Synthesis and neuroprotective effects of serofendic acid analogues

Analogues of serofendic acid were prepared and their protective effects against L-glutamate (Glu)-induced neurotoxicity were examined using primary cultures of rat cortical neurons. Some analogues exhibited similar neuroprotective activity to that of serofendic acid.     

Properties of two growth-stimulating proteins isolated from fetal calf serum

Two proteins (S1, S2) which stimulate cell growth in cultures of embryonic rat fibroblasts, have been isolated from fetal calf serum.S1 is a protein complex consisting of an α₂-macroglobulin and insulin, asc ould be shown by immunological methods. For growth-activating activity both components are essential; when added separately to the cultures, they are ineffective.S2 has a molecular weight of about 26 000 D and does not cross-react with S1 in immunological tests.For optimal stimulation of the cultures both factors must be present together in the medium. As demonstrated by autoradiographic studies, there probably exist two different cell populations in the rat fibroblast cultures which either respond to S1 or S2.S1 and S2 strongly influence the metabolism of isolated fat cells. S1 behaves like insulin; S2 stimulates glucose oxidation and lipogenesis, but, in contrast to insulin and non-suppressible insulin-like activity, also activates the adenylate cyclase in these cells.     

The molecular and catalytic properties of galactosyltransferase from fetal calf serum

A soluble galactosyltransferase (UDP-galactose:N-acetylglucosamine galactosyl-transferase) was purified to apparent homogeneity from fetal calf serum with an overall increase in specific activity of 19,600-fold. The enzyme exhibited the following properties: specific activity, 8.5 units/mg of protein; acceptor specificity, N-acetylglucosamine/ ovalbumin = 3.3; diffusion coefficient, 5.56; sedimentation coefficient, 3.2; and molecular weight, 47,800. Comparison of the structural and catalytic properties of the fetal calf serum enzyme with purified galactosyltransferase from bovine milk indicated that the enzymes from the two bovine sources are very similar and possibly identical.     

Isolation and characterization of viruses from fetal calf serum

Summary Ten lots of specially procured fetal calf serum collected under sterile conditions and not filtered and 16 lots of commercial fetal calf serum were tested for both human and bovine viral contamination. The presence of viruses was evaluated by observing for cytopathogenic effect (CPE), hemadsorption with guinea pig erythrocytes, and interference with cytopathogenic challenge viruses in both embryonic bovine trachea (EBTr) and human diploid lung (HDL) cells. Isolates were characterized by their cytopathogenicity, morphology, serology, and ability to propagate and produce cPE in a variety of bovine and nonbovine cells. One isolate was unequivocally identified as bovine herpes virus 1, and the other was presumptively identified as a bovine virus-diarrhea virus. This study was supported by The National Institutes of Health under Contract PH 43-66-539.     

Length of the polyadenylated sequence in cytoplasmic ribonucleic acid isolated from fetal calf myoblasts differentiating in vitro

Poly(adenylic acid) [poly(A)] containing cytoplasmic ribonucleic acid (RNA) in differentiating fetal calf myoblasts cultivated in vitro was examined by hybridization with radioactive poly(uridylic acid). The size distribution of the poly(A)-containing RNA after sucrose-gradient centrifugation was similar in cells before and after differentiation. There was no apparent correlation between the length of the poly(A) segment and the change in stability of messenger RNA which occurs on differentiation, nor with the polysomal or nonpolysomal localization of the RNA in the cytoplasm. The average length of the poly(A) segments in cytoplasmic RNA in the steady state was found to be dependent on the size of the RNA: the longer the RNA, the longer the average length of the poly(A) sequence. In contrast, in pulse-labeled RNa, the length of poly(A) is similar in all size classes of RNa.     

The ability of fetal calf serum, new-born calf serum and normal steer serum to promote the in vitro development of bovine morulae

The objective of this study was to compare fetal calf serum, new-born calf serum and normal steer serum as medium supplements in the development of bovine morulae in vitro . Bovine morulae were cultured in Hams F-10 tissue culture medium (HF-10) supplemented with 5% or 10% (v/v) fetal calf serum (FCS), new-born calf serum (NBCS) or normal steer serum (NSS). Embryos were recovered at slaughter from mixed bred donor cows of mixed breeding following estrus synchronization with prostaglandin and superovulation with follicle stimulating hormone. A total of 88 morulae were recovered, washed in HF-10 + 1% Bovine Serum Albumin and randomly assigned to treatments. Embryos were cultured in microdrops of medium under paraffin oil at 37 degrees C in a 5% CO(2) humidified atmosphere. Observations for stage of development were made every 24 hours. In vitro development was analyzed by assigning to each embryo a value of 0-5 based on the most advanced stage reached (0= no development, 5= development to a hatched blastocyst). Analysis of variance of these data revealed a significant treatment effect (P<.001) while no level effect or treatment x level interaction was apparent. Comparison of treatment means by Duncans new mulitple range test showed that NSS was superior to NBCS (P<.05) which was in turn superior to FCS (P<.05) as supplements of HF-10 in promoting the in vitro development of bovine morulae.     

Albuminamide: a novel peptide amide isolated from bovine serum

During a systemic search for peptides that possess the C-terminal amide structure, a novel heptapeptide with isoleucine amide was isolated from bovine serum by sequential steps of reversed phase HPLC. Microsequence and amino acid analyses revealed the structure: Asp-Thr-His-Lys-Ser-Glu-Ile-NH2. Since this peptide has the identical sequence to N-terminal (1-7) fragment of bovine serum albumin (BSA), we have designated it albuminamide. The final HPLC step yielded 10 micrograms of homogeneous peptide preparation from 1 liter of bovine serum.     

Multiple activities of a novel substance,dictyopyrone C isolated from Dictyostelium discoideum,in cellular growth and differentiation

Summary.  We report that a novel substance named dictyopyrone C (DPC) has remarkable effects on growth and differentiation of Dictyostelium discoideum Ax-2 cells, in a dose-dependent manner. In the presence of 3–15 μM DPC, differentiation of starving Ax-2 (clone MS) cells was greatly enhanced in submerged culture, when vegetative MS cells were harvested at the mid-late-exponential growth phase (>3 × 10⁶ cells per ml) and starved. In contrast, DPC above 30 μM markedly impaired the progression of differentiation including cell aggregation, most of starved cells being round after 3–4 h of DPC application and then lysed during further incubation. In the presence of 30 μM DPC however, MS cells that had been harvested at the early exponential growth phase (<5 × 10⁵ cells per ml) and starved became neither round nor lysed and exhibited rather enhanced differentiation. Essentially the same results were obtained in cultures of starved cells on nonnutrient agar. With respect to the DPC effect on MS cells growing in axenic medium, cell lysis and growth inhibition by DPC at concentrations higher than 15 μM were realized in the mid-late-exponential-growth-phase cells (>3 × 10⁶ cells per ml) but not in the early-exponential-growth-phase cells (<5 × 10⁵ cells per ml). Moreover, analysis using synchronized MS cells has demonstrated that the DPC effect changes in a cell-cycle-dependent manner. In contrast to such unique DPC actions, the pyrone ring of DPC had no effects on growth and differentiation within the range of 3–120 μM tested. These findings strongly suggested the importance of the combined structure of the pyrone ring and the linear carbon chain in revelation of the DPC activities. Received August 5, 2002; accepted November 11, 2002; published online April 8, 2003 RID="*" ID="*" Correspondence and reprints: Department of Developmental Biology and Neurosciences, Graduate School of Life Sciences, Tohoku University, Aoba, Sendai 980-8578, Japan. E-mail: ymaeda@mail.cc.tohoku.ac.jp     

Effect of fetal calf serum on the cadmium clastogenicity

Inconsistent results among reports on cadmium genotoxicity revealed that certain confounding factors might significantly influence the outcomes of assessment. In Chinese hamster ovary (CHO-W8) cells, chromosome aberration induced by six different cadmium compounds was found positively associated with intracellular cadmium concentration. A parallel association was also observed among different CHO strains treated with same cadmium compound, the cadmium acetate. Both the cadmium-induced chromosome aberration and cadmium uptake were influenced by the presence of fetal calf serum (FCS). The presence of 10% FCS during the 2 h treatment period greatly retarded the cellular cadmium uptake, and concurrently reduced the chromosome aberration induction. Other factors such as specific cadmium anion involved and the duration of cadmium treatment period in the investigation also influenced the assessment results of cadmium-induced chromosome aberration. In the protocol with a 2 h pulse treatment, cadmium acetate, chloride and sulfate induced more chromosome aberration than cadmium nitrate, carbonate and oxide. When cadmium was present in the culture of the entire treatment period for 18 h, the results went the opposite way. Cadmium nitrate, carbonate and oxide induced significant chromosome aberration, while other three cadmium compounds gave negative results. Cadmium compounds did not induce significant SCE at the same dose level that yielded significant chromosome aberration induction, either in the protocol with the short pulse or long treatment period.     

Cryopreserved substance, isolated from human fetal liver, restore biochemical processes in acute liver failure

Recovery processes dynamics in liver when treating experimental acute hepatic insufficiency (AHI) of various etiology by using cryopreserved biopreparations, obtained from human embryo liver of different terms of development (10-12 weeks), human fetuses (22-24 week of development).     

3-Phenyllactic acid,a root-promoting substance isolated from Bokashi fertilizer,exhibits synergistic effects with tryptophan

Bokashi fertilizer, an organic fertilizer made of plant residue, has been used in Japan not only to fertilize plants but to regulate their growth. Lactic acid bacteria have been found to play an important role in the fermentation process of Bokashi, but the relationship between these bacteria and plant growth activity has not been clarified. Using the adzuki rooting assay, this study identified 3-phenyllactic acid (PLA) produced by lactic acid bacteria as a root promoting compound in Bokashi. PLA showed synergistic effect with tryptophan, but no stem elongation activity. Lactic acid bacteria produced equal quantities of the L- and D-forms of PLA, which have similar root promoting activity. PLA did not significantly affect the amount of endogenous indole-3-acetic acid (IAA), although the chemical structure of PLA is highly similar to that of L-2-aminooxy-3-phenypropionic acid (L-AOPP), which inhibits IAA biosynthesis. These results indicate that the root promoting activity of PLA is not simply due to its increase in the amount of active auxin.     

Adsorption from fetal calf serum of collagen-like proteins which bind fibronectin and promote cell attachment

Baby hamster kidney cells were seeded onto Western blots of fetal serum proteins which had been extracted from several foreign surfaces. This revealed that the major cell adhesive proteins adsorbed onto these surfaces from fetal serum were (1) fibronectin of Mr 220,000 Da and (2) vitronectin of Mr 65,000 and 78,000 Da. Two minor bands of cell attachment were observed at Mr 153,000 and Mr 134,000 Da in the fetal serum proteins extracted from heparin-agarose and serotonin-agarose. However, by exposing the Western blots of separated proteins to a second round of serum proteins, prior to cell blotting, very strong cell adhesive bands were revealed at Mr 153,000, 134,000, and 120,000 Da. By (i) modifying the composition of the serum proteins used to treat the Western blots, (ii) using specific antibodies to fibronectin, and (iii) using radiolabeled fibronectin, it was conclusively demonstrated that the new cell adhesive bands owed their increased cell attachment activity to secondary binding of fibronectin. The new bands were shown (i) to be trypsin sensitive and collagenase sensitive and therefore to be collagen-like proteins and (ii) to react negatively in immunoblots using anti-fibronectin, anti-vitronectin, anti-fibrinogen, anti-fetuin or anti-thrombospondin. In SDS-PAGE (i) the Mr 120,000-Da protein comigrated with the alpha 2-chain of Type I collagen, (ii) the Mr 134,000-Da protein comigrated with the alpha 1-chain of Type I collagen, and (iii) the Mr 153,000-Da protein comigrated with the pN-alpha 1-chain of Type III collagen. Since the novel collagen-like proteins acted as strong sites of cell attachment on nitrocellulose blots by binding fibronectin, they might well promote cell attachment on the foreign surfaces from which they were extracted.