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ObjectiveTo investigate the effect of Jingangwan on the expression of osteoclast, c-Jun N-terminal kinase(JNK), p38 mitogen-activated protein kinase(p38 MAPK), and interleukin-1(IL-1) in the osteoporosis model rats, explore the mechanism of Jingangwan in the treatment of osteoporosis, and determine the optimal dosing concentration of Jingangwan. MethodFifty-six rats of SPF grade were randomized into a blank group,a sham operation group,a model group, model group,high-, medium-, and low-dose Jingangwan groups (0.72, 0.36, 0.18 g·kg-1·d-1, ig),and an estradiol valerate group (0.009 g·kg-1·d-1, ig), with eight rats in each group. The rats in the model group, the blank group, and the sham operation group received 3 mL of normal saline, respectively. Samples were collected 12 weeks after drug administration. The number of osteoclasts was observed by tartrate-resistant acid phosphatase (TRAP) staining. Serum levels of JNK, p38 MAPK, and IL-1 were detected by enzyme-linked immunosorbent assay (ELISA). The mRNA expression levels of p38 MAPK and JNK were detected by real-time quantitative polymerase chain reaction (Real-time PCR). ResultThe TRAP staining results showed that compared with the model group, the estradiol valerate group and the Jingangwan groups could inhibit the formation of osteoclasts to different degrees. As revealed by ELISA results, compared with the model group and the sham operation group, the model group showed increased serum levels of p38 MAPK, JNK, and IL-1 (P<0.01), while compared with the model group, all the groups with drug intervention showed decreased levels of p38 MAPK, JNK, and IL-1 (P<0.01). The serum levels of JNK and IL-1 in the high-dose Jingangwan group were lower than those in the estradiol valerate group (P<0.05). Real-time PCR results showed that compared with the blank group, the model group showed increased relative mRNA expression of p38 MAPK and JNK in the thighbone (P<0.01), while compared with the model group, all the groups with drug intervention showed decreased relative mRNA expression of p38 MAPK and JNK in the thighbone (P<0.01). ConclusionJingangwan can inhibit the formation of osteoblasts,reduce the diameter of the bone marrow cavity,improve bone quality,suppress the production of inflammatory factors,affect the metabolism of the MAPK signaling pathway,and blunt p38 MAPK and JNK activities to inhibit the differentiation and proliferation of osteoblasts and regulate bone metabolism, thereby preventing osteoporosis. Therefore,Jingangwan may be of application value in maintaining bone health and treating osteoporosis.
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CXC chemokine ligand 8 (CXCL8) is highly expressed in many human tumors including colorectal cancer, and it can promote the malignant progression of tumors. It was reported that M2 macrophages were abundant in colorectal cancer microenvironment, but whether CXCL8 affects the infiltration of M2 macrophages and its potential mechanism are not yet clear. The study aimed to investigate the effect of CXCL8 on M2 macrophage infiltration and chemotaxis in the colorectal cancer. Firstly, we analyzed the CXCL8 expression and immune cell infiltration in human colorectal cancer tissues from TCGA RNA-seq data. The expression of CXCL8 was verified by immunohistochemistry in tissues obtained from Shanxi Provincial Cancer Hospital. Then, Western blot and qRT-PCR were employed to detect CXCL8 expression in five colorectal cancer cell lines. THP-1 cells were allowed to differentiate into M2 macrophages via the phorbol myristate acetate (PMA) and IL-4 treatment, followed by detection of the chemotaxis of M2 macrophages towards HCT116, SW480 and CXCL8-HCT116, CXCL8-SW480 cell lines. HCT116 and SW480 cells were treated with interleukin 1β (IL-1β) to detect the expression of CXCL8, and co-cultured with M2 macrophages to analyze the chemotactic activity. The results revealed that the expression of CXCL8 was increased in pairs of CRC tissues versus normal adjacent tissues, and there were more M2 macrophage infiltration in cancer tissues with high expression of CXCL8. The mRNA and protein expression of CXCL8 in HCT116 and SW480 were increased after the IL-1β treatment (P < 0. 05). We confirmed that CXCL8 is a chemotactic factor for M2 macrophages by transwell assays (P<0. 05). In conclusion, CXCL8 in colorectal cancer cells can be induced by IL-1β in colorectal cancer cells and the upregulation of CXCL8 can promote the chemotaxis of M2 macrophages. The massive infiltration of M2 macrophages in colorectal cancer microenvironment may be related with the increased expression of CXCL8.
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Objective To explore the inhibitory effect of curcumin on LPS-induced inflammation and the activation of Toll-like receptor 4 (TLR4 )/NADPH oxidase/reactive oxygen species (ROS)signaling pathway in vascular smooth muscle cells (VSMCs).Methods Primary VSMCs were cultured and divided into control group, LPS group,LPS + curcumin 5 μmol/L group,LPS + curcumin 10 μmol/L group and LPS + curcumin 30 μmol/L group.Cell activity was observed by MTT assay.The secretion of tumor necrosis factor-α(TNF-α)and interleukin-1 (IL-1)was measured by enzyme linked immunosorbent assay (ELISA)kits.The mRNA expressions of TLR4 and p22phox were detected by real-time PCR.Expression of intracellular ROS was measured by flow cytometry. Results The activities of VSMCs were not significantly affected by curcumin at the concentration between 0 and 80 μmol/L.Curcumin (5,10 and 30 μmol/L)significantly inhibited LPS-induced oversecretion of TNF-αand IL-1, as well as overexpression of TLR4 and p22phox at the mRNA and protein levels,and ROS production in VSMCs in a concentration-dependent manner.Conclusion Curcumin has a concentration-dependent inhibitory effect on the secretion of inflammatory cytokine,overexpressions of TLR4 and p22phox,and production of ROS in VSMCs stimulated by LPS.Furthermore,curcumin may partly depend on TLR4/NADPH oxidase/ROS signaling pathways to inhibit inflammation in LPS-induced VSMCs.
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Las citoquinas pertenecientes a familia de la interleuquina-1 (IL-1) están codificadas por tres genes diferentes: IL-1A, IL-1B, e IL-1RN, los cuales codifican para IL-1 α, IL-1β, y el antagonista endógeno del receptor de IL-1 (IL-1ra), respectivamente. Las IL-1α e IL-1β actúan como citoquinas pro-inflamatorias, mientras que la IL-1ra se comporta como anti-inflamatoria. Han sido reportados varios polimorfismos bialélicos en los genes de IL-1B, incluyendo IL-1B-511(C/T) e IL-1B+3954(C/T), mientras que IL-1RN presenta en el intrón 2 un polimorfismo VNTR penta-alélico. Los polimorfismos funcionalmente relevantes de estos genes han sido correlacionados con un amplio conjunto de condiciones autoinmunes e inflamatorias crónicas, así como con cáncer. Con el fin de determinar la distribución de estos polimorfismos en la región centroccidental de Venezuela, se estudiaron 100 individuos no relacionados aparentemente sanos. Se extrajo ADN genómico a partir de sangre periférica, y se procedió a la tipificación de los polimorfismos IL-1B-511 e IL-1B+3954 por PCR-RFLP y VNTR de IL-1RN por PCR. Se determinaron las frecuencias alélicas y genotípicas con el programa Arlequín ver. 2.000. Se observó un predominio del alelo T (52%) y del alelo C (82%) en IL-1B-511 y IL-1B+3954, respectivamente. Mientras que para IL-1RN los genotipos más frecuente fueron el 1/1 (47%) y 1/2 (41%). Se compararon los resultados con las frecuencias poblacionales encontradas en otros países, destacándose diferencias significativas con poblaciones de diferente origen étnico. Los resultados podrían proporcionar una referencia valiosa para estudios futuros de asociación con cáncer y enfermedades inflamatorias en Venezuela.
The cytokines belonging to the family of interleukin-1 (IL-1) are encoded by three different genes: IL-1A, IL-1B, and IL-1RN, which encode for IL-1α, IL-1β and the endogenous receptor antagonist for IL-1 (IL-1Ra), respectively. IL-1α and IL-1β operate as pro-inflammatory cytokines, while the IL-1Ra as anti-inflammatory. It has been reported several biallelic polymorphisms in the genes of IL-1B, including IL-1B-511(C/T) and IL-1B+3954(C/T), while IL-1RN presents in intron 2 a penta-allelic VNTR polymorphism. The functionally relevant polymorphisms of these genes have been correlated with a wide range of chronic inflammatory and autoimmune conditions, as well as cancer. In order to determine the distribution of these polymorphisms in the Central-Western region of Venezuela, 100 unrelated apparently healthy individuals were studied. DNA was extracted from peripheral blood, and proceded to the characterization of polymorphisms IL-1B-511 and IL-1B +3954 by PCR-RFLP and VNTR IL-1RN by PCR. Allelic and genotypic frequencies were determined awith the program Arlequin v. 2.0. There was a predominance of T allele (52%) and the C allele (82%) for IL-1B-511 and IL-1B +3954, respectively. While for IL-1RN the more frequent genotypes were 1/1 (47%) and 1/2 (41%). We compare the results with the population frequencies found in other countries, highlighting differences with significant populations of different ethnic origin. These results could provide a valuable reference for future studies of association with cancer and inflammatory diseases in Venezuela.
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Osteoclasts are derived from pluripotent stem cells in bone marrow and spleen.They play a critical role in inflammation-induced bone loss and joint destruction because in the absence of them,bone de- struction does not occur even when inflammation exists.Synovioblasts in an inflamed joint can secrete numerous inflammatory factors,including tumor necrosis factor alpha(TNF-?)and interleukin-1(IL-1)which not only induce inflammatory reactions but also elevate osteoclast formation and function indirectly or directly through promoting RANKL expression.In this wdy the inflammatory reactions are associated with bone loss and destruction. In this article,we focus on the recent progress in study of TNF-?,IL-1 and osteoclast-target therapies in management of osteoclast-mediated inflammatory bone loss.TNF-?promotes differentiation of osteoclast precursor cells in the peripheral blood and spleen,which causes a marked increase in mature osteoclasts in a diseased joint.However, IL-I supports osteoblast survival and regulates the recombination of osteoclast cytoskeleton,which further stimulates bone resorption.Since osteoclast-target therapies may inhibit osteoclast formation and function,they are becoming more and more important for inflammation-induced bone loss and joint destruction.
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OBJECTIVE: Recently, cultured myoblast transplantation has been extensively studied as a gene complementation approach in such genetic diseases as Duchenne muscular dystrophy (DMD). In the present work we investigated the stimulating effects of the growth factors, such as basic fibroblast growth factor (bFGF), leukemia inhibitory factor (LIF) and interleukin-1 (IL-1), on growth rate and differentiation of myoblast. METHOD: Human myoblasts were cultured from biopsy and treated in vitro with various concentration of bFGF, LIF and IL-1. In serum-free defined medium the following observation were made to evaluate differentiation. RESULTS: bFGF and LIF except IL-1 were found to have stimulating effect of myoblast proliferation comparing to control group (p<0.05), yet there were no statistically significant differences among each growth factors (p<0.05). The most significant growth stimulation of myoblasts in culture was achieved by adding 3.0 ng/ml of bFGF, producing a stimulation effect up to 2.01-fold. All myoblasts treated with growth factors differentiated into myotube. CONCLUSION: Our findings indicate that bFGF and LIF stimulate the proliferation of myoblast, which may result in an effective way in producing large numbers of myoblasts for clinical myoblast transplantation in DMD patients.
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Humanos , Biopsia , Proteínas del Sistema Complemento , Factor 2 de Crecimiento de Fibroblastos , Factores de Crecimiento de Fibroblastos , Fibroblastos , Genes vif , Péptidos y Proteínas de Señalización Intercelular , Interleucina-1 , Factor Inhibidor de Leucemia , Leucemia , Fibras Musculares Esqueléticas , Distrofia Muscular de Duchenne , MioblastosRESUMEN
Collagen is the major component of scar tissue. Considerable progress of fibroblast growth kinetics and of collagen synthesis has been achieved in the past decade. We have been interested in fibroblasts activities as they are expressed by cells cultured in collagen substrate. This study is to examine the effects of collagen substrate and peptide growth factors In culture medium on DNA and protein synthesis of human dermal fibroblasts. Collagen, interleukin-1(IL-1) and transforming growth factor-beta(TGF-beta) were added to fibroblast culture media according to the designed experiment model and DNA and protein synthesis were measured by [3H]-thymidine, [3H]-leucine, and [3H]-proline incorporation method. The morphological features of fibroblasts were observed by light microscope. The results were as follows ; 1) There were significant decreases of DNA and protein synthesis of cultured fibroblasts in the presence of collagen substrate compared with those in Control groups(p<0.01). 2) DNA and protein synthesis were decreased as dose dependant manner of collagen density in culture media. 3) Morphological features of fibroblasts became less stellate and flat, more spindle-like in the presence of collagen. 4) In responsiveness to IL-1, collagen non-treated groups responded to IL-1 but collagen treated groups were unresponsive to IL-1 (P<0.05). 5) Cells In collagen non-treated groups responded to TGF-beta as dose-related manner(P<0.01). Collagen treated groups desponded to TGF-beta but did not show TGF-beta dose-dependant relationship. In Conclusion, collagen substrate in the culture medium could lower the DNA and protein synthesis of fibroblasts. Cells in collagen substrate were unresponsive or less responsive to peptide growth factors than those in non-collagen substrate.
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Humanos , Cicatriz , Colágeno , Medios de Cultivo , ADN , Fibroblastos , Péptidos y Proteínas de Señalización Intercelular , Interleucina-1 , Cinética , Factor de Crecimiento Transformador betaRESUMEN
The abnormal proliferation of vascular smooth muscle cells after endothe-lial injury is postulated to be the main pathophysiological process in atherosclerosis (AS). The effects of propylene glycol mannate sulfate(PGMS) on the proliferation of bovine cerebral microvessel smooth muscle cslls (BCMSMCs) induced by 10% fetal calf serum (FCS) and interleukin l(IL-1) were investigated in culture. 5 - 8 stage subcultured BCMSMCs were incubated into 96-well dish- With either 10% FCS or IL-1 (50u/ml) to produce BCMSMCs proliferation , the inhibitory effects of PGMS on proliferation of BCMSMCs were investigated. The results shows that PGMS could inhibit the proliferation of quiescent BCMSMCs induced by 10% PCS. The growth of cells was inhibited, comparing with normal control 72hours after the serum addition as determined by crystal violet stainning and MTTmethod. The proliferation of quiescent BCMSMCs induced by IL-1 (50u/ml) was also inhibited by PGMS as determined by crystal violet stainning and MTT method. The results suggested that PGMS inhibit the proliferation of BCMSMCs induced by 10% FCS and IL-1 ,and the use of PGMS may probably play an important role in the treatment of cerebrovascular disease.
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Objective:To explore the mechanism of action of thymosin ? 4 (T? 4) in septic shock by observing the 72h survival rate of septic shock mice treated with thymosin ? 4 and exploring the changes of TNF-? and IL-1?.Methods:LPS(lipopolysaccharide) was intraperitoneally injected into clean Kunming mice to establish the animal model.①The T? 4,between normal and septic shock mice were determined.②The survival rate of septic shock mice was studied by intraperitoneal injection of T? 4.Animals were divided into four groups of 30 and each received LPS(25mg/kg)intraperitoneal injection.Group Ⅰ was LPS control group.GroupⅡ,Ⅲ and Ⅳ were individually injected with T? 4 at 0h,0h,2h;0h,2h,4h post LPS through caudal vein.③TNF-? and IL-1? were examined in septic shock mice by ELISA.Results:T? 4 was obviously decreased in the blood of septic shock mice.T? 4 could raise the 72h survival rate of septic shock mice and down-regulate inflammatory mediators.Conclusion:The optimal protective methods in the mouse appears to be 5mg/kg T? 4 given immediately following(0h) and at 2h and 4h successively post LPS.T? 4 down-regulates the inflammatory mediators,which may be the mechanism of T? 4 in curing the septic shock.