Diaminobenzidine reactions in control and treated amoeba proteus

Summary Cytochrome oxidase activity was demonstrated in Amoeba proteus by diaminobenzidine (DAB) cytochemistry. Deposition of the reaction product occurred on the inner mitochondrial membranes and the cristae. The reaction was abolished by cyanide incubations. Positive reactions were produced with both unfixed and fixed cells: although staining potential was destroyed by any prefixatives which included glutaraldehyde. Cells prefixed with 4% formaldehyde, to raise structural preservation, retained staining ability.Amoebae subjected to prolonged anaerobiosis or to treatment with the carcinogen N-methyl-N-nitrosourethane (MNU) displayed a reduction in DAB reactivity. A positive reaction was only produced in incubations of unfixed cells and even in these the intensity of cristal staining was depleted. The possible use of DAB reactions where lesions in mitochondrial functioning have occurred is considered.     

Serum ionised calcium concentration: measurement versus calculation.

Four hundred and eighteen measurements of serum ionised calcium, total calcium, and protein concentrations were made from 47 normal volunteers, 104 patients with chronic renal failure (33 being treated conservatively and 71 with regular haemodialysis), and 83 renal transplant recipients. The serum ionised calcium concentration was measured with an Orion SS-20 meter and calculated from the total serum calcium and protein concentrations by using three formulae and a nomogram. In the normal subjects and patients undergoing regular haemodialysis, whose serum calcium concentrations were in or near the normal range, three of the calculations gave results similar to those obtained by direct measurement. In patients with conservatively treated chronic renal failure and those who had received renal transplants, however, there was poor aggrement between the methods. When patients with hypercalcaemia and hypocalcaemia from all the groups were considered separately there was again poor agreement between calculated and measured concentrations of serum ionised calcium. Of the patients whose measured concentrations of serum ionised calcium were high, 69-76% were classified as normal by the four indirect methods. We conclude that calculation of the serum ionised calcium concentrations is not an adequate substitute for direct measurement.     

Brief Communication Commercial Pha Preparations Contain Different Mitogenic Components

Three commercially available purified phytohaemagglutinin (PHA) preparations have been examined both chemically and biologically. Marked differences were observed in both composition and mitogenic effect on lymphocytes; the existence of at least two distinct mitogenic fractions in PHA was confirmed. A number of commercial preparations of PHA have become available for use as mitogens in lymphocyte cultures. Since there is no cross-standardization of biological activity between these preparations, workers have used their own discretion about which to use and whether further purification is desirable. Three commercial purified PHA preparations have been examined, and marked biological and chemical differences have been found.     

Synthesis and biological properties of 12-fluoroprostacyclins

The synthesis of the sodium salts of enantiomerically pure 12-fluoroPGI₂ (9), (±)-12-fluoroPGI₂ (9), (±)-15-epi-12-fluoroPGI₂ (10), (±)-12-fluoro-13,14-dihydroPGI₂ (11), (±)-12-fluoro-4(E)-isoPGI₂ (12), and (±)-5,6-dihydro-12-fluoroPGI₂ (13) is detailed starting from the corresponding derivatives of 12-fluoroPGF_2α methyl ester. Prostacyclins 9, (±)-9, (±)-10, (±)-11, (±)-12, and (±)-13 have been evaluated for their ability to inhibit human platelet aggregation and their effect on smooth muscle (isolated cat coronary artery).     

Formation of 6-keto-PGF1α by collecting tubule cells isolated from rabbit renal papillae

Homogeneous populations of collecting tubule epithelial cells have been isolated from rabbit renal papillae by a sequence of procedures involving: (a) dissociation of the tissue by mincing and treatment with trypsin; (b) destruction of contaminating non-collecting tubule cells by differential lysis in hypotonic media and (c) collection and washing by repeated centrifugation. The isolated cells have been characterized as being derived from the collecting tubules on the basis of anatomical source, size and histological staining for both NADH diaphorase activity and cyclooxygenase antigenicity. The cells are judged to be viable by several criteria including their ability to exclude both trypsin and vital dyes, their capacity to metabolize glucose and leucine and their ability to retain distinctive morphology following 10–14 days in culture media. Homogenates of freshly isolated collecting tubule cells when incubated with [³H]-arachidonic acid yielded radioactive products identified by thin-layer chromatographic behavior in multiple solvent systems as 6-keto-PGF_1α, PGF_2α, PGE₂, PGD₂ and a monohydroxy acid, probably HHT. No lipoxygenase-like activity was detected. At arachidonate concentrations of 2 or less, the major product was 6-keto-PGF_1α; while at substrate concentrations of greater than 10 , PGE₂ was the major radioactive prostaglandin formed. Similar distributions of products were observed when homogenates of dissociated renal papillae enriched in medullary interstitial cells were incubated with arachidonic acid. Our results indicate that collecting tubule cells do contain significant prostacyclin synthetase activity and suggest that PGI₂ plays a role in the function of mammalian collecting tubules.     

Variation in nuclear DNA in the genus Secale

Estimates of the 4C DNA amount per nucleus in 16 taxa of the genus Secale made by Feulgen microdensitometry ranged from 28.85 picograms (pg) in S. silvestre PBI R52 to 34.58 pg in S. vavilovii UM 2D49, compared with 33.14 pg in S. cereale cv. Petkus Spring which was used as a standard. Giemsa C-banding patterns showed considerable interspecific and intraspecific variation and several instances of polymorphism for large telomeric C-bands. The proportion of telomeric heterochromatin in the genome ranged from about 6% in S. silvestre and S. africanum to about 12% in cultivated rye. A detailed comparison of nine taxa showed no overall relationship between 4C DNA amount and the proportion of telomeric heterochromatin in the genome. However, evidence is presented which strongly supports the notion that the major evolutionary change in chromosome structure in Secale has involved the addition of heterochromatin at, or close to, the telomeres. It is suggested that saltatory amplification events at telomeres were initially responsible for each large increase in DNA amount. Subsequently unequal crossing over between homologues may have played an important secondary role by extending the range of variation in the amount of heterochromatin at a given telomere, while crossing over between non-homologues may have provided a useful mechanism allowing an increase in the DNA amount at one telomere to be distributed between chromosomes.     

Regenerative interactions between Drosophila imaginal discs of different types.

We have tested the ability of fragments of one type of imaginal disc to stimulate regeneration of another type. It has been shown by others that, when extreme proximal and distal fragments of the wing disc are combined, intercalary regeneration of the missing tissue ensues. Each fragment, if cultured alone, will merely duplicate its structures. We now find that distal fragments of other thoracic discs, haltere and leg, while retaining their autonomy for differentiation, also interact with proximal wing tissue to promote regeneration of more distal wing structures. The proximal wing tissue used in these experiments was the wingless abnormal wing disc which, in the absence of interaction, yields only proximal wing structures. These results suggest that spatial organization is controlled by similar systems in the various thoracic discs. In contrast, head and genital disc material provided no regenerative stimulus to the mutant wing disc tissue.