Dehydroepiandrosterone effects on the mRNA levels of peroxisome proliferator-activated receptors and their coactivators in human hepatoma HepG2 cells.

To investigate the effect of dehydroepiandrosterone (DHEA) on intracellular mRNA levels of peroxisome proliferator-activated receptors (PPARs) and PPAR-gamma coactivators (PGCs), we conducted a quantitative real-time RT-PCR study using HepG2 cells. Treatment with 100 micromol/l DHEA for 2-20 h caused a time-dependent elevation of mRNA levels in the cells. Upon 20 h, PPAR-alpha, -gamma1, and -gamma2 mRNAs and PGC-1alpha and -1beta mRNAs increased to 157, 161, 155, 656, and 475% of control levels, respectively (p < 0.05 each). Treatment with actinomycin D for 2.5-8 h revealed a significant stabilization effect of DHEA on PPAR-gamma1 and PGC-1alpha mRNAs at both 2.5 and 8 h incubation periods and a mild but significant stabilization effect on PGC-1beta mRNA at the 8 h incubation period suggesting that DHEA can modulate turnover of these mRNA transcripts. Basal mRNA levels of PPAR-alpha and PGC-1alpha were significantly suppressed upon 20 h treatment with cycloheximide, while those of PPAR-gamma1, -gamma2, and PGC-1beta were elevated. Cycloheximide also significantly reduced DHEA-induced accumulation of PPAR-alpha, -gamma1, -gamma2, and PGC-1alpha mRNAs, demonstrating the dependence of the DHEA action on de novo protein synthesis. The findings demonstrate that a supraphysiological concentration of DHEA can substantially influence gene expression of the PPAR signalling machinery at both transcriptional and posttranscriptional levels.

In vitro glycosidation potential towards olomoucine-type cyclin-dependent kinase inhibitors in rodent and primate microsomes.

Interspecies differences in glycosidation potential in mammalian tissues represent a factor contributing to ambiguity when endobiotic and/or xenobiotic metabolic pathways are extrapolated from animals to man. Using the TLC/autoradiographic technique, we conducted an in vitro investigation involving mouse, rat, monkey, as well as human liver and kidney microsomes to evaluate their glycoconjugation potential towards (3)H-labeled, purine-derived selective inhibitors of cyclin-dependent kinases such as olomoucine, bohemine, roscovitine, 6-(2-hydroxybenzyl)amino-2-(1-hydroxymethyl-2-methylpropyl)amino-9-isopropylpurine (compound A-4), and 6-(3-hydroxybenzyl)amino-2-[(1(R/S)-hydroxymethyl)propyl]amino-9-isopropylpurine (compound A-5) as aglycones. Principally, this study confirmed the aliphatic hydroxyl group of olomoucine-type inhibitors as a relatively suitable target for glucuronide, glucoside, xyloside, galactoside, and/or N-acetylaminoglucoside conjugation. Of the tissues examined, only the mouse microsomes were able to perform glucosidation and galactosidation reactions with the aglycones. On the other hand, monkey microsomes were superior to the mouse microsomes in a variety of glucuronide conjugates produced with compounds A-4 and A-5.

[Importance of reelin in the development of the central nervous system. II. Cyclin-dependent kinase 5 as an important element in the reelin signalling cascade]. / Význam reelinu pri vývoji centrálního nervového systému. II. Cyklin-dependentní kináza 5 jako významný clánek reelinové signalizacní kaskády.

Expression of cyclin-dependent kinase 5 (Cdk5) gradually increases during embryonal development until it reaches maximal levels that persist in the adult brain. Cdk5 represents a key component in the reelin signalling cascade. Activity of p35/Cdk5 is critical for the development of normal, six-layered structure of the mammalian brain cortex. Although it is not required for splitting of the cortical pre-plate into the marginal zone and sub-plate, a process that is affected in absence of the reelin and/or Dab1, it is substantial for the later-born neurons to bypass sub-plate neurons and to enter into the embryonal cortical plate. The defect of p35/Cdk5 disrupts the classical inside-out histogenetic pattern of neocortex. Neurons that fail to migrate past older layers of cells accumulate beneath the sub-plate thus forming an abnormal, inverted outside-in configuration in p35/Cdk5 defective animals. Exploration of neuronal migration disturbancies can shed new light on some diseases of human brain development including different heterotopias and lissencephaly. From an evolutionary point of view, the involvement of Cdk5 in the reelin signalling cascade could define a new step in the brain gray matter development.

In vitro biotransformation of 2,6,9-trisubstituted purine-derived cyclin-dependent kinase inhibitor bohemine by mouse liver microsomes.

1. Biotransformation pathways of the cyclin-dependent kinase inhibitor 6-benzylamino-2-(3-hydroxypropylamino)-9-isopropylpurine (bohemine) by mouse liver microsomes in vitro were investigated. 2. Metabolite profiles of [8-(3)H]-labelled bohemine were established by TLC/(3)H-autoradiography and enzymatic and MS analyses were used to elucidate the chemical structures of the metabolites. The structures of the main primary metabolites were confirmed by synthesis of authentic compounds. 3. A schema of the primary NADPH-dependent pathways has been proposed involving N(2)- and N9-dealkylation, N(6)-debenzylation, aromatic hydroxylation, and C2 side chain oxidation of bohemine. Three of the primary metabolites detected, 6-(benzylamino)-2-(3-hydroxypropylamino)purine (M4), 6-amino-2-(3-hydroxypropylamino)-9-isopropylpurine (M5) and 6-(4-hydroxybenzylamino)-2-(3-hydroxypropylamino)-9-isopropylpurine (M6), all retaining their parent primary hydroxyl group, were subsequently shown to be converted, by a liver cytosolic NAD(+)-dependent system, into their corresponding carboxylic acids. M6 was subject to microsomal glycosidations requiring UDP-sugar donors. NADPH-dependent conversion of M6 into M5 by microsomes was also demonstrated. 4. Cytochrome P450 (CYP) enzymes-selective inhibitors were used to identify CYPs involved in bohemine biotransformation. The findings suggested that CYP2a and CYP3a substantially contributed to the NADPH-dependent bohemine transformation in vitro. 5. The findings will facilitate experiments designed to dissect enzymatic systems catalysing clearance of C2,C6,N9-trisubstituted purine compounds from mammalian tissues.

[Precision-cut tissue sections--an important system for study of metabolism in vitro]. / Tkánové rezy o definované tloust'ce--významné systémy pro studie metabolismu in vitro.

Tissue slices represent a useful in vitro method for metabolic, pharmacologic and toxicologic studies. To their major advantage belong preserved cell to cell interactions, architecture and complexity of the tissue. They enable to reduce the number of laboratory animals, that are necessary for research purposes. Tissue slices are more similar to in vivo situation than other in vitro models such as microsomes, cell suspensions or cell cultures. Slices can be cultured up to 72 hours or even more. Preparation of the slices is rapid and easy. Slices can be prepared from any organ. There is a possibility of cross-species comparison, co-culturing of slices derived from different organs. Excess of the material can be cryopreserved for later studies.

Abdominal zygomycotic thromboangiitis in a patient with AML and t(1;13;14).

The case report of a 61 year-old man with AML M2 FAB, t(1; 13; 14) and zygomycotic mesenterial thromboangiitis is presented. Two induction cycles of chemotherapy were administered due to primary drug resistance. They were complicated by pneumonia, colonic pseudo-obstruction and perforation with peritonitis. The patient died on the 40th day of therapy, 4 days after undergoing palliative surgery. Zygomycotic thromboangiitis, which very probably contributed to the intestinal perforation, was confirmed morphologically at necropsy. the novel complex chromosomal translocation t(1; 13; 14) (q31; q32; q24) and the problems connected with the diagnosis of invasive fungal infections are discussed.

Proliferation of reticuloendothelial system (RES) in rats with altered vision or smell (to the hypothetic neural regulation of the RES).

A long-term administration of an azo-dye, trypan blue, induced a reactive proliferation of RES in rats. Reaction of the RES was followed in rats whose optical analyzer was eliminated by enucleation of bulbs after the birth, in rats whose smell analyzer was altered by a repeated long-term exposition to ammoniac and in normal rats. Blind rats showed a striking proliferation of histiocytes of the RES in the liver, while the reaction was weak in the spleen and lymph nodes. In contrary to that, rats with altered smell and also the normal rats exhibited standard reactive changes in the spleen and lymph nodes and distinctly weaker reaction in the liver. The obtained results support an idea that vegetative neural mechanisms play a role in control and coordination of RES reactions.

Regional lymph nodes in cancer III. B-lymphocyte region.

The available evidence on the reaction of B-cell zone, the role of B-cell response in interactions with T-cells, MPS cells, tumor antigens and antigen-antibody complexes is presented. The authors comment the interpretation of B-cell response in prognosis of cancer patients.

Regional lymph nodes in cancer. IV. T-lymphocyte region.

The data on the role of T-cell reaction in interactions with B-cell response and MPS reaction are summarized. The importance of paracortical activity (T-zone) in experimental and human tumors is discussed with regard to prognostic interpretation of T-cell response.

A complex choristoma of the conjunctiva.

The choristoma belongs to marginal problems in ophthalmology, nevertheless it is not negligible, and in the differential diagnosis of the tumors of the conjunctiva and the cornea we must take it into consideration especially in connection with the potential possibility of its malignant progression. The finding of a choristoma may signal a simultaneous occurrence of other lesions of an analogous origin in other localities than that of the bulbus or the orbit, which should lead the ophthalmologist to a complex examination of the patient and an interdisciplinary cooperation with the neurologist, oto-rhino-laryngologist, dermatologist, and others.

Regional lymph nodes in cancer. II. Macrophages.

The survey is given of the actual knowledge of mononuclear phagocyte system which is involved in reactive changes of regional lymph nodes in experimental and human tumors. The practical aspects of their role in the development of tumors are discussed.

[Morphological correlates of anti-tumor immunity. IV. Mastocytes in regional lymph nodes]. / Morfologické koreláty protinádorové imunity. IV. Mastocyty v regionálních lymfatických uzlinách.

Mastocytes of regional lymph nodes were studied in three groups of oncological patients suffering from cancer of stomach, breast or lung. Lymph nodes in all three groups showed varied numbers of mastocytes; top figures were found in stomach cancer. There was a convincing relation of lymph node mastocyte number neither to lymphatic tissue reaction (predominant lymphocytes, predominant germinal centres, lymphocyte depletion, sinus histiocytosis, unstimulated lymph node) nor to frequency of secondaries nor to patient survival rate.

[Morphologic correlations of anti-tumor immunity. III. Reactions of regional lymph nodes]. / Morfologické koreláty protinádorové imunity. III. Reakce regionálních lymfatických uzlin.

Regional lymph nodes were examined in 4 groups of oncologic patients (with cancer of oral cavity, breast, stomach and lung). Histological types of the lymph node reaction were correlated to patients' survival. Favourable prognosis in all four groups was related to lymphocytic predominance, especially when combined with sinus histiocytosis. The predominance of germinal centres (combined with sinus histiocytosis) was a favourable sign but in stomach cancer. A marker of unfavourable prognosis was lymphocytic depletion, especially when sinus histiocytosis was absent.

[Prognostic significance of reactive changes in regional lymph nodes in stomach and breast carcinoma]. / Prognostický význam reaktivnich zmen v regionálnich uzlinách u karcinomu zaludku a mammy.

The regional lymph nodes have been examined in 80 cases of carcinoma of the stomach and in 70 cases of cancer of the breast, and their reactive type has been determined and correlated with the duration of survival of the patients. A prognostically favorable sign in both groups appeared to be the finding of a predominance of lymphocytes combined with a sinus histiocytosis, in the case of cancer of the stomach, then also a reaction of the type of prevalence of germinae centers together with sinus histiocytosis. Prognostically unfavorable was apparently the finding of a depletion of lymphocytes, especially if sinus histiocytosis was also lacking. The isolated finding of a single lymph node metastasis, in itself, if we do not take into account the reactive type, could not be interpreted in the sense of a worse prognosis from the viewpoint of survival time.

Prognostic significance of reactive changes in regional lymph nodes in gastric and mammary carcinomas.

Regional lymph nodes in 80 cases of gastric and 70 cases of breast cancer have been examined and the reactive types have been correlated to the survival periods of the patients. In both groups of patients, a prognostically favorable sign was the detection of lymphocytic predominance combined with sinus histiocytosis, moreover, in patients with gastric cancer also the reactive type of predominance of germinal centers with sinus histiocytosis. Prognostically unfavorable sign was the detection of lymphocytic depletion, in particular if sinus histiocytosis was absent. The presence of metastases in the lymph nodes--apart from the reactive type--cannot be regarded as a sign of bad prognosis.