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Micrometer-scale patterns of a defined surface chemistry and structure were produced on both ultraflat Au(1 1 1) and on gold-coated monocrystalline silicon surfaces by a method combining microcontact printing, wet chemical etching and the replacement of etch-resist self-assembled monolayers (SAMs) by functionalized or reactive SAMs. Key steps in this methodology were characterized by X-ray photoelectron spectroscopy (XPS), ellipsometry and contact angle measurements. The covalent immobilization of (functional) biological systems on these surfaces was tested using an N-hydroxysuccinimide ester ω-functionalized disulphide (DSU), which covalently binds primary amines without the need for further activation steps. Atomic force microscope images of native collagen V single molecules immobilized on these patterned surfaces revealed both high spatial resolution and strong attachment to the monolayer/gold surface. Microcontact printing of DSU is shown to be feasible on specially prepared, ultraflat Au(1 1 1) surfaces providing a valuable tool for scanning probe experiments with biomolecules. The retention of enzymatic activity upon immobilization of protein was demonstrated for the case of horseradish peroxidase. The described approach can thus be used to confine biological activity to predetermined sites on microstructured gold/silicon devices – an important capability in biomedical and biomolecular research. © 1999 Kluwer Academic Publishers [ABSTRACT FROM AUTHOR]

Steroids are essential for the control of oedema in human malignant glioma patients but may interfere with the efficacy of chemotherapy. Boswellic acids are phytotherapeutic anti-inflammatory agents that may be alternative drugs to corticosteroids in the treatment of cerebral oedema. Here, we report that boswellic acids are cytotoxic to malignant glioma cells at low micromolar concentrations. In-situ DNA end labelling and electron microscopy reveal that boswellic acids induce apoptosis. Boswellic acid-induced apoptosis requires protein, but not RNA synthesis, and is neither associated with free radical formation nor blocked by free radical scavengers. The levels of BAX and BCL-2 proteins remain unaltered during boswellic acid-induced apoptosis. p21 expression is induced by boswellic acids via a p53-independent pathway. Ectopic expression of wild-type p53 also induces p21, and facilitates boswellic acid-induced apoptosis. However, targeted disruption of the p21 genes in colon carcinoma cells enhances rather than decreases boswellic acid toxicity. Ectopic expression of neither BCL-2 nor the caspase inhibitor, CRM-A, is protective. In contrast to steroids, subtoxic concentrations of boswellic acids do not interfere with cancer drug toxicity of glioma cells in acute cytotoxicity or clonogenic cell death assays. Also, in contrast to steroids, boswellic acids synergize with the cytotoxic cytokine, CD95 ligand, in inducing glioma cell apoptosis. This effect is probably mediated by inhibition of RNA synthesis and is not associated with changes of CD95 expression at the cell surface. Further studies in laboratory animals and in human patients are required to determine whether boswellic acids may be a useful adjunct to the medical management of human malignant glioma. [ABSTRACT FROM AUTHOR]

We studied the time of appearance of renin-immunoreactive cells in prenatal human kidneys by immunohistochemistry and transmission electron microscopy (TEM). Renin immunoreactivity was detectable as early as the 5 or 6 gestational week in the mesonephros and appeared at 8 weeks of gestation in the metanephros. In a 8 week old embryo 'renin granules' were seen by TEM in the juxtaglomerular epithelioid granular cells of metanephric tissue. In the embryos delivered by prostaglandin-induced abortion, granular renin immunoreactivity was also present in the cells of the proximal convoluted tubule. Our results support the contention that the renin-angiotensin system is active during fetal life and indicates a role for renin even during the embryonic period. In addition, they document a morphologically detectable effect of prostaglandins on the distribution of renin in the kidney. [ABSTRACT FROM AUTHOR]

A patient with central nervous system involvement by hairy cell leukemia is reported. Hairy cells were identified in the cerebrospinal fluid by electron microscopy and by tartrate-resistant acid phosphatase positive staining. Intrathecal treatment with methotrexate resulted in neurologic improvement, but was complicated by Cryptococcus neoformans meningitis. The leukemic phase of the disease was later successfully controlled by treatment with alpha interferon. Surface marker studies indicated a B- and T-cell phenotype of the hairy cells. [ABSTRACT FROM AUTHOR]

A gene encoding the p53 val135 mutant, which assumes mutant conformation at 38.5 degrees C and wild-type conformation at 32.5 degrees C, was introduced into p53-deficient K562 myeloid leukemia cells. Forced expression of wild-type, but not mutant, p53 resulted in growth arrest, accumulation of p21 and Bax proteins, and delayed cell death. Wild-type p53 enhanced the cytotoxic effects of some drugs and attenuated those of others. Compared with wild-type p53, mutant p53 induced much stronger sensitization to drug cytotoxicity. This occurred in the absence of effects on cell cycle progression or activation of several p53 target genes. Although both mutant and wild-type p53 induced changes of immunophenotype, no specific pattern of differentiation was associated with enhanced chemosensitivity. Thus, (1) induction of growth arrest and activation of p53 target genes such as p21 and bax are linked to the wild-type conformation of p53; (2) p53 induces immunophenotypic changes of myeloid leukemia cells suggestive of multidirectional differentiation in a conformation-dependent manner; and (3) (so-called) mutant p53 induces chemosensitization in the absence of effects on cell cycle progression, activation of bax, p21, gadd45 and mdm-2, or a specific pattern of differentiation; and (4) chemosensitization mediated by wild-type p53 may be masked by transcription-dependent induction of growth arrest. [ABSTRACT FROM AUTHOR]

Antidiuretic hormone (ADH; 2.5 x 10(-8) M vasotocin) produces a stimulation of apical fluid phase endocytosis, protein secretion and NaCl reabsorption in Xenopus laevis A6 distal nephron cell epithelia pretreated with aldosterone (10(-6) M). The increase of NaCl transport is mediated by a sequential opening of apical Cl and Na conductances. The aim of this study was to characterize the actin and tubulin cytoskeleton of A6 cells and to assess the impact of its disruption on baseline and ADH-induced apical vesicular membrane movements and ion transport to test for possible functional links. The microfilament (MF) and microtubule (MT) networks and their disruption were visualized by confocal laser microscopy. Conditions of depolimerization were selected, by cytochalasin D or cold and nocodazole, respectively. MF disruption produced an increase in baseline apical protein secretion (exocytic movements) (plus 18%) and a decrease of its induction by ADH (minus 35%). MF disruption also increased baseline horseradish peroxidase uptake (endocytic movements) (plus 21%), however, without affecting its ADH-induced increase. In the case of MT disruption, the ADH-induced stimulation of both protein secretion and fluid phase endocytosis was decreased by 70 and 44%, respectively. At the ion transport level, MF and MT disruption only insignificantly affected the ADH-induced Cl conductance, while they decreased the ADH-induced stimulation of Na transport (amiloride-sensitive short-circuit current and conductance) by a factor of 2 to 4. In conclusion, both MT and MF disruption decrease ADH-induced apical protein secretion and Na conductance, while the ADH-induced apical Cl conductance is not significantly affected. Taken together the data support the hypothesis that the modulation of Na channel expression by apical vesicular membrane movements plays a role in Na transport expression and its regulation by ADH. [ABSTRACT FROM AUTHOR]

Electron microscopic studies of the testis were performed on 12 human embryos and fetuses between 9 and 30 weeks post conceptionem. According to their ultrastructural features, the fetal germ cells could be divided into the following three stages of differentiation: 1) gonocytes, 2) intermediate cells, and 3) fetal spermatogonia. Sertoli cells were present among the germ cells in all the testes studied. The gonocytes showed the highest nucleo-cytoplasmic ratio. Their round nucleus contained a centrally located, prominent nucleolus. The cytoplasm displayed a well developed Golgi apparatus, lipid droplets and parallel arrays of short cisternae of the rough surfaced endoplasmic reticulum (rER). Microfilaments were numerous, particularly just beneath the cell membrane. The intermediate cells were found to extend several cytoplasmic processes and to contain a moderate number of long, branched and/or widened rER cisternae which were frequently connected to the perinuclear cisterna. Intermediate cells were often connected to one another by intercellular cytoplasmic bridges. The fetal spermatogonia also displayed cytoplasmic bridges. These cells showed the lowest nucleo-cytoplasmic ratio and a more condensed nuclear chromatin. The mitochondria were situated close to the nucleus. Many of them were connected by a cementing substance. Lipid droplets and rER cisternae were rare in these cells. Infoldings of the inner nuclear membrane were often present in the gonocytes and in the intermediate cells, but were rarely observed in the fetal spermatogonia. Glycogen particles, polyribosomes, and chromatoid bodies ('nuage') were present in all the three germ cell types. With the maturation of the fetus, the number of gonocytes was found to decrease, whereas the number of fetal spermatogonia increased. The Sertoli cells also changed their ultrastructure, showing an increase in the number of rER cisternae, as well as of microfilaments, lipid droplets, and secondary lysosomes. [ABSTRACT FROM AUTHOR]

Tubular aggregates in dilated cisternae of the rough endoplasmic reticulum were observed in various cell types of two human fetuses, the mothers of which had contracted rubella early in pregnancy. The virus was isolated from several of the fetal organs. It appears that rubella, like a number of other viruses, can stimulate the formation of such aggregates. In addition, our findings indicate that these structures may form not only in adult tissues under various pathological conditions, but also in diseased embryonic and fetal cells. [ABSTRACT FROM AUTHOR]