Oncotic Agents Research Articles

Authorizing of Immunogenicity of Concentrated and Purified Newcastle Disease Virus V4 Strain using Downstream Processing.

Newcastle disease virus (NVD) from the Paramyxoviridae family is a single-stranded negative-sense RNA virus. This infection can affect both domestic poultry and almost all other bird species. It has been considered a very severe difficulty for the poultry industry all over the world. Even though it remains a potential threat to poultry industries, this virus is a powerful oncolytic virus as well. In this study, a process was accomplished to achieve concentrated and highly purified NDV V4 strain particles. Downstream processing of Newcastle virus strain V4 was characterized by amplifying virus in embryonated chicken eggs. Through a sequence of steps, harvesting allantoic fluid, clarification by centrifuge, concentration by ultrafiltration, and size exclusion separation, the reduced volume and pure virus particles were considered for the amount of ovalbumin, hemagglutinin activity, transmission electron microscopy (TEM), electrophoresis, and additionally immunogenicity of prepared antigens. The results presented a high recovery of HA activity in concentrated and purified virus with the removal of ovalbumin and the typical morphology based on TEM. Sepharose CL-4B was determined as the best media among all used resins to purify the virus. Prepared formulations as vaccines demonstrated positive hemagglutinin inhibition for 6 months and stability for 2 years. Strong evidence from organized studies supports the effectiveness of this method in concentrating and purifying intact NDV, which could be valuable in vaccine research, antiserum preparation, or even as an alternative oncotic agent to traditional methods. Despite further studies being conducted, this method can be utilized particularly on a semi-industrial scale to produce various vaccine components.

Oncotic Agent(s) Coming to the Rescue of "Drowning" Cells?

Oncotic Agent(s) Coming to the Rescue of "Drowning" Cells?

Expanding the horizon: using fatty acids and mannitol as alternative fuels and oncotic agents for normothermic ex vivo heart perfusions in adult rats

Expanding the horizon: using fatty acids and mannitol as alternative fuels and oncotic agents for normothermic ex vivo heart perfusions in adult rats

Low-Fouling Zwitterionic Polymeric Colloids as Resuscitation Fluids for Hemorrhagic Shock.

Colloids, known as volume expanders, have been used as resuscitation fluids for hypovolemic shock for decades, as they increase plasma oncotic pressure and expand intravascular volume. However, recent studies show that commonly used synthetic colloids have adverse interactions with human biological systems. In this work, a low-fouling amine(N)-oxide-based zwitterionic polymer as an alternative volume expander with improved biocompatibility and efficacy is designed. It is demonstrated that the polymer possesses antifouling ability, resisting cell interaction and deposition in major organs, and is rapidly cleared via renal filtration and hepatic circulation, reducing the risk of long-term side effects. Furthermore, in vitro and in vivo studies show an absence of adverse effects on hemostasis or any acute safety risks. Finally, it is shown that, in a head-to-head comparison with existing colloids and plasma, the zwitterionic polymer serves as a more potent oncotic agent for restoring intravascular volume in a hemorrhagic shock model. The design of N-oxide-based zwitterionic polymers may lead to the development of alternative fluid therapies to treat hypovolemic shock and to improve fluid management in general.

Role of PEG35, Mitochondrial ALDH2, and Glutathione in Cold Fatty Liver Graft Preservation: An IGL-2 Approach.

The total damage inflicted on the liver before transplantation is associated with several surgical manipulations, such as organ recovery, washout of the graft, cold conservation in organ preservation solutions (UW, Celsior, HTK, IGL-1), and rinsing of the organ before implantation. Polyethylene glycol 35 (PEG35) is the oncotic agent present in the IGL-1 solution, which is an alternative to UW and Celsior solutions in liver clinical transplantation. In a model of cold preservation in rats (4 °C; 24 h), we evaluated the effects induced by PEG35 on detoxifying enzymes and nitric oxide, comparing IGL-1 to IGL-0 (which is the same as IGL-1 without PEG). The benefits were also assessed in a new IGL-2 solution characterized by increased concentrations of PEG35 (from 1 g/L to 5 g/L) and glutathione (from 3 mmol/L to 9 mmol/L) compared to IGL-1. We demonstrated that PEG35 promoted the mitochondrial enzyme ALDH2, and in combination with glutathione, prevented the formation of toxic aldehyde adducts (measured as 4-hydroxynonenal) and oxidized proteins (AOPP). In addition, PEG35 promoted the vasodilator factor nitric oxide, which may improve the microcirculatory disturbances in steatotic grafts during preservation and revascularization. All of these results lead to a reduction in damage inflicted on the fatty liver graft during the cold storage preservation. In this communication, we report on the benefits of IGL-2 in hypothermic static preservation, which has already been proved to confer benefits in hypothermic oxygenated dynamic preservation. Hence, the data reported here reinforce the fact that IGL-2 is a suitable alternative to be used as a unique solution/perfusate when hypothermic static and preservation strategies are used, either separately or combined, easing the logistics and avoiding the mixture of different solutions/perfusates, especially when fatty liver grafts are used. Further research regarding new therapeutic and pharmacological insights is needed to explore the underlying mitochondrial mechanisms exerted by PEG35 in static and dynamic graft preservation strategies for clinical liver transplantation purposes.

Glycocalyx as a Useful Marker of Endothelial Injury in Liver Transplantation: The Role of Preservation Solution.

The reported investigations from Schiefer et al,1 on the role of glycocalyx (GC) in orthotopic liver transplantation, propose GC as a relevant injury marker to be used in this field. GC is a complex sugar monolayer, which covers the endothelium and is thus exposed and susceptible to alteration as a consequence of liver ischemia-reperfusion (IR) injury associated with orthotopic liver transplantation. The authors have used histidine-tryptophan-ketoglutarate (HTK) solution in their study. However, other preservation solutions (PSs) with different compositions (such as University of Wisconsin [UW], Celsior, and IGL-1 PS) are commercially available and should be tested. Extension of the study with use of these alternative PS could provide important insights into organ PS. The presence of an oncotic agent (such as hydroxyethyl starch in the UW solution or polyethylene glycol 35 [PEG35] in IGL-1 solution) differentiates UW and IGL-1 from HTK and Celsior solutions, which do not contain oncotic agents. This difference is relevant because oncotic pressure, controlled by the oncotic agent, minimizes interstitial edema, which could directly affect GC. However, the oncotic agent increases viscosity, which may in turn affect the protective performance of the PS. Balancing the limitation of edema against viscosity during organ preservation is critical to GC integrity and graft status. In this context, our group reported that PEG35 used in a rinsing solution reduces liver graft damage after cold static preservation.2 The comparison of 2 PS, HTK, and IGL-1, in a rat liver cold preservation model, showed that the presence of PEG35 in the IGL-1 PS was the determinant of GC protection.3 These data support the concept that PEG35 reduces IR injury effects on the narrowing and the integrity of the GC. GC protection is linked to the mechanical properties that PEG35 confers on PS because of its viscosity, such as shear stress and PEG35 interacting with the cells causing remodeling of the cell’s actin cytoskeleton.4 It is acknowledged that IR injury is maximized in fatty livers partly because of sinusoidal narrowing and is accentuated by both normothermic and hypothermic machine perfusion.5 In addition, nitric oxide, a potent physiological vasodilator, is promoted by PEG35 through endothelial nitric oxide synthase activation.2,3 Therefore, PEG35 should be considered as an important component in any dynamic preservation strategy, such as hypothermic oxygenated perfusion, to preserve GC integrity and improve liver preservation, especially when fatty liver grafts are used.5 Thus, we support the relevance of the investigations reported by Schieffer et al, which point firmly, for the first time, to the utility of GC as a suitable marker of graft damage and preservation in orthotopic liver transplantation. Further investigation will be important to compare GC as an injury marker using different PS.

Polyethylene Glycol 35 as a Perfusate Additive for Mitochondrial and Glycocalyx Protection in HOPE Liver Preservation.

Organ transplantation is a multifactorial process in which proper graft preservation is a mandatory step for the success of the transplantation. Hypothermic preservation of abdominal organs is mostly based on the use of several commercial solutions, including UW, Celsior, HTK and IGL-1. The presence of the oncotic agents HES (in UW) and PEG35 (in IGL-1) characterize both solution compositions, while HTK and Celsior do not contain any type of oncotic agent. Polyethylene glycols (PEGs) are non-immunogenic, non-toxic and water-soluble polymers, which present a combination of properties of particular interest in the clinical context of ischemia-reperfusion injury (IRI): they limit edema and nitric oxide induction and modulate immunogenicity. Besides static cold storage (SCS), there are other strategies to preserve the organ, such as the use of machine perfusion (MP) in dynamic preservation strategies, which increase graft function and survival as compared to the conventional static hypothermic preservation. Here we report some considerations about using PEG35 as a component of perfusates for MP strategies (such as hypothermic oxygenated perfusion, HOPE) and its benefits for liver graft preservation. Improved liver preservation is closely related to mitochondria integrity, making this organelle a good target to increase graft viability, especially in marginal organs (e.g., steatotic livers). The final goal is to increase the pool of suitable organs, and thereby shorten patient waiting lists, a crucial problem in liver transplantation.

Aldehyde Dehydrogenase 2 (ALDH2) in Rat Fatty Liver Cold Ischemia Injury.

Institut George Lopez-1 (IGL-1) and Histidine-tryptophan-ketoglutarate (HTK) solutions are proposed as alternatives to UW (gold standard) in liver preservation. Their composition differs in terms of the presence/absence of oncotic agents such as HES or PEG, and is decisive for graft conservation before transplantation. This is especially so when fatty (steatotic) livers are used since these grafts are more vulnerable to ischemia insult during conservation. Their composition determines the extent of the subsequent reperfusion injury after transplantation. Aldehyde dehydrogenase-2 (ALDH2), a mitochondrial enzyme, has been reported to play a protective role in warm ischemia-reperfusion injury (IRI), but its potential in fatty liver cold ischemic injury has not yet been investigated. We evaluated the relevance of ALDH2 activity in cold ischemia injury when fatty liver grafts from Zucker Obese rats were preserved in UW, HTK, and IGL-1 solutions, in order to study the mechanisms involved. ALDH2 upregulation was highest in livers preserved in IGL-1. It was accompanied by a decrease in transaminases, apoptosis (Caspase 3 and TUNEL assay), and lipoperoxidation, which was concomitant with the effective clearance of toxic aldehydes such as 4-hydroxy-nonenal. Variations in ATP levels were also determined. The results were consistent with levels of NF-E2 p45-related factor 2 (Nrf2), an antioxidant factor. Here we report for the first time the relevance of mitochondrial ALDH2 in fatty liver cold preservation and suggest that ALDH2 could be considered a potential therapeutic target or regulator in clinical transplantation.

Comparison of IGL-1 Versus UW Ex Vivo Back Table Flush on Early Graft Function after Living Donor Kidney Transplantation

Background The Institut Georges Lopez (IGL-1) preservation solution contains the same buffer, osmotic agents and substances protective against ischemia reperfusion injuries as the University of Wisconsin solution. In contrast to UW solution, in the IGL-1 solution the oncotic agent is replaced through PEG 35 (1g/L) and a different intra/extracellular ionic composition is obtained. The aim of this study was to compare the effect of IGL-1 on early kidney graft function compared to UW solution in our living donor kidney transplantation (LDKT) program. Methods Between January 1, 2014 and August 30, 2017, 95 LDKT were performed at our center. Among them, 62 were flushed with UW (UW group) and 33 with IGL-1 (IGL group) preservation solution before transplantation. Donor and recipient characteristics were compared. Primary endpoints were 1) incidence of delayed graft function and time to reach an estimated glomerular filtration rate (eGFR) >40 ml/min/1.73m2. Secondary endpoints were 1) clinical biopsy proven acute rejection (BPAR) incidence and 2) 6 months and 1 year patient- and graft survival. Results The donor and recipient characteristics were comparable between the UW and the IGL group. No primary non-function or delayed graft function were observed in both study groups. Time to reach eGFR>40 ml/min/1.73m2 after transplantation was comparable in both study groups (p=0.107). Four (6.5%) and 2 (6.2%) patients never reached a GFR>40 ml/min/1.73m2 after LDKT, respectively in the UW versus IGL group (p=0.941). The incidence of BPAR during follow-up was 6 (9.7%) and 2 (6.1%) in the UW and IGL group (p=0.546), respectively. The 6 months and 1-year patient survivals were 100% in both study groups. Six months- and 1 year death-censored graft survivals were 100% and 98% versus 100% and 100%, respectively in the UW and IGL group (p=0.590). Discussion This series is the first in literature that demonstrates comparable results on early graft function using IGL-1 perfusion solution versus UW solution in a LDKT program. Conclusion Ex vivo flush and subsequent short static cold storage preservation by IGL-1 perfusion solution demonstrates comparable early graft function in our LDKT program compared to UW perfusion solution.

Low-Volume Resuscitation for Hemorrhagic Shock: Understanding the Mechanism of PEG-20k.

Hemorrhagic shock leads to cell and tissue swelling and no reflow from compressed capillaries. Cell impermeants, including polyethylene glycol-20,000 (PEG-20k), reverse ischemia-induced cell swelling, extend low-volume resuscitation (LVR) time after shock, and increase tolerance to the low-volume state. The purpose of this study was to explore the mechanisms of action of PEG-20k containing LVR solutions. We hypothesized that PEG-20k acts as both an oncotic agent and an impermeant in the microcirculation, which moves water out of the space and into the capillaries to affect peripheral capillary filling and enhanced perfusion during the low-volume state. Rats were hemorrhaged until arterial lactate reached 9-10 mM/liter. Then, saline-based LVR solutions containing various impermeant materials were administered (10% blood volume). The LVR times for these solutions were determined by measuring the amount of time required for plasma lactate to climb back to 9 to 10 mM after LVR administration (low-volume tolerance). Capillary blood flow was measured by colored microspheres, and blood volume was measured by fluorescein isothiocyanate-labeled albumin dilution. Gluconate (impermeant), albumin (colloid), and PEG-20k (hybrid) increased LVR time over saline by 4-, 3-, and 8-fold, respectively. The combination of impermeant + albumin produced a biologic effect that was similar to PEG-20k alone. Capillary blood flow and plasma volume were decreased after shock with saline LVR but increased with PEG-20k, relative to saline. These data are consistent with the hypothesis that PEG-20k may act by establishing multiple osmotic gradients in the microcirculation to drive cell-to-capillary water transfer during hypovolemic shock.

Innovative Pharmacological/Therapeutic Approaches against Hepatic Ischemia/Reperfusion Injury.

Prolonged oxygen deprivation corresponding to the ischemic period is observed during liver resection, transplantation, and trauma and subsequent oxygen restoration always leads to reperfusion injury. In particular, the ischemic period represents an inevitable event during conventional organ preservation before transplantation and the magnitude and severity of reperfusion injury depend on the time of liver preservation. Faced with the increasing shortage of donor organs for transplantation, there is renewed interest in marginal livers, such as those obtained from donation after circulatory death or fatty livers, which are particularly susceptible to ischemia/reperfusion (I/R) injury. The decreased tolerance towards I/R observed in fatty livers has not been fully elucidated yet, and underlying mechanisms appear different in lean and steatotic livers. The paper of A. Matsuda et al. proposed the use of activated protein C (APC), an anticoagulant, found to induce an initial attenuation of tissue damage by inhibiting inflammatory cell infiltration and sinusoidal endothelial injury in normal liver. Of interest, in steatotic mice livers, APC did not affect initial liver damage but attenuated late damage, suggesting the existence of an additional pathway to the anti-inflammatory cytoprotective effect of APC which may occur via activation of adenosine monophosphate-activated protein kinase (AMPK) phosphorylation. A further attempt to reduce the fatty liver susceptibility was reported by E. Pantazi et al. that used a peroxisome proliferator-activated receptor α (PPARα) agonist WY-14643. PPARα is a nuclear receptor highly expressed in liver and it functions as a lipid sensor. Treatment with WY-14643 reduced liver injury in fatty rat livers, enhanced the deacetylase enzyme sirtuin 1 (SIRT1), recently found to be a target for preventing I/R, and prevented endoplasmic reticulum stress. M. Bejaoui et al. investigated the use of polyethylene glycols (PEGs) that, besides their usefulness as oncotic agents in preservation solution, have been shown to protect against cold injury and ischemic damage. The intravenous PEG 35 administration by a unique dose protected steatotic rat liver grafts against the deleterious effects of cold storage and the subsequent reperfusion. The rationale for the intravenous administration of PEG 35 was to induce a pharmacological preconditioning effect; its protective mechanisms are related to preservation of mitochondria and the induction of protective cell signaling pathways (eNOS, Akt, and AMPK). An additional model of liver preconditioning by pharmacological induction of adenosine A2a receptor (A2aR) was reported by E. Alchera et al. The analysis of the molecular changes induced by A2aR stimulation revealed multiple mechanisms of liver cell protection that can be both immediate (early preconditioning) and delayed (late preconditioning). Note that the A2aR activation also protects against lipotoxicity by preventing lipoapoptosis in primary rat hepatocytes. Among the hepatic mechanisms involved in the development of I/R, mitochondrial dysfunction represents a crucial cellular event contributing to I/R injury. As reported by K. Go et al., the elimination of abnormal and dysfunction mitochondria by mitophagy appears to increase both the quality of the mitochondria and cell survival. Although the mechanisms underlying the onset and propagation of mitophagy remain elusive, this event represents an early adaptive response to facilitate cell survival during I/R. Articles published in this issue not only help to identify new pharmacological approaches for improving liver function during ischemia/reperfusion damage but also stimulate the identification of molecular mediators of hepatic injury. Mariapia Vairetti Hartmut Jaeschke Diethard Monbaliu Jae-Sung Kim Andrea Ferrigno

The Isolated Perfused Rat Kidney: A Technical Update

The isolated perfused kidney is commonly utilized as a screening tool for renal clearance and metabolism, and to correlate renal drug deposition to renal function. Here, we report on several aspects of this procedure that will facilitate a higher experimental success rate and lead to a reduction in animal use. First, we investigated the utility of inulin and creatinine as commonly used markers to measure glomerular filtration rate. For inulin, in the presence of either 20 mM glucose or 4.5% dextran in the buffer, significant interference was observed using an anthrone-based colorimetric assay. These findings suggest that caution needs to be exercised when using glucose or dextran and when inulin is quantitated using this method. Under these circumstances the use of alternative methods of inulin quantitation such as fluorescently tagged inulin is preferred. Second, we optimized bovine serum albumin (BSA) and BSA/dextran compositions that are routinely recommended as oncotic agents in the perfusion buffer and found that a 4% BSA/1.67% dextran composition had the best viability of kidney biomarkers in accordance with recommended threshold parameters. These considerations will be of particular relevance to researchers utilizing the isolated perfused kidney as a screening tool to measure renal biology and drug metabolism as well as applications to investigate diabetic nephropathy.

Risk Factors for Depressive Illness among Elderly Gopd Attendees at Upth

Depression among the elderly is associated with very high morbidity and suicide rates. In view of the fact that this cohort of patients often have concomitant psychosocial and biomedical health concerns it would be of interest to determine the risk factors for developing this debilitating illness. Objective: To determine the risk factors for depressive illness among elderly patients attending the GOPD of UPTH. Materials and Methods: A descriptive cross-sectional study was conducted among 150 elderly GOPD attendees at UPTH, Nigeria. The respondents completed a structured interviewer-administered questionnaire containing the Geriatric Depression Scale (a depression screening tool). Results: The male to female ratio was 1:1.5. Most of the patients fell into the 65-69 year age group. The men were significantly more educated than the women. 42 (28%) of the patients were found to be depressed. 34 (80.9%) of them were female. This difference was statistically significant. In addition, older age groups, urban dwelling (especially with children), poor family support, marital disharmony, widowhood, bereavement, poor educational background and low social status were found to be risk factors for depressive illness in the elderly. Prior history of depression, stressful life events, regular usage of antidepressant drugs, antihypertensives, antidiabetic and anti oncotic agents were also found to be risk factors. Ironically, unemployment and living alone were not found to be risk factors.

Renal excretion and accumulation kinetics of 2-methylbenzoylglycine in the isolated perfused rat kidney.

The effect of protein binding on kidney function has been studied by investigating the renal accumulation and secretion of the hippurate analogue 2-methylbenzoylglycine in the isolated perfused rat kidney in the absence and presence of bovine serum albumin (BSA). Experiments were performed with either 2.5% pluronic or a combination of 2.2% pluronic and 2% BSA as oncotic agents; a wide concentration range (1-190 micrograms mL-1) of 2-methylbenzoylglycine was studied. Tubular secretion appeared to be a function of the amount of unbound drug in the perfusate and was best described by a model consisting of a high and low affinity Michaelis-Menten term. Parameters obtained after the analysis of renal excretion data were maximum transport velocity for the high affinity site (TM,H) = 3.0 +/- 2.8 micrograms min-1, Michaelis-Menten constant for tubular transport for the high affinity site (KT.H) = 0.5 +/- 0.8 microgram mL-1, maximum transport velocity for the low affinity site (TM,L) = 250 +/- 36 micrograms min-1, and Michaelis-Menten constant for tubular transport for the low affinity site (KT,L) = 62 +/- 17 micrograms mL-1. The compound accumulated extensively in kidney tissue, ratios up to 175 times the perfusate concentration were reached. Accumulation data were best analysed by a two-site model similar to the model used to describe renal excretion. Calculated parameters were theoretical maximum capacity of the high affinity site (RM,H) = 26 +/- 23 micrograms g-1, affinity constant for renal accumulation at the high affinity site (KA,H) = 0.2 +/- 0.4 microgram mL-1, theoretical maximum capacity of the low affinity site (RM,L) = 1640 +/- 1100 micrograms g-1 and affinity constant for renal accumulation at the low affinity site (KA,L) = 60 +/- 58 micrograms mL-1. The very high accumulation in kidney tissue could be explained by active tubular uptake, mediated by the secretory mechanisms involved, and dependent on the amount of free drug in the perfusate. This study shows that anionic drugs, subject to active secretion, may reach high concentrations in tubular cells even at low plasma concentrations.

Disposition of 4-methylbenzoylglycine in rat isolated perfused kidney and effects of hippurates on renal mitochondrial metabolism.

Hippurates tend to accumulate within proximal tubule cells during renal secretion. High intracellular concentrations can alter proximal tubular function or lead to tubular toxicity. In this study we examined the renal disposition of the hippurate 4-methylbenzoylglycine, a compound known for its high renal intrinsic clearance in-vivo. The effect of intracellular accumulation on mitochondrial respiration was also measured in-vitro and compared with that of the 2-methyl and 4-amino analogues. Experiments were performed with either 2.5% pluronic or a combination of 2.2% pluronic and 2% bovine serum albumin (BSA) as oncotic agents. Within the concentration range studied (1-200 microg mL(-1)) tubular secretion seemed to be a function of the amount of unbound drug in the perfusate. Renal excretion data were best fitted by a model in which a Michaelis-Menten term was used to describe active secretion. Parameters obtained after the analysis of renal excretion data were the maximum transport velocity (TM = 55+/-2 microg min(-1)) and the Michaelis-Menten constant for tubular transport (KT = 4.2+/-0.8 microg mL(-1)). The compound accumulated extensively in kidney tissue, ratios up to 600 times the perfusate concentration were reached. Accumulation could be explained by active tubular uptake and data were analysed best by a model similar to the model used to describe renal excretion. Calculated parameters were theoretical maximum capacity (RM =300+/-210 microg g(-1)) and affinity constant for renal accumulation (KA = 5.0+/-4.4 microg mL(-1)). The high intracellular concentrations of 4-methylbenzoylglycine had no effect on kidney function and mitochondrial oxygen consumption. The 2-methyl analogue reduced mitochondrial respiration slightly, but 4-aminobenzoylglycine (p-aminohippurate) caused a significant reduction. In conclusion, this study shows that renal accumulation of a hippurate is determined by the efficiency of its tubular secretion. Whether the high intracellular concentrations affect tubular cell functioning depends on the analogue involved.

Comparing the effects of Dextran 70 and Hydroxyethyl starch in an intestinal storage solution

Introduction Our lab has developed a novel strategy for intestinal preservation involving the intraluminal delivery of a nutrient-rich preservation solution. The aim of this study was to compare the effectiveness of two impermeant agents for use in our solution: Dextran 70 (D70; Mw = 70 kDa) and Hydroxyethyl starch (HES; Mw = 2200 kDa). Methods Rat intestines were procured, including an intravascular flush with University of Wisconsin solution followed by a ‘backtable’ intraluminal flush with: UW solution (group 1, UW), or an amino acid-based nutrient-rich preservation solution (AA solution) containing either 5% D70 (group 2, AA-D70) or HES (group 3, AA-HES). Tissue samples ( n = 6) were taken at 2, 4, 8, and 12 h cold storage; histology, energetic, end-product, and oxidative parameters were assessed. In separate groups ( n = 4), D70 and HES were fluorescently labeled with fluorescein isothiocyanate (FITC) in order to directly observe mucosal penetration of the starch and dextran. Results Over the 12 h storage time-course, direct visualization of the fluorescently labeled D70 showed penetration of the mucosal layer as early as 2 h and progressively continued to do so throughout the 12 h period. In contrast, HES did not cross the mucosal barrier and remained captive within the lumen. As time of storage progressed, grade of injury increased in all groups, however, at 4 and 12 h the AA-HES treated tissues exhibited significantly less injury compared to UW and AA-D70, P < 0.05. AA-HES group showed on moderate villus clefting (median grade 2; P < 0.05) while the AA-D70 group exhibited complete villus denudation (grade 4) and the UW group had extensive injury into the regenerative cryptal regions (grade 6). Metabolic parameters revealed a preferential maintenance of ATP and Energy Charge; increases in lactate, alanine and ammonium supported the involvement of aerobic and anaerobic pathways for energy production. Conclusion The results of this study challenge the idea that oncotic support is not a fundamental requirement of static organ storage. Furthermore, our data suggests that HES is an effective oncotic agent for use in our intraluminal nutrient-rich preservation solution, while Dextran 70 is not.

Optimizing the concentration of hydroxyethylstarch in a novel intestinal-specific preservation solution

Introduction Our lab has developed an effective nutrient-rich solution that facilitates energy production and control of oxidative stress during static cold storage of the intestine; however, the requirement for oncotic agents, such as hydroxyethylstarch (HES), has not been evaluated. This study investigated the effectiveness and requirement for HES in an intraluminal preservation solution during a clinically relevant period of cold storage. Methods Rat intestines were procured, including an intravascular flush with University of Wisconsin solution followed by a ‘back table’ intraluminal flush with a nutrient-rich preservation solution containing varying amounts of HES ( n = 6 per group): Group 1, 0%; Group 2, 2.5%; Group 3, 5%; Group 4, 10%. Energetics, oxidative stress, and morphology were assessed over a 24 h time-course of cold storage. Results Overall, the 5% HES solution, Group 3, demonstrated superior energetic status (ATP and total adenylates) compared to all groups, P < 0.05. Malondialdehyde levels indicated a reduction in oxidative stress in Groups 3 and 4 ( P < 0.05). After 12 h, median modified Parks’ grades for Groups 2 and 3 were significantly lower than Groups 1 and 4, P < 0.05. Conclusion Our data suggests that when employing an intraluminal preservation solution for static organ storage, oncotic support is a fundamental requirement; 5% HES is optimal.

Importance of Organ Preservation Solution Composition in Reducing Myocardial Edema during Machine Perfusion for Heart Transplantation

Objective Machine perfusion preservation has been used experimentally to extend the storage interval of donor hearts. We previously demonstrated that machine perfusion with glucose-supplemented Celsior preservation solution led to superior reperfusion function but resulted in increased myocardial edema compared with conventional static preservation. We hypothesized that other solutions that contain an oncotic agent, such as University of Wisconsin Machine Perfusion Solution (UWMPS), might reduce graft edema development while maintaining myocardial oxidative metabolism during long-term storage. Methods Canine hearts were stored and perfused in a perfusion preservation device (LifeCradle; Organ Transport Systems) after cardioplegic arrest and donor cardiectomy. Hearts were perfused either with glucose-supplemented Celsior (which lacks an oncotic agent) or UWMPS (which contains hydroxyethyl starch) at 5°C in the perfusion device over 10 hours. Oxygen consumption (MVO 2), lactate accumulation, regional flow distribution, and myocardial water content were measured. Results Hearts in both groups continued to extract oxygen over the entire perfusion interval. Lactate accumulation was minimal in both groups. Both solutions delivered perfusate evenly to all regions of myocardium. Heart weight increase (Celsior 31.3 ± 4.3%, UWMPS −3.3 ± 1.9%) and final myocardial water content (Celsior 80.2 ± 1.3%, UWMPS 75.9 ± 0.3%) were higher in the Celsior group ( P < .005). Conclusions Donor hearts can be supported by a perfusion device over relatively extended storage intervals. These organs continue to undergo oxidative metabolism with little lactate accumulation. An oncotic agent appears to be important in limiting increases in myocardial water content. UWMPS appears to be superior for perfusion preservation of myocardium by reducing edema development during storage.

Isolation and Physicochemical Characterization of Assam Bora Rice Starch for Use as a Plasma Volume Expander

Water soluble polysaccharides are most effective oncotic agents which are used for treatment of intravascular volume deficiency. Nowadays, they are used as basic material for plasma volume expander. Plasma volume expander based on starch has lower tendency to remain in any major organ of body in comparison to other plasma volume expander. Branched component of starch amylopectin is very similar in structure to glycogen, the reserve polysaccharides of animal; for all this reason starch is compatible with body tissues. Physicochemical properties of raw starch and amylopectin, isolated from Assam Bora rice were characterized for use as plasma volume expander. Characterization involves the determination of ash value, weight average molecular mass, viscosity and resistance towards enzymatic (amylase) hydrolysis. Amylose content was almost negligible. The X-ray diffraction pattern of Assam Bora rice starch was typically A type. High degree of crystallinity of Assam Bora rice starch reflects its resistance towards enzymatic hydrolysis which is of therapeutic advantage for using it as a plasma volume expander.

The Use of 23.4% Hypertonic Saline for the Management of Elevated Intracranial Pressure in Patients With Severe Traumatic Brain Injury: A Pilot Study

Oncotic agents are a therapeutic mainstay for the management of intracranial hypertension. Both mannitol and varied concentrations of hypertonic saline (HTS) have been shown to be effective at reducing elevated intracranial pressure (ICP). We compared the safety and efficacy of 23.4% HTS to mannitol for acute management of elevated ICP after traumatic brain injury (TBI). After approval from our institutional review board, the records of patients admitted with severe TBI who received mannitol or HTS were reviewed. Demographic and physiologic data were recorded. ICP, cerebral perfusion pressure, reduction of ICP after dose administration, serum sodium, osmolality, and magnitude of dose response during the subsequent 60 minutes were analyzed. Efficacy was determined by comparison of proportion of patients with any response and mean change in ICP after dosing with either agent. Safety was determined by recording any new postinfusion electrolyte or neurologic anomalies. Data were compared using chi2 test, accepting p < 0.05 as significant. Twenty-two patients with severe TBI received 210 doses of either mannitol or HTS. All patients suffered severe blunt injury (mean Injury Severity Score 28 +/- 11). HTS patients had a significantly higher ICP at the initiation of therapy than that of mannitol group (30.7 +/- 7.94 mm Hg vs. 28.3 +/- 8.07 mm Hg, respectively). There was no difference in initial cerebral perfusion pressure. Mean ICP reduction in the hour after administration of 102 doses of mannitol and 108 doses of HTS was greater for patients receiving HTS (9.3 +/- 7.37 mm Hg vs. 6.4 +/- 6.57 mm Hg, respectively; p = 0.0028, chi2). More patients responded to HTS (92.6% HTS vs. 74% mannitol; p = 0.0002, chi2). There was no significant difference between groups in the duration of ICP reduction after dose administration (4.1 hours vs. 3.8 hours, respectively). No adverse events after administration of either agent were identified. Based on this retrospective analysis, 23.4% HTS is more efficacious than mannitol in reducing ICP. If these results are confirmed in a prospective, randomized study, 23.4% HTS may become the agent of choice for the management of elevated ICP after TBI.