Metformin alleviates vascular calcification induced by vitamin D3 plus nicotine in rats via the AMPK pathway

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牛磺酸对维生素D_3和尼古丁诱导的大鼠钙化血管精氨酸/NO途径的影响

目的　观察钙化血管精氨酸 /NO途径的改变和外源性牛磺酸 (taurine,Tau)对血管钙化及精氨酸 /NO途径的影响。方法　维生素D3 (vitaminD3 ,VitD3 )肌注和尼古丁(nicotine ,Nic)灌胃制备大鼠血管钙化模型 ,检测血管钙含量和血管碱性磷酸酶 (ALP)活性、血浆精氨酸和亚硝酸盐(NO-2 )含量、血管环磷酸鸟苷 (cGMP)含量、血管一氧化氮合酶 (NOS)活性及血管精氨酸 (arginine ,Arg)转运。结果　钙化组 (VDN)大鼠血管钙含量较对照组升高 6 6倍 (P <0 0 1) ,血管ALP活性高 12 6倍 (P <0 0 1) ;血浆NO生成减少 ,血管cGMP水平降低 ,血管NOS活性增高 ,其中cNOS活性高 18 9% (P <0 0 1) ,但精氨酸转运在血管平滑肌和内皮明显减少 (- 6 3 8% ,- 5 5 2 % ,P <0 0 1)。口服牛磺酸治疗的大鼠较单纯VDN组 ,血管钙含量降低 4 9 5 % (P <0 0 1) ,血管ALP活性明显降低 ,血浆NO生成增加 ,血管cGMP含量增加 2 9 1% (P均 <0 0 1) ,血管精氨酸转运在血管平滑肌和内皮分别增加 79 4 %和 5 5 1% (P <0 0 1)。结论　给予外源性牛磺酸可以减轻VitD3 加Nic大鼠的血管钙化程度 ,改善钙化血管L Arg/NOS/NO/cGMP途径紊乱 ;提示牛磺酸对防治动脉粥样硬化等疾病的血管钙化可能具有潜在临床意义     

维生素D_3联合尼古丁诱导的血管钙化亚慢性模型的建立

目的建立维生素D3(vitamin D3)联合尼古丁(nico-tine)诱导的大鼠血管钙化亚慢性动物模型。方法采用维生素D3(300 kIU.kg-1)肌注和尼古丁(25 mg.kg-1,5 ml.kg-1)灌胃制备大鼠血管钙化亚慢性模型,定期测定大鼠体重和尾动脉血压;在造模的第8周、第16周,分别测定大鼠心脏系数、血浆钙、磷含量、血管钙含量、血管碱性磷酸酶(alkaline phosphatase,ALP)活性和血管钙沉积等与血管钙化相关的指标,对胸主动脉进行Von Kossa染色。结果第3~8周,VDN(vitamin D3and nicotine)大鼠尾动脉压持续增加,与对照组相比差异均有显著性(P均<0.05)。第9周后,仍然维持在较高水平。与对照组相比,VDN组大鼠第8周心脏系数升高8.4%(P<0.01);血钙、血磷分别升高3.2%(P<0.01)和12.0%(P<0.05);血管组织钙含量升高27.0%(P<0.01);血管ALP活性与血管钙沉积分别高151.8%和68.8%(P均<0.01)。与对照组相比,第16周VDN组大鼠心脏系数升高7.7%(P<0.01),血钙升高3.2%(P<0.05),血磷降低1.4%;血管组织钙含量升高51.0%(P<0.01);血管ALP活性与血管钙沉积分别高87.2%(P<0.01),31.6%(P<0.05)。Von Kossa染色结果显示第8周和第16周VDN组大鼠胸主动脉中膜均有大量深色颗粒沉积。结论维生素D3和尼古丁诱导的16 wk大鼠血管钙化亚慢性模型可作为药物筛选和机制研究的疾病模型。     

Characteristics of tail-tremor induced by nicotine in rats

To characterize the tail-tremor and locomotor hyperactivity induced by repeated nicotine administration, the effects of nicotinic, -adrenergic and dopaminergic blockers were investigated in rats. Daily administration of nicotine (0.5 mg/kg, s.c.) induced tail-tremor from the 4th day, which became more marked in intensity by subsequent administration. Locomotor hyperactivity was also induced by nicotine, which was enhanced by daily administration. The tail-tremor and locomotor hyperactivity induced by repeated nicotine administration were inhibited by mecamylamine (0.1–1 mg/kg, i.p.) but not by hexamethonium (0.5 and 1 mg/kg, i.p.). Clonidine (0.02 and 0.04 mg/kg, i.p.) and prazosin (0.5 and 1 mg/kg, i.p.) reduced tail-tremor more markedly than hyperactivity. However, haloperidol (0.05–0.2 mg/kg, i.p.) and chlorpromazine (1–5 mg/kg, i.p.) reduced hyperactivity more markedly than tail-tremor. These results suggest that nicotine-induced tail-tremor and hyperactivity are due to an increased susceptibility of central nicotinic receptors of nicotine followed by catecholaminergic mechanisms, and that tail-tremor may be more associated with the noradrenergic system than the dopaminergic system. Correspondence to: Y. Gomita at the above address     

无机化合物CANA对大鼠血管钙化模型的改善作佑

目的:制备大鼠血管钙化模型并观察枸橼酸盐结构类似的无机化合物CANA对血管钙化的作用.方法:用肌注维生素D,和口服尼古丁诱导大鼠血管钙化模型.4周后,进行血管组织Yon Kossa染色,测定大鼠血管钙含量,碱性磷酸酶(ALP)活性、血磷、血钙含量以及心脏系数.结果:模型组大鼠血管VonKossa染色见平滑肌细胞内和细胞外基质有大量银染的深紫色颗粒聚集,主动脉血管钙含量以及ALP活性分别较对照组高66.9%(P＜0.001)和462.6%(P＜0.001),血磷和血钙含量及心脏系数较对照组比较无明显变化(P＞0.05);无机化合物CANA 740 mg·kg-1治疗组血管Von Kossa染色的深紫色颗粒有所减少,主动脉血管钙含量及ALP活性分别较模型组降低38.2%(P＜0.001)和22.0%(P＜0.05).结论:用维生素D3和尼古丁制备模型4周后,可造成血管组织钙化,但血磷和血钙含量及心脏系数无明显改变.无机化合物CANA对该血管钙化模型有改善作用.     

Hydrogen sulfide ameliorates disorders in the parafacial respiratory group region of neonatal rats caused by prenatal cigarette smoke exposure via an antioxidative effect

We previously found that hydrogen sulfide (H₂S) ameliorated the dysfunction of central chemoreception caused by prenatal cigarette smoke exposure (CSE). In the present study, we further explored whether the parafacial respiratory group (pFRG) is involved in the protection of central chemoreception by H₂S against prenatal CSE-induced injury. We found that NaHS, a donor of H₂S, restored the expression of Phox2b, which was downregulated by prenatal CSE, in the pFRG region of neonatal rats. NaHS also relieved the prenatal CSE-induced excitatory synapse disturbance in the pFRG region of neonatal rats. Additionally, NaHS prevented the increase in the malondialdehyde level and suppression of antioxidase activity in the pFRG region of neonatal rats induced by prenatal CSE. Furthermore, NaHS prevented the downregulation of the expression of antioxidases and Nrf2 in the pFRG region of neonatal rats with prenatal CSE. These results suggest that H₂S can protect the pFRG of neonatal rats against prenatal CSE-induced injury via an antioxidative effect.     

A comparative study of antioxidants S-allyl cysteine sulfoxide and vitamin E on the damages induced by nicotine in rats

The dietary consumption of antioxidant-rich fruits and vegetables is inversely correlated with the incidence of various diseases like cardiovascular diseases and lung cancer. We have tried to find out how far the S-allyl cysteine sulfoxide (SACS) isolated from garlic (Allium Sativum L.) can combat the nicotine-induced peroxidative damage in rats. The effects have been compared with the standard antioxidant vitamin E. Administration of SACS or vitamin E (100 mg/kg) to nicotine (0.6 mg/kg) treated rats for 21 days showed decreased concentrations of thiobarbituric acid reactive substances, hydroperoxides, and conjugated dienes in liver, lungs, and heart as compared with the values found in rats treated with nicotine alone. The activities of catalase and superoxide dismutase increased. The levels of the antioxidants like vitamins A, C, and E in the liver and glutathione in all tissues increased significantly in SACS-treated or vitamin E fed rats. However, the antioxidant status was higher when vitamin E was administered as compared with SACS administered to nicotine-treated rats.     

Drug actions in rats with arteriosclerosis induced by toxic doses of vitamin D2.

The vascular response of isolated perfused hind legs from normal and arteriosclerotic rats to noradrenaline, ATP, PGF2 alpha and vasopressin was determined. The increase of perfusion pressure to noradrenaline and ATP was reduced and that to PGF2 alpha and vasopressin was enhanced in arteriosclerotic rats in comparison to normal animals. The results indicate that the reactivity of the vascular smooth muscle of isolated hind legs of arterioslerotic rats is not only quantitatively but also qualitatively different in comparison with normal rats.     

Hydrogen sulfide facilitates carotid sinus baroreflex in anesthetized rats

AIM: To study effects of hydrogen sulfide (H2S) on the carotid sinus baroreflex (CSB). METHODS: The functional curve of the carotid sinus baroreflex was measured by recording changes in arterial pressure in anesthetized male rats with perfused carotid sinus. RESULTS: H2S (derived from sodium hydrosulfide) at concentrations of 25, 50, and 100 micromol/L facilitated the CSB, shifting the functional curve of the baroreflex downward and to the left. There was a marked increase in peak slope (PS) and reflex decrease in blood pressure (RD). Effects were concentration-dependent. Pretreatment with glibenclamide (20 micromol/L), a K(ATP) channel blocker, abolished the above effects of H2S on CSB. Pretreatment with Bay K8644 (an agonist of calcium channels; 500 nmol/L) eliminated the effect of H2S on CSB. An inhibitor of cystathionine gamma-lyase (CSE), DL-propargylglycine (PPG; 200 micromol/L), inhibited CSB in male rats and shifted the functional curve of the baroreflex upward and to the right. CONCLUSION: These data suggest that exogenous H2S exerts a facilitatory role on isolated CSB through opening K(ATP) channels and further closing the calcium channels in vascular smooth muscle. Endogenous H2S may activate the activity of the CSB in vivo.     

大鼠维生素D心肌损伤模型中VEGF的免疫组化研究

目的:探讨维生素D心肌损伤模型中血管内皮生长因子(vascular endothelial growth factor, VEGF)和蛋白的表达情况. 方法:大鼠维生素D灌胃,制作心肌损伤模型,制作心室肌病理切片,用免疫组化方法检测对照组和实验组VEGF蛋白的表达情况. 结果: 免疫组化结果显示,维生素D心肌损伤模型组VEGF蛋白表达较对照组显著增高. 结论:维生素D致心肌损伤可导致VEGF蛋白的表达水平增高.     

Hydrogen sulfide: a novel signaling molecule in the vascular system

Hydrogen sulfide (H2S) is an endogenous gasotransmitter produced in mammalian cells. It is responsible for physiological functions in many organs and systems, with attention focused mainly on the cardiovascular and nervous systems. In the vascular system, H2S produces biphasic effects in regulation of vascular tone. At lower concentrations, it induces vasoconstriction predominantly via decreasing cyclic adenosine monophosphate in smooth muscle cell and inhibiting the production and bioavailability of nitric oxide. At higher concentrations, it produces vasorelaxation mainly through opening of KATP channels and induction of intracellular acidification. Scavenging reactive oxygen species and elevation of cyclic guanosine monophosphate are also implicated in the vasorelaxant response. This review presents an overview of the current knowledge of H2S in the vascular system, with special emphasis and discussion on the involvement of various signaling pathways and ion channels based on current understanding and reported literature till date.     

Antagonism of a nicotine plus midazolam discriminative cue in rats

Rats were trained to discriminate nicotine (0.4mg/kg s.c.), midazolam (0.2mg/kg s.c.) or the combination of these drugs from saline (n = 10). The rats were trained to 95% accuracy in a two-bar operant procedure with a tandem schedule of food reinforcement. Testing with the individual drugs in the mixture-trained group showed that nicotine (85% drug-appropriate responding) was a more salient component than midazolam (47%) in the compound stimulus. The rats were tested with benzodiazepine and nicotine antagonists individually and in combination (mecamylamine 0.2-1.6mg/kg s.c.; flumazenil 2.5-20mg/kg i.p.). Results for the mixture-trained animals showed that flumazenil had no effect on its own, however mecamylamine on its own produced a significant but incomplete block in doses of 0.4-1.6mg/kg. The greater salience of the nicotine component of the cue would explain the block by mecamylamine but not flumazenil. The antagonists in combination produced greater blockade than mecamylamine on its own. The selectivity of the antagonist actions on the different cue components was also demonstrated. The results suggest that in drug discrimination experiments, "false negative" results may be obtained with antagonists when a training drug produces a stimulus with more than one component.     

Vitamin D ameliorates fluoride-induced embryotoxicity in pregnant rats

We have evaluated the ameliorative effect of vitamin D on fluoride-induced embryotoxicity in pregnant rats. Oral administration of sodium fluoride (NaF; 40 mg/kg body weight) from days 6 to 19 of gestation caused, as compared with control, significantly lowered body weight, feed consumption, absolute uterine weight and number of implantations. As compared with the control, higher incidence of skeletal (presence of wavy ribs, 14th rib, dumbbell-shaped 5th sternebrae, incomplete ossification of skull) and visceral (subcutaneous haemorrhage) abnormalities was recorded in the foetuses of fluoride-treated pregnant rat. Vitamin D (2 ng/0.2 ml olive oil/animal/day po) treatment significantly ameliorated the fluoride-induced reductions in body weight, feed consumption and absolute uterine weight. As compared with fluoride-treated alone, the total percentage of skeletal and visceral abnormalities observed in foetuses was significantly lowered in fluoride plus vitamin D-treated animals. These findings suggest that vitamin D treatment significantly reduced the severity and incidence of fluoride-induced embryotoxicity. The ameliorative effect of vitamin D against skeletal and visceral abnormalities could be due to stimulation of intestinal absorption of calcium and phosphate, thus raising the plasma calcium and phosphate concentrations.     

Effects of vitamin D derivatives on soft tissue calcification in neonatal and calcium mobilization in adult rats

The activity of 18 vitamin D analogs on soft tissue calcification and growth impairment in neonatal rats and their effect on bone calcium mobilization, intestinal calcium absorption and binding to intestinal 1,25-dihydroxyvitamin D₃ receptors in adult rats were compared. Depending on the chemical modification of the vitamin D parent compounds, they could be separated into active and inactive analogs. Cholecalciferol and ergocalciferol were similarly active, but epimerization of ergocalciferol at carbon 23 caused loss of activity. Hexafluorination at carbon 26 and 27 and the introduction of a double bond at carbon 22 or 23 had no or little effect on the activity. The loss of activity was caused by the introduction of a triple bond at carbon 23 and by hydroxylation at carbon 23, 26 or 28. The differentiation of human promyelocytic leukemia cells (HL-60) induced by these derivatives was used as a parameter for antitumour activity. All six analogs, which markedly affected calcium metabolism, were highly active in HL-60 cells. However, at least three derivatives were highly active in the antitumour test but failed to induce hypercalcemia. Thus, these results indicate that it could be possible to develop medically useful vitamin D derivatives devoid of hypercalcemic side-effects.     

Lobeline attenuates locomotor stimulation induced by repeated nicotine administration in rats

Lobeline inhibits [3H]nicotine binding to rat brain membranes and nicotine-induced [3H]dopamine release from superfused rat striatal slices, indicating that lobeline acts as a nicotinic receptor antagonist. To determine whether lobeline also inhibits the effects of nicotine in vivo, the present study assessed the effect of lobeline pretreatment on nicotine-induced hyperactivity and sensitization. For 12 consecutive days, rats were injected subcutaneously with lobeline (3 mg/kg) or saline, followed 10 min later by nicotine (0.3 mg/kg) or saline injection, and activity was monitored. To determine if lobeline inhibits induction of sensitization to nicotine, 1 or 28 days later, rats were pretreated with saline followed by nicotine or saline. Lobeline attenuated nicotine-induced hyperactivity when both drugs were administered repeatedly. Although an initial injection of lobeline produced hypoactivity, tolerance to this effect developed. Importantly, tolerance did not develop to the lobeline-induced attenuation of nicotine hyperactivity. Lobeline attenuated the induction of sensitization to nicotine 1 day, but not 28 days, after the cessation of lobeline treatment. These results demonstrate that systemic administration of lobeline attenuates the locomotor-activating effects of repeated nicotine injection and the sensitization to nicotine, consistent with lobeline inhibition of nicotinic receptors and/or neurotransmitter transporters.     

Effect of treatment with vitamin D plus calcium on oxidative stress in streptozotocin-induced diabetic rats

Background

In diabetes mellitus, uncontrolled hyperglycemia has been reported to induce oxidative stress, which may lead to health complications. Vitamin D, however, acts as a non-enzymatic antioxidant to protect cells against oxidative stress and damage.

Tripterygium wilfordii polyglycosidium ameliorates pouchitis induced by dextran sulfate sodium in rats

BackgroundThe aim of this study was to investigate the therapeutic effects of Tripterygium wilfordii polyglycosidium (TWP) to rats with dextran sulfate sodium (DSS)-induced pouchitis and its possible mechanism.MethodsSprague-Dawley rats underwent surgery of ileal pouch anal anastomosis (IPAA) and pouchitis was induced by DSS. Rats were randomly divided into no intervention (NI), normal saline (NS) and TWP groups. Rats were lavaged with normal saline (3 ml/day in NS group) or TWP (12 mg/kg/day in TWP group) for 7 days. General conditions of animals and histopathological examinations were evaluated. Interleukin (IL)-1β, IL-6, IL-10, and tumor necrosis factor (TNF)-α mRNA expression was measured. Levels of occludin and Zo-1 proteins were measured by immunohistochemistry. In addition, ALT and AST were assessed.ResultsTWP significantly attenuated the symptoms of pouchitis characterized by body weight loss, diarrhea, and bloody stool. Furthermore, TWP diminished histological damage compared with other groups. There was a significant reduction in levels of IL-1β, IL-6, and TNF-α, as well as an increase in IL-10 in the TWP group. The expression of tight junction proteins occludin and Zo-1 were increased in the TWP group. There were no statistical differences in serum ALT and AST among the three groups.ConclusionsTWP significantly ameliorated pouchitis and inhibited the production of IL-1β, IL-6, and TNF-α as well as increased the levels of IL-10, occludin, and Zo-1 protein in rats. These findings suggest TWP might be a potential therapeutic agent for patients with pouchitis.     

5HT3 receptor antagonists do not block nicotine induced hyperactivity in rats

Male Wistar rats with continuous access to 6% ethanol solution and water in their home cages were subjected to food restriction (FR). Reduction of body weight to 80% of normal was associated with a significant increase in ethanol drinking. It is known that the stress of FR gives rise to increased corticosterone secretion, and in line with these findings it was found that the weight of the thymus (whose size is inversely related to corticosterone levels) was reduced to 55% of normal in the present FR rats. Two subsequent experiments indicated that this adrenal activation contributed to the FR-induced enhancement of alcohol drinking. Firstly, adrenalectomized rats showed no evidence of enhanced alcohol drinking during food restriction, suggesting that adrenal corticosterone hypersecretion contributes to the enhanced ethanol consumption during FR. Secondly, treatment of FR rats with the enzyme inhibitor cyanoketone, which blocks stress-induced but not basal corticosterone secretion, at least partly prevented the FR-induced increase in ethanol drinking. These results add further evidence that sustained exposure to corticosterone facilitates ethanol consumption in the rat.     

Hydrogen sulfide alleviates apoptosis and autophagy induced by beryllium sulfate in 16HBE cells

Beryllium and its compounds are systemic toxicants that are widely applied in many industries. Hydrogen sulfide has been found to protect cells. The present study aimed to determine the protective mechanisms involved in hydrogen sulfide treatment of 16HBE cells following beryllium sulfate-induced injury. 16HBE cells were treated with beryllium sulfate doses ranging between 0 and 300 μM BeSO₄. Additionally, 16HBE cells were subjected to pretreatment with either a 300 μM dose of sodium hydrosulfide (a hydrogen sulfide donor) or 10 mM DL-propargylglycine (a cystathionine-γ-lyase inhibitor) for 6 hr before then being treated with 150 μM beryllium sulfate for 48 hr. This study illustrates that beryllium sulfate induces a reduction in cell viability, increases lactate dehydrogenase (LDH) release, and increases cellular apoptosis and autophagy in 16HBE cells. Interestingly, pretreating 16HBE cells with sodium hydrosulfide significantly reduced the beryllium sulfate-induced apoptosis and autophagy. Moreover, it increased the mitochondrial membrane potential and alleviated the G2/M-phase cell cycle arrest. However, pretreatment with 10 mM DL-propargylglycine promoted the opposite effects. PI3K/Akt/mTOR and Nrf2/ARE signaling pathways are also activated following pretreatment with sodium hydrosulfide. These results indicate the protection provided by hydrogen sulfide in 16HBE cells against beryllium sulfate-induced injury is associated with the inhibition of apoptosis and autophagy through the activation of the PI3K/Akt/mTOR and Nrf2/ARE signaling pathways. Therefore, hydrogen sulfide has the potential to be a promising candidate in the treatment against beryllium disease.     

硫化氢通过抗氧化作用改善脑缺氧导致的小鼠空间记忆障碍(英文)

目的观察外源性H2S对缺氧性脑损伤小鼠空间学习记忆障碍的影响,并探究其作用机制。方法连续4 d sc给予NaNO2 120 mg.kg-1.d-1制备缺氧模型;氢硫化钠(NaHS)治疗组在制备模型的同时ip给予NaHS 1 mg.kg-1.d-1。每天给药前进行Morris水迷宫实验,测定逃避潜伏期、原平台象限停留时间和穿越平台次数。比色法检测小鼠脑组织中超氧化物歧化酶(SOD)活性及丙二醛(MDA)含量。HE染色观察海马组织切片CA1区神经元形态学改变。结果水迷宫实验第3天和第4天,模型组小鼠逃避潜伏期分别为(26.0±7.3)s和(23.3±8.7)s,明显长于正常对照组的(16.1±9.6)s(P<0.05)和(11.1±6.2)s(P<0.01)。第5天,模型组小鼠穿越平台次数为4.1±1.9,在原平台象限停留时间为(20±8)s,与正常对照组穿越平台次数(7.2±1.6)次和在原平台象限停留时间(28±8)s比较明显减少(P<0.01)。与正常对照组相比,模型组小鼠脑组织中SOD活性降低12.6%(P<0.01),MDA含量升高43.9%(P<0.01)。在模型组小鼠海马CA1区,锥体细胞出现明显的核固缩、胞浆深染和排列紊乱等变性改变。与模型组比较,NaHS组小鼠在水迷宫实验的第3天和第4天逃避潜伏期明显缩短(P<0.05),分别为(17.9±7.0)s和(15.8±8.5)s;在平台所在象限停留时间和穿越平台次数明显增加(P<0.01),分别为(30±9)s和(6.7±2.5)次;SOD活性升高了8.9%(P<0.05),MDA含量显著下降了29.6%(P<0.01);海马CA1区神经元变性改变较模型组得到显著缓解。结论 NaHS减轻了脑缺氧损伤诱发的小鼠学习记忆的损害,其作用机制可能与H2S衰减海马区神经元损伤和抗氧化作用有关。