Binding of ATP to vascular endothelial growth factor isoform VEGF-A165 is essential for inducing proliferation of human umbilical vein endothelial cells

<p>Abstract</p> <p>Background</p> <p>ATP binding is essential for the bioactivity of several growth factors including nerve growth factor, fibroblast growth factor-2 and brain-derived neurotrophic factor. Vascular endothelial growth factor isoform 165 (VEGF-A₁₆₅) induces the proliferation of human umbilical vein endothelial cells, however a dependence on ATP-binding is currently unknown. The aim of the present study was to determine if ATP binding is essential for the bioactivity of VEGF-A₁₆₅.</p> <p>Results</p> <p>We found evidence that ATP binding toVEGF-A₁₆₅induced a conformational change in the secondary structure of the growth factor. This binding appears to be significant at the biological level, as we found evidence that nanomolar levels of ATP (4-8 nm) are required for the VEGF-A₁₆₅-induced proliferation of human umbilical vein endothelial cells. At these levels, purinergic signaling by ATP <it>via </it>P2 receptors can be excluded. Addition of alkaline phosphate to cell culture lowered the ATP concentration in the cell culture medium to 1.8 nM and inhibited cell proliferation.</p> <p>Conclusions</p> <p>We propose that proliferation of endothelial cells is induced by a VEGF-A₁₆₅-ATP complex, rather than VEGF-A₁₆₅alone.</p

Oxygen carriers affect kidney immunogenicity during ex-vivo machine perfusion

Normothermic ex-vivo machine perfusion provides a powerful tool to improve donor kidney preservation and a route for the delivery of pharmacological or gene therapeutic interventions prior to transplantation. However, perfusion at normothermic temperatures requires adequate tissue oxygenation to meet the physiological metabolic demand. For this purpose, the addition of appropriate oxygen carriers (OCs) to the perfusion solution is essential to ensure a sufficient oxygen supply and reduce the risk for tissue injury due to hypoxia. It is crucial that the selected OCs preserve the integrity and low immunogenicity of the graft. In this study, the effect of two OCs on the organ's integrity and immunogenicity was evaluated. Porcine kidneys were perfused ex-vivo for four hours using perfusion solutions supplemented with red blood cells (RBCs) as conventional OC, perfluorocarbon (PFC)-based OC, or Hemarina-M101 (M101), a lugworm hemoglobin-based OC named HEMO2life®, recently approved in Europe (i.e., CE obtained in October 2022). Perfusions with all OCs led to decreased lactate levels. Additionally, none of the OCs negatively affected renal morphology as determined by histological analyses. Remarkably, all OCs improved the perfusion solution by reducing the expression of pro-inflammatory mediators (IL-6, IL-8, TNFα) and adhesion molecules (ICAM-1) on both transcript and protein level, suggesting a beneficial effect of the OCs in maintaining the low immunogenicity of the graft. Thus, PFC-based OCs and M101 may constitute a promising alternative to RBCs during normothermic ex-vivo kidney perfusion

Acid-base and electrolyte status during normovolemic hemodilution with succinylated gelatin or HES-containing volume replacement solutions in rats.

BACKGROUND: In the past, several studies have compared different colloidal replacement solutions, whereby the focus was usually on the respective colloid. We therefore systematically studied the influence of the carrier solution's composition of five approved colloidal volume replacement solutions (Gelafundin, Gelafusal, Geloplasma, Voluven and Volulyte) on acid-base as well as electrolyte status during and following acute severe normovolemic hemodilution. The solutions differed in the colloid used (succinylated gelatin vs. HES) and in the presence and concentration of metabolizable anions as well as in their electrolyte composition. METHODS: Anesthetized Wistar rats were subjected to a stepwise normovolemic hemodilution with one of the solutions until a final hematocrit of 10%. Subsequent to dilution (162 min), animals were observed for an additional period (150 min). During dilution and observation time blood gas analyses were performed eight times in total. Additionally, in the Voluven and Volulyte groups as well as in 6 Gelafundin animals, electrolyte concentrations, glucose, pH and succinylated gelatin were measured in urine and histopathological evaluation of the kidney was performed. RESULTS: All animals survived without any indications of injury. Although the employed solutions differed in their respective composition, comparable results in all plasma acid-base and electrolyte parameters studied were obtained. Plasma pH increased from approximately 7.28 to 7.39, the plasma K(+) concentration decreased from circa 5.20 mM to 4.80-3.90 mM and the plasma Cl(-) concentration rose from approximately 105 mM to 111-120 mM. Urinary analysis revealed increased excretion of K(+), H(+) and Cl(-). CONCLUSIONS: The present data suggest that the carrier solution's composition with regard to metabolizable anions as well as K(+), Ca(2+) only has a minor impact on acid-base and electrolyte status after application of succinylated gelatin or HES-containing colloidal volume replacement solutions

Safety of poly (ethylene glycol)-coated perfluorodecalin-filled poly (lactide-co-glycolide) microcapsules following intravenous administration of high amounts in rats

The host response against foreign materials designates the biocompatibility of intravenously administered microcapsules and thus, widely affects their potential for subsequent clinical use as artificial oxygen/drug carriers. Therefore, body distribution and systemic parameters, as well as markers of inflammation and indicators of organ damage were carefully evaluated after administration of short-chained poly (vinyl alcohol, (PVA)) solution or poly (ethylene glycol (PEG))-shielded perfluorodecalin-filled poly (d,l-lactide-co-glycolide, PFD-filled PLGA) microcapsules into Wistar rats. Whereas PVA infusion was well tolerated, all animals survived the selected dose of 1247 mg microcapsules/kg body weight but showed marked toxicity (increased enzyme activities, rising pro-inflammatory cytokines and complement factors) and developed a mild metabolic acidosis. The observed hypotension emerging immediately after start of capsule infusion was transient and mean arterial blood pressure restored to baseline within 70 min. Microcapsules accumulated in spleen and liver (but not in other organs) and partly occluded hepatic microcirculation reducing sinusoidal perfusion rate by about 20%. Intravenous infusion of high amounts of PFD-filled PLGA microcapsules was tolerated temporarily but associated with severe side effects such as hypotension and organ damage. Short-chained PVA displays excellent biocompatibility and thus, can be utilized as emulsifier for the preparation of drug carriers designed for intravenous use

Influence of different volume replacement solutions on blood pressure during and subsequent to normovolemic hemodilution.

<p>Rats underwent normovolemic hemodilution to a final hematocrit of 10% for 162 min (phase of hemodilution: light grey) with either Gelafundin, Gelafusal or Geloplasma and were subsequently observed for 150 min. For analysis mean arterial blood pressure was monitored.</p

Effects of different volume replacement solutions on electrolyte and metabolic parameters as well as pH during and subsequent to normovolemic hemodilution in urine.

<p>Note: Calculations were made based on the values of the six individual animals. Therefore calculations using mean values may differ.</p

Chemical structure of the colloidal substances employed.

<p>1) Succinylated gelatin chain whose terminal carboxyl group ist deprotonated at physiological pH. 2) Hydroxyethyl starch whose single glucose subunits are additionally hydroxyethylated at carbon atoms two, three and six.</p

Effects of different volume replacement solutions on electrolyte parameters during and subsequent to normovolemic hemodilution.

<p>Rats underwent normovolemic hemodilution to a final hematocrit of 10% for 162 min (phase of hemodilution: light grey) with either Gelafundin, Gelafusal, Geloplasma, Voluven or Volulyte and were subsequently observed for 150 min. Electrolytes were determined in arterial blood samples at the points indicated. * <0.05 compared with the value measured at 3 min, in the respective group. # <0.05 (Voluven vs. Volulyte) ** <0.05 (Gelafundin vs. Gelafusal).</p

Composition of the five volume replacement solutions Gelafundin, Gelafusal, Geloplasma, Voluven and Volulyte.

<p>Note: The apparent anion gap in the case of Gelafundin, Gelafusal and Geloplasma is compensated by the negative charge of succinylated gelatin.</p