The polycyclic musk 7-acetyl-1,1,3,4,4,6-hexamethyl-1,2,3,4-tetrahydronaphthaline lacks liver tumor initiating and promoting activity in rats exposed to human-relevant doses.

7-Acetyl-1,1,3,4,4,6-hexamethyl- 1,2,3,4-tetra-hydronaphthaline (AHTN) is one of the two most widely used fragrances of a group of substances known collectively as the polycyclic musks. In the last few years evidence has been accumulating that AHTN is hepatotoxic when administered at high doses. In the present study the subchronic hepatotoxicity of AHTN administered to rats at doses within the human exposure range was evaluated. For this purpose female and male juvenile Wistar rats were exposed to AHTN (300 microg/kg body weight per day, i.p.) alone or to a single dose of diethylnitrosamine (DEN) (100 mg/kg body weight, i.p.) followed by AHTN (1, 10, 100 or 300 microg/kg body weight per day, i.p.) for 90 days. Thereafter the liver architecture as well as the presence of placental glutathione S-transferase (GST-P)-positive hepatic lesions was assessed. In male animals receiving AHTN alone or in combination with DEN the number of GST-P-positive single hepatocytes was similar to that in untreated rats, while GST-P-positive mini-foci and foci were not observed. In the case of female rats the number of GST-P-positive single hepatocytes and mini-foci in AHTN-treated rats was similar to that in untreated animals, whereas in those animals receiving AHTN either alone or in combination with DEN, GST-P-positive foci could not be detected or were present in a number as similar to that in untreated rats. In conclusion, in the present study it has been shown that AHTN administered over a 90-day period in concentrations similar to those taken up daily by humans does not lead to hepatotoxicity.

Quantitative detection of tumor M2-pyruvate kinase in plasma of patients with lung cancer in comparison to other lung diseases.

Lung cancer is one of the predominant causes of cancer death. The aim of this project is the development of a screening method in persons with high risk for developing lung cancer, based on the measurement of Tumor M2-pyruvate kinase (Tumor M2-PK). Tumor M2-PK is quantitatively detectable in ethylenediaminetetraacetic acid-plasma with a sensitive enzyme-linked immunosorbent assay. So far, 60 patients with newly diagnosed lung cancer were included. These were compared to 24 patients with acute inflammatory lung diseases, 56 patients with pneumoconiosis, 22 patients with obstructive airway diseases, and 28 healthy persons. Tumor patients and some individuals suffering from severe inflammatory lung diseases had significantly higher Tumor M2-PK concentrations in ethylenediaminetetraacetic acid-plasma than all the other groups. The histologic tumor type had no influence on the plasma levels of Tumor M2-PK. Tumor M2-PK concentrations correlate strongly with the tumor stage, showing significantly increasing concentrations with progressive tumor stages. The present data indicate that Tumor M2-PK could be a valuable tumor marker for the detection of lung cancer.

Tumor M2-PK and glutaminolytic enzymes in the metabolic shift of tumor cells.

The pyruvate kinase isoenzyme M2-PK is known to be associated with a metabolic shift that is characteristic for tumor cells. Meanwhile, the universal expression of this isoenzyme is the basis for the detection of various tumor diseases in human clinical diagnosis. Other enzymes which are known to be essential for this tumor specific metabolic shift in rat chemical carcinogenesis are the NADP-dependent enzymes malic enzyme, isocitrate dehydrogenase and glucose 6-phosphate dehydrogenase. To evaluate the role of these enzymes in human carcinogenesis, we compared their enzymatic activities in normal colon mucosa and tissues derived from primary colon tumors. Histochemical staining showed that the enzyme activities were restricted to mucosal colon cells and colon cancer cells. The enzymes were strongly but heterogeneously expressed in the tumor tissues, whereas staining of normal mucosa was weak. Tumor M2-PK showed the most prominent differences in normal colon mucosa and colon cancer cells.

Expression of pyruvate kinase M2 in preneoplastic hepatic foci of N-nitrosomorpholine-treated rats.

The expression of the pyruvate kinase (PK) isoenzymes L and M2 was analysed in the livers of rats treated with the hepatocarcinogenic agent N-nitrosomorpholine (NNM) in the drinking water. In control animals L-PK expression was restricted to liver parenchymal cells, whereas M2-PK was detected in bile duct epithelial, blood vessel wall, endothelial and Kupffer cells. In rats treated with NNM proliferating oval cells were consistently L-PK negative and M2-PK positive, while the ductal cells of cholangiofibroses were clearly L-PK positive and coexpressed M2-PK. However, no morphological differentiation of ductal cells into hepatocyte-like cells was observed. In the clear and acidophilic cell foci storing glycogen in excess strong staining for L-PK was observed. In glycogen-poor foci induced by NNM a shift from L-PK to M2-PK expression takes place.

Energy metabolism in the involuting mammary gland.

A strong and coordinated upregulation of the glycolytic, glutaminolytic and pentose phosphate pathway enzymes occurs during the onset of lactation in the normal mouse mammary gland. Induction of apoptosis by removing the pups led to an inactivation of the same enzymes with different time courses. While the ATP-consuming glycolytic 6-phosphofructo 1-kinase and mitochondrial bound hexokinase still remained high on days one and two of involution, the ATP-regenerating pyruvate kinase was immediately reduced. The enzymes of the pentose phosphate and glutaminolytic pathway were inactivated on the first two days of involution. In accordance with such an inactivation of the enzymes ATP, GTP, UTP, ADP, NAD NADH and lactate concentrations decreased. The synthetic product of UTP, UDP-N-acetylglucosamine, increased. AMP was found in the milk, not in the epithelial cells. The inactivation of the enzymes was caused by partial proteolysis or by a loss of the intact proteins from the cytosol without signs of proteolysis.

Neurotoxicity risk assessment of MPTP (N-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) as a synthetic impurity of drugs.

1-Methyl-4-phenyl-1,2,3,6 tetrahydropyridine (MPTP) induces symptoms indistinguishable from those of Parkinson's disease. It selectively destroys dopaminergic neurons in the substantia nigra and the globus pallidus. Death of these same neurons is apparently the cause of idiopathic Parkinson's disease. As phenyl-1,2,3,6 tetrahydropyridine is a commonly encountered subunit in heterocyclic drugs and because MPTP was found as a minor impurity in early batches of a candidate drug at Merck KGaA, it may be assumed that MPTP will also be present as an as yet undiscovered minor impurity in various existing drugs. A neurotoxicity risk assessment on MPTP has been conducted to define the risk of MPTP as an impurity in drugs that are used orally. This risk assessment has shown that compounds containing less than 5.0 p.p.m. MPTP administered orally will not cause a neurotoxicological health risk to patients treated with such a drug.

Expression of L- and M2-pyruvate kinases in proliferating oval cells and cholangiocellular lesions developing in the livers of rats fed a methyl-deficient diet.

Male outbred Sprague-Dawley rats were fed a choline-deficient diet containing 0.1% w/w DL-ethionine (CDE) for up to 22 weeks. The expression of the pyruvate kinase isoenzymes L (L-PK) and M2 (M2-PK) was immunohistochemically analyzed in liver slices from rats killed 4, 10, 14 and 22 weeks after starting the treatment. M2-PK was detected in bile duct epithelial cells of untreated rats and in proliferating oval cells, cholangiofibroses and cholangiofibromas of CDE-fed animals. Thus, M2-PK can be viewed as a positive marker of the bile duct epithelial/oval cell compartment. L-PK, a parenchymal cell-specific protein in untreated rat liver, was not present in proliferating oval cells, but was consistently observed in cells that were part of the ductal structures in the cholangiofibroses and cholangiofibromas. Based on their morphology, the L-PK-positive duct cells were undoubtedly part of the bile duct epithelial cell lineage and no L-PK-positive hepatocyte-like cells were observed in the ducts. Hence, this study clearly shows that the mere presence of a liver parenchymal cell marker in cells of the bile duct epithelial/oval cell compartment does not necessarily preclude that these cells are undergoing a differentiation into preneoplastic parenchymal cells, as has previously been suggested.

[The percutaneous extraction of an embolized Pudenz-Heyer catheter fragment from the pulmonary artery]. / Perkutane Extraktion eines embolisierten Pudenz-Heyer-Katheterfragmentes aus der Arteria pulmonalis.

A ventriculoatrial shunt had been placed 3 years previously in a now 17-year-old boy because of obstructive hydrocephalus of unknown cause. He presented with symptoms of elevated cerebrospinal fluid pressure and computed tomography demonstrated a dilated ventricular system. The chest X-ray film revealed a break in the shunt catheter at the level of the right clavicle with embolization of its 13 cm long distal part into the main stem and right branch of the pulmonary artery. The fragment was retrieved without complication with a percutaneously and transvenously introduced basket catheter. Centrally embolized catheter fragments should be removed as soon as possible. The percutaneous route is well tolerated by and of low risk for the patient. It should always be attempted before any surgical intervention.

An approach based on two-dimensional graph theory for structural cluster detection and its histopathological application.

An approach based on graph theory is described for detecting clusters of cells in tissue specimens (two-dimensional space). With a set of discrete basic elements (cell nuclei) having several measurable features (area, surface, main and minor axis of best-fitting ellipses) a graph is defined as having attributes associated with edges. Different minimum spanning trees (MSTs) can be constructed using different weight functions on the attributes (attributed MST). Analysis of the MST and of an attributed MST by use of a decomposition function allows detection of image areas with similar local properties. These clusters, which are then clusters of the tree, describe, for example, partial growth in different directions in a case of a human fibrosarcoma assuming that tumour cell nuclei are homogeneous with respect to their configuration and size. The model allows the separation of clusters of tumour cells growing in different directions and the approximation of the different growth angles. This decomposition also allows us to create new (higher) orders of structure (cluster tree).

Long distance image transfer: first results of its use in histopathological diagnosis.

Histopathological images were transferred by use of normal telephone lines between three pathology institutes located in three different cities in the FRG. Images were digitized using a colour TV camera and stored in a special computerized image transmission system. The stored image was transferred and visualized on a (receiver) colour TV screen after dialing the telephone number connected to the receiver image transmission system. An additional telephone dialogue was activated by use of a normal acoustic telephone, and the diagnostic difficulties of the underlying image were discussed. Diagnostic assistance was possible in all transferred cases as well as histopathological diagnosis. Resolution of the images was set at 512 x 512 pixel x 8 bit. Image transfer time was 3.2 min on average. The differences between the original and transferred image were measured by "retransfer" of the original image and by subtracting the two images from each other. No major transfer errors could be measured.

Biotinylated epidermal growth factor: a useful tool for the histochemical analysis of specific binding sites.

Epidermal growth factor (EGF) was labelled with biotin via modification of either the amino or carboxyl groups, using suitable reagents, namely biotinyl-N-hydroxysuccinimide ester or biotinamidocaproyl hydrazide. To assure that the specific binding capacity of EGF is retained despite its chemical modification, displacement of the EGF by biotinylated derivatives in a routine binding assay was performed. The inhibitory potency compared to unmodified EGF was only slightly reduced. This result is the prerequisite for testing the usefulness of biotinylated EGF in histochemistry. The biotinylated probes were applied to sections of human tumour tissue and of monkey organs (liver, kidney, uterus of Cynomolgus and Rhesus monkey) to localize the specific binding sites for EGF. Formalin-fixed, paraffin-embedded tissue sections were deparaffinized and incubated with the probes at a concentration of 10 micrograms ml-1 at room temperature for 60 min. Specific binding of the EGF was visualized by the avidin-biotin techniques (ABC). A positive reaction in conjunction with appropriate controls by competitive inhibition was seen for all monkey tissue sections and for the following number of cancer cases: breast carcinoma: 7/10; mesothelioma: 2/4; lung carcinoma: 1/3; colon carcinoma: 1/3. The staining properties were similar for both types of probes that differed in the functional group that is involved in modification by biotin attachment. However, the batches with modification of the amino groups stained more intensely and more distinctly than the carboxyl modified EGF. Overall, the data indicate that the ligand properties of the EGF are not impaired by biotinylation of the two types of functional groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Differentiation-related expression of epidermal growth factor receptors in human lung carcinoma demonstrated histochemically by biotinylated epidermal growth factor.

Biotin-labeled epidermal growth factor (EGF) was applied to routinely processed sections of 64 cases of human lung carcinoma as a histochemical tool for demonstrating EGF-specific receptors. Formalin-fixed, paraffin-embedded tissue sections were deparaffinized and incubated with the labeled EGF (10 micrograms/ml) for 60 min at room temperature. The specific binding of the growth factor to its receptor was visualized by the avidin-biotin complex (ABC) technique. Positive binding capacities were obtained for the following cases: 15/16 epidermoid carcinoma; 13/15 adenocarcinoma; 2/11 large cell anaplastic carcinoma; 12/20 small cell anaplastic carcinoma; 0/11 normal lung tissue; 0/6 main bronchi; 0/1 hamartoma; 0/1 primary fibrosarcoma of lung. In addition, a strong positive reaction was seen for neutrophilic granulocytes present within the tumorous tissue. Data indicate that EGF receptors are frequently expressed in more differentiated carcinoma in comparison with anaplastic carcinoma of lung.

[Ancylostomiasis in the mines. Apropos of 2 cases]. / L'ankylostomose dans les mines. A propos de deux cas.

The etiological importance of Ancylostomidae in miners' anaemia has been discovered in 1880 by Perroncito during the boring of the Saint-Gothard tunnel in a dreadful epidemic. Thereafter in France, the illness disappeared between 1930 and 1940 because of the preventive dispositions realized. Two present cases of contamination in the coal mines of Lorraine due to N. americanus drive us to remember the interest of systematic faecal examinations of the miners at the engagement and every five years.

Toxicological study on pramiverine (author's transl). / Toxikologische Prüfung von Pramiverin

4,4-Diphenyl-N-isopropyl-cyclohexylamine-hydrochloride (pramiverine, Sistalgin) was investigated alone and in combination (1 + 1000) with N-methyl-N-(2,3-dimethyl-5-oxo-1-phenyl-3-pyrazolin-4-yl)-aminomethanesulfonate (metamizole) in various species for acute toxicity after oral and intravenous administration, for local tolerance and subacute toxicity. In addition, long-term trials and reproduction toxicity studies were performed with pramiverine. Pramiverine was only slightly toxic and was well tolerated locally as an ampoule solution also in combination with metamizole. The acute trials of the two compounds in mice and rats like the subacute study in rats showed that the toxicity of metamizole was not increased by pramiverine. Rats tolerated in the long-term trial all pramiverine doses examined (0.5; 5.0; 50.0 mg/kg), while cholinolytic concomitant effects were observed in dogs under higher doses (5.0 and 20.0 mg/kg). Pramiverine did not have a foetotoxic and teratogenic effect in mice, rats and rabbits. In the perinatal and postnatal experiment in rats no effect on foetal development, viability and growth of the offspring as well as the course of labour and lactation ability of the dams was observed.