Sepsis and resulting multiple system organ failure are the leading causes of mortality in intensive care units. Hydroxyethyl starch (HES) 130/0.4 was a novel preparation, developed to improve the pharmacokinetics of current medium molecular weight HES solutions. This study was designed to explore the effects of HES 130/0.4 on pulmonary capillary permeability (PCP), production of cytokines, and activation of transcription factor in septic rats induced by cecal ligation and puncture (CLP).
Adult male Sprague Dawley rats were randomly divided into six groups (six rats/group): saline controls (30 ml/kg); CLP plus saline (30 ml/kg); CLP plus HES (7.5, 15, or 30 ml/kg, respectively), and HES alone (30 ml/kg). Mean arterial blood pressure and heart rate were monitored during the experiment process. Myeloperoxidase (MPO) activity, wet/dry weight ratio, PCP, tumor necrosis factor alpha (TNF-alpha), interleukin (IL)-6, IL-10, and nuclear factor-kappa B (NF-kappaB) were investigated at 6 h.
We demonstrated that CLP could provoke significant injury in lung, characterized by increase in PCP, wet/dry weight ratio, MPO activity, TNF-alpha, IL-6, and IL-10 level, and NF-kappaB activation. Without obvious influence on systemic macro-hemodynamics, HES 15 ml/kg and 30 ml/kg significantly reduced CLP-induced elevation of pulmonary capillary permeability, wet/dry weight ratio, and production of IL-6. Meanwhile, HES 15 ml/kg increased IL-10 level and HES 7.5, 15, and 30 ml/kg suppressed MPO activity, TNF-alpha level, and NF-kappaB activation.
HES 130/0.4 can inhibit CLP-induced PCP by attenuating pulmonary inflammation and NF-kappaB activation in vivo.
Hydroxyethyl starch (HES) is one of the most frequently used plasma substitutes, and could modulate inflammatory response in sepsis. Our aim of this study was to investigate the mechanism of the effect of HES 130/0.4 by studying plasma levels of inflammatory cytokines, nuclear factor-kappaB (NF-kappaB) activation, and Toll-like receptors (TLRs) expression in peripheral monocytes during polymicrobial sepsis.
Rats with sepsis induced by cecal ligation and puncture (CLP) were treated with HES130/0.4 (7.5, 15, or 30 mL/kg, intravenously); then, rat plasma and monocytes were isolated from blood 5 h later. The plasma level of cytokines (tumor necrosis factor [TNF]-alpha and interleukin [IL]-6), NF-kappaB activity, and mRNA and protein levels of TLRs (TLR2 and TLR4) in peripheral blood monocytes were determined by enzyme-linked immunosorbent assay, electrophoretic mobility shift assay, reverse transcription-polymerase chain reaction, and Western blotting, respectively.
HES130/0.4 dose-dependently reduced the plasma level of TNF-alpha and IL-6 in rats with sepsis. HES130/0.4 also significantly inhibited NF-kappaB activation, and TLRs mRNA and protein levels in peripheral monocytes.
During sepsis, HES130/0.4 can down-regulate the inflammatory response, possibly through inhibition of the TLRs/NF-kappaB signaling pathway, and could be one more appropriate plasma substitute in sepsis.
During perioperative management of patients with gastrointestinal cancer complicated by diabetes mellitus, adequate alimentation is required, but we often face difficulties associated with hyperglycemia and other accompanying complications. Recently, we investigated the effects of a novel palatinose based enteral formula (MHN-01) in suppressing post-prandial hyperglycemia and improving lipid metabolism in experimental animals and perioperative management of patients with esophageal cancer complicated by diabetes mellitus.
We gave normal rats and rats with type 2 diabetes mellitus a single oral dose of fluid diet, and analyzed comparatively the time course of blood glucose level in each group until 3 h after the dose. In both the normal rat group and the type 2 diabetes group, peak blood glucose level after the MHN-01 dose was significantly lower than after a dose of ordinary fluid diet and was comparable to the peak level after a dose of a fluid diet rich in MUFA (monounsaturated fatty acid). We allowed normal mice free access to fluid diet for 43 days, and measured their body fat levels. Fat accumulation was significantly lower in mice given MHN-01 than in mice given ordinary fluid diet. We also analyzed the respiratory quotient and resting energy expenditure of normal Sprague-Dawley rats fed by MHN-01 or an ordinary fluid diet. The respiratory quotient of the MHN-01 group was significantly lower than the ordinary fluid group, although the resting energy expenditure of both groups was almost the same level. The effect of MHN-01 was estimated to be based on improvement of lipid metabolism.
Between 2003 and 2005, among 164 patients who underwent radical thoracic esophagectomy and/or reconstruction for esophageal carcinoma at Okayama University Hospital, nine patients (5.5%) were diagnosed with diabetes mellitus in pre-operative screening and were treated with MHN-01. Clinical courses of two cases with severe status of diabetes mellitus were presented as successful case reports of MHN-01.
MHN-01 was very useful in perioperative management of patients complicated by diabetes mellitus, unable to ingest food p.o. such as esophageal cancer or other diseases.
The antimetastatic activity of a novel camptothecan conjugate, MEN4901/T-0128, in which 7-ethyl-10-aminopropyloxy-camptothecin (T-2513) is bound to a biodegradable carboxymethyldextran via a Gly-Gly-Gly linker, was observed in this study. High antimetastatic activity of MEN4901/T-0128 was demonstrated in a clinically-relevant orthotopic mouse model of human colon cancer. MEN4901/T-0128 and irinotecan were compared for anti-metastatic activity as well as efficacy against the primary tumor. An imageable, metastatic model was made by surgical orthotopic implantation (SOI) of the green fluorescent protein (GFP)-expressing HT-29 tumor in nude mice. MEN4901/T-0128 and irinotecan were administered intravenously at various doses and schedules. MEN4901/T-0128, with treatment beginning on d 49 after SOI, was highly effective on lymph node metastasis as well as against the primary tumor. Both GFP imaging and histology demonstrated a markedly lower metastatic incidence of lymph nodes in all MEN4901/T-0128 treated mice compared with irinotecan-treated and untreated mice. At the most efficacious dose of MEN4901/T-0128, only 1 of 12 animals had lymph node metastasis compared with 19 of 20 in the control group. The present study demonstrates the principle that when a camptothecan is conjugated to an appropriate polymer, the drug can become extremely effective with important clinical potential for antimetastatic therapy, a most urgent need.
Pneumoperitoneum may be responsible for ultra-structural alterations in the mesothelium during laparoscopy. To characterize the effect of pneumoperitoneum on the mesothelial cells with CO(2) and compressed air; and to compare to laparotomy and control group (anesthesia only).
Forty C-57 mice were divided in four groups of 10 animals each: CO(2), air, laparotomy, and control group. The animals were submitted to pneumoperitoneum at 8 mmHg during 30 min (CO(2) or compressed air). Five animals of each group were sacrificed 2 and 24 h after the procedure. Fragments of parietal peritoneum were collected and processed for scanning electron microscopy.
Control group revealed uninterrupted mesothelial cells, without any evidence of cellular limits; close contact between the cells; absence of intercellular clefts and presence of microvilli. In the laparotomy group, similar results to the control group, with decreased microvilli were noted. Air pneumoperitoneum was associated with alterations in the morphology of the mesothelial cells, clear cellular limits, and cells with spherical and fusiforme formats. CO(2) pneumoperitoneum showed mesothelial cells with clear cellular limits, predominantly spherical cellular format, and intercellular clefts that allowed the visualization of the exposed basal membrane. These alterations were more intense after 24 h. There was a statistical significance between CO(2) group (2 and 24 h) compared to the control group and laparotomy for cellular limits, intercellular clefts and microvilli, P < 0.0001.
Pneumoperitoneum causes damage in the mesothelial ultra-structure, which differs from the laparotomy group. CO(2) pneumoperitoneum is more harmful to the mesothelium than the air.
This study was designed to investigate whether or not a novel nonselective endothelin A/B (ETA/ETB) receptor antagonist (TAK-044) provides hepatoprotection during porcine liver transplantation. The grafts were stored in chilled Euro-Collins solution and recirculated following reflush with lactated Ringer's with (TAK group) or without (control group) TAK-044 (10 mg/kg). Intracellular (cytoplasma, mitochondria, and nucleus) calcium (Ca) concentrations were measured in the hepatic biopsy materials obtained serially at varying time point from donor laparotomy to recipient closure using an electron probe X-ray microanalyzer. Liver function tests also were determined. The cold and warm ischemia times of the grafts were comparable between the two groups. The peak endothelin-1 T-1) concentration after recirculation was significantly higher in the TAK group than in the control group (129 +/- 30 pg/ml vs 26 +/- 6.5 pg/ml). However, release of liver enzymes, increases in total bile acid, and deterioration of indocyanine green retention rate were significantly suppressed in the TAK group. In the control group, the intracellular Ca concentrations, especially in the mitochondrial fraction, were elevated markedly following recirculation of the hepatic arterial flow. In the TAK group, this effect was suppressed. Thus, the supplementary use of the nonselective ETA/ETB receptor antagonist TAK-044 via a rinse route may alleviate an early postreperfusion microcirculatory disturbance of the liver grafts without adverse effects by the increased ET-1 on the systemic circulation.
Ischemia reperfusion injury (IRI) contributes significantly to posttransplant graft dysfunction. An emphasis, therefore, has been directed toward the identification of novel renoprotective agents. In this study, the renoprotective effect of tetrodotoxin (TTX) alone, or in combination with a thromboxane synthetase inhibitor (OKY-046), was investigated in a 60-min warm ischemia, 72-h reperfusion, IRI rodent model. Unilateral nephrectomized rats were treated with the test vehicle alone, 1, 2, or 4 microgram/kg of TTX or 2 mg/kg of OKY-046 intravenously, either 15 min pre- or postischemia, or 2 microgram/kg TTX administered simultaneously with OKY-046 (2 mg/kg), following the ischemic interval. Baseline, 24, and 72 h mean plasma creatinine (Cr) and urea nitrogen (BUN) were compared. Maximal renoprotection was demonstrated by significantly improved 72-h Cr and BUN levels with the 2 microgram/kg of TTX or with 2 mg/kg of OKY-046, each administered after ischemia (ischemic control Cr = 8. 01 +/- 1.07 mg/dl vs TTX = 3.84 +/- 0.80 mg/dl, P = 0.008; vs OKY-046 = 4.0 +/- 1.5, P + 0.008; ischemic control BUN = 241.3 mg/dl +/- 32.8 vs TTX = 85.7 mg/dl +/- 18.7, P < 0.008; vs OKY-046 = 52.6 +/- 22.5, P = 0.008). The combination therapy utilizing TTX with OKY-046 resulted in reduced animal survival, demonstrating no renoprotection as measured with the biochemical parameters. These results support the renoprotective effects of TTX in a severe, rodent IRI model. The exact mechanism of action, as well as the therapeutic potential of TTX in preservation/transplantation, warrants further study.
Previous reports from other investigators demonstrate prolongation of allogeneic heart graft survival and decrease in CTL responses in rats treated with a small synthetic peptide corresponding to residues 75-84 of the human HLA-B7-01 molecule (Allotrap 07R). We wished to determine the efficacy of these peptides in the highly immunogenic ACI > LEW and LEW > ACI small bowel transplant models. Animals were divided into treatment groups: I, none; II, Allotrap (20 mg/kg/day on Days 0-4); III, cyclosporine (CsA; 10 mg/kg/day on Days 0-4); IV, Allotrap + CsA (as in groups II and III); V, Allotrap (40 mg/kg/day every other day on Days -19 to 4); VI, Allotrap + CsA (as in groups III and V); VII, Allotrap + CsA (as in groups III and V, with Allotrap administered intragraft Days 0-4). The animals were sacrificed at the time of graft rejection (defined by dusky, necrotic stoma and increased stomal output). Peripheral blood, spleen, native bowel, and allograft intraepithelial and lamina propria lymphocytes were harvested and mixed lymphocyte culture (MLC) reactivity against self, donor, and third-party splenocytes was assessed. Statistical analysis was performed by ANOVA with Dunnett's t for multiple comparisons against a control as a post hoc test. We found a very slight, but significant prolongation of graft survival in with treatment protocol V for both strain combinations. In addition, MLC response of splenocytes to donor antigen was decreased with combined CsA and Allotrap, but not with Allotrap alone. We conclude that Allotrap decreases response to alloantigens, and slightly, but significantly prolongs graft survival in the hihgly immunogenic small bowel transplant model.
Fibroblast survival in a three-dimensional collagen matrix is dependent in part upon the rigid anchorage of the matrix to tissue culture plastic. We hypothesized that focal adhesion kinase (FAK) and protein kinase B (Akt) would be activated and that the p53 level would be low in the rigidly anchored (attached) collagen matrix; loss of anchorage (detachment) was hypothesized to have the opposite effects.
Materials and methods:
Human foreskin fibroblasts were cultured in attached bovine collagen matrices for 48 h before detachment as free-floating matrices. At various time points postrelease, matrix lysates were blotted for the proteins of interest, and the terminal deoxynucleotidyltransferase-mediated dUTP nick-end label assay was performed on both whole matrices and cytospin preparations. Irradiated monolayer fibroblasts were used as positive controls for the amount of p53 protein.
Terminal deoxynucleotidyltransferase-mediated dUTP nick-end label positivity in attached versus detached matrices (at 24 h post detachment) was 0.7 +/- 03 versus 5.3 +/- 1.7% (P < 0.05, unpaired t test). FAK and Akt were phosphorylated (activated) in the attached matrix; there was a near complete of loss of both activated forms within 4 h of matrix detachment. Irradiated monolayer fibroblasts had increased levels of p53, mdm2, and p21. In contrast, the p53, mdm2, and p21 levels were just at the level of detection in the attached matrix, but were induced 5- to 10-fold within 2-4 h after matrix detachment.
FAK and Akt are activated in the attached fibroblast-populated collagen matrix whereas the p53 level is relatively low; matrix detachment downregulates FAK and Akt activity and induces p53. The state of mechanical anchorage of the collagen matrix regulates the survival of embedded fibroblasts through a mechanism which may involve FAK.
D-myo-Inositol-1,2,6-triphosphate (IP3) has been shown to reduce edema and progressive ischemia following experimental skin burns. The mechanism(s) are not identified but could be related to antiinflammatory effects of the agent. In the present ex vivo study we compared the effects of IP3 with those of saline and indomethacin on eicosanoid formation by normal and burned rat skin. In burned skin IP 3 treatment reduced the release of thromboxane B2 (TXB2) (P < 0.01) and leukotriene B4 (LTB 4) (P < 0.05) but there was only a weak trend for less prostaglandin E (PGE) compared to burned control animals receiving saline. Indomethacin reduced the release of TXB2 (P < 0.01), and PGE (P < 0.001), but not LTB 4 from burned skin compared to skin from saline-treated burned animals. In non-burned skin IP 3 increased the release of PGE (P < 0.01) and LTB 4 (P < 0.01), but did not significantly influence TXB2 accumulation in the incubation fluid compared to the saline-treated group. Indomethacin reduced the release of TXB2 (P < 0.001) and PGE (P < 0.001), but increased LTB 4 (P < 0.001) in normal skin compared to the saline-treated group. In conclusion, IP 3 inhibited the release of TXB2 and LTB 4 from burned skin ex vivo, but increased PGE and LTB 4 release from normal skin. These results suggest that the mode of action of IP 3 differs from that of nonsteroidal antiinflammatory drugs. The effects of IP 3 on the arachidonic acid cascade also seem to differ in burned versus normal skin.
1,25-dihydroxyvitamin D(3) (1,25-(OH)(2)D(3)) plays an important role in regulating immune responses, in addition to its effects on bone metabolism. The cytokine transforming growth factor beta (TGF-beta) regulates diverse biological processes, including cellular proliferation and differentiation, immune modulation, and modulation of extracellular matrix deposition. 1,25-(OH)(2)D(3) interacts in vitro with Smad proteins, important regulators of TGF-beta signal transduction. We hypothesized that exogenous 1,25-(OH)(2)D(3) would alter levels of TGF-beta(1) and TGF-beta(1) signaling proteins in renal tissue.
C57BL6 mice and Lewis rats were placed on diets with or without 1,25-(OH)(2)D(3) for 14 days. Renal lysates were examined for TGF-beta(1), vitamin D receptor (VDR), and Smad3 protein levels using a cell proliferation assay and Western blot analysis. Coimmunoprecipitation was used to determine if any interaction between VDR and Smad3 proteins occurs in vivo. Reverse transcription-polymerase chain reaction (RT-PCR) was used to assess messenger RNA (mRNA) levels for all of these molecules.
Vitamin D supplementation decreased VDR and Smad3 protein levels. Coimmunoprecipitation of VDR and Smad3 revealed a Smad3-VDR interaction in vivo. Vitamin D-treated rats had a significant (P = 0.001) reduction in bioactive renal TGF-beta(1). RT-PCR demonstrated no difference in mRNA expression for either VDR or TGF-beta(1).
Our results suggest that vitamin D has a significant effect in regulating levels of bioactive TGF-beta(1) and appears to affect aspects of the TGF-beta(1) signaling system. These effects, in combination with the immunomodulatory actions of vitamin D, may alter the evolution of chronic rejection in renal transplants.
The expression of vitamin D receptors (VDR) and growth inhibition induced by 1,25-dihydroxyvitamin D3 have been noted in certain human malignant melanoma cell lines. In this study, widely disparate levels of VDR mRNA expression were demonstrated in a panel of eight human malignant melanoma cell lines. Quantitation of receptor level by ligand binding assay showed a similar pattern. Proliferation and growth curve analysis was performed in two cell lines: RPMI 7951 (high VDR) and SK-MEL-28 (low VDR). Significant growth inhibition was noted in RPMI 7951 cells at 10(-9) M 1,25-dihydroxyvitamin D3. SK-MEL-28 cells, which express much lower levels of VDR, did not show any growth inhibition except at extremely high concentrations of 1,25-dihydroxyvitamin D3, namely 10(-5) M. These findings suggest a receptor-mediated mechanism of growth inhibition for 1,25-dihydroxyvitamin D3 and a role for this hormone in the growth of malignant melanoma cells.
Antibody-mediated rejection continues to be an obstacle for xenotransplantation despite development of α1,3-galactosyltransferase knockout (GTKO) pigs. Fibronectin (Fn) from GTKO pigs was identified as a xenoantigen in baboons. N-glycolylneuraminic acid (Neu5Gc), similar to galactose α1,3-galactose, is an antigenic carbohydrate found in pigs. We evaluated human antibody reactivity and performed initial antigenic epitope characterization of Fn from GTKO pigs.
Materials and methods:
GTKO pig aortic endothelial cells (AEC) were isolated and assessed for antibody-mediated complement-dependent cytotoxicity (CDC). Human and GTKO pig Fn were purified and analyzed using immunoblots. GTKO pig and human AEC absorbed human sera were assessed for CDC and anti-GTKO pig Fn antibodies. GTKO pig proteins were assessed for Neu5Gc. Immunoaffinity-purified human IgG anti-GTKO pig (hIgG-GTKOp) Fn using a GTKO pig Fn column were evaluated for cross-reactivity with other proteins.
GTKO pig AEC had greater human antibody binding, complement deposition and CDC compared with allogeneic human AEC. Human sera absorbed with GTKO pig AEC resulted in diminished anti-GTKO pig Fn antibody. Neu5Gc was identified on GTKO pig Fn and other proteins. The hIgG-GTKOp Fn cross-reacted with multiple GTKO pig proteins and was enriched with anti-Neu5Gc antibody.
Removal of antigenic epitopes from GTKO pig AEC would improve xenograft compatibility. GTKO pig Fn has antigenic epitopes, one identified as Neu5Gc, which may be responsible for pathology and cross-reactivity of hIgG-GTKOp Fn. Genetic knockout of Neu5Gc appears necessary to address significance and identification of non-Neu5Gc GTKO pig Fn antigenic epitopes.
Quinoxaline-1,4-di-N-oxide administered in divided doses to a total of 200 mg./kg. prior to irradiation stimulated leukopoiesis in autografted dogs, thus making their bone marrow more susceptible to the damaging effects of x-irradiation. The end result was a decrease in both mean survival time and total survival as compared to control dogs receiving the autograft alone.
The authors wish to thank Manuel Sanchez for his assistance during the course of this investigation.
Recently, hepatic surgery has made remarkable progress, and it is important to use appropriate liver perfusion. We evaluated the effect of normothermic liver perfusion with the addition of fructose-1, 6-bisphosphate (FBP) and oxygenation to maintain liver parenchymal, non-parenchymal, and Kupffer cell function.
The rats were divided into five groups according to the perfusate and continuous perfusion was performed: Control group = 4 degrees C lactate Ringer with 10% glucose (LRG) solution; normothermic group = 25 degrees C LRG solution; normothermic oxygenated group = 25 degrees C oxygenated LRG solution; normothermic FBP group = 25 degrees C LRG solution with addition of 10 mmol/L FBP; normothermic oxygenated FBP group = 25 degrees C oxygenated LRG solution with addition of 10 mmol/L FBP. Parameters under evaluation were oxygen consumption, liver energy level (adenosine triphosphate, total adenine nucleotide), glutathione, lipid peroxide, hyaluronic acid uptake ratio, apoptosis, and histomorphology. Moreover, we studied the effect of FBP and normothermia on Kupffer cells activation in vitro.
Liver energy level was lower in the normothermic group than the control group. But, it was improved by oxidation or addition of FBP, and it was satisfactorily maintained up to 120 min in the group with normothermic oxygenated FBP. Hyaluronic acid uptake was maintained highly at all times as measured in normothermic oxygenated FBP group. The uptake of lipopolysaccharide was significantly higher as a result of adding FBP, compared with that in the control group and the normothermic group. Moreover, the apoptotic index in the liver was decreased in normothermic FBP group compared to control group.
The normothermic liver perfusion under additional FBP and oxygenation protects both parenchymal and non-parenchymal cells from reperfusion injury.
Fructose 1,6-diphosphate (FDP) has been shown to attenuate tissue injury associated with ischemia and shock by enhancing the anaerobic carbohydrate utilization and by inhibiting oxygen-free-radical generation by the neutrophils. Previously, we have reported that FDP prevents ischemic renal failure if administered prior to the ischemic insult. The present study was designed to determine whether this agent could prevent renal damage when administered during the postischemic reperfusion period. Rats were subjected to 30 min of bilateral renal artery occlusion and infused with FDP (350 mg/kg body wt) beginning 10 min after release of the renal artery clamps. Control rats received an equal volume of glucose/saline solution. A third group of rats were sham operated. Twenty-four hours after injury, BUN, creatinine, and fractional sodium excretion values were less in FDP-treated rats than in control rats (P less than 0.001, P less than 0.005, and P less than 0.001, respectively) and not different from values observed in sham-operated rats. Inulin clearance was greater (P less than 0.001) in FDP-treated rats than in control rats (665 +/- 38 microliters/min/g kidney wt). Renal histology was also better preserved in the FDP-treated group. These data suggest that FDP infused after the initiation of an acute ischemic insult provides significant, but not complete, functional and histologic protection from renal damage.
Fructose-1,6-diphosphate (FDP) is reported to have a salutary effect in endotoxin shock and sepsis. This investigation describes the effect of FDP on pulmonary and systemic hemodynamics, lung lymph protein clearance, and leukocyte count in sheep infused with Escherichia coli endotoxin.
Anesthetized sheep (n = 18), some of which underwent thoracotomy to cannulate lymphatic nodes, were used in this study. After stabilization, all sheep received E. coli endotoxin, 5 microg/kg i.v. infusion over 30 min. Concomitant with the endotoxin infusion, half of the animals were randomly selected to receive an i.v. bolus of FDP (10%), 50 mg/kg, followed by a continuous infusion of 5 mg.kg(-1).min(-1) for 4 h; the rest were treated in the same manner with glucose (10%) in 0.9% NaCl.
Pulmonary artery pressure (PAP) and resistance in the glucose group increased from 20.8 +/- 1.6 to 36.7 +/- 3.2 mmHg (P < 0.007) and from 531 +/- 114 to 1137 +/- 80 dyn.s(-1).cm(-5), respectively (P < 0.005). Despite an increase during endotoxin infusion, these parameters in the FDP group returned to control values. There were no differences in left ventricular pressures, cardiac output, heart rate, and arterial oxygen tension between the groups. In the glucose group, lymph protein clearance was higher (P < 0.01) and blood leukocyte count was lower (P < 0.02). The wet/dry lung weight ratio (g/g) for the glucose group was 5.57 +/- 0.04 and for the FDP-treated group 4.76 +/- 0.06 (P < 0.0005).
FDP treatment attenuated significantly the characteristic pulmonary hypertension, lung lymph protein clearance, and pulmonary vascular leakage seen in sheep infused with endotoxin.
We hypothesized that the addition of fructose 1, 6-diphosphate (FDP) to a hypothermic heart preservation solution could improve metabolic recovery because it has several beneficial effects.
Twenty adult Sprague-Dawley rats were used to study hypothermic heart preservation. The hearts were removed under general anesthesia and preserved at 4 degrees C in Euro-Collins solution (30 ml/kg) for 8 h. In the study group (N = 10), FDP (5 mM) was added to the Euro-Collins solution. In the control group (N = 10), no FDP was added. Heart function was studied after preservation using a working heart model. The ability of various concentrations of fructose 1,6-phosphate to passively diffuse through an egg phosphatidylcholine multilamellar vesicle (MLV) membrane bilayer was examined.
Cardiac output ranged from 17.0 +/- 1.9 to 24.9 +/- 1.6 ml/min in the study group vs 2.0 +/- 1.0-12.3 +/- 1.7 ml/min for controls, average aortic flow was 10. 8 +/- 1.4 ml/min in the study group vs -1.3 +/- 1.6 ml/min for controls, and maximum LV generated power was 22.8 +/- 1.7 J/min vs 10.1 +/- 1.6 J/min for controls. Coronary flow, left ventricular stroke volume and stroke work, and myocardial oxygen consumption were much higher in the study group than in the control group. Coronary vascular resistance was lower in the study group than in the control group. Electron microscopic study indicated that many myocytes displayed patches of swollen mitochondria in the control group, but was rarely observed in the study group. The addition of 50 mM FDP caused substantial changes in MLV permeability. No dose of sucrose buffers outside the vesicles resulted in a significant changes of MLV permeability.
Our results indicate that the addition of FDP to Euro-Collins solution significantly improves hypothermic rat heart preservation, and FDP appeared to cross the membrane bilayer.
Gram-negative sepsis ranks as the leading cause of death in intensive care units, and its incidence is increasing steadily and mortality rates has not changed much over recent decades.
We investigated the efficacy of the amphibian peptide, citropin 1.1 alone and in combination with tazobactam-piperacillin (TZP) in two experimental mice models of gram-negative sepsis. Animals were given an intraperitoneal injection of (1) 1 mg Escherichia coli 0111:B4 LPS, and (2) 2×10(10) CFU of E. coli ATCC 25922. For each model, all animals were randomized to receive intraperitoneally isotonic sodium chloride solution, 1 mg/Kg citropin 1.1 and 120 mg/Kg of TZP, and finally 1 mg/Kg citropin 1.1 plus 60 mg/Kg of TZP. Lethality, bacterial growth in blood and peritoneum, and oxidative stress indices in plasma were evaluated.
All compounds reduced the lethality compared with controls. Treatment with citropin 1.1 resulted in significant decrease in plasma endotoxin and cytokine levels, while TZP exerted opposed effect. The combination between citropin 1.1 and TZP proved to be the most effective treatment in reducing all variables measured.
Due to its multifunctional properties, citropin 1.1 may become an important future consideration to treat conditions in which oxidative organ failure may be present.
Rabbit and dog kidneys were perfused for 30 min at 37°C with 1.4 M [3H]Me2SO in a K+-Mg2+−rich perfusate. Subsequently the kidneys were perfused for 30 min with Me2SO-free perfusate. The rate of Me2SO uptake and washout as well as Me2SO distribution in the tissue were determined. It was found that equilibrium conditions were achieved within 30 min for both uptake and washout with ratios approaching 1.0. The amount of Me2SO in the cortex and medulla of rabbit kidneys was not significantly different. The same experiment was repeated with dog kidneys at 25 and 10°C. At these lower temperatures the rate of uptake and washout was significantly less, but the final concentration achieved within 30 min was the same as at 37°C. Dog kidneys flushed with a K+−Mg2+−rich solution, with or without 1.4 M dimethyl sulfoxide (Me2SO), were kept at 10°C, then reimplanted in the autologous host, and an immediate contralateral nephrectomy was performed. Of the dogs receiving kidneys treated with Me2SO-free solution, 86% survived; of the dogs receiving Me2SO-treated kidneys, 75% survived. Dog kidneys were perfused for 30 min with a K+−Mg2+−rich solution, with or without 1.4 M Me2SO, at 25 or 37°C. All kidneys were then perfused for 30 min with a Me2SO-free solution at the same temperature used for the first perfusion. All kidneys were then reimplanted in the autologous host and an immediate contralateral nephrectomy was performed. Of the dogs receiving kidneys perfused at 25°C with Me2SO-free solution, 43% survived; of the dogs receiving kidneys perfused with Me2SO, 42% survived. Dog kidneys were also treated at 37°C in a manner similar to those at 25°C. Of the dogs receiving kidneys perfused at 37°C with Me2SO-free solution, 80% survived; of the dogs receiving kidneys perfused with Me2SO, 67% survived. Other results indicate that perfusion with a closed circuit is superior to perfusion with an open circuit. Also, gradual administration and washout of Me2SO gives better renal survival than rapid changes in Me2SO concentration.
Necrotizing enterocolitis (NEC) alters intestinal microvascular control mechanisms causing significant vasoconstriction. Our prior work with intraperitoneal 2.5% dextrose solution demonstrated increased intestinal perfusion in experimentally induced NEC. In the current study, we examine whether a buffered solution with lower glucose and osmolar loads similarly increases intestinal blood flow. We hypothesized that buffered 1.5% dextrose solution would increase ileal blood flow compared with baseline in NEC.
We randomly assigned pregnant Sprague-Dawley rats to control (n = 103) or NEC (n = 123) groups, by litter. We induced NEC by previously published methods. Control pups were vaginally delivered and dam-fed. We used laser Doppler flowmetry to evaluate perfusion in the terminal ileum at 12, 24, 48, 72, or 96 h after delivery at baseline and after application of topical 1.5% dextrose solution. We evaluated differences between groups and time points by analysis of variance and Tukey post hoc test.
Baseline blood flow in the terminal ileum increased with gestational age in both groups (P < 0.05). Control groups had significantly greater baseline blood flow than NEC groups (P < 0.05), and topical application of buffered 1.5% dextrose solution increased blood flow compared with baseline in both groups at all time points (P < 0.05).
Topical 1.5% dextrose solution significantly enhanced blood flow in the terminal ileum to the same degree as 2.5% dextrose solution. Thus, the use of buffered 1.5% dextrose solution might be more beneficial in treating clinical NEC, because it places a lower glucose and osmotic load on NEC-injured intestine.