Research in experimental medicine. Zeitschrift fur die gesamte experimentelle Medizin einschliesslich experimenteller Chirurgie最新文献

We examined the effects of methotrexate (MTX) on the level of nitric oxide (NO) produced by peritoneal macrophages from rats with adjuvant-induced arthritis (AA) ex vivo. During the development of AA, paw swelling increased and LPS enhanced the capacity of peritoneal macrophages to produce NO and prostaglandin E2 (PGE2). MTX (0.1 mg/kg, p.o.) treatment for 21 days reduced the paw swelling, and inhibited the increased NO and PGE2 production. However, when MTX (0.1 mg/kg, p.o.) was administered to rats with established AA, these parameters were not significantly influenced. In normal rats, MTX (0.1 mg/kg, p.o.) treatment for 21 days did not change NO and PGE2 production of LPS-stimulated macrophages. On the other hand, macrophages from normal and AA rats cultured in the presence of MTX (1, 10 and 100 microM), were activated by LPS in vitro. MTX did not influence NO or PGE2 production by LPS-stimulated macrophages in normal and AA rats. By contrast, indomethacin (IM) (1.0 mg/kg, p.o.) treatment for 21 days reduced the paw swelling, and inhibited NO and PGE2 production in AA rats. IM inhibited significantly PGE2 production, but did not influence NO production by LPS-stimulated macrophages in vitro. These results suggest that MTX treatment reduces NO production in peritoneal macrophages in AA rats, and these actions of MTX may have an inhibitory effect without the modulation of PGE2.

In heart transplantation, global ischemia of a graft is followed by reperfusion injury. The formation of oxygen free radicals induces arrhythmias and impairs functional recovery of the graft. This study was executed to evaluate the effect of the new antioxidant, nitecapone, on ischemia-reperfusion injury in heart transplantation in rats. Donor hearts were perfused and stored at +4 degrees C for 2 h in either Ringer's solution in the control group (C-group, n = 26) or Ringer's solution with nitecapone (NC) added (NC-group, n = 18). The donor aorta was anastomosed to the recipient's abdominal aorta and the pulmonary artery to the recipient's inferior vena cava. The grafts were classified into three categories based on the functional recovery. The rats in both groups were killed at 10, 30, or 60 min after release of the aortic clamp. Tissue samples for chemiluminescence were obtained from the left ventricle, the right ventricle, and the septum of the heart. All grafts in the NC-group (18/18) began beating after release of the aortic clamp, whereas only 50% (13/26) of the grafts in the C-group recovered (P < 0.0004). Chemiluminescence analysis showed lipid peroxidation values to be higher in the C-group than the NC-group up to 1 h after reperfusion. Also, the right ventricle samples showed lower chemiluminescence values in the NC-group than in the C-group. In conclusion, our results do not support the theory that different regions of the heart have different vulnerability to ischemia-reperfusion injuries. Nitecapone has a beneficial effect on the preservation of the grafts in terms of functional recovery.

An experimental study was planned to evaluate 99mTc-citrate, 67Ga-citrate and 99mTc(V) dimercaptosuccinic acid (DMSA) as agents for the visualization of acute appendicitis. Appendiceal ligation was performed through a midline incision in 24 rabbits. Twenty-four hours later the animals were divided into three equal groups. The rabbits were injected through the aurical vein with 1 mCi (37 MBq) 99mTc-citrate in group I, 0.5 mCi (18.5 MBq) 67Ga-citrate in group II and 1 mCi (37 MBq) 99mTc(V) DMSA in group III. After 3 h, static images of the rabbits were obtained with a gamma camera. There were positive images in seven, six and five rabbits in groups I, II and III respectively. The image quality was better in group I than in the other groups. Also, the mean uptake in group I was significantly higher than those of other two groups (P < 0.05). There was no significant difference between groups II and III (P > 0.05). All rabbits had appendicitis confirmed histologically. In conclusion, these results show that 99mTc-citrate is preferable to 67Ga-citrate and 99mTc(V) DMSA for the differential diagnosis of acute abdominal inflammations such as appendicitis, because of higher concentration ratios, simple and rapid preparation, low cost, excretion mainly through the kidneys and fast blood clearance.

Photodynamic therapy (PDT) could be a useful adjuvant in glioblastoma treatment. The fact that epidermal growth factor (EGF) and its receptor are involved in glioblastoma growth control led us to investigate the relationships between EGF and PDT with respect to three different glioma cell lines (C6, T98 G, U87 MG) responsive to growth stimulation by EGF. Flow cytometric analysis revealed that each cell line expressed EGF receptors. PDT was then applied to the cells using haematoporphyrin derivative (HPD) as photosensitizer and argon laser irradiation. When cells were incubated for 2 h with HPD (0.1-10 micrograms/ml) and then laser-irradiated (lambda = 514 nm; energy density 25 J/cm2), all three cell lines showed photosensitivity. The median lethal dose was respectively 3, 4.5 and 2.7 micrograms/ml for C6, T98 G and U87 MG. EGF (2-50 ng/ml) had no effect on HPD- and laser-induced toxicity when added to cells before PDT, whereas toxicity decreased for all three cell lines when EGF was added after PDT. HPD (1-2 micrograms/ml, incubation times 30-180 min) also induced an increase in EGF receptor expression for the C6 line.

That orally administered antigen was shown to induce gastrin release in immunized animals was a new aspect of gastrointestinal physiology. The mediators responsible for this immunological effect are still unclear. In an attempt to discover more about the mechanisms regarding antigen-induced gastrin release, we developed an in vitro system where fragments of rat antral mucosa were challenged. This makes it possible to determine the role of antigen-antibody complexes and the complement system in the mechanism of antigen-induced gastrin release. Wistar rats were immunized in vivo with NIP-OVA and mucosal fragments were challenge, in vitro with NIP-HGG. Gastrin was determined after a preincubation and a challenged incubation period without supernatants. After antigenic challenge, supernatants were used for in vitro challenge in order to rule out the presence of a soluble mediator and activation of complement. In a second group of experiments Wistar rats were used to study in vitro the release of specific antibodies after antigenic challenge. With this experimental design we were able to show increased gastrin secretion after antigenic challenge in vitro in the presence of intact tissue. It is shown that the increased gastrin release is most probably mediated by activation of the complement system in the presence of antigen-antibody complexes. These are built up by specific anti-NIP antibodies and NIP-HGG used for the challenge. The complement system might be the final pathway of the observed increased gastrin release.

Highly reactive oxygen metabolites play an important role in inflammatory processes in the lung. Ambroxol (2-amino-3,5-dibromo-N-[trans-4- hydroxycyclohexyl]benzylamine) has been shown to reduce oxidant-mediated cell damage. However, the mechanism of this effect remains unclear. In order to evaluate oxidant scavenger function increasing concentrations of ambroxol (0-10(-3) mol/l) were compared with equimolar concentrations of N-acetylcysteine (NAC) and glutathione (GSH) in vitro to reduce OH. (hydroxyl radical), HOC1 (hypochlorous acid), O-2 (superoxide anion) and H2O2 (hydrogen peroxide). OH. was measured spectrophotometrically (deoxyribose assay); O-2 (xanthine/x-oxidase), H2O2 and HOC1 (HOC1/OC1-) were determined by chemiluminescence. Ambroxol, NAC and reduced GSH scavenged OH. significantly at 10(-3) mol/l, while HOC1 was inhibited at concentrations > or = 10(-4) mol/l completely (P < 0.01). NAC and GSH had no anti-O-2 function, while ambroxol (10(-4) mol/l) reduced O-2 by 14.3 +/- 6.7%. In contrast, GSH and NAC scavenged H2O2 at > 10(-6) mol/l (P < 0.01), while ambroxol had no anti-H2O2 effect. Our data demonstrate direct oxidant-reducing capabilities of ambroxol, which may be directly related to the aromatic moiety of the molecule. However, high concentrations (micromolar concentrations) are needed. Due to differences in direct oxidant scavenger function, a combination of ambroxol and NAC could be beneficial in antioxidant therapy.

Background: The effect of selective bile duct obstruction (SBDO) on hepatic reserve function of the bile duct obstructed (BDO) and nonobstructed freely draining (FD) lobes of the liver is obscure.

Methods: The bile duct branches draining from the left lateral and median lobes of the liver were ligated for 4 and 10 days in rats, and hepatic reserve functions in BDO and FD lobes were assessed by microsomal cholesterol 7 alpha-hydroxylase activities and by hepatic adenine nucleotides and energy charge levels. The values were compared with those in the sham-operated control liver. Cholesterol 7 alpha-hydroxylase activities were determined by gas-liquid chromatography--mass spectrometry, and hepatic adenosine triphosphate (ATP), adenosine diphosphate (ADP), and adenosine monophosphate (AMP) levels with high-pressure liquid chromatography.

Results: The histological examination of the BDO lobes showed proliferation and formation of new bile ductules and fibrous connective tissues linking portal areas. Microsomal cholesterol 7 alpha-hydroxylase activities, hepatic energy charge and each adenine nucleotide level did not differ between FD and BDO lobes, and the values were similar to those in the sham-operated liver.

Conclusions: Selective bile duct obstruction shows no adverse effects on microsomal and mitochondrial functions in both the BDO and FD lobes of the liver.

The pathophysiological role of endogenous leukotrienes in cardiovascular control and the regulation of renal function in congestive heart failure is not known. Therefore, in six conscious dogs with or without heart failure induced by right ventricular pacing (270/min, 10 days) we studied the effects of the leukotriene receptor antagonist FPL55712 on hemodynamics, plasma hormones and renal function. In healthy dogs, FPL55712 (1 mg kg-1 + 0.01 mg kg-1 min-1 i.v.) had little effect on hemodynamics, only reducing heart rate by 11% and insignificantly increasing systemic vascular resistance. Plasma levels of norepinephrine (-57%), renin (-30%) and aldosterone (-24%) were significantly decreased. Renal function parameters were not changed. In dogs with heart failure, FPL55712 significantly increased systemic vascular resistance (+16%) and decreased cardiac output (-15%). Plasma hormone levels were not changed, but renal plasma flow was decreased (-13%) and glomerular filtration rate (+12%), renal vascular resistance (+13%) and filtration fraction (+23%) were increased. It is concluded that there is no evidence for a contribution of endogenous leukotrienes to the systemic vasoconstriction in experimental heart failure. Whether the increase in systemic and renal vascular resistance induced by the leukotriene antagonist in dogs with heart failure reflects a role for endogenous leukotrienes with vasodilator action is still unclear and deserves further investigation.

We tested the hypothesis that reduction of intramyocardial cyclic guanosine monophosphate (GMP) by methylene blue (MB) would improve mechanical dysfunction in stunned myocardium. Regional stunning was produced in nine open-chest anesthetized dogs by a 12-min left anterior descending coronary artery (LAD) occlusion. MB was infused into the LAD during reperfusion (1 mg/kg per min). Stunning reduced LAD force development, introduced a significant time delay between the onset of force and shortening (delay) and caused significant systolic bulging to occur. Stunning reduced systolic regional work (the integrated product of force and segment shortening during systole), but did not significantly alter regional oxygen consumption or cyclic GMP levels. MB decreased cyclic GMP (1.8 +/- 0.2 to 0.9 +/- 0.1 pmol/g) and increased peak force (36 +/- 5 to 55 +/- 10 g). However, MB increased delay (93.9 +/- 18.4 to 233 +/- 19 ms) and systolic bulging (5.9 +/- 2.1% to 9.3 +/- 2.8%) and further reduced systolic regional work (control; 4204 +/- 933 g x mm/min; stunned: 2191 +/- 542 g x mm/min; MB: 1153 +/- 516 g x mm/min). MB increased regional myocardial oxygen consumption (7.4 +/- 1.0 to 15.6 +/- 2.7 ml O2/min per 100 g). These results suggest that depressed contractility, while present in myocardial stunning, is not the primary cause of mechanical dysfunction.

The effects of myenteric neuronal denervation on smooth muscle thickening and epithelial cell proliferation were studied in the descending colon of rats treated by serosal application of 2 mM benzalkonium chloride (BAC) for 30 min. Control animals were treated with saline (0.9% NaCl). The animals were divided into six groups of 13 animals each and killed 10, 45 and 120 days after BAC treatment. A significant reduction in neuron number was observed in the myenteric plexus of animals treated with BAC, as well as smooth muscle thickening and an increase in crypt cell population, crypt cell production per crypt and a decrease in cell cycle time. These findings permit us to conclude that a relationship may exist between the increase of epithelial cell proliferation, smooth muscle thickening and myenteric neuron denervation in the descending colon caused by BAC, the latter probably playing an important role in the integration of the other two.