Transforming growth factor (TGF)-β signaling plays an important role in the pathogenesis of psoriasis. CD109, a novel TGF-β co-receptor, which inhibits TGF-β signaling by enhancing Smad7-dependent degradation of TGF-β type I receptor (TGF-β RI), is abnormally expressed in psoriasis. To date, the expression of Smad7 and the correlation between CD109 and Smad7 expression in psoriasis have not been fully elucidated. This study was designed to investigate the expression and the correlation of CD109 and TGF-β signaling associated proteins in psoriasis and their roles in the pathogenesis of psoriasis. Thirty-two psoriasis specimens were subjected to immunohistochemical staining for CD109, Smad7, TGF-β RI and Ki67. Ten normal skin (NS) specimens served as controls. The positive expression rate (% positive cells) of Smad7 and Ki67 in psoriasis was significantly higher than in NS (62.6%±19.9% vs. 17.2%±4.4%, and 50.7%±14.3% vs. 19.5%±3.2%, respectively, P<0.001), and the expression levels of CD109 and TGF-β RI were reduced significantly in psoriasis as compared with NS (8.1%±6.7% vs. 35.8%±6.7% and 27.3%±3.4% vs. 3.0%±3.4%, respectively, P<0.001). There were significantly negative correlations between CD109 and Smad7 (r=-0.831, P<0.01). These findings indicated that CD109 might play a certain role in the pathogenesis of psoriasis. Lower expression of CD109 and TGF-β RI was highly correlated with higher expression of Smad7 and Ki67, suggesting that CD109 may induce the pathogenesis of psoriasis through Smad7-mediated degradation of TGF-β RI, and lead to the termination of TGF-β signaling.
Adolescent ; Adult ; Antigens, CD ; genetics ; metabolism ; Case-Control Studies ; Down-Regulation ; Female ; GPI-Linked Proteins ; genetics ; metabolism ; Humans ; Male ; Middle Aged ; Neoplasm Proteins ; genetics ; metabolism ; Psoriasis ; metabolism ; pathology ; Signal Transduction ; Smad7 Protein ; genetics ; metabolism ; Transforming Growth Factor beta ; metabolism ; Up-Regulation

The deubiquitinating enzyme ubiquitin specific peptidase 15 (USP15) is regarded as a regulator of TGFβ signaling pathway. This process depends on Smad7, the inhibitory factor of the TGFβ signal, and type I TGFβ receptor (TβR-I), one of the receptors of TGFβ. The expression level of USP15 seems to play vital roles in the pathogenesis of many neoplasms, but so far there has been no report about USP15 in psoriasis. In this study, immunohistochemical staining of USP15, TβR-I and Smad7 was performed in 30 paraffin-embedded psoriasis specimens and 10 normal specimens to investigate the expression of USP15, TβR-I and Smad7 in psoriasis and to explore the relevance among them. And USP15 small interfering RNA (USP15 siRNA) was used to transfect Hacat cells to detect the mRNA expression of TβR-I and Smad7. Of 30 cases of psoriasis in active stage, 28, 24 and 26 cases were positive for USP15, TβR-I and Smad7 staining, respectively. The positive rates of USP15 and Smad7 were significantly higher in psoriasis specimens than in normal skin specimens (44.1%±26.0% vs. 6.1%±6.6%, 47.2%±27.1% vs. 6.6%±7.1%), and positive rate of TβR-I (20.3%±22.2%) in psoriasis was lower than that in normal skin specimens (46.7%±18.2%). There was a significant positive correlation between USP15 and Smad7 expression, and significant negative correlations between USP15 and TβR-expression, an I d between TβR- and Smad7 expression I in psoriasis. After transfection of USP15 siRNA in Hacat cells, the expression of TβR-mRNA was up I -regulated and that of Smad7 was down-regulated. It is concluded that USP15 may play a role in the pathogenesis of psoriasis through regulating the TβR-I/Smad7 pathway and there may be other cell signaling pathways interacting with USP15 to take part in the development of psoriasis.
Adult ; Cell Line ; Female ; Gene Expression ; Humans ; Immunohistochemistry ; Keratinocytes ; cytology ; metabolism ; Male ; Middle Aged ; Protein-Serine-Threonine Kinases ; biosynthesis ; genetics ; Psoriasis ; genetics ; metabolism ; RNA Interference ; Receptors, Transforming Growth Factor beta ; biosynthesis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Signal Transduction ; genetics ; Skin ; metabolism ; Smad7 Protein ; biosynthesis ; genetics ; Ubiquitin-Specific Proteases ; biosynthesis ; genetics ; Young Adult

MicroRNAs (miRNAs) have recently been recognized to have a role in human orthopedic disorders. The objective of our study was to explore the expression profile and biological function of miRNA-17-5p (miR-17-5p), which is well known to be related to cancer cell proliferation and invasion, in osteoblastic differentiation and in cell proliferation. The expression levels of miR-17-5p in the femoral head mesenchymal stem cells of 20 patients with non-traumatic osteonecrosis (ON) and 10 patients with osteoarthritis (OA) were examined by quantitative reverse transcription-PCR (qRT-PCR). Furthermore, the interaction between miR-17-5p and SMAD7 was observed. We found that in non-traumatic ON samples the level of mature miR-17-5p was significantly lower than that of OA samples (P=0.0002). By targeting SMAD7, miR-17-5p promoted nuclear translocation of beta-catenin, enhanced expression of COL1A1 and finally facilitated the proliferation and differentiation of HMSC-bm cells. We also demonstrated that restoring expression of SMAD7 in HMSC-bm cells partially reversed the function of miR-17-5p. Together, our data suggested a theory that dysfunction of a network containing miR-17-5p, SMAD7 and beta-catenin could contribute to ON pathogenesis. The present study prompts the potential clinical value of miR-17-5p in non-traumatic ON.
Adult ; Base Sequence ; Bone Morphogenetic Protein 2/metabolism ; Cell Differentiation ; Cell Line ; Cell Proliferation ; Female ; *Gene Expression Regulation ; Humans ; Male ; MicroRNAs/genetics/*metabolism ; Middle Aged ; Osteoarthritis/genetics/metabolism/pathology ; Osteoblasts/*cytology/metabolism/*pathology ; Osteogenesis ; Osteonecrosis/*genetics/metabolism/pathology ; Signal Transduction ; Smad7 Protein/*genetics/metabolism ; beta Catenin/metabolism

This study is to investigate the effect of artesunate on transforming growth factor-beta1 (TGF-beta1) induced epithelial-mesenchymal transition (EMT) and its possible mechanism. After the in vitro cultured RLE-6TN cells were treated with TGF-beta1 then artesunate intervened on it, after 24 h, expression of the markers of mesenchymal cell was assayed using Western blotting and real-time PCR analysis. Western blotting was also used to detect the effect of TGF-beta1 on the Smad3 and Smad7 expressions of RLE-6TN cells. Morphological alterations were examined by phase-contrast microscope, and ultrastructure changes by electron microscope. Incubation of RLE-6TN cells with TGF-beta1 resulted in the up-regulation of the expression of the mesenchymal cell markers, after artesunate intervened on it, resulted in the down-regulation of the expression. Meanwhile, incubation with artesunate intervened on RLE-6TN cells could lead to the apparent down-regulation of the expression of Smad3 and up-regulation of Samd7 and the transition of RLE-6TN cells to mesenchymal-like by TGF-beta1 induction, after artesunate intervened on it, RLE-6TN cells to epithelial-like. TGF-beta1 induced epithelial-mesenchymal transition process; artesunate can inhibit TGF-beta1-induced epithelial-mesenchymal transition process, the possible mechanism is up-regulation of the expression of Smad7 and down-regulation of the expression of Smad3, meanwhile inhibits phosphorylation of Smad3.
Actins ; genetics ; metabolism ; Animals ; Artemisia ; chemistry ; Artemisinins ; isolation & purification ; pharmacology ; Cell Line ; Cell Proliferation ; drug effects ; Epithelial Cells ; cytology ; metabolism ; Epithelial-Mesenchymal Transition ; drug effects ; Idiopathic Pulmonary Fibrosis ; pathology ; Plants, Medicinal ; chemistry ; Pulmonary Alveoli ; cytology ; RNA, Messenger ; metabolism ; Rats ; Smad3 Protein ; genetics ; metabolism ; Smad7 Protein ; genetics ; metabolism ; Transforming Growth Factor beta1 ; pharmacology ; Vimentin ; genetics ; metabolism

OBJECTIVETo discuss whether asiaticosides could effectively reduce the endothelial cell damage as a biochemical modulator, so as to further inhibit the post-stenting intima-media membrane hyperplasia.

METHODHuman aortic smooth muscle cells and aortic fibroblasts were selected and divided into the blank group, the rapamycin group and the asiaticoside group and the rapamycin and asiaticoside group. The expressions of muscle cells and fibroblasts TGF-beta1, Smad7 and I-collagen gene were determined by RT-PCR. The expression quantity of I-collagen protein was assayed by ELISA. The coefficient of drug interaction (CDI) between rapamycin and asiaticoside was calculated. Additionally, 16 Chinese mini-swines were randomly divided into group A and group B. One sirolimus drug-eluting stent of the same type was implanted after the high-pressure pre-expansion of anterior descending artery balloon. After the operation, the group A was intravenously injected with normal saline 30 mL x d(-1). Whereas the group B was intravenously injected with asiaticoside 30 mg x kg(-1) x d(-1)(diluted to 30 mL). The expressions of plasma vWF of the two groups were measured at the 7th and 14th days after the operation. At the 28th day after the operation, tissues of the stented vessel segments were sliced and stained to calculate the vessel area, inner stent area, lumen area and neointima area

RESULTCompared with the control group, the combination group showed significant up-regulation in smooth muscle cells and fibroblast Smad7 gene, down-regulation in TGF-beta, and obvious inhibition of I-collagen gene expression (P < 0.01). As for smooth muscle cells, there was no difference in the expression of I-collagen between the combination group and the rapamycin group, with CDI at 0. 83. As for fibroblasts, there was a significant difference in the expression of I-collagen between the combination group and the rapamycin group (P < 0.05), with CDI at 0.77. Plasma vWF of the group B was significantly lower than that of the group A (P < 0.05) at the 7th and 14th days after the operation. At the 28th day after the operation, no difference was observed in vessel area and stent area between the two groups. However, the lumen area in the group B was significantly larger than that of the group A(P < 0.05), and the neointima area of the group B was significantly smaller than that of the group A (P < 0.05).

CONCLUSIONAs an effective biochemical modulator for rapamycin, asiaticosides could inhibit TGF-beta expression, significantly decrease the synthesis and secretion of extracellular matrix, further inhibit the post-stenting intima-media membrane hyperplasia and reduce the endothelial cell damage by effectively up-regulate the expression of Smad7 protein.

Animals ; Collagen ; genetics ; metabolism ; Coronary Restenosis ; drug therapy ; prevention & control ; surgery ; Drugs, Chinese Herbal ; administration & dosage ; Humans ; Hyperplasia ; drug therapy ; genetics ; metabolism ; prevention & control ; Smad7 Protein ; genetics ; metabolism ; Stents ; adverse effects ; Swine ; Transforming Growth Factor beta1 ; genetics ; metabolism ; Triterpenes ; administration & dosage

OBJECTIVETo evaluate the effect of Notoginseng Radix on hepatic expression of transforming growth factor beta1 (TGF-beta1) and connective tissue growth factor (CTGF) in rats with alcoholic liver disease (ALD), in order to discuss its protective effect on alcoholic cirrhosis.

METHODFifty SD male rats were divided into the normal control group, the model group, the high-dose and low-dose Notoginseng Radix groups (3.0, 12.0 g x kg(-1)) and the magnesium isoglycyrrhizinate group (24 mg x kg(-1)), with 10 rats in each group. Apart from the control group, other groups were administered with ethanol-cornoil-pyrazole for 14 weeks to establish the alcoholic liver disease model. During the establishment of the model, the high-dose and low-dose Notoginseng Radix groups were administered with 12 g x kg(-1) x d(-1) Notoginseng Radix for 14 weeks, once everyday. Efforts were made to detect liver function, pathology with Masson staining, and the expressions of TGF-beta1, Smad3, Smad7 and CTGF mRNA.

RESULTCompared with the rats in model group, rats in Notoginseng Radix groups showed significant reduction in liver ALT, AST, collagen fiber deposition, and TGF-beta1, Smad3 and CTGF mRNA expressions in liver tissues, with the increase in the expression quantity of Smad7 mRNA. There were differences between the Notoginseng Radix groups. No significant difference was observed between the high-dose Notoginseng Radix group and the magnesium isoglycyrrhizinate group.

CONCLUSIONNotoginseng Radix can affect TGF-beta1/Smads signaling pathway and reduce the expression of CTGF.

Animals ; Connective Tissue Growth Factor ; genetics ; metabolism ; Drugs, Chinese Herbal ; administration & dosage ; Gene Expression ; drug effects ; Humans ; Liver Diseases, Alcoholic ; drug therapy ; genetics ; metabolism ; Male ; Panax notoginseng ; chemistry ; Rats ; Rats, Sprague-Dawley ; Smad3 Protein ; genetics ; metabolism ; Smad7 Protein ; genetics ; metabolism ; Transforming Growth Factor beta1 ; genetics ; metabolism

OBJECTIVETo study the mechanism of Huogu I formula (I) in treating osteonecrosis of femoral head.

METHODSForty-eight healthy female Leghorn chickens were randomly divided into control group, model group and Huogu I group, and each group consisted of 16 chickens. At the meantime of model establishment, chickens of the Huogu I group were administrated with decoction, while the model and control group with distilled water by gavage. At the 8th and 16th week after medication, blood samples were obtained for blood lipid detection while both sides of femoral head were harvested for the rest of examinations. Specifically, expressions of bone morphogenetic protein-2 (BMP2), transforming growth factor beta1 (TGFβ(1)), Smad4 and Smad7 were evaluated by immunohistochemistry, while expression of osteoprotegerin/receptor activator of nuclear factor kappaB ligand (OPG/RANKL) mRNA was detected by in situ hybridization.

RESULTSCompared with the control group, serum levels of total cholesterol (TC), triglyceride (TG) and low-density lipoprotein cholesterol (LDL-C) in the model group rose significantly. Positive cell counting of BMP2, TGFβ(1), Smad4 and OPG in femoral head of the model group dropped prominently. Positive cell counting of Smad7 and RANKL increased dramatically. In contrast with the model group, levels of TC, TG and LDL-C in Huogu I group reduced significantly. Positive cell counting of BMP2, TGFβ(1), Smad4 and OPG in femoral head of the Huogu I group increased prominently. Indices of Smad7 and RANKL both decreased significantly. Especially at the 8th week, these variations were more significant.

CONCLUSIONHuogu I formula is effective in promoting repair of necrotic femoral head by regulating the expressions of BMP2, TGFβ(1), Smads and OPG/RANKL of osteoclast in femoral head.

Animals ; Bone Morphogenetic Protein 2 ; metabolism ; Bone Regeneration ; drug effects ; physiology ; Chickens ; Chondrocytes ; metabolism ; Disease Models, Animal ; Drugs, Chinese Herbal ; pharmacology ; Female ; Femur Head Necrosis ; chemically induced ; drug therapy ; metabolism ; Lipid Metabolism ; physiology ; Osteocytes ; metabolism ; Osteoprotegerin ; genetics ; metabolism ; Receptor Activator of Nuclear Factor-kappa B ; genetics ; metabolism ; Smad4 Protein ; metabolism ; Smad7 Protein ; metabolism ; Steroids ; pharmacology ; Transforming Growth Factor beta1 ; metabolism

Idiopathic pulmonary fibrosis (IPF) is a lethal parenchymal lung disease characterized by myofibroblast proliferation. Alveolar epithelial cells (AECs) are thought to produce myofibroblasts through the epithelial to mesenchymal transition (EMT). Receptor for advanced glycation end products (RAGE) is a member of the immunoglobulin superfamily of cell surface receptors whose activation is associated with renal fibrosis during diabetes and liver fibrosis. RAGE is expressed at low basal levels in most adult tissues except the lung. In this study, we evaluated the interaction of ligand advanced glycation end products (AGE) with RAGE during the epithelial to myofibroblast transition in rat AECs. Our results indicate that AGE inhibited the TGF-beta-dependent alveolar EMT by increasing Smad7 expression, and that the effect was abolished by RAGE siRNA treatment. Thus, the induction of Smad7 by the AGE-RAGE interaction limits the development of pulmonary fibrosis by inhibiting TGF-beta-dependent signaling in AECs.
Animals ; Epithelial Cells/cytology ; Epithelial-Mesenchymal Transition/*drug effects ; Glycosylation End Products, Advanced/genetics/*metabolism ; Idiopathic Pulmonary Fibrosis/metabolism ; Pulmonary Alveoli/cytology ; RNA, Small Interfering/genetics ; Rats ; Receptors, Immunologic/genetics/*metabolism ; Smad7 Protein/genetics/*metabolism ; Transforming Growth Factor beta/genetics/metabolism

BACKGROUNDTransforming growth factor β (TGFβ) is one of the most important growth factors in the development of fibrosis and scarring on cornea. Smad7, an inhibitory Smad, can inhibit TGFβ signal transduction. In recent years, effects of lentiviral-mediated Smad7 on inhibition of fibrosis on some organs have been studied, while little is known about the effects on cornea. This study aimed to determine the effects of lentiviral-mediated Smad7 gene expression on keratocyte proliferation and fibrosis induced by TGF β2 in vitro.

METHODSKeratocytes were cultured from corneal tissue isolated from Sprague-Dawley (SD) rats and transfected with Smad7 expressing lentiviral vector (Lv-Smad7) or non-functioning control vector (Lv-blank). Following the exposure to TGFβ2, keratocytes were processed for immunoblotting to assess the phosphorylation of Smad2 as down-stream event of TGFβ/Smad signaling. Expression of fibrotic markers α-smooth muscle actin (α-SMA), type III collagen (collagen III) were measured by Western blotting and quantitative real time polymerase chain reaction (PCR). Overall cell proliferation was determined by 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and the expression of cell cycle-related marker Ki67 at both mRNA and protein levels.

RESULTSThe Smad7 gene transfer suppressed TGFβ/Smad signaling in keratocytes by down-regulating phosphorylation of Smad2. Markers of cell proliferation and fibrosis including Ki67, α-SMA, collagen III were inhibited by introduction of Smad 7 into TGFβ exposed keratocytes. Consequently, the rate of cell proliferation was attenuated.

CONCLUSIONSmad7 gene transfer inhibited fibrogenic responses of keratocytes to TGFβ2.

Actins ; genetics ; metabolism ; Animals ; Blotting, Western ; Cell Proliferation ; Cells, Cultured ; Collagen Type III ; genetics ; metabolism ; Corneal Keratocytes ; cytology ; drug effects ; metabolism ; Genetic Vectors ; genetics ; Ki-67 Antigen ; genetics ; metabolism ; Lentivirus ; genetics ; Rats ; Rats, Sprague-Dawley ; Real-Time Polymerase Chain Reaction ; Reverse Transcriptase Polymerase Chain Reaction ; Signal Transduction ; drug effects ; genetics ; Smad7 Protein ; genetics ; metabolism ; pharmacology ; Transforming Growth Factor beta2 ; pharmacology

OBJECTIVETo explore the mechanism of action of Longbixiao Capsule (LBXC, a Chinese herbal preparation) on human prostatic stromal cell cultured in vitro.

METHODSNine Japan rabbits were assigned to 3 groups. The high, low dose group was given LBXC [2.0 g/(kg x d), 1.0 g/(kg x d)] by gastro gavage respectively, while equal volume of normal saline was given by gastrog avage 60 rats in the control group, all twice a day with an infeval of 12 h, for 3 successive days. The serum collected at 3 h after the last gastro gavage was added into cell culture fluid. Rabbit's serum containing LBXC was incubated with the cultured stromal cells, and the levels of cell proliferation and apoptosis were determined using relative techniques as TUNEL, ELISA, and immunocytochemistry. Besides, Real-time RT-PCR was applied to detect the mRNA expressions of TGF-beta1, and Smad7 in the stromal cells.

RESULTSThe cell proliferation showed culture time dependence in all groups. The proliferation in the drug-serum treated groups was lower than that in the control group, and it was lower in the high dose treated group than in the low dose treated group (all P < 0.01). The unfavorable growth did not occur morphologically after being cultured for 48 h and showed insignificant difference between various groups. Cell apoptosis was not found excepting for a few appeared in the high dose treated group (with a little amount of apoptotic cells occurring). After treatment, the expressions of TGF-beta, and Smad7 were lower in the low dose treated group and high dose treated group than in the control group (P < 0.01). There was not statistical difference between the low dose treated group and high dose treated group (P > 0.05).

CONCLUSIONLBXC could reduce the expressions of TGF-beta1, and smad7 mRNA in stromal cells and inhibit the stromal cell proliferation, but its effect on promoting cell apoptosis is unobvious.

Animals ; Apoptosis ; drug effects ; Capsules ; Cell Proliferation ; drug effects ; Cells, Cultured ; Drugs, Chinese Herbal ; pharmacology ; Humans ; Male ; Prostate ; cytology ; RNA, Messenger ; genetics ; metabolism ; Rabbits ; Random Allocation ; Serum ; Smad7 Protein ; genetics ; metabolism ; Stromal Cells ; cytology ; Transforming Growth Factor beta1 ; genetics ; metabolism