A comparison of the isoelectric points of mouse liver UDP-glucuronosyltransferase enzymes conjugating the twelve benzo[a]pyrene phenols

Solubilized mouse liver microsomes were subjected to chromatofocusing using a pH 9.5 to 6.0 gradient. UDP-glucuronosyltransferase activity was assayed using 12 benzo[a]pyrene phenols as substrates. The rank of microsomal activity for the phenols was as follows: 12 > 10 > 4 > 1 > 7 > 5 > 8 > 9 > 3 > 11 > 6 > 2. Fractions separated on chromatofocusing according to isoelectric point indicated that 3-, 10-, 11-, and 12-hydroxybenzo[a]pyrene were conjugated primarily by a high pI (～8.5) activity(s), 2-, 6-, 8-, and 9-hydroxybenzo[a]pyrene were conjugated primarily by a low pI (～6.7) activity(s), and 1-, 4-, 5-, and 7-hydroxybenzo[a]pyrene were conjugated equally well by high and low pI forms.     

Physiological and morphological acid resistance ofCandida boidinii

The growth ofCandida boidinii strain 2 in a methanol-limited chemostat at a dilution rate of 0.1/h and a low extracellular pH (2.8–4.0) is characterized by a maximum yield coefficient referred to the methanol consumedY _S of 0.4 g/g and a maximum cell content of nitrogenous compounds of 60%. The cell proteins are rich in essential amino acids. At pH<2.6 or >4.0 the cell concentration decreases due to lower growth rate, accompanied by increased metabolic quotientsQ _S,Q _CO2 andQ _form, and increased activities of dissimilating dehydrogenases. The activity of alcohol oxidase (AO) in intact cells (0.54 IU/mg protein) was unaffected by pH 2.8–3.8 although in a cell-free extract the AO activity decreased at these low pH values after a 10-min incubation. The lower AO activity in cells at pH<2.8 and pH>3.8 brought about increased residual methanol levels in the medium, and also an increased level of riboflavin phosphate, arising probably by the release of FAD from active AO. Catalase activity was completely pH-independent. Cell morphology also showed no changes at pH 2.8–4.2, formation of cell chains being observed only at pH<2.8. However, the ultrastructure of cells grown in the chemostat at pH 2.6, however, did not evince any changes as compared with cells grown, at higher pH apart from a lag in cytokinesis. These findings, which point to acid resistance of strain 2, make it possible to produce biomass from methanol, with a high content of valuable proteins and AO, under nonsterile conditions.     

The proliferation of mouse mammary epithelial cells in response to specific mitogens is modulated by the mammary fat pad in vitro

Summary The ability of the murine mammary fat pad to directly stimulate the growth of mammary epithelial cells and to modulate the effects of various mammogenic agents has been investigated in a newly described, hormone- and serum-free coculture system. COMMA-1D mouse mammary epithelial cells were cultured for 5 or 7 d with various supplements in the absence or presence of epithelium-free mammary fat pad explants from virgin female BALB/c mice. Cocultured fat pad stimulated increases in the DNA content of COMMA-1D cultures by two- to threefold or six-to eightfold after 5 or 7 d, respectively. The mitogenic effect was additive to that of 10% fetal calf serum and could not be attributed to the release of prostaglandin E₂ or synthesis of prostaglandins by epithelial cells. In addition, bovine serum albumin attenuated (P<0.05) the mitogenic effect of cocultured mammary fat pad. Added alone, insulinlike growth factor-I, epidermal growth factor, and insulin increased (P<0.05) total DNA of COMMA-1D cultures by 2.5-, 3.7-, and 2.3-fold, respectively. Cocultured mammary fat pad markedly interacted (P<0.01) with these mitogens to yield final DNA values that were 21.2-, 13.3-, and 22.1-fold greater than in basal medium only. Associated with this proliferation was the formation of numerous domes above the COMMA-1D monolayer. There was no proliferative response to growth hormone or prolactin in the absence or presence of cocultured fat pad (P>0.05). Whereas hydrocortisone did not alter cell number, it attenuated (P<0.05) the mitogenic effect of cocultured mammary fat pad. These results indicate that the murine mammary fat pad is not only a direct source of mitogenic activity, but also modulates the response of mammary epithelial cells to certain mammogens.     

Polymorphism identification in GDF9 gene and its association analysis with reproduction traits in Jinghai Yellow chicken

Abstract

GDF9 (growth differentiation factor 9) belongs to the transforming growth factor-β (TGF-β) superfamily and plays an irreplaceable role in female fertility. To reveal its genetic effects on productivity performance in chickens, 373 Jinghai Yellow chickens were chosen randomly to detect SNPs in GDF9 by PCR-SSCP and DNA sequencing methods. Eventually, four SNPs (g.2053G?>?A, g.2275T?>?C, g.2338C?>?T, g.2420T?>?C) in total had been detected. Amongst them, g.2420T?>?C was first found significantly associated with reproduction trait in chickens and heterozygous type C₂T₂ had higher average egg weight at 300?days of age (AEWD300) than T₂T₂ (p?<?0.01). Least squares analysis showed that age at first laying (AFE) of H1 and H1H1 chickens were significantly earlier than that of H7 and H7H7 ones, respectively (p?<?0.05). H1H5 hens showed higher AEWD300 than H4H7 ones (p?<?0.05). For total egg number at 300?days of age (END300), mean of H5H5 was significantly higher than that of H4H4 (p?<?0.05). Hence, the study suggested that hybrid vigor at g.2420T?>?C could be utilized in practice. H1H1, H1H5 and H5H5 could be the dominant diplotypes for chicken breeding. The study may contribute to the breeding progress of productive chickens and supply reference for oviparous animal production practice.     

Liver-Associated Natural Killer Activity in Cirrhotic Rats

An impaired host defense mechanism is well known in patients with liver cirrhosis (LC). Using a sinusoidal lavage method, lymphocytes were obtained from LC rats that were administered thioacetamide, and natural killer (NK) activity was measured by ^5lCr-release assay. The NK cell count was measured by flow cytometric analysis using monoclonal antibody (Mab) 3.2.3 and/or CD 3-8+ as markers for NK cells, and by immunohistochemical staining using Mab 3.2.3. Furthermore, interferon (IFN) α was administered to LC rats and the subsequent changes in hepatic NK activity and NK cell count were observed. In the large granular lymphocyte (LGL)-rich fraction (Fr.1, LGLs: 60-90%), the NK activity was significantly lower in the LC rats (40.0±3.8%) compared to that in the control rats (48.4±4.3%) (P<0.005). In addition, the number of NK cells in the liver tissues of the LC rats was significantly lower compared to that in the liver tissues of the control rats by morphometric analysis (P<0.05). For LC rats, NK activity of the Fr.1 24 hr after IFNα administration (5×10⁴ IU / 100 g body weight) increased significantly (P<0.005). Hepatic NK activity and NK cell count were reduced in the LC rats, and recovered following IFNα administration. The results obtained in this study may give clues to better understanding the impaired host defense mechanism in LC patients.     

Characterization of Water-Soluble Complexes of Polyacrylic Acid with α-Amylase from <Emphasis Type="Italic">Aspergillus oryzae</Emphasis>

Polycomplex formation of α-Amylase from Aspergillus oryzae (TAKA) with polyacrylic acid (PAA) was studied by pH titration, fluorescence, and high performance liquid chromatography (HPLC) methods in water solutions. Acording to the our results, the complex formation and its solubility were depended on nature of enzyme and the pH of solutions. Both of them correlates isoelectric points (PI). The stability of PAA–amylase complexes was negligibly weak at pH 7 [pH > pI (isoelectric pH)]. Stable water-soluble polycomplexes were formed at pH 5 (pI ~4.5) and coexisted with free protein molecules. Insoluble complexes has been observed at pH < 4.5. The frozen storage stabilities of the obtained complexes were also studied by measuring the activities at different pH.     

Effect of yeast inoculation rate on the metabolism of contaminating lactobacilli during fermentation of corn mash

Two separate 4 (bacterial concentrations)×6 (yeast concentrations) full factorial experiments were conducted in an attempt to identify a novel approach to minimize the effects caused by bacterial contamination during industrial production of ethanol from corn. Lactobacillus plantarum and Lactobacillus paracasei, commonly occurring bacterial contaminants in ethanol plants, were used in separate fermentation experiments conducted in duplicate using an industrial strain of Saccharomyces cerevisiae, Allyeast Superstart. Bacterial concentrations were 0, 1×10⁶, 1×10⁷ and 1×10⁸ cells/ml mash. Yeast concentrations were 0, 1×10⁶, 1×10⁷, 2×10⁷, 3×10⁷, and 4×10⁷ cells/ml mash. An increased yeast inoculation rate of 3×10⁷ cells/ml resulted in a greater than 80% decrease (P<0.001) and a greater than 55% decrease (P<0.001) in lactic acid production by L. plantarum and L. paracasei, respectively, when mash was infected with 1×10⁸ lactobacilli/ml. No differences (P>0.25) were observed in the final ethanol concentration produced by yeast at any of the inoculation rates studied, in the absence of lactobacilli. However, when the mash was infected with 1×10⁷ or 1×10⁸ lactobacilli/ml, a reduction of 0.7–0.9% v/v (P<0.005) and a reduction of 0.4–0.6% v/v (P<0.005) in the final ethanol produced was observed in mashes inoculated with 1×10⁶ and 1×10⁷ yeast cells/ml, respectively. At higher yeast inoculation rates of 3×10⁷ or 4×10⁷ cells/ml, no differences (P>0.35) were observed in the final ethanol produced even when the mash was infected with 1×10⁸ lactobacilli/ml. The increase in ethanol corresponded to the reduction in lactic acid production by lactobacilli. This suggests that using an inoculation rate of 3×10⁷ yeast cells/ml reduces the growth and metabolism of contaminating lactic bacteria significantly, which results in reduced lactic acid production and a concomitant increase in ethanol production by yeast.     

Clearance rates of sessile rotifers: in vitro determinations

We measured laboratory clearance rates of 10 rotifer and one unidentified bryozoan species from 3 different lakes using ³²P labeled algae (Chlamydomonas) or yeast (Rhodotorula). Clearance rates for all rotifers fed yeast ranged from < 2.0 to > 260 µl · animal^–1 · h^–1 depending on species. The in vitro clearance rates of two sessile rotifers (Ptygura crystallina and P. pilula) were not significantly different from previously measured in situ rates (Wallace and Starkweather 1983). Clearance rates for 5 rotifers fed algae ranged from < 5.0 to > 90.0 µl · animal^–1 · h^–1. Ptygura beauchampi, P. crystallina, P. pilula, Floscularia conifera, and F. melicerta ingested both cell types but their clearance rates varied substantially among species and between cell types. There was a substantial time-dependent loss of 32P from formalin-fixed animals (Sinantherina socialis) awaiting processing. This loss stabilized at approximately 20 hours and was estimated to be about 40% of the initial ingested label. Clearance rates for the bryozoan fed yeast or algae were highly variable, ranging from < 1.0 to > 3 000 µl · animal^–1 · h^–1.     

The seasonal abundance and vertical distribution of the <3-μm phytoplakton in the north basin of Lake Biwa

The seasonal changes in the size-fractionated chlorophylla concentrations (<3 μm, 3 to 25 μm, and >25 μm) were investigated at a pelagic site of the north basin of Lake Biwa during June to December 1985. Autofluorescing plankton cells in the <3-μm fractions were also examined using the fluorescein isothiocyanate staining epifluorescence microscopic technique. The <3-μm phytoplankton (usually dominated by chroococcoid cyanobacteria except for a few cases dominated by small eukaryotes) showed a clearly different pattern of seasonal change compared with the larger fractions. That is, from August to early September, chlorophylla of the larger fractions declined considerably, while the <3-μm chlorophylla did not decrease significantly. Moreover, cyanobacterial cell density in the <3-μm fraction showed a maximum value (2–3.5×10⁵ cells·ml⁻¹) during this period. The relative contribution of the <3-μm chlorophylla to the total chlorophylla increased from <5% to 45% during the course of this change. No clear vertical trend in the distribution and composition of the <3-μm phytoplankton was found, except that relatively large cyanobacteria (>4 μm³) appeared at a depth of 15m but not at 0,5 and 10 m from late July to August. These large cells were also found in November and December. The drastic seasonal change of phytoplankton size structure occurring in this basin was discussed in relation to grazing, nutrient depletion and sinking. Contribution from Otsu Hydrobiological Station, Kyoto Univeristy (No. 308, foreign language series).     

Arsenic Uptake and Accumulation in Rice (Oryza sativa L.) at Different Growth Stages following Soil Incorporation of Roxarsone and Arsanilic Acid

Experiments were conducted with rice (Oryza sativa L.) by adding 0, 10, 20, 30, 40, 50 mg kg^-1 of arsenic (As) to soil (with roxarsone and arsanilic acid, presented as As concentrations) at a field with an isolation chamber. The aims were to evaluate the effects of As- (roxarsone or arsanilic acid) contaminated soil on rice agronomic parameters and uptake of As in different plant parts of the rice plant. The results showed that As (roxarsone or arsanilic acid) could significantly reduce plant height, effective tiller number, straw weight and grain yield (P < 0.01). As concentrations in different parts of the plant varied with the growth stages, and behaved similarly. At the maturing stage, the level in different parts peaked in all treatments, with tissue As concentrations showing the pattern: root > leaf > stem > husk > grain. In addition, at the mature stage, the As concentrations in different parts of the rice plant increased with increasing concentrations of roxarsone and arsanilic acid. The highest concentration of As found in grain was 0.82 mg kg^-1, which did not exceed the statutory permissible limit for rice grain (1.0 mg As kg^-1), and in the leaf and stem it was approximately 6.0 mg kg^-1, which was significantly higher than that in the controls. The results showed that rice could accumulate As from contaminated soil (roxarsone or arsanilic acid), which may be transferred to human beings via the food chain.     

Cloning and functional expression of the d-β-hydroxybutyrate dehydrogenase gene of Rhodobacter sp. DSMZ 12077

Nucleotide sequence and biochemical analysis of d-β-hydroxybutyrate dehydrogenase (EC 1.1.1.30), isolated from Rhodobacter sp., indicate functional oligomers composed of subunits of 257 amino acids with a calculated M _r of 26,800 and a pI of 5.90. Compared to mammalian short-chain alcohol dehydrogenases, the bacterial enzyme lacks a C-terminal lipid anchor domain and was found to be highly active upon expression in Escherichia coli even without lipid supplement. The recombinant enzyme could be highly enriched using a single chromatography step and was shown to be stable over a broad range of pH and temperature. Received: 1 April 1999 / Received last revision: 11 June 1999 / Accepted: 11 June 1999     

Amino Acid Substitution of the Largest Subunit of Yeast RNA Polymerase II: Effect of a Temperature-Sensitive Mutation Related to G1 Cell Cycle Arrest

A mammalian temperature-sensitive mutant tsAF8 shows cell cycle arrest at nonpermissive temperatures in mid-G₁ phase. DNA sequence comparison of the largest subunit of RNA polymerase II (Rpb1) from the wild-type and the mutant shows that the mutant phenotype results from a (hemizygous) C-to-A variation at nucleotide 944 in one rpb1 allele, giving rise to an Ala-to-Asp substitution at residue 315 in the protein. This amino acid substitution was introduced into the Schizosaccharomyces pombe rpb1 gene. Whereas tsAF8 cells showed growth defects and altered Rpb1 distribution at nonpermissive temperatures, yeast cells harboring this amino acid substitution did not show apparent temperature sensitivity. The effect of another temperature-sensitive Rpb1 mutation was also small. These results suggest that mutation of the rpb1 gene, which is critical in mammalian cells, may not be deleterious in yeast cells. RID= ID= <E5>Correspondence to: </E5>K. Sugaya; <E5>email:</E5> k_sugaya&commat;nirs.go.jp Received: 2 September 2002 / Accepted: 7 October 2002     

Use of preparative isoelectric focusing in a Sephadex gel slab to separate immunizing and growth facilitating moieties in crude 3 M KCl extracts of a murine fibrosarcoma

Summary Crude 3 M KCl extracts of the methylcholanthrene-induced fibrosarcoma of C3H/HeJ mice, MCA-F, were demonstrated to contain two fractions, one inducing tumor resistance and the other facilitating the outgrowth of neoplastic cell challenge. In immunoprotection tests in syngeneic C3H/HeJ mice, optimal doses of crude solubilized tumor antigen afforded only a 28% reduction in growth compared with saline-treated controls. When crude extracts were fractionated by preparative isoelectric focusing (pIEF) in a slab of superfine Sephadex G-75, significant biologic activity was demonstrated in two fractions. Fraction (Fr) 1, pI 2.5–3.6, induced potent tumor facilitation, increasing the tumor size by more than 100%, while Fr 15, pI 5.8–6.0, engendered resistance that reduced their respective biological effects to MCA-F, but not the antigenically unrelated MCA-D tumor. Thus 3 M KCl extracts contain at least two biologically active components, one immunoprotective and one tumor-facilitating. Since the weak immunoprotective activity of crude materials may represent the vectorial effect of these antagonistic components, subsequent molecular characterization of both moieties may afford insight into the complex response of hosts toward tumors. Furthermore, TSTA purified by the rapid method of isoelectric focusing may be a more suitable reagent for immunotherapy than the parent crude 3 M KCl extracts by virtue of the absence of facilitating antigens.Abbreviations CE crude 3 M KCl extract - pIEF preparative isoelectric focusing - Fr fraction from pIEF - MCA-F and MCA-D antigenically different methylcholanthrene-induced fibrosarcomas of C3H/HeJ mice - TSTA tumor specific transplantation antigens     

A combined treatment of ionomycin with ethanol improves blastocyst development of bovine oocytes harvested from stored ovaries and microinjected with spermatozoa

Regardless of the presence of sperm-borne oocyte-activating factors, activation of bovine oocytes with exogenous activation stimuli is required for further development after intracytoplasmic sperm injection (ICSI). The current study was designed to develop a new activation regimen for improving the blastocyst yield after ICSI of bovine oocytes harvested from ovaries stored at 10 to 12 °C for 24 h. After ICSI, oocytes were treated with 5 μM ionomycin for 5 min, 7% ethanol for 5 or 10 min, ionomycin followed by ethanol (5 or 10 min), ionomycin followed by 10 μg/mL cycloheximide for 5 h, or ionomycin followed by 1.9 mM 6-dimethylaminopurine for 3 h. Across the activation regimens, the cleavage rates of ICSI oocytes (45% to 77%) were higher than those of parthenogenetically activated oocytes (11% to 21%; P < 0.05). Activating the ICSI oocytes with ionomycin plus ethanol improved the blastocyst yield (29% to 30%) compared with that of nontreated oocytes (12%; P < 0.05), but the other regimens did not improve the blastocyst yield (9% to 18%; P > 0.05). Higher blastocyst yields were due to increasing the proportion of ICSI oocytes that passed through the early postfertilization events until cleavage. None of the regimens have any adverse effect on the quality of the blastocysts regarding the total cell number or the proportion of the inner cell mass cells. Thus, a new activation regimen using two triggers for single calcium increase effectively improved blastocyst yield after bovine ICSI using oocytes harvested from stored ovaries.     

Effects of doxorubicin on human dental pulp cells in vitro

There is substantial information concerning the effects of continuous exposure to supratherapeutic or therapeutic concentrations of doxorubicin on human molar pulpal cells; the effects of continuous exposure to subtherapeutic concentrations of this agent are undetermined. To this end, we studied the proliferation of human fibroblasts and pulpal cells and their pattern of mineralized nodule deposition in vitro. Cell proliferation was assessed at 1, 3, 5, and 7 days from populations with either no exposure (control) or exposure to 10⁻⁶–10⁻⁹ mol/L doxorubicin. Mineralized nodule deposition and calcium-45 incorporation were assessed at 7 and 21 days of culture. Data were compared by factorial ANOVA and a post-hoc Tukey test. 10⁻⁶ and 10⁻⁷ mol/L doxorubicin significantly reduced the total number of viable pulpal cells in cultures from days 1 to 3 (p < 0.05); doxorubicin 10⁻⁶–10⁻⁹ mol/L significantly inhibited cell proliferation (p < 0.05) and DNA synthesis 5 days after plating (p < 0.001). After 21 days, doxorubicin 10⁻⁶–10⁻⁸ mol/L significantly decreased calcium-45 incorporation into pulpal cultures (p < 0.001); all dilutions significantly reduced the number of mineralized nodules within the 21-day pulpal cultures (p < 0.05). In addition, all dilutions of doxorubicin significantly inhibited fibroblast cell proliferation and incorporation of [³H]thymidine. In contrast, the fibroblast cultures did not produce mineralized nodules, suggesting that the mineralized nodules within the pulpal cell cultures did not result from dystrophic calcification. Thus, exposure to subtheraputic doxorubicin concentrations has potential adverse effects on mineralized tissue formation within the pulp, which could affect the rates of reparative dentin deposition within the tooth pulps of patients receiving this chemotherapeutic agent.     

Relationship between substrate inhibition and maintenance energy ofChlamydomonas reinhardtii in heterotrophic culture

The relationship between substrate inhibition and maintenance energy ofChlamydomonas reinhardtii grown heterotrophically on acetate was investigated. At low acetate concentrations (<0.4 g l^–1), where no inhibition of cell growth was observed, the cell growth yield and specific growth rate could be represented by the Pirt model, 1/Y=1/Y _g +m/ with a constant value of maintenance energy coefficient m. However, at high acetate concentrations (>0.4 g l^–1), inhibition of cell growth occurred, in which m became variable and dependent on the acetate concentration. A simple mathematical model was proposed to predict the actual maintenance energy coefficient m in the inhibited cultures and experimentally validated.Author for correspondence     

Improved yields of the extracellular domain of the epidermal growth factor receptor produced using the baculovirus expression system by medium replacement following infection

The extracellular domain of the epidermal growth factor receptor (EGFR) was expressed using the baculovirus expression vector system. The maximum level of the EGFR extracellular domain secreted into the medium in Sf-9 (Spodoptera frugiperda or fall armyworm) cell batch culture was approximately 2.5 g ml^–1. In order to increase this yield, a process was developed that included the following sequence of steps: batch growth to maximum cell density, infection of the cells with recombinant virus, and replacement of spent medium. By using this process, the specific yield of recombinant protein, which in batch culture drops when infection is carried out at densities greater than 3 × 10⁶ cells ml^–1, can be maintained at a maximum in cultures infected at densities of 10⁷ cells ml^–1 or greater. The process, when applied to 3-1 and 11-1 bioreactor cultures, allowed a maximum volumetric yield of triple the maximum value attainable in batch culture. Spent-medium analysis indicates that medium replacement provides certain nutrients that could otherwise be limiting for recombinant protein production.     

Preliminary assessment of protein associated with airborne particles in Mexico City

The protein associated with airborne particles was measured during 1991 as an indicator of airborne biological material in different outdoor urban environments. Fifty air samples were collected simultaneously at three sampling sites, located in the north, south and downtown Mexico City, using a PM10 high-volume sampler (particles<10 μm). The air filters were weighed and protein extracted using a phosphate buffer. Protein concentrations were determined by Lowry assay. The extracts were also analysed by SDS electrophoresis and IEF using a Phastsystem. High concentrations of airborne particles were recorded at the sampling sites with a geometric mean of 70.2 μg/m³ in the south (residential area), 95.5 μg/m³ in the center (urban-commercial area), and with the highest value of 108.9 μg/m³ in the north (urban-industrial area). No statistically significant difference (P>0.05) was observed among the protein concentrations from the sampling sites and the concentrations ranged from non-detectable to 2.54 μg/m³. However, the protein concentrations presented significant difference (P<0.05) with respect to rainy and dry seasons. The Spearman correlation coefficient between protein concentration and airborne particles concentration was statistically significant (r=0.50). The molecular weights (MW) and isoelectric points (pI) for the proteins present in some of the extracts were determined. The values ranged from approximately 8000 to 106 000 Da and the pI values from nearly 4.0 to 5.85. This is important because the major allergens from inhalants are mostly acidic proteins with molecular weights in the range of 20 000–40 000 Da.     

In vitro development and nutrient uptake by embryos derived from oocytes of pre‐pubertal and adult cows

The present study compared the developmental potential and uptake of nutrients by embryos from pre‐pubertal and adult cows. Oocytes retrieved from ovaries of 5 to 7 month old calves and adult cows were matured and fertilized in vitro. Embryos were cultured in SOFaa to the blastocyst stage (7 days post‐insemination). At successive stages of development, rates of glucose and pyruvate uptake were measured non‐invasively by microfluorescence for individual embryos. Fertilization was equivalent in embryos from pre‐pubertal and adult cows (P > 0.05), however development to blastocyst was significantly lower in embryos from pre‐pubertal cows (9.8% versus 33.7%, respectively; P < 0.05). Total blastocyst cell number was not different between pre‐pubertal and adult material (P > 0.05). Glucose uptake was exponential (pre‐pubertal, r = 0.82; adult, r = 0.82; P < 0.05), with an increase in uptake beyond the 8‐ to 16‐cell stage. Glucose uptake was significantly lower in embryos from pre‐pubertal cows at the 2‐ to 4‐cell stages (1.5 versus 3.0 pmoles/embryo/hr; P < 0.05), but was equivalent to the adult cow at all other stages of development (P > 0.05). Pyruvate uptake was low until the blastocyst stage. Pyruvate uptake by embryos from pre‐pubertal cows was significantly different to adult cows at the 1‐cell stage (2.7 versus 4.6 pmoles/embryo/hr, respectively; P < 0.05) and 2‐ to 4‐cell stages (4.9 versus 3.6 pmoles/embryo/hr, respectively; P < 0.05). Pyruvate uptake was equivalent in the two groups in the later stages of development (P > 0.05). Perturbations in the uptake of nutrients by embryos from pre‐pubertal cows were most likely due to the presence of a high proportion of developmentally incompetent embryos. Further, embryos from pre‐pubertal cows that did develop to the blastocyst were as viable as blastocysts from adult cows with respect to nutrient uptakes and total cell number. Mol. Reprod. Dev. 54:49–56, 1999. © 1999 Wiley‐Liss, Inc.     

INFLUENCE OF LOW-FREQUENCY ELECTRIC FIELDS ON PROLIFERATION AND IL-8 RELEASE OF HL-60 CELLS

The influence of capacitively coupled extremely low-frequency (ELF) electric fields on proliferation and on interleukin (IL)-8 release of exponentially growing HL-60 cells was examined. The cell suspensions were treated with the field component of interferential current (IFC) using different exposure protocols. Modulation frequencies of 10 and 100 Hz were applied with field strengths between 0.075 and 11.54 V_pp/cm for 48 hr using a 5-min exposure time at every 3 hr. At a field strength of 1 V_pp/cm, the influence of the time between two exposure sessions was examined for different modulation frequencies. All exposure protocols applied have no effect on cell proliferation (p>0.05), but statistical significant reduction (p<0.05) of the IL-8 release at selected modulation frequencies and interval times could be observed.