Hepatic bile formation in the rat

While changes in gastric, pancreatic, and intestinal secretion in response to more recently identified gastrointestinal peptides have been characterized, there has been less investigation into effects of these hormones on hepatic bile production. The isolated perfused rat liver model has been used to examine effects of vasoactive intestinal peptide (VIP), somatostatin, bombesin, and thyrotropin-releasing hormone (TRH) on bile flow and bile acid transport. No changes were seen following bolus administration of bombesin (3×10^–8–1.5×10^–6 M) or TRH (3×10^–7–3×10^–6 M), while somatostatin (6×10^–6 M) produced a small decrease in bile flow without any change in bile acid output. VIP (3×10^–7 M) caused a highly significant increase in both volume of bile flow (0.85±0.8 to 1.11±0.09 l/min/g liver,P<0.001) and bile acid output (31.6±1.5 to 43.2±1.7 nmol/min/g liver,P<0.001). Elimination of Ca²⁺ from liver perfusate did not prevent VIP-induced increases in bile flow and bile acid output, and no synergistic effect of concomitant theophylline administration was observed. While effects of VIP on bile flow appear to be due to alterations in hepatic transport of bile acids, the exact mechanism(s) producing these changes remains to be elucidated.This work was supported by National Institutes of Health grant AM-32208 and the Research Service of the Veterans Administration.     

Inhibitory neuropeptides and intrinsic inhibitory innervation of descending human colon

The effects of aging on inhibitory neuropeptide concentrations and intrinsic inhibitory innervation of circular muscle were investigated using normal descending colon obtained at surgery. Immunoreactive vasoactive intestinal peptide, peptide histidine-methionine, met⁵-enkephalin, neuropeptide Y, and somatostatin were extracted from specimens of muscularis externa (patient ages: 19–84 years) and measured by radioimmunoassay. Intracellular electrical activity was recorded from strips of circular muscle (patients ages: 49–84 years) using glass microelectrodes; inhibitory junction potentials were evoked by electrical field stimulation. There were no significant differences (t tests:P>0.05) between neuropeptide concentrations in patients<70 years old (N=28) compared to patients70 years old (N=12). However, the amplitude of inhibitory junction potentials declined with increasing patient age (r=–0.58,P=0.02,N=16), with no change in resting membrane potentials (r=0.22;P>0.05). The decline in amplitude in women (r=–0.68,P=0.03,N=9) preceded the decline in men (r=–0.62,P=0.10,N=7). Age-related decline in inhibitory junction potentials could be related to decreased: density of inhibitory nerves, release of inhibitory neurotransmitter, density of binding sites for inhibitory neurotransmitter on smooth muscle, or a combination thereof. Alternatively, this decline might represent a change in interaction of inhibitory neurotransmitter with the smooth muscle membrane, such as a change in coupling of binding site with the potassium channel, decreased number of potassium channels, or altered permeability of the potassium channel.This study was supported in part by the National Institutes of Health (DK 17238 and DK 34988), and by VA Medical Research funds.     

Decreased vasoactive intestinal peptide levels and glutathione depletion in acquired megacolon

We reported decreased vasoactive intestinal peptide levels in acquired megacolon. The origin of altered neuropeptide levels is unknown, but recent work suggested that tissue antioxidants may function as neuroprotectants. Our hypothesis was that altered levels of inhibitory neurotransmitters in human colon are associated with depletion of the tripeptide thiol, glutathione. Normal colon samples (N=10; from patients 41–80 years old) and acquired megacolon samples (N=10; from patients 31–98 years old) were obtained at surgery. Vasoactive intestinal peptide levels were decreased in muscularis externa from acquired megacolon (P=0.01), while there was a modest increase in NADPH diaphorase activity in muscularis externa from megacolon (P=0.10). Glutathione in acquired megacolon was detectable in muscularis externa from only five specimens (P<0.05), but was not significantly different (P>0.05) in the mucosal-submucosal layer. The results supported the presence of vasoactive intestinal peptide and NADPH diaphorase in distinct subpopulations of nerves in human colon. The results also supported the hypothesis that glutathione functions as a neuroprotectant in a subset of patients with acquired megacolon.Supported by VA Medical Research Funds.     

Enteric neural modulation of slow-wave activity in cat colon

These studies test the hypothesis that the generation of colonic slow waves can be modulated by stimulation of intrinsic enteric nerves and attempt to identify a neurotransmitter that may be responsible for this change in slow-wave activity. Isolated segments from the mid-colon of the cat generated regular, continuous slow waves at 6.5±1.1 cpm. Activation of the intrinsic nerves by electrical field stimulation transiently reduced the rate of slow-wave generation to 4.7±0.7 cpm (P<0.001). The response to electrical stimulation was blocked by tetrodotoxin and -chymotrypsin. The following antagonists were not effective in blocking the response: atropine, hexamethonium, phenoxybenzamine, propranolol, methysergide, naloxone, or imidazole. Vasoactive intestinal polypeptide (5×10^–7 M) decreased slow wave frequency to 4.5±0.4 cpm. Vasoactive intestinal polypeptide (VIP) fragment 10-28 inhibited the effect of electrical field stimulation but also decreased the slow-wave frequency. VIP-immunoreactive nerves were much more abundant in the plexus submucosus extremus than in the circular muscle of the muscularis externa. Thus, pacemakers for colonic slow waves may be modulated by intrinsic colonic nerves, and vasoactive intestinal polypeptide may be the neurotransmitter responsible for this modulation.This work was supported by NIH grants DK 1750 and Digestive Diseases Core Center grant AM 34986 and a grant from the Veterans Administration.     

Myoelectric and contractile effects of motilin on dog small intestinein vivo

The effects of the close intraarterial administration of motilin on intestinal myoelectric and contractile activities were examined in 37 dogs. After anesthetization or decerebration, a segment of proximal jejunum was instrumented serosally with electrodes and stain gauges. A mesenteric artery supplying a short length of this segment was cannulated for the injection of motilin and other agents. Motilin (0.03–0.3 g) caused: (1) a series (1–5 min) of phasic contractions and electrical response activity (ERA) bursts locally; (2) a short (15–60 sec) series of phasic contractions and ERA bursts aborally followed by relaxation; and (3) a series of phasic contractions and ERA bursts whose onset migrated 3.7±1.0 cm orally. The length of orad response increased to 6.6±1.9 cm in the decerebrate dogs (P<0.01). No other tested agent, including serotonin, bethanechol, morphine, dopamine, substance P, neurotensin, somatostatin, vasoactive intestinal peptide, bombesin, pentagastrin, cholecystokinin octapeptide, prostaglandin F₂ or leucine-enkephalin, cuased similar responses. All motilin responses were mediated by neural pathways consisting of both nicotinic and muscarinic receptors. The similarity of responses and mechanisms of action of the motilin-activated contractile response with the intrinsic mucosal reflex suggested that motilin may mediate this reflex.This work was supported in part by VA grant 7722-01P.     

Role of vasoactive intestinal peptide (VIP) in pathogenesis of ethanol-induced gastric mucosal damage in rats

To elucidate the possible role of vasoactive intestinal peptide (VIP) in the pathogenesis of acute gastric mucosal damage, rats were treated intragastrically with 1.0 ml 96% ethanol with or without intravenous or intraperitoneal coadministration of VIP (1 nmol/liter to 1 mol/liter/100 g). VIP was found to double the mean lesion area when compared with that induced by ethanol alone (P<0.05), an effect that was prevented by VIP antagonist (1 mol/liter/100 g). A substance P antagonist (1 mol/liter/100 g) also reduced the extent of gastric damage induced by coadministration of VIP and ethanol. VIP antagonist or substance P antagonist significantly reduced ethanol-induced gastric mucosal damage. Gastric mucosal levels of LTB₄, LTC₄, VIP, and substance P were significantly increased in ethanol-treated rats as compared with saline-treated animals (P<0.05). The augmentation of ethanol-induced damage by VIP was associated with increased gastric mucosal levels of LTB₄. In VIP-treated rats, gastric mucosal levels of substance P were found to be significantly increased compared with control rats (P<0.05). Administration of VIP to pyloric-ligated rats significantly increased gastric acid output and blood pepsinogen A levels as compared with saline treated rats (P<0.05). Ketotifen, a mast cell stabilizer (100 g/100 g), administered orally 30 min before damage induction by ethanol, with or without VIP, totally abolished the damage of the surface epithelium of the entire gastric mucosa and significantly reduced the mucosal levels of LTC₄ and LTB₄ (P<0.05). It is suggested that VIP is involved in the pathogenesis of acute ethanol-induced gastric mucosal damage. The effective mucosal protection by ketotifen suggests a role for mast cells and their mediators in the pathogenesis of acute gastric mucosal damage.     

Changes in the somatostatin,substance P and vasoactive intestinal polypeptide content of the gastrointestinal tract following streptozotocin-induced diabetes in the rat

Summary Rats with streptozotocin-induced diabetes of 10 weeks' duration showed significant changes in the total content of somatostatin, substance P and vasoactive intestinal polypeptide in the stomach and small intestine compared with control animals. An increase (p<0.05) in the concentration and total content of gastric somatostatin and a decrease (p<0.05) in the concentration and content of gastric substance P were seen in the streptozotocin-treated rats. The increase in the vasoactive intestinal polypeptide (VIP) content (54%, p<0.05) and the decrease in the substance P content (35%, p<0.05) of the gut may contribute to the impaired intestinal motility observed in animals with experimentally produced diabetes. Both the diabetogenic effect of streptozotocin and the changes in regulatory peptide concentrations were prevented by injection of nicotinamide before streptozotocin suggesting that the changes did not arise from a non-specific toxic effect of streptozotocin upon gastrointestinal neurones and/or endocrine cells.     

Human intraepithelial lymphocytes

This study evaluates the immunomodulation and receptor binding of vasoactive intestinal peptide on human peripheral blood lymphocytes and intraepithelial lymphocytes. Vasoactive intestinal peptide (VIP, 10^–8 and 10^–12 M) had no effect on the concanavalin A-induced proliferation or the spontaneous cytotoxicity against K-562 targets by either lymphocyte type. Human peripheral blood lymphocytes had a mean of 927 vasoactive intestinal peptide receptors per cell with a K_d of 1.12×10^–10 M, as demonstrated by the competitive displacement of [¹²⁵I]peptide by unlabeled peptide using Scatchard analysis. In contrast, intraepithelial lymphocytes had no high-affinity receptors as shown by the negligible binding of 50 pM [¹²⁵I]VIP. Peptide binding by peripheral blood lymphocytes, although reduced by exposure to dithiothreitol and ethylenediamine tetraacetic acid, was still greater than binding by intraepithelial lymphocytes. As intraepithelial lymphocytes are mainly CD8⁺ T cells, the possibility that this phenotype may not bind VIP at all was tested by specifically depleting peripheral blood lymphocytes by antibody and complement lysis. Peripheral blood lymphocytes expressing CD8, CD4, and/or CD2 were responsible for most of the binding, indicating that CD8⁺ T lymphocytes in the peripheral blood and in the intestinal epithelium differ in their capacity to bind VIP.This work was supported by grants from the National Foundation for Ileitis and Colitis and from the National Institutes of Health (RO1DK42166).     

Mechanism of bradykinin-induced cyclic GMP accumulation in bovine tracheal smooth muscle

Bradykinin (10^–8-10^–5M) caused a concentration-dependent increase in cyclic GMP (cGMP) production in bovine tracheal smooth muscle in the absence of epithelium. The effect was calcium-dependent and was inhibited by pyrogallol (10 M) and methylene blue (10 M). The inhibition of pyrogallol was reversed by superoxide dismutase (100 Usnowml). Nitric oxide (NO) synthase inhibitors, N ^G-methyl-l-arginine (10–100 M) and N ^G-nitro-l-arginine (10–100 M) reduced cGMP accumulation induced by bradykinin in a concentration-dependent fashion, and the inhibition was reversed by l-arginine. Immunohistochemistry with a specific antibody against neuronal NO synthase from rat cerebellum showed positive staining localized in some nerve fibers. Bradykinin-induced cGMP accumulation appears to be related to the release of NO, part of which is probably synthesized in nonadrenergic noncholinergic nerve in bovine trachea.Offprint requests to: Dr. Hong Sheng     

Gastric mucosal cell protection by epidermal growth factor in primary monolayer culture of guinea pig gastric mucous cells

Background. Epidermal growth factor (EGF) has an anti-ulcer effect, but the mechanisms of this gastric mucosal protection are incompletely understood. We have suggested the importance of mucin as a mucosal protectant. We investigated whether increased mucin biosynthesis might be involved in the gastric mucosal protection conferred by EGF. Methods. EGF and then ethanol were added to primary monolayer cultures of guinea pig gastric mucous cells, in which factors such as gastric acid and gastrointestinal hormones were excluded. Mucin and prostaglandin E₂ (PGE₂) were assayed. Results. Cytoprotection induced by EGF was demonstrated. Mucin biosynthesis and PGE₂ release were both significantly increased by EGF. When endogenous PGE₂ synthesis was inhibited by pretreatment with 10^–5M or 10^–4M indomethacin (IND), mucin biosynthesis was still significantly increased by EGF. Ethanol-induced cell damage was concentration-dependent in cultures with no other additions (normal PGE₂ and mucin biosynthesis). Damage by ethanol was decreased by EGF pretreatment (increased PGE₂ and mucin biosynthesis). Damage by ethanol was increased by 10^–5M IND pretreatment (decreased PGE₂; normal mucin biosynthesis) and by 10^–4M IND pretreatment (decreased PGE₂ and mucin biosynthesis). Ethanol-induced damage was decreased by EGF pretreatment even in the presence of 10^–5M and 10^–4M IND (decreased PGE₂; increased mucin biosynthesis). Conclusions. Increased mucin biosynthesis, induced by EGF independently of PGE₂, protects gastric mucous cells.     

α-Adrenergic responses of isolated canine coronary microvessels

Although -adrenergic activation is known to increase coronary microvascular resistance in vivo, the magnitude of its segmental microvascular consequences is not well understood. Quantification of these effects in vivo is hindered by escape mechanisms that minimize the influences of constrictors, and alterations in flow and pressure, which effect microvascular tone by shear stress-dependent and myogenic mechanisms, respectively. To eliminate these confounding influences, we have studied responses in vitro under conditions with these variables controlled. We evaluated the diameter changes of isolated canine coronary arterioles (110±12 m, n=35) and venules (98±7 m, n=9) in response to -adrenergic activation by norepinephrine (10^–10 to 10^–4 M) in the presence of -adrenergic blockade by alprenolol (10^–6 M). In contrast to the situation in vivo, -adrenergic activation did not constrict isolated coronary arterioles, but constricted isolated coronary venules in a dose-dependent manner over a range of 10^–10 to 10^–4 M (–27 ±3% maximum diameter change). Coronary arteriolar -adrenergic constriction was not promoted by 1) subthreshold or vasoactive doses of the vasoconstrictors KCl, angiotensin II, U46619, endothelin-1, neuropeptide Y or arginine vasopressin, 2) inhibition of the presynaptic uptake of norepinephrine by imipramine (10^–6 M), 3) inhibition of EDRF synthesis by N^g-monomethyl-L-arginine (10^–5 M) or 4) inhibition of prostaglandin synthesis by indomethacin (10^–5 M). Furthermore, -adrenergic activation did not modify microvascular dilatation by adenosine (10^–9 to 10^–4 M) or nitroglycerin (10^–9 to 10^–4 M), suggesting that -adrenergic constriction in vivo is not due to attenuation of cAMP or cGMP-dependent mechanisms of coronary dilatation. In contrast to the lack of constriction in coronary arterioles, canine skeletal muscle arterioles exhibited significant -adrenergic constriction (–80±4%), maximum diameter change). The coronary venular -adrenergic constriction was significantly inhibited by both the ₁-and ₂-adrenergic receptor antagonists, prazosin (10^–8 M) and rauwolscine (10^–7 M), indicating a mixed population of ₁-and ₂-adrenergic receptors. These results suggest that coronary arterioles, but not venules, lose -adrenergic responsiveness during isolation and cannulation, or that the primary coronary microvascular response to -adrenergic activation is venular constriction.     

Cytoreductive effect of recombinant alpha interferon in patients with myelofibrosis with myeloid metaplasia

Summary In an attempt to reduce myeloproliferation, we administered recombinant -2b interferon (r-INF) to ten patients with myelofibrosis with myeloid metaplasia (MMM) in a hypercellular phase, as part of a phase II trial. Two patients experienced severe side effects and stopped treatment before completion of the first week. In the eight evaluable patients, r-INF was given for 16 weeks at an initial dosage of 3×10⁶ U/day, with monthly increments in the case of response failure, i.e. a decrease in WBC or platelet count of less than 25% of the initial value. Two cases responded at the starting dosage, while the effective dosage was 5×10⁶ U/day in the others. At the end of the 16th week, Hb showed minor changes: from an initial value of 12.08 g/dl, range 8.3–17.3, to 11.6 g/dl, range 7.7–18 (P=0.12); WBC were reduced from 54×10⁹/l, range 6.4–69.4, to 17.5×10⁹/l, range 5–39 (P=0.09, 4/8 responses); platelets decreased from 775×10⁹/l, range 215–1748, to 403×10⁹/l, range 118–730 (P=0.008, 8/8 responses). Minor changes in spleen size were also noted, while no significant changes in bone marrow fibrosis occurred. Influenza-like symptoms and fatigue were common side effects. In conclusion, r-INF has a role as a non-leukemogenic cytoreductive agent in the therapy of MMM, especially for cases with thrombocytosis.     

Expression of functional Y1 receptors for neuropeptide Y in human Ewing's sarcoma cell lines

Summary In the human Ewing's sarcoma cell line WE-68, saturation analysis using³H-labelled neuropeptide Y ([³H]NPY) as the radioligand disclosed a homogeneous population of binding sites with a dissociation constant (K _d ) of 4.5 nM and maximal binding capacity (B _max) of 712 fmol/mg cell protein. Besides the WE-68 cell line, ten other human Ewing's sarcoma cell lines (FM-62, HS-80, HT-78, HT-M1-78, NT-68, RM-82, RS-63, VH-64, WE-M1-68, WE-M2-68) were also found to display NPY receptors withK _d varying from 3.5 nM to 10.7 nM andB _max=247–3744 fmol/mg cell protein. NPY, its natural analogues and the Y₁-receptor-specific peptide ligand [Leu³¹, Pro³⁴]NPY inhibited [³H]NPY binding in the potency order: [Leu³¹,Pro³⁴]NPYhuman NPYpeptide YY (PYY)>> salmon pancreatic polypeptide (PP) > human PP>porcine NPY_13–36NPY_22–36. In the Ewing's sarcoma cell lines NPY provoked inhibition of forskolin-stimulated cyclic AMP formation by up to 98%. Pertussis toxin alleviated the cyclic-AMP-inhibitory response to NPY. In isolated Ewing's sarcoma plasma membranes pertussis toxin [³²P]ADP-ribosylated a 41-kDa protein. The ability of NPY and analogues to inhibit cyclic AMP accumulation paralleled their potencies in displacing radioligand binding. By contrast, a cell line derived from an atypical form of Ewing's sarcoma did not express specific and functional NPY receptors. These results demonstrate that conventional Ewing's sarcoma cells possess G_i-potein-coupled NPY receptors of the Y₁ type, which upon interaction with NPY, PYY, and PP mediate inhibition of cyclic AMP generation.Abbreviations NPY neuropeptide Y - PP pancreatic polypeptide - PYY peptide YY - VIP vasoactive intestinal peptide     

Time-related interference of misoprostol with experimental gastric cancer formation induced byN-methyi-N′-nitro-N-nitrosoguanidine in the rat

Summary The purpose of this study was to investigate the effect of long-term misoprostol administration, at non-antisecretory doses, onN-methyl-N-nitro-N-nitrosoguanidine(MNNG)-induced gastric carcinogenesis. The incidence of gastric carcinomas and precancerous lesions was evaluated in 50 male 250-g Sprague-Dawley rats after 52 weeks of continuous oral administration of MNNG (120 mg/l;n=20), MNNG plus misoprostol (2 mg kg^–1 day^–1;n=20) or tap water (n=10) (experiment 1), and in 30 rats treated with MNNG for 30 weeks followed by tap water (n=15) or by misoprostol (n=15) for 22 weeks; a third group (n=10) received tap water only for 52 weeks (experiment 2). After sacrifice, gastric mucosal lesions were macroscopically evaluated and their histology obtained. MNNG consumption was comparable in all groups (6.5±1.1 mg rat^–1 day^–1). Misoprostol consumption was 180±0.25 mg kg^–1 day^–1 rat^–1. In experiment 1 the incidence of gastric carcinomas was 60% in the MNNG group and 25% in the group treated with MNNG plus misoprostol (P<0.05). Cytotoxic and hyperplastic gastric mucosal lesions were also significantly reduced by misoprostol. In experiment 2 the incidence of carcinomas was 31% and 38.6% respectively. Misoprostol significantly decreased the incidence of gastric cancer formation when given from the beginning of the experiment. By contrast, when administered after 30 weeks of MNNG treatment it did not interfere with experimental gastric cancer formation. Exogenous prostaglandins are able to prevent the early MNNG-induced gastric mucosal lesions, thus interfering with gastric carcinogenesis.Abbreviation MNNG N-methyl-N-nitro-N-nitrosoguanidine This work is part of the Panisperna Project on Cancer and was supported by grants from the Ministry of Public Education and the National Research Council     

Nitric oxide mediates inhibitory nerve effects in human esophagus and lower esophageal sphincter

The effect of inhibition of nitric oxide synthase on nonadrenergic, noncholinergic nerve-mediated responses in circular smooth muscle of the human esophageal body and lower esophageal sphincter (LES) was examinedin vitro. Tissues were obtained from 10 patients (eight esophageal resection for cancer, two transplant donors). Muscle strips from the LES developed significant spontaneous tension (11.6 ± 2.1 mN/mm²,N=6) and relaxed in response to electrical stimulation. The nitric oxide synthase inhibitor,N -nitro-l-arginine (NNA), at 10^–5 M, inhibited the relaxation, but had no significant effect on the spontaneous tension (13.0 ± 2.6 mN/mm²,P=0.07). Esophageal body strips developed little spontaneous tension, demonstrated an off contraction following the cessation of the electrical stimulus, and when contracted with 10^–5 M carbachol, relaxed during electrical stimulation. NNA (10^–5 M) inhibited the off contraction and the relaxation seen after carbachol and unmasked a prominent intrastimulus contraction. This intrastimulus contraction was enhanced by eserine and inhibited by atropine and tetrodotoxin. NNA showed similar potency in the esophageal body and LES and its effects were reversed byl-arginine, but notd-arginine. The results indicate that nitric oxide is an important mediator for nonadrenergic, noncholinergic nerve effects in the human esophagus and lower esophageal sphincter.This research was supported in part by an ICI Pharma/Medical Research Council of Canada Research Fellowship grant awarded to H.G. Preiksaitis and a Medical Research Council Program Grant PG8.     

Nitric oxide-mediated neurotransmission is attenuated in the anococcygeus muscle from diabetic rats

Summary The effect of STZ-induced diabetes of 8-weeks duration was examined on nitric oxide-mediated neurotransmission in the rat anococcygeus muscle. In the presence of noradrenergic blockade and raised tissue tone, relaxant responses to nerve stimulation (0.5–5 Hz, for 10 s), sodium nitroprusside (5 and 10 nmol/l) and nitric oxide (1 and 3 mol/l) were significantly reduced in anococcygeus muscles from diabetic rats compared to responses from control rats (p <0.05). In contrast, relaxations to papaverine (3 and 10 mol/l were not reduced in tissues from diabetic rats. The nitric oxide synthesis inhibitor NOLA (100 mol/l) abolished relaxant responses to nerve stimulation but had no effect on responses to any of the relaxant agents used. Exposure to NOLA at 10 mol/l reduced stimulation-induced relaxations; this reduction was significantly greater in tissues from the diabetic group than from the control group (p <0.05), probably as a consequence of the smaller relaxant responses in muscles from diabetic rats. Contractile responses to nerve stimulation (1–10 Hz, for 10 s), but not noradrenaline (0.03–30 mol/l), were significantly greater in anococcygeus muscles from diabetic rats than from control rats (p <0.05). NOLA (100 mol/l) significantly enhanced stimulation-induced contractions (p <0.05), however the enhancement was significantly less in tissues from diabetic rats (p <0.05). The results suggest that STZ-induced diabetes impairs smooth muscle reactivity to nitric oxide in the rat anococcygeus muscle.Abbreviations STZ Streptozotocin - NOLA N^G-nitro-l-arginine - NANC nonadrenergic noncholinergic - ANOVA analysis of variance     

Analysis of the inhibitory effect of biguanides on glucose absorption: Inhibition of active sugar transport

Summary The effect of biguanides (phenethylbiguanide, butylbiguanide and dimethylbiguanide) on absorption of actively transported sugars was examined by incubating rings of hamster small intestinein vitro. Biguanides inhibited transport of D-glucose, D-galactose and 3-0-methyl-D-glucose but had no effect on the transport of D-fructose. Inhibition of D-xylose transport could only be demonstrated if concentrations far below halfmaximal saturation concentration (Km) were used (10^–5M), but not with concentrations approaching concentrations during a D-xylose tolerance test (18 mM). Formation of lactate by intestinal tissue was increased in presence of biguanides using D-glucose or D-fructose as substrates. The minimal inhibitory concentrations on transport of D-galactose were 10^–3M for phenethylbiguanide, 2×10^–3M for butylbiguanide and 6×10^–3M for dimethylbiguanide. The metabolite of phenethylbiguanide, 1-(4-hydroxy--phenethyl)-biguanide, did not affect glucose uptake but increased glucose metabolism to some extent. The demonstrated inhibition of active intestinal transportin vitro may be the mechanism for the decreased absorption of glucose observed by other authorsin vivo in man and animals after biguanides.Part of this work has been presented at the 5th annual meeting of the German Diabetes Society, Bad Godesberg, May 1970 and the 7th Congress of the International Diabetes Federation, Buenos Aires, August 1970.     

Sulphonylurea stimulates glucose uptake in rats through an ATP-sensitive K+ channel dependent mechanism

Summary We studied the effect of gliclazide, a second-generation sulphonylurea, on rat skeletal muscle glucose uptake using perfused hindquarter muscle preparations. Gliclazide at concentrations of 10 to 1000 g/ml increased (p<0.05) the basal glucose uptake. The effect of gliclazide on glucose uptake was immediate and dose-dependent, reaching a plateau at a concentration of 300 g/ml; the half-maximal effect was obtained between 25 and 50 g/ml. The glucose uptake stimulated by gliclazide (300–1000 g/ ml) did not differ from that achieved by 10^–9 mol/l insulin, and was lower (p<0.05) than that obtained with 10^–7 mol/l insulin. The combination of gliclazide (300 g/ml) and 10^–9 mol/l insulin produced an increase in glucose uptake (7.7±0.6 mol · g^–1 · h^–1, n=8, mean±SEM) which was higher (p<0.05) than that achieved with 10^–9 mol/l insulin (5.6±0.7 mol · g^–1 · h^–1, n=11) and not different from that obtained with 10^–7 mol/l insulin (9.8±1.0 mol · g^–1 · h^–1, n=11). Diazoxide (100 mol/l), an ATP-sensitive K⁺ channel opener, reversed the stimulatory effect of gliclazide (100 g/ml) on muscle glucose uptake from 3.1±0.4 to 0.5±0.2 mol · g^–1 · h^–1, (n=7, p<0.001). The addition of diazoxide prior to gliclazide into the perfusion medium blocked the gliclazide-induced glucose uptake by the hindquarter muscle preparations. In conclusion, gliclazide alone has an immediate stimulatory effect on glucose uptake by skeletal muscle and together with insulin has an additive effect on muscle glucose uptake. The effect of gliclazide on muscle glucose uptake seems to be due to the inhibition of ATP-sensitive K⁺ channels.Abbreviations NIDDM Non-insulin-dependent diabetes mellitus - GLUT glucose transporter     

Acute renal effects of angiotensin converting enzyme inhibition in microalbuminuric type 1 diabetic patients

The renal effects of intravenous injection of 40 mg enalapril were investigated in 16 normotensive microalbuminuric type 1 (insulin-dependent) diabetic patients. After enalapril the following changes were observed: fractional albumin clearance ( Alb) decreased from 9.9 (3.0–23.8) to 8.2 (2.0–18.3)×10^–6 (2P<0.01); filtration fraction (FF) decreased from 0.260 (0.225–0.312) to 0.253 (0.190–0.297) (2P<0.01); renal plasma flow (RPF) increased from 565 (411–690) to 623 (449–785) (2P<0.01); and glomerular filtration rate (GFR) remained stable at 149 (128–181) versus 150 (124–185) ml · min^–1 (NS). These values were unchanged after placebo (n=8), except for RFP which decreased from 606 (401–701) to 559 (381–677) ml · min^–1 (2P<0.05) and GFR which was reduced from 148 (111–173) to 138 (111–167) (2P<0.05). A reduction in mean blood pressure from 94 (87–103) to 89 (79–101) mmHg (2P<0.05) was found in the enalapril group and a minor reduction in the placebo group from 97 (83–106) to 96 (81–104) mmHg (2P<0.05) was also noted. The relative changes in systolic blood pressure in the enalapril group correlated with changes in Alb (Spearman'sr=0.66, 2P<0.02) and FF (r=0.53, 2P<0.05). Acute inhibition of angiotensin converting enzyme does not reduce the pathological hyperfiltration in these patients and a reduction in Alb and FF can not be dissociated from the reduction in blood pressure.     

Effect of protein kinase C activation and inhibition on rat hepatic stellate cell activation

Protein kinase C (PKC) may play a role in the intracellular signaling pathways responsible for transforming hepatic stellate cells into myofibroblasts. This study examined the effects of inhibitors and activators of PKC on hepatic stellate cell activation. Stellate cells isolated from normal rats were incubated with either 10^–5 M chelerythrine, 10^–7 M bisindolylmaleimide I hydrochloride (BIM), or 10^–6 M staurosporine (PKC inhibitors), or 10^–7 M phorbol myristate acetate (PMA) or 10^–6 M thymeleatoxin (PKC activators). Chelerythrine suppressed -smooth muscle actin expression and proliferation by 49% and 33%, respectively. BIM inhibited -smooth muscle actin expression by 60%, but had no significant effect on proliferation. Staurosporine decreased proliferation by 86% and completely prevented -smooth muscle actin expression. PKC activators had divergent effects on proliferation and -smooth muscle actin expression. PMA and thymeleatoxin caused a 2.8- to 3.2-fold increase in proliferation, while suppressing -smooth muscle actin expression by 50–70%. The demonstration that hepatic stellate cell activation can be suppressed by PKC inhibitors suggests a role for PKC in the regulation of hepatic stellate cell activation.