RNAi沉默PLCε对Crocin抑制人膀胱癌BIU-87细胞增殖作用调控的研究

目的:观察RNAi沉默膀胱癌BIU-87细胞磷脂酶C-ε(Phospholipase C epsilon,PLCε)表达后对藏花素(Crocin)抑制人膀胱癌BIU-87细胞增殖作用的影响。方法:将携有siRNA基因的真核表达质粒PGenesil-PLCε转染BIU-87细胞后,分为5组:未转染组、pGenesil-NP阴性质粒组、pGenesil-PLCε质粒组、Crocin组、pGenesil-PLCε联合Crocin组。MTT法观察不同处理组对BIU-87细胞的生长抑制作用。RT-PCR法检测转染后对PLCεmRNA表达的影响及各处理组PCNA(Proliferating cell nuclear antigen)、CyclinD1 mRNA表达。Western-blot检测PCNA、CyclinD1蛋白表达。结果:RNAi可抑制BIU-87细胞78.06%的PLCεmRNA表达。pGenesil-PLCε联合Crocin组可增强Crocin对BIU-87细胞的生长抑制作用,48h抑制率达45.30%,且与pGenesil-PLCε质粒组和Crocin组相比较有显著性差异(P<0.01)。RT-PCR和Western-blot结果显示与RNAi沉默PLCε组比较,RNAi PLCε联合Crocin组明显下调了PCNA、Cyclin D1基因和蛋白的表达(P<0.05);较单独Crocin组比较,RNAi PLCε联合Crocin组显著降低了PCNA基因和蛋白的表达(P<0.05),降低了Cyclin D1蛋白的表达(P<0.05),Cyclin D1 mRNA表达变化两组间无显著性差异(P>0.05)。结论:沉默PLCε基因可增强Crocin对BIU-87细胞的抑制作用,其机制与进一步下调PCNA、CyclinD1的表达有关。

Beneficial effects of saffron(Crocus sativus L.) in ocular pathologies, particularly neurodegenerative retinal diseases

Saffron(Crocus sativus L.)has been traditionally used in food preparation and as a medicinal plant.It currently has numerous therapeutic properties attributed to it,such as protection against ischemia,as well as anticonvulsant,antidepressant,anxiolytic,hypolipidemic,anti-atherogenic,anti-hypertensive,antidiabetic,and anti-cancer properties.In addition,saffron has remarkable beneficial properties,such as anti-apoptotic,anti-inflammatory and antioxidant activities,due to its main metabolites,among which crocin and crocetin stand out.Furthermore,increasing evidence underwrites the possible neuroprotective role of the main bioactive saffron constituents in neurodegenerative diseases,such as Parkinson’s and Alzheimer’s diseases,both in experimental models and in clinical studies in patients.Currently,saffron supplementation is being tested for ocular neurodegenerative pathologies,such as diabetic retinopathy,retinitis pigmentosa,age-related macular degeneration and glaucoma,among others,and shows beneficial effects.The present article provides a comprehensive and up to date report of the investigations on the beneficial effects of saffron extracts on the main neurodegenerative ocular pathologies and other ocular diseases.This review showed that saffron extracts could be considered promising therapeutic agents to help in the treatment of ocular neurodegenerative diseases.

Effect of crocin carotenoid on BDNF and CREB gene expression in brain ventral tegmental area of morphine treated rats

Objective: To investigate the effect of crocin carotenoid on BNDF and CREB gene expression in the brain ventral tegmental area(VTA) and the serum level of BDNF in morphine-treated rats compared to control. Methods: In this study, 40 male Wistar rats(200-250 g) were used in 5 experimental groups: 1) non morphine treat rats(control); 2) non morphine-treated rats with 25 mg/kg crocin carotenoid(i.p., for 21 d); 3) morphine treated rats(10 mg/kg twice a day, s.c., 21 d); 4 and 5) morphine-treated rats with 12.5 and 25 mg/kg crocin carotenoid, respectively. By the end of research, BDNF and CREB expression was determined by real-time-PCR method. ELISA analysis was also applied for assessing the serum BDNF level. Results: The data indicated that morphine treatment could cause a significant decrease in BDNF and CREB gene expression(P<0.01 and P<0.001, respectively) in brain VTA as well as serum level of BDNF(P<0.01) in comparison to control group. Treatment with 25 mg/kg crocin carotenoid caused a significant enhancement in BDNF and CREF gene expression(P<0.01 and P<0.05, respectively) and serum level of BDNF(P<0.01) in morphine-treated rats in comparison to morphine-treated group. Conclusions: Regarding to obtained results, crocin carotenoid can inhibit unfavorable effects of morphine on the neural system to some extent through enhancing BDNF and CREB gene expression in brain VTA and serum level of BDNF.

Effects of Crocin on Nox2 Expression and ROS Level of Hypoxia/Reoxygenation-induced Injury of Cardiomyocytes

[Objectives]To explore the protection mechanism of crocin against ischemia-reperfusion injury of myocardial cells.[Methods]Newborn male SD rats were selected,left ventricular cardiomyocytes(CMs)were isolated,and a hypoxia/reoxygenation model of CMs was established to simulate the process of ischemia/reperfusion injury.The cells were randomly divided into four groups:normal cell group(control group),crocin group),hypoxia/reoxygenation group(H/R group),hypoxia/reoxygenation+crocin group(H/R+crocin group).H/R+crocin group selected the concentration of crocin 1,10,and 100μmol/L,and determined the optimal concentration of crocin by detecting the cell proliferation ability.After the cells were pretreated using the optimal concentration of crocin,the levels of superoxide anion,cell proliferation,apoptosis and Nox2 levels in each group of cells were detected.[Results]Compared with the control group,the proliferation ability of CMs after hypoxia-reoxygenation injury was reduced(P<0.05),while cell apoptosis and intracellular superoxide anion levels were significantly increased(P<0.01);the CMs pretreated with crocin can reduce the level of Nox2(P<0.01),increase the cell proliferation ability of CMs,reduce cell apoptosis,and accordingly reduce the level of superoxide anion in the cell(P<0.05).[Conclusions]Crocin protects CMs from hypoxia/reoxygenation injury through down-regulating the level of Nox2 and reducing oxidative stress injury.

Hepato-and reno-protective effects of thymoquinone,crocin,and carvacrol:A comprehensive review

Medicinal plants are rich in nutrients and phytochemicals which prevent and treat a wide range of ailments.Accumulating experimental studies exhibit that some bioactive ingredients extracted from medicinal plants have suitable therapeutic effects on hepatic and renal injuries.This review focuses on the hepato-and reno-protective effects of thymoquinone,crocin,and carvacrol.The relevant literature was retrieved from PubMed,Scopus,Web of Science,and Google Scholar databases from the beginning of 2015 until the end of November 2021.According to the scientific evidence,the considered phytochemicals in this review have been applied with useful therapeutic effects on hepatic and renal damage.These therapeutic effects were mainly mediated through the amelioration of oxidative stress,suppression of inflammatory responses,and inhibition of apoptosis.Intracellular signaling pathways linked to nuclear factor kappa B(NF-κB),adenosine monophosphate-activated protein kinase,c-jun N-terminal kinase,and extracellular signal-regulated kinase 1/2 and Toll-like receptors are the most important pathways targeted by these phytochemicals.Up-regulation of transcription factor Nrf2 and down-regulation of transforming growth factor-beta 1 by these natural compounds also contribute to the alleviation of hepatic and renal injuries.

Dietary crocin reverses melanoma metastasis

Crocus sativus and its bioactive constituent crocin are well known for anti-tumor potential in different models.However, the efficacy of crocin on in-vivo melanoma metastasis is not yet reported. In this study, melanoma metastatic model was developed by tail vein injection of B16 F-10 cells in to C57 BL/6 mice. Metastatic mice treated with two different doses of crocin（250 and 500 μg/kg of bodyweight） for 10 days and parameters such as lung metastasis inhibition, mean survival time, lung hydroxyproline, uronic acid and hexosamine levels were analyzed after 21 days of treatment. Then blood was collected and serum gamma glutamyl transpeptidase（γ-GGT）, sialic acid,tumor necrosis factor alpha（TNF-a）, interleukin 10（IL-10）, IL-6, IL-2, and TIMP-1 levels were measured. Further, a lung histological examination was done in crocin treated metastatic mice. Subsequently hallmark metastatic parameters such as matrix metalloproteinases（MMPs）, extracellular regulated kinase 2（ERK2）, vascular endothelial growth factor（VEGF）, and K-ras gene expression were investigated in the lungs of crocin treated metastatic mice.Further, in-vitro adhesion, invasion and migration of B16 F-10 cells were examined after 24 hours of crocin（5 and 10μg/mL） treatment. Administration of crocin to tumor bearing C57 BL/6 mice reduced the lung metastasis by 85%.Elevated levels of hydroxyproline, uronic acid, hexosamine, serum sialic acid and y-GGT in metastatic control were found to be significantly reduced in crocin treated mice. Crocin also inhibited expression of MMP-2, MMP-9, ERK-2,K-ras, and VEGF. Crocin reduced the ability of B16 F-10 cells invasion（P〈0.05）, migration（P〈0.05） and adhesion by upregulating E-cadherin expression. In conclusion, crocin elicited marked anti-metastatic potential by regulating the metastasis induced biomarkers.

Experimental Investigation of Inhibition Efficiency of Crocin for Chloride-Induced Corrosion of Aluminum Alloys

The use of Crocin, derived from the flowers of Crocus sativus, is investigated as corrosion inhibitor for the AA1050, AA5083, AA5754 and AA6082 aluminum alloys in chloride ions environment. Aluminum and aluminum alloys are subjected to corrosion in the aggressive environment of chlorides, so several green corrosion inhibitors, mostly of plant origin, with minimum impact on health and the environment have been examined. In this study, the inhibition efficiency of 1.25 mM Crocin in a 0.01 M NaCl corrosive solution was assessed via electrochemical corrosion techniques and gravimetric mass loss measurements of the aluminum alloys. The surface of the specimens was examined using Scanning Electron Microscopy, Energy Dispersive Spectroscopy, Stereomicroscopy and Glossiness measurements. Experimental results reveal the protective anticorrosive action of Crocin for all aluminum alloys in the sodium chloride medium.

Crocus genome reveals the evolutionary origin of crocin biosynthesis

Crocus sativus (saffron) is a globally autumn-flowering plant, and its stigmas are the most expensive spice and valuable herb medicine. Crocus specialized metabolites, crocins, are biosynthesized in distant species, Gardenia (eudicot) and Crocus (monocot), and the evolution of crocin biosynthesis remains poorly understood. With the chromosome-level Crocus genome assembly, we revealed that two rounds of lineage-specific whole genome triplication occurred, contributing important roles in the production of carotenoids and apocarotenoids. According to the kingdom-wide identification, phylogenetic analysis, and functional assays of carotenoid cleavage dioxygenases (CCDs), we deduced that the duplication, site positive selection, and neofunctionalization of Crocus-specific CCD2 from CCD1 members are responsible for the crocin biosynthesis. In addition, site mutation of CsCCD2 revealed the key amino acids, including I143, L146, R161, E181, T259, and S292 related to the catalytic activity of zeaxanthin cleavage. Our study provides important insights into the origin and evolution of plant specialized metabolites, which are derived by duplication events of biosynthetic genes.

Formulation Containing Phytosomes of Carotenoids from Nyctanthes arbor-tristis and Tagetes patula Protect D-galactose Induced Skin Aging in Mice

Background:Carotenoids play important role in delay of aging process.Orange coloured tubular calyx of flowers of Nyctanthes arbor-tristis contains crocin,an apocarotenoid which forms a major component of stigma of saffron.Due to presence of crocin in orange coloured tubular calyx of Nyctanthes arbor-tristis,it can be used as an economical substitute to saffron for its medicinal and cosmetic utilities.Lutein from flower petals of Tagetes patula L.,is another popular carotenoid which has antioxidant effect and many health benefits.The carotenoids are highly unstable when exposed to atmosphere.One of the ways to improve stability of these phytoconstituents,is their entrapment in phytosomes.Preparation of phytosomes will have dual advantages of improving stability as well as bioavailability of molecule.Objective:In the present study crocin and lutein rich extracts were entrapped into phytosomes to improve stability and efficacy of topical preparation.Methods:The phytosomes of Carotenoid rich extract of tubular calyx of Nyctanthes arbor-tristis L.and the petals of Tagetes patula L.(standardized for crocin and lutein content)were prepared using lipid film hydration technique and these phytosomes were then incorporated into gel base.The gel formulation was evaluated for stability as per ICH guidelines.Efficacy of formulation was evaluated by d-galactose induced aging model.Aging in skin was induced by administration of d-galactose(100 mg/kg bwsc.)to albino mice for 42 days.The gel formulation was applied topically for 42 days.Then the effect of formulation on skin aging was evaluated by estimation of biochemical parameters viz.glutathione and malondialdehyde(MDA)and histopathological studies of treated skin samples.Expression of COL type I and elastin genes in skin samples was also carried out by RT-PCR.Results:Percent entrapment(%w/w)of crocin and lutein in phytosomes were found to be 60.20%and 50.81%,respectively.Accelerated stability studies showed improvement in stability of carotenoids viz.crocin and lutein and the content of crocin and lutein in formulation was found in the range of 99.98%w/w to 99.85%w/w at the end of three months.The formulation containing extract of Phytosomes of carotenoid rich extracts of Nyctanthes arbor-tristis L.and the petals of Tagetes patula L.exhibited potent antiaging activity through significant(P<0.05)increase in dermal and epidermal layers,and increase in GSH levels of skin as compared to the untreated group.The treatment with the gel formulation revealed upregulation of collagen type I and elastin genes.There was significant reduction in lipid peroxidation as revealed through reduction in MDA levels as compared to untreated group.Conclusion:Crocin and lutein have potential to prevent skin aging via upregulation of collagen type I gene and elastin gene.Upregulation of genes resulted into increase in the thickness of epidermal and dermal layer along with reduction in oxidative stress in skin.Entrapment of carotenoid rich extracts of Nyctanthes arbor-tristis and Tagetes patula in phytosomes enhanced the stability and efficacy of the formulation.

Therapeutic Effect of Crocin on Diabetic Retinopathy in Rats Based on TLR4/My D88/NF-κB Pathway

Objective:To study the therapeutic effect of crocin on diabetic retinopathy(DR)in rats based on toll-like receptor 4(TLR4)/myeloid differentiation factor 88(My D88)/nuclear factor-κB(NF-κB)pathway.Methods:Thirty SPF SD rats were used in the experiment,which were randomly divided into DR group,control group and crocin group,with 10 rats in each group.The DR rat model was established by feeding the rats in both the DR group and crocin group with a high glucose and high fat diet,along with intraperitoneal injection(IP)of streptozotocin.Crocin IP was administered to the rats in the crocin group,whereas the rats in the DR group and control group received an equivalent dosage of saline IP for 12 weeks.A comparison was made among the three groups regarding retinal thickness,vascular permeability,expression of TLR4/My D88/NF-κB pathway protein,levels of inflammatory factors,and levels of Bcl-2,Bax,and Bcl-2/Bax.Results:The DR group and crocins group exhibited a lower retinal thickness compared to the control group,while the crocins group displayed a higher thickness than the DR group.The DR group and crocins group had higher retinal vascular permeability than the control group,and the crocins group had lower retinal vascular permeability than the DR group(P<0.05).TLR4,My D88,and P-NF-κB relative expressions were higher in the DR and crocin groups than in the control group,whereas TLR4,My D88,and P-NF-κB relative expressions were lower in the crocin group than in the DR group(P<0.05).The DR group and crocin group exhibited elevated levels of inflammatory cytokines compared to the control group,while the crocin group displayed decreased levels in comparison to the DR group(P<0.05).The DR group and crocin group exhibited lower levels of Bcl-2 and Bcl-2/Bax compared to the control group,whereas the control group displayed higher levels of Bax.The crocin group exhibited elevated levels of Bcl-2 and Bcl-2/Bax compared to the DR group,whereas the DR group displayed diminished levels of Bax(P<0.05).Conclusion:Crocin has the potential to enhance the retinal thickness and vascular permeability of DR rats,and the inhibition of the TLR4/My D88/NF-κB pathway by crocin could play a crucial role in impeding the advancement of DR.

Research and application progress of Gardenia jasminoides

Fruits of Gardenia jasminoides（Zhizi in Chinese） have been used as dietary supplement and traditional Chinese medicine for thousands of years. Recent studies on Gardenia jasminoides reveal that the extracts or the active components play an essential role in oxidative stress, inflammation, hepatoprotection,neuroprotection, anti-diabetes, antitumor and so on. Here, we reviewed G. jasminoides in the aspects of botanical identity, chemical ingredients, pharmacology and pharmacokinetics, safety and toxicity evaluation, clinical application, etc.

Effects of Gardenia jasminoides extracts on cognition and innate immune response in an adult Drosophila model of Alzheimer's disease

Herbal extracts have been extensively used worldwide for their application on memory improvement, especially among aged and memory-deficit populations. In the present study, the memory loss induced by human Abeta protein over-expression in fruitfly Alzheimer's disease(AD) model was rescued by multiple extracts from Gardenia jasminoides. Three extracts that rich with gardenia yellow, geniposide, and gardenoside components showed distinct rescue effect on memory loss. Further investigation on adding gardenoside into a formula of Ganoderma lucidum, Panax notoginseng and Panax ginseng(GPP) also support its therapeutic effects on memory improvement. Interestingly, the application of GPP and gardenoside did not alter the accumulation of Abeta proteins but suppressed the expression of immune-related genes in the brain. These results revealed the importance and relevancy of anti-inflammation process and the underlying mechanisms on rescuing memory deficits, suggesting the potential therapeutic use of the improved GPP formulation in improving cognition in defined population in the future.

Effects of Culture Conditions, Carbon Source and Regulators on Saffron Callus Growth and Crocin Accumulation in the Callus

There are many factors influencing the growth and secondary metabolites of callus and saffron callus. In this paper, the effects of culture conditions, including culture temperatures, light levels, the carbon source and its concentration, and the preserve of regulators (mainly hormones), are studied for callus cultures. All the experiments used Murashige and Skoog (MS) solid medium as the basic medium with 10 g/L agar, pH 5.75.8. Saffron callus was cultured at 20℃ in the dark, with a sucrose concentration of 45 g/L (or starchy hydrolysate concentration of 40 g/L), but 30 g/L sucrose was best for the synthesis of crocin (for starchy hydrolysate the concentration can range from 20 to 40 g/L). To promote callus growth, the best auxin was α-naphthaleneacetic acid (NAA) and the optimum ratio of NAA (mg/L) to benzylaminopurine (BA) (mg/L) was 2∶0.25. Indole-3-acetic acid (IAA) (4 mg/L), gibberellin(GA3) (2 mg/L), and uniconazole (S-07) (1.25 mg/L) increased the crocin content remarkably as analyzed by high performance liquid chromatography (HPLC). NAA (2 mg/L) promoted the growth of saffron callus but had no benefit and may inhibit crocin synthesis while S-07 (1.25 mg/L) had the opposite effect. GA3 promoted both growth and synthesis.