Intra-renal microcirculatory alterations on non-traumatic hemorrhagic shock induced acute kidney injury in pigs.

Acute kidney injury (AKI) is frequently seen in patients with hemorrhagic shock due to hypotension, tissue hypoxia, and inflammation despite adequate resuscitation. There is a lack of information concerning the alteration of renal microcirculation and perfusion during shock and resuscitation. The aim of this study was to investigate the possible role of renal microcirculatory alterations on development of renal dysfunction in a pig model of non-traumatic hemorrhagic shock (HS) induced AKI.Fully instrumented female pigs were divided into the two groups as Control (n = 6) and HS (n = 11). HS was achieved by withdrawing blood until mean arterial pressure (MAP) reached around 50 mmHg. After an hour cessation period, fluid resuscitation with balanced crystalloid was started for the duration of 1 h. The systemic and renal hemodynamics, renal microcirculatory perfusion (contrast-enhanced ultrasound (CEUS)) and the sublingual microcirculation were measured.CEUS peak enhancement was significantly increased in HS during shock, early-, and late resuscitation indicating perfusion defects in the renal cortex (p < 0.05 vs. baseline, BL) despite a stable renal blood flow (RBF) and urine output. Following normalization of systemic hemodynamics, we observed persistent hypoxia (high lactate) and high red blood cell (RBC) velocity just after initiation of resuscitation resulting in further endothelial and renal damage as shown by increased plasma sialic acid (p < 0.05 vs. BL) and NGAL levels. We also showed that total vessel density (TVD) and functional capillary density (FCD) were depleted during resuscitation (p < 0.05).In this study, we showed that the correction of systemic hemodynamic variables may not be accompanied with the improvement of renal cortical perfusion, intra-renal blood volume and renal damage following fluid resuscitation. We suggest that the measurement of renal injury biomarkers, systemic and renal microcirculation can be used for guiding to the optimization of fluid therapies.

Sublingual Microcirculatory Evaluation of Extracorporeal Hemoadsorption with CytoSorb® in Abdominal Sepsis: A Case Report.

Cytokemia is associated with microcirculatory alterations often with persistent loss of coherence between the micro- and macrocirculation, linked to organ failure and poor outcome of septic patients. Addition of a hemoadsorbant filter to an extracorporeal circuit next to conventional treatment of septic shock results in the hematological clearance of cytokines, hypothetically leading to normalization of the microcirculation and thus organ perfusion. Bedside sublingual microcirculatory assessment using handheld vital microscopy allows real-time direct visualization of the microcirculation and its response to therapy. This is demonstrated in the present case report of an 83-year-old man admitted to our intensive care unit after surgical repair of a colonic perforation for fecal soiling after a low anterior resection for a rectum carcinoma, with leakage of bowel content at the resection site. The clinical course of this patient can be described as having undergone adequate surgical treatment taking away the source of the disease, followed by optimal support including antibiotic treatment in the ICU. However, during the course of his stay in the ICU, his condition deteriorated with symptoms consistent with septic shock. Our report shows that the addition of a hemoadsorbent (CytoSorb) to the continuous renal replacement therapy circuit was associated with an improvement in the condition of our severely ill patient with abdominal sepsis. Parallel to the clinical improvement of our patient, the functional parameters of the microcirculation also showed improvement suggesting that such a noninvasive real-time evaluation of the status of the microcirculation may be a sensitive diagnostic tool to monitor the effectiveness of hemoadsorbent therapy.

Increased Hepatic Microvascular Density, Oxygenation, and VEGF in the Hypertrophic Lobe following Portal Vein Embolization in Rabbits.

INTRODUCTION: The microvascular events following portal vein embolization (PVE) are poorly understood despite the pivotal role of the microcirculation in liver regeneration and tumor progression. We aimed to assess the changes in hepatic microvascular perfusion and neo-angiogenesis after experimental PVE. METHODS: PVE of the cranial liver lobes was performed in 12 New Zealand White rabbits divided into 2 groups of permanent (P-PVE) and reversible PVE (R-PVE), respectively. Hepatobiliary scintigraphy and CT were used to evaluate hepatic function and volume. Hepatic microcirculation was assessed using a handheld vital microscope (Cytocam) to measure microvascular density (total vessel density; TVD) before PVE, right after PVE, and 20 min after PVE, as well as at 14 days (D14 post-PVE) and 35 days (D35 post-PVE). Additionally, on D35, microvascular PO2 and liver parenchymal VEGF were assessed. RESULTS: Eleven rabbits were included after PVE (R-PVE, n = 5; P-PVE, n = 6). TVD in the nonembo-lized (hypertrophic) lobes was higher than in the embolized (atrophic) lobes of the P-PVE group at D35 post-PVE (36.7 ± 7.2 vs. 23.4 ± 4.9 mm/mm2; p < 0.05). In the R-PVE group, TVD in the nonembolized lobes was not increased at D35. Function and volume were increased in the nonembolized lobes of the P-PVE group compared to the embolized lobes, but not in the R-PVE group. Likewise, the mmicrovascular PO2 and VEGF staining rate were higher in the nonembolized lobes of the P-PVE group at D35 post-PVE. DISCUSSION/CONCLUSION: Successful volumetric and functional hypertrophy of the nonembolized lobe was accompanied by microvascular alterations featuring increased neo-angiogenesis, microvascular density, and microvascular oxygen pressure following P-PVE.

Capillary Leukocytes, Microaggregates, and the Response to Hypoxemia in the Microcirculation of Coronavirus Disease 2019 Patients.

OBJECTIVES: In this study, we hypothesized that coronavirus disease 2019 patients exhibit sublingual microcirculatory alterations caused by inflammation, coagulopathy, and hypoxemia. DESIGN: Multicenter case-controlled study. SETTING: Two ICUs in The Netherlands and one in Switzerland. PATIENTS: Thirty-four critically ill coronavirus disease 2019 patients were compared with 33 healthy volunteers. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: The microcirculatory parameters quantified included total vessel density (mm × mm-2), functional capillary density (mm × mm-2), proportion of perfused vessels (%), capillary hematocrit (%), the ratio of capillary hematocrit to systemic hematocrit, and capillary RBC velocity (µm × s-1). The number of leukocytes in capillary-postcapillary venule units per 4-second image sequence (4 s-1) and capillary RBC microaggregates (4 s-1) was measured. In comparison with healthy volunteers, the microcirculation of coronavirus disease 2019 patients showed increases in total vessel density (22.8 ± sd 5.1 vs 19.9 ± 3.3; p < 0.0001) and functional capillary density (22.2 ± 4.8 vs 18.8 ± 3.1; p < 0.002), proportion of perfused vessel (97.6 ± 2.1 vs 94.6 ± 6.5; p < 0.01), RBC velocity (362 ± 48 vs 306 ± 53; p < 0.0001), capillary hematocrit (5.3 ± 1.3 vs 4.7 ± 0.8; p < 0.01), and capillary-hematocrit-to-systemic-hematocrit ratio (0.18 ± 0.0 vs 0.11 ± 0.0; p < 0.0001). These effects were present in coronavirus disease 2019 patients with Sequential Organ Failure Assessment scores less than 10 but not in patients with Sequential Organ Failure Assessment scores greater than or equal to 10. The numbers of leukocytes (17.6 ± 6.7 vs 5.2 ± 2.3; p < 0.0001) and RBC microaggregates (0.90 ± 1.12 vs 0.06 ± 0.24; p < 0.0001) was higher in the microcirculation of the coronavirus disease 2019 patients. Receiver-operating-characteristics analysis of the microcirculatory parameters identified the number of microcirculatory leukocytes and the capillary-hematocrit-to-systemic-hematocrit ratio as the most sensitive parameters distinguishing coronavirus disease 2019 patients from healthy volunteers. CONCLUSIONS: The response of the microcirculation to coronavirus disease 2019-induced hypoxemia seems to be to increase its oxygen-extraction capacity by increasing RBC availability. Inflammation and hypercoagulation are apparent in the microcirculation by increased numbers of leukocytes and RBC microaggregates.

Resuscitation with PEGylated carboxyhemoglobin preserves renal cortical oxygenation and improves skeletal muscle microcirculatory flow during endotoxemia.

PEGylated carboxyhemoglobin (PEGHbCO), which has carbon monoxide-releasing properties and plasma expansion and oxygen-carrying properties, may improve both skeletal microcirculatory flow and renal cortical microcirculatory Po2 (CµPo2) and, subsequently, limit endotoxemia-induced acute kidney injury. Anesthetized, ventilated Wistar albino rats (n = 44) underwent endotoxemic shock. CµPo2 was measured in exposed kidneys using a phosphorescence-quenching method. Rats were randomly assigned to the following five groups: 1) unresuscitated lipopolysaccharide (LPS), 2) LPS + Ringer's acetate (RA), 3) LPS + RA + 0.5 µg·kg·-1min-1 norepinephrine (NE), 4) LPS + RA + 320 mg/kg PEGHbCO, and 5) LPS + RA + PEGHbCO + NE. The total volume was 30 mL/kg in each group. A time control animal group was used. Skeletal muscle microcirculation was assessed by handheld intravital microscopy. Kidney immunohistochemistry and myeloperoxidase-stained leukocytes in glomerular and peritubular areas were analyzed. Endotoxemia-induced histological damage was assessed. Plasma levels of IL-6, heme oxygenase-1, malondialdehyde, and syndecan-1 were assessed by ELISA. CµPo2 was higher in the LPS + RA + PEGHbCO-resuscitated group, at 35 ± 6mmHg compared with 21 ± 12 mmHg for the LPS+RA group [mean difference: -13.53, 95% confidence interval: (-26.35; -0.7156), P = 0.035]. The number of nonflowing, intermittent, or sluggish capillaries was smaller in groups infused with PEGHbCO compared with RA alone (P < 0.05), while the number of normally perfused vessels was greater (P < 0.05). The addition of NE did not further improve CµPo2 or microcirculatory parameters. Endotoxemia-induced kidney immunohistochemistry and histological alterations were not mitigated by PEGHbCO 1 h after resuscitation. Renal leukocyte infiltration and plasma levels of biomarkers were similar across groups. PEGHbCO enhanced CµPo2 while restoring skeletal muscle microcirculatory flow in previously nonflowing capillaries. PEGHbCO should be further evaluated as a resuscitation fluid in mid- to long-term models of sepsis-induced acute kidney injury.

Leukocyte-Endothelium Interaction in the Sublingual Microcirculation of Coronary Artery Bypass Grafting Patients.

OBJECTIVE: The aim of this study was to apply an innovative methodology to incident dark-field (IDF) imaging in coronary artery bypass grafting (CABG) patients for the identification and quantification of rolling leukocytes along the sublingual microcirculatory endothelium. METHODS: This study was a post hoc analysis of a prospective study that evaluated the perioperative course of the sublingual microcirculation in CABG patients. Video images were captured using IDF imaging following the induction of anesthesia (T0) and cardiopulmonary bypass (CPB) (T1) in 10 patients. Rolling leukocytes were identified and quantified using frame averaging, which is a technique that was developed for correctly identifying leukocytes. RESULTS: The number of rolling leukocytes increased significantly from T0 (7.5 [6.4-9.1] leukocytes/capillary-postcapillary venule/4 s) to T1 (14.8 [13.2-15.5] leukocytes/capillary-postcapillary venule/4 s) (p < 0.0001). A significant increase in systemic leukocyte count was also detected from 7.4 ± 0.9 × 109/L (preoperative) to 12.4 ± 4.4 × 109/L (postoperative) (p < 0.01). CONCLUSION: The ability to directly visualize leukocyte-endothelium interaction using IDF imaging facilitates the diagnosis of a systemic inflammatory response after CPB via the identification of rolling leukocytes. Integration of the frame averaging algorithm into the software of handheld vital microscopes may enable the use of microcirculatory leukocyte count as a real-time parameter at the bedside.

Effect of Polyethylene-glycolated Carboxyhemoglobin on Renal Microcirculation in a Rat Model of Hemorrhagic Shock.

BACKGROUND: Primary resuscitation fluid to treat hemorrhagic shock remains controversial. Use of hydroxyethyl starches raised concerns of acute kidney injury. Polyethylene-glycolated carboxyhemoglobin, which has carbon monoxide-releasing molecules and oxygen-carrying properties, was hypothesized to sustain cortical renal microcirculatory PO2 after hemorrhagic shock and reduce kidney injury. METHODS: Anesthetized and ventilated rats (n = 42) were subjected to pressure-controlled hemorrhagic shock for 1 h. Renal cortical PO2 was measured in exposed kidneys using a phosphorescence quenching method. Rats were randomly assigned to six groups: polyethylene-glycolated carboxyhemoglobin 320 mg · kg, 6% hydroxyethyl starch (130/0.4) in Ringer's acetate, blood retransfusion, diluted blood retransfusion (~4 g · dl), nonresuscitated animals, and time control. Nitric oxide and heme oxygenase 1 levels were determined in plasma. Kidney immunohistochemistry (histologic scores of neutrophil gelatinase-associated lipocalin and tumor necrosis factor-α) and tubular histologic damages analyses were performed. RESULTS: Blood and diluted blood restored renal PO2 to 51 ± 5 mmHg (mean difference, -18; 95% CI, -26 to -11; P < 0.0001) and 47 ± 5 mmHg (mean difference, -23; 95% CI, -31 to -15; P < 0.0001), respectively, compared with 29 ± 8 mmHg for hydroxyethyl starch. No differences between polyethylene-glycolated carboxyhemoglobin and hydroxyethyl starch were observed (33 ± 7 mmHg vs. 29 ± 8 mmHg; mean difference, -5; 95% CI, -12 to 3; P = 0.387), but significantly less volume was administered (4.5 [3.3-6.2] vs. 8.5[7.7-11.4] ml; mean rank difference, 11.98; P = 0.387). Blood and diluted blood increased the plasma bioavailability of nitric oxide compared with hydroxyethyl starch (mean rank difference, -20.97; P = 0.004; and -17.13; P = 0.029, respectively). No changes in heme oxygenase 1 levels were observed. Polyethylene-glycolated carboxyhemoglobin limited tubular histologic damages compared with hydroxyethyl starch (mean rank difference, 60.12; P = 0.0012) with reduced neutrophil gelatinase-associated lipocalin (mean rank difference, 84.43; P < 0.0001) and tumor necrosis factor-α (mean rank difference, 49.67; P = 0.026) histologic scores. CONCLUSIONS: Polyethylene-glycolated carboxyhemoglobin resuscitation did not improve renal PO2 but limited tubular histologic damages and neutrophil gelatinase-associated lipocalin upregulation after hemorrhage compared with hydroxyethyl starch, whereas a lower volume was required to sustain macrocirculation.

Investigating the physiology of normothermic ex vivo heart perfusion in an isolated slaughterhouse porcine model used for device testing and training.

BACKGROUND: The PhysioHeart™ is a mature acute platform, based isolated slaughterhouse hearts and able to validate cardiac devices and techniques in working mode. Despite perfusion, myocardial edema and time-dependent function degradation are reported. Therefore, monitoring several variables is necessary to identify which of these should be controlled to preserve the heart function. This study presents biochemical, electrophysiological and hemodynamic changes in the PhysioHeart™ to understand the pitfalls of ex vivo slaughterhouse heart hemoperfusion. METHODS: Seven porcine hearts were harvested, arrested and revived using the PhysioHeart™. Cardiac output, SaO2, glucose and pH were maintained at physiological levels. Blood analyses were performed hourly and unipolar epicardial electrograms (UEG), pressures and flows were recorded to assess the physiological performance. RESULTS: Normal cardiac performance was attained in terms of mean cardiac output (5.1 ± 1.7 l/min) and pressures but deteriorated over time. Across the experiments, homeostasis was maintained for 171.4 ± 54 min, osmolarity and blood electrolytes increased significantly between 10 and 80%, heart weight increased by 144 ± 41 g, free fatty acids (- 60%), glucose and lactate diminished, ammonia increased by 273 ± 76% and myocardial necrosis and UEG alterations appeared and aggravated. Progressively deteriorating electrophysiological and hemodynamic functions can be explained by reperfusion injury, waste product intoxication (i.e. hyperammonemia), lack of essential nutrients, ion imbalances and cardiac necrosis as a consequence of hepatological and nephrological plasma clearance absence. CONCLUSIONS: The PhysioHeart™ is an acute model, suitable for cardiac device and therapy assessment, which can precede conventional animal studies. However, observations indicate that ex vivo slaughterhouse hearts resemble cardiac physiology of deteriorating hearts in a multi-organ failure situation and signalize the need for plasma clearance during perfusion to attenuate time-dependent function degradation. The presented study therefore provides an in-dept understanding of the sources and reasons causing the cardiac function loss, as a first step for future effort to prolong cardiac perfusion in the PhysioHeart™. These findings could be also of potential interest for other cardiac platforms.

Glycocalyx Degradation Is Independent of Vascular Barrier Permeability Increase in Nontraumatic Hemorrhagic Shock in Rats.

BACKGROUND: Glycocalyx shedding after traumatic hemorrhagic or septic shock, as well as different resuscitation fluids, has been causally linked to increased vascular barrier permeability (VBP) resulting in tissue edema. In nontraumatic hemorrhagic shock (NTHS), it remains questionable whether glycocalyx degradation in itself results in an alteration of VBP. The composition of fluids can also have a modulatory effect on glycocalyx shedding and VBP. We hypothesized that the shedding of the glycocalyx during NTHS has little effect on VBP and that the composition of fluids can modulate these effects. METHODS: Fully instrumented Wistar-albino rats were subjected to a pressure-controlled NTHS (mean arterial pressure of 30 mm Hg) for 60 minutes. Animals were fluid resuscitated with Ringer's acetate, balanced hydroxyethyl starch (HES) solution, or 0.9% normal saline to a mean arterial pressure of 80 mm Hg and compared with shams or nonresuscitated NTHS. Glycocalyx shed products were determined at baseline and 60 minutes after fluid resuscitation. Skeletal muscle microcirculation was visualized using handheld vital microscopy. VBP changes were assessed using plasma decay of 3 fluorescent dyes (40- and 500-kDa dextran and 70-kDa albumin), Evans blue dye exclusion, intravital fluorescence microscopy, and determination of tissue edema (wet/dry weight ratio). RESULTS: All glycocalyx shedding products were upgraded as a result of NTHS. Syndecan-1 significantly increased in NTHS (mean difference, -1668; 95% confidence interval [CI], -2336 to -1001; P < .0001), balanced crystalloid (mean difference, -964.2; 95% CI, -1492 to -436.4; P = .0001), and HES (mean difference, -1030; 95% CI, -1594 to -465.8; P = .0001) groups at the end of the experiment compared to baseline. Hyaluronan levels were higher at the end of the experiment in nonresuscitated NTHS (-923.1; 95% CI, -1216 to -630; P = .0001) and balanced crystalloid (-1039; 95% CI, -1332 to -745.5; P = .0001) or HES (-394.2; 95% CI, -670.1 to -118.3; P = .0027) groups compared to controls. Glycocalyx shedding resulted in microcirculation alterations as observed by handheld video microscopy. Total vessel density was altered in the normal saline (mean difference, 4.092; 95% CI, 0.6195-7.564; P = .016) and hemorrhagic shock (mean difference, 5.022; 95% CI, 1.55-8.495; P = .0024) groups compared to the control group, as well as the perfused vessel density and mean flow index. Despite degradation of endothelial glycocalyx, VBP as determined by 4 independent assays remained intact and continued to be so following fluid resuscitation. CONCLUSIONS: NTHS induced glycocalyx shedding and microcirculation alterations, without altering VBP. Fluid resuscitation partially restored the microcirculation without altering VBP. These results challenge the concept that the glycocalyx barrier is a significant contributor to VBP.

Assessment of hepatic microvascular flow and density in patients undergoing preoperative portal vein embolization.

BACKGROUND: The microvascular effects occurring after unilateral preoperative portal vein embolization (PVE) are poorly understood. The aim of this study was to assess the microvascular changes in the embolized and the non-embolized lobes after right PVE. METHODS: Videos of the hepatic microcirculation in patients undergoing right hemihepatectomy following PVE were recorded using a handheld vital microscope (Cytocam) based on incident dark field imaging. Hepatic microcirculation was measured in the embolized and the non-embolized lobes at laparotomy, 3-6 weeks after PVE. The following microcirculatory parameters were assessed: total vessel density (TVD), microcirculatory flow index (MFI), proportion of perfused vessel (PPV), perfused vessel density (PVD), sinusoidal diameter (SinD) and the absolute red blood cell velocity (RBCv). RESULTS: 16 patients after major liver resection were included, 8 with and 8 without preoperative PVE. Microvascular density parameters were higher in the non-embolized lobes when compared to the embolized lobes (TVD: 40.3 ± 8.9 vs. 26.8 ± 4.6 mm/mm2 (p < 0.003), PVD: 40.3 ± 8.8 vs. 26.7 ± 4.7 mm/mm2 (p < 0.002), SinD: 9.2 ± 1.7 vs. 6.3 ± 0.8 µm (p < 0.040)). RBCv, PPV and the MFI were not significantly different. CONCLUSION: The non-embolized lobe has a significantly higher microvascular density, however without differences in microvascular flow. These findings indicate increased angiogenesis in the hypertrophic lobe.

Dynamic Contrast-Enhanced Ultrasound Identifies Microcirculatory Alterations in Sepsis-Induced Acute Kidney Injury.

OBJECTIVES: We developed quantitative methods to analyze microbubble kinetics based on renal contrast-enhanced ultrasound imaging combined with measurements of sublingual microcirculation on a fixed area to quantify early microvascular alterations in sepsis-induced acute kidney injury. DESIGN: Prospective controlled animal experiment study. SETTING: Hospital-affiliated animal research institution. SUBJECTS: Fifteen female pigs. INTERVENTIONS: The animals were instrumented with a renal artery flow probe after surgically exposing the kidney. Nine animals were given IV infusion of lipopolysaccharide to induce septic shock, and six were used as controls. MEASUREMENTS AND MAIN RESULTS: Contrast-enhanced ultrasound imaging was performed on the kidney before, during, and after having induced shock. Sublingual microcirculation was measured continuously using the Cytocam on the same spot. Contrast-enhanced ultrasound effectively allowed us to develop new analytical methods to measure dynamic variations in renal microvascular perfusion during shock and resuscitation. Renal microvascular hypoperfusion was quantified by decreased peak enhancement and an increased ratio of the final plateau intensity to peak enhancement. Reduced intrarenal blood flow could be estimated by measuring the microbubble transit times between the interlobar arteries and capillary vessels in the renal cortex. Sublingual microcirculation measured using the Cytocam in a fixed area showed decreased functional capillary density associated with plugged sublingual capillary vessels that persisted during and after fluid resuscitation. CONCLUSIONS: In our lipopolysaccharide model, with resuscitation targeted at blood pressure, contrast-enhanced ultrasound imaging can identify renal microvascular alterations by showing prolonged contrast enhancement in microcirculation during shock, worsened by resuscitation with fluids. Concomitant analysis of sublingual microcirculation mirrored those observed in the renal microcirculation.

Mycophenolate mofetil improves renal haemodynamics, microvascular oxygenation, and inflammation in a rat model of supra-renal aortic clamping-mediated renal ischaemia reperfusion injury.

Ischaemia/reperfusion (I/R) is one of the main causes of acute kidney injury (AKI), which is characterized by sterile inflammation and oxidative stress. Immune cell activation can provoke overproduction of inflammatory mediators and reactive oxygen species (ROS), leading to perturbation of the microcirculation and tissue oxygenation associated with local and remote tissue injury. This study investigated whether the clinically employed immunosuppressant mycophenolate mofetil (MMF) was able to reduce I/R-induced renal oxygenation defects and oxidative stress by preventing sterile inflammation. Rats were divided into three groups (n=6/group): (1) a sham-operated control group; (2) a group subjected to renal I/R alone (I/R); and (3) a group subjected to I/R and MMF treatment (20 mg/kg prior to I/R) (I/R+MMF). Ischaemia was induced by a vascular occluder placed on the abdominal aorta for 30 minutes, followed by 120 minutes of reperfusion. Renal I/R deteriorated renal oxygenation (P<.001) and oxygen delivery (P<.01), reduced creatinine clearance (P<.01) and tubular sodium reabsorption (P<.001), and increased iNOS, renal tissue injury markers (P<.001), and IL-6 (P<.001). Oral MMF administration prior to insult restored renal cortical oxygenation (P<.05) and iNOS, renal injury markers, and inflammation parameters (P<.001) to near-baseline levels without affecting renal function. MMF exerted a prophylactic effect on renal microvascular oxygenation and abrogated tissue inflammation and renal injury following lower body I/R-induced AKI. These findings may have clinical implications during major vascular or renal transplant surgery.

Effects of a human recombinant alkaline phosphatase on renal hemodynamics, oxygenation and inflammation in two models of acute kidney injury.

Two small clinical trials indicated that administration of bovine intestinal alkaline phosphatase (AP) improves renal function in critically ill patients with sepsis-associated acute kidney injury (AKI), for which the mechanism of action is not completely understood. Here, we investigated the effects of a newly developed human recombinant AP (recAP) on renal oxygenation and hemodynamics and prevention of kidney damage and inflammation in two in vivo AKI models. To induce AKI, male Wistar rats (n=18) were subjected to renal ischemia (30min) and reperfusion (I/R), or sham-operated. In a second model, rats (n=18) received a 30min infusion of lipopolysaccharide (LPS; 2.5mg/kg), or saline, and fluid resuscitation. In both models, recAP (1000U/kg) was administered intravenously (15min before reperfusion, or 90min after LPS). Following recAP treatment, I/R-induced changes in renal blood flow, renal vascular resistance and oxygen delivery at early, and cortical microvascular oxygen tension at late reperfusion were no longer significantly affected. RecAP did not influence I/R-induced effects on mean arterial pressure. During endotoxemia, recAP treatment did not modulate the LPS-induced changes in systemic hemodynamics and renal oxygenation. In both models, recAP did exert a clear renal protective anti-inflammatory effect, demonstrated by attenuated immunostaining of inflammatory, tubular injury and pro-apoptosis markers. Whether this renal protective effect is sufficient to improve outcome of patients suffering from sepsis-associated AKI is being investigated in a large clinical trial.

Blood transfusion improves renal oxygenation and renal function in sepsis-induced acute kidney injury in rats.

BACKGROUND: The effects of blood transfusion on renal microcirculation during sepsis are unknown. This study aimed to investigate the effect of blood transfusion on renal microvascular oxygenation and renal function during sepsis-induced acute kidney injury. METHODS: Twenty-seven Wistar albino rats were randomized into four groups: a sham group (n = 6), a lipopolysaccharide (LPS) group (n = 7), a LPS group that received fluid resuscitation (n = 7), and a LPS group that received blood transfusion (n = 7). The mean arterial blood pressure, renal blood flow, and renal microvascular oxygenation within the kidney cortex were recorded. Acute kidney injury was assessed using the serum creatinine levels, metabolic cost, and histopathological lesions. Nitrosative stress (expression of endothelial (eNOS) and inducible nitric oxide synthase (iNOS)) within the kidney was assessed by immunohistochemistry. Hemoglobin levels, pH, serum lactate levels, and liver enzymes were measured. RESULTS: Fluid resuscitation and blood transfusion both significantly improved the mean arterial pressure and renal blood flow after LPS infusion. Renal microvascular oxygenation, serum creatinine levels, and tubular damage significantly improved in the LPS group that received blood transfusion compared to the group that received fluids. Moreover, the renal expression of eNOS was markedly suppressed under endotoxin challenge. Blood transfusion, but not fluid resuscitation, was able to restore the renal expression of eNOS. However, there were no significant differences in lactic acidosis or liver function between the two groups. CONCLUSIONS: Blood transfusion significantly improved renal function in endotoxemic rats. The specific beneficial effect of blood transfusion on the kidney could have been mediated in part by the improvements in renal microvascular oxygenation and sepsis-induced endothelial dysfunction via the restoration of eNOS expression within the kidney.

The renal microcirculation in sepsis.

Despite identification of several cellular mechanisms being thought to underlie the development of septic acute kidney injury (AKI), the pathophysiology of the occurrence of AKI is still poorly understood. It is clear, however, that instead of a single mechanism being responsible for its aetiology, an orchestra of cellular mechanisms failing is associated with AKI. The integrative physiological compartment where these mechanisms come together and exert their integrative deleterious action is the renal microcirculation (MC). This is why it is opportune to review the response of the renal MC to sepsis and discuss the determinants of its (dys)function and how it contributes to the pathogenesis of renal failure. A main determinant of adequate organ function is the adequate supply and utilization of oxygen at the microcirculatory and cellular level to perform organ function. The highly complex architecture of the renal microvasculature, the need to meet a high energy demand and the fact that the kidney is borderline ischaemic makes the kidney a highly vulnerable organ to hypoxaemic injury. Under normal, steady-state conditions, oxygen (O2) supply to the renal tissues is well regulated; however, under septic conditions the delicate balance of oxygen supply versus demand is disturbed due to renal microvasculature dysfunction. This dysfunction is largely due to the interaction of renal oxygen handling, nitric oxide metabolism and radical formation. Renal tissue oxygenation is highly heterogeneous not only between the cortex and medulla but also within these renal compartments. Integrative evaluation of the different determinants of tissue oxygen in sepsis models has identified the deterioration of microcirculatory oxygenation as a key component in the development AKI. It is becoming clear that resuscitation of the failing kidney needs to integratively correct the homeostasis between oxygen, and reactive oxygen and nitrogen species. Several experimental therapeutic modalities have been found to be effective in restoring microcirculatory oxygenation in parallel to improving renal function following septic AKI. However, these have to be verified in clinical studies. The development of clinical physiological biomarkers of AKI specifically aimed at the MC should form a valuable contribution to monitoring such new therapeutic modalities.

Effects of Crataegus microphylla on vascular dysfunction in streptozotocin-induced diabetic rats.

Vascular dysfunction plays a key role in the pathogenesis of diabetic vascular disease. In this study, we aimed to investigate whether chronic in vivo treatment of Crataegus microphylla (CM) extract in diabetic rats induced with streptozotocin (STZ, intraperitoneal, 65 mg/kg) preserves vascular function and to evaluate whether the reduction of inducible nitric oxide synthase (iNOS), proinflammatory cytokines, and lipid peroxidation mediates its mechanisms of action. Starting at 4 weeks of diabetes, CM extract (100 mg/kg) was administrated to diabetic rats for 4 weeks. In aortic rings, relaxation to acetylcholine and vasoreactivity to noradrenaline were impaired, whereas aortic iNOS expression and plasma tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), total nitrite-nitrate, and malondialdehite levels were increased in diabetic rats compared with controls. Chronic CM treatment significantly corrected all the above abnormalities in diabetic rats. In comparison, pretreatment of the aorta of diabetic rats with N-[3(aminomethyl) benzyl]-acetamidine, dihydrochloride (10(-5) M), a selective inhibitor of iNOS, produced a similar recovery in vascular reactivity. These results suggest that chronic in vivo treatment of CM preserves endothelium-dependent relaxation and vascular contraction in STZ-induced diabetes, possibly by reducing iNOS expression in the aorta and by decreasing plasma levels of TNF-α and IL-6 and by preventing lipid peroxidation.

Herbal haemorrhoidal cream for haemorrhoids.

Although hemorrhoids are one of the most common diseases in the world, the exact etiology underlying the development of hemorrhoids is not clear. Many different ointments are currently used to treat hemorrhoids; however, there is little evidence of the efficacy of these treatments to support their use. The aim of this study was to compare different herbal creams used for the treatment of hemorrhoids. Twenty-eight male Wistar albino rats, 6-8 weeks old and weighing 160-180 g, were used in this study as 1-control, 2-croton oil, 3-croton oil+fig leaves+artichoke leaves+walnut husks and 4-croton oil+fig leaves+artichoke leaves+walnut husks+horse chestnut fruit. After 3 days of croton oil application, rats were treated with 0.1 ml of cream or saline twice a day for 15 days by syringe. Tissue and blood samples were collected for histological, immunohistochemical and biochemical studies. Statistical significance was determined using one-way ANOVA followed by Tukey's multiple comparison tests. Croton oil administration resulted in severe inflammation. The third group showed partial improvement in inflammation; however, the greatest degree of improvement was seen in the fourth group, and some recovered areas were observed. Myeloperoxidase immunoreactivity was found to be decreased in the third and fourth groups compared to the second group. Additionally, biochemical analyses (Myeloperoxidase, Malondyaldehyde, nitrate/nitrite and nitrotyrosine levels and Superoxide Dismutase activity) were in agreement with the histological and immunohistochemical results. In conclusion, croton oil causes inflammation in the anal area and results in hemorrhoids. Treatment with our herbal hemorrhoid creams demonstrated anti-inflammatory and anti-oxidant effects in this model.

Furosemide exacerbated the impairment of renal function, oxygenation and medullary damage in a rat model of renal ischemia/reperfusion induced AKI.

BACKGROUND: Perioperative acute kidney injury (AKI) caused by ischemia-reperfusion (IR) is a significant contributor to mortality and morbidity after major surgery. Furosemide is commonly used in postoperative patients to promote diuresis and reduce tissue edema. However, the effects of furosemide on renal microcirculation, oxygenation and function are poorly understood during perioperative period following ischemic insult. Herein, we investigated the effects of furosemide in rats subjected IR insult. METHODS: 24 Wistar albino rats were divided into 4 groups, with 6 in each; Sham-operated Control (C), Control + Furosemide (C + F), ischemia/reperfusion (IR), and IR + F. After induction of anesthesia (BL), supra-aortic occlusion was applied to IR and IR + F groups for 45 min followed by ongoing reperfusion for 15 min (T1) and 2 h (T2). Furosemide infusion was initiated simultaneously in the intervention groups after ischemia. Renal blood flow (RBF), vascular resistance (RVR), oxygen delivery (DO2ren) and consumption (VO2ren), sodium reabsorption (TNa+), oxygen utilization efficiency (VO2/TNa+), cortical (CµO2) and medullary (MµO2) microvascular oxygen pressures, urine output (UO) and creatinine clearance (Ccr) were measured. Biomarkers of inflammation, oxidative and nitrosative stress were measured and kidneys were harvested for histological analysis. RESULTS: IR significantly decreased RBF, mainly by increasing RVR, which was exacerbated in the IR + F group at T2 (2198 ± 879 vs 4233 ± 2636 dyne/s/cm5, p = 0.07). CµO2 (61.6 ± 6.8 vs 86 ± 6.6 mmHg) and MµO2 (51.1 ± 4.1 vs 68.7 ± 4.9 mmHg, p < 0.05) were both reduced after IR and did not improve by furosemide. Moreover, VO2/TNa+ increased in the IR + F group at T2 with respect to the IR group (IR: 3.3 ± 2 vs IR + F: 8.2 ± 10 p = 0.07) suggesting a possible deterioration of oxygen utilization. Ccr did not change, but plasma creatinine increased significantly in IR + F groups. Histopathology revealed widespread damage both in the cortex and medulla in IR, IR + F and C + F groups. CONCLUSION: Renal microvascular oxygenation, renal function, renal vascular resistance, oxygen utilization and damage were not improved by furosemide administration after IR insult. Our study suggests that furosemide may cause additional structural and functional impairment to the kidney following ischemic injury and should be used with caution.