Ola Carlsson

Transendothelial transport: the vesicle controversy.

The relative contribution of transcytosis vs. large pore transport to the passage of macromolecules across microvascular endothelia has been a controversial issue for nearly half a century. To separate transcytosis from ‘porous’ transport, the transcytosis inhibitors N-ethylmaleimide (NEM) and filipin have been tested in in situ or ex vivo perfused organs with highly conflicting results. In continually weighed isolated perfused organs, where measurements of pre- and post-capillary resistances, capillary pressure and capillary filtration coefficients can be repeatedly performed, high doses of NEM and filipin increased the bulk transport of macromolecules from blood to tissue, despite producing vasoconstriction. By contrast, in in situ perfused organs, marked reductions in the tissue uptake of albumin tracer have been observed after NEM and filipin. When tissue cooling has been employed as a means of inhibiting (active) transcytosis, results have invariably shown a low cooling sensitivity of albumin transport, compatible with passive transendothelial passage of albumin. This observation is further strengthened by the commonly observed dependence of albumin transport upon the capillary pressure and the rate of transcapillary convection. For low-density lipoprotein (LDL), a cooling-sensitive, non-selective transport component has been discovered, which may be represented by filtration through paracellular gaps, lateral diffusion through transendothelial channels formed by fused vesicles, or by transcytosis. From a physiological standpoint there is little evidence supporting active transendothelial transport of most plasma macromolecules. This seems to be supported by studies on caveolin-1-deficient mice lacking plasmalemmal vesicles (caveolae), in which there are no obvious abnormalities in the transendothelial transport of albumin, immunoglobulins or lipoproteins. Nevertheless, specific transport in peripheral capillaries of several hormones and other specific substances, similar to that existing across the blood-brain barrier, still remains as a possibility.

The effects of low-sodium peritoneal dialysis fluids on blood pressure, thirst and volume status

Background. Poor ultrafiltration is associated with worse outcomes in peritoneal dialysis (PD) patients. This might in part reflect problems associated with salt and water excess. Increasing the diffusive component of peritoneal sodium removal using low-sodium PD fluids might have beneficial effects on blood pressure (BP), thirst and fluid status that could translate into clinical benefits. Methods. Using a multicentre, prospective, baseline controlled (1 month), non-randomized intervention (2 months) design, two novel solutions designed from predictions using the three-pore model were investigated. In group A ([Na+] = 115 mmol/l), the glucose (G) was increased to 2.0% to compensate for reduced osmolality whereas in group B ([Na+] = 102 mmol/l), it was unchanged (2.5%). Both solutions were substituted for one 3- to 5-h exchange per day and no change was made to the rest of the dialysis regime. Results. Ten patients in group A and 15 in group B completed the study. Both solutions resulted in significant increases (30–50 mmol/dwell) in diffusive sodium removal during the test exchanges, P < 0.001. Ultrafiltration was maintained in group A but reduced in group B. Ambulatory nocturnal mean BP fell in group A [93.1 ± 10.6 mmHg (±SD) versus 85.1 ± 10.2 mmHg, P < 0.05], but was stable in group B (95.4 ± 9.4 versus 95.1.1 ± 10.7 mmHg, NS). Thirst reduced independent of appetite and mood in both groups by 2 months, more markedly in group A. Indices of fluid status, including TBW by bioimpedance and D dilution also improved in group A, P < 0.05, whereas weight increased in group B. Conclusions. Increasing the diffusive component of sodium removal whilst maintaining ultrafiltration is associated with improvements in BP, thirst and fluid status. The lack of effect seen with uncompensated low-sodium dialysate suggests that these benefits cannot be achieved by manipulation of dialysate sodium removal alone. These observations provide valuable information of the design of future randomized studies to establish the clinical role for low-sodium dialysis fluids.

Citrate treatment reduces endothelial death and inflammation under hyperglycaemic conditions

Hyperglycaemia and glucose degradation products (GDPs) are closely associated with oxidative stress and inflammation in diabetic patients, a condition that leads to endothelial dysfunction and cardiovascular problems. We evaluated the effect of citrate and gluconate on glucose- and GDP-induced endothelial inflammation by measuring changes in viability, inflammation and function in primary human umbilical vein endothelial cells (HUVECs). The extent of apoptosis/necrosis was measured by flow cytometry and visualised with confocal microscopy by staining with annexin V or propidium iodide, respectively. Protein kinase C-βII (PKC-βII) activation was evaluated with Western blotting. Incubation with glucose (30 mM) and GDP (50 µM) significantly increased PKC-βII expression, endothelial cell death and inflammation. The addition of citrate decreased hyperglycaemia-induced apoptosis (p = 0.021), necrosis (p = 0.04) and reduced PKC-βII expression (p = 0.021) down to background levels. Citrate improved endothelial function by reducing the inflammatory markers (p = 0.01) and by decreasing neutrophil diapedesis (p = 0.012). These results suggest that citrate may have therapeutic potential by reducing hyperglycaemia-induced endothelial inflammation and abolishing endothelial dysfunction.

Transvascular Passage of Macromolecules into the Peritoneal Cavity of Normo- and Hypothermic Rats in vivo: Active or Passive Transport?

During the last decades there has been a debate regarding whether transvascular protein transport is an active (transcytosis) or a passive (porous) process. To separate cooling-sensitive transcytosis from passive transport processes between blood and peritoneal fluid, we induced hypothermia in rats in vivo, reducing their body temperature to 19°C. Control rats were kept at 37°C. Either human albumin, or IgG, or IgM, or LDL, radiolabeled with 125I, was given intra-arterially together with 51Cr-EDTA. During tracer administration, a 2-hour peritoneal dialysis dwell was performed. Clearance of the tracers to dialysate, and the permeability-surface area coefficient (PS) for 51Cr-EDTA and glucose were assessed. During cooling, mean arterial blood pressure (MAP) was reduced to 40% of control and plasma viscosity increased by 48.5%, while peritoneal blood flow was reduced to 10%. At 19°C, clearance of albumin to dialysate fell from 9.30 ± 1.62 (SEM) to 3.13 ± 0.28 µl/min (p < 0.05), clearance of IgG from 6.33 ± 0.42 to 2.54 ± 0.12 µl/min (p < 0.05), clearance of IgM from 3.65 ± 0.33 to 1.10 ± 0.12 µl/min (p < 0.05), and clearance of LDL from 3.54 ± 0.20 to 0.73 ± 0.06 µl/min (p < 0.05). The fall in PS for 51Cr-EDTA was from 0.320 ± 0.01 to 0.075 ± 0.003 ml/min (p < 0.05), and that for glucose from 0.438 ± 0.02 to 0.105 ± 0.01 ml/min (p < 0.05). Tissue cooling reduced large solute transport largely in proportion to the cooling-induced reductions of MAP (to 40%), and the concomitant increase in viscosity (to 67%), i.e. to ≈20–30% (0.40 × 0.67) of control, though LDL clearance was reduced further. The fall in small solute PS, in excess of the viscosity effect, mirrored the fall in peritoneal blood flow occurring during hypothermia. In conclusion, the good correlation of predicted to calculated changes suggests that the overall transendothelial macromolecular passage in vivo occurs passively, and not due to active processes.

Enhanced Peritoneal Diffusion Capacity of 51Cr-EDTA during the Initial Phase of Peritoneal Dialysis Dwells: Role of Vasodilatation, Dialysate ‘Stirring’, and of Interstitial Factors

Mass transfer area coefficients (PS) to small solutes are usually markedly increased during the first 0–15 min of peritoneal dialysis (PD) dwells. This phenomenon may be due to, for example, initial arteriolar vasodilatation and, hence, recruitment of capillary surface area. Other possibilities are an initial discharge (or saturation) of solutes from (in) the interstitium or an increased mixing, i.e., ‘macrostirring’ caused by the exchange procedure per se. We have investigated these possibilities during acute PD in rats, by assessing PS for 51Cr-EDTA as a function of time [PS(t)]. The discharge effect was studied by saturating the peritoneal interstitium with 51Cr-EDTA by intravenous tracer infusion prior to each dwell and the results compared to those obtained in rats when tracer infusion and dwells were started simultaneously. The potential effect of initial vasodilatation was studied by adding isoproterenol to the dialysis fluid. Finally, the potential influence of an increased interstitial ‘microstirring’, induced by high glucose concentrations, was investigated by comparison of results for 1.36% Dianeal® with those for 3.86% Dianeal. In nonvasodilated rats there was a significant drop in PS(t) between 2.5 and 15 min regardless of whether the rats were preloaded with tracer or not. However, there were no significant changes in PS(t) within the isoproterenol-treated group, indicating that vasodilatation plays a crucial role for the high PS initially. Furthermore, there was no difference in overall PS for 51Cr-EDTA among 1.36 and 3.86% Dianeal dwells. In conclusion, we have found that vasodilatation, but not interstitial discharge (or loading), may explain the inflation of PS occurring during the initial part of PD dwells. In addition, ‘macrostirring’, induced by the exchange procedure per se, may also be important.

Peritoneal lymphatic absorption and solute exchange during zymosan-induced peritonitis in the rat

Lymph flow is elevated in most inflammatory conditions. However, a few previous studies have indicated that peritoneal lymph flow may actually fall during acute peritonitis. This study was performed to explore this issue further and to study the pathophysiology of peritoneal exchange during peritonitis. Therefore, we wanted to assess the total peritoneal clearance (Cl) and the clearance from peritoneum to plasma (Cl → P) of 125I-labeled albumin (125I-albumin) as well as plasma-to-dialysate clearance (Cl → D) of Evans blue-labeled albumin together with peritoneal ultrafiltration (UF) profiles and mass transfer area coefficients of51Cr-EDTA and glucose in rats after acute peritonitis induced by zymosan. Zymosan incubation of the peritoneal cavity (120 mg) for 4 h generally led to a 4- to 10-fold increase in peritoneal fluid white blood cell count, indicating that acute peritonitis had been induced. Then 16 ml of 3.86% Dianeal and125I-albumin were instilled intraperitoneally, whereas Evans blue-labeled albumin and51Cr-EDTA were given as infusions intravenously. Compared with control, mass transfer area coefficients for glucose and 51Cr-EDTA increased markedly from 0.43 ± 0.06 and 0.25 ± 0.04 to 0.91 ± 0.06 and 0.59 ± 0.05 (SE) ml/min, respectively, during peritonitis, whereas Cl and Cl → D increased from 32.8 ± 5.6 and 8.6 ± 1.6 to 74.5 ± 7.3 and 12.9 ± 1.0 μl/min, respectively. The UF profile in peritonitis indicated type I loss of UF (resulting from the increases in permeability-surface area product for glucose). However, the Cl → P declined to 5.9 ± 1.0 μl/min from 7.9 ± 0.8 μl/min ( P < 0.05) in control. In conclusion, despite marked effects on peritoneal solute transport and on UF, conceivably resulting from vasodilatation and increases in capillary permeability, zymosan-induced peritonitis did not cause any acute increases in direct peritoneal lymphatic absorption.Lymph flow is elevated in most inflammatory conditions. However, a few previous studies have indicated that peritoneal lymph flow may actually fall during acute peritonitis. This study was performed to explore this issue further and to study the pathophysiology of peritoneal exchange during peritonitis. Therefore, we wanted to assess the total peritoneal clearance (Cl) and the clearance from peritoneum to plasma (Cl --> P) of 125I-labeled albumin (125I-albumin) as well as plasma-to-dialysate clearance (Cl --> D) of Evans blue-labeled albumin together with peritoneal ultrafiltration (UF) profiles and mass transfer area coefficients of 51Cr-EDTA and glucose in rats after acute peritonitis induced by zymosan. Zymosan incubation of the peritoneal cavity (120 mg) for 4 h generally led to a 4- to 10-fold increase in peritoneal fluid white blood cell count, indicating that acute peritonitis had been induced. Then 16 ml of 3.86% Dianeal and 125I-albumin were instilled intraperitoneally, whereas Evans blue-labeled albumin and 51Cr-EDTA were given as infusions intravenously. Compared with control, mass transfer area coefficients for glucose and 51Cr-EDTA increased markedly from 0.43 +/- 0.06 and 0.25 +/- 0.04 to 0.91 +/- 0.06 and 0.59 +/- 0.05 (SE) ml/min, respectively, during peritonitis, whereas Cl and Cl --> D increased from 32.8 +/- 5.6 and 8.6 +/- 1.6 to 74.5 +/- 7.3 and 12.9 +/- 1.0 microl/min, respectively. The UF profile in peritonitis indicated type I loss of UF (resulting from the increases in permeability-surface area product for glucose). However, the Cl --> P declined to 5.9 +/- 1.0 microl/min from 7.9 +/- 0.8 microl/min (P < 0.05) in control. In conclusion, despite marked effects on peritoneal solute transport and on UF, conceivably resulting from vasodilatation and increases in capillary permeability, zymosan-induced peritonitis did not cause any acute increases in direct peritoneal lymphatic absorption.

Phoxilium(®) reduces hypophosphataemia and magnesium supplementation during continuous renal replacement therapy.

Abstract Background Although associated with severe clinical complications, phosphate remains a neglected ion. Additionally, phosphate balance during continuous renal replacement therapy (CRRT) is complex and multifunctional. The present retrospective study investigated the effects of phosphate-containing CRRT fluid on phosphate homeostasis. Methods We retrospectively analysed 112 patients treated with CRRT at Skåne University Hospital, Sweden. The control group was treated with Hemosol® B0 (no phosphate; n = 36) as dialysis and replacement fluid, while the study group received Phoxilium® (phosphate; n = 76) as dialysis fluid and Hemosol® B0 as replacement fluid. Results Hypophosphataemia (<0.7 mM) occurred in 15% of the treatment days in the control group compared with 7% in the study group (P = 0.027). Magnesium substitution was reduced by 40% in the study group (P < 0.001). No differences in acid–base parameters were detected between the groups. Conclusions In this larger cohort, we could confirm that Phoxilium® reduced the episodes of hypophosphataemia during CRRT. A beneficial effect on magnesium balance could also be observed.

Simplified Citrate Anticoagulation for CRRT Without Calcium Replacement

Since 2012, citrate anticoagulation is the recommended anticoagulation strategy for continuous renal replacement therapy (CRRT). The main drawback using citrate as anticoagulant compared with heparin is the need for calcium replacement and the rigorous control of calcium levels. This study investigated the possibility to achieve anticoagulation while eliminating the need for calcium replacement. This was successfully achieved by including citrate and calcium in all CRRT solutions. Thereby the total calcium concentration was kept constant throughout the extracorporeal circuit, whereas the ionized calcium was kept at low levels enough to avoid clotting. Being a completely new concept, only five patients with acute renal failure were included in a short, prospective, intensely supervised nonrandomized pilot study. Systemic electrolyte levels and acid–base parameters were stable and remained within physiologic levels. Ionized calcium levels declined slightly initially but stabilized at 1.1 mmol/L. Plasma citrate concentrations stabilized at approximately 0.6 mmol/L. All postfilter ionized calcium levels were <0.5 mmol/L, that is, an anticoagulation effect was reached. All filter pressures were normal indicating no clotting problems, and no visible clotting was observed. No calcium replacement was needed. This pilot study suggests that it is possible to perform regional citrate anticoagulation without the need for separate calcium infusion during CRRT.