In situ localization of pea enation mosaic virus double-stranded ribonucleic acid

Precursor ([³H]uridine) incorporation studies in the presence or absence of actinomycin D (AMD) showed that AMD-insensitive nucleic acid synthesis occurred only in the nuclei in pea enation mosaic virus (PEMV)-infected pea plant tissues. Ferritin-labeled antibody studies showed that ds-RNA was present only in nuclei. In vitro hybridization of the nucleic acid from various infected or healthy cell fractions with [¹²⁵I]PEMV-ss-RNA indicated that PEMV-ds-RNA was primarily associated with nuclei-enriched fractions from infected cells.An in situ hybridization technique, which utilized autoradiography to detect the subcellular location of material which hybridized with [¹²⁵I]PEMV-ss-RNA, was developed. This method confirmed that PEMV-ds-RNA was localized in the nuclei of PEMV-infected tissues.     

3种钙激动剂促培养的大鼠血管平滑肌细胞增殖

目的：探讨不同来源的细胞内钙离子（[Ca²⁺]i）对血管平滑肌细胞（vascular smooth muscle cells, VSMCs）丝裂素活化蛋白激酶（MAPK）介导的增殖反应的作用。方法：以培养的大鼠VSMCs为靶细胞,用血管紧张素II（Ang Ⅱ）剌激VSMCs跨膜Ca²⁺内流、三磷酸肌醇（IP₃）和雷尼丁（RY）剌激胞内Ca²⁺释放,[γ-³² P]-ATP掺入法和免疫印迹（Western blot）测MAPK活性及蛋白含量,氚-亮氨酸（[³H]-Leu）、氚-胸腺嘧啶（[³H]-TdR）掺入量作为VSMCs增殖的指标。结果：Ang Ⅱ、IP₃和RY均能显著增加VSMCs的[Ca²⁺]i浓度、MAPK活性及蛋白含量,并提高[³H]-Leu、[³H]-TdR掺入量,与对照组VSMCs相比差异显著（P<0.01）。结论：钙激动剂诱导的MAPK活性及含量的增加参与了VSMCs的增殖,VSMCs的肥大增殖与[Ca²⁺]i浓度增加有关,而与[Ca²⁺]i的来源无关。     

Comparison of the Sensititre YeastOne dilution method with the Clinical Laboratory Standards Institute (CLSI) M27-A3 microbroth dilution reference method for determining MIC of eight antifungal agents on 102 yeast strains

The Clinical Laboratory Standards Institute ([CLSI] formerly NCCLS) reference broth microdilution testing method (protocol M27-A3) was compared with a commercially available methods (Sensititre YeastOne^®) by testing two quality control strains and 102 isolates of Candida sp. and Cryptococcus sp. against fluconazole, itraconazole, ketoconazole, posaconazole, voriconazole, flucytosin, amphotericin B and caspofungin. Minimal inhibitory concentrations (MIC) endpoints were determined after 24 h of incubation for Sensititre YeastOne^® and after 24 and 48 h for CLSI microdilution method. Essential agreements between methods vary from 70.6 to 92.2%. Categorical agreements vary from 94.1% for 5FC to 72.6% for AMB. Sensititre YeastOne^® reading appears to be useful for avoiding very major errors and this confirms the interest of this method for evaluating new antifungals activity in vitro.     

Threshold increases in plasma growth hormone in relation to plasma catecholamine and blood lactate concentrations during progressive exercise in endurance-trained athletes

Plasma human growth hormone ([HGH]), adrenaline ([A]), noradrenaline ([NA]) and blood lactate ([La^–]_b) concentrations were measured during progressive, multistage exercise on a cycle ergometer in 12 endurance-trained athletes [aged 32.0 (SEM 2.0) years]. Exercise intensities (3 min each) were increased by 50 W until the subjects felt exhausted. Venous blood samples were taken after each intensity. The [HGH] and catecholamine concentrations increased negligibly during exercise of low to moderate intensities revealing an abrupt rise at the load corresponding to the lactate threshold ([La^–]-T). Close correlations (P < 0.001) were found between [La^–]_b and plasma [HGH] (r = 0.64), [A] (r = 0.71) and [NA] (r = 0.81). The mean threshold exercise intensities for [HGH], [A] and [NA], detected by log-log transformation, [154 (SEM 19) W, 162 (SEM 15) W and 160 (SEM 17) W, respectively] were not significantly different from the [La^–]-T [161 (SEM 12) W]. The results indicated that the threshold rise in plasma [HGH] followed the patterns of plasma catecholamine and blood lactate accumulation during progressive exercise in the endurancetrained athletes.     

Local and systemic effects on blood lactate concentration during exercise with small and large muscle groups

To evaluate the relationship between lactate release and [lac]_art and to investigate the influence of the catecholamines on the lactate release, 14 healthy men [age 25±3 (SE) year] were studied by superimposing cycle on forearm exercise, both at 65% of their maximal power reached in respective incremental tests. Handgrip exercise was performed for 30 min at 65% of peak power. In addition, between the tenth and the 22nd minute, cycling with the same intensity was superimposed. The increase in venous lactate concentration ([lac]_ven) (rest: 1.3±0.4 mmol·l⁻¹; 3rd min: 3.9±0.8 mmol·l⁻¹) begins with the forearm exercise, whereas arterial lactate concentration ([lac]_art) remains almost unchanged. Once cycling has been added to forearm exercise (COMB), [lac]_art increases with a concomitant increase in [lac]_ven (12th min: [lac]_art, 3.2±1.3 mmol·l⁻¹; [lac]_ven, 5.7±2.2 mmol·l⁻¹). A correlation between oxygen tension (P_vO₂) and [lac]_ven cannot be detected. There is a significant correlation between [lac]_art and norepinephrine ([NE]) (y=0.25x+1.2; r=0.815; p<0.01) but no correlation between lactate release and epinephrine ([EPI]) at moderate intensity. Our main conclusion is that lactate release from exercising muscles at moderate intensities is neither dependent on P_vO₂ nor on [EPI] in the blood.     

Effect of contraction on lymphatic,venous, and tissue electrolytes and metabolites in rabbit skeletal muscle

Summary The effect of muscle contraction on lymphatic and plasma [K⁺], [Na⁺], [Ca²⁺], [Mg²⁺], [Cl^–], [P_i], [lactate] ([Lac^–]); [creatine] ([Cr]), ideal osmolality (OSM), and [protein] was evaluated in femoral venous blood and lymph specimens sampled from the calf muscles of rabbits before, in the course of, and after contractions. In addition, total [K⁺], [Na⁺], [Ca²⁺], [Mg²⁺], [Cl^–], and [H₂O] were analyzed in the muscle tissue. To facilitate lymph sampling both hind limbs were passively flexed and extended, in imitation of natural running movements, by an electrically driven crank. The muscles of one side also performed superimposed rhythmic isotonic contractions. Before contractions, lymphatic [K⁺], [Na⁺], [Ca²⁺], [Mg²⁺], [Lac^–], [Cr], and OSM did not significantly differ from corresponding femoral venous concentrations, [Cl^–], and [P_i] were significantly higher, [protein] significantly lower in the lymph than in the plasma. During contractions lymphatic [K⁺], OSM, [Lac^–], and [P_i] were raised significantly more in the lymph compared with the plasma concentrations. [Na⁺], [Cl^–], [Ca²⁺], and [Mg²⁺] showed only small changes in the course of contractions and thereafter, and they were altered in a similar way in the lymph and plasma. It was suggested that lymphatic and interstitial concentrations were in equilibrium. Comparing inactive with active muscles, the latter lost K⁺ but gained Na⁺, Cl^–, and H₂O, whereas minimal changes occurred in the [Ca²⁺] and [Mg²⁺]. The changes were discussed in connection with the hypothesis that electrolyte shifts might be involved in the activation of the muscular non-proprioceptive interstitial nerve endings which appear to play a role in reflexogenic cardiovascular and respiratory control.A preliminary report of this work has been given elsewhere [33]Supported by Deutsche Forschungsgemeinschaft     

Changes in plasma volume and red cell formation after a marathon competition

Summary The purpose of this study was to investigate middle-term influences of a marathon race on plasma volume (PV) and red cell production. We performed the following measurements in the blood of 15 male athletes: haemoglobin ([Hb]), haematocrit (Hct), plasma protein concentration ([Prot]), plasma osmolality, sodium concentration ([Na⁺]), potassium concentration ([K⁺]), aldosterone concentration ([Aldo]), haptoglobin concentration ([Hpto]), and the reticulocyte count, as well as the calculation of relative changes in PV, 3 days before and on 3-consecutive days after a marathon race. By the 2nd day of recovery PV had increased by 16%. Plasma osmolality and [K⁺] remained constant, whereas [Na⁺] had decreased slightly 2 days after the competition and [Aldo] tended to be elevated 1 day after the competition. [Hpto] was low before and 1 day after the competition and increased on the following days. Reticulocyte count was unaffected 1 day after the race, but increased by 106% on the 2nd day and was still elevated after 3 days. The causes for higher post-marathon plasma volumes and reticulocyte counts could be in the complex variations in hormonal regulation, which have not yet been sufficiently investigated. A preliminary report was presented at the 64^th congress of the German Physiological Society in 1987     

Human cytomegalovirus

Summary Exposure of human lung fibroblasts to human cytomegalovirus (HCMV) stimulated a rapid increase in the release of [³H] from cells prelabelled with radiolabelled arachidonic acid ([³H]AA). Maximum stimulation of [³H] release was observed at 20 min postinfection and was quantitatively similar to that induced by the tumor promoter 12-O-tetradecanoylphorbol 13-acetate (TPA: 10nM) or fetal calf serum (5%). The level of [³H] release was dependent on the multiplicity of infection, and appeared to be mediated by a component(s) of the virion, since the findings from three series of experiments suggested that neither infectious virus, nor HCMV-specific macromolecular synthesis was required for stimulation of [³H] release. (1) Inactivation of HCMV infectivity with ultra-violet (UV) light (254nm, 4.80 × 10⁴ ergs/mm²) did not diminish the stimulation of [³H] release. (2) Significant reduction in the level of [³H] release was not observed when infected cells were maintained in the presence of a protein synthesis inhibitor, cycloheximide (50 µg/ml), or an inhibitor of mRNA synthesis, 3-deoxyadenosine (cordycepin, 50 µg/ml). (3) No correlation was established between the expression of HCMV immediate early (IE) antigens and the induction of [³H] release, since there was little, if any, synthesis of HCMV IE antigen detectable by anticomplement immunofluorescence through the first 30 min postinfection. These findings suggesting that the HCMV particle rapidly stimulates AA metabolism are consistent with the view that the interaction of a HCMV virion component(s) with the cell surface may initiate membrane-associated events similar to those induced by growth factors.     

Characterization of levamisole binding sites in Trichinella spiralis

Characterization of the levamisole receptor was performed with total extracts of Trichinella spiralis muscle larvae using binding assays with tritiated levamisole ([³H]LEV, 291 GBq/mmol). We detected a specific [³H]LEV binding activity with a dissociation constant (Kd) of 4.76 μM and a receptor density (Bmax) of 2.14 pmol/mg of protein. In inhibition studies, only dimethylphenylpiperazinium iodide (DMPP) and hexamethonium were found to be competitive inhibitors of the [³H]LEV binding with an inhibition constant (Ki) of 31.04 and 4.43 μM, respectively, whereas d-tubocurarine and α-bungarotoxine had no effect on [³H]LEV binding activity, and procaine and atropine potentiated the [³H]LEV-receptor binding. All these data support the idea that levamisole acts as a cholinergic agonist in T. spiralis. Received: 6 April 1998 / Accepted: 24 April 1998     

Study on the cationic polymerization mechanism of 1,3-dioxepane in the presence of ethylene glycol

The cationic polymerization of 1,3-dioxepane (DOP) initiated with trifluoromethanesulfonic acid (I) in the presence of ethylene glycol (EG) was investigated. At sufficiently low concentration of the initiator ([I] > 0.01 mol/L vs. [EG] < 0.20 mol/L), the molecular weights of the obtained polyacetal oligodiols are controlled by the mole ratio of consumed DOP to initial EG. Gel-permeation chromatography studies revealed that the concentration of cyclic oligomers in the products are negligible. The mechanism of the polymerization was investigated by means of kinetic studies. The results showed that the polymerization proceeds according to the active chain end mechanism (ACF) in combination with the activated monomer mechanism (AM); thus the cyclic oligomer in the obtained polymer is reduced, and intermolecular chain transfer to EG in ACE is dominant. It was also demonstrated that as [DOP]²[I]/[EG] decreases the contribution of ACE to the polymerization decreases and that of AM increases. In addition, ¹H and ¹³C NMR data illustrated that each macromolecule of polyDOP oligodiols contained one EG unit on average and that no EG end groups exist.     

Analysis of Human Serum Immunoglobulin G against O-Acetyl-Positive and O-Acetyl-Negative Serogroup W135 Meningococcal Capsular Polysaccharide

The capsular polysaccharide of Neisseria meningitidis serogroup W135 is expressed in both O-acetyl-positive (OA⁺) and O-acetyl-negative (OA⁻) forms. This study investigates the impact of OA status (OA⁺ versus OA⁻) on serological measurements of anti-W135 immunoglobulin G (IgG) antibodies in immunized adults. W135-specific serum antibody assignments were made for 28 postimmunization sera from adults by enzyme-linked immunosorbent assay using the meningococcal standard reference serum CDC1992. The established IgG concentration in micrograms per milliliter ([IgG]μg/ml) for CDC1992 against OA⁺ antigen (16.2 μg/ml) was used as a reference to assign a concentration of 10.13 μg/ml IgG against OA⁻ antigen by cross-standardization. Overall, the IgG assignments for these sera were higher against OA⁺ antigen (geometric mean concentration [GMC] = 7.16 μg/ml) than against OA⁻ antigen (GMC = 2.84 μg/ml). However, seven sera showed higher specific [IgG]μg/ml values against the OA⁺ antigen than against the OA⁻ antigen. These sera were also distinguished by the inability of fluid-phase OA⁻ antigen to compete for antibody binding to OA⁺ solid-phase antigen. Although there was no overall difference in functional activity measured by complement-mediated serum bactericidal assay (SBA) against OA⁺ and OA⁻ target bacteria (geometric mean titers of 9,642 and 9,045, respectively), three serum specimens showed a large difference in SBA antibody titers against OA⁺ versus OA⁻ W135 target bacteria, which may reflect different epitope specificities for these sera. Our data indicate that, for some sera, the agreement in anti-OA⁺ versus anti-OA⁻ W135 IgG assignments is serum specific and does not reflect the functional (killing) activity in vitro.     

The intracellular localization of two antigens after uptake in vivo by peritoneal macrophages from normal mice

Normal mouse macrophages, which had ingested ferritin labelled with fluorescein isothiocyanate and human serum albumin labelled with tetramethylrhodamine isothiocyanate in vivo, were fixed in formalin and embedded for electron microscopy. The examination of sections 1–2 μ thick and adjacent ultrathin sections showed that the yellow-green fluorescent droplets (due to ferritin-FITC) seen by fluorescence microscopy were in the same position as the ferritin-containing phagolysosomes as seen by electron microscopy.

Normal mouse macrophages, which had ingested ¹²⁵I-labelled human serum albumin ([¹²⁵I]HSA) and unlabelled ferritin, were investigated by electron microscopic autoradiography. Both antigens were found to be situated within the same lysosomes.

     

Erythropoietin concentrations during 10 days of normobaric hypoxia under controlled environmental circumstances

Serum erythropoietin levels (s‐[epo]), haemoglobin concentration ([Hb]), haematocrit (hct), and ferritin concentration ([fer]) were measured in seven healthy male volunteers (20–23 years) exposed continuously to hypoxia (PO₂ 14 kPa) for 10 days. Serum erythropoietin concentration increased significantly from 9.5 ± 3.51 to 33.6 ± 11.64 U L^–1 (P < 0.05) after 2 days of hypoxia. Thereafter, s‐[epo] decreased. However, after 10 days s‐[epo] was 18.7 ± 5.83 U L^–1 which was still increased above the pre‐hypoxia level (P < 0.05). Serum haemoglobin concentration and hct increased over the 10 days of hypoxia, [Hb] from 152 ± 8.9 to 168 ± 9.2 gL^–1 (P < 0.001), and hct from 43 ± 2.4 to 49 ± 2.6% (P < 0.001). Ferritin concentration decreased significantly during the hypoxic exposure from 82 ± 46.9 to 44 ± 31.7 mmol L^–1 after 10 days (P < 0.01). In conclusion, the initial increase of s‐[epo] under controlled normobaric hypoxia was marked, 353%, and levelled off after 5–10 days at 62–97% above normoxia level. There was also a significant increase in [Hb] and hct and a decrease in [fer] after 10 days of exposure to normobaric hypoxia.     

Haematological and iron-related parameters of male endurance and strength trained athletes

Summary To obtain more information on the effects of long-lasting endurance and strength training on the constituents of the blood, several haematological and iron-related parameters were measured at rest in 39 male athletes from the Polish team who participated in the Olympics in Seoul in 1988. The athletes were divided into two groups: endurance-trained subjects (group E, cyclists, canoeists and rowers; n=22) and strength-trained subjects (group S, wrestlers and judo; n=17). The control group was composed of untrained male subjects (n=48). Blood samples were taken from an antecubital vein with the subject at rest for determinations of haemoglobin concentration ([Hb]), packed cell volume (PCV), erythrocyte (RBC) and reticulocyte count, plasma free haemoglobin concentration, haptoglobin concentration, serum iron, transferrin concentration and ferritin concentrations ([Ferr]); red blood cells were used for estimation of glutamato-oxalate transaminase (GOT) activity and free erythrocyte protoporphyrin concentration ([FEP]). The mean [Hb], PVC, RBC measured in the E athletes were significantly lower than in the control group but were comparable to those obtained in the S athletes. There were no significantly differences in the haematological indices [mean corpuscular volume (MCV), mean copuscular haemoglobin and mean corpuscular haemoglobin concentration] between the groups of athletes and the control group. A significant increase in reticulocytosis and GOT activity was observed in the endurance-trained athletes. No impairment of erythropoiesis was observed as indicated by several sensitive markers of haemoglobin formation (FEP, MCV and inspection of blood smears) in the athletes. The athletes from group E had mean serum [Ferr] below 50 g·l^–1 which was significantly lower than [Ferr] in the serum of subjects from the control group and the strength-trained athletes. The results of the present investigation showed that some haematological parameters and the iron status of the endurance athletes differed from the untrained subjects as well as the strength-trained athletes.     

Effect of acute mild hypoxia during exercise on plasma free and sulphoconjugated catecholamines

Catecholamine (CA) response to hypoxic exercise has been investigated during severe hypoxia. However, altitude training is commonly performed during mild hypoxia at submaximal exercise intensities. In the present study we tested whether submaximal exercise during mild hypoxia compared to normoxia leads to a greater increase of plasma concentrations of CA and whether plasma concentration of catecholamine sulphates change in parallel with the CA response. A group of 14 subjects [maximal oxygen uptake, 62.6 (SD 5.2) ml · min^–1 · kg^–1 body mass] performed two cycle ergometer tests of 1-h duration at the same absolute exercise intensities [191 (SD 6) W] during normoxia (NORM) and mild hypoxia (HYP) followed by 30 min of recovery during normoxia. Mean plasma concentrations of noradrenaline ([NA]), adrenaline ([A]), and noradrenaline sulphate ([NA-S]) were elevated (P < 0.01) after HYP and NORM compared with mean resting values and were higher after HYP [20.9 (SEM 3.1), 2.2 (SEM 0.24), 8.12 (SEM 1.5) nmol · 1^–1, respectively] than after NORM [(13.7 (SEM 0.9), 1.5 (SEM 0.14), 6.8 (SEM 0.7) nmol · 1^–1, respectively P < 0.01]. The higher plasma [NA-S] after HYP (P < 0.05) were still measurable after 30 min of recovery. From our study it was concluded that exercise at the same absolute submaximal exercise intensity during mild hypoxia increased plasma CA to a higher extent than during normoxia. Plasma [NA-S] response paralleled the plasma [NA] response at the end of exercise but, in contrast to plasma [NA], remained elevated until 30 min after exercise.     

Sustained noradrenaline sulphate response in long-distance runners and untrained subjects up to 2 h after exhausting exercise

Summary We investigated the response of plasma and platelet free catecholamine ([CA]) and sulphated catecholamine ([CA-S]) concentrations after an incremental treadmill test to exhaustion and during recovery. In triathletes (n = 9) plasma and platelet [CA] and [CA-S] were measured before, immediately after and 0.5 and 24 h after exercise. In long-distance runners (n = 9) and in controls (n = 10) plasma [CA] and [CA-S] were determined 2 h instead of 24 h after exercise. Platelet [CA] and [CA-S] remained unchanged throughout the study. Plasma [CA] increased after exercise in all groups (P<0.05) and returned to pre-exercise values within 0.5 h of recovery. Plasma sulphoconjugated noradrenaline concentration ([NA-S]) was elevated after exercise in the triathletes, long-distance runners and in controls [9.96 (SEM 0.84) nmol·1^–1, 11.8 (SEM 1.19) nmol·1^–1, 9.53 (SEM 1.10) nmol·l^–1, respectively;P<0.05] compared with resting values [7.13 (SEM 1.04) nmol·l^–1, 6.19 (SEM 0.56) nmol·l^–1, 6.76 (SEM 0.67) nmol·1^–1, respectively] and remained elevated after 0.5 h of recovery [9.94 (SEM1.14) nmol·l^–1, 10.96 (SEM 0.80) nmol·l^–1, 8.95 (SEM 0.99) nmol·l^–1, respectively;P<0.05]. In the long-distance runners and controls plasma [NA-S] remained elevated during 2 h of recovery [9.96 (SEM 0.76) nmol·l^–1, 9.03 (SEM 0.88) nmol·l^–1, respectively]. These results would indicate that plasma [NA-S] increases after sympathetic nervous system activation by an exhausting incremental exercise test and remain elevated up to 2 h after exercise.     

A study of the initial interaction of the Sendai virus with cells,using the method of autoradiography

Summary A study was made of localization in the cellular structures of the Sendai virus with labeled nucleic (with the aid of uracil-C¹⁴ and radiophosphorus) or protein (with the aid of methionine and S³⁵-cystine) components. Autoradiography was employed for this purpose. As revealed, during the action of virus preparations labeled with C¹⁴-uracil and P³⁵ on the chick embryo cells and human amniotic cells, radioactive granules localized in the nucleoli and nuclei during the first hour then partially passed into the perinuclear zone of the cytoplasm. With the use of S^–35 labeled preparations radioactive granules were evenly distributed in the cells. Basing upon the data obtained it may be concluded that the nucleic component of the virus penetrates into the nuclei and nucleoli whereas the protein component or its considerable part remains on the cell membranes or in the ectoplasm.Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 56, No. 7, pp. 67–73, July, 1963     

The role of posture on the changes in plasma atrial natriuretic factor and arginine vasopressin levels during immersion

Summary In seven healthy male volunteers we investigated changes in plasma atrial natriuretic factor ([ANF]), arginine vasopressin ([AVP]) and plasma volume (PV) during supine immersion. Twenty minutes head-out water immersion in a supine position in a thermo-neutral water bath attenuated the increase in PV induced by 20 min in a supine position in air, but increased the mean plasma [ANF] from 32.0 pg · ml^–1, SEM 5.1 to 53.3 pg · m^–1, SEM 3.6 and decreased the mean plasma [AVP] from 1.4 pg · ml ^–1, SEM 0.1 to 0.9 pg · ml^–1, SEM 0.04. Simultaneously, diuresis and natriuresis increased markedly. During a 20-min control period in the supine posture without immersion, PV, plasma [ANF] and [AVP] remained unaffected while diuresis and natriuresis did not increase to the same extent. These data suggest that an increase in the central blood volume induced by a weak external hydrostatic pressure during supine immersion triggered the changes in plasma [ANF] and [AVP] and that the increase was probably due to a shift of blood volume from peripheral to central vessels. The changes in plasma [ANF] contributed to the changes in natriuresis.     

Identification ofβ 2-adrenoceptors on guinea pig alveolar macrophages using (−)-3-[125I]iodocyanopindolol

The -adrenoceptor antagonist (–)-3-[¹²⁵I]iodocyanopindolol ([¹²⁵I]ICYP) binds with high affinity and in a saturable way to membranes of guinea pig alveolar macrophages. The equilibrium dissociation constant for [¹²⁵I]ICYP is 24.3 ± 1.2 pM, and the number of binding sites is 166.3 ± 13.7 fmol/mg protein (N=4, ±SEM). Displacement studies with selective antagonists showed that [¹²⁵I]ICYP labels ₂-adrenoceptors on guinea pig alveolar macrophages.