Acyl,pseudotetra-, tri- and dipeptide active-core analogs of insect neuropeptides

Pseudopeptides of the achetakinin insect neuropeptide family were synthesized by replacing the amino acid blocks Phe-, Phe-Tyr-, and Phe-Tyr-Pro- of the active-core pentapeptide Phe-Tyr-Pro-Trp-GIy-NH₂ with hydrocinnamic acid, 6-phenylhexanoic acid, and both 9-phenylnonanoic and 6-phenylhexanoic acid, respectively. All four of these analogs retained myotropic activity, demonstrating that the active core could be reduced from a pentapeptide to a modified dipeptide. Most notable of these was the pseudotetrapeptide hydrocinnamyl-Tyr-Pro-Trp-Gly-NH₂, which retained 70% of the potency and over 85% of the maximal activity of the parent pentapeptide. The N-terminal amino group, the phenol ring of the Tyr residue, the sulfate moiety and the Gly residue of the insect sulfakinin active core Tyr(SO₃H)-Gly-His-Met-Arg-Phe-NH₂ were all replaced by dodecanedioic acid. The resulting pseudotetrapeptide, dodecandioyl-His-Nle-Arg-Phe-NH₂, elicited myostimulatory activity. Conversely, the related acyl pseudopentapeptide azelayl-Gly-His-Nle-Arg-Phe-NH₂ proved myoinhibitory. A possible explanation for these disparate biological responses is discussed. These acyl pseudopeptides are important advances towards the eventual development of stable, potent mimetic agonists and antagonists of insect neuropeptides.     

An active pseudopeptide analog of the leucokinin insect neuropeptide family

A pseudopeptide analog of the active core of the leucokinin insect neuropeptide family was synthesized and found to retain myotropic activity. No reports of active pseudopeptide analogs of an insect or other invertebrate neuropeptide have previously appeared in the literature. The pseudopeptide (Pheψ[CH₂-NH] Phe-Ser-Trp-Gly-NH₂) contains a reduced-amide linkage between the two N-terminal Phe residues. Unlike its amide-bond containing counterpart, the activity of the pseudopeptide was not destroyed upon exposure to aminopeptidase M.     

Effect of repeated doses of netazepide,a gastrin receptor antagonist,omeprazole and placebo on 24 h gastric acidity and gastrin in healthy subjects

Aim

To administer repeated oral doses of netazepide to healthy subjects for the first time, to assess safety, tolerability, pharmacokinetics and effect on 24 h gastric pH and plasma gastrin.

Method

We did two randomized, double-blind, parallel group studies. The first compared netazepide 25 and 100 mg 12 hourly, omeprazole 20 mg once daily and placebo for 7 days. On day 7 only, we measured pH and assayed plasma gastrin. The second study compared netazepide 5, 10 and 25 mg and placebo once daily for 14 days. We measured pH on days 1, 7 and 14 and assayed plasma gastrin on days 1 and 14. We compared treatments by time gastric pH ≥ 4 during 0–4, 4–9, 9–13 and 13–24 h after the morning dose, and by plasma gastrin. P < 0.05 was significant.

Results

Netazepide was well tolerated. On day 7 of the first study, netazepide increased pH significantly only during 9–13 h after the 100 mg dose, whereas omeprazole raised pH significantly during all periods. Both netazepide and omeprazole increased plasma gastrin significantly. Netazepide had linear pharmacokinetics. In the second study, netazepide caused dose-dependent, sustained increases in pH on day 1, but as in the first study, netazepide had little effect on pH on days 7 and 14. Again, netazepide increased plasma gastrin significantly.

Conclusion

Although repeated doses of netazepide led to tolerance to its effect on pH, the accompanying increase in plasma gastrin is consistent with continued inhibition of acid secretion, via gastrin receptor antagonism and gene up-regulation.     

Synthesis of analogues of the Des-Phe-NH2C-terminal hexapeptide of cholecystokinin showing gastrin antagonist activity

Four analogues of Z-CCK-27–32-NH₂, Z-Tyr(SO₃-)-Met-Gly-Trp-Met-Asp-NH₂, a cholecystokinin receptor antagonist have been synthesized by solution methodology. In these analogues, Z-Tyr(SO₃-)-Nle-Gly-Trp-Met-Asp-NH₂ 16, Z-Tyr(SO₃-)-Nle-Gly-Trp-Nle-Asp-NH₂ 17, BOC-Tyr(SO₃-)-Met-Gly-Trp-Met-Asp-NH₂ 24 and Boc-Tyr(SO₃-)-Met-Gly-Trp-Nle-Asp-NH₂ 25 methionyl residues were replaced by norleucyl residues. Preliminary biological activity on gastrin-induced acid secretion, in rat, are reported. These derivatives proved to antagonize the action of gastrin, with ED 50 of between 0.5 and 3 mg/kg.     

菊粉硫酸酯的制备及其抑真菌活性研究

目的研究菊粉硫酸酯(inulin sulfate,IS)的制备及其抑制真菌活性。方法采用氯磺酸-吡啶法对菊粉(inulin)进行硫酸化修饰,得到IS;采用生长速率法对IS的抑制真菌活性进行测定。结果IS产率为菊粉投料量的128%左右,水溶性好,呈中性,硫取代度(DS)为2.22左右,硫含量为18.2%左右。结论抑菌实验表明IS具有一定抑制真菌的活性。     

胃泌素对人结肠癌细胞Colo320WT中β-catenin/Tcf-4通路的影响

目的探讨胃泌素对结肠癌细胞Colo320WT中β-catenin/Tcf-4通路影响。方法10-8mol·L-1胃泌素分别于0、1、6、12、24、48h时间点干预Colo320WT细胞,同时先用10-6mol·L-1胃泌素受体拮抗剂L365,260预干预Co-lo320WT细胞30min,再加10-8mol·L-1胃泌素干预至12h。采用免疫共沉淀和免疫印迹检测Colo320WT中TX-100可溶性部分(胞质部分)和不可溶部分(细胞骨架结合部分)中β-catenin的表达及β-catenin/Tcf-4复合物的表达。免疫印迹观察c-Myc和cyclinD1的表达。结果TX-100可溶性部分中β-catenin的量随着胃泌素干预时间呈递减趋势,12h达最小;而TX-100不溶解部分却随时干预时间的呈递增趋势,12h达最大。免疫共沉淀发现胃泌素干预后β-catenin/Tcf-4复合物表达量明显增多。免疫印迹检测到胃泌素干预下的细胞中c-Myc和cyclinD1表达明显增加。胃泌素受体拮抗剂阻断后,则可阻断胃泌素引起的上述效应。结论胃泌素与其受体作用后使结肠癌细胞中β-catenin分布发生转移和活化β-catenin/Tcf-4通路来降低结肠癌细胞的黏附,增加肿瘤细胞的侵袭和转移。     

纤维素硫酸酯体内抗凝血活性的实验研究

目的:研究纤维素硫酸酯对动物体内的抗凝血活性。方法:用试管法测凝血时间(CT),用凝固法测定活化部分凝血活酶时间(APTT)、凝血酶原时间(PT)和凝血酶时间(TT),以肝素和硫酸化右旋糖酐为阳性对照,考察其抗凝血活性的量效性和时效性。结果:纤维素硫酸酯能显著延长CT,APTT和TT,且呈剂量依赖性,但在所试剂量范围内对PT无明显影响;其抗凝血活性在给药2h时达到最大。结论:纤维素硫酸酯具有明显抗凝血作用,强于现有同类抗凝血药硫酸化右旋糖酐,略低于150IU·mg-1的肝素。     

Visualisation of the gastrin receptor within rat mucosa using a biotinylated gastrin antagonist

We have previously demonstrated that the peptide Boc-l -Trp-l -Leu-β-Ala is a potent and specific antagonist of pentagastrin-stimulated acid secretion in both the rat and the dog. Using conventional solution phase methodology, the analogue biotinyl-l -Trp-l -Leu-β-Ala was prepared in reasonable yield and purity and applied to cryostat sections of rat intestinal and other tissues. The sections were exposed to 5–10 μg of peptide and the bound analogue was visualised using streptavidin-fluorescein. The binding of the analogue was demonstrated in sections from fundus, duodenum, ileum, colon, and lung. However, the analogue failed to bind to tissue from the pancreas, heart, kidney, or liver. The binding of the probe was greatly reduced or completely inhibited by preincubation with Boc-l -Trp-l -Leu-β-Ala, pentagastrin, or gastrin 1–17. The distribution of the cells recognised by the probe was consistent with the distribution of histamine-containing enterochromaffin-like cells. The results of this study may have some bearing on current theories of the mechanism of gastrin-stimulated acid release.     

单甘酯对实验性血栓形成的影响

目的　研究单甘酯 (glycolmannatesulfate ,GMS)对血栓形成的影响。方法　结扎大鼠下腔静脉观察对静脉血栓的形成 ,Chandler方法观察体外血栓形成 ,全自动凝血仪测定凝血指标和纤维蛋白原及ATⅢ、Ⅱ、Ⅱa活性。结果　GMS 2 0、40mg·kg-1对大鼠静脉血栓的形成有抑制作用 (P<0 0 1) ;2 5mg·kg-1即能明显抑制家兔体外血栓的形成 ,随剂量的增加而增强。GMS可使家兔TT、CT、APTT、RT及PT明显延长 ,降低大鼠血浆纤维蛋白原含量及降低因子Ⅱ及因子Ⅱa活性 ,升高ATⅢ活性。结论　GMS具有明显的抗血栓形成作用 ,其作用机制与其抗凝血作用有关     

Detection of nascent polyproline II helices in solution by NMR in synthetic insect kinin neuropeptide mimics containing the X-Pro-Pro-X motif

The conformations of three synthetic peptide analogs containing the dPro-dPro-dXaa motif (dXaa=dThr, dGlu, dAsn) in aqueous solution were studied by a combination of NMR and molecular modeling simulations. The three compounds were identified from a random d -amino acid tripeptide library on the basis of their ability to either mimic or block the diuretic activity of neuropeptides of the insect kinin family. TOCSY and ROESY correlations, as well as abnormal secondary chemical shifts for protons on the d -proline residues were employed to obtain conformational ensembles consistent with the experimental NMR data for the three analogs using an in vacuo simulated annealing protocol. Similar secondary structures were found for the three molecules after refinement, in agreement with the similarities observed between their NMR spectra. Unrestrained molecular dynamics simulations with explicit water representation indicate that the structural motifs found in vacuo are stable in aqueous solution. The three analogs can be considered initiators of right-handed poly d -proline II helices, mirror images of the poly l -proline II left-handed helical motifs normally found in proline-rich proteins. The role of these secondary folds on binding of the analogs to the kinin receptors is discussed.     

Effect of nifedipine on gastric acid secretion and gastrin release in healthy man

Summary Nifedipine, a calcium-channel antagonist widely used in cardiovascular disease, has recently been reported to be effective in the treatment of oesophageal motor disorders. The effect of a single therapeutic dose of nifedipine (20 mg p.o.) has been evaluated on basal and submaximal pentagastrin-stimulated gastric secretion and meal-stimulated gastrin release in healthy man. In comparison with placebo, nifedipine significantly decreased both basal and stimulated gastric acidity and juice volume, whereas only a slight but insignificant reduction in meal-stimulated gastrin levels was observed after drug administration. The results are in agreement with previous reports that calcium is involved in stimulus-secretion coupling in the human parietal cell. They do not confirm the effect of calcium on G-cells, although it is likely that doses of nifedipine higher than those commonly used might be effective in the reduction of gastrin secretion.     

硫酸卡那霉素体外抑制呼吸道合胞病毒作用的研究

目的评价硫酸卡那霉素体外抗呼吸道合胞病毒作用机制。方法采用细胞培养技术观察硫酸卡那霉素对细胞的毒性和抗呼吸道合胞病毒作用。结果用空斑减数实验测得半数有效剂量(EC50)为(1.71±0.23)mg/ml,MTT法测定其半数中毒浓度(CC50)为(12.36±0.85)mg/ml,SI为7.2,SI>4(有意义);在不同时间给药对病毒的抑制实验中,呼吸道合胞病毒感染喉癌上皮细胞(Hep-2)细胞后1、2、4、6、8和10h给药均有抑制作用(P<0.05),穿入和吸附抑制实验表明其对病毒穿入过程有明显地抑制作用(P<0.05),但是对吸附过程没有抑制作用。硫酸卡那霉素对RSV病毒没有直接的灭活作用。结论硫酸卡那霉素在体外实验中对呼吸道合胞病毒穿入细胞及在细胞内复制的过程均有抑制作用。     

硫酸镁联合硫酸沙丁胺醇对重度支气管哮喘急性发作患儿肺功能的影响

目的 探讨硫酸镁联合硫酸沙丁胺醇对重度支气管哮喘急性发作患儿肺功能的影响.方法 选择本院2015年1月至2016年6月期间收治的60例重度支气管哮喘急性发作患儿作为观察对象,随机分为观察组和对照组,每组30例.对照组采用硫酸沙丁胺醇治疗,观察组予以硫酸镁联合硫酸沙丁胺醇治疗,治疗1周后比较两组治疗效果、康复指标(哮鸣音、气短、咳嗽)、住院时间、肺功能指标[最大呼气流量(PEF)、1秒用力呼气容积(FEV1)、用力肺活量(FVC)]与不良反应.结果 观察组总有效率为96.67％,显著高于对照组的70.00％,差异有统计学意义(x 2=7.680,P＜0.05);治疗后与对照组相比,观察组哮鸣音消失时间[(1.04±0.35)d]、气短消失时间[(2.11±1.25)d]、咳嗽消失时间[(3.75±1.08)d]及住院时间[(5.28±1.53)d]均较短,差异均有统计学意义(均P＜0.05);两组治疗前的肺功能指标比较差异均无统计学意义(均P＞ 0.05);治疗后两组FEV1、PEF、FVC水平均显著升高,且观察组上升趋势更为显著,差异均有统计学意义(均P＜ 0.05);两组患儿不良反应发生率比较差异无统计学意义(x 2=0.267,P＞0.05).结论 与单纯采用硫酸沙丁胺醇治疗重度支气管哮喘急性发作相比,硫酸镁联合硫酸沙丁胺醇治疗效果更加确切,且不良反应较少,对改善患儿临床症状与肺功能均具有积极作用,值得临床进一步推广应用.     

胃泌素在胃肠道肿瘤中的作用及其信号转导途径的研究进展

胃泌素对于胃肠道肿瘤的促进作用已经被很多研究所证实。胃泌素与细胞表面受体结合后,通过一定的信号转导途径来调节某些基因转录与表达,从而促进肿瘤发生与发展。通过对胃泌素在胃肠道肿瘤中作用机制及其信号转导途径研究,有望找到更有效的胃肠道肿瘤治疗方法。     

硫酸依替米星与硫酸阿米卡星治疗下呼吸道细菌性感染临床评价

目的：进一步评价硫酸依替米星治疗下呼吸道细菌感染的安全有效性。方法：采用随机对照开放试验方法。硫酸依替米星组100mg溶于生理盐水100ml中静脉滴注，每日2次。硫酸阿米卡星400mg溶于生理盐水500ml中静脉滴注，每日1次，疗程均为5－10d。结果：硫酸依替米星组和硫酸阿米卡星组各有30例评价疗效。两组有效率分别为90．0％及86．7％，细菌清除除率分别为88．5％及88．0％，两组安全性评价各为30例，耳聋性不良反应分别为3．3％和3．3％，肾毒性不良反应分别为3．3％和3．3％。两组经统计学处理差异无显著性（P＞0．05）。结论：采用硫酸依替米星治疗下呼吸道细菌性感染安全有效。     

Netazepide,a gastrin/CCK2 receptor antagonist,causes dose-dependent,persistent inhibition of the responses to pentagastrin in healthy subjects

Aims

To confirm by means of pentagastrin, a synthetic gastrin agonist, that netazepide is a gastrin/CCK₂ receptor antagonist in healthy subjects, and that antagonism persists during repeated dosing.

Methods

We did two studies in which we infused pentagastrin (0.6 μg kg⁻¹ h⁻¹ intravenously), aspirated gastric secretion and measured the volume, pH and H⁺ secretion rate of the gastric aspirate. First, we did a double-blind, five-way crossover study (n = 10) to assess the effect of single oral doses of netazepide (1, 5, 25 and 100 mg) and placebo on the response to pentagastrin. Then, we did a single-blind, placebo-controlled study (n = 8) to assess the effect of the first and last oral doses of netazepide (100 mg) twice daily for 13 doses on the response to pentagastrin.

Results

Netazepide was well tolerated. After placebo, pentagastrin increased the volume and H⁺ secretion rate and reduced the pH of gastric aspirate. Compared with placebo, single doses of netazepide caused dose-dependent inhibition of the pentagastrin response (P < 0.02); netazepide (100 mg) abolished the response. After 13 doses, the reduction in volume and H⁺ secretion rate persisted (P < 0.001), but the pH effect was mostly lost.

Conclusions

Netazepide is an orally active, potent, competitive antagonist of human gastrin/CCK₂ receptors. Antagonism is dose dependent and persists during repeated dosing, despite tolerance to the effect on pH. Further studies are required to explain that tolerance. Netazepide is a tool to study the physiology and pharmacology of gastrin, and merits studies in patients to assess its potential to treat gastric acid-related conditions and the trophic effects of hypergastrinaemia.     

Effect of two antiserotoninergic drugs,methysergide and metergoline,on gastric acid secretion and gastrin release in healthy man

Summary The effects of acute oral administration of the antiserotoninergic drugs methysergide (3 mg) and metergoline (4 mg) on basal, submaximal (0.6 µg/kg i. m.) and maximal (6 µg/kg) pentagastrinstimulated gastric acid secretion, as well as on basal and food-induced gastrin release, have been evaluated in healthy volunteers. Methysergide significantly increased basal and submaximal pentagastrin-stimulated gastric acid secretion, and metergoline significantly inhibited gastric acidity in all experiments. Basal and stimulated serum gastrin concentrations were not modified by either drug. The effect of methysergide on gastric acid secretion was opposed to that of serotonin and was probably dependent on its antiserotoninergic action, but the decrease in gastric acidity caused by metergoline is not easily explained. Although the effect is similar to that of a dopamine infusion, it does not depend on dopamine receptor stimulation, since it was not influenced by pretreatment with metoclopramide. It is suggested that it might be due to the weak anticholinergic and/or antihistaminic properties of metergoline.     

The effect of selective and non-selective cholinergic blockade on bombesin- and peptone-stimulated gastrin release

Summary We have studied the effects of the selective muscarinic M₁-receptor antagonist pirenzepine and the non-selective muscarinic antagonist atropine on bombesin- and peptone-stimulated gastrin release in healthy subjects.Pirenzepine (i. v. bolus 0.6 mg/kg) and atropine (i. v. bolus 15 µg/kg, followed by an infusion of 5 µg·kg^–1·h^–1) were given in doses equipotent in terms of reduction of gastric acid secretion. Neither affected bombesin- or peptone-stimulated gastrin release.These findings do not support the involvement of M₁-receptors in the cholinergic regulation of gastrin release and suggest that the reduction in acid secretion caused by pirenzepine is not mediated by inhibition of gastrin release.     

Effect of omeprazole treatment on plasma concentrations of the gastric peptides,xenin, gastrin and somatostatin,and of pepsinogen

Abstract: The peptide xenin 25 is a gastric mucosal constituent like gastrin, somatostatin and pepsinogen. Gastrin and pepsinogen plasma concentrations increase when the secretion of gastric acid is reduced by proton pump inhibitors. In the present investigation, treatment with omeprazole led to an increase in fasting and postprandial plasma concentrations of xenin, gastrin and pepsinogens A and C (P < 0.05, in each instance), whereas somatostatin plasma levels remained unchanged. Because subcutaneous injection of pentagastrin did not raise xenin plasma concentrations, a direct effect of gastrin on xenin production seems unlikely. This study indicates that xenin plasma concentrations are regulated by intragastric pH, as are those of gastrin and pepsinogen.