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1.
Article in Chinese | WPRIM | ID: wpr-928157

ABSTRACT

This study aims to explore the effect of icariin(ICA) on mitochondrial dynamics in a rat model of chronic renal failure(CRF) and to investigate the molecular mechanism of ICA against renal interstitial fibrosis(RIF). CRF was induced in male Sprague-Dawley(SD) rats with 5/6(ablation and infarction, A/I) surgery(right kidney ablation and 2/3 infarction of the left kidney). Four weeks after surgery, the model rats were randomized into the following groups: 5/6(A/I) group, 5/6(A/I)+low-dose ICA group, and 5/6(A/I)+high-dose ICA group. Another 12 rats that received sham operation were randomly classified into 2 groups: sham group and sham+ICAH group. Eight weeks after treatment, the expression of collagen-Ⅰ(Col-Ⅰ), collagen-Ⅲ(Col-Ⅲ), mitochondrial dynamics-related proteins(p-Drp1 S616, p-Drp1 S637, Mfn1, Mfn2), and mitochondrial function-related proteins(TFAM, ATP6) in the remnant kidney tissues was detected by Western blot. The expression of α-smooth muscle actin(α-SMA) was examined by immunohistochemical(IHC) staining. The NRK-52 E cells, a rat proximal renal tubular epithelial cell line, were cultured in vitro and treated with ICA of different concentration. Cell viability was detected by CCK-8 assay. In NRK-52 E cells stimulated with 20 ng·mL~(-1) TGF-β1 for 24 h, the effect of ICA on fibronectin(Fn), connective tissue growth factor(CTGF), p-Drp1 S616, p-Drp1 S637, Mfn1, Mfn2, TFAM, and ATP6 was detected by Western blot, and the ATP content and the mitochondrial morphology were determined. The 20 ng·mL~(-1) TGF-β1-stimulated NRK-52 E cells were treated with or without 5 μmol·L~(-1) ICA+10 μmol·L~(-1) mitochondrial fusion promoter M1(MFP-M1) for 24 h and the expression of fibrosis markers Fn and CTGF was detected by Western blot. Western blot result showed that the levels of Col-Ⅰ, Col-Ⅲ, and p-Drp1 S616 were increased and the levels of p-Drp1 S637, Mfn1, Mfn2, TFAM, and ATP6 were decreased in 5/6(A/I) group compared with those in the sham group. The levels of Col-Ⅰ, Col-Ⅲ, and p-Drp1 S616 were significantly lower and the levels of p-Drp1 S637, Mfn1, Mfn2, TFAM, and ATP6 were significantly higher in ICA groups than that in 5/6(A/I) group. IHC staining demonstrated that for the expression of α-SMA in the renal interstitium was higher in the 5/6(A/I) group than in the sham group and that the expression in the ICA groups was significantly lower than that in the 5/6(A/I) group. Furthermore, the improvement in the fibrosis, mitochondrial dynamics, and mitochondrial function were particularly prominent in rats receiving the high dose of ICA. The in vitro experiment revealed that ICA dose-dependently inhibited the increase of Fn, CTGF, and p-Drp1 S616, increased p-Drp1 S637, Mfn1, Mfn2, TFAM, and ATP6, elevated ATP content, and improved mitochondrial morphology of NRK-52 E cells stimulated by TGF-β1. ICA combined with MFP-M1 further down-regulated the expression of Fn and CTGF in NRK-52 E cells stimulated by TGF-β1 compared with ICA alone. In conclusion, ICA attenuated RIF of CRF by improving mitochondrial dynamics.


Subject(s)
Adenosine Triphosphate/pharmacology , Animals , Female , Fibrosis , Flavonoids , Humans , Infarction/pathology , Kidney , Kidney Failure, Chronic , Male , Mitochondrial Dynamics , Rats , Rats, Sprague-Dawley , Renal Insufficiency, Chronic , Transforming Growth Factor beta1/metabolism
2.
Article in Chinese | WPRIM | ID: wpr-923498

ABSTRACT

@#To explore the effects and molecular mechanism of icariin on the vascular function of mice with type 1 diabetes induced by alloxan, type 1 diabetic mice model was established by intraperitoneal injection with 200 mg/kg alloxan.After oral administration with icariin (60, 120 mg/kg) daily for 2 weeks, blood glucose, body weight, food intake and water intake were detected.To evaluate the impact of icariin on the function of isolated vascular ring contraction and relaxation, thoracic aortas of mice were removed and the Ach-induced vascular ring relaxation, Phe-induced vascular ring contraction, SNP-induced vascular ring relaxation and KCl-induced vascular ring contraction response were detected.To further confirm the mechanism of icariin to improve vascular function, human umbilical vein endothelial cells (HUVECs) were induced by high glucose (HG) in vitro.Western blot was used to detect the effect of icariin on eNOS, p-eNOS, p38 MAPK and p-p38 MAPK expressions in HG-induced human umbilical vein endothelial cells (HUVECs).The results indicated that icariin significantly ameliorated the weight loss and dampened the increase in water intake of the diabetic mice.Meanwhile, icariin had a certain ameliorative effect on blood glucose and food intake without significant difference.The results of isolated thoracic aortas vascular rings contraction and vasodilation function indicated that icariin significantly improved Phe-induced vascular contraction and Ach‐induced vascular relaxation.Meanwhile, icariin had a certain ameliorative effect on KCl-induced vascular contraction response without significant difference.However, no significant change was observed on endothelium‐independent vascular rings relaxation response induced by SNP after treatment with icariin.Results of Western blot showed that icariin inhibited the expression of p-p38 MAPK and induced expression of p-eNOS in the high glucose-induced HUVECs cell model.Therefore, icariin may attenuate alloxan-induced type 1 diabetic mice vascular diastolic function by inhibiting expression of p-p38 MAPK and inducing expression of p-eNOS.

3.
Article in Chinese | WPRIM | ID: wpr-921702

ABSTRACT

This study evaluated the effects of epimedium polysaccharide(EPS) on the solubility of icariin and baohuoside Ⅰ so as to preliminary explore its solubilization function and the underlying mechanism. The solubility of these two insoluble flavonoids in water and polysaccharide solutions was compared by high performance liquid chromatography, and the mechanism was investigated by diffe-rential scanning calorimetry(DSC) and critical micelle concentration determination. The results indicated that their solubilization in crude EPS solutions was concentration-dependent. The solubility of icariin and baohuoside Ⅰ in 20 mg·mL~(-1) EPS-1-1 was 9.05 times and 5.76 times that in water, respectively; while their solubility in 20 mg·mL~(-1) EPS-2-1 was 10.55 and 8.39 times that in water, respectively. The change of the DSC thermograms suggested the formation of new complexes from icariin and baohuoside Ⅰ with polysaccharides. The critical micelle concentrations proved the micellar properties of both EPS-1-1 and EPS-2-1. In short, EPS can significantly increase the solubility of icariin and baohuoside Ⅰ, the mechanism of which may be related to the formation of micellar complexes between EPS and insoluble flavonoids.


Subject(s)
Epimedium , Flavonoids , Polysaccharides , Solubility
4.
China Pharmacy ; (12): 225-230, 2021.
Article in Chinese | WPRIM | ID: wpr-862648

ABSTRACT

OBJECTIVE:To establish the method for content determination of 6 components in Fuzheng guben granules ,such as 2,3,5,4′-tetrahydroxystilbene glucoside ,baicalin,icariin,scutellarin,baicalein and wogonin. METHODS :HPLC method was adopted. The determination was performed on Dikma Diamonsil C 18 column with mobile phase consisted of acetonitrile- 0.1% phosphoric acid aqueous solution (gradient elution )at the flow rate of 1.0 mL/min. The detection wavelengths were set at 275 nm (0-8 min),320 nm(8-9 min)and 275 nm(9-33 min). The column temperature was set at 25 ℃,and sample size was 10 μL. With baicalin as reference material ,the relative corr ection factors (fk/s) of other five components were calculated by multi-point correction method and slope correction method ;the retention time difference method was used to locate the chromatographic peaks ; the calculation values obtained by above 2 QAMS were compared with measured values of external standard method. RESULTS : The linear range of 2,3,5,4′-tetrahydroxystilbene glucoside ,baicalin,icariin,scutellarin,baicalein and wogonin were 0.053-2.12, 0.163-6.52,0.059-2.36,0.021 6-0.864,0.03-1.2,0.021-0.84 μg(r>0.999),respectively. RSDs of precision ,stability(12 h)and reproducibility tests were all lower than 3%. Average recoveries were 98.72%-99.82%(RSDs were 0.89%-1.24%,n=9). Using baicalin as reference material ,fk/s of multi-point correction method for 2,3,5,4′-tetrahydroxystilbene glucoside ,icariin,scutellarin, baicalein and wogonin were 1.172,0.528,1.479,1.820 and 2.534,respectively;fk/s of slope correction method were 1.234, 0.550,1.559,1.939,2.664. RSDs of 6 components in 10 batches of Fuzheng guben granules by 3 methods were 0.29%-2.77% (n=10),respectively. Pearson correlation coefficient was not lower than 0.999 9(P<0.001)in measured values between QAMS and external standard method. CONCLUSIONS :QAMS method is established successfully for simultaneous determination of 6 components in Fuzheng guben granules.

5.
China Pharmacy ; (12): 812-818, 2021.
Article in Chinese | WPRIM | ID: wpr-875813

ABSTRACT

OBJECTIVE:To study the improvement effects of ica riin(ICA)on cognitive function in schizophrenia model rats and its mechanism. METHODS :SD rats were divided into blank control group ,model group ,ICA low-dose ,medium-dose and high-dose groups (15,30,60 mg/kg). Except for blank control group ,other groups were given N-methyl-D-aspartate receptor antagonist MK- 801(0.2 mg/kg)intraperitoneally to induce schizophrenia rats models ,once a day ,for consecutive 14 days. After modeling,ICA groups were intragastrically administered with the corresponding drugs ,while blank control group and model group were intragastrically administered with the same volume of water ,once a day ,for consecutive 7 days. The behavioral com changes of rats were detected by Morris water maze test ,open field test , forced swimming test and Y maze test pathological changes of hippocampus were observed by Nissl staining;the levels of cholinergic indexes [acetylcholine (Ach),choline acetyltransferase (ChAT) and acetylcholinesterase (AchE)] in cerebral tissues were detected by ELISA. The expression of BDNF ,ERK and CREB mRNA in cerebral tissue were detected by RT-PCR ;expression or phosphorylation level of BDNF ,ERK,CREB protein ,apoptosis related proteins (Bcl-2,Bax and Caspase- 3)were detected by Western blot. RESULTS :Compared with blank control group ,escape latency ,distance at T 1-T3, cumulative immobility time and the expression of Caspase- 3 protein in cerebral tissues were significantly increased in model group (P<0.05);the times of crossing platform ,alternation rate ,the number of Nissl staining positive neurons in hippocampus tissues , the levels of Ach and ChAT in cerebral tissues ,Bcl-2/Bax ratio ,mRNA and protein expression of BDNF ,mRNA expression of ERK and CREB ,the phosphorylation of ERK 1/2 and CREB were significantly decreased (P<0.05).Compared with model group , escape latency ,distance at T 1-T3,cumulative immobility time ,the number of Nissl staining positive neurons ,AchE level in cerebral tissues and relative expression of Caspase- 3 protein were significantly decreased in ICA high-dose group (P<0.05);the times of crossing platform ,alternation rate ,levels of Ach and ChAT in cerebral tissues ,Bcl-2/Bax ratio ,mRNA and protein expression of BDNF ,mRNA expression of ERK and CREB ,the phosphorylation of ERK 1/2 and CREB were increased significantly(P<0.05). Above indexes in ICA low-dose and medium-dose groups were partially improved significantly than model group(P<0.05). CONCLUSIONS :ICA can improve cognitive function in schizophrenia model rats.Its mechanism may be related to regulating cholinergic system ,inhibiting neuronal apoptosis ,and promoting the expression of BDNF/ERK/CREB signaling pathway.

6.
Article in Chinese | WPRIM | ID: wpr-888151

ABSTRACT

This study aimed to observe the inhibitory effect of icariin against oxidative stress-induced calcification in aortic vascular smooth muscle cells(VSMCs) and elucidate the molecular mechanism of icariin in inhibiting endoplasmic reticulum stress(ERS)-mediated atherosclerotic calcification, so as to provide new ideas for exploring the anti-atherosclerotic mechanism of Epimedii Folium. The VSMCs in rat thoracic aorta were subjected to adherent culture and then treated with the complete calcification DMEM containing high glucose and hydrogen peroxide(H_2O_2) for three weeks. The resulting calcified VSMCs were divided into different treatment groups. Icariin was added one week after calcification induction for protecting the VSMCs, whose viability was then detected using cell counting kit-8(CCK-8). Alizarin red-S staining was conducted to observe the calcification degree. The activity of alkaline phosphatase(ALP) in VSMCs was measured using the disodium phenyl phosphate substrate and the calcium content was measured by arsenazo Ⅲ method. The mRNA expression levels of ossification-related factors including osteocalcin(OC), osteopontin(OPN), Runt-related transcription factor 2(Runx2), and type Ⅰ collagen(Col Ⅰa) were detected by real-time PCR. Western blot was carried out to determine the protein expression levels of α-smooth muscle actin(α-SMA), Runx2, activating transcription factor 4(ATF4), and eukaryotic translation initiation factor(eIF)-2α. The results showed that H_2O_2 significantly induced the calcification of VSMCs, increased the ALP activity and calcium content in VSMCs, promoted OC, OPN, Runx2, and Col Ⅰa mRNA expression and Runx2 protein expression, and reduced α-SMA protein expression. The ATF4 protein expression and eIF2α phosphorylation were also elevated significantly. Icariin reversed the calcification of VSMCs induced by H_2O_2, inhibited ALP activity and calcium content in VSMCs, down-regulated the mRNA expression levels of OC, OPN, Runx2 and Col Ⅰa and Runx2 protein expression, and relatively up-regulated the expression of α-SMA. The expression of ATF4 and phosphorylation of eIF2α also declined significantly. All these have demonstrated that icariin inhibited VSMCs calcification by down-regulating the ossification-related factors and lowering ALP activity and calcium content in VSMCs. Besides, the down-regulation of Runx2 expression and the inhibition of ATF4 and eIF2α-mediated cellular calcification pathway in ERS might also be involved in such calcification-suppressing process.


Subject(s)
Animals , Cells, Cultured , Flavonoids/pharmacology , Muscle, Smooth, Vascular/metabolism , Myocytes, Smooth Muscle , Oxidative Stress , Rats
7.
Article in Chinese | WPRIM | ID: wpr-848023

ABSTRACT

BACKGROUND: Icariin is the main effective component of Epimedium, which functions to tonify the kidney, and strengthen tendons and bones. In recent years, a large number of studies have found that icariin plays a significant role in the treatment of osteoarthritis. OBJECTIVE: To review the research progress in the molecular mechanism of icariin in the treatment of osteoarthritis. METHODS: The first author used “Icariin, Osteoarthritis, Cartilage, Subchondral bone, Synovial membrane, synovium, Inflammation" as search words in English and Chinese to search PubMed, CNKI, WanFang, and VIP databases. According to the inclusion criteria and exclusion criteria, 42 articles were included for final analysis. RESULTS AND CONCLUSION: Icariin can promote the cartilage differentiation of bone marrow mesenchymal stem cells and enhance the proliferation of cartilage cells and osteoblasts, to inhibit the degradation of cartilage extracellular matrix, reduce the activity of osteoclasts and alleviate synovial inflammation caused by inflammatory factors. It is an effective treatment for osteoporosis. However, the optimal effective dose and concentration safety of icariin still need a large number of experimental studies. Currently, most of the experiments are still in animal and tissue cell experiments. Numerous clinical studies are needed to continue to explore its specific mechanism in order to provide evidence-based medical evidence for icariin in the treatment of osteoarthritis.

8.
Article in Chinese | WPRIM | ID: wpr-847356

ABSTRACT

BACKGROUND: Nonunion is a common clinical complication in orthopedics, which seriously impacts the physical and mental health and quality of life of patients. In recent years, a large number of studies have found that icariin plays a significant role in promoting fracture healing and treating bone defects. Bone nonunion and fracture healing coexist, and the research on the mechanism of fracture healing actually focuses on the treatment of bone nonunion. OBJECTIVE: To review the research progress in the molecular mechanism of icariin in the treatment of bone nonunion. METHODS: The first author used “icariin, bone nonunion, bone marrow mesenchymal stem cells, periosteal cell, osteoblasts, osteoclast” as key words in English and Chinese to search PubMed, CNKI, WanFang and VIP databases. A total of 542 articles were retrieved and screened manually according to the selection criteria and exclusion criteria. Finally, 44 articles were included for result analysis. RESULTS AND CONCLUSION: Icariin can effectively promote fracture healing and treat bone nonunion by promoting the proliferation and differentiation of bone marrow mesenchymal stem cells and periosteal cells, promoting the proliferation and maturation of osteoblasts and inhibiting the osteoclast effect of osteoclasts. However, most of the experiments are still in the basic experimental research, and there is still a need for a large number of clinical studies as well as studies on related proteins and genes, to provide a new idea for the clinical use of Chinese herbs in the treatment of bone nonunion.

9.
Article in Chinese | WPRIM | ID: wpr-847318

ABSTRACT

BACKGROUND: Bone tissue engineering has provided a novel ideal for treating bone defects in clinic. This study is the first to combine traditional Chinese medicine with the nanostructures of tissue-engineered scaffolds in order to explore and construct a new bone tissue substitute material for the treatment of bone defects. OBJECTIVE: To investigate the osteogenic activity of icariin (ICA)/hydroxyapatite (HA)/poly(lactic-co-glycolic acid) (PLGA) composite scaffolds. METHODS: A HA/PLGA composite scaffold was prepared by physical blending of HA and PLGA, and was then soaked in ICA solution of different concentrations to obtain the HA/ICA/PLGA scaffold. Rabbit bone marrow mesenchymal stem cells were used to evaluate the cell adhesion, proliferation, osteogenesis and cytotoxicity of the composite scaffold. The cell adhesion, proliferation and cytotoxicity were detected by MTT method. The activities of alkaline phosphatase and osteocalcin were detected by ELISA. The expression levels of osteogenic genes and proteins were detected by fluorescence quantitative PCR and western blot assay, respectively. RESULTS AND CONCLUSION: Adding appropriate amount of HA into PLGA could improve the mechanical strength of the scaffold, and 10% HA had the best effect with tensile strength of (1.67±0.37) MPa, and compression modulus of (4.17±1.62) MPa, and nanostructure would be formed on the surface of the scaffold. The nanostructure could promote the adhesion of bone marrow mesenchymal stem cells on the surface of the scaffold. ICA did not affect the proliferation of bone marrow mesenchymal stem cells on the composite scaffold. However, the HA/PLGA composite scaffold soaked in 1.00 µmol/L ICA aqueous solution had the optimal osteogenic differentiation function, and the expression levels of alkaline phosphatase, osteocalcin, osteogenic related genes and proteins (Runx-2 and COL I) were increased. The ICA/HA/PLGA scaffold had no cytotoxicity. These results suggest that HA (10%)/ICA (1.00 µmol/L)/PLGA scaffold has good mechanical properties, osteogenesis and biocompatibility, which has the potential to be a favorable scaffold for bone tissue engineering.

10.
Article in Chinese | WPRIM | ID: wpr-846707

ABSTRACT

Objective: To compare the flavonoids contained in different parts of different botanical origins of Epimedii Folium, and provide a basis for the quality evaluation of Epimedii Folium and the reasonable selection of medicinal parts. Methods: The aerial parts of 13 batches of Epimedii Folium were collected and divided into three parts: leaf, petiole and stem. The HPLC fingerprint and content of five flavonoids, including epimedin A, epimedin B, epimedin C, icariin and baohuoside I, were analyzed. Then the analysis of variance and the similarity evaluation software of traditional Chinese medicine chromatographic fingerprint were used. Combined with cluster analysis (HCA), the content differences of flavonoids in leaf, petiole and stem of Epimedii Folium were evaluated. Results: The fingerprints showed that the chemical constituents in Epimedii Folium leaves were richer than stems and petioles, and the chemical constituents in petioles and stems were basically the same. The content of five components in leaves was significantly higher than that of petiole and stem, even up to 10 times. Cluster analysis also showed that the leaves were obviously distinct from the petiole and stem. Conclusion: The quality differences of Epimedii Folium leaves, petioles and stems were clarified, and this study can provide the scientific evidence for the selection of medicinal parts and quality control of Epimedii Folium.

11.
Article in Chinese | WPRIM | ID: wpr-846383

ABSTRACT

Objective: Based on UPLC-Q/TOF-MS technology, the fingerprint of Epimediumkoreanum before and after processing was established to analyze the whole composition and find out the iconic chemical composition to clarify the change rule of flavonoids. Methods: The data of E.koreanum raw and processed products were collected in positive ion mode using UPLC-Q/TOF-MS technology, and orthogonal partial least least squares-discriminant analysis (OPLS-DA) method was used to explore the differences in chemical composition of E.koreanum before and after processing in nine different origins and batches. Results: Nine iconic chemical constituents from E.koreanum raw and processed products were found and identified, including 8-ethylene-kaempferol, icaritin, icariin I, icartin-3-O-glucoside, isoamyl alcohol sagittatoside B,1,3-prenyl epimedin C, 1,3-prenyl-sagittatoside B-7-O-glucuronic acid, 3-O-((4-acetoxy) rhamnose-2-O-(m-2acetoxy) glucose) icariin and its isomers. Conclusion: The structures of E.koreanum's flavonoids changed after the processing, the secondary glycosides of flavonoids increased, the polyglycosides decreased, and the flavonoids were generally converted to low glycoside components, which further clarified the change rule of E.koreanum's flavonoids after processing.

12.
Article in Chinese | WPRIM | ID: wpr-846206

ABSTRACT

Objective: To investigate the best technological conditions for the purification of epimedin and icariin from Epimedii Folium by macroporous resin, and preliminarily characterize the purification fraction of the best technological conditions. Methods: Five kinds of macroporous resins were screened by static adsorption experiment with the content of epimedin A1, epimedin A, epimedin B, epimedin C and icariin as indexes. The best purification conditions were optimized by the concentration of upper column solution, the maximum sample volume, the upper column flow rate, the volume of water washing, the concentration of removing impurity ethanol and elution ethanol, the volume of removing impurity ethanol and elution ethanol, the column diameter-height ratio through dynamic adsorption experiment. Finally, UPLC-Q-TOF/MS, HPLC and ultraviolet spectrophotometry were used to characterize the purification fraction of the best technological conditions. Results: The best macroporous resin was AB-8, column diameter-height ratio was 1:7, 6 BV of upper column solution (crude drug 0.5 g/mL) was used for dynamic adsorption at a flow rate of 6 BV/h, 5 BV of water and 5 BV of 20% ethanol were used for impurity removal, and 6 BV of 50% ethanol was used for elution. The flow rate of impurity removal and elution was 6 BV/h. After purification, the total flavonoids content was 63.29%, the total content of epimedin A1, A, B, C and icariin was 40.48%, the content of epimedin A1, epimedin A, epimedin B, epimedin C and icariin was 1.63%, 2.52%, 16.36%, 5.51% and 14.46%, respectively. Conclusion: The purification process of epimedin and icariin from Epimedii Folium by AB-8 macroporous resin is stable, reasonable and feasible. The chemical characterization indicated that the purification fraction was mainly flavonoids, mainly consisting of epimedin and icariin. The optimized purification process can be used for the purification and enrichment of such ingredients.

13.
Article in Chinese | WPRIM | ID: wpr-845983

ABSTRACT

Objective: In order to provide a scientific basis for the quality control of Kunxian Capsules (KC), HPLC characteristics chromatogram combined with multi-components determination was established. Methods: The analysis was performed on Agilent Zorbax SB-C18 column (250 mm × 4.6 mm, 5 μm), using acetonitrile and 0.1% phosphoric acid solution as the mobile phase at a flow rate of 0.8 mL/min for gradient elution, the column temperature was 33 ℃, and the detection wavelength was 270 nm. The fingerprints of 15 batches of KC were established and evaluated by the similarity evaluation system of TCM (2012A version), hierarchical cluster analysis and discriminant analysis of partial least squares. Furthermore, the content of hyperoside, epimedin A, epimedin B, epimedin C, icariin and baohuoside Ⅰ were determined. Results: The HPLC fingerprint with 21 common peaks of KC was established, and the similarities of samples were over 0.9. The linearity relationships separated with hyperoside, epimedin A, epimedin B, epimedin C, icariin and baohuoside Ⅰ were good, and the contents of the above-mentioned components in 15 batches of preparations were 2.817-7.527, 7.287-9.103, 8.730-18.675, 33.377-70.371, 35.297-50.291 and 4.059-9.079 mg/g, respectively. Conclusion: The combination methods of HPLC characteristic chromatograms and simultaneous determinations of multiple components are rapid, simple and reproducible, which can provide methodological reference for the quality control of KC.

14.
Article in Chinese | WPRIM | ID: wpr-873350

ABSTRACT

Objective:To study the effect of icariin on renin homologous protein A (RhoA)/Rho-related kinase (ROCK) pathway in rats with nephrotic syndrome (NS) and its protective mechanism. Method:Totally 54 clean-grade male SD rats were tested and randomly divided into normal group, model group, RhoA inhibitor group (Rhosin, 40 mg·kg-1·d-1) and three doses of icariin groups (low, medium and high corresponding dose, 30, 60, 120 mg·kg-1·d-1). Adriamycin hydrochloride 6.5 mg·kg-1 was given in tail vein of rats to induce NS model in rats. After the model was established, peritoneal administration was carried out. The normal group and the model group were given saline 2.5 mL·d-1, and the inhibitor group and all of dose groups were given corresponding doses of Rhosin and icariin for intervention. Total urinary protein (Alb), creatinine (Cre), total urinary protein/creatinine ratio (A/C) kit were detected in rats, ultrastructure of kidney was identified by transmission electron microscopy (TEM), and Real-time fluorescent quantitative polymerase chain reaction (Real-time PCR) and Western blot were used to detect the mRNA and proteins expressions of RhoA, ROCK1, ROCK2. Result:TEM showed that the basement membrane was intact and the foot process was regular in the normal group, in model group, basement membrane was damaged seriously, foot process disappeared, and fusion was serious, in the low-dose group, the basement membrane injury was alleviated, the number and density of foot process were improved, and the fusion was obvious, in the middle-dose group and the inhibitor group, the basement membrane thickening was alleviated, and the foot process was slightly fused, in the high-dose group, the basement membrane structure was more complete, and podocytes were longer and arranged tightly. Compared with the normal group, the levels of Alb, Cre and A/C in urine, and RhoA, ROCK1 and ROCK2 mRNA and protein expressions in kidney tissue of rats of the model group were significantly higher (P<0.05). Compared with model group, the levels of Alb, A/C in urine and RhoA, ROCK1, ROCK2 mRNA and protein expressions in kidney tissue in the inhibitor group and low, medium and high-dose groups, and Cre in urine in inhibitor group and high-dose group decreased significantly (P<0.05). Compared with the inhibitor group, the levels of Alb, Cre in urine and RhoA protein in kidney tissue in the high-dose group were significantly decreased (P<0.05), the levels of Alb, Cre, A/C in urine and RhoA, ROCK1, ROCK2 mRNA and protein expressions in kidney tissue of the low-dose group, and the levels of RhoA, ROCK1 and ROCK2 mRNA expressions in kidney tissue of the middle-dose group were significantly increased (P<0.05). Compared with the low-dose group, the levels of Alb, A/C in urine, and RhoA, ROCK1, ROCK2 mRNA and protein expressions in kidney tissue in the middle and high-dose groups, Cre in urine of the high-dose group were significantly decreased (P<0.05). Compared with the middle-dose group, the levels of Alb, Cre in urine, and RhoA, ROCK1, ROCK2 mRNA and protein expressions, ROCK2 mRNA expression in kidney tissue in the high-dose group were significantly decreased (P<0.05). Conclusion:Icariin may protect glomerular endothelium and podocyte by affecting RhoA/ROCK pathway in the treatment of NS rats.

15.
Article in Chinese | WPRIM | ID: wpr-872791

ABSTRACT

Objective::To observe the effect of icariin on damaged neurons from the perspective of endoplasmic reticulum stress, in order to explore some mechanisms for repairing damaged neurons. Method::PC12 cells were induced by nerve growth factor (NGF) to differentiate into neurons, and the positive rate of microtubule associated protein-2 (MAP2) and neuron-specific enolase (NSE) expressions was determined by flow cytometry. The experiment was divided into 4 groups, blank control group: PC12 induced differentiation into neuronal cells, solvent control group: PC12 induced differentiation into neurons+ 0.1% dimethyl sulfoxide (DMSO), thapsigargin group: PC12 induced differentiation into nerves Yuan+ 2 μmol·L-1 thapsigargin, and icariin group: PC12 induced differentiation into neurons+ 2 μmol·L-1 thapsigargin+ 0.1 μmol·L-1 icariin. The proliferation of the cells was detected by using cell counting kit-8(CCK-8) method, the apoptosis of the cells was detected by flow cytometry, the protein expressions of CCAAT/enhace-binding protein homologous protein(CHOP) and glucoseregulated protein 78(Grp78) were detected by Western blot, and the mRNA expressions of CHOP and Grp78 were detected by real-time quantitative PCR (Real-time PCR). Result::Compared with the solvent control group, the thapsigargin group inhibited the proliferation of neuron-like PC12 cells induced by NGF, promoted apoptosis, and up-regulated the expressions of CHOP and Grp78 (P<0.05, P<0.01). Compared with the thapsigargin group, the icariin group can alleviate the inhibition of neurotrophic activity by thapsigargin, reduce neuronal apoptosis, and down-regulate the expressions of CHOP and Grp78 (P<0.05, P<0.01). Conclusion::Icariin can inhibit endoplasmic reticulum stress by down-regulating the expressions of CHOP and Grp78 and promote the repair of damaged neurons.

16.
Article in Chinese | WPRIM | ID: wpr-872728

ABSTRACT

Objective:To investigate the effect of icariin on neuroprotection in cerebral ischemia-reperfusion rats and microglia toll-like receptor 4 (TLR4)/nuclear transcription factor-κB (NF-κB) pathway. Method:In the blank group, blood vessels were only isolated but not ligated and blocked,and the rats were injected intraperitoneally with the same volume of normal saline. After successful modeling, they were randomly divided into model group, butyphthalide group (6 mg·kg-1), and high, medium and low (40,20,10 mg·kg-1)-dose icariin group,and abdominally administered with drugs at 5,12, 24 h after ischemia, respectively. The nerve function scores were detected, 2,3,5-triphenyltetrazole chloride (TTC) staining was used to measure the cerebral infarction rate,immunohistochemical assay was performed to detect the expressions of microglial markers ionized calcium binding adapter molecule 1(Iba1) and TLR4 in the rat brain cortex, Western blot immunoassay was used to detect the expression of NF-κB p65 in the cerebral cortex, and enzyme-linked immunosorbent assay (ELISA) was used to detect interleukin-1α (IL-1α) and tumor necrosis factor-α (TNF-α) content. Result:Compared with the sham-operation group, the nerve score, the cerebral infarction rate, the activations of Iba1 and TLR4 in microglial cells, the protein expression of NF-κB p65(P<0.01), and the contents of inflammatory factors IL-1α and TNF-α in the model group increased significantly(P<0.01). After treatment with icariin, compared with the model group, the neurological function score and the cerebral infarction rate of rats were improved, whereas the activations of Iba1 and TLR4 in microglia, the protein expression of NF-κB p65, and the contents of inflammatory factors IL-1α and TNF-α decreased obviously(P<0.05,P<0.01). Conclusion:Icariin may inhibit the activations of TLR4 and its downstream NF-κB signaling pathway and reduce the expression of relevant inflammatory factors IL-1α and TNF-α by regulating the activation of microglia, so as to play a protective role in the brain after stroke.

17.
Chinese Pharmacological Bulletin ; (12): 675-681, 2020.
Article in Chinese | WPRIM | ID: wpr-856972

ABSTRACT

Aim To study the protective effect of icariin on the impairment of rat bone mesenchymal stem cells (BMSCs) induced by 5-fluorouracil (5-FU) and the underlying mechanism. Methods The proliferation of BMSCs was evaluated by MTT assay, the morphological change of apoptotic cells was tested by using the chromatin dye Hoechst 33258, and the apoptotic rate was analyzed by flow cytometry. The proteins expression of cleaved caspase-3 was detected with Western blotting, the activity of caspase-3 was detected by Activity Assay Kit; iNOS and TNOS were detected by nitric oxide synthase (NOS) assay kit. Results MTT detection showed that 5-FU could reduce the survival rate of BMSCs, 10, 20, 40 μmol · L-1ICA could promote the proliferation of BMSCs, and blocked the inhibition of BMSCs cells growth induced by 5-FU. When BMSCs cells were treated by both ICA (10, 20 p, mol · L-1) and 5-FU (2.5×l0-2g· L-1) for 48 hours, compared with model group, the early apoptotic rates of ICA test groups were significantly reduced (P < 0. 01), the expression of cleaved caspase-3 and the activity of caspase-3 were significantly reduced (P < 0. 01), and the activity of iNOS and TNOS was inhibited in a certain degree (P < 0. 05). Conclusions ICA could protect BMSCs against 5-FU-induced damage. The protective role of ICA is probably related to reducing the activity of iNOS and TNOS, inhibiting the activation of caspase-3, and inhibiting the occurrence of apoptosis.

18.
Chinese Pharmacological Bulletin ; (12): 616-620, 2020.
Article in Chinese | WPRIM | ID: wpr-856961

ABSTRACT

Aim To investigate the effect of icariin (ICA) on the proliferation and differentiation of MC3T3-E1 pre-osteoblast cell line. Methods CCK-8 assay was used to detect the proliferation activity of MC3T3-E1 cells. Alizarin red staining (AR-S) assay was used to detect osteoblast differentiation and the formation of calcium nodules. Western blot assay was used to examine the protein expression of osteocalcin, ALP and shh. ALP assay was used to detect its activity. Results The effect of CCK-8 assay showed that ICA (5 ∼ 40 μmol · L-1) significantly promoted MC3T3-E1 cell proliferation compared with control group. The activity and protein expression of ALP and the quantitative analysis of mineralization deposition significantly increased in ICA group compared with control group. The ALP activity, protein expression level of shh and the quantification of alizarin red staining showed a significant decrease in ICA + CYC group compared with ICA group. Conclusions ICA promotes osteoblast proliferation and differentiation through the Hedgehog signaling pathway.

19.
Article in Chinese | WPRIM | ID: wpr-856392

ABSTRACT

Objective: To explore the effect of icariin on early steroid-induced osteonecrosis of the femoral head in rabbits. Methods: Fifty mature New Zealand rabbits (weighing, 2.5-3.0 kg) were randomly divided into control group ( n=10), model group ( n=20), and experimental group ( n=20). The rabbits of model and experimental groups were injected with lipopolysaccharide and methylprednisolone to establish the animal model of early steroid-induced osteonecrosis of the femoral head. The rabbits of experimental group were feeded with icariin solution once a day for 6 weeks since the first injection of methylprednisolone, whereas the rabbits of control and model groups were given normal saline at the same time points. The left femoral heads were removed after 6 weeks and gross morphological features were evaluated. Micro-CT scan was performed to analyze the trabecular microstructure with the following parameters: trabecular bone volume to total volume (BV/TV), trabecular number (Tb.N), trabecular thickness (Tb.Tn), and trabecular separation (Tb.Sp). The Micro-CT scan was also converted to three-dimensional reconstruction images for observation. HE staining was applied to observe the trabecular structure and morphological changes of osteocytes and marrow adipocytes. It was also used to determine whether the samples of femoral heads occurred osteonecrosis based on the criteria for pathological diagnosis, and calculate the rate of empty lacunae. Results: Seven rabbits died during the study, and 9, 16, and 18 rabbits in the control, model, and experimental groups, respectively, enrolled the final analysis. Compared with control group, the femoral head collapse and trabecular breaks were more obvious, and the trabeculae were sparse with irregular arrangement in the model group according to the results of gross observation, Micro-CT scan, and three-dimensional reconstruction images. But in the experimental group, the surface of femoral head was slight shrinking without obvious collapse, and the degeneration of trabecular structure was mild. According to bone microstructures analysis, the Tb.N, Tb.Tn, and BV/TV of femoral head in model and experimental groups were lower than those in control group, while the Tb.Sp in the model and experimental groups were significantly higher. The Tb.N, Tb.Tn, and BV/TV of femoral head in experimental group were higher than those in model group, while the Tb.Sp in the experimental group was significantly lower. The differences between groups were all significant ( P<0.05). In the model group, HE staining showed that the number of osteocytes reduced, the number of empty lacunae increased, and the marrow adipocytes piled up in the space between femoral trabeculae, some even mashed together like a cyst. In the experimental group, the trabecular structure was still relatively complete compared with model group, no obvious apoptosis of osteocytes was observed, the size and number of adipocytes were basically normal. None of the animals in control group occurred osteonecrosis of the femoral head based on the criteria for pathological diagnosis, and the incidence of osteonecrosis were 81.3% (13/16) in the model group and 66.7% (12/18) in the experimental group, and the difference was not significant ( P=0.448). The rate of empty lacunae of osteonecrotic femoral heads in the model group was 33.1%±1.4%, which was higher than that in experimental group (18.9%±0.8%) and in control group (12.7%±1.5%), and the differences between groups were significant ( P<0.05). Conclusion: The icariin has a protective effect on the early steroid-induced osteonecrosis of the femoral head in rabbits, which can decrease osteocytes apoptosis, improve the bone microstructure, and delay such disease processes.

20.
Article in Chinese | WPRIM | ID: wpr-828053

ABSTRACT

In this study, the growth index including plant height, compound leaf area, specific leaf area, leaf water content, number of branches, and leaf biomass per plant and the icariin flavonoids such as epimedin A, epimedin B, epimedin C and icariin of Epimedium pseudowushanense were determined on 30 d and 60 d under light intensity(18.2±2.5) μmol·m~(-2)·s~(-1)(L1) and(90.9 ±2.5) μmol·m~(-2)·s~(-1)(L2), and white light as control, red light, blue light and yellow light were used as three light quality treatments, to study the effect of light quality on the growth and flavonoids accumulation of E. pseudowushanense. The E. pseudowushanense was sui-table for growth under L1 light intensity, the blue light treatment significantly reduced the leaf area, but had little effect on the stem height, the red light treatment and the yellow light treatment had no obvious effect on the stem height and leaf area, but the yellow light treatment significantly increased the germination of new branches, and had a sustained promoting effect, and the biomass was significantly higher than the white light treatment at 60 d. The content of icariin flavonoids in red light, blue light and yellow light treatment was higher than that in white light treatment at 30 d and 60 d under L1 light intensity, while yellow light treatment promoted the synthesis of icariin flavonoids to the largest extent, which was 1.8 and 1.9 times of white light treatment(30 d and 60 d).Under L2 light intensity, the effect of strong light on promoting stem germination became the main factor, while the yellow light treatment showed no significant effect on promoting stem germination, and the red light treatment exhibited a significant effect on reducing leaf area. Icariin flavonoids under red light, blue light and yellow light treatment were all lower than that under white light treatment, that is, the effect of white light treatment on the synthesis of icariin flavonoids is better than red light, blue light and yellow light treatment. When the time of strong light treatment was longer, the degradation range of icariin flavonoids in other light treatment appeared, while red light treatment promotes the synthesis of icariin flavonoids. Therefore, the influence of light quality on E. pseudowushanense is quite different under different light intensity, no matter from growth index or flavonoid content index. The results support that the biomass and icariin flavonoid content can be increased by providing appropriate red and yellow light.


Subject(s)
Drugs, Chinese Herbal , Epimedium , Flavonoids , Plant Leaves
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