Effect of ultrafilterable fragments from chondroitinase and protease-treated aggrecan on calcium phosphate precipitation in liposomal suspensions

A liposome-centered endogenous precipitation method was used to investigate the effect of ultrafilterable fragments from the enzymatic digestion of rat chondrosarcoma aggrecan on the formation of insoluble calcium phosphate salts in buffered solutions at pH 7.4 and 22°C. Unlike the intact aggrecan and its major chondroitin sulfate and core protein components, disaccharide units from chondroitinase degradation of the aggrecan and small (<3kg/mol molecular weight) fragments from protease digestion of the core structure were found to be only weakly inhibitory toward mineral formation. Corresponding reductions in Ca²⁺-binding indicate that these fragments were unable to adsorb to active sites on the apatite surface for long enough periods to significantly hinder crystal growth. The data suggest that controlled enzymatic breakdown of aggrecan may be one possible mechanism by which the calcification of growth plate cartilage is allowed to advance in vivo.The commercial materials and equipment identified in this paper do not imply recommendation or endorsement by the National Institute of Standards and Technology nor is the material and equipment necessarily the best available for the purpose.     

钩藤碱对心血管系统部分药理作用研究

探讨钩藤碱对钙离子的拮抗作用以及对脑血流量和氧消耗的作用。方法观察钩藤碱对豚鼠心肌的兴奋性、功能性不应期、正阶梯现象的作用，以及对去甲肾上腺素诱发的兔主动脉条Ⅰ、Ⅱ相收缩的作用，同时观察钩藤碱对小鼠氧消耗和大鼠脑血流量的影响。结果钩藤碱能提高心肌兴奋性，延长功能性不应期，抑制正阶梯现象和兔主动脉条Ⅰ、Ⅱ相收缩；减慢小鼠氧消耗速度，对大鼠脑血流量无影响。结论钩藤碱具有许多钙拮抗剂的共同特点，提示其在抗心律失常方面会有一定作用。     

Relative weight of glucose, insulin and parathyroid hormone in the urinary loss of phosphate by chronically diabetic rats

This report deals with the relationships between glucose (G) and insulin on the tubular transport of phosphate (P) in chronically diabetic rats with high plasma levels of parathyroid hormone (PTH). Alloxan-induced diabetes leads to phosphorus depletion of the soft tissues. This phenomenon appears associated with weight loss and negative P balances caused by the increased urinary P excretion. Administration of 2 IU of insulin/100 g body weight (bw) to diabetic rats normalized their P balance and body weight. The effect of parathyroid function on the P metabolism of diabetic rats was investigated with balance experiments. Diabetic rats, intact or thyroparathyroidectomized (TPTX), have a greater urinary excretion of P than their controls. However, in control rats, the ratio intact:TPTX for urinary P is 1.0:0.76, showing the antiphosphaturic effect of parathyroid ablation. For diabetic animals, on the other hand, the ratio is 1.0:1.44. The simultaneous deficit of insulin and PTH thus quadruples the urinary P loss, instead of compensating for each other. The contribution of insulin deficit and hyperglycemia to the defect in tubular reabsorption (TRP) was investigated with clearance experiments (done on anesthetized, perfused rats). Five experimental groups were used: Controls (C), diabetics (D), controls+glucose (C+G), diabetics+insulin (D+I) and diabetics+insulin+glucose (D+I+G). All experimental groups showed a linear relationship between the TRP of P and G. The regression equation for C is significantly different (F=40.1, P<0.001) from that of D animals. The slope value measure the number of μmoles of P per μmol of G reabsorbed. For C and D rats, the ratio P:G approximates 1:4 and 1:20, respectively. The increase in P:G ratios represents the competition between both substrates for tubular resorption. Glycemias up to 11 mM (C and D+I) exist concurrent with the P:G ratio 1:4. Glycemias above 25 mM (D, C+G and D+I+G) produce a P:G ratio of 1:20. Fractional excretion of P (FEP) increased significantly in untreated, chronically diabetic rats (0.47± 0.12 vs controls=0.05±0.01, P<0.001). After a single intramuscular injection of insulin, the FEP decreased as a function of insulin levels. To normalize the FEP of diabetic rats in short-term experiments, insulin had to be administered in doses that produce plasma insulin levels 25 times greater than normal. The general information afforded by the present experiments shows that in untreated, chronically diabetic rats, insulin deficit plays an indirect role. The absence of PTH enhances the effect of hyperglycemia. The latter and the concurrent tubular overload of glucose are the cause of hyperphosphaturia in these animals. Received: 10 September 1996 / Accepted in revised form: 18 April 1997     

Fmoc/solid-phase synthesis of Tyr(P)-containing peptides through t-butyl phosphate protection

The synthesis is of Tyr(P)-containing peptides by the use of Fmoc-Tyr(PO₃Me₂)-OH in Fmoc/solid phase synthesis is complicated since, firstly, piperidine causes cleavage of the methyl group from the -Tyr(PO₃ Me₂)-residue during peptide synthesis and, secondly, harsh conditions are needed for its final cleavage. A very simple method for the synthesis of Tyr(P)-containing peptides using t-butyl phosphate protection is described. The protected phosphotyrosine derivative, Fmoc-Tyr(PO₃^tBu₂)-OH was prepared in high yield from Fmoc-Tyr-OH by a one-step procedure which employed di-t-butyl N,N-diethyl-phosphoramidite as the phosphorylation reagent. The use of this derivative in Fmoc/solid phase peptide synthesis is demonstrated by the preparation of the Tyr(P)-containing peptides, Ala-Glu-Tyr(P)-Ser-Ala and Ser-Ser-Ser-Tyr(P)-Tyr(P).     

Altered calcium metabolism in red blood cells of hypertensives: Persistent marker or sequel of essential hypertension?

Summary The characteristics of the increased calcium (Ca) influx observed in metabolically depleted red blood cells (RBCs) of hypertensive patients were investigated. Twenty-four normotensives, 16 untreated essential hypertensives, and 10 essential hypertensives under sufficient blood pressure control by 50–100 mg/day atenolol were studied. Free intracellular concentrations of Ca, sodium (Na), and potassium (K) were assessed using ion-selective electrodes in freeze-thawed RBCs, which were metabolically depleted by 30 mM desoxy-glucose at 37°C for 48 h. In the treated hypertensives values for Ca and K at 24 and 48 h were not different from values for the normotensives, whereas elevated Ca was found in RBCs of untreated hypertensives. Na in treated hypertensives was significantly increased at 0 and 48 h, thus, being similar to values for untreated hypertensives. Additionally, RBCs of six normals were stressed in a glass/teflon potter. Before metabolic depletion electrolytes were not affected by this procedure, while Ca at 24 and 48 h of metabolic depletion increased to significantly higher values for the hypertensive patients as compared to the controls. These results suggest that the altered Ca metabolism in the RBCs of hypertensives may reflect a secondary phenomenon due to the mechanical damage to RBCs by the elevated blood pressure.     

TD清洗消毒剂对口腔科器械消毒效果探讨

通过实验室和口腔科临床观察表明:TD清洗消毒剂(氯化磷酸三钠)对金黄色葡萄球菌、绿脓杆菌和枯草杆菌黑色变种的平均杀菌率为100.0％,对口腔科器械消毒无菌率为100％,对被HBsAg阳性血清污染的器械浸泡30分钟转阴率为98.1％。其消毒效果优于新洁尔灭,与戊二醛相同。在口腔科使用两年,未见金属器械有锈斑生成。     

The effects of brisk walking on markers of bone and calcium metabolism in postmenopausal women

Weight-bearing exercise has been shown to maintain or increase bone mass in younger as well as older individuals but the mechanisms by which mechanical loading affects bone metabolism are not known in detail. Twelve postmenopausal women participated in a single bout of brisk walking (50% of VO_{2 max}) for 90 minuttes. Calciotropic hormones and markers of type I collagen formation (PICP) and degradation (ICTP) were measured before the exercise, and 1, 24, and 72 hours following the exercise. Total body bone mineral content (BMC) and density (BMD) were measured by dual energy X-ray absorptiometry (DXA). Brisk walking did not induce any significant changes in the concentrations of ionized calcium, parathyroid hormone (PTH), calcitonin, or osteocalcin. A significant increase of PICP was noted 24 and 72 hours (P<0.01) after exertion and a significant decrease in the concentration of serum ICTP at 1 hour (P<0.05) was followed by an increase at 72 hours (P<0.001). There was no significant difference between the increases in the concentrations of PICP and ICTP at 72 hours. Strong inverse correlations between the basal levels of PTH and BMD (r=−0.78;P<0.01) as well as between osteocalcin and BMD (r=−0.83;P<0.01) were noticed. The changes in serum levels of bone collagen markers indicate an altered bone collagen turnover due to this moderate endurance exercise. The results also support the fact that serum levels of PTH as well as those of osteocalcin are associated with total body BMD in postmenopausal women.     

Niflumic acid-induced increase in potassium currents in frog motor nerve terminals: effects on transmitter release

The actions of the nonsteroidal antiinflammatory drug niflumic acid were studied on frog neuromuscular preparations by conventional electrophysiological techniques. Niflumic acid reduced the amplitude and increased the latency of endplate potentials in a concentration-dependent manner. Neuromuscular junctions pretreated with niflumic acid (0.05–0.5 mM) showed much less depression than control when they were stimulated with trains of impulses. Inhibition of acetylcholine release was reverted by raising the extracellular Ca²⁺ concentration but not by simply washing out the preparations with niflumic acid-free solutions. Pretreatment with indomethacin (0.1 mM), another nonsteroidal antiinflamatory drug, did not affect the niflumic acid-induced inhibition of evoked responses. Niflumic acid (0.1 mM) did not change the amplitude of miniature endplate potentials and had a dual action on the frequency of miniatures: it decreased their frequency at 0.1 mM whereas it produced an enormous increase in the rate of spontaneous discharge at 0.5 mM. Niflumic acid (0.1–1 mM) reversibly increased the amplitude and affected the kinetics of presynaptic voltage-activated K⁺ current and Ca²⁺-activated K⁺ current in a concentration-dependent manner. Niflumic acid (0.1–1 mM) irreversibly decreased the amplitude and reversibly affected the kinetics of the nodal Na⁺ current. Indomethacin (0.1 mM) had no effect on presynaptic currents. In conclusion, niflumic acid reduces acetylcholine release by increasing presynaptic K⁺ currents. This may shorten the depolarizing phase of the presynaptic action potential and may reduce the entry of Ca²⁺ with each impulse.     

Cardioprotective Effect of Succinate Against Ischemia/Reperfusion Injury

(Received for publication on Sept. 12, 1996; accepted on May 12, 1997)     

Activity-dependent survival and enhanced turnover of calcium in cultured rat cerebellar granule neurons

Neurons survive when their activity is maintained. An influential hypothesis on the cellular mechanism underlying this phenomenon is that there is an appropriate range of intracellular Ca²⁺ concentration ([Ca²⁺]i) for survival. The rat cerebellar granule neuron in culture serves as the most often used model system for the analysis of activity-dependent survival, since it does not survive unless an excitant (KCl or glutamate) is added to the culture medium. Against the above-mentioned hypothesis, we found in our previous examination no difference between steady-state [Ca²⁺]i in granule neurons cultured under high KCl (i.e., survival) and low KCl (i.e., death) conditions. In this report, we present the quantitative background of unchanged [Ca²⁺]i between the two culture conditions. Influx of Ca²⁺ due predominantly to L-type voltage-dependent calcium channels was higher in high KCl cultures than in low KCl cultures. At the same time, efflux of Ca²⁺ due to the activity of Ca²⁺/Na⁺ antiport was also higher in high KCl cultures. Additionally, we found that the endocytotic activity was greater in high KCl cultures than in low KCl cultures, as monitored by the rate of uptake of horseradish peroxidase added to medium. Since the uptake was blocked by an internal Ca²⁺ chelator, the increased endocytotic activity in high KCl cultures might be a consequence of the enhanced Ca²⁺ turnover.