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At least 973 records · Page 54Linked to original sources
PubMed1996

Group B streptococcal beta-hemolysin expression is associated with injury of lung epithelial cells.

Group B streptococci (GBS) are the leading cause of serious bacterial infection in newborns. Early-onset disease is heralded by pneumonia and lung injury, and the lung may serve as a portal of entry for GBS into the bloodstream. To examine a potential role for GBS beta-hemolysin in lung epithelial injury, five wild-type strains varying in beta-hemolysin expression were chosen, along with five nonhemolytic (NH) and five hyperhemolytic (HH) variants of these strains derived by chemical or transposon mutagenesis. Monolayers of A549 alveolar epithelial cells were exposed to log-phase GBS or stabilized hemolysin extracts of GBS cultures, and cellular injury was assessed by lactate dehydrogenase (LDH) release and trypan blue nuclear staining. Whereas NH strains produced no detectable injury beyond baseline (medium alone), hemolysin-producing strains induced LDH release from A549 cells in direct correlation to their ability to lyse sheep erythrocytes. HH strains were also associated with marked increases in trypan blue nuclear staining of A549 monolayers. The extent of LDH release produced by HH strains was significantly reduced in the presence of dipalmitoyl phosphatidylcholine, a known inhibitor of hemolysin and the major phospholipid component of human surfactant. Electron microscopic studies of A549 cell monolayers exposed to HH GBS mutants revealed global loss of microvillus architecture, disruption of cytoplasmic and nuclear membranes, and marked swelling of the cytoplasm and organelles. We conclude that GBS hemolysin expression correlates with lung epithelial cell injury and may be important in the initial pathogenesis of early-onset disease, particularly when pulmonary surfactant is deficient.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1998

Gentle in situ liver manipulation during organ harvest decreases survival after rat liver transplantation: role of Kupffer cells.

BACKGROUND: The etiology of primary graft nonfunction and dysfunction is unknown but most likely involves Kupffer cell-dependent reperfusion injury. However, the donor operation and surgical technique may also have an effect on the outcome after transplantation. Because liver manipulation during harvest cannot be prevented completely with standard procedures, its effect on survival was assessed here. METHODS: Donor livers were harvested from female Sprague-Dawley rats (200-230 g). Briefly, after minimal dissection during the first 12 min, livers were either manipulated gently or left alone for 13 subsequent minutes. At 25 min, all livers were perfused with cold University of Wisconsin solution via the portal vein, and transplantation was performed after cold storage (1 hr). In some rats, Kupffer cells were destroyed with gadolinium chloride or inactivated with dietary glycine before harvest. Survival, proteolytic activity in the rinse effluent, serum transaminases, trypan blue distribution to index microcirculation, and histology were compared. RESULTS: In the nonmanipulated group, survival was 100% after transplantation; however, gentle manipulation decreased survival by 70%. Further, manipulation elevated transaminases fivefold and caused about 200% necrosis. At harvest, proteolytic activity and the time for trypan blue to distribute homogeneously were elevated three- to eightfold by manipulation. Gadolinium chloride and glycine prevented the effects of manipulation on all parameters studied. CONCLUSION: These data indicate for the first time that brief, gentle manipulation of the donor liver has a marked detrimental effect on survival by priming or activating Kupffer cells. This may represent an important early event in pathogenesis, because Kupffer cells play an important role in primary graft nonfunction.

Adenosine↗
PubMed1984

Spontaneous duplications and transpositions of a large chromosome segment in Aspergillus nidulans.

Spontaneous revertants of the leaky adE20 mutant of Aspergillus nidulans were obtained as vigorous sectors emerging from stunted colonies on adenine-free medium. Among the genetically heterogeneous sectors up to about 20% were recognized unequivocally as having an additional chromosome segment bearing adE20; two doses of this leaky allele permitted growth without added adenine. Eleven spontaneous duplication strains of independent origin were analysed genetically. Eight carried the duplicate segment on chromosome IIR; three of these, phenotypically similar to all eight, were analysed in detail and were shown-within the limits of such genetic analysis - to have a large, terminal segment of IR duplicated and attached terminally and uninverted to IIR. One strain had a duplication, possibly tandem, on IR and two had duplications attached elsewhere in the genome. The results suggested a preferential site for the initiation of duplicate segments in this system, as well as a preferential site for their attachment. Agents known to modify instability of a previously studied Dp(IR----IIR) strain affected the frequency of duplications among selected adE20 revertant sectors and/or the genomic locations of duplicated segments. Trypan blue and coumarin, which enhance Dp(IR----IIR) instability in a specific way, and Co2+, which stabilizes Dp(IR----IIR), gave 14, 50 and 62% duplication sectors respectively, among revertants. Duplications selected in the presence of Co2+ had mainly IIR attachments; of those from trypan blue and coumarin, about one-quarter were attached to IIL and none to IIR.

Aspergillus nidulans↗
PubMed1993

Egg viability in urinary schistosomiasis. I. New methods compared with available methods.

A new method of quantifying viability and hatchability of Schistosoma haematobium eggs in urine has been developed. It consists of filtering urine samples through Nytrel filters followed by mercurochrome staining to assist in the detection of viable (motile) miracidia in egg shells, and iodine staining to show hatched miracidia. The new method is compared with two existing techniques for viability/hatchability determination, centrifugation hatching and Nuclepore filtration trypan blue staining. Seventy-three egg positive urine samples are compared; the overall proportions of viable eggs are very close for centrifugation hatching and the new method, but 33% of eggs are lost in centrifugation and 19% of eggs are unclassifiable by Nuclepore filtration trypan blue staining. The main advantages and disadvantages of the new and existing methods are described and discussed; the cheapness, simplicity and ease of performance of the new method make it suitable for large-scale field use. Suggestions for improvements and standardization of the new method are made.

Animals↗
PubMed1985

Halothane hepatitis. Detection of a constitutional susceptibility factor.

We studied susceptibility to halothane hepatitis with an in vitro test that detects cell damage from electrophilic drug intermediates. Metabolites of phenytoin were generated by incubation of phenytoin with rat hepatic microsomes in the presence of the epoxide hydrolase inhibitor 1,1,1-trichloropropene oxide (TCPO), which prevents the further metabolism of phenytoin to an inert metabolite. In lymphocytes exposed to this system, cytotoxicity was measured by trypan blue dye exclusion and was expressed as the percentage increase in trypan blue-positive cells after the addition of TCPO. In the presence of TCPO, lymphocytes from 11 patients with halothane hepatitis exhibited an increase in cytotoxicity at 0.06 mM phenytoin that was eight times greater than the increase in healthy controls (54 +/- 10 per cent [mean +/- S.E.M.] vs. 7.1 +/- 2.2 per cent, P less than 0.0001). Patients with other liver diseases and persons recently exposed to halothane without adverse effects did not differ from healthy controls. In three patients with halothane hepatitis who were studied serially, the lymphocyte abnormality was still present after 13 months. Family studies revealed abnormal results on 10 cytotoxicity tests among 19 members of four families. We propose that there is a familial, constitutional susceptibility factor that predisposes persons to halothane hepatitis.

Adult↗
PubMed2000

Reperfusion injury is dramatically increased by gentle liver manipulation during harvest.

Kupffer cell-dependent injury in livers gently manipulated during harvest develops upon reperfusion. The purpose of this study was to characterize this injury and to detect underlying mechanisms. Livers from female Sprague-Dawley rats were harvested for transplantation within 25 min. Minimal dissection was performed during the first 12 min, including freeing the liver from ligaments. After this, for further 13 min, livers were either left alone or manipulated gently. The livers were then cold-stored for 24 h in University of Wisconsin (UW) solution and perfused with oxygen-saturated Krebs-Henseleit buffer at 37 degrees C. Trypan blue in the buffer was used to index microcirculation. Cell damage was assessed with histology. Initial dissection during harvest and cold storage had minimal effects on sinusoidal lining cells; in contrast, the subsequent gentle organ manipulation dramatically increased cell death 6.5-fold, while the time for complete trypan blue distribution increased 2.3-fold (P < 0.05). Manipulation increased proteolysis 2-fold (P < 0.05). At harvest, manipulation increased portal venous pressure significantly by 68%. Treatment of donors with gadolinium chloride, a selective Kupffer cell toxicant, or with dietary glycine, an inhibitor of Kupffer cell activation, prevented effects of organ manipulation on all parameters studied. These findings demonstrate Kupffer cell-dependent reperfusion injury of sinusoidal lining cells caused by manipulation of the liver during its recovery. The mechanisms are those of proteolysis and impaired hepatic microcirculation.

Animals↗
PubMed1996

Pre-treatment with trimetazidine increases sarcolemmal mechanical resistance in reoxygenated myocytes.

OBJECTIVE: Cytoskeletal and sarcolemmal fragility secondary to anoxia may contribute to sarcolemmal rupture and cell death during reoxygenation of cardiomyocytes. This study investigated the influence of trimetazidine (TMZ), a drug with effects on lipid metabolism and cell membranes, on reoxygenation-induced sarcolemmal rupture. METHODS: Isolated adult rat myocytes were submitted to 60 min of metabolic inhibition and 5 min of hypo-osmotic reoxygenation to simulate reperfusion edema in situ. Cells were allocated to 3 groups of treatment: in one group, TMZ 100 mumol/l was added to both the metabolic inhibition and reoxygenation buffers (group TMZ); another group was submitted to the same treatment but cells had previously been incubated with TMZ 100 mumol/l for 3 h (group TMZ-Pre); a control group underwent metabolic inhibition and hypo-osmotic reoxygenation without any treatment. Cell morphology was monitored throughout the experiment and sarcolemmal integrity was assessed by quantification of LDH activity and trypan blue exclusion test. RESULTS: After 60 min of metabolic inhibition most cells (83.1 +/- 2%) presented rigor contracture without between-group differences. Reoxygenation resulted in hypercontracture of 84.2 +/- 2.3, 91.2 +/- 1.4 and 84.1 +/- 2.1% of cells in TMZ, TMZ-Pre and control groups, P = NS. The trypan blue exclusion test revealed a higher proportion of cells with sarcolemmal integrity in TMZ and TMZ-Pre groups than in controls (12.7 +/- 2.0, 10.0 +/- 1.5 and 6.3 +/- 0.8%, respectively, P = 0.002). No between-group differences in LDH activity in the extracellular medium were observed at the onset or at the end of metabolic inhibition. However, LDH release was significantly lower (P = 0.002) in the TMZ-Pre group (1.6 +/- 0.1 IU/1000 cells) than in the TMZ and control groups (1.9 +/- 0.2 and 2.2 +/- 0.1 IU/1000 cells). CONCLUSION: Preincubation of cardiomyocytes with TMZ does not prevent rigor contracture induced by metabolic inhibition or hypercontracture during subsequent reoxygenation, but does improve sarcolemmal resistance to reoxygenation-induced mechanical stress. This could help to explain the beneficial effect of TMZ on infarct size.

Animals↗
PubMed1988

Hepatic reperfusion injury following orthotopic liver transplantation in the rat.

At 24 hr following orthotopic transplantation, rat liver grafts were perfused in situ for 7 min with trypan blue, a vital dye that provides information on hepatic microcirculation and stains nuclei of nonviable cells. Spotty and uneven dye distribution was observed indicating that hepatic microcirculation was disturbed 24 hr following transplantation surgery. Under these conditions, 15-20% of the hepatocytes were nonviable as assessed from trypan blue staining and frank necrosis. In contrast, perfusion of livers from untransplanted rats or liver explants exposed to cold ischemia for 60 min were judged normal by the criteria of uniform distribution of dye in the organ and absence of necrosis and nuclear dye uptake. Thus the observed damage was associated with reintroduction of blood and can therefore be classified as a reperfusion injury. The altered microcirculation and cell death following the operation was reduced markedly by perfusion of the cold, ischemic explant with nitrogen-saturated but not with oxygen-saturated buffer for 5 min prior to the implantation operation. Protection was even greater if the perfusion medium contained verapamil (20 micrograms/ml), a Ca++ channel blocker. We conclude that reperfusion of the stored liver causes an oxygen-dependent alteration in hepatic microcirculation that leads to hypoxia and scattered hepatocellular necrosis in the implanted graft. Brief perfusion of the hypoxic implant under anaerobic conditions may remove substrates involved in oxygen radical generation and prevent reperfusion injury upon introduction of oxygen into the graft via the blood. Taken together, these results suggest that removal of Euro-Collins' solution under anaerobic conditions may be beneficial clinically in preventing injury of surgical explants.

Animals↗
PubMed1982

Electrical determination of viability in saline-treated mouse myeloma cells.

Suspension of mouse myeloma cells in phosphate buffered saline (PBS) induced a significant amount of cell death. The lethal effects of PBS include an increase in cell lysis, a decreased ability of cells to exclude trypan blue, and a decrease in the colony-forming ability of these cells. Dead cells were also detected on a Coulter counter by the increase in the fraction of cells with a smaller electrical size distribution (ESD). Comparing mixtures of live and dead cells by ESD and trypan-blue exclusion showed a high correlation of electrical size with viability (correlation coefficient = 0.98). Sizing of PBS-treated cells by light microscopy suggested that the altered ESD of the PBS-treated cells was due to a downward shift in the volume distribution. Light-scattering experiments also suggested a decrease in the size of cells after PBS treatment. An increase in permeability of the cell membrane may also contribute to these results. We conclude that electrical sizing is an excellent indicator of physical changes that result from PBS-induced cell death, and is an effective method for distinguishing live and dead mouse myeloma cells after PBS treatment.

Animals↗
PubMed1986

Atrial natriuretic factor does not stimulate prostaglandin synthesis in isolated rat glomeruli.

The effect of alpha-human atrial natriuretic polypeptide (alpha-hANP) on the synthesis of prostaglandins was studied in isolated rat glomeruli. Glomeruli were isolated by a passive sieving technique according to the method of Misra and were incubated at 37 degrees C for 60 min in the presence (Group II: 10(-6) M, Group III: 10(-5) M) or absence (Group I: control) of alpha-hANP. Furthermore, glomeruli were incubated with arachidonic acid (10(-5), 10(-4), and 10(-3) M) and at the end of the incubation period trypan blue was added to the glomerular suspension. The presence of alpha-hANP (10(-6) and 10(-5) M) caused no significant difference in prostaglandin synthesis as compared with the control. On the other hand, arachidonic acid stimulated prostaglandin synthesis and the glomerular preparation was not stained by trypan blue, indicating that they remained viable. These results suggest that alpha-hANP does not directly affect the prostaglandin synthesis in isolated rat glomeruli.

6-Ketoprostaglandin F1 alpha↗
PubMed1987

Cytotoxic effects of ether lipids and derivatives in human nonneoplastic bone marrow cells and leukemic cells in vitro.

The effects of 2-lysophosphatidylcholine (2-LPC), the alkyl lysophospholipid derivatives (ALP) 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine (ET-18-OCH3) and 1-O-hexadecyl-sn-glycero-3-phospho-trimethyl-ammonio-hexanol, the 2-acetamide analog of platelet-activating factor (PAF) 1-O-octadecyl-2-acetamide-sn-glycero-3-phosphocholine, the thioether lysophospholipid derivative (TLP) BM 41.440 and the ether-linked lipoidal amine CP-46,665 on tritiated thymidine uptake and trypan blue dye exclusion were tested in vitro in various freshly explanted cell samples from human nonneoplastic bone marrow and human leukemias. In both assay systems, a dose range of 1-20 micrograms/ml of the compounds was tested after 24, 48 and 72 hr of coincubation with the cells. The trypan blue dye exclusion revealed statistically significant preferential cytotoxicity in leukemic cells for three compounds with the order of quantitative selectiveness: ET-18-OCH3 greater than BM41.440 greater than 2-acetamide analog of PAF. CP-46,665 was the most toxic compound, but did not reveal significant differences between nonneoplastic bone marrow and leukemic cells when added in concentrations greater than 1 microgram/ml. The trimethyl-ammonio-hexanol compound showed only minor activity in the majority of tests, when added at concentrations less than 20 micrograms/ml. 2-LPC was rather ineffective. The tritiated thymidine uptake showed only preferential antiproliferative effects towards leukemic cells of ET-18-OCH3 and, sometimes, within the dose time frame tested of BM 41.440. All compounds tested except 2-LPC and the trimethyl-ammonio-hexanol compound were active also in this assay (inhibition of uptake greater than 50% of the controls).(ABSTRACT TRUNCATED AT 250 WORDS)

Antineoplastic Agents↗
PubMed2002

Cytotoxicity of flutamide and 2-hydroxyflutamide and their effects on CYP1A2 mRNA in primary rat hepatocytes.

AIM: To compare the cytotoxicity of flutamide and its active metabolite 2-hydroxyflutamide and their effects on cytochrome P-450 1A2 mRNA in primary rat hepatocytes. METHODS: After the isolation of hepatocytes a nd t he primary incubation for 4 h, flutamide and 2-hydroxyflutamide were added respectively to the medium at the concentration of 10, 20, and 50 mg/L and incubated for 8 h. Cytotoxicity of hepatocytes was assessed by Trypan blue exclusion, lactate dehydrogenase (LDH) leakage, percentage of alanine aminotransferase (ALT) or aspartate aminotransferase (AST) release, and reduced glutathione (GSH). The effect of flutamide and 2-flutamide on the CYP1A2 mRNA level was further analyzed by Northern blot. RESULTS: After incubation for 8 h, cell viability was observed by Trypan blue exclusion. The increase of ALT and AST activity and the decrease of glutathione content were also noted at 10, 20, and 50 mg/L of flutamide and 50 mg/L of 2-hydroxyflutamide as compared with normal rat hepatocytes. Induction of CYP1A 2 mRNA were 2-, 5-, and 7.5-fold at 10, 20, and 50 mg/L of flutamide and 3.5-fold at 50 mg/L of 2-hydroxyflutamide. CONCLUSION: Cytotoxicity of flutamide and its effect on CYP1A2 mRNA were stronger than those of its active metabolite 2-hydroxyflutamide in primary rat hepatocytes.

Androgen Antagonists↗
PubMed2003

[Effect of echo-contrast agent on rat vascular smooth muscle cell proliferation during ultrasound exposure].

OBJECTIVE: To investigate the effect of echo-contrast agent on rat vascular smooth muscle cell (VSMC) proliferation in the presence of ultrasound exposure. METHODS: The VSMCs of rats were cultured in 6-well plates with or without the echo-contrast agent and stained with trypan blue at 3, 24, and 48 h respectively after ultrasound exposure for 1 min at 2 MHz, 0.25 mechanical index (MI). At each time point as indicated after the exposure, the cells in parallel wells were trypsinized for cell counting using Coulter counter. RESULTS: No cytotoxicity was found in trypan blue staining. Compared with the control cells, the proliferation of VSMCs was inhibited by echo-contrast agent and adjunctive ultrasound within 24 h, but another 24 h later, the effect failed to be observed. CONCLUSIONS: The proliferation of VSMC was inhibited by with low mechanical index cavitation activity of contrast agents.

Angioplasty, Balloon, Coronary↗
PubMed1998

The effects of four mycotoxins on the mitogen stimulated proliferation of bovine peripheral blood mononuclear cells in vitro.

The effects of four mycotoxins, T-2 toxin, deoxynivalenol (DON), ochratoxin A (OTA) and fumonisin B1 (FB1) on the response of bovine peripheral blood mononuclear cells (PBM) in vitro to the mitogens concanavalin A (Con A), phytohaemagglutinin A (PHA) and pokeweed mitogen (PWM) were assayed after 4 days' incubation using 3H-thymidine uptake and the MTT bioassay. The concentrations of mycotoxin required to reduce the proliferative response of PBM by 50% for Con A, PHA and PWM as measured by 3H-thymidine incorporation was for T-2 toxin 0.30, 0.40 and 0.18 ng ml-1; for DON 0.07, 0.09 and 0.04 microgram ml-1; for OTA 0.10, 0.20 and 0.15 microgram ml-1, and for FB1 35, 18 and 11 micrograms ml-1 respectively. The concentrations of mycotoxin required to reduce the proliferative response of PBM by 50% for Con A, PHA and PWM as measured by the MTT bioassay were for T-2 toxin 2.0, 2.0 and 1.0 mg ml-1; for DON 0.70, 0.50 and 0.50 microgram ml-1; OTA 1.5, 1.5 and 1.5 micrograms ml-1; and FB1 > 50, > 50 and 20 micrograms ml-1 respectively. Further cytotoxicity assays including the LDH bioassay and Trypan blue exclusion were performed only on Con A-stimulated PBM cells after 72 h incubation. With the LDH-bioassay the 50% inhibition levels were T-2 toxin 0.3 ng ml-1, DON 0.4 microgram ml-1, OTA 1.4 micrograms ml-1 and FB1 3.5 micrograms ml-1; for Trypan blue uptake the 50% inhibition levels were T-2 toxin 5 ng ml-1, DON 2.3 micrograms ml-1 and OTA 4 micrograms ml-1 respectively.

Animals↗
PubMed2002

Evaluation of the acrosome reaction and viability in buffalo spermatozoa using two staining methods: the effects of heparin and calcium ionophore A23187.

In this study, we investigated the effect of heparin and calcium ionophore A23187 on in vitro induction of buffalo sperm acrosome reaction (AR). Two methods for detection of the AR and viability were employed. Fluorescein isothiocyanate-conjugated Arachis hypogea agglutinin (FITC-PNA) was used as a vital stain in combination with ethidium homodimer-1 (EthD-1) to determine the acrosome status of viable spermatozoa. In another experiment, trypan blue replaced EthD-1 to differentiate live and dead spermatozoa having undergone AR. The results from the two methods were significantly correlated (r > 0.9). Four different staining patterns were found in both methods. The FITC-PNA intensely labels the acrosome region of acrosome-intact spermatozoa. EthD-1 and trypan blue stained red and blue at the post-acrosomal region of dead spermatozoa, respectively. Spermatozoa incubated with heparin showed a significant increase ( p < 0.05) in the percentage of live acrosome-reacted sperm after 30 min incubation period. This trend continued and was significantly different over the entire incubation period when compared with the control group at the same interval. In the ionophore-treated group, the proportion of changes in live acrosome-intact and live acrosome-reacted spermatozoa was statistically significantly different ( p < 0.001) when compared with those treated with heparin at the same interval. The AR occurred sooner and to a greater extent when incubated with the ionophore but at 5 h of incubation the percentage of false acrosomal reaction was significantly higher than those in the control and heparin-treated groups. The results in this study indicated that the in vitro induction of AR by heparin and calcium ionophore evaluated by both methods could be used to assess sperm fertilizing capacity for in vitro fertilization of this species.

Acrosome Reaction↗
PubMed1991

Skin banking: a simple method for cryopreservation of split-thickness skin and cultured human epidermal keratinocytes.

A simple unit has been developed for the simultaneous passive cooling of small and large amounts of allograft or autograft split-thickness skin, as well as cultured human epidermis. An expanded polystyrene box of variable size, aluminum plates, and cellulose tissue are fused. This unit is cooled in a -70 degrees C constant-temperature mechanical refrigerator. Maximal cooling rates of -1.3 degrees C min-1 are obtained in a box with a constant wall thickness of cellulose tissue. The cooling rate can be varied by altering the number of cellulose layers. Exothermic temperature plateaus associated with skin cooled in this unit last for less than 0.3 minutes. The viability of the cryopreserved skin was determined by using up to four methods simultaneously: a dye-exclusion test with trypan blue; glucose consumption; production of lactate; and carbon-13 nuclear magnetic resonance spectroscopy. Using a cryoprotectant medium with 15% (vol/vol) glycerol for split-thickness skin 0.25 mm thick, a storage time of up to 509 days at -70 degrees C was observed, with only a small decrease in viability (trypan blue, 62.5%; glucose consumption, 71 to 90% compared with freshly harvested skin). Storage in liquid nitrogen did not significantly improve results (p greater than 0.05).

Adult↗
PubMed2001

Mesenteric lymph from rats subjected to trauma-hemorrhagic shock are injurious to rat pulmonary microvascular endothelial cells as well as human umbilical vein endothelial cells.

Previously, we have documented that gut-derived lymph from rats subjected to trauma/hemorrhagic shock (T/HS) is injurious to human umbilical vein endothelial cells (HUVEC). To verify these findings in an all rat systems, the ability of T/HS lymph to increase rat pulmonary microvascular endothelial cell (RPMVEC) monolayer permeability and kill RPMVEC was compared with that observed with HUVECs. RPMVEcs isolated from male rats or HUVECs were grown in 24-well plates for the cytotoxicity assays or on permeable filters in a two-chamber system for permeability assays. Mesenteric lymph was collected from male rats subjected to trauma (laparotomy) plus hemorrhagic shock (T/HS group) or to a laparotomy plus sham-shock (T/SS group). The T/HS group had their mean arterial pressure decreased to 30 mmHg and kept there for 90 min. Lymph samples centrifuged to remove the cellular component were incubated with the RPMVECs or HUVECs at a 10% concentration. Neither T/SS lymph nor post-T/HS portal vein plasma was toxic to or increased the permeability of the RPMVECs or HUVECs. The pattern of cytotoxicity observed in the HUVECs incubated with T/HS mesenteric lymph was similar to that observed in the RPMVECs, as reflected by trypan blue dye exclusion, with more than 95% of the HUVECs and RPMVECs being killed after a 16-h incubation with T/HS mesenteric lymph. However, at earlier time points the amount of LDH released from the HUVEC cells incubated with T/HS lymph was greater than that observed with the PRMVEC, although trypan blue dye exclusion was similar. Similarly, incubation with 10% T/HS lymph increased the permeability of both HUVEC and RPMVEC monolayers more than 2-fold, even with an incubation period as short as 1 h. In conclusion, these results provide further evidence that T/HS lymph, but not T/SS lymph or post-T/HS portal vein plasma, is injurious to endothelial cells and that RPMVECs are as susceptible to injury as HUVECs. Additionally, these studies support the emerging concept that gut-induced distant organ injury is mediated by factors contained in mesenteric lymph.

Animals↗
PubMed1990

Ethanol potentiates oxygen uptake and toxicity due to menadione bisulfite in perfused rat liver.

Menadione bisulfite is a hepatotoxicant that damages periportal regions of the lobule in perfused liver in an oxygen-dependent manner. The effect of ethanol on menadione bisulfite toxicity was examined in perfused rat liver. Addition of menadione bisulfite (3 mM) alone to the perfusate increased oxygen uptake by 20-30 mumols/g/hr. Lactate dehydrogenase was released into the effluent after 60 min of perfusion and reached values around 100 units/g/hr. Under these conditions, trypan blue was taken up exclusively in periportal regions of the liver lobule; 44% of periportal cells were stained. In the presence of ethanol, maximal increases in oxygen uptake due to menadione bisulfite were much larger (about 90 mumols/g/hr), and lactate dehydrogenase release occurred earlier and reached higher maximal values (330 units/g/hr). Trypan blue staining was also more extensive; 90% of periportal cells were stained. The effect of ethanol on menadione bisulfite-induced oxygen uptake required metabolism via alcohol dehydrogenase (ADH), because ethanol increased oxygen uptake due to menadione bisulfite from 44 to 81 mumols/g/hr in deermice with ADH but had no effect in deermice lacking ADH. Other agents that increase NADH (xylitol and 2-ethyl-1-hexanol) also potentiated the stimulation of oxygen uptake due to menadione bisulfite, suggesting that ethanol was working by increasing the NADH redox state. Cyanide abolished the increase in oxygen uptake due to menadione bisulfite, both in the absence and in the presence of ethanol, supporting the hypothesis that the effect of ethanol on menadione bisulfite-mediated oxygen uptake involves the mitochondrial respiratory chain. Further, the stimulation of oxygen uptake by menadione bisulfite in isolated mitochondria was enhanced when matrix NADH was increased by addition of beta-hydroxybutyrate. These data indicate that ethanol potentiates oxygen uptake and toxicity due to menadione bisulfite most likely by generation of NADH for redox cycling of this model quinone.

Adenosine Triphosphate↗