated with oligomerization of Bax at mitochondria and caspase activation. Bax antagonism prevented caspase activation, apoptosis and cell death. The pancaspase inhibitor zVAD was also protective. 3,4-DGE and peritoneal dialysis solutions with a high content in glucose degradation products induce mesothelial cell apoptosis by a Bax-dependent mechanism. This could contribute to chronic demesothelization in peritoneal dialysis.

Reduced systemic advanced glycation end products in children receiving peritoneal dialysis with low glucose degradation product content

Glucose degradation products (GDP) in peritoneal dialysis (PD) solutions are toxic to the peritoneal membrane and promote the formation of advanced glycation end products (AGE), which contribute to accelerated atherosclerosis and amyloidosis. Double chamber PD solutions have a markedly reduced GDP content. We analysed GDP and AGE kinetics in 21 children (7 months to 18 years) on automated PD in a prospective multicentre trial with randomized administration of single chamber, high-GDP and double-chamber, low-GDP dialysis solution for 12 weeks each. Total AGE fluorescence, carboxymethyllysine (CML, ELISA) and 3-deoxyglucosone (3-DG, HPLC) were measured in plasma and PD effluent during a 4 h peritoneal equilibration test. Plasma AGE profiles were assessed by size selective gel permeation chromatography and compared with 23 healthy controls. Initial effluent 3-DG concentrations were 140+/-55 and 25+/-4 micromol/l with high- and low-GDP PD fluid, respectively and declined to 53+/-32 and 7+/-2 micromol/l within 4 h dwell time (P<0.001). The ex vivo AGE generating capacity was three times higher with the high-GDP solution and decreased significantly with dwell time. Plasma AGE levels were 1.8-7.4-fold above those of healthy controls; the elevation was most marked for the small molecular fraction (<2 kDa). Plasma AGE and CML levels were significantly higher after 12 weeks exposure to high-GDP solution (20991+/-4145 AU and 1505+/-617 ng/ml) than following treatment with low-GDP fluid (17518+/-4676 AU and 1151+/-438 ng/ml; both P<0.05). Four hour AGE clearance was higher with low-GDP solution (0.74+/-0.3 vs 0.44+/-0.15 ml/min*1.73 m2, P<0.01). GDP are rapidly absorbed from the peritoneal cavity. Administration of PD solutions with low-GDP content reduces plasma AGE levels and may thus improve the cardiovascular risk profile of dialysed children.

Biocompatibility of a Bicarbonate/Lactate-Buffered PD Fluid Tested with a Double-Chamber Cell Culture System

In peritoneal dialysis (PD), neutrally buffered PD fluids with lower concentrations of glucose degradation products (GDP) have tested superior to conventional fluids in terms of biocompatibility. However, conventional in vitro studies provoke debate because, due to the lack of subsequent equilibration with the blood, they do not resemble the true intraperitoneal situation of PD. We established a double-chamber cell culture system with peritoneal mesothelial cells seeded on top of a permeable membrane, with a physiological buffer below. Thus adequately reflecting the in vivo equilibration pattern, we compared a conventional fluid with a neutral bicarbonate/lactate-buffered PD solution. Using an exchange pattern adapted from an 8-hour continuous ambulatory PD regimen, cell viability was assessed with an MTT assay, and cell function via constitutive and stimulated interleukin (IL)-6 release. As an indicator of potential induction of fibrosis and as a parameter of mesothelial cell integrity, respectively, transforming growth factor-beta 1 (TGF-beta1) generation and cancer antigen 125 (CA125) release were measured. The conventional solution significantly compromised mesothelial cell viability and function in terms of mitochondrial activity (p < 0.05) and stimulated IL-6 release (p < 0.05). The bicarbonate/lactate fluid had no effect on cell viability or IL-6 release and turned out to be equivalent to the properties of the growth medium. Whereas lactate-incubated cells did not respond to IL-1beta stimulation, bicarbonate/lactate-treated cells adequately increased IL-6 release after stimulation (p < 0.0005). Release of TGF-beta1 and CA125 did not differ between the different fluids and the control. Due to the sustained equilibration process, the double-chamber cell culture model allows a more realistic insight into mesothelial cell viability and function in terms of PD. As in classic in vitro studies, an adverse effect of conventional PD solutions on mesothelial cells was overt in the present cell culture system. The neutral bicarbonate/lactate-buffered fluid with low GDP content, however, did not interfere with mesothelial cell vitality or function, indicating superior biocompatibility.

La biocompatibilité des solutions de dialyse péritonéale

Repeated and long-term exposure to conventional glucose-based peritoneal dialysis fluids (PDFs) with poor biocompatibility plays a central role in the pathogenesis of the functional and structural changes of the peritoneal membrane. We have used immortalized human peritoneal mesothelial cells in culture to assess in vitro the biocompatibility of PDFs. Low pH, high glucose concentration and heat sterilization represent major factors of low biocompatibility. Two recent groups of glucose derivatives have been described. Glucose degradation products (GDPs) are formed during heat sterilization (glycoxidation) and storage. GDPs can bind protein and form AGEs (Advanced Glycation End-products), which can also result from the binding of glucose to free NH 2 residues of proteins (glycation). The physiological pH, and the separation of glucose during heat sterilization (low GDP content) in the most recent PDFs dramatically increase the biocompatibility. The choice of PD programs with high biocompatibility PDFs allows preserving the function of the peritoneal membrane. Improvement of PDF biocompatibility may limit the occurrence of chronic chemical peritonitis and may allow long-term PD treatment.

Differential expression of receptors for advanced glycation end-products in peritoneal mesothelial cells exposed to glucose degradation products

Autoclaving peritoneal dialysate fluid (PDF) degrades glucose into glucose degradation products (GDPs) that impair peritoneal mesothelial cell functions. While glycation processes leading to formation of advanced glycation end-products (AGE) were viewed commonly as being mediated by glucose present in the PDF, recent evidence indicates that certain GDPs are even more powerful inducers of AGE formation than glucose per se. In the present study, we examined the expression and modulation of AGE receptors on human peritoneal mesothelial cells (HPMC) cultured with GDPs, conventional PDF or PDF with low GDP content. HPMC cultured with GDPs differentially modulated AGE receptors (including RAGE, AGE-R1, AGE-R2 and AGE-R3) expression in a dose-dependent manner. At subtoxic concentrations, GDPs increased RAGE mRNA expression in HPMC. 2-furaldehyde (FurA), methylglyoxal (M-Glx) and 3,4-dideoxy-glucosone-3-Ene (3,4-DGE) increased the expression of AGE-R1 and RAGE, the receptors that are associated with toxic effects. These three GDPs up-regulated the AGE synthesis by cultured HPMC. In parallel, these GDPs also increased the expression of vascular endothelial growth factor (VEGF) in HPMC. PDF with lower GDP content exerted less cytotoxic effect than traditional heat-sterilized PDF. Both PDF preparations up-regulated the protein expression of RAGE and VEGF. However, the up-regulation of VEGF in HPMC following 24-h culture with conventional PDF was higher than values from HPMC cultured with PDF containing low GDP. We have demonstrated, for the first time, that in addition to RAGE, other AGE receptors including AGE-R1, AGE-R2 and AGE-R3 are expressed on HPMC. Different GDPs exert differential regulation on the expression of these receptors on HPMC. The interactions between GDPs and AGE receptors may bear biological relevance to the intraperitoneal homeostasis and membrane integrity.

The Effects of Peritoneal Dialysis Solutions on Peritoneal Host Defense

Conventional peritoneal dialysis fluid (PDF) is a bioincompatible solution owing to the acidic pH, the high glucose concentrations and the associated hyperosmolarity, the high lactate concentrations, and the presence of glucose degradation products (GDPs). This unphysiologic composition adversely affects peritoneal host defense and may thus contribute to the development of PD-related peritonitis. The viability of polymorphonuclear leukocytes, monocytes, peritoneal macrophages, and mesothelial cells is severely depressed in the presence of conventional PDF. In addition, the production of inflammatory cytokines and chemoattractants by these cells is markedly affected by conventional PDF. Further, conventional PDF hampers the recruitment of circulating leukocytes in response to an infectious stimulus. Finally, phagocytosis, respiratory burst, and bacterial killing are markedly lower when polymorphonuclear leukocytes, monocytes, and peritoneal macrophages are exposed to conventional PDF. Although there are a few discrepant results, all major PDF components have been implicated as causative factors. Generally, novel PDF with alternative osmotic agents or with alternative buffers, neutral pH, and low GDP content have much milder inhibitory effects on peritoneal host defense. Clinical studies, however, still need to demonstrate their superiority with respect to the incidence of PD-related peritonitis.

What Can We Do to Preserve the Peritoneum?

Long-term peritoneal dialysis may lead to peritoneal membrane failure. Loss of ultrafiltration is the most important clinical abnormality. Loss of ultrafiltration is associated with an increased number of peritoneal blood vessels, with fibrotic alterations, and with loss of mesothelium. Continuous exposure to bioincompatible dialysis solutions is likely to be important in the pathogenesis of these alterations. This article reviews the toxicity of various constituents of dialysate, current assessments of interventions, and the results of interventions aimed at preserving the peritoneum. Glucose, possibly in combination with lactate, and glucose degradation products (GDPs) are likely to be the most toxic constituents of dialysate. Diabetiform peritoneal neoangiogenesis is likely to be mediated by vascular endothelial growth factor (VEGF). Release of VEGF might be influenced by glucose-induced cellular pseudohypoxia, which is likely to be increased by exposure to lactate. Glucose and GDPs are both toxic to peritoneal cells. Glucose degradation products induce the formation of advanced glycosylation end-products at a much faster rate than does glucose itself, but the relative importance of GDPs and glucose in clinical PD has not been clarified. The effects of interventions should first be assessed in long-term animal models, followed by clinical studies on peritoneal transport and on effluent markers that may reflect the status of the peritoneum. Possible interventions aim at reducing peritoneal exposure to glucose, GDPs, and lactate. Techniques include peritoneal resting, replacing some glucose-based exchanges with amino acid-based and icodextrin-based dialysate, using bicarbonate as a buffer, and administering solutions that have a low GDP content. Exposure to various dialysis solutions with a reduced GDP content has resulted in an increase in the effluent concentration of the mesothelial cell marker CA125, irrespective of the buffer used. Experimental studies in a long-term peritoneal exposure model in rats showed that the combination of a reduction in the concentration of lactate and replacement of lactate with pyruvate resulted in a reduction of the number of peritoneal blood vessels. Results of drug therapy have been studied in various animal models. Their use in patients is still experimental. Strategies to preserve the peritoneum aim at reducing membrane exposure to bioincompatible solutions. Currently available dialysis fluids that are more biocompatible are likely to have some beneficial effects. Further research on the development of dialysis solutions that use combinations of osmotic agents and alternative buffers is necessary.

Effects of conventional and new peritoneal dialysis fluids on leukocyte recruitment in the rat peritoneal membrane.

Peritonitis remains an important cause of morbidity and technique failure in peritoneal dialysis (PD). Conventional peritoneal dialysate fluids (PDF) inhibit peritoneal leukocyte function in vitro and may thus adversely affect the immune response to peritonitis. New PDF have been designed with neutral pH, low glucose degradation product (GDP) contents, and bicarbonate as buffer. The present intravital microscopy study examined the effects of conventional and new PDF on leukocyte behavior in the peritoneal microcirculation of Wistar rats. The visceral peritoneum was superfused by a control solution (EBSS), a conventional (CAPD), or a new bicarbonate-buffered PDF with neutral pH and low GDP content (CAPD BicaVera). In addition, spent conventional and new PDF were tested. The number of rolling, adhering, and extravasated leukocytes and leukocyte rolling velocity were assessed at different time intervals after exposure to lipopolysaccharide (LPS) or cell-free supernatants of coagulase-negative staphylococci (CNS-CFS). Exposure to LPS or CNS-CFS dissolved in EBSS dramatically increased the number of rolling, adhering and extravasated leukocytes and decreased leukocyte rolling velocity. Superfusion by CAPD abolished the LPS- or CNS-CFS-induced leukocyte recruitment, whereas CAPD BicaVera had significantly fewer depressant effect. Spent PDF affected the leukocyte response in a similar way as fresh PDF. High lactate concentrations, GDP, and hypertonicity appeared to be mainly responsible for the inhibition of leukocyte recruitment. In conclusion, conventional PDF abolish in vivo leukocyte recruitment in response to potent inflammatory stimuli. Bicarbonate-buffered pH-neutral PDF with low GDP contents have fewer depressant effects and may therefore contribute to a better preservation of peritoneal host defense.

Hemodynamic effects of peritoneal dialysis solutions on the rat peritoneal membrane: role of acidity, buffer choice, glucose concentration, and glucose degradation products.

Conventional peritoneal dialysis fluids (PDF) are unphysiologic because of their hypertonicity, high glucose and lactate concentrations, acidic pH, and presence of glucose degradation products (GDP). Long-term exposure to conventional PDF may cause functional and structural alterations of the peritoneal membrane. New PDF have a neutral pH, a low GDP content, and contain bicarbonate or lactate as the buffer. Intravital microscopy was used to analyze the vasoactive effects of conventional and new PDF on the rat peritoneal membrane. A conventional, acidic pH, lactate-buffered 4.25% glucose PDF induced maximal vasodilation of mesenteric arteries, resulting in a doubling of the arteriolar flow and a 20% increase of the perfused capillary length per area. The hemodynamic effects of conventional PDF were similar after pH-adjustment with NaOH, indicating that acidity per se is not essential for the changes. Superfusion by a pH-neutral, lactate-buffered PDF with low GDP content caused only a transient arterial vasodilation despite continuous exposure, with a commensurate effect on arteriolar flow and capillary recruitment. Application of a pH-neutral, bicarbonate-buffered PDF with low GDP content did not affect the hemodynamic parameters. Resterilization of the bicarbonate solution increased GDP levels and completely restored the vasodilatory capacity. The corresponding 1.5% glucose PDF induced similar but less pronounced changes. Conventional PDF have important vasoactive effects on the peritoneal circulation, mainly because of the presence of GDP and transiently because of high lactate concentrations. Capillary recruitment may increase effective peritoneal vascular surface area. In addition, chronic vasodilation may induce structural adaptations in the blood vessel wall, contributing to vascular sclerosis. PDF with reduced GDP content induce no major hemodynamic effects and may thus have the potential to better preserve peritoneal vascular integrity.

Prolonged preservation of rat peritoneal mesothelial cells (RPMC) and rat peritoneal fibroblasts (RPFB) with neutral pH peritoneal dialysis solution depleted of glucose degradation products (GDPs)

Background. In-vitro biocompatibility tests were conducted to verify the assumption that glucose degradation products (GDPs) and pH could be factors that reduce peritoneal cell viability. Methods. Peritoneal mesothelial cells and fibroblasts isolated from the omentum of male Wistar rats were used in all experiments. Peritoneal dialysis solutions (PDSs) to be tested varied in GDP content, pH, and glucose concentration (1.36%, 2.27%, and 3.86%). Subconfluent cells were exposed to the dialysis solutions over time courses of 30 min to 6 h. Cell viability was assessed by MTT evaluation and the gene expression of interleukin 6 (IL-6) was determined by reverse transcription (RT)-polymerase chain reaction (PCR). Peritoneal dialysis solutions (PDSs) tested were: (I) PDS autoclaved (PDS-AC), neutral pH, +GDP; (II) PDS filtered (PDS-Filt), neutral pH, −GDP; (III) Gambrosol autoclaved (Gsol-AC), acidic pH, ++GDP; and (IV) Gambrosol filtered (Gsol-Filt), acidic pH, −GDP. Results. After 30-min exposure to the dialysis solutions, cell viability was greatest in cells exposed to group II solutions, followed by groups I, IV, and III. With increased exposure time (6 h), this difference in viability was further enhanced, showing that cells exposed to high GDP and acidic pH had a significant reduction in viability as compared with cells exposed to low GDP and neutral pH. The expression of IL-6 mRNA in mesothelial cells was much more susceptible to pH than this expression in peritoneal fibroblasts, as was also seen in the MTT evaluation. Conclusions. These results indicate that dialysis solutions with neutral pH and low GDP content provide a more biocompatible environment than dialysis solutions with acidic pH and high GDP content in which to maintain the viability of resident peritoneal cells.