The sugar permeability test reflects disease activity in children and adolescents with inflammatory bowel disease

OBJECTIVES: To investigate the relationship of intestinal permeability in children and adolescents with inflammatory bowel disease (IBD) to disease activity, disease extent, and response to therapy. Study design: Patients with new and established diagnoses of IBD (12 Crohn's disease [CD] and 18 ulcerative colitis [UC]) were studied. Intestinal permeability was evaluated by measuring with high-performance liquid chromatography 5-hour urinary excretion ratio of lactulose/L-rhamnose (L/Rh). RESULTS: In 8 of 9 patients with active CD, the L/Rh ratio was higher than the reference range (0.006 to 0.074, n = 36). In inactive CD (n = 3) the L/Rh ratio was within the reference range. In 6 of 7 patients with active extensive UC, the L/Rh ratio was elevated. In inactive extensive UC (n = 6) the normal permeability ratio was shown. In both active CD and active extensive UC, the frequency of elevated intestinal permeability was significantly greater than values in both inactive forms. The permeability ratio was normal in 4 of 5 patients with active left-sided colitis. In 5 of 7 patients (3 CD, 4 UC), repeat permeability values entered the reference range after acute phase therapy. Two patients with persistently elevated intestinal permeability (1 CD, 1 UC) had a disease flare-up within 6 months. CONCLUSIONS: Intestinal permeability is a marker of disease activity in CD and extensive UC. Serial permeability test may be useful in monitoring disease activity.     

Thiolmethyltransferase activity in the human colonic mucosa: implications for ulcerative colitis

Ulcerative colitis is associated with a selective reduction of n-butyrate oxidation by the colonic epithelial cells although the reason for this has been unclear. Colonic epithelial cell n-butyrate oxidation can be inhibited in vitro by incubation with sulphide but the role of mucosal detoxification of sulphide in the metabolic welfare of the colonic mucosa has not been examined. This study aimed to assess the role mucosal detoxification of sulphide by thiolmethyltransferase (TMT)-mediated methylation may play in protecting the healthy colonic mucosa from the adverse effects of luminal sulphide. Colonic epithelial cell suspensions from healthy human proximal (n = 9) and distal colon (n = 10) were incubated in the presence of 14C-labelled n-butyrate (5 mmol/L) alone, butyrate plus sodium hydrogen sulphide (NaHS) (1.5 mmol/L), or butyrate plus NaHS plus S-adenosyl-methionine 1,4 butane disulphonate (SAMe) (5 mmol/L). Study end points were metabolic performance (14CO2 production) and mucosal TMT activity. Incubation with NaHS induced a significant inhibition of 14CO2 production compared with control incubations (P < 0.001) which was similar for proximal and distal colonic cell suspensions. S-adenosyl-methionine 1,4 butane disulphonate reversed this effect completely in proximal but not in distal cell incubations, suggesting a greater susceptibility of the distal colon to the sulphide effect. Although median whole mucosal TMT values did not differ between proximal and distal colonic mucosa, a non-normal distribution of distal TMT values was observed. However, neither the degree of sulphide inhibition of control 14CO2 production nor the degree to which SAMe reversed this inhibition correlated with whole mucosal TMT activity. The study concluded that regional variation exists in TMT activity in the human colon but whilst methylation appears to protect colonic epithelial cells against sulphide-induced inhibition of n-butyrate oxidation, this cannot be directly correlated with mucosal TMT activity.     

Measurement of contribution from intracellular cysteine to sulfate in phosphoadenosine phosphosulfate in rat ovarian granulosa cells

Synthesis of the large dermatan sulfate (DS) proteoglycan by rat ovarian granulosa cells was studied using metabolic radiolabel precursors in culture media with varying concentrations of environmental sulfate (20-800 microM) and cysteine (130 and 650 microM). Experiments using [3H]glucosamine and [35S]sulfate showed that the average size of the DS chains and the rate of DS proteoglycan synthesis were independent of the sulfate and cysteine concentrations in the medium. Experiments with [35S]cysteine were then used to determine the contribution that metabolic conversion of cysteine sulfur to sulfate makes to the 3'-phosphoadenosine 5'-phosphosulfate (PAPS) pool which provides the substrate for sulfoester formation in DS synthesis. When 35S in cysteine is metabolized into [35S]PAPS, the specific activity is reduced from that of the [35S]cysteine pool, by dilution with other sulfur sources such as extracellular sulfate, and this dilution factor directly reflects the contribution of cysteine to the PAPS pool. The decreases of 35S specific activity were measured under various sulfate-depleted and cysteine-supplemented conditions by comparing the specific activity of [35S]sulfate ester in the DS chains with that of [35S]cysteine residues in the core protein of the DS proteoglycan. The contribution of sulfur in cysteine to the intracellular PAPS pool was 0.03% in culture medium with normal sulfate (800 microM). Depleted environmental sulfate (20 microM) and increased cysteine supply (650 microM) only increased the sulfur contribution from cysteine to PAPS up to 0.74 and 1.5%, respectively, even though the DS chains were greatly undersulfated (55 and 82% of the control value). Thus, the source of sulfur in the intracellular pool of PAPS was mainly derived from environmental sulfate, and the contribution from cysteine was minimal in these cells.     

Detoxification of optically active bay- and fjord-region polycyclic aromatic hydrocarbon dihydrodiol epoxides by human glutathione transferase P1-1 expressed in Chinese hamster V79 cells

Dihydrodiol epoxides (DEs) are important carcinogenic metabolites of polycyclic aromatic hydrocarbons (PAHs). The metabolic formation of four stereoisomeric DEs (a pair of optically active diastereomers termed as syn- and anti-form) is possible. Glutathione tranferases (GSTs) have been demonstrated to catalyze the detoxification of DEs. Purified GSTs display remarkable differences in catalytic efficiencies towards bay- and fjord-region DEs along with a high degree of regio- and stereoselectivity. Here we determined to which extent heterologously expressed human GSTP1-1, a major GST isoform in lung, affects the mutagenicity of stereoisomeric bay-region DEs of benzo[a]pyrene in Chinese hamster V79 cells. To evaluate the influence of sterical crowding in the substrate on the activity of GSTP-1, the study was extended to the strongly mutagenic fjord-region (-)-anti-DEs of benzo[c]phenanthrene and dibenzo[a,l]pyrene. GSTP1-1,reduced preferentially the mutagenicity (studied at the hprt locus) of (+)-anti and (+)-syn-DEs of benzo[a]pyrene (by 66 and 67%) as compared with the corresponding (-)-anti- and (-)-syn-enantiomers (by 15 and 13%). These results are in line with previous studies on the enantioselectivity of purified GSTP1-1 towards the DE isomers of benzo[a]pyrene and benzo[c]phenanthrene showing that enantiomers with (R)-configuration at the benzylic oxiranyl carbon are better substrates than those with (S)-configuration. Interestingly, the (-)-anti-DEs of benzo[c]phenanthrene and dibenzo[a,l]pyrene were efficiently detoxified by GSTP-1-1 in the constructed cell line (reduction of mutagenicity by 66 and 64%). This study demonstrates that differences in the caalytic activity seen for purified GST towards individual mutagens do not necessarily reflect the detoxification of DEs by the same enzyme in a living cell and provides further evidence that specific human GSTs play a role in the detoxification of DEs of PAHs.     

Women and menopause: beliefs, attitudes, and behaviors. The North American Menopause Society 1997 Menopause Survey

It has been demonstrated that exposure to mercury or cadmium compounds causes alterations in the glutathione system in a model glial cell line, C6. Here we report that two organic tin compounds, triethyltin (TET) and trimethyltin (TMT), are also toxic to these cells with EC50 values for cell death of c. 0.02 microM and 0.8 microM respectively. Exposure for 24 h to either of these compounds at sub-toxic concentrations caused increases in the amount of reduced glutathione (GSH) per cell. Increases in glutathione-S-transferase enzyme activity were also demonstrated after TET or TMT exposure. This suggests that glutathione increases occur in glial cells after toxic insults below that required to cause cell death, possibly acting as a protective mechanism. To test whether GSH plays a role in organotin-induced cell death we manipulated GSH in the culture media or via intracellular GSH and looked at the effects on sensitivity to TET or TMT toxicity. Adding GSH to the culture media did not protect the cells. Depletion of intracellular GSH with buthionine-[S,R] sulphoximine did not alter cytotoxicity of TET or TMT. However, pre-treatment with (-)-2-oxo-4-thiazolidine carboxylic acid (OTC), which increases intracellular GSH levels, protected the cells against both compounds. The EC50 for TMT was increased from 0.77 to 1.8 microM, a 2.3-fold shift, whereas the EC50 for TET was increased > 20-fold, from 0.022 to 0.47 microM. One interpretation of these results is that GSH protects cells against the toxicity of organic tin compounds without reacting directly with them to any significant extent. Under conditions where GSH is depleted, additional protective mechanisms may be active.     

Relationships among lactate concentration, blood flow and histopathologic profiles in rat C6 glioma

Increased capacity for glycolytic metabolism is a well-known characteristic of neoplastic cells. Because lactic acid is the end product of glycolysis, in vivo MRS measurements of tumor lactate concentration ([lac]t) may provide valuable information about tumor metabolism, which will aid the development of therapies and the clinical diagnosis and treatment of tumors. In the present study, several hemodynamic and histologic parameters were evaluated with respect to their influence on [lac]t. Pronounced differences in [lac]t in two distinct populations of tumors suggested a putative perfusion threshold. Above this threshold, [lac]t was independent of hemodynamic and histologic factors including tumor blood flow (measured using MRS and the method of D2O washout), extent of necrosis and inflammatory cell infiltrate. Thus, for most tumors, [lac]t was not determined by any one single factor such as hypoxia, venous clearance, glucose supply, extent of necrosis or degree of inflammatory cell infiltrate. Rather, [lac]t may be equilibrated, at least in part, by an interplay of forces involving hemodynamics and substrate supply. In general, the data are consistent with the hypothesis that elevated lactate in most tumors is related to the high glycolytic activity of adequately perfused, viable neoplastic cells.     

Polymorphic expression of the glutathione S-transferase P1 gene and its susceptibility to Barrett's esophagus and esophageal carcinoma

Factors determining individual susceptibility to esophageal cancer or premalignant Barrett's epithelium are still largely unclear. An imbalance between phase I drug metabolism [e.g., cytochrome P450 (CYP)] and phase II detoxification [e.g., glutathione S-transferase (GST)] may contribute to the development of these diseases. Polymorphic variants in the CYP1A1 gene were described leading to increased levels of bioactive compounds, whereas polymorphisms in GST genes often resulted in impaired detoxification. We studied the frequencies of polymorphic variants in CYP1A1, GSTP1, GSTT1, and GSTM1 genes in 98 patients with Barrett's epithelium and 34 patients with esophageal cancer. The results were compared with those obtained from 247 healthy blood donors. DNA was extracted, and PCR-RFLP methods were used to detect genetic polymorphisms. Chi2 analysis, Spearman rank correlation, and Wilcoxon rank sum tests were used for statistical evaluation. Polymorphisms in CYP1A1, GSTM1, and GSTT1 occurred at an equal frequency in patients and controls. Occurrence of the polymorphic GSTP1b variant in the GSTP1 gene resulted in a significantly lower GST enzyme activity (P < 0.05), and GSTP1b was found significantly more often in patients with Barrett's epithelium (70%; P < 0.001) and patients with esophageal adenocarcinoma (76%; P = 0.005), as compared to healthy blood donors (41%). In conclusion, presence of the GSTP1b allele leads to lower GST enzyme activity levels and, consequently, impaired detoxification. This most important esophageal GST isoform may, therefore, contribute to the development of Barrett's epithelium and adenocarcinoma.     

An increase in fetal plasma cortisol but not dehydroepiandrosterone sulfate is followed by the onset of preterm labor in patients with preterm premature rupture of the membranes

OBJECTIVE: The role of steroid hormones in the control of human parturition has been a subject of debate. Activation of the fetal hypothalamic-pituitary-adrenal axis leading to an increase in plasma cortisol is followed by the onset of parturition in sheep. In contrast, androgens, specifically, dehydroepiandrosterone sulfate, have been implicated in the control of parturition in nonhuman primates. The purpose of this study was to determine the relationship between human fetal plasma cortisol and dehydroepiandrosterone sulfate and the onset of preterm labor in patients with preterm premature rupture of the membranes. STUDY DESIGN: Fetal blood sampling was performed in 51 patients with preterm premature rupture of membranes who were not in labor on admission. Amniotic fluid was cultured for aerobic and anaerobic bacteria and mycoplasmas. Corticosteroids had not been administered before fetal blood sampling. Cortisol and dehydroepiandrosterone sulfate were measured with sensitive and specific immunoassays. Analysis was conducted with nonparametric statistics and survival analysis. RESULTS: (1) Patients who went into spontaneous labor and delivered within 7 days of cordocentesis had a significantly higher median level of fetal plasma cortisol but not of dehydroepiandrosterone sulfate than those delivered after 7 days (for fetal plasma cortisol: median 8.35 [4.7 to 12.4] micrograms/dL vs median 4.75 [3.0 to 10.4] micrograms/dL, P <.0001; for fetal plasma dehydroepiandrosterone sulfate: median 154.4 [8.6 to 333.8] micrograms/dL vs median 194.6 [96.7 to 402.5] micrograms/dL, P =.09). (2) The cordocentesis-to-delivery interval was significantly shorter in patients with a fetal plasma cortisol value of >/=7 micrograms/dL (derived by receiver-operating characteristic curve analysis) than in those with fetal cortisol <7 micrograms/dL (median 49 [4 to 1849] hours vs median 325 [11 to 2590] hours, P <.001). (3) Fetal plasma cortisol, but not maternal cortisol, was an independent predictor of the duration of pregnancy after we adjusted for gestational age and the results of amniotic fluid culture (hazards ratio 2.9, P <.05). (4) There was a significant correlation between fetal plasma cortisol and fetal plasma interleukin-6 (r = 0.3, P <.05). (5) A strong relationship was found between the fetal plasma cortisol/dehydroepiandrosterone sulfate ratio and the interval to delivery (P <.005). CONCLUSION: An elevation in fetal plasma cortisol but not dehydroepiandrosterone sulfate was followed by the onset of spontaneous preterm labor in patients with preterm premature rupture of the membranes.     

Activity of the adenylyl cyclase in lymphocytes of male alcoholic patients is state dependent

A decreased basal and/or stimulated activity of the G-protein/adenylyl cyclase (AC) system in peripheral blood cells has been proposed to represent a trait marker for alcoholism. However, AC activity may underlie state-dependent changes, which may impair a proper interpretation of AC activity measurements. Our study examined systematically the AC activity in peripheral lymphocytes of 73 male alcohol-dependent patients (according to DSM-IV criteria) at three different time points of measurement during the clinical course of detoxification (day 0 = at admission, while still ethanol-affected; day 2 = at the presumed peak of withdrawal symptoms; day E = after detoxification). Basal and stimulated (with GTPgammaS and forskolin) AC activities were measured. AC activities were compared to those of a control group of 44 healthy male age-matched volunteers. As our main finding, we detected a significant decrease in AC activity from day 0 to day 2 (during withdrawal), with lowered AC activities in a vast majority of patients. This effect resolved after detoxification, as AC activities showed a significant increase from day 2 to E. No significant difference was detected between day 0 and E in AC activities of the patients. Compared with controls, AC activities in patients were significantly lower at day 2, but not at day 0 and E. Taken together, our results indicate rapid and marked state-dependent changes of AC activities in alcohol-dependent patients during the course of detoxification. For an adequate interpretation of AC activities in alcoholic patients, their clinical status must be taken into consideration.     

Thrombophilia and inflammatory bowel disease: does factor V mutation have a role?

BACKGROUND: An increased tendency for thromboembolism is a well known problem of inflammatory bowel disease (IBD). Microvascular thrombosis has also been claimed as a pathogenic factor in IBD. Recently a point mutation in the gene coding factor V (FV Leiden) has been identified in various thromboembolic diseases, but the role in IBD is unknown. OBJECTIVE: To determine the frequency of FV Leiden in IBD patients and compare with a group of controls. METHODS: Sixty-three IBD patients [43 ulcerative colitis (UC) patients and 20 Crohn's disease (CD) patients] and 36 healthy controls were included in the study. Only one of the UC patients had a history of cerebral thromboembolism. The extracted DNA from frozen blood was subjected to polymerase chain reaction for the amplification of FV gene. The amplicons were hybridized both with the mutant and wild-type probes to detect FV mutation. Readings of optical density above 0.3 were considered as positive results. According to the patterns of ELISA, heterozygosity and homozygosity for normal and mutant alleles were determined. RESULTS: Eight (18%) of UC patients were heterozygous normal and one (2%) patient had homozygous mutation. Eight (45%) of the 20 CD patients had a heterozygous pattern and one (5%) had a homozygous pattern. In the control group four (11%) subjects showed a heterozygous genotype. FV Leiden was found to be statistically more frequent in CD patients (P < 0.005) (odds ratio 6.5, 95% confidence interval 1.3-18.), but not in the UC patients as compared with controls (P> 0.05). There was no significant correlation between FV Leiden presence and disease activity, gender or disease duration for both UC and CD. CONCLUSION: The results suggest that FV Leiden is more frequent in CD patients, but not in the UC patients as compared with controls. The high rate of factor V mutation in our CD patients suggests the need for further studies to confirm a relationship between this mutation and aetiology of the disease.     

Carbonic anhydrase I reduction in colonic mucosa of patients with active ulcerative colitis

Ulcerative colitis (UC) is associated with low intracolonic pH and unbalanced transmucosal ionic exchanges. Along the gastrointestinal tract carbonic anhydrase isoenzyme I (CA-I) is specifically expressed in colon epithelium and is involved in mucosal control of ion, fluid, and acid-base balance. Since altered CA-I expression may play some role in UC, CA-I was measured at the mRNA and protein level and carbonic anhydrase (CA) enzyme activity was determined in colon biopsies of 14 UC patients (6 remission, 4 mild, 4 moderate UC) and of 12 healthy subjects. Patients with mild or moderate UC showed a significant reduction of CA-I mRNA and protein and of total CA activity in the inflamed mucosa compared to controls. Patients with UC in remission showed a pattern of CA-I expression and CA activity similar to controls. This is the first report showing a reduction in the expression of CA-I in active UC.     

Interfacial effects of surface-active agents under zinc pressure leach conditions

Liquid sulfur-zinc sulfate solution interfacial tensions and liquid sulfur-zinc sulfate solution-zinc sulfide (marmatite) contact angles were measured in the absence and presence of surface-active agents. Interfacial tensions measured varied between 54 ± 1 mN/m in the surfactant-free system and 20 ± 1 mN/m in the presence of a surfactant. The liquid sulfur-zinc sulfide mineral-zinc sulfate solution contact angle varies between 80 ± 5 deg, in the absence of any surfactant, and 148 ± 5 deg, depending on the surfactant used. The surface-active agents were used as dispersants for sulfur in bench-scale zinc pressure-leaching experiments. The observed extent of zinc extraction depends on the surfactant and varies from 40 to 96 pct.     

Noninvasive measurement of tissue magnesium and correlation with cardiac levels

BACKGROUND: Intracellular magnesium ([Mg]i) plays an important role in the regulation of myocardial metabolism, contractility, and the maintenance of transsarcolemmal and intracellular ionic gradients. An understanding of the role of magnesium in the clinical setting, however, is hampered by the lack of an assay of intracellular tissue magnesium levels. METHODS AND RESULTS: We used energy-dispersive x-ray analysis to measure [Mg]i in sublingual epithelial cells and to correlate the level with those in atrial biopsy specimens from the same patients during cardiopulmonary bypass. Levels were also measured in acute myocardial infarction (AMI) patients before and after intravenous magnesium sulfate administration and compared with those from intensive care unit (ICU) patients and healthy individuals. A strong correlation between sublingual epithelial cell (mean, 32.1 +/- 0.3 mEq/L) and atrial tissue (mean, 32.1 +/- 0.3 mEq/L) [Mg]i was present in 18 cardiac surgery patients (r = .68, P < .002). Epithelial and atrial [Mg]i levels were lower than in healthy individuals (33.7 +/- 0.5 mEq/L, P < .01) studied at that time and correlated poorly with serum magnesium. Mean [Mg]i in 22 AMI patients was 30.7 +/- 0.4 mEq/L, which was significantly lower than in 21 ICU patients and 15 healthy individuals (35.0 +/- 0.5 mEq/L and 34.5 +/- 0.7 mEq/L, respectively, P < .001). Intravenous magnesium sulfate was administered to most of the AMI patients (mean dose, 36 +/- 6 mmol). [Mg]i rose significantly in the AMI patients over the first 24 hours, and the magnitude of the increase was greater in those who received higher doses of intravenous magnesium sulfate. CONCLUSIONS: Sublingual epithelial cell [Mg]i correlates well with atrial [Mg]i but not with serum magnesium. [Mg]i levels are low in patients undergoing cardiac surgery and those with AMI. Intravenous magnesium sulfate corrects low [Mg]i levels in AMI patients. Energy-dispersive x-ray analysis determination of sublingual cell [Mg]i may expedite the investigation of the role of magnesium deficiency in heart disease.     

Feasibility of a rapid method to evaluate platelet survival time

The purpose of this study was to evaluate the feasibility of a shorter method of performing platelet kinetic studies with respect to the conventional 8-9-day approach. METHODS: We studied 41 patients (28 women, 13 men; mean age 52 yr) with primary idiopathic thombocytopenic purpura (ITP) (n = 20), secondary ITP (n = 9), HCV associated thrombocytopenia (n = 9), splenectomy (n = 1) and hairy-cell leukemia (n = 1). The patients were in a steady-state of platelet turnover. Initial platelet counts ranged from 19 to 302 x 10(9)/liter (mean value = 83). Platelet survival times (PST) were measured from the blood radioactivity disappearance curve of 111In-oxine-labeled autologus platelets following the recommendations of the International Committee for Standardization in Haematology: blood samples were taken at 30 min and 2 and 4 hr and thereafter daily for 7 days. PST was calculated by the weighted mean method and ranged from 18 to 219 hr (mean value = 98). PST was also calculated using only the data collected at 2, 48 and 96 hr. If the radioactivity in the blood at 96 hr exceeded 10% of the 2-hr value, the additional point at 168 hr was used. RESULTS: By using this reduced dataset, we obtained a correlation of r = 0.97 with the PST obtained from the whole dataset. In 24 patients, the difference was between +/- 10 hr and exceeded 1 day in only 4. CONCLUSION: About 94% of the data may be recovered with only three or four blood samples and the duration may be shortened to 4 days in a significant proportion of patients (48% of ITP patients). This approach offers the advantages of increased patient throughput, compliance and reduced examination costs.     

Heparan sulfate upregulates platelet-derived growth factor receptors on human lung fibroblasts

Heparan sulfate is a molecule that possesses a large structural variability and which has been shown to inhibit the proliferation of fibroblasts in vitro. The aim of this study was to determine whether the anti-proliferative effects of heparan sulfate were exerted by regulation of the activity of the platelet-derived growth factor and/or of the platelet-derived growth factor receptors. Both l-iduronate-rich, anti-proliferative and the l-iduronate-poor, non-anti-proliferative heparan sulfate species, were incubated with confluent human embryonic lung fibroblasts for 24 h. The mRNA levels for PDGF-AA, PDGF-BB, and their receptors were measured. Binding studies were performed with [125I]-PDGF-BB and [125I]-EGF for 2 h at 4 degreesC in cultures preincubated with both types of heparan sulfate for 24 h. In separate experiments, cultures were incubated together with heparan sulfate and [125I]-PDGF-BB for 2 h at 4 degreesC. Increases of two- to threefold in the mRNA levels for both the alpha- and the beta-receptors of PDGF was obtained after treatment with both types of heparan sulfate, whereas the mRNA levels of both the PDGF-AA and the PDGF-BB were essentially unaffected. A sixfold increase in binding was only noted for [125I]-PDGF-BB in cultures pre-treated with the anti-proliferative heparan sulfate for 24 h, whereas no effect was noted with use of the non-anti-proliferative heparan sulfate. Incubating the [125I]-PDGF-BB and the anti-proliferative heparan sulfate together for 2 h resulted in a smaller, threefold increase in binding. This indicates that the anti-proliferative heparan sulfate both stabilizes and increases expression of the PDGF receptors. To investigate whether the increased number of PDGF receptors could affect cell activity, cells were preincubated with anti-proliferative heparan sulfate and then treated with PDGF-BB. This resulted in an increase in mitogenicity compared to cells treated only with PDGF-BB. Neither an increase in binding for [125I-EGF] nor an increase in the mitogenic response of EGF could be observed in cultures pre-treated with the anti-proliferative heparan sulfate. The results indicate that the extracellular matrix itself may regulate important biological phenomena such as cell proliferation and matrix production through affecting the expression of receptors of PDGF, which initiate both stimulatory and inhibitory signals.     

N-Acetylgalactosamine 4,6-bissulfate in rat urine. II. Occurrence in rat cartilage and blood

The intraperitoneal injection of inorganic [35S]sulfate to rat was followed by the rapid appearance in urine of a labeled compound which behaved as N-acetylgalactosamine 4,6-bissulfate on paper chromatography and paper electrophoresis and when treated with two sulfatases with a high degree of specificity toward the sulfate bonds at positions 4 and 6, respectively. Enzymatically-prepared N-acetylgalactosamine 4,6-[6-35S]bissulfate was injected intravenously into rats. Of the injected dose, 90% was excreted unchanged in the urine during the subsequent 12 h, suggesting that the urinary N-acetylgalactosamine 4,6-bissulfate may derive from blood as renal filtrate. Examination of the rats injected with inorganic [35S]sulfate revealed the presence of labeled N-acetylgalactosamine 4,6-bissulfate at significant levels in the blood and cartilage, but at much lower levels in the liver. The cartilage component was highest in its rate of 35S uptake, suggesting that the blood component may derive at least in some part from the cartilage. Exposure of surviving cartilage slices to inorganic [35S]sulfate, followed by extraction of the slices with hot 50% ethanol yielded a number of radioactive compounds, of which three were characterized as UDP-N-acetylgalactosamine-4,6-[35S]bissulfate, N-acetylgalactosamine-1-phosphate 4,6-[35S]bissulfate and N-acetylgalactosamine 4,6-[35S]bissulfate. By subjecting the prelabeled tissue to chase incubation, it was possible to show that the UDP-N-acetylgalactosamine-4,6-bissulfate in the tissue disappeared with an approximate half-life of 10 min with a concomitant appearance in the medium of N-acetylgalactosamine 4,6-bissulfate and its 1-phosphate ester. These results suggest the occurrence in cartilage of an enzymatic system which is responsible for rapid turnover of UDP-N-acetylgalactosamine-4,6-bissulfate and possibly required for the rapid secretion of N-acetylgalactosamine 4,6-bissulfate into extracellular field.     

Ulcerative colitis: patterns of involvement in colorectal biopsies and changes with time

Chronic inflammation, both endoscopic and histologic, in a contiguous and symmetric distribution is said to be important in distinguishing ulcerative colitis (UC) from Crohn's disease. Little is known whether this rule holds during the course of the disease and whether endoscopic/histologic correlation persists. In this study, we analyzed histologic patterns of UC in sequential sets of biopsy specimens to assess whether endoscopic and histologic findings correlate with time and treatment and to see whether distribution changes. Two hundred seventeen sets of colorectal biopsy specimens from 797 sites from 41 patients with clinical UC were studied and correlated with endoscopic findings. Each biopsy specimen was classified as definite or suspicious for chronic colitis or normal. Two histologic patterns of disease were identified: (1) diffuse, when all areas in all pieces from a biopsy segment had clear-cut colitis and (2) nondiffuse, when not all pieces were involved or single pieces had disease and normal mucosa both. Of 41 patients, the maximal extent of histologic disease was pancolitis in 30; 25 had less extensive disease at some point in the course. The maximal extent was left-sided in eight patients, seven of whom had less extent at some point. Of the three patients in whom the maximal extent was proctosigmoiditis, in one the inflammation disappeared. Seventy percent of the biopsy sites had diffuse patterns and 30% had nondiffuse. Histologic and endoscopic disease reverted to normal in 22 and 24 of 41 patients, respectively. Endoscopic and histologic findings were similar in 65% of the biopsy sites. Our results indicate that in long-standing UC (1) histologic disease may revert to normal mucosa, (2) because endoscopy alone may be insufficient to identify the mucosa as normal, biopsies should also be performed on the endoscopically normal mucosa, (3) the full extent of UC often is not established by a single set of biopsies, and (4) nondiffuse chronic inflammation and rectal sparing occurs in UC and are not necessarily markers of Crohn's disease.     

Expression of E-selectin, sialyl Lewis X, and macrophage inflammatory protein-1alpha by colonic epithelial cells in ulcerative colitis

The pathogenic significance of cell adhesion molecules (CAMs) in ulcerative colitis (UC) is largely unknown. Colonic expression of E-selectin, sialyl Lewis X (sLe(x)), and macrophage inflammatory protein-1x (MIP-1alpha) as well as serum concentrations of E-selectin and MIP-1alpha in UC were studied. Thirty patients with UC, 10 patients with irritable bowel syndrome, and 10 healthy subjects were included. Colonic biopsies were stained immunohistochemically, and blood concentrations were measured with an ELISA technique. Soluble E-selectin did not correlate with diagnosis or disease activity. MIP-1alpha was below the detection limit. Epithelial cells expressed all three molecules, both on surface membranes and intracellularly. sLe(x) staining was weaker (P = 0.0002) and MIP-1alpha staining stronger (P = 0.014) in UC patients than in controls. Leukocyte MIP-alpha staining correlated with diagnosis (P = 0.021), sLe(x) staining (P = 0.023), and colonoscopy (P = 0.018). It is shown that E-selectin, sLe(x), and MIP-1alpha are synthesized and expressed by epithelial cells, indicating that CAMs are not only involved in leukocyte extravasation and migration, but also in the interaction between leukocytes and colonic epithelium. This knowledge might contribute to the development of improved treatments in UC.     

Bioactivation of benzylic and allylic alcohols via sulfo-conjugation

Although sulfo-conjugation, in general, has been regarded as a detoxification process in the xenobiotic metabolism, there is a substantial body of data supporting that the same reaction can also lead to activation of certain types of chemical carcinogens and mutagens. Examples include some aromatic amines and amides, alkenylbenzenes, methyl-substituted polyaromatic hydrocarbons, nitrotoluenes and nitrosamines. The N- or O-hydroxy derivatives of these compounds undergo sulfonation to form extremely reactive sulfuric acid esters that can play a role as ultimate carcinogenic/mutagenic metabolites. Previous studies from several laboratories have shown that hydroxymethyl polyarenes, such as hydroxymethylbenz[a]anthracenes, 6-hydroxymethylbenzo[a]pyrene, and 1-hydroxymethylpyrene, are activated to reactive benzylic sulfuric acid esters, preferentially by rat hepatic hydroxysteroid sulfotransferase. Some aromatic hydrocarbons bearing the secondary benzylic hydroxy functionality can also yield electrophilic sulfate esters in the presence of hepatic sulfotransferase activity. Thus, benzylic mono- and dihydroxy derivatives of cyclopenta[cd]pyrene form mutagenic and DNA binding species when incubated with rat liver cytosol and the sulfo-group donor, 3'-phosphoadenosine-5'-phosphosulfate. 1-Hydroxy-3-methylcholanthrene that also possesses the cyclopenta-fused ring system appears to be metabolically activated through sulfo-conjugation. Likewise, benzo[a]pyrene tetraol might be activated through sulfuric acid esterification at one of two benzylic hydroxyl groups. Methylene-bridged polyarenols represent another potential group of cyclic secondary benzylic alcohols that can be activated by sulfotransferases. Certain non-polycyclic aromatic type benzylic alcohols have also been proposed to undergo sulfotransferase-mediated activation. Besides benzylic sulfonation, sulfuric acid esterification of certain allylic alcohols can produce reactive species.     

Physiological significance of plasma sulfoconjugated dopamine in patients with hypertension--clinical and experimental studies

Sulfoconjugated catecholamines, especially dopamine sulfate, have recently attracted much attention because of the possibility of their conversion to active free dopamine by tissue arylsulfatase. In the present study, we have measured the plasma levels of free and sulfoconjugated dopamine in patients with hypertension and have investigated the physiological significance of sulfoconjugation. Results showed that the plasma level of dopamine sulfate in patients with essential hypertension was higher than the level in control subjects, and was highest in patients with renal hypertension. However, the plasma level of free dopamine showed no significant difference between patients with hypertension and normal subjects. Moreover, after normalization of blood pressure in hypertensive patients with medication, the plasma levels of conjugated dopamine decreased to almost the control value. In the experimental study, dopamine sulfate inhibited angiotensin II-induced aldosterone release from bovine adrenal cortical cells to a similar extent as produced by free dopamine. From these results, we have concluded that plasma sulfoconjugated dopamine may regulate free dopamine in the plasma of patients with hypertension, and it may have some physiological effects on blood pressure regulation.