Rat epididymis-specific sperm maturation antigens. I. Evidence that the 26 kD 4E9 antigen found on rat caudal epididymal sperm tail is derived from a protein secreted by the epididymis

Monoclonal antibody 4E9, which was raised against a partially purified detergent extract of rat caudal epididymal sperm, recognizes the tail of sperm from the cauda, but not from caput epididymidis, as well as epithelial cells in a restricted region of the distal caput/corpus epididymidis and proteins in epididymal fluid from corpus and cauda epididymidis. The antigen is apparently a glycoprotein, since it is retained on a Ricinus communis agglutinin l lectin column. Epididymal fluid antigens have apparent M_rS of 38–26 kD, whereas the memrane-associated form of the molecule has an M_r of 26 kD. Immunocytochemical data and Western immunoblot data suggest that the membrane antigen is derived from the fluid antigen, which, in turn, is secrteted by the epididymal epithelium. Characterization of the membrane antigen indicates that it is tightly associated with the sperm surface, behaving as though it is an integral membrane protein. The antigen persists on ejaculated sperm. © 1994 Wiley-Liss, Inc.     

Neither Moderate Hypoxia nor Mild Hypoglycaemia Alone Causes Any Significant Increase in Cerebral [Ca2±i: Only a Combination of the Two Insults Has This Effect.: A 31P and 19F NMR Study

(1) The energy state and free intracellular calcium concentration ([Ca^2±_i) of super-fused cortical slices were measured in moderate hypoxia (～65 μM O₂), in mild hypoglycaemia (0.5 mM glucose), and in combinations of the two insults using ¹⁹F and ³¹P NMR spectroscopy. (2) Neither hypoxia nor hypoglycaemia alone caused any significant change in [Ca^2±_i. Hypoxia caused a 40% fall in phosphocreatine (PCr) content but not in ATP level, and hypoglycaemia produced a slight fall in both (as expected from previous studies). These changes in the energy state recovered on return to control conditions. (3) A combined sequential insult (hypoxia, followed by hypoxia plus hypoglycaemia) produced a 100% increase in [Ca^2±, and a decrease in PCr level to ～25% of control. The reverse combined sequential insult (hypoglycaemia, followed by hypoglycaemia plus hypoxia) had the same effect. On return to control conditions there was some decrease in [Ca^2±_i and a small increase in PCr content, but neither recovered to control levels. (4) Exposure of the tissue to the combined simultaneous insult (hypoxia plus hypoglycaemia) immediately after the control spectra had been recorded resulted in a fivefold increase in [Ca^2±_i and a similar decrease in PCr level to 20–25% of control. There was little if any change of [Ca^2±_i or PCr level on return to control conditions. (5) These results are discussed in terms of metabolic adaptation of some but not all of the cortical cells to the single type of insult, which renders the tissues less vulnerable to the combined insult.     

Phloretin as an Antagonist of Prostaglandin F2α Receptor in Cultured Rat Astrocytes

Abstract: The effect of phloretin on prostaglandin (PG) F_2α-induced phosphoinositide hydrolysis and elevation of intracellular Ca²⁺ concentration was examined in cultured rat astrocytes. Phloretin inhibited PGF_2α (1 μ M )-induced phosphoinositide hydrolysis in a concentration-dependent manner with an IC₅₀ value of 16 μ M . The inhibitory action of phloretin was specific for PGs. The addition of increasing concentrations of phloretin caused progressive shifts of the dose-response curves of PGF_2α to the right. In digitoninpermeabilized astrocytes, phloretin (100 μ M ) inhibited the stimulation induced by PGF_2α (1 μ M ) plus GTPγS (50 μ M ) without affecting that induced by GTPγS alone. PGF_2α at 1 μ M transiently increased astrocytic intracellular Ca²⁺ concentration in 39% of the cells tested. The response was completely blocked by 100 μ M phloretin and the calcium response recovered again after washing out phloretin. These results suggest that phloretin is an antagonist of PGF_2α receptor linked to phospholipase C in astrocytes.     

Does Gγ/Aγ ratio and Hb F level influence the severity of sickle cell anaemia

Sickle cell anaemia (SCA) exhibits significant variations in clinical presentation in different populations for which several genetic factors including SCA-associated -and -thalassaemias, G-6-PD deficiency and elevated Hb F level have been implicated as possible ameliorating factors. Saudi Arabia is unique in that mild and severe forms of the disease occur at a high frequency. We investigated the G/A ratio and Hb F level and correlated these values with the severity of SCA. The results showed that Hb F level varies significantly in both groups of patients with no evident correlation with the mild clinical manifestations. However, G/A ratio correlated significantly with the disease severity where a high ratio was observed in patients with the mild and a low ratio in patients with the severe disease. The results are evaluated and discussed in the light of correlation studies and regression analysis.     

Conformation and activity ofPhaseolus coccineus var. rubronanus lectin

The conformation of native and denaturedPhaseolus coccineus var. rubronanus lectin was studied by circular dichroism (CD) and correlated to the hemagglutinating activity. The far-UV CD spectrum at 25°C showed a broad, negative band around 223 nm and a positive one at 196 nm. CD data analysis of the lectin indicated a -sheet-rich protein. At high temperatures, the spectrum was blue-shifted with increasing magnitude; these changes correlated well with the loss of the activity. The conformation of lectin betweenpH 2 and 10 remained essentially unchanged. AtpH 13 the CD spectrum resembled that of unordered form with a negative band near 200 nm and the activity was completely lost. The denatured lectin in 6 M guanidine hydrochloride would be renatured upon diluting the denaturant to 0.75 M; the changes in CD spectrum again correlated well with the loss of the activity. The effect of sodium dodecyl sulfate on the lectin was drastic; it sharply increased thea-helix at the expense of the -sheet and reduced the activity; the changes reached a plateau above 20 mM surfactant.     

A carbon-13 nuclear magnetic resonance investigation of the metabolic fluxes associated withy glucose metabolism in human erythrocytes

We have used [2-¹³C]d-glucose and carbon-13 nuclear magnetic resonance (NMR) spectroscopy to investigate metabolic fluxes through the major pathways of glucose metabolism in intact human erythrocytes and to determine the interactions among these pathways under conditions that perturb metabolism. Using the method described, we have been able to measure fluxes through the pentose phosphate pathway, phosphofructokinase, the 2,3-diphosphoglycerate bypass, and phosphoglycerate kinase, as well as glucose uptake, concurrently and in a single experiment. We have measured these fluxes in normal human erythrocytes under the following conditions: (1) fully oxygenated; (2) treated with methylene blue; and (3) deoxygenated. This method makes it possible to monitor various metabolic effects of stresses in normal and pathological states. Not only has ¹³C-NMR spectroscopy proved to be a useful method for measuring in vivo flux through the pentose phosphate pathway, but it has also provided additional information about the cycling of metabolites through the non-oxidative portion of the pentose phosphate pathway. Our evidence from experiments with [1-¹³C]-, [2-¹³C]-, and [3-¹³C]d-glucoses indicates that there is an observable reverse flux of fructose 6-phosphate through the reactions catalyzed by transketolase and transaldolase, even in the presence of a net flux through the pentose phosphate pathway.     

放线菌素D和环已亚胺对伊贝母胚性愈伤组织中胚性细胞团和球形胚发生及大分子代谢动态的影响

伊贝母(F.pallidiflora Schrenk)胚性愈伤组织接种于NAA 1.0mg/L+6-BA2.0 mg/L的MS培养基上,在培养10天前可产生大量单细胞到多细胞胚性细胞团,培养10至15天,逐渐形成大量球形胚。利用这样一个实验体系,在培养0、1、2、3和4天后加入放线菌素D(AMD,20μg/ml)和环己亚胺(CHM,20μg/ml),继续培养至第6天,分析大分子代谢动态和观察胚性细胞团的形成情况;培养6和10天后加入同样浓度的AMD和CHM。继续培养至第15天,分析大分子代谢动态及观察球形胚形成情况。结果表明:(1)培养0、1、2、3和4天加入AMD的分别抑制胚性细胞团的100%、63%和45%,加入CHM的抑制100%、85%和75%,培养6和10天后加入CHM抑制球形胚的100%和75%;(2)DNA、RNA和蛋白质在胚性细胞团和球形胚形成时出现两个峰值,其中RNA变化剧烈,最早出现峰值。AMD和CHM分别抑制RNA和蛋白质的合成;(3)过氧化物酶同工酶带在胚性细胞团和球形胚形成过程中顺序表达,AMD和CHM分别在转录和转译水平上对其进行规律性抑制。根据以上结果,本文对伊贝母体细胞胚胎发生的机制进行了初步讨论。     

动物细胞在鼓泡式生物反应器中的死亡速率

通过实验测定,证明生物反应器中细胞死亡速率与气体鼓泡速率成正比而与反应器体积成反比。实验发现气泡大小对细胞死亡速率具有两种作用,一种作用在于影响气泡表面积生成速率;另一种作用则在于影响细胞在气泡表面的吸附程度,其最佳直径为5mm左右。血清和Pluronic F68能显著降低细胞死亡速率,当Pluronic F68浓度达到0．1％时,kd趋于零。所有这些实验结果均与前文提出的生物反应器设计模型具有很好的一致性。     

凤眼莲根分泌物氨基酸组分对根际肠杆菌属F2细菌的趋化作用

凤眼莲(Eichhornia crassipes)的根分泌物中含有Met等多种氨基酸,其中Met、GABA、Gly、Ala、Asp、Ser、Val和Leu(10^-7～10^-2mol·L^-1)均对凤眼莲的根际肠杆菌属F₂(Enterobacter sp.F₂)细菌有强烈的正趋化作用;Glu、Thr和His(10^-7～10^-3mol·L^-1)也对该菌有一定的正趋化作用;而Lys、Cys、Arg、Tyr、Pro、Asn、Gln、Ile、Phe和Typ则对该菌表现出一定的负趋化作用.对细菌的正趋化作用存在一个趋化物的最适浓度范围.具有正趋化作用的氨基酸在凤眼莲根际的浓度都较高,而具有负趋化作用的浓度则较低,这正是凤眼莲与该根际细菌结合为根际微生态系统的原因之一.