Toward understanding the influence of soil metals and sulfate content on plant thiols.

Plants respond to increased concentrations of metals by a number of mechanisms, including chelation with phytochelatins (PCs). Soil specimens and plants (Veronica anagalis-aquatica, Typha domingensis, Cynodon dactylon, Chenopodium album, Rumex dentatus, Amaranthus gracilis, Chenopodium murale, Inula viscosa) leaves were collected from two sites in northern Jordan and subsequently metals (cadmium, copper, and lead), sulfate, and PC (from leaves) levels were determined. One of these sites was contaminated with metals and the other served as a control site. The contaminated site had elevated cadmium, copper, lead, and sulfate levels. This increase of metal and sulfate levels in the soil at the contaminated site correlated with a rise in plant total glutathione (GSH(T)) and cysteine (CYS(T)). These increases were not attributed to an elevation in total phytochelatin levels. However, a significant increase in the ratio of short-chain phytochelatins to the total phytochelatin stores was observed. The individual effects of metals and sulfate on glutathione, short-chain PCs and long-chain PCs levels were dissimilar.

Polyamines protect against radiation-induced oxidative stress.

Astronauts and cosmonauts are exposed to a wide variety of different hazards while in space that include radiation, which presents one of the most critical barriers to long-term missions. A major deleterious effect directly associated with ionizing radiation is the production of reactive oxygen species (ROS) such as peroxides and hydroxyl radicals. The free radicals generated by ultraviolet (UV) or ionizing radiation can attack cellular lipids, proteins and DNA. Endogenous free radical scavengers such as glutathione and the polyamines (e.g, spermidine and spermine) can inhibit the action of ROS. In particular, heme oxygenase-1 (HO-1), the enzyme involved in heme protein metabolism, can provide antioxidant protection through the production of the antioxidant bilirubin. Furthermore, polyamines have been shown to indirectly increase HO-1 content and antioxidant protection. The beta2-adrenoceptor agonist clenbuterol has been shown to stimulate polyamine synthesis and by extension, might provide a margin of antioxidant protection through increasing HO-1 content. However, it is unclear whether the polyamines are acting as a tertiary messengers for antioxidant protection in the be beta2-adrenoceptor signal transduction pathway. The purpose of this study was to study the role of the polyamine pathway in attenuating free radical-induced damage.

Relation of endothelial nitric oxide synthase gene to plasma nitric oxide level, endothelial function, and blood pressure in African Americans.

BACKGROUND: The role of eNOS gene polymorphisms on plasma nitrite or nitrate (NOx) level, endothelial function, and blood pressure (BP) remains unclear. METHODS: We estimated the relationship of eNOS polymorphisms (the T(-786)C in the 5'-flanking promoter region, T(-786)C; 27-bp repeat in intron 4, eNOS4; and Glu298Asp in exon 7, G894T) with plasma NOx level, brachial endothelial function assessed by ultrasound measure of brachial artery flow-mediated dilation (FMD), and BP in 60 healthy African Americans, 30 men and 30 women aged 18 to 73 years. RESULTS: Among them, 73.1%, 23.9%, and 3.0% carried TT, TC, and CC of T(-786)C, respectively, 14.5%, 27.5%, 53.6%, and 1.4% carried aa, ab, bb, and bc of eNOS4 polymorphism, respectively, and 70.4%, 23.9%, and 5.6% carried GG, GT, and TT of G894T, respectively. G894T and eNOS4 were observed in linkage disequilibrium. Mean values of age, plasma NOx, FMD, systolic and diastolic BPs were not significantly different (P >.05) by eNOS polymorphisms. Plasma NO(x) level was found to be associated with systolic BP (r = 0.51, P =.03), and diastolic BP (r = 0.41, P =.08), but not with FMD, in individuals with "a" allele of eNOS4 polymorphism after adjustment for age, body mass index, serum glucose, and smoking status. CONCLUSIONS: We reveal a positive association between plasma NOx level and BP in normotensive African Americans who carry the "a" allele of eNOS4. Because the frequency of the rare allele "a" is significantly higher in African Americans than in other ethnic groups, this finding may provide a clue to understanding the genetic susceptibility to hypertension in African Americans.

Changes in muscle proteins and spermidine content in response to unloading and clenbuterol treatment.

Anabolic agents such clenbuterol (Cb) are useful tools for probing the mechanisms by which muscles respond to disuse. Cb was examined under different loading conditions with respect to its effects on muscle mass, protein (myofibrillar and cytosolic), and spermidine content in mature male rats. Compared with control treatment, Cb significantly increased loaded and unloaded soleus, plantaris, and extensor digitorum longus (EDL) mass. Likewise, Cb significantly increased loaded and unloaded soleus (24.8 and 21.6%, respectively), plantaris (12.1 and 22.9%, respectively), and EDL (22.4 and 13.3%, respectively) myofibrillar protein content. After unloading, cytosolic proteins significantly increased in the EDL but decreased in the soleus and plantaris. Cb significantly increased cytosolic protein levels in all loaded muscles, while only causing increases in unloaded soleus. When compared with controls, unloading caused significant reductions in spermidine levels in the soleus (40.4%) and plantaris (35.9%) but caused increases in the EDL (54.8%). In contrast, Cb increased spermidine levels in unloaded soleus (42.9%), plantaris (102.8%), and EDL (287%). In loaded muscles, Cb increased spermidine levels in all three muscles, but to a lesser degree than under unloading conditions. Nonlinear regression analyses indicated that the plantaris behaves like a slow-twitch muscle under unloading conditions and like a fast-twitch muscle when loaded. This suggests that the responses of these muscles to unloading and (or) Cb treatment might be influenced by factors beyond fiber type alone.

Muscle-specific effects of hindlimb suspension and clenbuterol in mature male rats.

Anabolic agents are useful tools for probing the mechanisms by which muscle fibers perceive and respond to disuse. beta(2)-Adrenergic agonists exert protective, and/or reparative, effects on atrophying muscle tissue. The effects of one such agent, clenbuterol (Cb), were examined on muscle mass, total protein content, and myofibrillar protein content in selected hindlimb muscles [adductor longus (ADL), extensor digitorum longus (EDL), plantaris (PLAN), soleus (SOL)] of mature male rats, under different loading conditions. Pair-fed rats were divided into four experimental groups: vehicle- and Cb-treated nonsuspended, vehicle- and Cb-treated hindlimb suspended (HLS). Experiments lasted 14 days, during which the rats received subcutaneous injections of 1 mg/kg Cb or 1 ml/kg vehicle. HLS induced significant atrophy in all muscles, except the EDL, in a generally fiber type-related pattern. However, myofibrillar protein content was affected in a more regional pattern. Cb treatment of nonsuspended rats induced hypertrophy in all muscles, in a generally uniform pattern. However, myofibrillar protein content was affected in a more fiber type-related pattern. Cb treatment of HLS rats reduced or eliminated HLS-induced atrophy in all muscles, in a muscle-specific pattern. Overall, the ADL and SOL were most susceptible to HLS-induced atrophy. The PLAN had the greatest magnitude of Cb-induced sparing of atrophy. The results show that, in mature male rats, Cb exerts anabolic effects that are load-dependent and muscle-specific. Responses to this drug cannot be reliably predicted by fiber-type composition alone.

Effect of hindlimb suspension and clenbuterol treatment on polyamine levels in skeletal muscle.

Polyamines are unbiquitous, naturally occurring small aliphatic, polycationic, endogenous compounds. They are involved in many cellular processes and may serve as secondary or tertiary messengers to hormonal regulation. The relationship of polyamines and skeletal muscle mass of adductor longus, extensor digitorum longus, and gastrocnemius under unloading (hindlimb suspension) conditions was investigated. Unloading significantly affected skeletal muscle polyamine levels in a fiber-type-specific fashion. Under loading conditions, clenbuterol treatment increased all polyamine levels, whereas under unloading conditions, only the spermidine levels were consistently increased. Unloading attenuated the anabolic effects of clenbuterol in predominately slow-twitch muscles (adductor longus), but had little impact on clenbuterol's action as a countermeasure in fast- twitch muscles such as the extensor digitorum longus. Spermidine appeared to be the primary polyamine involved in skeletal muscle atrophy/hypertrophy.

High performance liquid chromatographic assay for the quantitation of total glutathione in plasma.

A simple and widely used homocysteine HPLC procedure was applied for the HPLC identification and quantitation of glutathione in plasma. The method, which utilizes SBDF as a derivatizing agent utilizes only 50 microl of sample volume. Linear quantitative response curve was generated for glutathione over a concentration range of 0.3125-62.50 micromol/l. Linear regression analysis of the standard curve exhibited correlation coefficient of 0.999. Limit of detection (LOD) and limit of quantitation (LOQ) values were 5.0 and 15 pmol, respectively. Glutathione recovery using this method was nearly complete (above 96%). Intra-assay and inter-assay precision studies reflected a high level of reliability and reproducibility of the method. The applicability of the method for the quantitation of glutathione was demonstrated successfully using human and rat plasma samples.