Seminal prostaglandins in men with subnormal sperm motility and therapeutic treatment

The contents of prostaglandins in seminal plasma from a total of 73 men were evaluated. The subjects were grouped as follows: normospermic men, patients with impaired motility, patients with small untreated varicocelle and patients with impaired motility and Kallikrein therapy. Sperm density, morphology and motility were examined. High performance reversed phase liquid chromatography (HPLC) in combination with specific radioimmunoassays were used for the determination of PGE₂, PGI₂ and PGF_2α. There was a significant difference (p < 0, 025; F-test) between the PGI₂ concentrations in patients with impaired motility (5,6 ± 1,4 pg/mg protein) and normal men (8,8 ± 3,7 pg/mg protein). PGE₂ and PGF_2α were significantly different in patients with varicocele (p < 0,025, F-test). Wide ranges of prostaglandins occured in the Kallikrein-group with no significant differences. We conclude that: a) PGI₁ is an additional prostaglandin compound in seminal plasma. b)its measurement may not be useful as diagnostic parameter in subfertile men and c) Kallikrein has no influence on the prostaglandin content in seminal plasma and other seminal parameters as motility, motility index and sperm counts.     

Alcaligenes eutrophus CH34 is a facultative chemolithotroph with plasmid-bound resistance to heavy metals.

Alcaligenes eutrophus strain CH34, which was isolated as a bacterium resistant to cobalt, zinc, and cadmium ions, shares with A. eutrophus strain H16 the ability to grow lithoautotrophically on molecular hydrogen, to form a cytoplasmic NAD-reducing and a membrane-bound hydrogenase, and most metabolic attributes; however, it does not grow on fructose. Strain CH34 contains two plasmids, pMOL28 (163 kilobases) specifying nickel, mercury, and cobalt resistance and pMOL30 (238 kilobases) specifying zinc, cadmium, mercury, and cobalt resistance. The plasmids are self-transmissible in homologous matings, but at low frequencies. The transfer frequency was strongly increased with IncP1 plasmids RP4 and pUZ8 as helper plasmids. The phenotypes of the wild type, cured strains, and transconjugants are characterized by the following MICs (Micromolar) in strains with the indicated phenotypes: Nic+, 2.5; Nic-, 0.6; Cob+A, 5.0; Cob+B, 20.0; Cob-, less than 0.07; Zin+, 12.0; Zin-, 0.6; Cad+, 2.5; and Cad-, 0.6. Plasmid-free cells of strain CH34 are still able to grow lithoautotrophically and to form both hydrogenases, indicating that the hydrogenase genes are located on the chromosome, in contrast to the Hox structural genes of strain H16, which are located on the megaplasmid pHG1 (450 kilobases).     

Action of neurotensin on size, composition, and growth of pancreas and stomach in the rat

Since the gastrointestinal peptide neurotensin has a stimulatory effect on the secretion of the exocrine pancreas and an inhibitory effect on secretion and motility of the stomach, we investigated whether chronic parenteral administration of neurotensin would affect pancreatic and gastric growth. We therefore infused synthetic neurotensin subcutaneously (dose, 43 and 282 pmol X kg-1 X min-1) in 20 Wistar rats for 2 weeks using Alzet osmotic minipumps and compared pancreatic weight, DNA, RNA, protein, lipase, amylase, pancreatic polypeptide and insulin with these parameters in 10 control rats from the same litter with subcutaneously implanted plastic cylinders approximately the size of the minipumps. In another experiment, synthetic neurotensin (836 pmol X kg-1) was injected intraperitoneally three times a day for 3 days in 12 rats. Thereafter, we measured pancreatic DNA and in vitro incorporation of [3H]thymidine into pancreatic DNA. These effects were compared with the actions of caerulein and normal saline. Long term infusion of the high neurotensin dose induced an increase of pancreatic weight (control: 0.87 g, neurotensin: 1.02 g) and of DNA (control: 2.5 micrograms; neurotensin: 3.5 micrograms) and pancreatic polypeptide (control: 2.4 ng; neurotensin: 7.4 ng) contents, whereas pancreatic protein, RNA, amylase and lipase contents were not stimulated. In relation to DNA, these parameters even were significantly depressed. Insulin remained unchanged. Intraperitoneal injection of neurotensin induced an increase of pancreatic DNA content and stimulated [3H]thymidine incorporation into DNA (control: 11 000 dpm/g; neurotensin: 15 800 dpm/g pancreas). Moreover, long-term neurotensin infusion with the high dose led to a rise in protein concentration and an increase in the thickness of the gastric antrum; antral DNA concentration was insignificantly stimulated. Parenteral neurotensin in the doses and at the times administered, led therefore, to hyperplasia of the pancreas and induced growth of the gastric antrum. It is concluded that neurotensin can act as a trophic factor on pancreas and gastric antrum of the rat. It remains to be determined whether this represents a physiological effect of neurotensin.     

Late enzymes of vindoline biosynthesis

From differentiated plants of Catharanthus roseus (L.) G. Don we have isolated a specific enzyme of the vindoline biosynthetic pathway catalysing the S-adenosylmethionine-dependent methylation of 11-O-demethyl-17-O-deacetyl-vindoline. The enzyme we named S-adenosyl-L-methionine : 11-O-demethyl-17-O-deacetylvindoline 11-O-methyltransferase. This transferase exhibits a high substrate specificity. Obviously the O-methylation at C-11 precedes the O-acetylation at the C-17 position during the biosynthesis of vindoline.A second enzyme was detected which hydrolyses the acetyl function of vindoline. The distribution of this acetylesterase in C. roseus plants demonstrates that the enzyme is not specifically associated with the vindoline distribution in the plant material. Most probably this enzyme plays no essential role in the biosynthesis of vindoline.     

Late enzymes of vindoline biosynthesis. Acetyl-CoA: 17-O-deacetylvindoline 17-O-acetyl-transferase

From differentiated plants of Catharanthus roseus (L.) G. Don, a specific enzyme was isolated and named acetyl-CoA : 17-O-deacetylvindoline 17-O-acetyltransferase, acting on the biosynthetic formation of the Aspidosperma type alkaloid vindoline.The enzyme shows a high selectivity towards different substrates. The acetyl-CoA-dependent transferase also catalyses the reverse reaction by hydrolysis of the 17-O-acetyl group of vindoline in the presence of free CoA. This enzyme is localized only in vindoline-containing plant parts, but was so far not detectable in cell suspension cultures of C. roseus. The enzyme allows the synthesis of labelled vindoline with high specific activity, applicable for instance as tracer for radioimmunoassays of vindoline.     

Disruption of phospholipid asymmetry in erythrocyte vesicles deficient in spectrin

The transbilayer distribution of phospholipids in right-side-out and inside-out vesicles derived from human erythrocytes was studied by phospholipase A2 digestion assays and by staining with the fluorescent dye merocyanine 540. In both types of vesicles, the normal asymmetric distribution of phospholipids characteristic of intact cells was disrupted. Because both types of vesicles are deficient in spectrin, the major protein of the cytoskeletal network which normally underlies the membrane, these results support the contention that spectrin is involved in the maintenance of phospholipid asymmetry.     

The persistence of micronucleated erythrocytes in the peripheral circulation of normal and splenectomized Fischer 344 rats: implications for cytogenetic screening

Micronucleated erythrocytes are selectively removed from the peripheral circulation of normal rats. Splenectomy prevents this selective removal. In normal rats treated daily for 20 days with 0.2 mg/kg triethylenemelamine (TEM), micronucleated normochromatic (mature) erythrocytes did not accumulate in peripheral blood. In these same animals, the frequencies of micronucleated cells among polychromatic (newly formed) erythrocytes increased from 0.21 to 5.25 per thousand in peripheral blood and from 1.75 to 31.5 per thousand in bone marrow. Since both control and induced frequencies in peripheral blood were approximately 15% of those in bone marrow, the removal appears to be equally efficient for cells containing either spontaneously occurring or clastogen-induced micronuclei. In splenectomized rats treated daily for 11 days with 0.2 mg/kg TEM, the frequency of micronucleated normochromatic erythrocytes (NCEs) in the peripheral blood rose rapidly to 9 times the control value and remained elevated for 50-55 days, indicating a life span approximately equivalent to that of normal erythrocytes. Among splenectomized rats exposed to either 0.15 mg/kg triethylenemelamine, 6.5 mg/kg cyclophosphamide, or 300 mg/kg urethane for periods exceeding the erythrocyte life span, the incidences of micronucleated NCEs in the peripheral blood rose steadily from a control value of 1.0 per thousand to maximum values of 15.0, 12.7 and 8.9 per thousand, respectively. During these extended exposures, the mean frequencies of micronucleated polychromatic erythrocytes (PCEs) in peripheral blood increased from a spontaneous value of 0.9 per thousand to 23.0, 13.0 and 6.6 per thousand, respectively, reflecting the frequencies among PCEs in the bone marrow and approximating the maximum values among NCEs in the peripheral blood. Thus, the frequency of micronucleated erythrocytes in the peripheral blood of splenectomized rats can be used as an index of both acute and cumulative chromosomal damage, while in normal rats the use of peripheral blood for cytogenetic monitoring is restricted by the selective removal of these micronucleated cells.     

Characterization of a native subunit of the NAD-linked hydrogenase isolated from a mutant of Alcaligenes eutrophus H16

The cytoplasmic, NAD-linked hydrogenase of Alcaligenes eutrophus H16 consists of four non-identical subunits. From the mutant strain HF14, defective in this enzyme, a protein was isolated that reacted with specific antibodies raised against the wild-type hydrogenase; the reaction type was of partial identity. The same protein was also tested with specific antibodies raised against each of the four denatured subunits of the wild-type hydrogenase and was found to be completely identical with the second largest subunit; it reacted weakly with the antibody against the largest subunit and not at all with the antibody against the small subunits. In SDS-polyacrylamide gel electrophoresis the protein of the mutant migrated as a single polypeptide and corresponded to the second largest subunit of soluble hydrogenase with Mr = 56,000. The mutant enzyme strongly differed from the wild-type hydrogenase in its binding behaviour to chromatographic gels. It had pronounced hydrophobic properties and bound strongly to phenyl-Sepharose; it had high affinity to triazin dye gels. Enzymatically the polypeptide was totally inactive with NAD as electron acceptor, but showed weak residual activities with methylene blue, ferricyanide and cytochrome c. The protein also contained nickel; however, because of the instability of this polypeptide the amount varied between 0.2-1.4 nickel per enzyme molecule. As shown by ESR studies, the iron is organized in a [4Fe-4S] cluster but is partially present also in the 3Fe-form. No nickel signal could be detected. The role of the polypeptide subunit for hydrogen activation in the intact hydrogenase is discussed.     

The dopaminergic innervation of the goldfish pituitary

Summary The dopaminergic innervation of the goldfish pituitary gland was studied by immunocytochemistry at the electron-microscope level using highly specific antibodies against dopamine coupled to bovine serum albumin with glutaraldehyde. A satisfactory preservation of the tissue was achieved after immersion in 5% glutaraldehyde in phosphate buffer containing sodium metabisulfite to prevent oxidation of the endogenous dopamine. The immunocyto-chemical procedure was performed on Vibratome sections using the preembedding method. Immunoreactivity was restricted to part of the neurosecretory type-B fibers (diameter of the secretory vesicles lower than 100 nm) in which it was found to occupy the whole cytoplasm. Labeled fibers were observed within the neurohypophysis in the different parts of the gland and in the adenohypophyseal tissue where immunoreactive profiles were detected in close apposition to the different cell types. These data are in agreement with previous results obtained by means of radioautography and further support a role for dopamine in the neuroendocrine regulation of pituitary functions in teleosts.     

On the Phospholipid Metabolism of Glial Cell Primary Cultures.

Abstract: Primary cultures were prepared from newborn rat brain. After 16-18 days, they consisted mainly of mature and immature astrocytes and oligodendrocytes, as judged by immunohistochemistry. To study the metabolism of ethanolamine glycerophospholipids, the cells were incubated with 1-[1-³H]alkyl- sn -glycero-3-phosphoethanolamine (1-alkyl-GPE), for 1–20 h. Five main products were formed: 1-alkyl-2-acyl-GPE; 1-alkyl-2-acyksn-glycero-3-phosphocholine (1-alkyl-2-acyl-GPC); 1-alkenyl-2-acyl-GPE (ethanolamine plasmalogen); 1-alkenyl-2-acyl-GPC (choline plasmalogen); and 1-alkyl-glycerol. Acylation of the substrate was the main reaction during the first 3 h of incubation, whereas desaturation to plasmaiogen reached a maximum after 12 h. Greater amounts of radioactivity were observed in the phosphatidylcholine fraction after longer incubation times. Only small amounts of choline plasmalogen were observed. The phosphatidylethanolamine fraction consisted of 26.5% diacyl-, 27.5% alkyl-acyl-, and 46.0% alkenyl-acyl- compounds, whereas the corresponding data for the phosphatidylcholine fraction were 78.5, 16.4, and 5.1%, respectively, after 20 h of incubation. Hydrolysis of the substrate to 1-alkyl-glycerol was a minor reaction.