Ghrelin receptor is activated by naringin and naringenin,constituents of a prokinetic agent Poncirus fructus

Ethnopharmacological relevance

Poncirus fructus (PF), also known as a dried immature fruit of Poncirus trifoliata (L.) Raf. (Rutaceae), has long been traditionally used for the various gastrointestinal disorders in Eastern Asia.

Aim of study

The aqueous extract of PF (PF-W) has the strong prokinetic effect, yet the underlying mechanism is still elusive. The present study investigated whether PF-W has any effect on motilin receptor or ghrelin receptor, since these receptors enhance intestinal motility when activated.

Materials and methods

The effect of PF-W and its components on motilin or ghrelin receptor was determined by calcium imaging and whole-cell patch clamp methods.

Results

PF-W activates the ghrelin receptor, but not the motilin receptor, resulting in a transient increase of intracellular calcium levels. Furthermore, among various constituents of PF, only naringin and naringenin evoked the intracellular calcium augmentation via the ghrelin receptor. Moreover, cortistatin-8 – a ghrelin receptor inhibitor – specifically blocked naringin- and naringenin-induced calcium increases. In addition, naringin and naringenin induced inward currents in ghrelin receptor-expressing cells under whole-cell patch clamp configuration.

Conclusion

PF-W activates the ghrelin receptor, and naringin and naringenin are key constituents responsible for the activation of ghrelin receptor. Therefore, the present study suggests that the ghrelin receptor is a molecular entity responsible for the strong prokinetic activity of PF-W.     

Poncirus trifoliate fruit modulates pacemaker activity in interstitial cells of Cajal from the murine small intestine

Ethnopharmacological relevance

Poncirus fructus (PF) has been widely used as a traditional medicine in Eastern Asia, especially to ameliorate the symptoms of gastrointestinal (GI) disorders related to abnormal GI motility.

Aim of the study

Poncirus fructus (PF), also known as Poncirus trifoliata (L.) Raf. (Rutaceae), is widely used as a traditional medicine in Eastern Asia mainly to ameliorate the symptoms of gastrointestinal (GI) disorders related to abnormal GI motility. In a previous study, a methanol extract of PF was found to have particularly potent gastroprokinetic effects. Interstitial cells of Cajal (ICCs) are pacemaker cells in the gastrointestinal tract, but the action mechanisms of PF extract in mouse small intestinal ICCs have not been investigated. Therefore, in the present study, we investigated the effects of a methanol extract of PF (MPF) in mouse small intestinal ICCs. In addition, we sought to identify the receptors involved.

Materials and Methods

Enzymatic digestions were used to dissociate ICCs from small intestines. The whole-cell patch-clamp configuration was used to record potentials (current clamp) from cultured ICCs. In addition, we analyzed intracellular Ca²⁺ concentrations ([Ca²⁺]_i).

Results

MPF decreased the amplitudes of pacemaker potentials in ICCs, and depolarized resting membrane potentials in a concentration dependent manner. Y25130 (a 5-HT₃ receptor antagonist) and RS39604 (a 5-HT₄ receptor antagonist) blocked MPF-induced membrane depolarizations, whereas SB269970 (a 5-HT₇ receptor antagonist) did not. Pretreatment with Na⁺ or Ca²⁺-free solution or thapsigargin (a Ca²⁺-ATPase inhibitor in endoplasmic reticulum) abolished the generation of pacemaker potentials and suppressed MPF-induced activity. [Ca²⁺]_i analysis showed that MPF increased [Ca²⁺]_i. Furthermore, treatments with PD 98059, SB203580, or JNK II inhibitor blocked MPF-induced membrane depolarizations in ICCs.

Conclusion

These results suggest that MPF modulates pacemaker potentials through 5-HT₃ and 5-HT₄ receptor-mediated pathways via external Na⁺ and Ca²⁺ influx, and via Ca²⁺ release from internal stores in a mitogen-activated protein kinase dependent manner. The study shows MPF is a good candidate for the development of a gastroprokinetic agent. In view of the effects of MPF on ICCs, further research is required, particularly to identify the active compound(s) involved and to determine their action mechanisms.     

An aqueous extract of Poncirus fructus activates the prokinetic activity of 5-HT receptor subtype 4 without hERG interaction

Aim of the study

Poncirus fructus (PF) - also known as the dried, immature fruit of Poncirus trifoliata (L.) Raf. (Rutaceae) - is a natural substance that has long been used for various gastrointestinal disorders in eastern Asia. An aqueous extract of PF (PF-W) has particularly potent gastroprokinetic effects, but its molecular mechanism was not well understood. Identification of the underlying prokinetic mechanism of PF-W was pursued in the present study.

Materials and methods

Changes in in vitro cAMP levels and in vivo intestinal transit rate (ITR) caused by PF-W were measured after pretreatment with GR125487, an antagonist for serotonin receptor subtype 4 (5-HT4R). An [³H] astemizole binding assay and electrophysiology experiments were performed to determine if PF-W has any interaction with the human ether-à-go-go related gene (hERG) potassium channel.

Results

PF-W induced an increase in intracellular cAMP in 5-HT4R-expressing HEK293T cells, indicating that PF-W does activate 5-HT4R. Moreover, pretreatment with GR125487 successfully blocked the increase, suggesting that the response was 5-HT4R-specific. More importantly, pretreatment of GR125487 in rats inhibited the elevation of ITR by PF-W, indicating that the prokinetic effect of PF-W was indeed exerted via 5-HT4R. On the other hand, both [³H]-astemizole binding assay and electrophysiological experiments revealed that PF-W did not interfere at all with the hERG channel.

Conclusion

It was found that PF-W exerts its prokinetic activity through a 5-HT4R-mediated pathway, with no interaction with hERG channels. Therefore, PF-W is a good candidate that might be developed as a prokinetic agent with fewer expected cardiac side effects.     

Protective effects of neohesperidin and poncirin isolated from the fruits of Poncirus trifoliata on potential gastric disease

The effects of Poncirus trifoliata (P. trifoliata) (Ponciri Fructus, PF) extract and its constituents such as neohesperidin and poncirin on gastritis in rats and human gastric cancer cells were investigated. The PF 70% ethanol extracts (1 g) showed approximately 11.38% of acid‐neutralizing capacities and cytotoxicity (IC₅₀ = 85.39 µg/mL) against human AGS gastric cancer cells. In addition, neohesperidin exhibited antioxidant activity (IC₅₀ = 22.31 µg/mL) in the 1,1‐diphenyl‐2‐picryldydrazyl (DPPH) radical‐scavenging assay. Neohesperidin (50 mg/kg) and poncirin (100 mg/kg) significantly inhibited 55.0% and 60.0% of HCl/ethanol‐induced gastric lesions, respectively, and increased the mucus content. In pylorus ligated rats, neohesperidin (50 mg/kg) significantly decreased the volume of gastric secretion and gastric acid output, and increased the pH. From these results, it could be suggested that neohesperidin and poncirin isolated from PF may be useful for the treatment and/or protection of gastritis. Copyright © 2009 John Wiley & Sons, Ltd.     

绿衣枳实的化学成分

目的：研究绿衣枳实（Poncirus trifoliata（L.）Raf.）的化学成分。方法：对绿衣枳实95%乙醇提取物的氯仿部分进行色谱分离,根据光谱数据和理化性质确定各化合物的结构。结果：分离得到8个已知化合物,分别为：21α-methylmelianodiol（1）,21β-methylmelianodiol（2）,21α,25-dimethylmelianodiol（3）,21α-methyltoosen-danpentol（4）,21R,23R-epoxy-21α-methoxy-7,24,25-trihydroxy-14-apotirucallen-3-one（5）,21S,23R-epoxy-21β-methoxy-7,24,25-trihydroxy-14-apotirucallen-3-one（6）,21R,23R-epoxy-21α-ethoxy-24S,25-dihy-droxy-apotirucalla-7-en-3-one（7）,ichanexic acid（8）.。结论：化合物4-8为首次从枳属中分得。     

Hexane extract of Poncirus trifoliata (L.) Raf. stimulates the motility of rat distal colon

Aim of the study

Poncirus trifoliata (L.) Raf. (Rutaceae, PT) has been commonly used for treating gastrointestinal (GI) disorders in Korean traditional medicine, but its pharmacological roles in the regulation of colonic motility have not been clarified. This study investigated the regulatory effects of PT on the colonic motility.

Materials and methods

Immature fruits of PT were sequentially partitioned with MeOH, n-hexane, CHCl₃, EtOAc, n-BuOH and H₂O, and the effects of PT extracts on the contractility of colonic strips and colonic luminal transit in rats were measured in vitro and in vivo, respectively.

Results

Among six different extracts, only hexane extract of PT (PTHE) dose-dependently increased the low frequency contraction of longitudinal muscle in distal colonic strips, and the ED₅₀ value was revealed to be 0.71 μg/ml. The contractile patterns induced by PTHE were remarkably different from those caused by acetylcholine (ACh) and serotonin (5-HT). The stimulatory effects of PTHE on the whole distal colonic strips were more prominent than on the mucosa/submucosa-denuded segments. The M₂ receptor-preferring, methoctramine (0.5 μM), and M₃ receptor-preferring antagonist, 4-DAMP (0.5 μM) significantly blocked the PTHE (1 μg/ml)-induced contraction of distal colon longitudinal muscles, whereas the 5-HT receptor antagonists (1.0 μM, alone or in combination) selective for 5-HT₃ (ondansetron), 5-HT₄ (GR113808) and 5-HT_{1, 2, 5–7} (methysergide) receptors did not change the PTHE (1 μg/ml)-induced contractility of distal colon longitudinal muscles. SNAP (0.1 mM), a NO donor, enhanced the stimulatory effects of PTHE on the longitudinal muscle of distal colon, but l-NAME (0.1 mM), a NO synthesis inhibitor, had no effects. PTHE (10–100 mg/kg) caused a dose-dependent increase of colonic luminal transit.

Conclusions

Collectively, these findings suggest that PTHE specifically acts on the longitudinal muscle of distal colon in rats, and these stimulatory effects are likely mediated, at least, by activation of acetylcholinergic M₂ and M₃ receptors.     

仲景方中枳实用药剂量古今折算研究

采用文献考证和实物测量的方法对仲景方中枳实进行了系统研究。从诸多本草文献的记载和相关附图的研究发现,仲景方所用枳实并非今之所用枳实,而是种属为芸香科植物枸橘的枳壳。随后对选用产自河南驻马店的枸橘枳壳作为仲景方枳实的实测药材进行了实物测量,得出枸橘枳壳一枚为12g。结合现代道地枳实、枳壳实测结果的对比,并运用所得实测结果对小承气汤全方剂量配伍比例的分析,认为仲景方中枳实为枸橘枳壳,其一枚的剂量为12g是合理的。该研究为今后建立仲景方的药物剂量和配伍比例标准提供研究基础。     

Review of the Pharmacological Effects of Vitis vinifera (Grape) and its Bioactive Constituents: An Update

Vitis vinifera fruit (grape) contains various phenolic compounds, flavonoids and stilbenes. In recent years, active constituents found in the fruits, seeds, stems, skin and pomaces of grapes have been identified and some have been studied. In this review, we summarize the active constituents of different parts of V. vinifera and their pharmacological effects including skin protection, antioxidant, antibacterial, anticancer, antiinflammatory and antidiabetic activities, as well as hepatoprotective, cardioprotective and neuroprotective effects in experimental studies published after our 2009 review. Clinical and toxicity studies have also been examined. Copyright © 2016 John Wiley & Sons, Ltd.     

枳壳组织培养和植株再生研究

以枳壳PoncirustrifoliataRaf．实生苗上胚轴为材料进行离体培养，BA1mg／L时，出芽率最高（86．0％），BA浓度升高，出芽率随之下降；苗龄为20d的实生苗，其外植体易于出芽；实生苗培养中照光与否对出芽无太大影响；外植体取材部位，以实生苗的中部及下部较好，出芽率可达95％以上。     

Pharmacological properties of Visnea mocanera L. f. extracts: Screening for analgesic,antipyretic, antiinflammatory and ulcerogenic activities in rodents

Different extracts of Visnea mocanera L. f. (Theaceae) were evaluated for analgesic, antipyretic, antiinflammatory and ulcerogenic properties in mice and rats. The acetone extract from the leaves and the syrup from the fruits exerted central analgesic properties, associated with significant but transient antiinflammatory effects on acute inflammatory processes. The acetone extract also possessed topical antiinflammatory effects. The extracts assayed did not display antipyretic and ulcerogenic activity. Sub-acute toxicological experiments indicated a very low toxicity in mice.