The present study was undertaken to determine the effect of epigallocatechin gallate (EGCG) on lipopolysaccharide (LPS)-induced production of inflammatory chemokine regulated upon activation normal T cell expressed and secreted (RANTES) in human retinal endothelial cells (HRECs) and to explore the underlying regulatory mechanism. HRECs were stimulated with LPS in the presence or absence of EGCG at various concentrations (100, 50, 25, 12.5, 6.25 μmol/L). The optimum concentration of drug was determined by a real-time cell-electronic sensing (RT-CES) system, and MTS chromatometry was used to detect the toxicity of LPS and EGCG on HRECs. RANTES production in the culture supernatant was measured by ELISA. The expression levels of Akt and phosphorylated Akt were examined by Western blot assay. The result showed that LPS markedly stimulated RANTES secretion from HRECs. EGCG treatment significantly suppressed LPS-induced RANTES secretion in a dose-dependent manner. Furthermore, EGCG exhibited a dose-dependent inhibitory effect on LPS-induced phosphorylation of Akt. Taken together, our data suggest that EGCG suppresses LPS-induced RANTES secretion, possibly via inhibiting Akt phosphorylation in HRECs.
Catechin ; analogs & derivatives ; pharmacology ; Cells, Cultured ; Chemokine CCL5 ; metabolism ; Endothelial Cells ; metabolism ; Humans ; Lipopolysaccharides ; Phosphorylation ; Proto-Oncogene Proteins c-akt ; metabolism ; Retina ; cytology

OBJECTIVETo explore the effects of three Chinese herbal antidotes, i.e. Herba Artemisiae annuae (A), Herba Hedyotis diffusae (H) and Rhizoma Cimicifugae (C), all were ingredients of Jiedu Quyu Ziyin Recipe, for adjusting the regulated on activation normal T cell expressed and secreated (RANTES), gene expression in serum and renal tissue of MRL/lpr mice.

METHODSFifty-four MRL/lpr mice were randomized into 9 groups, with 6 in each, and intragastrically infused with A, H, C, A+H, H+C, A+C, A+H+C (all in dosage-form of decoction), prednisone suspension and physiological saline, respectively for 12 weeks. RANTES expression in serum and renal tissue of animals were detected with ELISA and RT-PCR at the end of the study.

RESULTSLevels of RANTES expression was significantly reduced in the prednisone treated group after treatment. Excepting no significant change being observed in the groups treated with A and C, the changes in the other groups were all milder than those in the group treated with A+H+C.

CONCLUSIONChinese herbal antidotes A, H and C in combination can significantly inhibit the RANTES expression in serum and renal tissue of MRL/lpr mice.

Animals ; Artemisia annua ; chemistry ; Chemokine CCL5 ; blood ; metabolism ; Cimicifuga ; chemistry ; Drugs, Chinese Herbal ; pharmacology ; Female ; Hedyotis ; chemistry ; Kidney ; metabolism ; Mice ; Mice, Inbred MRL lpr ; T-Lymphocytes ; immunology ; metabolism

OBJECTIVETo evaluate the effects of Artesunate on tumor necrosis factor alpha (TNF-alpha), monocyte chemoattractant protein (MCP-1), and on reduced activation normal T cell expressed and secreted (RANTES) in the serum and the synoviocyte culture supernate of collagen-induced arthritis (CIA) rats.

METHODSEighty male Wistar rats were selected to establish the CIA rat model. On the 6th day after modeling, 60 rats with the sum of arthritis index of right metapedes and two propodium > or = 6 were selected, and randomly divided into 6 groups (n = 10), i.e., the blank control group, the CIA model control group (treated with normal saline, abbreviated as the CIA group), the MTX positive control group (abbreviated as the MTX group), the large dose Artesunate group (at the daily dose of 20 mg/kg), the moderate dose Artesunate group (at the daily dose of 10 mg/ kg), and the small dose of Artesunate group (at the daily dose of 2.5 mg/kg). Mice were sacrificed 7 days of immune injection and their venous blood was collected to obtain the serum. Meanwhile, the synovial tissues of the knee joint were taken by aseptic techniques and primary cultured for 48 h. The supernate was collected by centrifuge. The changes of MCP-1, RANTES, and TNF-alpha in the serum and the synoviocyte culture supernate were observed in each group before and after treatment using ELISA.

RESULTSArtesunate significantly decreased the expressions of TNF-alpha in the serum and the synoviocyte culture supernate, showing significant difference when compared with the model control groups (P < 0.05). There was no statistical difference in the large dose Artesunate group and the moderate dose Artesunate group when compared with the MTX group (P > 0.05). But statistical difference existed in the large dose Artesunate group, the moderate dose Artesunate group, and the MTX group when compared with the small dose Artesunate group (P < 0.05). Artesunate could significantly decrease the expressions of MCP-1 and RANTES in the serum and the synoviocyte culture supernate, showing statistical difference when compared with the model control group (P < 0.05). But no statistical difference existed when compared with the MTX group (P > 0.05).

CONCLUSIONThe mechanism of anti-inflammatory action and immune regulation of Artesunate might be correlated with the inhibition of inflammatory factor TNF-alpha and chemotactic factors MCP-1 and RANTES.

Animals ; Artemisinins ; pharmacology ; Arthritis, Experimental ; blood ; metabolism ; Cells, Cultured ; Chemokine CCL2 ; blood ; metabolism ; Chemokine CCL5 ; blood ; metabolism ; Disease Models, Animal ; Male ; Rats ; Rats, Wistar ; Synovial Fluid ; cytology ; metabolism ; Tumor Necrosis Factor-alpha ; blood ; metabolism

OBJECTIVEA study was conducted to investigate the effects of Porphyromonas gingivalis lipopolysaccharide (Pg-LPS) on the expression of regulated upon activation normal T-cell expressed and secreted (RANTES) and fractalkine in human umbilical vein endothelial cells (HUVECs).

METHODSHUVECs were incubated with different concentrations of Pg-LPS (200, 500, and 1000 ng x mL(-1)) for 1, 6, 12, and 24 h, respectively. Then real time quantitative polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent method (ELISA) were adopted to detect the protein levels and mRNA levels of RANTES and fractalkine.

RESULTSThe RANTES protein levels and mRNA levels, as well as fractalkine mRNA levels, were significantly higher in all experimental groups of 1, 6, and 12 h than in the control group (P<0.05), except the expression of RANTES mRNA in 200 ng x mL(-1) group of 12 h and RANTES protein in 200 ng x mL(-1) group of 1 h. The expression levels of RANTES mRNA and fractalkine mRNA were highest in 1000 ng x mL(-1) group of 6 h and were 4.88- and 6.20-fold higher, respectively, than those in the control group. The expression levels of RANTES protein, mRNA, and fractalkine mRNA decreased 6 h after stimulation, and were significantly higher than those in the control group (P<0.05) in the RANTES and fractalkine in HUVEC, and such expression is important in the development of atherosclerosis 500 ng x mL(-1) group of 24 h. There was a significant difference between the expression of fractalkine mRNA in 1000 ng x mL(-1) group of 6 and 12 h than in the control group (P<0.05).

CONCLUSIONPg-LPS infection might up-regulate the expression of RANTES and fractalkine in HUVEC, and such expression is important in the development of atherosclerosis.

Atherosclerosis ; Cells, Cultured ; Chemokine CCL5 ; genetics ; metabolism ; Chemokine CX3CL1 ; analysis ; genetics ; metabolism ; Enzyme-Linked Immunosorbent Assay ; Human Umbilical Vein Endothelial Cells ; metabolism ; Humans ; Lipopolysaccharides ; pharmacology ; Porphyromonas gingivalis ; immunology ; isolation & purification ; RNA, Messenger ; analysis ; Reverse Transcriptase Polymerase Chain Reaction ; Up-Regulation

This study aimed to explore the protective effect of Reduning Injection(RDN) on mice infected by influenza virus A/PR/8(PR8) and its immune regulatory roles during viral infection. In in vivo experiments, female C57 BL/6 mice were randomly divided into phosphate buffered saline(PBS) group, PR8-infected group, oseltamivir treatment group(OSV) and RDN treatment group. After 2 h of PR8 infection, mice in the oseltamivir group were gavaged with oseltamivir 30 mg·kg~(-1), and those in the RDN treatment group were injected intraperitoneally with RDN 1.5 mL·kg~(-1)once per day for seven consecutive days. The body weight of mice in each group was recorded at the same time every morning for 16 consecutive days. The line chart of body weight change was created to analyze the protective effect of RDN on flu-infected mice. The relative mRNA expression of different cytokines(IL-6, TNF-α, MCP-1, IL-1β, MIP-2, IP-10 and IL-10) in lung samples of flu-infected mice was detected by PCR. Flow cytometry was utilized to analyze the composition of immune cells of mouse BALF samples on day 5 after infection. Mouse macrophage cell line RAW264.7 was planted and treated by different concentrations of RDN(150, 300, 600 μg·mL~(-1)) for 24 h or 48 h, and cell proliferation was detected by CCK-8 assay. RAW264.7 cells and mouse primary peritoneal macrophages were stimulated with synthetic single stranded RNA(R837), which elicited the inflammatory response by mimicking the infection of single-stranded RNA viruses. The expression of cytokines and chemokines in the supernatants of above culture system was detected by ELISA and qPCR. On days 4, 5, 6, 7 and 15 after infection, the body weight loss of mice in the RDN treatment group was alleviated compared with that of PR8-infected mice(P<0.05). RDN treatment obviously reduced lung index and the production of IL-6, TNF-α, MCP-1 and MIP-2 in lung tissues of flu-infected mice(P<0.05). The proportions of macrophages, neutrophils and T cells in mouse BALF samples were analyzed by flow cytometry, and compared with PR8-infected mice, RDN decreased the proportion of macrophages in BALF of flu-infected mice(P<0.05), and the proportion of T cells was recovered dramatically(P<0.001). In CCK-8 assay, the concentrations of RDN(150, 300, 600 μg·mL~(-1)) failed to cause cytotoxicity to RAW264.7 cells. In addition, RDN lowered the expression of inflammatory cytokines such as IL-6, TNF-α,MCP-1, IL-1β, RANTES, and IP-10 and even anti-inflammatory cytokine IL-10 in R837-induced macrophages. RDN reduced the infiltration of inflammatory macrophages and the production of excessive inflammatory cytokines, alleviated the body weight loss of flu-infected mice. What's more, RDN restored the depletion of T cells, which might prevent secondary infection and deteriorative progression of the disease. Taken together, RDN may inhibit cytokine production and therefore down-regulate cytokine storm during the infection of influenza virus.
Animals ; Anti-Inflammatory Agents/pharmacology* ; Body Weight ; Chemokine CCL5/pharmacology* ; Chemokine CXCL10/pharmacology* ; Cytokine Release Syndrome ; Cytokines/genetics* ; Drugs, Chinese Herbal ; Female ; Imiquimod/pharmacology* ; Interleukin-10 ; Interleukin-6 ; Lung ; Mice ; Mice, Inbred C57BL ; Oseltamivir/pharmacology* ; Phosphates/pharmacology* ; RNA ; RNA, Messenger ; Sincalide/pharmacology* ; Tumor Necrosis Factor-alpha/genetics* ; Weight Loss

OBJECTIVETo investigate the effects of progesterone and progestin on the expressions of regulated on activation, normal T cell expressed and secreted (RANTES) in eutopic endometrium from patients with endometriosis.

METHODSWe collected the samples of endometrium from patients with endometriosis before operation or after insertion of levenorgestrel releasing intrauterine system (LNG-IUS), administration of oral medroxyprogesterone (MPA), or injection of gonadotrophic hormone releasing hormone agonist (GnRHa). Reverse transcription-polymerase chain raction was used to assay the expression of RANTES mRNA. On the other hand, progesterone (Po) and tumor necrosis factor-alpha (TNFalpha) of different concentrations and different manners were used to treat cultured cells in vitro. RANTES secretion was evaluated in the culture medium using ELISA. In order to evaluate the effect of Po on the secretion of RANTES under stimulation of TNFalpha, the cells were cultured in medium containing 100 U/ml TNFalpha and Po of different concentrations for 24 hours. After the pretreatment of Po for 48 hours at different concentrations, TNFalpha (100 U/ml, 16 h) was added to observe whether Po inhibits RANTES or not.

RESULTSThe expression of RANTES mRNA in eutopic endometrium of patients with endometriosis was significantly higher than in control group (28.0 +/- 9.0 vs. 22.0 +/- 5.6, P < 0.05). Following the exposures to LNG-IUS (24.0 +/- 4.2 vs. 25.9 +/- 4.2, P > 0.05) or GnRHa (23.0 +/- 12.9 vs. 26.9 +/- 5.2, P > 0.05), the expression of RANTES mRNA had no change. MPA significantly increased the expression of RANTES mRNA (42.6 +/- 3.1 vs. 24.3 +/- 5.7, P < 0.05). Po itself had no significant effect on the secretion of RANTES. Stimulated by Po and TNFalpha at the same time, the secretion of RANTES significantly increased. After pretreatment with Po for 48 hours, the reaction of RANTES to the stimulating effect of TNFalpha was down-regulated.

CONCLUSIONThe eutopic endometrium of patients with endometriosis has high chemotactic activity. It may be feasible to prevent and treat endometriosis with progestins.

Cells, Cultured ; Chemokine CCL5 ; biosynthesis ; Endometriosis ; drug therapy ; metabolism ; Endometrium ; drug effects ; metabolism ; Female ; Gonadotropin-Releasing Hormone ; agonists ; Humans ; Intrauterine Devices, Medicated ; Levonorgestrel ; therapeutic use ; Medroxyprogesterone ; therapeutic use ; Progesterone ; pharmacology ; therapeutic use ; Progestins ; therapeutic use ; Transforming Growth Factor alpha ; pharmacology

AIM: To investigate the effect of DT-13 on gastric cancer cell migration, and to explore the possible mechanisms underlying the anti-metastasis activity of DT-13. METHODS: Growth inhibition of DT-13 was analyzed by the MTT assay. Cell migration was measured by the scratch-wound assay and transwell double chamber assay. To investigate the possible mechanisms underlying the anti-metastasis activity of DT-13, chemokine receptors that are involved in cancer metastasis (CCR2, CCR5, CCR7, CXCR4, and CXCR6) were detected by conventional PCR. The effect of DT-13 on CCR5 and CXCR4 expression was further evaluated by quantitative PCR and Western blot, respectively. The secretion of CCL5 (ligand of CCR5) and SDF-1 (ligand of CXCR4) were detected by enzyme-linked immunosorbent assay (ELISA). RESULTS: DT-13 inhibited BGC-823 and HGC-27 cell growth in a dose dependent manner, and the estimated IC50 value for 24 h treatment was 23.5 ± 5.1 μmol·L(-1) for BGC-823 cells and 35.6 ± 7.6 μmol·L(-1) for HGC-27 cells. DT-13 also significantly decreased gastric cancer cell migration. DT-13 significantly decreased the gene expression of CCR5 in both BGC-823 and HGC-27 gastric cancer cells, and moderately reduced the expression of CXCR4. Similar to the results of gene expression, significant down-regulation of CCR5 protein was observed, but CXCR4 protein levels were much less affected. CCL5 secretion, but not SDF-1 production, was inhibited by DT-13. CONCLUSION DT-13 inhibited gastric cancer cell migration by down-regulation of the CCR5-CCL5 axis.
Antineoplastic Agents, Phytogenic ; pharmacology ; Cell Movement ; drug effects ; Chemokine CCL5 ; analysis ; Down-Regulation ; Humans ; Neoplasm Metastasis ; drug therapy ; Receptors, CCR5 ; analysis ; Saponins ; pharmacology ; Stomach Neoplasms ; pathology ; Tumor Cells, Cultured

OBJECTIVETo investigate the effect of curcumin on the injury in hippocampal neurons and the expression of regulated upon activation nonnal T-cell expressed and secreted (RANTES) in hippocamp during cerebral ischemia/reperfusion (I/R) in rats with spontaneous hypertension (SH).

METHODSMale Wistar-Kyoto (WKY) rats and spontaneous hypertension rats (SHR) were randomly divided into five groups (n = 6): sham group (W-Sham and S-Sham group), ischemia/reperfusion group (W-/R and S/R group), curcumin group (S-Cur group) . Each group was splitted into 5 subgroups of 3 h,12 h, 1 d, 3 d and 7 d according to the time interval before reperfusion. Global brain ischemia/reperfusion model was established by 4-VO method. Hematoxylin-eosin staining (HE staining) was used to observe the vertebral cell morphology in hippocampal CA1 region. Nissl staining was applied to detect the average density of cone cells in hippocampal CA1 region. The expression of RANTES in hippocamp was determined by ELISA. The behavior of the rats was evaluated at 7 days after reperfusion. Results: Compared with the sham group rats, the ability of learning and memory was significantly decreased in ischemia/reperfusion group rats, the number of injured neurons were greatly elevated , the protein expression levels of RANTES was significantly increased (P < 0.05). Compared with W-I/R group rats, the ability of learning and memory in S-I/R group rats was greatly reduced, the number of injured neurons increased extremely, the protein expression level of RANTES was significantly enhanced( P <0.05). The number of injured neurons declined significantly in S-Cur group rats, the ability to learn and remember of these rats was improved and the RANTES protein content decreased significantly (P < 0.05).

CONCLUSIONSHR are more susceptible to ischemia/reperfusion induced hippocampal neuronal injury which may be improved by curcu min. Its underlying mechanism is possibly associated with the inhibition of RANTES protein expression level.

Animals ; Brain Ischemia ; metabolism ; pathology ; physiopathology ; Chemokine CCL5 ; metabolism ; Cognition ; drug effects ; Curcumin ; pharmacology ; Hippocampus ; cytology ; metabolism ; pathology ; Hypertension ; metabolism ; pathology ; physiopathology ; Male ; Neurons ; drug effects ; metabolism ; pathology ; Rats ; Rats, Inbred SHR ; Rats, Inbred WKY ; Reperfusion Injury ; metabolism

OBJECTIVETo investigate the effect of vitamin D on the expression of chemokine regulated on activation, normal T cells expressed and secreted (RANTES) in the lung tissue of asthmatic rats, and the role of vitamin D in the control of asthmatic airway inflammation and the synergistic action of hormones.

METHODSForty female Wistar rats were randomly and equally divided into normal control, asthma, vitamin D intervention, budesonide intervention, and budesonide+vitamin D intervention groups. Hematoxylin and eosin staining was used to observe pathological changes in the lung tissue. Immunohistochemistry was used to measure the protein expression of RANTES in lung tissue. Enzyme-linked immunosorbent assay was used to measure the level of RANTES in bronchoalveolar lavage fluid (BALF). Real-time quantitative PCR was used to measure the mRNA expression of RANTES.

RESULTSThe asthma group showed the most significant pathological changes in the lung tissue, including inflammatory cell infiltration, bronchial stenosis and distortion and smooth muscle rupture, while the intervention groups showed fewer pathological changes. Of the intervention groups, the budesonide intervention group showed fewer pathological changes than the vitamin D intervention group, and the budesonide+vitamin D intervention group showed the mildest pathological changes, which were similar to those observed in the normal control group. Protein expression of RANTES in the lung tissue and BALF was significantly higher in the asthma group than in the normal control group (P<0.05), while it was lower in the intervention groups than in the asthma group, exhibiting significant differences between each intervention group and the asthma group (P<0.05) (except the difference in protein expression of RANTES in BALF between the vitamin D intervention and asthma groups). The budesonide+vitamin D intervention group showed less protein expression of RANTES in the lung tissue and BALF than both the budesonide intervention and vitamin D intervention groups (P<0.05). The mRNA expression of RANTES was significantly higher in the asthma group than in the normal control group (P<0.05), while it was significantly lower in three intervention groups than in the asthma group (P<0.05), however no significant difference was found between the intervention groups in this regard. The budesonide+vitamin D intervention group showed the lowest level of RANTES mRNA, with no significant difference from the normal control group.

CONCLUSIONSThe mRNA and protein expression of RANTES in BALF and lung tissue increases significantly in asthmatic rats. Vitamin D intervention can decrease the expression of RANTES, suggesting that vitamin D can reduce airway inflammation by regulating the expression of RANTES. Vitamin D can be used together with budesonide to further decrease the mRNA and protein expression of RANTES.

Animals ; Asthma ; drug therapy ; metabolism ; Bronchoalveolar Lavage Fluid ; chemistry ; Budesonide ; therapeutic use ; Chemokine CCL5 ; analysis ; genetics ; Enzyme-Linked Immunosorbent Assay ; Female ; Immunohistochemistry ; Lung ; metabolism ; pathology ; Rats ; Rats, Wistar ; Vitamin D ; pharmacology

OBJECTIVETo explore the effect of Toll-like receptor 4 (TLR4) activation on the migration of asthmatic airway smooth muscle cell (ASMCs) induced by airway epithelial cells.

METHODSPrimary ASMCs were cultured by the method of cell digestion. Cell culture supernatant of RTE cells were collected by TNF-alpha stimulation of epithelial cells. Detected the IL-8 and RANTES levels in the supernatant. The transmembrane migration of asthmatic ASMCs were detected by Modified Boyden chemotaxis chamber. The effect of TLR4 on the migration of asthmatic ASMCs induced by epithelial cells with TLR4 antibody drugs as a tool.

RESULTSThe levels of IL-8 and RANTES in the supernatant of TNF-alpha groups were significantly increased, and that in the 20 ng/ml group was significantly higher than other groups (P < 0.01). The transmembrane migration of asthmatic ASMCs groups was increased than that of control group. The transmembrane migration of asthmatic ASMCs from asthma group and TNF-alpha + TLR4 antibody group was significantly decreased compared with that in TNF-alpha group (P < 0.01). The migration of asthma ASMCs from TNF-alpha + TLR4 antibody group was increased than that of asthma group (P < 0.05).

CONCLUSIONTLR4 in the surface of asthmatic ASMCs may be activated by cytokines secreted by the airway epithelial cells and enhance the transmembrane migration of asthmatic ASMCs induced by airway epithelial cells so that it plays a role in airway remodeling of asthma.

Animals ; Asthma ; metabolism ; Cell Movement ; Cells, Cultured ; Chemokine CCL5 ; metabolism ; Epithelial Cells ; metabolism ; Interleukin-8 ; metabolism ; Myocytes, Smooth Muscle ; cytology ; metabolism ; Rats ; Rats, Sprague-Dawley ; Toll-Like Receptor 4 ; metabolism ; Tumor Necrosis Factor-alpha ; pharmacology