Biphasic action of phospholipase A in collagen-stimulated rat platelets

The formation of thromboxane A2 (TXA2) in collagen-stimulated rat platelets was successfully divided into two stages, an initial and a second one, by the specific TXA2 receptor antagonist, ONO3708. In the presence of this antagonist, only the initial TXA2 production was observed, without the subsequent platelet shape change and aggregation. Collagen causes the specific cleavage of arachidonic acid from phosphatidylinositol (PI) in the initial stage, whereas in the absence of the antagonist, it caused decrease in the arachidonic acid levels in phosphatidylethanolamine (PE) and PI with concomitant formation of the respective lyso-forms. These results demonstrate that phospholipase A (PLA) preferentially acts on PI to release arachidonic acid which leads to the initial TXA2 production, which might be a trigger for the second release of arachidonic acid from PE and PI.     

Isolation, purification, and characterization of the pellicle of Euglena gracilis z

The pellicle was isolated from the cell homogenate obtained on sonication of Euglena gracilis z grown aerobically under illumination and purified by a combination of differential and sucrose density gradient centrifugations. The purity and homogeneity of the pellicle fragments were determined by an electron microscopic method and biochemical analysis of the components. The protein, lipid, and sugar contents of the purified pellicle were 68.7, 17.9, and 13.5%, respectively. The equilibrium density of pellicle fragments was 1.21 g/cm3. SDS-polyacrylamide gel electrophoresis revealed that the pellicle contained 50 mol% of nonpolar amino acids. The constituents of the lipid and sugar were very different from those of the cell membrane of other organisms.     

Purification and characterization of aquacobalamin reductase (NADPH) from Euglena gracilis

Euglena aquacobalamin reductase (NADPH: EC 1.6.99.-) was purified, and its subcellular distribution was studied to elucidate the mechanism of the conversion of hydroxocobalamin to 5'-deoxyadenosylcobalamin. The enzyme was found in the mitochondria. It was purified about 150-fold over the Euglena mitochondrial extract in a yield of 38%. The purified enzyme was homogeneous in polyacrylamide gel electrophoresis. Spectra of the purified enzyme showed that it was a flavoprotein. The molecular weight of the enzyme was calculated to be 66,000 by Sephadex G-100 gel filtration and 65,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme was specific to NADPH with an apparent Km of 43 microM and to hydroxocobalamin with an apparent Km of 55 microM. The enzyme did not require FAD or FMN as a cofactor. The optimum pH and temperature were 7.0 and 40 degrees C, respectively.     

A case of von Recklinghausen's disease associated with pheochromocytoma and papillary carcinoma of the thyroid gland

A 58-year-old woman was admitted to our hospital complaining of headache, dizziness and intermittent elevation of blood pressure. Multiple café-au-lait spots and neurofibromas had appeared on the back and the limbs since the age of 30 years. At the age of 54 years she underwent total thyroidectomy because of papillary carcinoma of the thyroid gland. On admission, the levels of plasma norepinephrine and epinephrine, urinary norepinephrine and normetanephrine were all within the normal range. However, urinary excretion of metanephrine was markedly increased to 1.49 +/- 0.45 (Mean +/- SD) mg/day and that of epinephrine was also slightly increased. The computed tomographic scans of the abdomen and the scintigraphy with 131I-metaiodobenzylguanidine revealed a tumor mass in the region of the right adrenal gland. The tumor was histologically confirmed to be pheochromocytoma at the operation. In her family history, her mother and one of her two sisters had von Recklinghausen's disease and another sister suffered from follicular carcinoma of the thyroid gland. As far as we know, this paper is the first report of a patient with von Recklinghausen's disease associated with both pheochromocytoma and non-medullary carcinoma of the thyroid gland, and her family.     

The highly modified membrane of cornified cells in stratified squamous epithelia: A comparison of heterogenous deposits in keratinized and nonkeratinized epithelia

Summary The chemical nature of the thickened plasma membrane of cornified cells in stratified squamous epithelium was investigated in comparison with that in noncornified epithelium. Localizations of transglutaminase, molecular weight 92000 daltons, and detection of epidermal cysteine proteinase inhibitor were effected with a monoclonal antibody and a monospecific rabbit anti-inhibitor immunoglobulin, respectively, directed to the antigens. N-(7-dimethylamino-4-methylcoumarinyl) maleimide was used to demonstrate S-S cross-linking. In all keratinizing epithelia, the enzyme and inhibitor were deposited on membranes of granular cells. S-S bonds were formed in cornification with the appearance of electron-dense material by the inner leaflet. Both enzyme and inhibitors occurred on the corneal epithelium, but S-S linkage and the thickened plasma membrane did not form even at the last stage of maturation. On the other hand, the internal vaginal epithelium in the proestrous stage without keratinization contained the enzyme, but neither inhibitor nor S-S linkage. Both antigens and S-S bonds were detected when keratinization proceeded during estrus. The staining patterns in the epithelium near the vaginal introitus were identical to those in the skin. Cuboidal and simple epithelia exhibited none of those constituents. The findings indicated that heterogenous components contribute to modification of the plasma membrane of cornified cells, but S-S cross-linkages are associated exclusively with formation of the ultrastructurally unique membrane structure. In addition, findings suggested hormonal regulation in the chemical modification of the membrane in estrogen-sensitive internal vaginal epithelium.     

The different effects of recombinant human tumor necrosis factor on rat fibrosarcoma sublines

Summary The antitumor effect of recombinant human tumor necrosis factor (rH-TNF) on two clones of rat fibrosarcoma with different metastatic potential to lymph nodes was examined. The colony formation of clone A, which has high metastatic potential, was completely inhibited by continuous exposure to rH-TNF at 50 U/ml. In contrast, colony formation of clone G, which has low metastatic potential, was not inhibited by high concentrations of rH-TNF (10,000 U/ml). The inhibitory effect of rH-TNF on colony formation by clone A was also observed with a 1-h exposure to rH-TNF. This effect was time and concentration dependent, as determined by the colony assay, ³H-thymidine uptake assay, and ⁵¹Cr-release assay. ³H-thymidine and ³H-uridine uptake per cell of clone A exposed to rH-TNF was not decreased. This suggests that the mechanisms of the antitumor effect of rH-TNF were not due to inhibition of DNA and RNA synthesis of tumor cells. In vivo growth and lymph node metastases of clone A inoculated i.p. to Donryu strain rats were completely suppressed by 14 consecutive i.p. injections of 10⁵ or 10⁶ U/kg per day of rH-TNF. On the other hand the growth of clone G was not influenced by rH-TNF administration.     

Protective role of potentially lethal damage repair in the neoplastic transformation of Balb/c 3T3 cells treated with N-methyl-N'-nitro-N-nitrosoguanidine

To determine the role of repair of potentially lethal damage (PLD) in the initiation process of neoplastic transformation, Balb/c 3T3 cells treated with N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) were temporarily exposed to conditioned medium obtained from density-inhibited Chinese hamster cell cultures, as a post-treatment for the induction of PLD repair. With or without this exposure, cell survival and transformation frequencies were simultaneously determined by colony-formation and focus-formation assays, respectively. Temporary exposure to conditioned medium resulted in a 20-30% increase in cell survival compared with no exposure. Post-treatment with conditioned medium resulted in a 60-65% reduction in transformation frequencies. At the molecular level, the repair of MNNG-induced single-strand breaks of DNA occurred much more rapidly in conditioned medium. These data suggest that PLD repair reduces the in vitro neoplastic transformation through excision repair operative during the cessation of DNA replication. Thus, PLD repair appears to be preventive against neoplastic fixation in initiation of neoplastic development.     

Scanning electron microscopic observation on the distal part of the male urethra of the mouse

In the proximal end of the navicular fossa, the stratified columnar epithelium lining the spongy part of the urethra abruptly changes into the stratified squamous one, presenting a sharp demarcation. It is considered that the junction between these 2 epithelia corresponds to the former site of the junction of the entoderm and ectoderm. The width of the microridges in the distal part of the navicular fossa was about doubled compared to that in the proximal part. It is suggested that the epithelium in the distal part is keratinized, while that in the proximal part is nonkeratinized.     

Effect of histamine H1-receptor antagonist on the regulation of carbohydrate and lipid metabolism in rat liver

In order to elucidate the role of histamine in the liver, we studied the effect of a histamine H1-receptor antagonist on the carbohydrate and lipid metabolism in the rat liver. The administration of the H1-receptor antagonist decreased significantly the contents of glycogen and malonyl-CoA in the liver. However, it did not affect the levels of serum glucose and free fatty acid. These results suggest that histamine may play a part in the regulation of metabolism of carbohydrates and lipids in the liver.     

Prolactin secretion and its response to stress during the estrous cycle of the rats

Serum and pituitary prolactin (PRL) concentrations were measured during the estrous cycle of the rat with particular attention to the afternoons of the days of proestrus and estrus. Homogenizing machines, a Polytron and Sonifier, were used to extract PRL from the pituitary gland. The effects of ether anesthesia and restraint were also examined on the afternoons of both proestrus and estrus. The occurrence of a surge in PRL secretion during proestrus was confirmed with a peak at 1500 h, and this was accompanied by a decline in pituitary PRL content. A relatively high level of serum PRL was observed in the afternoon of estrus, during which time pituitary PRL content increased progressively. Ether anesthesia had no effect on the proestrus PRL surge, while restraint enhanced it. On the afternoon of estrus, restraint completely suppressed the rise in serum PRL, but ether anesthesia failed to suppress it completely. From these results, the following conclusions were drawn: 1) the PRL surge on the afternoon of proestrus occurs without synthesis of the hormone in the pituitary; 2) PRL secretion on the afternoon of estrus is accompanied by its synthesis in the gland; 3) the PRL response is distinct for each type of stress applied; and 4) PRL secretion is thus regulated by different mechanisms in proestrus and estrus.