【Abstract】ObjectiveTo explore the effect of hepatocyte growth factor/scatter factor (HGF/SF) on apoptosis of colorectal cancer cells induced with curcumin. MethodsMTT assay was used to evaluate the cytotoxicity of curcumin to colorectal cancer cells. Flow cytometry was used to detect the antiapoptosis effect of HGF. ResultsFlow cytometry showed only 64 μg/ml curcumin could play the proliferationinhibiting role in Caco-2 cells leading to their apoptosis; at the same time, different concentrations of HGF could antagonize this inhibitory effect resulting in the decrease of apoptosis, but HGF worked without a concentration-dependent manner. The study on MAPK pathway showed that the protective effect of HGF on the apoptosis of Caco-2 cells was not influenced by inhibiting p42/p44 MAPK and p38 MAPK pathway. ConclusionHGF/SF antagonizes the apoptosis of Caco-2 cells induced with curcumin, but MAPK signaling pathway might not participate in this process.
Objective To study the inhibitory effects of curcumin on bleomycin-induced pulmonary fibrosis in rats at the fibrosing stage and explore its possible mechanism.Methods 96 male SD rats were randomly divided into a normal control group，a fibrosis model group，a fibrosis model treated with prednisone group and a fibrosis model treated with curcumin group.Pulmonary fibrosis were induced by instilled bleomycin through tracheal.From day 15 after bleomycin administration，the curcumin group and prednisone group were given curcumin（300 mg/kg） or prednisone（5 mg/kg） per day by intragastric administration，respectively.The normal control group and fibrosis model group were given 1% sodium carboxymethyl cellulose（10 mL/kg） as control.Six rats of each group were randomly sacrificed on day 21，28，42 and 56 after bleomycin administration，respectively.The histological changes of the lung were evaluated by HE and Masson’s trichrome staining.Lung expressions of transforming growth factor-β1（TGF-β1） and hydroxyproline were assessed by immuno-histochemistry and digestion method，respectively.Results Pulmonary fibrosis and hydroxyproline level in the curcumin group were significantly reduced as compared with those in the model group on day 42 and 56.The expession of TGF-β1 in the curcumin group was significantly lower than that in the model group on day 28，42 and 56，and was not significantly different from the normal group on day 56.Conclusion Curcumin could alleviate bleomycin-induced pulmonary fibrosis in rats at the fibrosing stage by inhibiting the expressions of TGF-β1.
Objective To investigate the proliferation inhibitory effect and to explore the molecular mechanism of curcumin on pulmonary fibroblasts. Methods Fibroblasts derived from lung tissue of patients with idiopathic pulmonary fibrosis ( IPF) was cultured in vitro and incubated with curcumin at different concentrations for different time. Fibroblasts were randomized into 5 groups, ie. a control group and 4 curcumin groups ( intervened by 5, 10, 20, 40 μmol / L curcumin, respectively) . MTT assay was used to determine the inhibitory rate of curcumin on the proliferation of pulmonary fibroblasts. Apoptosis and the Caspase-3 expression of pulmonary fibroblasts were identified by flow cytometry ( FCM) . Variables were compared with One-Way ANOVA. The correlations between variables were analyzed using Pearson’scorrelation coefficient. Results Curcumin inhibited pulmonary fibroblasts proliferation in a dose-dependent and time-dependent manner( r =0. 886, r = 0. 832, respectively, all P lt; 0. 01) . Apoptosis rate of pulmonary fibroblasts in 4 curcumin groups was ( 29. 58 ±2. 13) % , ( 64. 36 ±3. 92) %, ( 72. 98 ±4. 42) % , ( 83. 14 ±2. 51) % , respectively, which was significantly higher than that in the control group[ ( 3. 84 ±1. 88) % , P lt;0. 01] . The positive expression rate of apoptosis-regulating protein caspase-3 was ( 26. 24 ±3. 64) % ,( 44. 87 ±5. 31) % , ( 57. 44 ±4. 23) % , ( 73. 65 ±5. 01) % , respectively, which was significantly higher than that of the control group[ ( 4. 02 ±0. 62) % , P lt; 0. 01] . Conclusions In vitro, curcumin can significantly inhibit proliferation and induce apoptosis of pulmonary fibroblasts of patients with IPF. The mechanism maybe associated with up-regulating expression of Caspase-3.
Objective To investigate the effect of curcumin on calcitionin gene related peptide (CGRP) expression after spinal cord injury (SCI) in rats. Methods A total of 200 rats, weighing 250-300 g, were randomly divided into 4 groups (n=50): sham-operation group, normal saline (NS) group, low-dose curcumin group (30 mg/kg), and high-dose curcumin group (100 mg/kg). In sham-operation group, only vertebral lamina excision was performed without SCI; the SCI model was established in the other 3 groups. At immediate after modeling, 30 mg/kg and 100 mg/kg curcumin were injected intraperitoneally in 2 curcumin groups, equivalent NS was given in NS group (30 mg/kg), but no treatment in sham-operation group. At 1, 3, 7, 14, and 21 days after operation, the motor neural function was evaluated by the inclined plane test and Basso-Beattie-Bresnahan (BBB) scores; immunohistochemical staining and Western blot assay were used to observe CGRP expression. Results BBB score and inclined plane test score of NS group, low-dose curcumin group, and high-dose curcumin group were significantly lower than those of sham-operation group at each time point (P lt; 0.05). BBB score of low-dose curcumin group and high-dose curcumin group was significantly higher than that of NS group at 3, 7, 14, and 21 days after SCI (P lt; 0.05), and the score of high-dose group was significantly higher than that of low-dose curcumin group at 7, 14, and 21 days after SCI (P lt; 0.05). Inclined plane test score of low-dose curcumin group and high-dose curcumin group was significantly higher than that of NS group at 7, 14, and 21 days after SCI (P lt; 0.05), and the score of high-dose curcumin group was significantly higher than that of low-dose curcumin group at 7, 14, and 21 days after SCI (P lt; 0.05). Immunohistochemical staining results showed that the CGRP positive cells of sham-operation group was significantly more than those of the other 3 groups, and the CGRP positive cells of high-dose curcumin group were significantly more than those of low-dose curcumin group at each time point (P lt; 0.05); the CGRP positive cells of low- and high-dose curcumin groups were significantly more than those of NS group at 3, 7, 14, and 21 days after SCI (P lt; 0.05). Western blot assay results showed that the CGRP protein expressed at each time point after SCI in sham-operation group; the CGRP protein expression gradually decrease with time passing in NS group; but the CGRP protein expression gradually increased with time passing in low- and high-dose curcumin groups, and reached the peak at 14 days, then maintained a high level. Conclusion After SCI in rats, 30 mg/kg curcumin can improve rats’ motor function, and 100 mg/kg curcumin effect is more obvious, especially in promoting the expression of CGRP. That may be the mechanism of protection of the nervous system.
To observe the effect of different dosage of curcumin on expression of MMP-2 and MMP-9 in the tissue of cystiform in air-pouch mouse models after the injection of polyethylene wear particles, and to investigate its mechanism of intervening inflammatory response induced by wear particles. Methods Seventy-two kunming strain mice were used to establ ish air-pouch animal models by referring to the method of Yang et al. and injecting 3 mL suspension of ultrahigh molecular weight polyethylene wear particles (concentration 1 × 108 cells/mL) into dorsal cyst cavity. Then the animals were randomized into 3 groups (n=24 per group): group A (control group), 0.6 mL/day normal sal ine by gavage; group B(low-dosage experimental group), 0.6 mL/day curcumin solution at a concentration of 1.6 mg/mL by gavage; group C (highdosage experimental group), 0.6 mL/day curcumin solution at a concentration of 3.2 mg/mL by gavage. General condition of the animals was observed after operation. The mice were killed 3, 7 and 14 days after operation (8 mice per group at a time), the tissue of cystiform was harvested to receive gross, histology and immunohistochemistry observation, as well as RT-PCR and Western blot detection. Results All mice survived till the end of experiment. White cystiform tissue was evident on the back of mice subcutaneously in each group. For diameter of the cyst cavity at each time point, group A was obviously greater than groups B and C, and group C was significantly less than group B. Microscope observation showed that inflammatory response in group A was ber than that of groups B and C, and group C was obviously less than group B at 7 and 14 days. There was a significant difference between groups B and C and group A in terms of MMP-2 and MMP-9 expression at 7 and 14 days after curcumin del ivery (P lt; 0.05), and no significant differences were evident at 3 days (P gt; 0.05). There was no significant difference between group B and group C in MMP-2 expression at 7 days after curcumin del ivery (P gt; 0.05), and significant difference was evident at 14 days (P lt; 0.05). There was significant difference bewteen group B and group C in MMP-9 expression at 7 and 14 days after curcumin del ivery (P lt; 0.05). Nuclear translocation of NF-κB P65 was inhibited remarkably after curcumin del ivery,and there were significant differences among three groups at 7 and 14 days (P lt; 0.05), and no significant differences were evident at 3 days (P gt; 0.05). Conclusion Ultra-high molecular weight polyethylene wear particles can stimulate expression of MMP-2 and MMP-9 in cystiform tissue. Curcumin can restrain expression of MMP-2 and MMP-9 in cystiform tissue of air-pouch animal models, and expression of MMP-2 and MMP-9 may be regulated by the activation of NF-κB.
Objective To investigate the effects and mechanism of curcumin on the retinal neovasularization in mice with oxygeninduced retinopathy (OIR). Methods A total of 72 C57BL/6J mice were divided into normal, OIR model, vehicle control ［dimethyl sulphoxide (DMSO)］, and curcumin group (100, 50, and 10 mg). The mice in normal group lived in normoxia condition; OIR model was set up according to standard methods in the literature. Five days after OIR establishment, the mice in curcumin group received an intraperitoneal (IP) injection of 0.1 ml curcumin (100, 50, and 10 mg), and the mice in DMSO group received an IP injection of 0.1 ml 1permil; DMSO. All of the mice were executed at the age of postnatal day 17 (P17) and the eyeballs were collected. Endothelial cell nuclei breaking through the internal limiting membrane were counted after stained with hematoxylin and eosin (HE). The expression of vascular endothelial growth factor-A (VEGF-A), vascular endothelial growth factor receptor-2 (VEGFR-2), endostatin (ES), and phosphorylated p38 mitogen-activated protein kinase (p-p38MAPK) in the retina in each group were measured by real-time polymerase chain reaction (RT-PCR) and Western blot methods.Results Compared with the normal group, retinal neovascularization was found in OIR model group (P＜0.05). The number of endothelial cell nuclei was 46.00plusmn;16.00 in OIR model group and 0.17plusmn;0.41 in normal group (P＜0.05). The expression of VEGF-A, ES, and p-p38MAPK in 100 mg curcumin group differed statistically from which in 50 and 10 mg curcumin group (P＜0.05). The expression of VEGFR-2 was same in the three curcumin groups (P＞0.05). Conclusion Curcumin can inhibit the formation of retinal neovascularization; the mechanism may be associated with inhibiting the expression of VEGFA and VEGFR-2, increasing the expression of ES, and inhibiting the p38MAPK signal transduction pathway.
Objective To observe the inhibition effect of curcumin on the proliferation of rabbit retinal pigment epithelial (RPE) cells and investigate its mechanism. Methods The 4th generation of RPE cells were selected and divided into curcumin group and blank control group. The concentration of curcumin included 10, 15, and 20 mu;g/ml. The MTT assay was used to evaluate the inhibition effect on the proliferation of RPE cells at the 24th, 48th, 72nd and 96th hour after cultured with curcumin (10, 15, and 20 mu;g/ml). The IC50 value of curcumin at different time points were calculated by Linear Regression. Flow cytometry was used to detect the effect on the cell cycle at the 72nd hour after cultured with curcumin (15 mu;g/ml); the expression and apoptosis of proliferating cell nuclear antigen (PCNA) were also determined at the 24th,48th, and 72nd hour after cultured with curcumin (15 mu;g/ml) respectively. The configuration of RPE cells were observed by transmission electron microscope. Results The IC50 value of curcumin at the 24th,48th, 72nd and 96th hour was 29.31, 17.50, 13.24, and 10.99 mu;g/ml respectively. Cell cycel analysis indicated that curcumin blocked cells in G0/G1 phase. At the 24th, 48th, and 72nd hour after cultured with curcumin (15 mu;g/ml), the expression of PCNA of RPE cells were 565.04plusmn;23.60, 473.61plusmn;36.88, and 396.15plusmn;32.45; the apoptosisrate were （12.83plusmn;0.13）%，（32.27plusmn;4.51）%，（56.81plusmn;8.67）%, respectively. The differeces of curcumin groups compared with the control group were significant (P＜0.05). Apoptosis of RPE cells was observed under transmission electron microscope. Conclusions Curcumin can inhibite the proliferation of RPE cells by inhibit the synthesization of PCNA and inducing the apoptosis of RPE cells. Curcumin may become a potential drug to prevent and treat PVR.
Objective To investigate the effects of curcumin on oxidative stress in the co-culture system including human fetal lung fibroblasts and A549 cells, and discuss the potential and protective mechanism of the prophylactic effect of curcumin on pulmonary fibrosis. Methods The human fetal lung fibroblasts co-cultured with A549 cells were divided into five groups. The cells in the control group were cultured in DMEM without TGF-β1 or curcumin. The cells in the TGF-β1 group were cultured in DMEM containing 5 ng/mL TGF-β1 . In three TGF-β1 + cucurmin treatment groups, the cells were cultured in DMEM containing 5 ng/mL TGF-β1 and three different concentration of curcumin( 5, 10, 20 μmol /L, respectively) . ELISA was used to analyze the content of TNF-α. Serum level of MDA and SOD were tested by spectrophotometric analysis. Intracellular ROS production was detected by flow cytometry. NF-κB was measured by western blot. Results The serum MDA, intracellular ROS, the content of TNF-αand NF-κB protein expression in the TGF-β1 group were significantly increased while the activity of SOD was significantly decreased( P lt; 0. 01) , suggesting that the oxidative level of human fetal lung fibroblasts was obviously increased after TGF-β1 stimulation. After intervening by different concentration of curcumin, the serum MDA, intracellular ROS, content of TNF-αand NF-κB were significantly decreased while the activity of SOD was obviously increased( P lt;0.01) . Conclusion Low concentration of curcumin can reduce the oxidative level of human fetal lung fibroblasts co-cultured with A549 after TGF-β1 stimulation, and significantly increase the level of SOD, implying that curcumin may intervene pulmonary fibrosis by reduce oxidative level.
ObjectiveTo prepare curcumin loaded monomethoxyl poly(ethylene glycol)-poly(lactic-co-glycolicacid) (mPEG-PLGA) nanopaticles (CUR-NPs), investigate the effect of curcumin (CUR) and CUR-NPs on reversing corticosteroid resistance induced by cigarette smoke extract (CSE), and compare biological function between CUR and CUR-NPs in macrophages RAW264.7. MethodsmPEG-PLGA nanoparticles loaded with CUR were prepared via emulsion solvent evaporation.In lipopolysaccharide (LPS) stimulated macrophages RAW264.7, budesonide (BUD) was used to treat macrophages RAW264.7.In LPS and CSE stimulated macrophages RAW264.7, BUD (10-10-10-5 mol/L), CUR(10-10-10-5 mol/L), CUR(10-7 mol/L)+BUD(10-9-10-5 mol/L), CUR(10-9-10-5 mol/L)+BUD(10-7 mol/L), and CUR-NPs(10-9-10-5 mol/L)+BUD(10-7 mol/L) were respectively used to treat macrophages RAW264.7 activated.The level of IL-8 in cell culture supernatant was measured by ELISA.In CSE stimulated macrophages RAW264.7, CUR(10-7 and 10-6 mol/L) and CUR-NPs(10-7 and 10-6 mol/L) were used to treat macrophages RAW264.7.The mRNA level of HDAC2 was measured by real-time PCR, the protein level of HDAC2 was measured by Western blot.Cellular uptake of CUR and CUR-NPs in macrophages RAW264.7 was determined by cellular fluorescence intensity observed and detected by laser confocal microscopy imaging. ResultsThe morphology of CUR-NPs was spherical and the mean particle size was (356.4±146.6)nm.Compared with LPS stimulation, co-stimulation of LPS and CSE led to a significant decrease in the maximum inhibitory rate of BUD on IL-8 (P < 0.05) and a significant increase in the 50% inhibitory concentration (IC50) of BUD on IL-8 (P < 0.05).When using LPS+CSE to stimulate, compared with BUD (10-10-10-5 mol/L) group, the maximum inhibitory rate of BUD in CUR (10-7 mol/L)+BUD (10-9-10-5 mol/L) group on IL-8 was significantly higher (P < 0.05) and the IC50 of BUD decreased significantly (P < 0.05).When using LPS+CSE to stimulate, CUR and CUR-NPs in 10-9, 10-8 and 10-7 mol/L concentration, the inhibitory rate of CUR-NPs+BUD (10-7 mol/L) on IL-8 was significantly higher than that of CUR+BUD (10-7 mol/L) (P < 0.05). CSE stimulation induced a significant decrease in the mRNA and protein expression of HDAC2. Compared with CSE group, the mRNA and protein levels of HDAC2 of CUR(10-7 and 10-6 mol/L) group and CUR-NPs(10-7 and 10-6 mol/L) group were significantly higher (P < 0.05).In 10-7 mol/L concentration, the mRNA and protein levels of HDAC2 in CUR-NPs group were significantly higher than those in CUR group.In 10-7 mol/L concentration, cellular uptake of CUR in CUR-NPs was significantly higher than the native CUR. ConclusionsCUR and CUR-NPs can reverse the corticosteroid resistance induced by CSE.CUR-NPs can improve the cellular uptake of CUR.In the case of low concentration, CUR-NPs have more biological activity than CUR.
ObjectiveTo investigate the effect of Curcumin combined with Rhodiola on rats with severe acute pancreatitis (SAP) associated renal injury and explore the possible mechanisms. MethodsA total of 24 rats were randomly divided into SAP with renal injury group (SAP group, n=8), Curcumin group (n=8), Curcumin combined with Rhodiola group (n=8).The SAP group was given 1.5 mL saline through intragastric administration before operation while the Curcumin group was fed with same amount of Curcumin diluent.The Curcumin combined with Rhodiola group was given 1.5 mL Curcumin diluent through intragastric administration and 6 g/kg Rhodiola diluent through intraperitoneal injection before operation.The pancreas and pancreatic tail-segment was dissociated and the head of pancreas were occluded in rats to make the model, blood vessel forceps was loosed after three hours.All the rats were sacrificed at 18 h after modeling.The levels of serum amylase, creatinine, blood urea nitrogen were detected and pathological changes of pancreas and the left kidney were observed under the light microscope.The cell apoptosis was analyzed using TUNEL staining.The serum levels of interleukin (IL)-1β, IL-6, and IL-10 among the three groups were detected by enzyme-linked immunosorbent assay.The expression of inducible nitric oxide synthase (iNOS) mRNA in the right kidney was detected by real-time polymerase chain reaction.The superoxide dismutase (SOD) activity of the renal tissue was determined by hydroxylamine method. ResultsCompared with the SAP group, the levels of serum amylase, creatinine, blood urea nitrogen, IL-1β, IL-6, the cell apoptosis index, and the expression of iNOS mRNA were significantly decreased, the serum level of IL-10 and the activity of SOD were significantly increased (P < 0.05), the pancreas and the kidney damaged more slightly in the Curcumin group and Curcumin combined with Rhodiola group.Compared with the Curcumin group, the above situations were more better in the Curcumin combined with Rhodiola group. ConclusionsCurcumin combined with Rhodiola has a better protective effect on SAP associated renal injury.It might be through inhibiting the expressions of IL-1β, IL-6, stimulating the expression of IL-10, down-regulating the expression of iNOS mRNA, and improving the activity of SOD.It could reduce the cell apoptosis and necrosis of the kidney and improve the ability of the kidney to tolerate hypoxia.