Immunocytochemical characterization of porcine zona pellucida during follicular development

Sections of bovine ovaries fixed in Bouin's fluid or methanol-acetic acid and embedded in paraffin were incubated with chicken polyclonal antibodies to HPLC-purified zona glycoproteins ZP3 and ZP3. Oocytes of primordial follicles as well as of primary follicles showed weak labelling with anti-ZP3 and anti-ZP3. No immunostaining could be observed in the follicle cells. The ZP of primary follicles displayed distinct immunoreactivity for both ZP3 and ZP3. In secondary follicles, distinct labelling with anti-ZP3 and weak labelling with anti-ZP3 could be seen in the oocyte. The ZP showed immunoreactivity with antibodies to ZP3 and ZP3. Both antibodies labelled single follicle cells. In tertiary follicles, the oocytes were weakly labelled with anti-ZP3 and anti-ZP3. Some granulosa cells showed staining for ZP3 and ZP3. The ZP displayed strong immunoreactivity for ZP3 and ZP3. Cells of the corona radiata were strongly immunopositive for ZP3 and ZP3. Similar histotopography of immunoreactive cells could be seen in preovulatory follicles. The characteristic pattern observed for the distribution of ZP3 and ZP3 strongly suggests that in the porcine ovary both the oocyte and the follicle cells contribute to the synthesis of the ZP, perhaps in sequence.     

Distribution of renal integrin receptors and their ligands in congenital nephrotic syndrome of the finnish type

The aim of this study was to examine the distribution of 1 and v integrins (Ints) and some of their ligands in the kidneys of patients with congenital nephrotic syndrome of the Finnish type (CNF) and in controls using indirect immunofluorescence with monoclonal antibodies. The mesangial reactivity of Int 1 and Int 1 subunits was more variable and an increased glomerular reactivity with Int 3 and Int-6 antibodies was found in CNF kidneys than in controls. Int 2 subunit was either completely missing from or found in significantly lesser amounts in CNF kidney glomeruli. The immunoreactivity for Int v was more variable, fainter and also more granular in CNF samples than in control kidneys. The glomerular reactivity for Int 5 was more diffuse and weaker, and in sclerotic Bowman's capsules more intense in CNF kidneys than in controls. Immunoreactivity for Int 6 was restricted and was comparable in extent in CNF and control kidneys. Of the extracellular matrix components studied, the expression of EDAFn, EDBFn, OncFn, Ln 2 chain, Ln 1 chain and tenascin was increased. This is also seen in several glomerular diseases with inflammation and sclerosis. Immunoreactivity for vitronectin was decreased. Several differences were found in the intensity or location of the immunostaining for the 1 and v Ints and their ligands in CNF kidneys compared with controls, which have not been found in any other proteinuric disease. Disturbed Int expression pattern in CNF may specifically reflect the disturbance of glomerular function caused by the primary defect in this disease.     

Antigenic similarity between the protein neurotoxin α-bungarotoxin and neuromuscular blocking drugs

The snake neurotoxins -bungarotoxin (-BGT) and -bungarotoxin (-BGT) which act at the neuromuscular junction, were found to bind to IgE antibodies directed against neuromuscular blocking (NMB) drugs in the sera of two patients who had experienced lifethreatening anaphylactic reactions to succinylcholine. -BGT inhibited IgE-binding to a choline-Sepharose solid support in one patient better than the NMB drug alcuronium, choline, triethylcholine and -BGT. IC₅₀s for -BGT and succinylcholine were 16 and 10 nmol respectively for one patient and 34 and 6.0 nmol for the other.Recognition of the NMB drugs and -BGT by the same antibody is the first demonstration of an antigenic similarity between these drugs and the protein toxin.     

Differential effects of interleukin-1α and β on the arachidonic acid cascade in human synovial cells and chondrocytes in culture

The effects of interleukin-1 and were tested on the [³H]-arachidonic acid release and the prostaglandin synthesis by human cultured synovial cells and chondrocytes. Both forms of interleukin-1 stimulated the arachidonic acid release but interleukin-1 was more potent than IL-1. Human synovial cells and chondrocytes synthesized three types of prostaglandins upon stimulation with interleukin-1 or : prostaglandin E₂, F₂ and 6-keto-prostaglandin F₁. Regarding the synthesis of these prostaglandins, IL-1 was again more potent than IL-1. A comparison between interleukin-1-stimulated synovial cells and chondrocytes revealed neither significant quantitative nor qualitative differences in both the arachidonic acid release and the prostaglandin synthesis.     

Synoviocytes in chronic synovitis in situ and cytokine stimulated synovial cells in vitro neo-express α1, α3 and α5 chains of β1 integrins

The expression of the 1 integrins was examined immunohistochemically in synoviocytes from normal synovial membrane and from chronic synovitis of different aetiology and intensity. Normal synoviocytes were 61-positive but lacked 1 through 5. In mild inflammation type A synoviocytes neo-expressed 1, 3, and 5 chains. In severe inflammation both type A and B synoviocytes expressed 3, 4, 5, and 6 chains. The effects of inflammatory cytokines, as single agents or in combination, on the 1 integrin expression in cultured normal synoviocytes was determined by immunocytochemistry and flow cytometry. The 1 chain, while absent in unstimulated synoviocytes, was induced by interleukin-1 (IL-1), tumour necrosis factor- (TNF-), and interferon- (INF-). This effect was enhanced by combining IL-1 and TNF-. Expression of the 3 chain was up-regulated by IL-1 and, more intensely, by IFN-. Transforming growth factor (TGF-) inhibited the up-regulating effect of IL-1 and antagonized the effect of IFN- on 3 chain expression. Expression of the 5 chain was up-regulated significantly by co-stimulation through IL-1 together with TGF- or TNF-. Thus, the 1 integrin profile of cytokine activated synoviocytes in vitro resembled that of synoviocytes in synovitis in situ. These data suggest that IL-1, TNF-, IFN-, and TGF- are likely to be among the effectors regulating 1 integrin expression in synoviocytes in vivo.     

Die Nebennierenrindenfunktion unter Applikation von 9α-Fluorcortisol

Zusammenfassung Untersuchungen über den Einfluß von 9-Fluorcortisol auf die Nebennierenrindenfunktion ergaben — in Verbindung mit in der Literatur mitgeteilten Werten — eine dosisabhängige Einschränkung der Ausscheidung von Nebennierenrindenhormonen. Die Ansprechbarkeit der Nebennierenrinde auf exogenes ACTH bleibt erhalten. Es ist daher eine Hemmung der hypophysären ACTH-Sekretion anzunehmen, die durch die Struktur des synthetischen Steroids erklärbar ist. — In geringer Dosierung, wie sie als Erhaltungsdosis bei Langzeittherapie verabfolgt wird, verursacht 9-Fluorcortisol keine wesentliche Einschränkung der Hormonausscheidung.

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Effect of 9-fluorocortisol on adrenocortical function

Summary Investigation of adrenal cortical function during administration of 9-fluorcortisol revealed—in connection with results obtained from the literature—a dose-related inhibition of the secretion of adrenocortical hormones. Adrenal cortical response to exogenous ACTH remains unaffected. An inhibition of hypophyseal ACTH-secretion is therefore assumed, caused by the structure of the synthetic steroid. At low dosage, as applied in long-term treatment, no significant alteration of steroid excretion patterns was observed.

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Astonin-H, Hersteller: Fa. E. Merck A.G., Darmstadt.

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In der Arbeit werden folgende Abkürzungen verwendet: 17-KS=17-Ketosteroide; 17-OH-CS=17-Hydroxycorticosteroide; F=Cortisol=Pregn-4-en-11,17,21-triol-3,20-dion; E=Cortison=Pregn-4-en-17,21-diol-3,11,20-trion; THF=Tetrahydrocortisol=5-Pregnan-3,11,17,21-tetrol-20-on; allo-THF=allo-Tetrahydrocortisol=5-Pregnan-3,11,17,21-tetrol-20-on; THE=Tetrahydrocortison=5-Pregnan-3,17,21-triol-11,20-dion; Andro=Androsteron=5-Androstan-3-ol-17-on; Ätio=Ätiocholanolon=5-Androstan-3-ol-17-on; DHA=Dehydroepiandrosteron=Androst-5-en-3-ol-17-on.

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Herrn Prof. Dr. med. H. Franke zum 60. Geburtstag gewidmet.     

β-Subunits co-determine the sensitivity of rat neuronal nicotinic receptors to antagonists

We have investigated the effect of 4 ganglionic cholinergic antagonists (hexamethonium, mecamylamine, pentolinium, trimetaphan) on rat 32 and 34 neuronal nicotinic acetylcholine receptors (nAChRs) expressed in Xenopus oocytes. Current responses were elicited by fast application of acetylcholine on voltage-clamped oocytes (holding potential Vinh = -80mV). Concentration-inhibition curves were used to get estimates of IC₅₀, the antagonist concentration yielding 50% reduction of the peak current. The K_B's of the antagonists were calculated using estimates of the apparent K_D of acetylcholine. The order of affinity of the antagonists was similar for both receptor subtypes: mecamylamine pentolinium > hexamethonium > trimetaphan. However, 34 neuronal nAChRs were 9 to 22 times more sensitive to each of the 4 antagonists than 32 receptors. These results further underline the importance of the -subunit as co-determinant of the functional properties of neuronal nAChRs.     

Distinct mechanism of human neuroblastoma cell adhesion to fibronectin

We investigated the adhesion of three morphologically distinct human neuroblastoma cell lines (NCG, GOTO and SK-N-DZ) to intact fibronectin, central cell binding domain fragment (CBF) and CS peptide-IgG conjugates in the fibronectin molecule. Each cell line was found to express different integrin fibronectin receptors ( _{3 1}, _{4 1} and ₅₁), although similarly attached on intact fibronectin. To CBF, NCG attached well, while GOTO moderately and SK-N-DZ poorly attached. Only GOTO adhered to CS1-IgG. RGDS inhibited the spreading of NCG and SK-N-DZ on intact fibronectin, but it barely inhibited that of GOTO. The analysis by fluorescence-activated cell sorting (FACS) revealed that NCG expressed abundant ₃₁ and ₅₁, but little ₄₁, while GOTO expressed a large amount of ₄₁ as well as ₅₁. SK-N-DZ was undetectable in any of these molecules, but expressed _v1, which was identified by immunoprecipitation and immunoblotting. Polyclonal antibody to _v3 inhibited the adhesion of SK-N-DZ but not that of NCG or GOTO on intact fibronectin. These results suggest the existence of a distinct mechanism of cell adhesion to fibronectin among human neuroblastoma cell lines. It remains to be determined if such heterogeneous adhesion properties are related to the unique metastatic character of human neuroblastoma.     

Modification of the Na+ current conducted by the rat skeletal muscle α subunit by coexpression with a human brain β subunit

Sodium channels are multimeric structures composed of and subunits. Oocytes injected with RNA encoding only the subunit express voltage-gated Na⁺ currents. The kinetics of inactivation, however, are abnormal. Co-injection of rat brain and subunits modifies inactivation of I_Na such that it closely resembles endogenous currents [1]. Here we show that a subunit derived from human brain directs the same functional modification of I_Na expressed by a rat skeletal muscle subunit. This implies that functional domains for the interaction of and subunits are highly conserved across both tissues and species.     

In situ expression of β1, β3 and β4 integrin subunits in non-neoplastic endothelium and vascular tumours

Endothelial cells play an important role in adhesive interactions between circulating cells and extracellular matrix proteins. In vitro studies have shown that many of these processes are mediated by a superfamily of heterodimeric transmembrane glycoproteins called integrins. The distribution patterns of 1, 3 and 4 integrin subunits in endothelial cells (EC) in situ were examined immunohistochemically on serial forzen sections of a wide range of non-neoplastic tissues and of vascular tumours, both benign and malignant. Expression of the 1 subunit was a constitutive feature of EC. Among the 1-associated subunits, 5 and 6 were broadly distributed in EC, irrespective of vessel size and microenvironment. The 3 subunit displayed intermediate levels of expression with a slight preference for small vessel EC. Presence of 1 was confined to EC of capillaries and venules/small veins. Expression of 2 in EC was inconsistent. With rare exceptions, the 4 chain was absent in EC. The 3 and v subunits were expressed in most EC, though not always concomitantly. In contrast to the 1 chain, however, these integrin subunits were absent in EC of glomerular capillaries and were expressed variably in sinusoidal EC. The 4 chain was evenly present in the great majority of EC, except for those of large vessels. In vascular tumours, the patterns of 1 and 1 to 6 subunit expression generally corresponded to those found in their non-neoplastic counterparts. Expression of 3, v and 4 chains, however, decreased in neoplasia, especially in angiosarcomas. These data show that EC dispose of broad and at the same time differential repertoires of integrin subunits that presumably reflect vessel-type associated functional differences among these cells. In vascular tumours, the orthologous distribution patterns of 1 and 1 to 6 chains are conserved in most instances while the amounts of 3, v and 4 subunits expressed in EC tend to decrease in the course of malignant transformation.Dedicated to Prof. Dr. med. Dres. h.c. Wilhelm Doerr on the occasion of his 80th birthday