Showing papers in "Research communications in chemical pathology and pharmacology in 1993"

Antioxidant activity of melatonin in mice.

Abstract: Melatonin (in gum tragacanth as solvent) was administered to mice in the dose range of 100 to 450 mg/kg intraperitoneally. It prevented the increase in plasma glucose resulting from pancreatic toxicity caused by the intravenous administration of alloxan at 40 mg/kg. This action of melatonin was significant and dose-dependent. In parallel work using mouse brain homogenates, melatonin and more so its principal hepatic metabolite, 6-hydroxymelatonin, inhibited the formation of colored products reacting with thiobarbituric acid. Again, this inhibition was significant and dose-dependent. Alloxan-induced diabetes and lipoperoxidation induced by thiobarbituric acid are imputed to the production of oxygen free radicals. The consistent results obtained using these two experimental models show the antioxidant activity of melatonin, both in vivo and in vitro. This effect may be reasonably attributed to the indole structure of the molecule.

Binding of taxol to human plasma, albumin and alpha 1-acid glycoprotein.

Abstract: The binding of taxol to human plasma and to individual plasma proteins was studied by equilibrium dialysis. Taxol was found to bind extensively (about 95%) without a significant difference between healthy volunteers and cancer patients. At clinically relevant concentrations (0.1-6 microM), the binding was found to be concentration independent, indicating nonspecific hydrophobic binding. Human serum albumin and alpha 1-acid glycoprotein were found to contribute about equally to the binding, with a minor contribution from lipoproteins. None of the drugs commonly coadministered with taxol (dexamethasone, diphenhydramine, ranitidine, doxorubicin, 5-fluorouracil and cisplatin) altered the binding of taxol significantly. The protein binding of taxol was found to dramatically decrease the red blood cell uptake of taxol.

Strength and endurance in the therapeutic evaluation of prednisolone-treated mdx mice

Abstract: The dystrophin-deficient, X-linked dystrophic mouse (mdx) was used to evaluate the efficacy of prednisolone treatment. A test protocol was used to take advantage of the quantifiable weakness and disability as well as molecular genetic defect shared with the X-linked Duchenne muscular dystrophy (DMD). Whole-body weakness and fatigue were determined by non-invasive force-transducer physiographic and variable-speed treadmill techniques, respectively. Other measurements included hind-limb muscle protein, calcium, and histomorphology. Subcutaneously-administered prednisolone elicited significant improvements in whole body strength throughout a two-month test period. Increases in strength were also accompanied by measurable increments in running endurance. In fact, prednisolone treatment appeared to protect mdx mice from the stressful effect of continuous running as determined by strength and muscle fiber diameter. Test results from this study support the limited therapeutic benefit observed previously in DMD patients treated with the glucocorticoid.

Effects of twenty-three drugs on the metabolism of FK506 by human liver microsomes.

Abstract: We investigated the effects of 23 drugs on the metabolism of FK506 by human liver microsomes. Acyclovir, amphotericin B, cefixime, cefotaxime, ciprofloxacin, cyclosporin A, diltiazem, enoxacin, erythromycin, ethinyl estradiol, fluconazole, fosfomycin, kanamycin, lincomycin, loxoprofen, minocyclin, nifedipine, nilvadipine, norethindrone, ofloxacin, phenobarbital, prednisolone, or rifampicin was added to the reaction media at equimolar or at ten times an excess molar ratio of the substrate concentration; their effects on FK506 metabolism were examined. Drugs known to be the substrate of cytochrome P-450 3A inhibited the metabolism of FK506, and among the drugs tested, the inhibition by cyclosporin A and nifedipine was the strongest.

Second branchial arch anomalies induced by fluconazole, a bis-triazole antifungal agent, in cultured mouse embryos.

Abstract: The teratogenic potential of the novel bis-triazole antifungal agent fluconazole (F) was investigated on mouse embryos during early organogenesis using the whole-embryo culture system. Early somite stage embryos (4-5 somites) were continuously cultured for 48 hours in rat serum added with 0, 25, 50 or 75 micrograms/ml of F. Doses of 50 and 75 micrograms/ml of F significantly (p < 0.001) impaired morphogenesis, resulting in a 83.3% and 91.6% of embryos showing second branchial arch anomalies, respectively. These findings demonstrate that F induces second branchial arch anomalies in cultured mouse conceptuses. Furthermore, these results may suggest an useful model for the study of arches-derived malformations.

Competitive irreversible inhibition of dopamine uptake by 6-hydroxydopamine.

Abstract: We examined the effects of 6-hydroxydopamine (6-OHDA) treatment on the human neuroblastoma cell line SK-N-SH-SY5Y (SY5Y) and the rat pheochromocytoma cell line, PC12. Structural and metabolic integrity was tested by measuring the ability of cells to transport the non-metabolizable amino acid analogue [3H]-alpha-aminoisobutyric acid (AIB). We determined that treatment with 6-OHDA at concentrations of 49 microM and 62 microM inhibited 50% of the AIB uptake in SY5Y and PC12 cells, respectively. Inhibition of AIB uptake was prevented by the addition of catalase, but was not influenced by the addition of 1 mM dopamine. This indicated that cell damage resulted from the generation of H2O2 and was independent of the catecholamine uptake system. Effects directly on the catecholamine uptake system were observed by measuring the uptake of 3H-dopamine. In contrast to the effects on amino acid uptake, dopamine uptake was significantly inhibited by 6-OHDA treatment, and this inhibition was not prevented by the addition of catalase. The results indicate a Ki of 430 microM for inhibition of dopamine uptake by 6-OHDA treatment of PC12 cells. The results are consistent with a competitive irreversible inhibition of the dopamine uptake sites by 6-OHDA or one of its metabolites. Thus, the lack of a catecholamine uptake-dependent cellular toxicity appears to result from the direct inactivation of catecholamine uptake sites. Similarly, the inhibition of dopamine uptake in vivo by 6-OHDA may be explained, at least in part, by direct inactivation of dopamine uptake sites rather than exclusively by intracellular transport and action of 6-OHDA.

Effect of NS-398, a new nonsteroidal anti-inflammatory agent, on gastric ulceration and acid secretion in rats

Abstract: The ulcerogenic activity of NS-398 was compared with that of indomethacin and the effects of NS-398 on stress-induced ulceration, gastric acid secretion and gastric mucosal prostaglandin (PG) contents were investigated in rats. NS-398 in a single dose of up to 1,000 mg/kg, p.o. did not significantly cause gastric ulceration while other nonsteroidal anti-inflammatory drugs such as, loxoprofen, indomethacin, diclofenac and ibuprofen, produced distinct gastric lesions. In cases of stress-induced ulcerations, NS-398 at 30 mg/kg, p.o. had no significant influence while indomethacin markedly potentiated the ulceration in a dose dependent manner. In basal gastric secretion studies, both NS-398 and indomethacin decreased secretion volume and acidity. However, in the 2-deoxy-D-glucose-stimulated gastric acid secretion study, NS-398 had no significant influence on gastric secretions while indomethacin significantly potentiated the secretion. Both NS-398 and indomethacin to much the same extent significantly decreased prostaglandin E2 (PGE2) contents in inflammatory tissue. However, with respect to gastric mucosal PGE2 contents, NS-398 did not decrease PGE2 contents while indomethacin significantly decreased the contents. It would thus appear that the absence of ulcerogenic properties of NS-398 is due to a relative lack of activity in inhibiting gastric PG synthesis.

Melatonin receptors in purified cell nuclei of liver

Abstract: Specific binding of [125I]iodomelatonin to homogenates from purified rat liver nuclei was characterized. The binding is rapid, reversible, saturable and of high affinity. Specific binding seems to be found in the nuclear protein fraction, since after precipitation of the proteins with trichloroacetic acid, the specific binding disappeared. The Kd (180 +/- 20 pM) and Bmax (9.1 +/- 0.03 fmol/mg protein) values (mean +/- S.E.M.) agree with the melatonin concentration in nuclei and may imply a physiological locus for melatonin action.

Effect of vanadyl sulfate feeding on susceptibility to peroxidative change in diabetic rats.

Abstract: The effects of vanadyl sulfate treatment on susceptibility to oxidative stress were investigated in streptozotocin-diabetic Wistar rats. A 2 x 2 factorial design was employed, with four groups of animals: 1) untreated, non-diabetic; 2) vanadyl-treated, non-diabetic; 3) untreated, diabetic; and 4) vanadyl-treated, diabetic. Vanadyl sulfate was administered as a 1.00 to 1.25 mg/ml solution in drinking water. Cataract development was entirely suppressed in vanadyl-treated compared to untreated, diabetic rats. STZ-induction of diabetes diminished glutathione (GSH) levels in liver homogenates; whereas vanadyl treatment resulted in restored levels of this nonenzymatic antioxidant. Thiobarbituric acid reactive substances (TBARS), both basal and iron-stimulated, were significantly elevated in all vanadyl-treated animals. Vanadyl treatment lowered liver glutamine synthetase activities in diabetic rats, but not in non-diabetic animals. Thus, vanadyl treatment was antioxidant in terms of cataract formation and reduced glutathione concentration in liver homogenates, pro-oxidant by reason of iron-stimulated TBARS formation and inconclusive with respect to glutamine synthetase activity. These results highlight the importance of using multiple indicators of peroxidative change in evaluating new pro-oxidant/antioxidant treatment regimens.

Effect of various dietary constituents on gastrointestinal absorption of aluminum from drinking water and diet.

Abstract: The influence of some frequent dietary constituents on gastrointestinal absorption of aluminum from drinking water and diet was investigated in mice. Eight groups of male mice received lactic (57.6 mg/kg/day), tartaric (96 mg/kg/day), gluconic (125.4 mg/kg/day), malic (85.8 mg/kg/day), succinic (75.6 mg/kg/day), ascorbic (112.6 mg/kg/day), citric (124 mg/kg/day), and oxalic (80.6 mg/kg/day) acids in the drinking water for one month. At the end of this period, animals were killed and aluminum concentrations in liver, spleen, kidney, brain, and bone were determined. All the dietary constituents significantly increased the aluminum levels in bone, whereas brain aluminum concentrations were also raised by the intake of lactic, gluconic, malic, citric, and oxalic acids. The levels of aluminum found in spleen were significantly increased by gluconic and ascorbic acids, whereas gluconic and oxalic acids also raised the concentrations of aluminum found in kidneys. Because of the wide presence and consumption of the above dietary constituents, in order to prevent aluminum accumulation and toxicity we suggest a drastic limitation of human exposure to aluminum.

Anxiolytic activity of melatonin in mice: involvement of benzodiazepine receptors.

Abstract: The anxiolytic properties of melatonin are revealed by two behavioral studies. In a free exploratory situation, the holeboard test, melatonin decreased head-dip performance. In an unconditioned conflict test, the light/dark box choice situation, melatonin increased the time spent in the lit box as well as the number of transitions between the two compartments. Melatonin was given in a dose range from 0.5 to 5.0 mg/kg body weight i.p. 30 minutes before testing in daytime. Moreover, the anxiolytic activity of diazepam (2.5 mg/kg i.p.) was evaluated and found to be completely inhibited by the specific benzodiazepine antagonist flumazenil (10 mg/kg i.p. 30 minutes before). In the same manner flumazenil counteracted melatonin activity in the two tests. Involvement of the benzodiazepine/GABAergic system in the anxiolytic activity of melatonin is discussed.

Possible action of human placental aminopeptidase N in feto-placental unit.

Abstract: Aminopeptidase N purified from human placenta actively hydrolyzed various immunomodulating peptides from their N-terminus such as splenopentin, thymopentin, thymic humoral factor gamma 2, tuftsin and rigin in vitro. Aminopeptidase N also actively hydrolyzed neuropeptide hormones (met-enkephalin, somatostatin and neurokinin A) and vasoactive peptides (lysyl-bradykinin and angiotensin III) from their N-terminus. In addition, angiotensin II, secretin, thymopoietin II peptide fragment, motilin, endothelin-I and insulin were tested for hydrolysis by aminopeptidase N. Km and Vmax values for the N-terminal amino acid, Thr, a liberation from tuftsin were 267 microM and 8.33 mumol/min/mg protein, respectively. L-Leucyl-p-nitroanilidase activity in the human placental membrane fraction was almost completely neutralized by anti-aminopeptidase N antibody. Our present study suggests that possible roles for surface enzyme aminopeptidase N in the human placenta would be to down-regulate the action of immunomodulating peptides as well as vasoactive and neuropeptide hormones, and to control both immunology and endocrinology of pregnancy.

Antiproliferative and antimetastatic effects of gossypol on Dunning prostate cell-bearing Copenhagen rats.

Abstract: Gossypol, a polyphenolic aldehyde naturally present in cottonseed, has long been recognized as a male contraceptive and recently as a potential anticancer agent. Our study used a rodent model to evaluate gossypol's potential for the treatment of human prostatic carcinoma. Two-month-old Copenhagen male rats received subcutaneous implants of a subpassage of MAT-LyLu prostatic cancer line, a highly metastatic, androgen-independent Dunning prostate tumor subline that specifically metastasizes to lymph nodes and lungs of recipients. After 2 weeks of gossypol treatment (0 or 12.5 mg/kg B.W./day S.C.) initiated immediately after transplantation, the rats were sacrificed and evaluated for prostate tumor growth and metastasis. Testosterone and gossypol levels in tumor tissue and various reproductive organs and serum potassium level were measured by radioimmunoassay (RIA), high pressure liquid chromatography (HPLC) and atomic emission spectroscopy (AES), respectively. Gossypol-treated rats exhibited weight reductions in developed MAT-LyLu prostate tumor mass and prostate of 24% (p < 0.05) and 31% (p < 0.05), respectively; whereas testicular and epididymal weights were not significantly affected. Few metastases (20%) were observed in either lymph nodes or lungs of gossypol-treated recipients. The control rats, however, had a much higher rate of lung (60%) and lymph node metastasis (40%). Testicular testosterone levels, as measured by RIA, were significantly lower in gossypol-treated rats than in controls (p < 0.05), but serum testosterone levels were not different. Extractable gossypol content in the prostate tumor, as measured by HPLC, reached 19.67 ng/gm and was 1.28 times higher than in liver, 1.98 times higher than in testes, but was 3.3% of that in prostate. Moreover, serum had the highest gossypol content (10.7 micrograms/ml). Serum potassium levels, as measured by AES, were significantly higher in gossypol-treated individuals than controls (p < 0.05). Our results indicate for the first time that gossypol has antiproliferative and antimetastatic effects on MAT-LyLu prostate cancer cells and can be explored as a potential therapeutic agent for androgen-independent human prostatic carcinoma.

The inhibitory actions of protease inhibitors on the production of polymorphonuclear leukocyte elastase and interleukin 8.

Abstract: The inhibitory actions of Ulinastatin, which is a protease inhibitor, on the production of polymorphonuclear leukocyte elastase (PMN-elastase) and interleukin 8 (IL-8) in vascular endothelial cells were evaluated. Our findings suggest that IL-8 plays a role in the production of PMN-elastase. Ulinastatin inhibited the lipopolysaccharide (LPS)-stimulated activity of polymorphonuclear leukocytes (PMN) and the production of IL-8 in vascular endothelial cells. Ulinastatin also inhibited the LPS-stimulated production of PMN-elastase in PMN.

beta-Casomorphin-immunoreactivity in the brain stem of the human infant.

Abstract: Using a peptide extraction procedure, reversed phase high performance liquid chromatography, and a radioimmunoassay that utilized an antibody raised specifically against human beta-casomorphin-8 (BC8), BC-immunoreactivity (BCIR) was detected in rostrocaudally increasing levels in nineteen microscopically distinct and functionally relevant areas of mesencephalon, pons cerebri, and medulla oblongata of eight infants. On the basis of the methodology used, it can be concluded, that the BCIR present in their brain stem was due to BC8 and/or to some of its congeners. Data in the literature together with those of this study indicate that beta-casomorphins could be transported by specific mechanisms from the blood into the brain stem and that they could play a role in the central regulation of various physiological phenomena.

Protective actions of YM737, a new glutathione analog, against cerebral ischemia in rats.

Abstract: Effects of YM737[N-(N-r-L-glutamyl-L-cysteinyl)glycine 1-isopropyl ester sulfate monohydrate], a new glutathione (GSH) analog more readily transported into cells than GSH, on cerebral ischemia were compared with those of GSH and some other drugs in rats subjected to occlusion of the bilateral carotid arteries. YM737 significantly reduced lethality, increased brain-water levels as measured by both dry-wet and NMR methods, and increased malondialdehyde (MDA) levels in the cerebral ischemic rats. On the other hand, pharmacological actions of GSH itself was less than those of YM737. In the ischemic rats used in the present study, there was no significant difference in 31P-NMR signals between the normal and the cerebral ischemic rats. These results suggest that YM737 showed anti-cerebral ischemic effects presumably due, in part, to inhibition of lipid peroxidative responses.

The role of lipid peroxidation and calcium in galactosamine induced toxicity in the rat liver.

Abstract: Liver homogenate and subcellular fractional levels of lipid peroxides, homogenate glutathione (GSH) and calcium contents and hepatic plasma membrane Ca 2+ -ATPase activity were determined in rats, 12, 18, 24 and 48 hours after treatment with a single dose of 1 g/kg galactosamine. Lipid peroxides and calcium levels in liver homogenate and subcellular fractions were found to increase, but hepatic GSH content and Ca 2+ -ATPase activity were observed to decrease reversibly

Role of Na(+)-K(+)-ATPase in airway smooth muscle relaxation by vasoactive intestinal peptide and pituitary adenylate cyclase activating peptide.

Abstract: To elucidate the effects of vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase activating peptide (PACAP) on airway smooth muscle function, we studied rabbit isolated tracheal segments under isometric conditions in vitro. Addition of VIP and PACAP dose-dependently relaxed tracheal smooth muscle precontracted with acetylcholine, with the order potency being PACAP (1) > or = VIP (0.78), accompanied by the corresponding increase in intracellular cyclic AMP levels. The VIP- and PACAP-induced muscle relaxations were significantly inhibited by ouabain and K(+)-free medium. Incubation of tissues with VIP reduced the contractile responses to electrical field stimulation, whereas PACAP had no effect. These results suggest that VIP and PACAP may cause bronchodilation through activation of Na(+)-K(+)-ATPase and that VIP but not PACAP inhibits the release of acetylcholine from the cholinergic nerve terminals.

Increased excretion of proline-containing peptides in dipeptidyl peptidase IV-deficient rats.

Abstract: We have reported that a substrain of Fischer 344 rat exhibits a deficiency of dipeptidyl peptidase IV (Watanabe et al. Experientia 43: 400-401, 1987). In this study, this rat model was found to excrete large amounts of proline-containing peptides in the urine. The peptide fraction also contained other amino acids, such as hydroxyproline and glycine. The results indicate that the enzyme might be involved in the metabolism of collagens and related peptides.

Insulin like effects of lithium and vanadate on the altered antioxidant status of diabetic rats.

Abstract: Lithium is widely used for treatment of behavioral disorders and has been shown to possess insulin-mimetic properties. The present study examines the in vivo effects of lithium alone, as well as in combination with vanadate (a potent insulin-mimetic agent), on the altered antioxidant status in the liver and kidney of diabetic rats. The elevated blood glucose levels in diabetic rats were about 50% restored by oral administration of lithium (0.3 mg/ml) and were completely normalized following vanadate addition (0.05 mg/ml) to lithium. Lithium therapy effectively normalized the decreased activities of catalase (CAT) and glutathione peroxidase (GSH-PX) but could not restore the lowered superoxide dismutase (SOD) in the liver of diabetic rats; while in kidney, the treatment proved to be ineffective. Inclusion of vanadate produced synergistic effect and caused partial restoration of the altered CAT, GSH-PX and CuZn-SOD levels in diabetic kidney and the depressed SOD activity in diabetic liver. These results suggest that lithium therapy may prove effective in improving the impaired antioxidant status during diabetes and vanadate supplementation at a low dose potentiates the effectiveness of lithium action.

Inhibitory effect of urate on oxidative damage induced by adriamycin-Fe3+ in the presence of H2O2.

Abstract: Urate strongly inhibited DNA damage induced by adriamycin (AD)-Fe3+ in the presence of H2O2. Deoxyribose (DOR) was degraded during the interaction of Ad-Fe3+ with H2O2, and urate strongly inhibited the DOR degradation. However, hydroxyl radical (HO.) scavengers did not always prevent the DOR degradation. ESR signal of 5,5-dimethyl-1-pyroline N-oxide-hydroxyl radical adduct was detected during the interaction of Ad-Fe3+ with H2O2. Mannitol and DNA strongly diminished the ESR signal but urate only poorly did. These results suggested that inhibitory effect of urate on the H2O2-dependent AD-Fe(3+)-induced DNA damage was not always due to trap only HO.. Lipid peroxidation of erythrocyte membrane was induced by AD-Fe3+ alone and H2O2 slightly inhibited the peroxidation reaction. Urate could not inhibit the Ad-Fe(3+)-induced lipid peroxidation. These results indicate that oxygen radical species which causes the lipid peroxidation does not involve in the H2O2-dependent Ad-Fe(3+)-induced DNA damage.

Vitamin A pretreatment and bleomycin induced rat lung injury.

Abstract: Possible antioxidant effects of pretreatment with vitamin A on bleomycin-induced rat lung injury were studied. Intratracheal bleomycin was administered to rats pretreated with vitamin A (50,000 IU/day) or vehicle control. Analysis of bronchoalveolar lavage fluid (BAL) total and differential cell counts, lung weight, lung pathology, and alveolar macrophage superoxide anion production were performed before and at various time points after the instillation of bleomycin. Bleomycin with vehicle raised total BAL cell count and the per cent of BAL neutrophils at day 7 post injury. The percent of lung involved with pneumonitis, the lung wet weight/body weight ratio and the alveolar macrophage superoxide anion production were also increased after bleomycin alone compared to the group pretreated with vitamin A. Rats pretreated with vitamin A demonstrated a statistically significant reduction in total BAL cell count and in alveolar macrophage superoxide anion production 7 days after bleomycin compared with vehicle control. Lung wet weight/body weight ratio 7 days after bleomycin was reduced in the vitamin A treated rats. There was a trend to less pneumonitis in the vitamin A pretreated group. These data suggest that vitamin A attenuates bleomycin induced pulmonary damage by a mechanism which involves inhibition of bleomycin-induced alveolar macrophage superoxide anion production.

Immunoreactive hepatocyte growth factor is present in tissue extracts from human breast cancer but not in conditioned medium of human breast cancer cell lines.

Abstract: Hepatocyte growth factor (HGF) is a novel mitogen for mature hepatocytes. In the present study, we have measured immunoreactive (ir)-HGF concentration in tumor extracts of 82 primary human breast cancers using an enzyme-linked immunosorbent assay (ELISA). Ir-HGF was detectable in all tissue extracts, the concentration ranging from 1.4 to 306.5 ng/100 mg protein (median value: 11.2 ng/100 mg protein). Correlation analyses between ir-HGF concentration and clinicopathological factors showed that the ir-HGF level was significantly higher in tumors with sizes of more than 5.0 cm compared with those less than 5.0 cm. In contrast, no detectable amount of ir-HGF was secreted into culture medium of two breast cancer cell lines, MCF-7 and ZR-75-1, suggesting that the cancer cell itself has no ability to produce ir-HGF.

Inhibition by methylene blue of the L-arginine metabolism to L-citrulline coupled with nitric oxide synthesis in cultured endothelial cells.

Abstract: The effect of methylene blue on the metabolism of L-arginine to L-citrulline coupled with nitric oxide synthesis in cultured endothelial cells was investigated. The addition of the calcium ionophore ionomycin (10(-8)-10(-5) M) to endothelial cells stimulated L-citrulline formation from L-arginine. Ionomycin-stimulated L-citrulline formation was inhibited by NG-nitro-L-arginine (10(-6)-10(-4) M), a potent inhibitor of nitric oxide synthase. These results indicate that ionomycin-stimulated L-citrulline formation was coupled to nitric oxide synthesis. Therefore, L-citrulline formation from L-arginine was used as a marker for nitric oxide synthesis. Methylene blue (10(-4)-10(-3) M) inhibited ionomycin-stimulated L-citrulline formation in bovine aortic endothelial cells. Moreover, the formation of L-citrulline by the particulate fraction obtained from bovine cultured endothelial cells was also inhibited by methylene blue concentrations in excess of 10(-5) M. Although the addition of methylene blue to various tissues has been reported to release superoxide anions (Wolin et al., 1990; Marczin et al., 1992), inhibition of L-citrulline formation by methylene blue was not affected by the presence of superoxide dismutase (100 micrograms/mL) in either cultured endothelial cells or the particulate fraction. These findings suggest that methylene blue is a direct inhibitor of nitric oxide synthase in bovine aortic endothelial cells.

Buccal administration of human insulin in streptozocin-diabetic rats.

Abstract: This study investigates the use of the buccal route of administration in the delivery of human insulin in rats. Streptozocin-induced diabetic female Wistar rats were used in this study. Insulin (100 U) either free (i.e., insulin solution) or associated with a carrier, namely erythrocyte-ghosts (EG) and liposomes-vesicles (LEV), was administered buccally. Blood samples were collected from the tail over a period of 5 hr. These results indicate that insulin absorption occurred, as evidenced from a decrease in blood glucose concentration, and in the case of free insulin and erythrocyte-ghosts-insulin (EG-INS). The magnitude of the blood glucose level decline was at its maximum of 39.53 mg/dl (at 2 hr) and 26.23 mg/dl (at 4 hr) for free insulin and EG-INS, respectively. No significant difference in the blood glucose level profile was observed after either LEV or liposomes-vesicles-insulin (LEV-INS). This study demonstrates the ability of human insulin to be absorbed from the mouth cavity when it is instilled in the form of a simple solution or EG-INS suspension. This absorption resulted in a definite pharmacological effect but not a significant therapeutic effect.

Dexamethasone treatment in utero enhances neonatal cholinergic nerve terminal development in rat brain.

Abstract: Fetal glucocorticoid administration has been proposed to elicit both promotional and inhibitory effects on neuronal development. In the current study, pregnant rats were given 0.05, 0.2 or 0.8 mg/kg of dexamethasone on gestational days 17, 18 and 19, and the effects on development of central cholinergic projections was assessed on postnatal day 1 by measuring the specific binding of [3H]hemicholinium-3 to the high affinity choline transporter localized in cholinergic nerve terminal membranes. Dexamethasone produced a dose-dependent retardation of brain region growth, but enhanced [3H]hemicholinium-3 binding in both the forebrain and the midbrain + brainstem. At the highest dose, the promotional effect on [3H]hemicholinium-3 binding was lost in the forebrain, a region that is particularly sensitive during late gestation to inhibitory effects of glucocorticoids on neuronal development. These results indicate that, even in the face of growth retardation, glucocorticoids promote the development of central cholinergic projections; however, at high doses, inhibitory actions of the steroid can offset the promotional effects in some regions.

Inhibition of cholinesterases by histamine 2 receptor antagonist drugs

Abstract: Many studies have demonstrated that histamine 2 receptor antagonists (H2RA) have in vitro anticholinesterase effects, but discrepancies about type and potency of this inhibitory effect exist among published results. Moreover, cholinesterase inhibition has not been shown in patients receiving H2RA. These discrepancies led us to study the in vitro antibutyryl- and in vitro antiacetylcholinesterase activities of ranitidine, cimetidine, nizatidine comparatively to pyridostigmines. Plasma cholinesterase activity (PCEA), erythrocyte cholinesterase activity (ECEA) and plasma ranitidine levels were measured in six patients before and during continuous IV infusion (150 or 200 mg/d) of ranitidine. Our in vitro results confirm the weak anticholinesterase activity of H2RA. Ranitidine is the most potent inhibitor of butyrylcholinesterase (Ki = 61 microM). Ranitidine and nizatidine are the most potent inhibitors of acetylcholinesterase (Ki' = 2.1 microM, Ki' = 5.1 microM, respectively) but one thousand times less effective than pyridostigmine (Ki = 0.003 microM). The results in patients show no statistically significant difference between PCEA and ECEA measured before and during ranitidine infusion (plasma ranitidine levels between 0.31 and 1.25 microM).

Fate of cadmium bound to phytochelatin in rats.

Abstract: The fate cadmium(Cd) bound to phytochelatin [PC, (gamma-Glu-Cys)n-Gly)] was studied in rats using synthesized 109Cd-PC. Less Cd was absorbed through the digestive tracts than CdCl2, but the ratio of renal Cd to hepatic Cd was higher. After parenteral administration of Cd-PC, Cd was distributed mainly in the liver, kidney, small intestine and pancreas. More Cd was found in the kidney than the liver after Cd-PC (n = 5) administration. Most of the Cd was bound to the high molecular weight fraction in the hepatic cytosol 0.5 hr after administration and moved to the metallothionein fraction at 6 hr. The tissue distribution of Cd was not affected even when free PC (n = 5) was administered 3 hr after or before Cd injection. The distribution in the kidney increased only in the case of the simultaneous administration of Cd with PC. These findings show that the absorbance of Cd bound to PC from the alimentary tract is lower than that of CdCl2 although absorbed Cd is distributed to the kidney more than CdCl2, and Cd is liberated from PC soon after uptake by the cells.

Metabolism of trimipramine in vitro by human CYP2D6 isozyme.

Abstract: In vitro metabolism of the tricyclic antidepressant trimipramine using a commercial preparation of human CYP2D6 isozyme expressed in a human cell line is described. 2-Hydroxytrimipramine and a previously unreported metabolite, 2,10- or 2,11-dihydroxytrimipramine were isolated. Their structures were determined by gas chromatography/mass spectroscopy of underivatized and derivatized extracts. Acetylation of the new metabolite resulted in dehydration at C10 to give 10,11-dehydro-2-acetoxytrimipramine. No N-dealkylation of trimipramine was observed. Prior administration of quinidine produced a large reduction in the metabolic oxidation of trimipramine with CYP2D6 while prior administration of quinine had no effect. The use of this CYP2D6 isozyme preparation in vitro is of value in the identification of possible in vivo substrates for the human CYP2D6 isozyme.

Differential effects of fatty acyl coenzyme A derivatives on citrate synthase and glutamate dehydrogenase.

Abstract: We investigated the hypothesis that one mechanism underlying fatty acid toxicity is the selective inhibition of rate-limiting and/or regulated tricarboxylic acid cycle and related enzymes by fatty acyl coenzyme A (CoA) derivatives by examining the effects of several fatty acyl CoAs on purified citrate synthase (CS) and glutamate dehydrogenase (GDH). The results indicate that, at pathophysiological levels, palmitoyl CoA, a long-chain acyl CoA, is a potent inhibitor of CS and GDH with IC50 values of 3-15 microM. At much higher levels (in the pathological and toxicological range), octanoyl and decanoyl CoA (medium-chain acyl CoAs) inhibited both enzymes with IC50 values of 0.4-1.6 mM. Butyryl CoA, a short-chain acyl CoA, inhibited CS (IC50 = 0.9 mM) at toxicological levels but inhibited GDH poorly. These results suggest that the long-chain fatty acyl CoA inhibition of CS and GDH may assume some pathophysiological importance in fatty acid toxicity and in metabolic encephalopathies in which organic acidemia is persistent. The findings also provide additional support for the original hypothesis.