改构型酸性成纤维细胞生长因子干预慢性衰老模型大鼠脑及肝组织和血液ATP酶的活力

背景：有研究表明改构型酸性成纤维细胞生长因子是多功能生长因子,但其抗衰老作用至今尚未有报道。 目的：观察改构型酸性成纤维细胞生长因子对D-半乳糖致衰老大鼠脑组织、肝组织及红细胞膜Na⁺-K⁺-ATP酶活力和Ca²⁺-Mg²⁺-ATP酶活力的影响。 方法：将Wistar大鼠采用皮下注射D-半乳糖建立衰老模型,建模成功后随机分为模型组、生理盐水对照组和改构型酸性成纤维细胞生长因子组,另设正常对照组。改构型酸性成纤维细胞生长因子组按12 µg/kg剂量肌肉注射改构型酸性成纤维细胞生长因子,生理盐水对照组肌肉注射等量的生理盐水,模型组不干预。 结果与结论：与正常对照组相比,模型组大鼠脑组织、肝组织及红细胞膜Na⁺-K⁺-ATP酶活力和Ca²⁺-Mg²⁺-ATP酶活力均明显著降低(P < 0.01)。改构型酸性成纤维细胞生长因子组脑组织、肝组织及红细胞膜Na⁺-K⁺-ATP酶活力和Ca²⁺-Mg²⁺-ATP酶活力均明显高于模型组和生理盐水对照组(P < 0.05)。结果提示,改构型酸性成纤维细胞生长因子能提高衰老大鼠脑组织、肝组织及红细胞膜Na⁺-K⁺-ATP酶活力和Ca²⁺-Mg²⁺-ATP酶活力,具有延缓衰老作用。     

Effects of free Ca on the [Ca + Mg-Dependent adenosinetriphosphatase (ATPase) of Alzheimer and normal fibroblasts

Differences between Alzheimer and control fibroblasts [Ca²⁺ + Mg²⁺]-dependent ATPase activity at free Ca²⁺ concentration considerably higher than physiologic concentrations were observed. At 50 μM free Ca²⁺, Alzheimer and control fibroblast homogenates exhibited maximum velocity values ranging from 8 to 25 nmoles phosphate released/min/mg protein. Higher free Ca²⁺ (350 μM) inhibited control fibroblast ATPase activity approximately 77%; whereas, Alzheimer fibroblasts retained greater than 75% starting activity. Although the pathophysiological significance of these findings is at present unclear, these data suggest the Ca²⁺ pump of Alzheimer fibroblasts behaves differently in the presence of high free Ca²⁺. Such behavior may be of potential diagnostic value.     

The Na/K-ATPase as [K]o sensor: Role in cardiovascular disease pathogenesis and augmented production of endogenous cardiotonic steroids

Current evidence demonstrates that augmented production of endogenous cardiotonic steroids (CTS) such as ouabain and marinobufagenin is involved in the pathogenesis of hypertension and other cardiovascular diseases associated with volume expansion. It is also well documented that the development of hypertension and the cardiovascular complications of this disease are provoked by hypokalemia and suppressed by high-K⁺ diet. We hypothesized that altered extracellular K⁺ (K⁺)_o handling contributes to the pathogenesis of hypertension via modulation of interaction of endogenous CTS with Na⁺/K⁺-ATPase. To examine this hypothesis, experiments were performed with C7-Madin-Darby canine kidney epithelial cells at [K⁺]_o detected in plasma under control conditions (4.5 mM), severe hypokalemia (2 mM), and hyperkalemia (7 mM). Elevation of [K⁺]_o from 2 to 7 mM increased the threshold of modulation of intracellular (Na⁺)_i and (K⁺)_i content by ouabain from 1 to 10 nM, which corresponds to the range of endogenous CTS detected in plasma from patients with volume-expanded disorders. In control medium, 30% activation of cell proliferation was observed with 3 nM ouabain, whereas the addition of 0.3 nM ouabain was sufficient to induce about the same increment of cell proliferation in K⁺-depleted medium. [K⁺]_o elevation up to 7 mM completely abolished the proliferative effect of ouabain. At [K⁺]_o = 2, 4.5 and 7 mM, the death of ouabain-treated cells was indicated in the presence of 10, 30 and 300 nM ouabain, respectively. In conclusion, our results showed that modulation of [K⁺]_o in a pathophysiologically reasonable range sharply affected efficacy of endogenous CTS in the elevation of the [Na⁺]_i/[K⁺]_i ratio and in triggering (Na⁺)_i,(K⁺)_i-independent signaling resulting in cell proliferation and death. We propose that Na⁺/K⁺-ATPase may be considered as a [K⁺]_o sensor involved in the crosstalk of (K⁺)_o handling with the pathogenesis of cardiovascular diseases.     

妊高征肾病患者血清CysC水平与红细胞膜ATP酶活性的关系

目的：探讨了妊高征肾病患者血清胱抑素蛋白酶抑制素C（CysC）水平与红细胞膜Na^＋、K^＋-ATP酶和Ca^2＋、Mg^2＋-ATP酶活性改变参与妊高征肾病发生的可能机制。方法：应用乳胶免疫增强比浊法和Reinil制膜法测定了32例妊高征肾病患者血清CysC水平与红细胞膜Na^＋、K^＋-ATP酶和Ca^2＋、Mg^2＋-ATP酶含量,并与35名正常孕妇组作比较。结果：妊高征肾病患者血清CysC水平非常显著地高于正常孕妇组（P〈0．01）,而红细胞膜Na^＋、K^＋-ATP酶和Ca^2＋、Mg2^2＋-ATP酶含量显著地低于正常孕妇组（P〈0．01）。结论：妊高征肾病的发生发展与血清CysC水平和红细胞膜Na^＋、K^＋-ATP酶和Ca^2＋、Mg^2＋-ATP酶活性有密切的关系。     

依达拉奉对断肢再植后肢体的保护作用

背景：怎样减轻断肢再植后缺血再灌注损伤的严重程度,目前尚无有效方法。 目的：探讨依达拉奉能否对断肢再植术后肢体起到保护作用。 方法：根据不同的干预方法,将18只健康兔随机数字表法均分为3组(n=6),分别为依达拉奉组、缺血再灌注组和空白对照组。建立左下肢的离断肢体模型,其中,依达拉奉组于再灌注前给予以依达拉奉为主的灌注液进行灌注(1.5 mg/kg),分别于2,4,8,12 h于兔的胫骨前肌取材,分别测定骨骼肌超氧化物歧化酶、髓过氧化物酶、丙二醛、Ca²⁺-ATP酶、Na⁺-K⁺-ATP酶,湿质量/干质量比值,光镜下观察骨骼肌结构的病理变化。 结果与结论：缺血再灌注后兔骨骼肌的肌纤维水肿,损伤明显,大量炎性细胞浸润,依达拉奉干预后损伤程度减轻,但均随时间增加而加剧。依达拉奉能够明显提高缺血再灌注后机体内超氧化物歧化酶、过氧化物酶、Na⁺-K⁺-ATP酶、Ca²⁺-ATP酶含量,降低丙二醛、湿质量/干质量比值,各项指标差异均存在显著性意义(P < 0.05),可见依达拉奉对断肢再植后肢体有着明显的保护作用。依达拉奉为主的混合灌注液能够有效地预防再植后肢体(特别是骨骼肌)的缺血再灌注损伤。 中国组织工程研究杂志出版内容重点：组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程全文链接：     

Endogenous protein phosphorylation in chick and rat brain synaptic membranes

The protein kinase activities endogenous to synaptic membranes prepared by an identical procedure from avian (chick) and mammalian (rat) brains were compared. Both species showed similar responses towards both protein kinase effector molecules cyclic adenosine monophosphate and Ca²⁺. K_app for cyclic adenosine monophosphate-dependent protein kinase activity occurred at 0.4–0.8μM cAMP and K_app for Ca²⁺-dependent, calmodulin-requiring protein kinase activity occurred at 1–2μM Ca²⁺ (free ion concentration) both in the absence or presence of calmodulin added to the reaction mixture. This suggests that endogenous calmodulin in these membranes was able to modulate the Ca²⁺-dependent, calmodulin requiring protein kinase activity. After EGTA-treatment of the membranes to remove endogenous Ca²⁺ and calmodulin, no significant response towards Ca²⁺ on the phosphorylation of the membrane polypeptides was measured unless exogenous calmodulin was added after which the K_app for Ca²⁺ was increased to 15μM Ca²⁺ (free ion concentration). There was a difference in the maximal levels of kinase activity in these membranes with chick membranes containing 57% less cyclic adenosine monophosphate-dependent protein kinase activity, but 65% more Ca²⁺-dependent, calmodulin-requiring protein kinase activity than the rat membranes. Similar results were determined when either low (5 μM) or high (5.8mM) concentrations of adenosine 5′-triphosphate were added to the reaction mixtures.

Besides certain species differences in the molecular weights of the resulting phosphoproteins, we observed several major differences with respect to the absence or presence of some of the phosphoproteins. Chick synaptic membranes may lack the cyclic adenosine monophosphate-requiring, microtubule-associated phosphoprotein, MAP₂, one of the 2 neurospecific, cyclic adenosine monophosphate-requiring and Ca²⁺, calmodulin-requiring phosphoproteins (Protein Ib, although Protein Ia apparently is present), and the Ca²⁺-requiring, calmodulin-independent, ACTH-sensitive phosphoprotein, B50.

The phenothiazines, trifluoperazine, fluphenazine and chlorpromazine were found to inhibit the Ca²⁺-dependent, calmodulin-requiring protein kinase activities of both the chick and rat synaptic membranes. This inhibition appeared to be specific for calmodulin because at the same concentrations the phenothiazine analogue, chlorpromazine-sulfoxide, had no effect on this activity. Also found to inhibit Ca²⁺-dependent calmodulin-requiring protein kinase activity were dibucaine and adrenocorticotropin.

These data suggest that rat forebrain synaptic plasma membranes are activated by cyclic adenosine monophosphate to a greater extent than are chick forebrain synaptic plasma membranes while the chick membranes are activated to a greater extent by Ca²⁺, calmodulin than are the rat membranes, though similarities dominate comparison of the two membrane systems.     

Effects of Escherichia coli Endotoxin on Structure and Permeability of Myocardial Capillaries

A study was carried out to investigate changes in myocardial capillaries induced by endotoxin, in order to clarify the pathogenesis of myocardial damage in endotoxemia. Wis-tar rats were injected intraperitoneally with 100 mg/kg Escherichia coli lipopolysaccharide and then sacrificed at 1, 2, 3,4, 5,6,8, and 24 h after injection. The myocardium was observed by electron microscopy with histochemistry using horseradish peroxidase and immunocytochemistry for Na⁺, K⁺ ATPase. The earliest evident endothelial alterations were swelling, increased numbers of pinocytotic vesicles, and formation of cytoplasmic projections. Interstitial edema and focal detachment of the endothelial cells from the basement membrane occurred with time. Vascular permeability was increased after endotoxin injection. Activity of Na⁺, K⁺-ATPase was reduced on the plasma membrane of the endothelial cells. It is concluded that endotoxin induces structural and enzymatic changes in the myocardial capillary endothelium and an increase of capillary permeability. Acta Pathol Jpn 41: 12–18, 1991.     

Mechanism of histamine-induced calcium efflux from cultured bovine adrenal chromaffin cells: possible involvement of an Na/Ca exchange mechanism

The effect of stimulation of the histamine receptor on Ca²⁺ mobilization in cultured bovine adrenal chromaffin cells was examined. Histamine (10⁻⁵ M) increased the intracellular free Ca²⁺ ([Ca²⁺]_i) to a peak in the presence or absence of extracellular Ca²⁺, followed by decrease with time. Histamine (10⁻⁸–10⁻⁵ M) also stimulated ⁴⁵Ca²⁺ efflux from cultured bovine adrenal chromaffin cells in a concentration dependent manner. Its stimulatory effect on ⁴⁵Ca²⁺ efflux was inhibited by the specific histamine H₁ receptor antagonist mepyramine. The increase in histamine-stimulated ⁴⁵Ca²⁺ efflux was inhibited by deprivation of extracellular Na⁺ and by the Na⁺/Ca²⁺ exchange inhibitor amiloride. In addition, histamine stimulated ²²Na⁺ influx into the cells, and this action was inhibited by amiloride. These results suggest that stimulation of the histamine H₁ receptor regulates Na⁺/Ca²⁺ exchange in cultured bovine adrenal chromaffin cells.     

线粒体酶活性及线粒体肿胀与不同运动强度的关系

背景：关于不同运动强度对机体的影响早有研究,并且取得了不少有价值的成果。但是对于不同运动强度对线粒体Na⁺,K⁺-ATP酶、Ca2⁺-ATP酶活性以及线粒体肿胀程度影响的研究较少。 目的：观察不同运动强度对大鼠骨骼肌线粒体Na⁺,K⁺-ATP酶和Ca2⁺-ATP酶以及线粒体肿胀的影响。 方法：将24只SD大鼠随机分为正常对照组,中等强度运动组和高强度运动组。正常对照组大鼠正常笼内生活,不运动。另2组游泳训练方式,分别建立中强度和高强度运动模型,进行相应的运动锻炼,训练4周。 结果与结论：中强度运动组线粒体Na⁺,K⁺-ATP酶、Ca2⁺-ATP酶活性与正常对照组相比均增高(P < 0.05),线粒体肿胀程度降低(P < 0.05);而高强度运动组线粒体Na⁺,K⁺-ATP酶、Ca2⁺-ATP酶活性与正常对照组相比均降低(P < 0.05),线粒体肿胀程度增大(P < 0.05)。实验结果表明中强度运动可以保护线粒体Na⁺,K⁺-ATP酶、Ca2⁺-ATP酶的活性,提高线粒体功能;而高强度运动则降低了线粒体的功能。     

Alterations in in vitro function and protein oxidation of rat sarcoplasmic reticulum Ca2+-ATPase during recovery from high-intensity exercise

The hypothesis tested in this study was that the extent to which sarcoplasmic reticulum (SR) Ca²⁺-ATPase is oxidized would correlate with a decline in its activity. For this purpose, changes in the SR Ca²⁺-sequestering ability and the contents of carbonyl and sulfhydryl groups during recovery after exercise were examined in the superficial portions of vastus lateralis muscles from rats subjected to 5 min running at an intensity corresponding to maximal oxygen uptake (50 m min⁻¹, 10% gradient). A single bout of exercise elicited a 22.4% reduction ( P < 0.05) in SR Ca²⁺-ATPase activity. The decreased activity progressively reverted to normal levels during recovery after exercise, reaching normal levels after 60 min of recovery. This change was paralleled by a depressed SR Ca²⁺-uptake rate, and the proportional alteration in these two variables resulted in no change in the ratio of Ca²⁺-uptake rate to Ca²⁺-ATPase activity. The contents of SR Ca²⁺-ATPase protein and sulfhydryl groups in microsomes were unchanged after exercise and during recovery periods. In contrast, the content of carbonyl groups in SR Ca²⁺-ATPase behaved in an opposite manner to that of SR Ca²⁺-ATPase activity. An approximately 80% augmentation ( P < 0.05) in the carbonyl group content occurred immediately after exercise. The elevated carbonyl content decreased towards normal levels during 60 min of recovery. These results are strongly suggestive that oxidation of SR Ca²⁺-ATPase is responsible, at least in part, for a decay in the SR Ca²⁺-pumping function produced by high-intensity exercise and imply that oxidized proteins may be repaired during recovery from exercise.     

钙离子感受蛋白STIM1与Orai1在钙敏感受体介导钙内流及NO生成中的相互作用

目的:研究Ca~(2+)感受蛋白[基质相互作用分子1(STIM1)与钙释放激活钙通道蛋白1(Orai1)]在人脐静脉内皮细胞(HUVECs)钙敏感受体(Ca SR)介导的钙内流和一氧化氮(NO)生成中的相互作用。方法:将Ca SR激动剂精胺[钙池操纵性钙通道(SOC)和受体操纵性钙通道(ROC)均激活]单用或与ROC模拟剂12-O-十四烷酰佛波醇-13-醋酸酯(TPA)+Ca SR负性变构调节剂Calhex 231(激活ROC、阻断SOC)、蛋白激酶C(PKC)抑制剂Ro 31-8220和PKCα/β1选择性抑制剂Go 6976(激活SOC、阻断ROC)联合孵育HUVECs;利用免疫荧光技术检测HUVECs中STIM1和Orai1的蛋白表达和共定位;免疫共沉淀法检测STIM1和Orai1之间的相互作用;取2~3代HUVECs随机分为特异性的质粒转染组(sh STIM1+sh Orai1组)、空质粒组(vehicle-STIM1+vehicle-Orai1组)和未转染组(control组),将3组细胞分别加入上述4组不同药物刺激,采用荧光探针Fura-2/AM检测HUVECs中Ca~(2+)浓度的变化,NO荧光探针DAF-FM DA负载方法同步检测HUVECs中NO生成的变化。结果:STIM1和Orai1蛋白表达共定位于胞浆,与control组相比,加入Calhex 231+TPA、Ro 31-8220和Go 6976刺激后,STIM1和Orai1在胞浆中的定位均减少,且二者的相互作用均减弱;在4种不同处理因素作用下,sh STIM1+sh Orai1组细胞内Ca~(2+)浓度和NO净荧光强度均明显降低(P0.05)。结论:STIM1与Orai1以二元复合物的形式共同调节Ca SR,并通过激活SOC和ROC介导钙内流及NO生成。     

体外诱导人肾间质成纤维细胞成骨分化

 目的: 观察在体外培养条件下,使用成骨诱导培养液或不同浓度钙离子诱导人肾间质成纤维细胞发生成骨分化的效果,初步探讨肾脏Randall斑形成可能的细胞机制。方法: 体外培养人肾间质成纤维细胞,实验分为5组:成骨诱导组(加成骨诱导液)、CaⅠ组(加0.5 mmol/L Ca²⁺液)、CaⅡ组(加1.5 mmol/L Ca²⁺液)、Ca Ⅲ 组(加2.5 mmol/L Ca²⁺液)和对照组(加PBS)。各组细胞分别培养至第1、3、6、9天时,采用MTT法检测细胞活力;诱导至第9天时,用细胞钙茜素红染色液和钙钴法磷酸酶染色液对各组细胞进行染色,观察钙结节形成和碱性磷酸酶的表达情况。另外,分别采用real-time PCR和Western blot检测各组细胞不同时间点Runt相关转录因子2(Runx2)的 mRNA和蛋白表达水平。 结果: 在1.5 mmol/L 和2.5 mmol/L Ca²⁺浓度条件下细胞的活力明显受抑制。细胞染色结果证实成骨诱导液组和钙离子组均可以见到典型的红色钙结节和黑色块状的硫化钴沉淀物。成骨诱导组细胞Runx2的mRNA和蛋白相对表达量(0~9 d)逐渐升高(P<0.05);诱导至第9天时,3个钙离子组细胞Runx2的mRNA和蛋白表达呈浓度依赖性升高(P<0.05)。结论: 在体外培养环境下,人肾成纤维细胞可以在成骨诱导培养液的刺激作用下发生成骨分化;另外,在高钙离子环境,人肾成纤维细胞也发生了类似的成骨分化。肾乳头组织中的成纤维细胞在高钙离子等因素的作用下发生成骨分化,这可能是肾脏Randall斑形成的细胞学基础。     

Modulation of Ca channels by activation of adenosine A1 receptors in rat striatal glutamatergic nerve terminals

We determined that activation of adenosine A₁ receptors in striatal synaptosomes with 100 nM N⁶-cyclopentyladenosine (CPA) inhibited both the release of endogenous glutamate and the increase of intracellular free Ca²⁺ concentration ([Ca²⁺]_i), due to 4-aminopyridine (4-AP) stimulation, by 28 and 19%, respectively. Furthermore, CPA enhanced the inhibition of endogenous glutamate release due to ω-conotoxin GVIA (ω-Cgtx GVIA), ω-Cgtx MVIIC or ω-Cgtx GVIA plus ω-Cgtx MVIIC. Similar effects were observed in the [Ca²⁺]_i signal. The inhibitory effects of CPA and ω-Cgtx GVIA were additive, but the effects of CPA and ω-Cgtx MVIIC were only partially additive. These results suggest that P/Q-type Ca²⁺ channels and other type(s) of Ca²⁺ channel(s), coupled to glutamate release, are inhibited subsequently to activation of adenosine A₁ receptors.     

Modulation of the dihydropyridine-insensitive Ca influx by 8-bromo-guanosine-3′:5′-monophosphate, cyclic (8-Br-cGMP) in bovine adrenal chromaffin cells

Pretreatment of chromaffin cells with the permeable analogue of cGMP, 8-Br-cGMP (100 μM), leads to a reduction (35%) of depolarization-evoked intracellular calcium concentration ([Ca²⁺]_i) increases. There is evidence that bovine adrenal chromaffin cells are provided with both dihydropyridine-sensitive and -resistant voltage-sensitive Ca²⁺ influx pathways. Combined incubations with nifedipine 10 μM and 8-Br-cGMP reduced KCl-evoked intracellular Ca²⁺ concentration to a greater extent that each compound separately. Moreover, 8-Br-cGMP failed to affect the [Ca²⁺]_i transient induced by the L-type Ca²⁺ channel agonist Bay K 8644 (1μM) under conditions of low depolarization. Neomycin (0.2 mM) and θ-Aga Toxin-IVA (AgTx) (1μM) inhibited the calcium transient to a similar extent, and this inhibition was not enhanced by the presence of 8-Br-cGMP. It is concluded that 8-Br-cGMP modulated the dihydropyridine-insensitive Ca²⁺ influx pathway in the chromaffin cell.     

P2 receptors in satellite glial cells in trigeminal ganglia of mice

There is strong evidence for the presence of nucleotide (P2) receptors in sensory neurons, which might play a role in the transmission of pain signals. In contrast, virtually nothing is known about P2 receptors in satellite glial cells (SGCs), which are the main glial cells in sensory ganglia. We investigated the possibility that P2 receptors exist in SGCs in murine trigeminal ganglia, using Ca²⁺ imaging, patch-clamp recordings, and immunohistochemistry. We found that ATP caused an increase in intracellular Ca²⁺ concentration ([Ca²⁺]_i) in SGCs. As adenosine had no effect on [Ca²⁺]_i, and the P2 receptor antagonist pyridoxalphosphate-6-azophenyl-2′,4′-disulfonic acid largely blocked the response to ATP we conclude that P1 receptors did not contribute to the responses. We obtained the following evidence that the responses to ATP were mediated by metabotropic P2Y receptors: (i) persistence of the responses in Ca²⁺-free solution, (ii) inhibition of the response by cyclopiazonic acid, (iii) [Ca²⁺]_i increases in response to the P2Y agonists uridine triphosphate, adenosine thiodiphosphate, and 2-methylthio ADP, and (iv) failure of the P2X agonist ,β-methylene ATP to elicit a response. Agonists of P2Y₁ receptors and uridine triphosphate, an agonist at P2Y₂ and P2Y₄ receptors, induced [Ca²⁺]_i increases suggesting that at least these P2Y receptor subtypes are present on SGCs. Using an antibody against the P2Y₄ receptor, we found immunopositive SGCs. Patch-clamp recordings of SGCs did not reveal any inward current due to ATP. Therefore, there was no evidence for the activation of ionotropic P2X receptors under the present conditions.

The results indicate the presence of functional nucleotide (P2Y) receptors in SGCs.     

Role of Ca-dependent K channels in the membrane potential and contractility of uterus from spontaneously hypertensive rats

Membrane potentials and the contractile responses to prostaglandin F₂, oxytocin, bradykinin, 5-hydroxytryptamine and acetylcholine were determined in uteri from spontaneously hypertensive rats (SHR) and normotensive Wistar Kyoto rats (WKY), both in the absence and in the presence of the Ca²⁺-dependent K⁺ channel blocker apamin. Compared to WKY, the uterine smooth muscle from SHR was more reactive to all the agonists tested. In the presence of apamin, the contractile responses of WKY uteri were enhanced, whereas the responses of SHR uteri were either unaltered or decreased, so that the WKY responses became similar or even greater than those of SHR. Smooth muscle cell membrane potentials were lower in SHR than in WKY. Treatment with apamin did not affect the membrane potential of SHR cells, but caused depolarization of WKY cells. It is concluded that a reduced activity of Ca²⁺-dependent K⁺ channels in SHR uteri contributes to their lower membrane potentials and higher responsiveness to contractile stimuli by agonists, It is possible that uterine quiescence, essential for the normal fetal development, is impaired in SHR, resulting in lower uterine blood fluxes with consequent lower fetal and litter size and weight observed in this strain.     

The angiotensin receptor type 1-Gq protein-phosphatidyl inositol phospholipase Cbeta-protein kinase C pathway is involved in activation of proximal tubule Na+-ATPase activity by angiotensin(1-7) in pig kidneys

In a previous study, we observed that angiotensin(1–7) (Ang(1–7)) stimulates proximal tubule Na⁺-ATPase activity through the angiotensin receptor type 1 (AT₁R). Here we aimed to study the signalling pathways involved. Our results show that the stimulatory effect of Ang(1–7) on Na⁺-ATPase activity through AT₁R involves a G_q protein–phosphatidyl inositol-phospholipase Cβ(PI-PLCβ) pathway because: (1) the effect was reversed by GDPβS, a non-hydrolysable GDP analogue, and by a monoclonal G_q protein antibody; (2) the effect was similar and not additive to that of GTPγS, a non-hydrolysable GTP analogue; (3) Ang(1–7) induced a rapid decrease (30 s) in phosphatidylinositol 4,5-bisphosphate levels, a PI-PLCβ substrate; and (4) U73122, a specific inhibitor of PI-PLCβ, abolished Ang(1–7)-induced stimulation of Na⁺-ATPase activity. Angiotensin(1–7) increased the protein kinase C (PKC) activity similarly to phorbol-12-myristate-13-acetate (PMA), an activator of PKC. This effect was reversed by losartan, a specific antagonist of AT₁R. The stimulatory effects of Ang(1–7) and PMA on Na⁺-ATPase activity are similar, non-additive and reversed by calphostin C, a specific inhibitor of PKC. A catalytic subunit of PKC (PKC-M) increased the Na⁺-ATPase activity. These data show that Ang(1–7) stimulates Na⁺-ATPase activity through the AT₁R–G_q protein–PI-PLCβ–PKC pathway. This effect is similar to that described for angiotensin II, showing for the first time that these compounds could have similar effects in the renal system.     

The physiological impact of proinsulin C-peptide

The proinsulin C-peptide fulfills an important function in the biosynthesis of insulin by facilitating the formation of the correct secondary and tertiary structure of the hormone. C-peptide and insulin are released in equimolar amounts to the circulation but C-peptide has generally been considered to be biologically inert. However, recent studies indicate that C-peptide administration to type 1 diabetes patients is accompanied by improved renal function, amelioration of autonomic dysfunction, stimulated tissue glucose utilization and augmented skin and muscle blood flow. The cellular mechanisms underlying these effects may be related to C-peptide’s capacity to stimulate both Na⁺, K⁺-ATPase and endothelial nitric oxide synthase (eNOS). The possibility that C-peptide may be beneficial in the long term treatment of patients with type 1 diabetes should be considered.     

Disruption of ionic and cell volume homeostasis in cerebral ischemia: The perfect storm

The mechanisms of brain tissue damage in stroke are strongly linked to the phenomenon of excitotoxicity, which is defined as damage or death of neural cells due to excessive activation of receptors for the excitatory neurotransmitters glutamate and aspartate. Under physiological conditions, ionotropic glutamate receptors mediate the processes of excitatory neurotransmission and synaptic plasticity. In ischemia, sustained pathological release of glutamate from neurons and glial cells causes prolonged activation of these receptors, resulting in massive depolarization and cytoplasmic Ca²⁺ overload. High cytoplasmic levels of Ca²⁺ activate many degradative processes that, depending on the metabolic status, cause immediate or delayed death of neural cells. This traditional view has been expanded by a number of observations that implicate Cl⁻ channels and several types of non-channel transporter proteins, such as the Na⁺,K⁺,2Cl⁻ cotransporter, Na⁺/H⁺ exchanger, and Na⁺/Ca²⁺ exchanger, in the development of glutamate toxicity. Some of these ion transporters increase tissue damage by promoting pathological cell swelling and necrotic cell death, while others contribute to a long-term accumulation of cytoplasmic Ca²⁺. This brief review is aimed at illustrating how the dysregulation of various ion transport processes combine in a ‘perfect storm’ that disrupts neural ionic homeostasis and culminates in the irreversible damage and death of neural cells. The clinical relevance of individual transporters as targets for therapeutic intervention in stroke is also briefly discussed.