Effect of aspirin on prostaglandin E2 and leukotriene B4 production in human colonic mucosa from cancer patients.

Results from epidemiological studies indicate that chronic administration of aspirin reduces the incidence of colon cancer. The mechanism that accounts for this reduction is not known, but it may be related to the decreased production of prostanoids that results from aspirin inhibition of cyclooxygenase. However, it is not known whether aspirin has a local effect on prostanoid production in the colonic mucosa and whether this effect is dose dependent. In this study, we determined the effect of oral administration of aspirin on the production of the prostanoid prostaglandin E2 (PGE2) in the intact human colonic mucosa. Inhibition of cyclooxygenase could result in an increased availability of arachidonic acid and a corresponding increase in production of other eicosanoids. To determine whether such an effect occurs, we also quantitated the concentration of leukotriene B4 (LTB4) in colonic mucosal samples. Mucosal samples were obtained during sigmoidoscopy from the colons of 17 subjects with a history of colonic cancer prior to and following 60 days of self-administration of 325 mg aspirin/day and again 60 days after administration of 650 mg aspirin/day. PGE2 and LTB4 concentrations were determined by enzyme immunoassay for tissue samples that were flash frozen after removal from the biopsy forceps and also in medium that was collected from tissue samples that were incubated for 4 h following removal from the subject. PGE2 concentrations were decreased significantly in samples collected after 60 days of consumption of 325 mg aspirin. An additional 60 days of consuming 650 mg aspirin/day did not result in a further significant decrease relative to that attained after consumption of 325 mg/day. Similar results were obtained using colonic explants, and the addition of aspirin to medium further reduced PGE2 production. LTB4 in tissue and medium was not significantly different in pre-versus post-aspirin samples, with the exception of an increased concentration in medium samples collected after consumption of 650 mg/day relative to pre-aspirin samples. The results indicate that aspirin affects eicosanoid production in the colonic mucosa of humans, but the effect is most likely restricted to products of the cyclooxygenase-dependent pathway. It appears that 325 mg of aspirin is sufficient to affect PGE2 production and that increasing the dosage to 650 mg daily provides an additional decrease in PGE2 synthesis. However, the higher dosage was associated with a considerable increase in complaints of gastric discomfort. Additional study is needed to establish whether doses less than 325 mg also provide a significant decrease in PGE2 production.

Photoaffinity labelling of the phenylalkylamine receptor of the skeletal muscle transverse-tubule calcium channel.

The tritiated arylazido phenylalkylamine (-)-5-[(3-azidophenethyl)[N-methyl-3H]methylamino]-2-(3,4, 5-trimethoxyphenyl)-2-isopropylvaleronitrile was synthesized and used to photoaffinity label the phenylalkylamine receptor of the membrane-bound and purified calcium channel from guinea-pig skeletal muscle transverse-tubule membranes. The photoaffinity ligand binds reversibly to partially purified membranes with a Kd of 2.0 +/- 0.5 nM and a Bmax of 17.0 +/- 0.9 pmol/mg protein. Binding is stereospecifically regulated by all three classes of organic calcium channel drugs. A 155 kDa band was specifically photolabelled in transverse-tubule particulate and purified calcium channel preparations after ultraviolet irradiation. Additional minor labelled polypeptides (92, 60 and 33 kDa) were only observed in membranes. The heterogeneous 155 kDa region of the purified channel was resolved into two distinct silver-stained polypeptides after reduction (i.e. 155 and 135 kDa). Only the 155 kDa polypeptide carries the photoaffinity label and it is concluded that the 135 kDa polypeptide (which migrates as a 165 kDa band under alkylating conditions) is not a high-affinity drug receptor carrying subunit of the skeletal muscle transverse-tubule L-type calcium channel.

Cytotoxic effect of beta-carotene in vitro is dependent on serum concentration and source.

In a previous study we reported that beta-carotene solubilized in tetrahydrofuran (THF) is toxic for human colonic tumor cells in vitro using media containing 10% fetal calf serum (FCS). Cytotoxicity was evident using beta-carotene concentrations that are achieved in human serum as a result of supplementation with 30 mg beta-carotene/day. In an attempt to determine the mechanism for this toxicity we investigated the effect of beta-carotene when present in human serum as a result of dietary supplementation. This effect was compared to that observed for cells incubated in THF-solubilized beta-carotene. The results indicate that human serum from subjects with a high concentration of beta-carotene is not cytotoxic. Subsequent analysis revealed that in contrast to the results using FCS, THF-solubilized beta-carotene is not cytotoxic in the presence of human serum. In addition, the effect observed with FCS is blunted with increasing FCS concentration from > or = 90% cytotoxicity using 10% FCS to 36% using 100% FCS. The difference in results obtained using FCS and human serum may be due to a serum component that is relatively lacking in FCS as compared to human serum.

Effects of pregnancy on the cytochrome P-450 system in mice.

The effects of pregnancy on the hepatic cytochrome P-450-dependent mixed-function monooxygenase system (P-450) from day 6 to day 18 of gestation were examined in the C57BL/6J mouse. Pregnancy induced an initial increase and then a decrease in total P-450 content, a decrease in microsomal aminopyrine-N-demethylase activity, and had no effect on microsomal ethylmorphine-N-demethylase activity. Pregnancy also induced in the C57BL/6J and the DBA/2J mice a new major isozyme of P-450 (P-450gest) as determined by high performance liquid chromatography and gel electrophoresis.

The caffeine breath test and caffeine urinary metabolite ratios in the Michigan cohort exposed to polybrominated biphenyls: a preliminary study.

A field biochemical epidemiology study was conducted using the Michigan cohort consisting of 51 rural residents exposed to polybrominated biphenyls (PBB). The study had three major objectives: a) to determine the serum half-life of the major PBB congener, hexabromobiphenyl (HBB), in the human, b) to determine if the PBB-exposed subjects had elevated cytochrome P-450I function as determined by the caffeine breath test (CBT) and the caffeine urinary metabolite ratio (CMR), and c) to determine the applicability of the CBT and CMR in field studies. PBB serum levels were detected in 36 of the 51 PBB-exposed subjects. The serum half-life of HBB was determined by comparing the current serum HBB values to the subject's previous serum values obtained 5 to 8 years earlier. The median HBB half-life was 12 years (range 4-97 years). The CBT and CMR were elevated in the subjects exposed to PBBs as compared to the values obtained from urban nonsmokers and were similar to those found in adults who smoke. A gender effect was seen in the PBB-exposed subjects, the median CBT and CMR values of the females being lower than the values of males. There was a correlation between the CBT and the HBB serum values (r2 = 0.2, p = 0.01) but not between CMR and HBB serum values. The CBT and CMR were easily conducted in the field and appear to be useful metabolic probes of cytochrome P-450I activity in human environmental toxicology.

Soluble adhesion molecules in the HIT syndrome: pathophysiologic role and therapeutic modulation.

Heparin-induced thrombocytopenia pathophysiology is now known to be a complex process that involves platelets, vascular endothelium, and leukocytes/lymphocytes. The activation products from these sites also contribute to the activation of coagulation and fibrinolytic deficit. While many of the markers of hemostatic activation processes have been found to be increased during the acute phase of heparin-induced thrombocytopenia syndromes, the circulating levels of soluble adhesion molecules such as the P, E, and L selectins, and intracellular and vascular cell adhesion molecules have not been reported. Since the pathophysiology of heparin-induced thrombocytopenia involves the activation of platelets, endothelium, and leukocytes, it is expected that the activation products related to these hemostatic systems including soluble selectins and cellular adhesion molecules will also be increased in circulating blood. These alterations may also provide an index of the pathophysiologic process. With the availability of highly sensitive enzyme-linked immunosorbent assays for soluble P, E, and L selectins, intracellular and vascular cell adhesion molecules, it is now possible to measure these adhesion molecules in biological fluids. This study reports on the circulating levels of various adhesion molecules in patients with heparin-induced thrombocytopenia and their modulation after therapeutic interventions by the use of direct thrombin inhibitors. With the availability of recombinant hirudin, it is now possible to treat these patients with alternate antithrombin agents. However, the immunoactivation of platelets and other cells as shown here indicates the possible need for additional adjunct therapeutic approaches to suppress their participation in the thrombotic process. The reported increase in the circulating levels of the soluble adhesion molecules during the heparin-induced thrombocytopenia and heparin-induced thrombocytopenia with thrombosis syndrome suggests that the antiheparin platelet factor 4 antibody is capable of modulating their regulation. The prognostic role of these mediators in the management of heparin-induced thrombocytopenia syndrome warrants further investigation.

Fractures in the under-3-year-old age cohort.

One hundred twenty-four urban children under 3 years of age admitted for fractures were retrospectively reviewed to determine the frequency of accidental and nonaccidental causes in this population. The fractures were categorized according to their mechanisms: motor-vehicle passenger or pedestrian accident, other accidents, or child abuse. There were no differences in the frequency of fractures by race, date of birth, or season in which the injury occurred. Skull fractures were most frequent (62%), followed by femur fractures (11%). There was a 26% increase in fractures between 1987 and 1989, especially in the non-motor vehicle cohort. Caretaker ignorance and/or carelessness was a common cause of fractures in the infant and toddler age group. Injuries were still occurring in spite of infant care seat use. The American public must be educated in preventive medicine and safety to decrease the senseless morbidity of our greatest resource.

The metabolic fate of 2H-labelled propafenone in man.

The metabolism of propafenone was studied in three humans by using 300 mg of the deuterated compound given orally. The excretion of propafenone and its metabolites was assessed by measuring the deuterium content in the excretion products by means of a microwave plasma detector. At selected times the metabolic pattern in plasma, urine, bile and feces was determined. The metabolites were isolated and the structure of the main compounds was characterized. Propafenone is absorbed completely and quantitatively metabolized. Unchanged propafenone is excreted neither in the urine nor in the feces in amounts of more than 1% the dose. The percentage of free unconjugated propafenone of the total deuterium content in the plasma was about 10% 3 and 6 hours after application. Conjugates of hydroxylated derivatives of propafenone are present predominantly in the plasma. The excretion of the metabolites takes place mainly by way of the feces, 53% of the dose is excreted by this route within 48 hours. Urinary elimination accounted for 18.5 and 38% of the dose within 48 hours in two humans. It was possible to elucidate the structure of 11 metabolites, accounting for more than 90% of the dose administered. The major metabolites are conjugates of 5-hydroxypropafenone and hydroxy-methoxy-propafenone with glucuronic and sulphuric acid and propafenone glucuronide. Furthermore, metabolic products of oxydative deamination pathway were identified, i.e. a glycol and a lactic acid derivative. C-C splitting yields a relatively large amount of 3-phenyl-propionic acid, while cleavage of the ether group leads to a phenolic product and is only of minor importance.

[Results of the Hamburg ovarian blastoma study]. / Ergebnisse der Hamburger Ovarialblastom-Studie.

A randomized chemotherapy trial is forthwith reported on, in which therapy A (vincristine/ifosfamide/tegafur) was compared with therapy B (fluorouracil/trofosfamide/methotrexate/melphalan). Both patient groups received trofosfamide and oral progesterone as maintenance treatment until the end of the second therapy year. Forty-six patients were randomized into group A, 63 into group B. The evaluation of the survival curves suggests a division of the patients into 3 groups with different mortality risks.

[Mysterious segmental stenosis of the small intestine (author's transl)]. / Rätselhafte segmentale Dnndarmstenosen. Morphologische und katammestische Analyse von 12 Krankheitsfällen.

Within a period of four years twelve cases of short small bowel stenoses were seen requiring surgical removal by resection because of acute ileus. Two of these cases were considered examples of an eosinophilic enteritis. The remaining cases were mostly men of an average age of 63 years. With one exception, they had been treated for cardiovascular diseases with various drugs, mostly potassium, digitalis and reserpine preparations. The intestinal stenoses were 1-3.5 cm in length with a constriction down to 1 mm diameter. Obviously, these obstructions develop from an infarct-like necrosis of the mucosa via a circular ulcer; the narrowing of the intestinal lumen is chiefly due to fibromuscular hyperplasia of the submucous coat. The morphological changes agree with the findings known from ischaemic intestinal lesions as well as with those in jejunal ulcus simplex. Hence, it appears justified to interpret them as ischaemic intestinal stricture. However, an additional and as yet unknown damaging factor seems to be involved.

Endodermal sinus tumour of the ovary: a comparative light and electron microscopic study.

An endodermal sinus tumour of the ovary from a 12 year old girl is analysed light- and electron microscopically. The histological appearance is characterized by glandular-papillary structures and microcystic-reticular areas with inclusion of occasional endodermal sinuses. By electron microscopy immature and highly differentiated cells can be distinguished. In analogy to certain structures in the human yolk sac the most differentiated cells in the tumour are regarded as neoplastic endoderm. On the basis of transitional forms between immature and differentiated cells a development of the latter from the former is suggested. The cells in mesenchyme-like areas differ from those of solid and glandular parts merely in the degree of cytoplasmic differentiation but are otherwise believed to represent the same "cell line". In addition to other possible cell functions the cytoplasmic features of the differentiated cell indicates a protein synthesis and secretion. From our observations it is concluded that the endodermal sinus tumour originates from germ cells and differentiates into yolk sac endoderm. The ultrastructural differences between this tumour and clear cell carcinomas of the ovary are discussed.

Expression of mRNA for the gap-junctional protein connexin43 in human colonic tissue is variable in response to beta-carotene supplementation.

Increased expression of the gap-junctional protein connexin43 (Cx43) is reported to be increased in mouse and human dermal fibroblasts in vitro in response to beta-carotene treatment. In the present study, we determined the level of Cx43 mRNA expression in colonic mucosa from normal subjects and subjects with a prior history of colonic polyps or cancer before and after three months of administration of a placebo or beta-carotene. RNA was reverse transcribed and used in a polymerase chain reaction assay employing primers selected from the human Cx43 gene sequence. Cx43 mRNA expression was normalized on the basis of beta 2-microglobulin mRNA expression. The beta-carotene concentration in colonic mucosa, as well as serum and diet, was also determined to establish a correlation between Cx43 expression and beta-carotene concentration. In an initial analysis of samples collected from 10 subjects before supplementation, the quantity of Cx43 mRNA was variable and did not correlate with beta-carotene intake or the concentration of beta-carotene in tissue or serum. In samples collected at zero and three months from eight subjects who were controls or received a placebo, there was no correlation between Cx43 mRNA level and tissue or serum beta-carotene concentration. In samples collected from subjects before and after three months of beta-carotene supplementation, there was a significant increase in tissue and serum beta-carotene concentration in all subjects and an increase in Cx43 mRNA expression after supplementation relative to baseline in four of six samples. The high variability in Cx43 expression indicates that induction of Cx43 mRNA expression is not solely dependent on the concentration of beta-carotene in diet, serum, or tissue. However, the results from subjects supplemented with beta-carotene suggest that induced expression may occur in colonic tissue of some individuals. some individuals.

Studies on the metabolism of propafenone. 3rd Comm.: Isolation of the conjugated metabolites in the dog and identification using fast atom bombardment mass spectrometry.

The metabolism and urinary and biliary excretion of propafenone (2-(2'-hydroxy-3'-propylamino-propoxy)-omega-phenyl-propiophenone hydrochloride) were studied in the dog. Approximately 20% of the dose was excreted in 24 h with the urine and about 65% with the bile. Propafenone was extensively metabolized. Less than 4% of the dose was recovered unchanged in urine and bile. The metabolites were mainly excreted as conjugates. Free metabolites accounted for less than 20% of the dose. Separation methods were developed to isolate and purify the conjugated metabolites. Fractionation on an Al2O3-column yielded a sulphate and a glucuronide fraction, further separation and purification was done by TLC and HPLC. Positive and negative ion fast atom bombardment mass spectra (FAB/MS) were obtained of the purified glucuronides. The structures of two hydroxylated propafenone derivatives and two O-methylated catechol-like derivatives were definitely proven by FAB/MS, the glucuronic acid moiety being conjugated to the hydroxyl function in the different aromatic rings. Two isomeric propafenone glucuronides were found in the bile, probably diastereomeric forms of the O-glucuronide. Thus FAB/MS proved to be a complementary method to electron impact ionization mass spectrometry (EI/MS) for studying drug metabolism. The structures of free and conjugated metabolites can be defined from the combination of both mass spectrometric techniques.

In vitro beta-carotene toxicity for human colon cancer cells.

Experiments were conducted to determine the effect of beta-carotene on human colon cancer cells in vitro. beta-Carotene solubilized in tetrahydrofuran (THF) was determined to be cytotoxic for three different cell lines: LS 180, SW 620, and HCT-15. The number of LS 180 and SW 620 cells surviving treatment with 2.9 microM beta-carotene was significantly reduced relative to THF-treated cells, and a similar reduction was achieved in HCT-15 cells with use of 5.8 microM beta-carotene. These concentrations are in the range achieved in serum of individuals supplemented with beta-carotene at 30 mg/day. There was no beta-carotene cytotoxicity in the concentration range that characterizes serum of unsupplemented individuals. Vitamin E at > 200 microM was not cytotoxic and at higher concentrations slightly stimulated proliferation of all three cell lines. Exposure of cells to vitamin E did not diminish the cytotoxicity of beta-carotene, suggesting that the toxic effect of beta-carotene is not due to prooxidant activity. Percent cytotoxicity was increased by extending the duration of exposure of cells to beta-carotene. Interestingly, beta-carotene cytotoxicity decreased with increasing cell density. This density-dependent toxicity was attributable to a higher beta-carotene concentration per cell for cells plated at lower densities. Thus toxicity of beta-carotene for colon cancer cells is dose, time, and cell density dependent and occurs in vitro at concentrations that can be achieved safely in humans.

[Labelling meproscillarin with tritium at the C-19 position (author's transl)]. / Radioaktive Markierung von Meproscillarin in der C-19-Methylgruppe mit Tritium.

Labelling of 14-hydroxy-3beta-[4-O-methyl-alpha-L-rhamnopyranosyl)oxy]-14beta-bufa-4,20,22-trienolide (meproscillarin, Clift) by introduction of dueterium and tritium at the metabolically stable C-19 position is described.

Studies on the metabolism of propafenone. 1st Comm.: synthesis and chromatographic/mass spectrometric properties of the labelled compound and of the reference substances.

The synthesis of 14C-propafenone and 2H-propafenone (propafenone: 2-(2'-hydroxy-3'-propylamino-propoxy)-omega-phenyl-propiophenone hydrochloride) and some reference compounds is described. The thin-layer chromatographic, high-performance liquid and gas chromatographic properties of the substances are described. Propafenone and the reference substances were studied by mass spectrometry and compared with each other, with respect to structural elucidation of the metabolites. The chromatographic and mass spectrometric data (key ions) enables the metabolites of propafenone to be identified in biological material.

Studies on the metabolism of propafenone. 2nd comm.: studies on the biotransformation in the dog.

Following administration of 14C- and 2H-labelled propafenone hydrochloride (2-(2'-hydroxy-3'-propylaminopropoxy)-omega-phenyl-propiophenone hydrochloride) to beagle dogs, the metabolites were isolated from urine and bile and analysed by mass spectrometry. Reference substances were synthesized on the basis of the structures postulated from the mass spectra and compared with the substances isolated from the biological material. Propafenone was absorbed completely following i.d. administration and eliminated mainly with the bile. Within 28 h 10% of the dose was excreted with the urine and 87% with the bile. Propafenone was extensively metabolized. Less than 1% of the dose was recovered as unchanged substance in urine and bile. The urinary and biliary metabolites were almost exclusively conjugated. The main metabolite, accounting for more than 30% of the dose, was propafenone glucuronide, followed by conjugates of hydroxylated propafenone derivatives with glucuronic acid and sulphuric acid. 5-Hydroxy-propafenone, a propafenone derivative hydroxylated in the middle aromatic ring, and a derivative hydroxylated in the omega-phenyl ring each accounted for about 15% of the dose. Besides these monohydroxy metabolites, two other O-methylated catechol-like derivatives, substituted in the different aromatic rings were recovered. The remainder of the metabolic products identified were mainly substances resulting from degradation of the propoxyamine side chain.

[Malignant non-epithelial tumors of the pancreas]. / Maligne nichtepitheliale Tumoren des Pankreas.

Nonepithelial malignant tumors of the pancreas are extremely rare neoplasms with a frequency of approximately 0.6%. They are always explored because of a suspected diagnosis of pancreatic carcinoma. Amongst the more than 600 primary pancreatic neoplasms in our pancreatic tumor archive only 5 neoplasms were of nonepithelial origin (one was a malignant peripheral nerve sheath tumor [MPNST], one a leiomyosarcoma, one a malignant mesothelioma, and two were peripheral neuroectodermal tumors [PNET]. The differential diagnosis includes secondary infiltration of the pancreas by mesenchymal tumors of the retroperitoneum, undifferentiated pancreatic carcinoma and, especially in the case of PNET, malignant lymphoma. Preoperative chemotherapy and down-staging can improve the operability and prognosis, especially in PNET.

[Disseminated lipogranulomatosis (Farber disease) with hydrops fetalis]. / Disseminierte Lipogranulomatose (M. Farber) mit Hydrops fetalis.

We report on a female preterm infant of 29 weeks' gestation with severe hydrops fetalis who died 3 days post natum as a result of disseminated intravascular coagulation. Autopsy findings included anasarca, bilateral pleural effusions, ascites and hepatosplenomegaly as well as multiple, up to pinhead sized, white granulomas on the surface of liver, spleen and lungs. Microscopy revealed storage macrophages of the reticuloendothelial system, especially in liver, spleen and bone marrow, the lymphatic organs, the salivary glands, the thyroid gland and the suprarenal medulla. Cerebrum, heart, kidneys, intestines and placenta were not afflicted. Atrophy of the lymphatic compartments in the spleen, lymph nodes and thymus, as well as disorder of the liver texture, are presumably a secondary result. The diagnosis of Farber's disease was established biochemically by the demonstration of ceramide depositions in the spleen, and in fibroblast cultures in situ by the accumulation of ceramide released from loaded radioactive glucosylceramide. Ultrastructurally, corresponding storage lysosomes were found in macrophages. To our knowledge this is the first account of Farber's disease in a preterm infant with hydrops fetalis.

Solid and cystic acinar cell tumour of the pancreas. A tumour in young women with favourable prognosis.

The clinico-pathological features of five cases with a distinctive pancreatic tumour are presented. The tumours, which occurred only in young women and an adolescent girl, were of large size (2.5-10 cm), had an uncharacteristic symptomatology and showed fibrous encapsulation with no evidence of metastases. The histological features include (1) solid areas with a monomorphic cell pattern and intracellular PAS positive globules, and (2) large foci of degeneration with cystic necroses, haemorrhages and cholesterol granulomas. Some tumour cells were positive for alpha1-antitrypsin. The ultrastructural demonstration of zymogen-like granules suggests an acinar origin for the tumours. We therefore propose the term solid and cystic acinar cell tumour. This tumour resembles the so called pancreatoblastomas in small children in some respects. It must be clearly distinguished, on the other hand, from acinar cell carcinoma with its acinic structures and poor prognosis. This lesion is not included in the WHO classification of pancreatic neoplasms.