Antioxidant properties of colchicine in acute carbon tetrachloride induced rat liver injury and its role in the resolution of established cirrhosis.

Antioxidant and antifibrotic properties of colchicine were investigated in the carbon tetrachloride (CCl(4)) rat model. (1) The protective effect of colchicine pretreatment on CCl(4) induced oxidant stress was examined in rats subsequently receiving a single lethal dose of CCl(4). Urinary 8-isoprostane, kidney and liver malondialdehyde and kidney glutathione levels increased following CCl(4) treatment, but only the rise in kidney malondialdehyde was significantly inhibited by colchicine pretreatment. Serum total antioxidant levels were significantly higher in the colchicine pretreatment group. (2) The long term effects of colchicine treatment on CCl(4) induced liver damage were investigated using liver histology and biochemical markers (hydroxyproline and type III procollagen peptide). Co-administration of colchicine with sub-lethal doses of CCl(4) over 10 weeks did not prevent progression to cirrhosis. However, rats made cirrhotic with repeated CCl(4) challenge and subsequently treated with colchicine for 12 months, all showed histological regression of cirrhosis. (3) The antioxidant effect of colchicine in vitro was evident only at very high concentrations compared to other plasma antioxidants. In summary, colchicine has only weak antioxidant properties, but does afford some protection against oxidative stress; more importantly, long term treatment with this drug may be of value in producing regression of established cirrhosis.

Longitudinal variation in the activities of mucosal enzymes in the small intestine of suckling rats.

The extent of positional variation in mucosal enzyme activity along the small intestine was investigated in 14-day-old suckling rats. Samples were taken from ten equally spaced sites along the intestine in 11 rat pups and the activities of the enzymes alkaline phosphatase, neutral aminopeptidase, gamma-glutamyl transferase, lactase and sucrase were measured. All the enzymes except sucrase were subject to considerable positional variation. Alkaline phosphatase and aminopeptidase activities were distributed throughout the intestine, with a broad maximum in the distal intestine. Lactase was also broadly distributed but with greatest activity in the mid intestine. gamma-glutamyl transferase exhibited a novel profile, with a very high proportion of the total activity (78%) present in the distal intestine. Sucrase was essentially absent throughout the intestine.

Effects of urogastrone-epidermal growth factor and age at administration on five enzymes in the small intestine of suckling rats.

Suckling rats were given urogastrone-epidermal growth factor (EGF: 1,000 micrograms/kg body weight) or vehicle by gavage at one of three stages of development: 8 to 10, 11 to 13 or 14 to 16 days of age. Intubation was carried out at 8-hourly intervals over these periods. Fourteen to 16 h after the last intubation the rats were killed; that is, at 11, 14 and 17 days respectively. Samples of proximal and distal small intestine (SI) were taken for enzyme analysis. Five enzymes were assayed; sucrase, lactase, gamma-glutamyl transferase, alkaline phosphatase and neutral amino-peptidase, and their activities expressed per g protein. Treatment with EGF had no effect on body weight or on the length of the small intestine at any age. The nature of the effects on enzyme activities depended on the specific enzyme concerned, the site within the small intestine and the timing of the treatment. Lactase was increased by EGF at both sites only on day 14, whereas gamma-glutamyl transferase was increased in proximal samples at 11 and 14 days, and in distal samples at 17 days. Nor was the outcome always to increase activity. On day 11 alkaline phosphatase was increased in proximal SI, but decreased in distal SI; and so too was aminopeptidase N decreased in distal SI at 11 days. Sucrase showed no response at all. The pattern is complex. Certainly it does not indicate accelerated functional maturation.

Simultaneous assessment of intestinal permeability and lactose tolerance with orally administered raffinose, lactose and L-arabinose.

1. In order to develop an improved differential sugar absorption test for simultaneously assessing intestinal permeability and lactose intolerance, methods were established for determining raffinose, lactose and L-arabinose in human urine. Using NAD(P)H-coupled enzymatic assays and fluorimetry, each sugar was measurable over a concentration range of approximately 3-300 mumol/l in diluted urine specimens. 2. After an overnight fast, 40 normal volunteers drank an iso-osmotic solution containing raffinose, lactose and L-arabinose. The median 5 h urinary sugar excretion was 0.26% of the ingested raffinose, 0.05% of lactose and 17.5% of L-arabinose. 3. In 143 patients with gastrointestinal disease, excretion of both ingested raffinose and lactose was significantly increased in coeliac disease in relapse or in partial remission and in Crohn's disease, but not in the irritable bowel syndrome, coeliac disease in remission or ulcerative colitis. Excretion of lactose, but not raffinose, was increased in patients with mucosal lactase deficiency, whereas excretion of L-arabinose was reduced in all disease groups except ulcerative colitis. 4. Discrimination between diseases was poor when based on individual sugar recoveries, but improved dramatically when excretion was expressed relative to that of L-arabinose. The raffinose/L-arabinose excretion ratio, an index of intestinal permeability, was greater than 0.08 in 15/15 untreated coeliac patients but less than 0.06 in all normal subjects and in 9/9 lactase-deficient patients, 15/16 recovered coeliac patients, 5/6 patients with ulcerative colitis, 13/16 patients with Crohn's disease and 61/62 patients with irritable bowel syndrome.

Changes in jejunal permeability and passive permeation of sugars in intestinal biopsies in coeliac disease and Crohn's disease.

1. The relative effects of changes in mucosal surface area and mucosal permeability on the passive uptakes of mannitol and raffinose have been studied in vitro using jejunal biopsies from 48 controls, 32 patients with coeliac disease and 11 patients with Crohn's disease. Total mucosal permeation was corrected for surface area measured morphometrically to provide an index of mucosal permeability. 2. In untreated coeliac disease, permeation of mannitol was reduced by 35% (P = 0.006) and that of raffinose was increased by 66% (P = 0.0095) compared with controls, whereas mucosal permeability to mannitol was increased twofold (P = 0.009) and to raffinose fivefold (P = 0.0001). Mucosal permeability was similar for each sugar. 3. In treated coeliac disease, permeation and permeability for mannitol were normal, but remained elevated for raffinose by 23% (P = 0.036) and 41% (P = 0.024), respectively. 4. In Crohn's disease, permeation of mannitol was reduced by 21%, but that of raffinose and mucosal permeability to both sugars were normal. 5. These findings suggest that surface area is quantitatively more important than mucosal permeability in determining the total permeation of mannitol, while the converse is true for raffinose. The findings are compatible with paracellular uptake of raffinose, but with both paracellular and transcellular uptake of mannitol. Both pathways are affected in coeliac disease, whereas only transcellular uptake is affected in Crohn's disease.

The kinetics of monosaccharide absorption by human jejunal biopsies: evidence for active and passive processes.

The kinetics of initial rates of uptake of glucose, galactose, arabinose and mannitol have been measured in jejunal biopsies from normal subjects in order to investigate the existence of multiple uptake systems. Glucose kinetics fitted best a model of a saturable uptake system (app Kt = 2.06 +/- 0.33 mM, app Jmax = 93.85 +/- 1.19 nmol/10 min/mg dry weight), together with a linear uptake indistinguishable from the passive uptake of arabinose and mannitol (app Kd = 0.80 +/- 0.05 nmol/10 min/mg dry weight/mM for glucose, 0.75 +/- 0.03 for arabinose, and 0.83 +/- 0.03 for mannitol). The saturable uptake, but not the linear uptake, was inhibited by phlorizin and by the absence of sodium. Cytochalasin B and phloretin had no effect on overall uptake. Galactose kinetics in the absence of inhibitors fitted best a model of a single saturable uptake system (app Kt = 11.05 +/- 0.12, app Jmax = 201.9 +/- 1.13) with no evidence of linear uptake. In the presence of phlorizin, or in the absence of sodium, uptake was predominantly linear with app Kds of 0.84 +/- 0.03 and 0.79 +/- 0.01, not significantly different from the linear component of glucose uptake. We conclude that hexose uptake in human jejunum in vitro occurs by both active and passive routes, and that the active uptake of galactose appears to be inhibited at high galactose concentration.

Lactose digestion by human jejunal biopsies: the relationship between hydrolysis and absorption.

The relationship between lactose hydrolysis and absorption of released glucose was investigated by determining the kinetics of lactose digestion by jejunal biopsies incubated in vitro. Lactase activity in intact biopsies correlated with conventional assay of tissue homogenates (r = 0.85, p less than 0.001), and glucose uptake from 28 mM lactose was directly proportional to lactase activity (r = 0.95, p less than 0.001) in 21 subjects with normal lactase levels, six with hypolactasia (primary or secondary to coeliac disease) and two with lactose intolerance but normal lactase activity. Kinetic analysis at 0.56-56 mM lactose in five normal subjects showed saturable kinetics for hydrolysis (app Km = 33.9 +/- 2.2 mM; app Vmax = 26.5 +/- 1.1 nmol/min/mg dry weight) but glucose uptake could be fitted to a model either of saturable uptake (app Kt = 47.2 +/- 0.3 mM; app Jmax = 14.1 +/- 0.2 nmol/min/mg) or saturable uptake plus a linear component (app Kt = 21.3 +/- 1.15; app Jmax = 4.59 +/- 0.12; app Kd = 0.093 +/- 0.010 nmol/min/mg/mM). The proportion of glucose taken into the tissue did not significantly exceed 50% of the total released at any lactose concentration suggesting the lack of an efficient capture mechanism for the released glucose. The results suggest that lactose hydrolysis is the rate limiting step in the overall absorption of glucose from lactose in vitro, and that the relationship between hydrolysis and absorption is the same in normal subjects and in hypolactasic subjects.