Immunohistochemistry of anterior proliferative vitreoretinopathy

Anterior proliferative vitreoretinopathy is characterized by epiretinal proliferation that extends anteriorly over the vitreous base, and may, in addition to the cells usually contributing to proliferative vitreoretinopathy, also contain cells of ocular structures located in that area. We examined 11 complete globes with aPVR that were enucleated after previous severe trauma or perforating injuries (n=8) and complicated retinal detachment (n=3) by a panel of immunohistochemical markers. We found presence of RPE, glial cells, macrophages and fibrocytes, as consistently reported in PVR membranes. In addition, T-cell lymphocytes were present in 6 of the cases, and cells expressing the common leucocyte antigen on 8 cases. Cells staining positive for the intracytoplasmic contractile filament-smooth muscle actin were present in 5 cases and cells staining for desmin in one case. Collagen type IV was part of most of the membranes, and vessels with leakage of plasma factors were present in more than half of the cases.This paper was presented in part at the First Annual Meeting of the European Community Ophthalmic Research Association (ECORA), Bonn, Germany, 6.10.1993.     

H3 receptor-mediated inhibition of noradrenaline release: an investigation into the involvement of Ca2+ and K+ ions,G protein and adenylate cyclase

The effects of ₂-adrenoceptor agonists (dexmedetomidine, oxymetazoline), alone or in combination with various -adrenoceptor subtype-selective antagonists (CH-38083, idazoxan, WB4101, BRL44408, ARC-239, prazosin), on noradrenaline- and isoprenaline-induced lipolysis were investigated in human isolated abdominal subcutaneous fat cells. The rank order of potency of antagonists in preventing dexmedetomidine- and oxymetazoline-evoked suppression of isoprenaline-induced lipolysis was (pA₂-values): CH-38083 (7.69 and 7.48) idazoxan (7.5 and 7.41) > BRL 44408 (7.23 and 7.19) WB 4101 (7.13 and 7.12) > prazosin (5.18 and 5.17) > ARC-239 (4.72, 4.9). While CH-38083 and idazoxan, non-subtype selective ₂-adrenoceptor antagonists and BRL44408, a selective a_2A-adrenoceptor antagonist as well as WB4101 potentiated the lipolytic effect of noradrenaline, ARC-239, the selective _2B-adrenoceptor antagonist failed to affect it. In addition since the _2A-adrenoceptor selective agonist, oxymetazoline concentration dependently inhibited the lipolytic effect of isoprenaline, and WB4101 and BRL44408 (a_2A-adrenoceptor antagonists) antagonised the effect of oxymetazoline in a competitive manner, it is concluded that the a_2A-adrenoceptor subtype is involved in antilipolysis. In addition, functional evidence was obtained that there is an interaction between _2A- and -adrenoceptors located on the cell surface of adipocytes, through which locally released noradrenaline and/or circulating circulating adrenaline influence lipolysis.On leave from Institute of Medical Pharmacology, University of Pisa, Via Roma 55, I-5626 Pisa, Italy Correspondence to: E. S. Vizi at the above address     

Signal transduction pathway of the muscarinic receptors mediating gallbladder contraction

In gallbladder smooth muscle, carbachol interacts with M₃ receptors to mediate contraction. To examine components of the intracellular second messenger system that is coupled to these receptors we have tested whether carbachol stimulates the formation of inositol phosphates (IP) to cause contraction. Guinea pig gallbladder muscle strips were prelabeled with [³H]inositol and were incubated with 0.1 mmol/l carbachol, a concentration causing maximal contraction. [³H]inositol monophosphates, [³H]inositol bisphosphates and [³H]inositol trisphosphates and contraction were measured at various times (0–90 s). To examine whether a pertussis toxin-sensitive guanine nucleotide binding protein is coupled to the muscarinic receptors, guinea pigs were pretreated with pertussis toxin (180 g/kg i.v./24 h). The effectiveness of pertussis toxin treatment was determined by measuring [³²P]ADP-ribosylation of a –40/41 kDa protein from gallbladder homogenates. Carbachol caused a significant time-dependent increase in the formation of [³H]inositol monophosphates, [³H]inositol bisphosphates and [³H]inositol trisphosphates. The time course of [³H]inositol trisphosphate turnover caused by carbachol was biphasic, and was detectable at 15 s and maximal at 60 s; at 75 s and 90 s formation of [³H]inositol trisphosphates decreased, whereas the time course of carbachol-induced contraction of the gallbladder smooth muscle strips reached a plateau after 90 s. The effects of carbachol on [³H]inositol trisphosphates and on contraction were abolished by atropine. Pretreatment with pertussis toxin resulted in ADP-ribosylation of a 40/41 kDa protein from gallbladder cell membranes but did not affect the concentration-response or time course of carbachol-induced contraction. These results indicate that carbachol-induced contraction of gallbladder smooth muscle cells is accompanied by the activation of inositol phosphate turnover and does not involve a pertussis toxin-sensitive G-protein.This article is based in part on the doctoral thesis of Burkhard Mackensen at the Faculty of Medicine, University of Hamburg, Germany. Some of the results were presented at the meeting of the American Gastroenterological Association (AGA) in San Francisco 1992 (von Schrenck et al. 1992) Correspondence to: T. von Schrenck at the above address     

Pharmacokinetics and bioavailability of benperidol in schizophrenic patients after intravenous and two different kinds of oral application

Pharmacokinetics and bioavailability of benperidol were determined in 13 schizophrenic patients after acute administration of 6 mg benperidol as an intravenous (i.v.) bolus injection, orally as liquid, and orally as tablets using a partially randomized cross-over design. Drug plasma levels were determined by high performance liquid chromatography with electrochemical detection and subjected to model independent pharmacokinetic analyses. After i.v. dosing the geometric means (mean-g) were 3.2 min for the distribution half-life, 5.80 h for the elimination half-life (t _1/2), 4.21 l/kg for the distribution volume, 7.50 h for the mean residence time (MRT), and 0.50 l/(h*kg) for the clearance. After oral administration as liquid and as tablet mean-g data for the time lag until the first appearance of measurable plasma concentrations were 0.33 and 1.1 h, mean-g t _1/2 values were 5.5 and 4.7 h, respectively, mean-g t _max data were 1.0 h and 2.7 h, mean-g MRT values were 8.44 and 8.84 h, and mean-g C _max ^maxvalues were 10.2 and 7.3 ng/ml. Differences between liquid and tablet administration were statistically significant for time lag,t _max, andC _max. Mean-g absolute bioavailabilities were computed as 48.6% after liquid and 40.2% after tablet administration respectively. All parameters studied exhibited large intersubject variation. The plasma concentrations of the presumed metabolite reduced benperidol were found to be very low.     

How to improve histopathological results in the biopsy diagnosis of gut dysganglionosis

In a methodological survey, the technical prerequisites for optimal histopathological diagnosis of gut dysganglionosis are presented. To make a proper diagnosis, the pediatric surgeon or gastroenterologist and the pathologist must consider certain preconditions. The most important steps for the optimal biopsy diagnosis of an aganglionosis, an ultrashort Hirschsprung segment, a intestinal neuronal dysganglionosis (IND), a ganglioneuromatosis, a hypogenesis, or immaturity of the vegetative gut innervation are: (1) taking 3–4 biopsies the size of a peppercorn (3–5 mm³) with submucosa; (2) the best instruments for taking rectal mucosal biopsies are forceps and scissors or a conventional large biopsy forceps; and (3) biopsies may be taken 1 cm, 3–4 cm, 6–9 cm, and 9–12 cm (or from a preternatural anus) above the pectinate line. A biopsy containing mucosa, muscularis mucosae, and submucosa guarantees a satisfying histopathological diagnosis. The native biopsies can be transported on water-ice if the distance to the pathologist takes no longer than 4–6 h. For long distances, biopsies have to be frozen on dry ice (CO₂ –80 °C) and transported in a sufficient amount of dry ice (adapted to the time of transportation). For biopsy processing, the following points are important: a total of 122 to 160 15-m-thick native cryostat serial sections have to be cut per biopsy and distributed on four microscope slides. Forty sections are used for lactic dehydrogenase reactions, 32 for succinic dehydrogenase reactions, and the rest for an acetylcholinesterase (AChE) reaction. An AChE reaction alone is sufficient for the diagnosis of Hirschsprung's disease (HD), but never for IND or other developmental malformations of the submucous and myenteric plexuses. Enzymehistotopochemical reactions allow the assessment of functional parameters. These reactions, in contrast to immunohistochemical staining, offer information about the functional activity of special gut structures, e. g., increased AChE activity in nerve fibers of the rectal wall in HD or a lack of dehydrogenase activity in immature ganglia.     

Interferon gamma and interleukin 10 levels in preimplantation embryo culture media

Purpose The aim of our study is to elucidate whether human oocyteslembryos secrete IFN and/or IL-10 and whether the fertilization process depends on the balance between these cytokines.Methods A total of 142 embryo culture media from 24 patients were collected and the cytokine levels were tested with ELISA.Results IFN and IL-10 were detectable in 40.1% and 29.6% of culture media respectively. The difference of IFN and IL-10 levels in media from fertilized oocytes between day 1 and day 2 are significant (0.46 vs. 1.47 and 34.2 vs. 12.7, respectively). However there was no significant difference between the IFN levels of the media from fertilized and nonfertilized oocytes 0.46 vs. 0.85 at day 1 and 1.47 vs. 1.49 at day 2, as well as IL-10 levels 34.2 vs. 30.9 at day 1 and 12.7 vs. 9.58 at day 2 respectively.Conclusions Human preimplantation embryos secrete the cytokines IFN and IL-10. No effect of these cytokines on fertilization process could be shown.Presented at the IXth World Congress on In Vitro Fertilization and Alternate Assisted Reproduction, April 3–7, 1995, Vienna, Austria.     

CD44: physiological expression of distinct isoforms as evidence for organ-specific metastasis formation

Continuous progress has been achieved during recent decades in the therapy of metastasizing malignancies by improving chemotherapeutic strategies and new approaches in radiation therapy. Genetic manipulation of tumor cells and of the tumor fighting immune system is hoped to add significant contributions to curative interventions in disseminated tumors. That we are still far from eradicating death by malignant growth is due ultimately to our limited understanding of the cascade of events resulting in metastasis formation, which until recently was believed to rely on multiple rounds of mutation and selection processes. This implies an individually specific history of each metastatic tumor, which would rule out uniform diagnostic and therapeutic concepts. When it was noted in a rat tumor model that the transfer of cDNA of a single gene, a CD44 variant isoform (CD44v) covering the exons v4–v7, sufficed to initiate metastasis formation of a locally growing tumor, hope was created that a metastogene may have been identified. Although the idea of CD44v expression as a unifying concept for tumor progression was not sustained, the discovery of CD44v-initiated metastatic spread allowed a conceptually new hypothesis on tumor progression as a consequence of the reactivation of genetic programs of ontogeny, stem cell differentiation, and/or lymphocyte activation. Since distinct CD44 isoforms play an important role in these processes, unraveling the functions of this family of molecules can indeed provide a cornerstone in the understanding of tumor progression. This article summarizes briefly the present knowledge on known functions of CD44 isoforms with particular focus on parallels between physiological programs and tumor progression.Abbreviations CD44s CD44 standard isoform - CD44v CD44 variant isoforms - HA Hyaluronate