The murine bone marrow macrophage,a sensitive indicator cell for murine migration inhibitory factor and a new method for their harvest

Murine bone marrow macrophages grown on Teflon-coated petri dishes for a period of 8–16 days can be removed with a yield of 90–95% and a viability greater than 95% following incubation in 1 mM EDTA. Bone marrow cells cultured on Teflon-coated dishes did not differ in their replication rate, peroxidase and nonspecific esterase content, pinocytosis, secretion of lysozyme and neutral proteinases from bone marrow cells cultured on plastic dishes. Murine bone marrow macrophages were found to be sensitive indicator cells for mouse migration inhibitory factor (MIF). Large numbers of cells for the MIF assay can be obtained, since their yield is 10–15 times higher than the yield of oil-induced peritoneal exudate macrophages from the same number of mice.     

Investigations on the lectin-producing cells in the sponge Axinella polypoides (Schmidt)

Summary The localization of two carbohydrate binding proteins, so-called lectins, was studied in the sponge tissue of Axinella polypoides by light and immunofluorescence microscopy. They do not occur at the cellular surface of any cell type, but they are stored in vesicles of the spherulous cells. After short formaldehyde fixation spherulous cells can be isolated and they release the active lectins upon lysis in distilled water.Electron microscopical studies of spherulous cells show that they contain almost nothing else but a small nucleus and vesicles of different size and number. Small vesicles are full of an electron dense material, whereas the content of large vesicles has a fluffy and fibrillar structure. Spherulous cells are large and tightly packed in the outer layer of the ectosome and in the mesh work of the spongin fibres of the central axis. They are small and scattered in the inner layer of the ectosome, and they are found throughout the choanosome. The function of the lectins is not clearly defined, and different alternatives such as participation in glycoprotein synthesis, immunological defense, or carbohydrate transport are possible.This study was supported by a grant from the Deutsche ForschungsgemeinschaftWe are gratefully indebted to Dr. D. Keyser for his help in our electron microscopical studies     

Preliminary studies of plasma growth hormone releasing activity during medical therapy of acromegaly.

The in vitro growth hormone releasing activity of plasma obtained from six acromegalic subjects was measured before and during therapy. In five subjects, plasmas were obtained before and during successful medical therapy with medroxyprogesterone acetate (MPA). The sixth subject was sampled before and after transphenoidal Sr90-induced hypopituitarism. All subjects had a decrement in fasting growth hormone levels with respective therapies (29-88%). The in vitro growth hormone released from Rhesus monkey anterior pituitaries was assessed after incubating one lateral half in control plasma (pre-therapy) and the contralateral pituitary half in plasma obtained during or after therapy. Studies with plasmas obtained from the five patients successfully treated with MPA showed a decreased in growth hormone releasing activity during therapy in all (18-57%). Plasma obtained after Sr90 pituitary ablation in the sixth subject had 35% more growth hormone releasing activity than obtained before therapy. These results suggest that active acromegalics who respond to MPA with significantly lowered growth hormone levels may actually achieve this response because of a decrease in growth hormone releasing factor measured peripherally. The opposite response in one acromegalic subject, following Sr90 pituitary ablation and hypopituitarism, suggests that growth hormone releasing factor secretion may increase when growth hormone levels are lowered by ablative therapy.     

Analysis of gene function of bacteriophage phi 29 of Bacillus subtilis: identification of cistrons essential for viral assembly.

Restrictive infection of Bacillus subtilis by suppressor-sensitive (sus) mutants of phi 29 has been used to search for cistrons that function in viral assembly. The products of cistrons 7, 9, 10, and 16 are necessary for head morphogenesis. The neck upper collar protein P10 and the tail protein P9 must be present for DNA packaging to occur. The protein P7 must be present for phage-related particles to form. A prohead-like particle has been isolated during 16-restrictive infection. The particle is composed of the proteins Hd, P10, F, and P7. P16 must function for DNA-filled particles to accumulate. A DNA-containing particle produced in the absence of the cistron 11 product may be an intermediate in the phi 29 assembly pathway. The protein P13 interacts with P9 and P11 to form a stable DNA-filled particle. The products of cistrons 2 and 3 are essential for viral DNA synthesis, and in their absence virus-related particles are not detected.     

On the dynamics of DNA in aqueous solution. Pulse radiolysis studies using the light scattering detection method

Summary The kinetics of DNA chain breakage in solution induced by 2 µs pulses of 15 MeV electrons were investigated by light scattering. On irradiating native calf thymus DNA at room temperature the decrease of light scattering intensity (LSI) - due to double strand ruptures - shows a fast decay with a half life _1/2 of about 30 ms as well as a slow decay with _1/2 of about 10 s. With increasing temperature (20–40° C) both the total degree of degradation and the fraction of the fast decay increase due to the facilitated melting of segments between two single strand breaks on alternate strands forming a double strand break. Above 40° C a third mode of LSI decay with _1/2 of 5–10 s arises, indicating detachment of relatively long segments.The total relative decrease of LSI after irradiation A, which can be taken as a measure of the degree of degradation, follows the square of the absorbed dose in the case of native DNA, whereas on irradiating denatured DNA A rises linearly with dose. The decay of LSI due to the degradation of denatured DNA is much faster than that of native DNA with _1/2 down to 150 µs, depending on the absorbed dose. The half lives are interpreted in terms of the separation of fragments by diffusion and of the melting of double strand segments between two single strand breaks.     

Isolation of super-repressor mutants in the histidine utilization system of Salmonella typhimurium.

Two super-repressor mutations in the histidine utilization (hut) operons of Salmonella typhimurium are described. Cells bearing either of these mutations have levels of hut enzymes that do not increase above the uninduced levels when growth is in the presence of either histidine or the gratuitous inducer imidazole propionate. Both mutations lie in the region of the gene for the hut repressor, hutC, and reverse mutations of both are to the constitutive (repressor-negative) rather than to the inducible (wild type) phenotype. In hybrid merodiploid strains the super-repressor mutations are dominant over either wild-type (hutC+) or repressor-negative (hutC-) alleles. Whereas both super-repressor mutations cause the uninducible synthesis of hut enzymes, the degree of repression is different. One mutation causes repression of enzyme synthesis in one of the two hut operons to a level below the basal, uninduced level of wild-type cells. The other mutation causes repression to a lesser degree than in wild-type cells, so that the hut enzymes are present at a level above the normal basal level; this partially constitutive synthesis is greater for the enzymes of one of the hut operons than for the enzymes of the other. Thus, both mutations apparently result in repressors with altered operator-binding properties, in addition to altered inducer-binding properties.     

Changes in DNA secondary structure afterγ-irradiation

Native calf thymus DNA was gamma-irradiated at 500 mug/ml in 0.01 M NaCl in the presence or absence of oxygen. By irradiation, an increasing amount of DNA becomes reactive with a water-soluble carbodiimide-derivative (CMEC). In the DNA sections reactive with CMEC the nucleotide strands are separated, a phenomenon previously described as radiation-induced denaturation. The dose-effect curve for the formation of denatured DNA shows an upward-bent form; a distinct oxygen effect of about 2 is observed. By a comparative study with DNA samples, degraded partially with DNAse I, it was shown that a minor part of the radiation-induced denaturation results from the formation of the radiation-induced single strand breaks, whereas the major part is a local denaturation independent of the strand breaks. In these locally denatured regions 20 to 50 nucleotide pairs are separated.     

Specific features of ex‐obese patients significantly influence the functional cell properties of adipose‐derived stromal cells

Adipose‐derived stromal cells (ADSC) are increasingly used in clinical applications due to their regenerative capabilities. However, ADSC therapies show variable results. This study analysed the effects of specific factors of ex‐obese patients on ADSC functions. ADSC were harvested from abdominal tissues (N = 20) after massive weight loss. Patients were grouped according to age, sex, current and maximum body mass index (BMI), BMI difference, weight loss method, smoking and infection at the surgical site. ADSC surface markers, viability, migration, transmigration, sprouting, differentiation potential, cytokine secretion, telomere length and mtDNA copy number were analysed. All ADSC expressed CD73, CD90, CD105, while functional properties differed significantly among patients. A high BMI difference due to massive weight loss was negatively correlated with ADSC proliferation, migration and transmigration, while age, sex or weight loss method had a smaller effect. ADSC from female and younger donors and individuals after weight loss by increase of exercise and diet change had a higher activity. Telomere length, mtDNA copy number, differentiation potential and the secretome did not correlate with patient factors or cell function. Therefore, we suggest that factors such as age, sex, increase of exercise and especially weight loss should be considered for patient selection and planning of regenerative therapies.     

Phytochemical composition of Potentilla anserina L. analyzed by an integrative GC-MS and LC-MS metabolomics platform

Potentilla anserina L. (Rosaceae) is known for its beneficial effects of prevention of pre-menstrual syndrome (PMS). For this reason P. anserina is processed into many food supplements and pharmaceutical preparations. Here we analyzed hydroalcoholic reference extracts and compared them with various extracts of different pharmacies using an integrative metabolomics platform comprising GC-MS and LC-MS analysis and software toolboxes for data alignment (MetMAX Beta 1.0) and multivariate statistical analysis (COVAIN 1.0). Multivariate statistics of the integrated GC-MS and LC-MS data showed strong differences between the different plant extract formulations. Different groups of compounds such as chlorogenic acid, kaempferol 3-O-rutinoside, acacetin 7-O-rutinoside, and genistein were reported for the first time in this species. The typical fragmentation pathway of the isoflavone genistein confirmed the identification of this active compound that was present with different abundances in all the extracts analyzed. As a result we have revealed that different extraction procedures from different vendors produce different chemical compositions, e.g. different genistein concentrations. Consequently, the treatment may have different effects. The integrative metabolomics platform provides the highest resolution of the phytochemical composition and a mean to define subtle differences in plant extract formulations.