OBJECTIVETo investigate the effects of spironolactone on the concentration of collagen type I, III in the myocardium of spontaneous hypertension rats (SHR).

METHODSTwenty 8-week male SHR were assigned randomly into spironolactone (SHR-SPIRO, n=10) and control groups (SHR-CON, n=10), sex-age matched Wistar Kyoto rats (WKY group, n=7) were also served as controls. The rats of SHR-SPIRO group were given 20 mg/(kg*d) of spironolactone, the rats of SHR-CON and WKY groups were given the same volume of distilled water. After 16 weeks, the concentration of collagen type I was analyzed with Western blot. The areas of collagen type I and III were observed under polarized light microscopy and the ratio of type I/III collagen was calculated through accumulation score.

RESULTSCompared with WKY group,the concentration of collagen type I in SHR-CON group was significantly higher (1.87 ±0.2 Compared with 1.21 ±0.7, P<0.05). After 16 weeks of treatment the concentration of collagen type I (1.42 ±0.05 Compared with 1.87 ±0.2, P<0.05) and I/III ratio in SHR-SPIRO group were significantly reduced (15.64 ±1.34 Compared with 20.8 ±3.04, P<0.05) compared with SHR-CON group; but there were no differences in accumulation area scores of collagen type III among three groups (368.3 ±30.2 Compared with 481.6 ±32.4 Compared with 406.2 ±45.3, P>0.05).

CONCLUSIONThe deposition of collagen type I in myocardium may be involved in myocardial fibrosis of SHR, and spironolactone can decrease the concentration of collagen type I, which may be one of the mechanisms for its therapeutic effects.

Animals ; Collagen Type I ; metabolism ; Collagen Type III ; metabolism ; Male ; Mineralocorticoid Receptor Antagonists ; pharmacology ; Myocardium ; metabolism ; Rats ; Rats, Inbred SHR ; Rats, Inbred WKY ; Spironolactone ; pharmacology

Although many different attempts have been made to reduce adhesions after tendon surgery, this complicated problem continues to be foremost among factors compromising tendon surgery and repair. The results of the finest tendon surgery and repair are frequently compromised by adhesions that restrict motion, decrease function and often lead to permanent deformation. Many substances and materials have been introduced into the area around primary flexor tendon repairs in an effort to prevent or diminish the adhesion formation. Various biochemical agents including antihistamines, anaboilic agents, lathyrogenic drug, betaamino-propionitrile and steroids, have been investigated and material such as nylon, cellophane, polyethylene film and silastic have been used to form pseudosheath. However the results with all of these method have been disappointing. Recently, interest has grown in a mucopolysaccharide found in synovial fluid hyaluronic acid. Synovial fluid normally contains a concentration of 2-3 mg/ml of hyaluronic acid. Preliminary investigations suggest that hyaluronic acid placed between the tendon and its sheath promote tendon healing and decrease adhesion formation. With this in mind, the present study was designed to examine the effect of hyaluronic acid on injured chicken flexor tendon healing and to determine the quantity and quality for adhesions in hyaluronic acid treated chicken compared to control. The stereomicroangiographic technique has enabled us to study the vascular process inside the tendon and surrounding tissues after injury of the tendon. For the study, the deep flexor tendon of the third toe of adult chicken with initial weight of about 2kg was used. Animals were divided into three groups and treated with different levels of hyaluronic acid. After 1,3,7,21,35 and 42 days postoperation, the animals were sacrificed and specimen were prepared. Obtained specimen were examined macroscopically and microscopically. At 1,3,5, and 7 weeks postoperation, microangiographic study were done and examind specimen by stereomicroscope. The results were as follows: 1. Though both the treated and control groups showed adhesion, hyaluronic acid treated tendon showed less adhesion tissue and better sliding properties as compared to the controls. 2. During the first week after operation, microangiographic studies showed failure of filling of all the vessels within the injured tendon. At 3 weeks highly vascular adhesions which extended throughout the injured site were observed. After then, the amount of vascularity decreased. 3. There were no difference microscopically between hyaluronic acid treated and control group at first week. After one week hyaluronic acid treated group showed less granulation tissue and less formation of collagen fibers. 4. There were no difference of healing process between hyaluronic acid treated and control group. From this results, it is suggested that hyaluronic acid is effective in reducing the adhesions after the tendon surgery.
Adult ; Animals ; Cellophane ; Chickens ; Collagen ; Granulation Tissue ; Histamine Antagonists ; Humans ; Hyaluronic Acid ; Methods ; Nylons ; Polyethylene ; Steroids ; Synovial Fluid ; Tendons ; Toes

BACKGROUNDRepeated attacks of bronchial asthma lead to different degrees of airway remodeling, the mechanism of which is not yet clear. Some evidences indicate that it is related to the excessive expression of some growth promotion factors. Angiotensin II is a polypeptide that may be involved in airway remodeling. To evaluate its role in airway remodeling in asthma, we observed the effects of an angiotensin II type 1 receptor antagonist (valsartan) on the expression of collagen III, collagen V, and transforming growth factor beta1 (TGF-beta1) mRNA and protein in the airway walls of sensitized rats.

METHODSForty Wistar rats were randomly divided into 5 groups: control group, sensitized group, and valsartan groups 1, 2, and 3. The rats in the sensitized group and in valsartan groups 1, 2, and 3 were sensitized and challenged with ovalbumin. Rats in control group were sensitized and challenged with 0.9% NaCl. Rats from valsartan groups 1, 2, and 3 were drenched with valsartan (10 microg, 20 microg, or 30 microg, respectively) at the time of the ovalbumin challenges. The expression of collagen III, collagen V, and TGF-beta1 protein were detected using immunohistochemical method in combination with image analysis methods. The expression of TGF-beta1 mRNA was detected by in situ hybridization.

RESULTSThe expression in the airways of collagen III and collagen V was significantly higher in rats from the sensitized group (7.73 +/- 0.81, 1.34 +/- 0.28) and from valsartan groups 1, 2, and 3 (5.73 +/- 0.64, 1.13 +/- 0.15; 4.96 +/- 0.51, 0.98 +/- 0.08; 4.43 +/- 0.35, 0.93 +/- 0.06, respectively) than those in the control group (2.65 +/- 0.38, 0.67 +/- 0.08, P < 0.05). In addition, collagen levels were significantly lower in valsartan groups 1, 2, and 3 than those from the sensitized group (P < 0.05). The expression of TGF-beta1 mRNA and protein in the airways was significantly higher in rats from the sensitized group (20.49% +/- 3.46%, 29.73% +/- 3.25%) and from valsartan groups 1, 2, and 3 (16.47% +/- 1.94%, 19.41% +/- 1.87%; 14.38% +/- 1.58%, 18.29% +/- 1.43%; 12.96% +/- 1.73%, 18.63% +/- 1.11%, respectively) than that from the control group (7.84% +/- 1.61%, 5.63% +/- 1.07%, P < 0.05). TGF-beta1 mRNA and protein levels were significantly lower in valsartan groups 1, 2, and 3 than that in the sensitized group (P < 0.05).

CONCLUSIONSAngiotensin II receptor antagonist valsartan can suppress synthesis of collagen III and collagen V by downregulating TGF-beta1 mRNA and protein expression. Valsartan can decrease airway remodeling and could play a role in asthma therapy.

Angiotensin Receptor Antagonists ; Animals ; Asthma ; physiopathology ; Bronchi ; metabolism ; Collagen Type III ; analysis ; Collagen Type V ; analysis ; Immunization ; Male ; Ovalbumin ; RNA, Messenger ; analysis ; Random Allocation ; Rats ; Rats, Wistar ; Tetrazoles ; pharmacology ; Transforming Growth Factor beta ; analysis ; Valine ; analogs & derivatives ; pharmacology ; Valsartan

OBJECTIVETo observe the effect of compound Puerarin on collagen IV of streptozotocin-induced diabetic rats.

METHODSDiabetic nephropathy rats were induced by intraperitoneal injection of streptozotocin (STZ). Rats were allocated randomly to control group (10), diabetes model group (10), Vitamin C group (10), Puerarin group (10), vitamin C plus Puerarin group (10). The study period lasted for 12 weeks. During and after the treatment, the general state, blood glucose levels, glycosylated hemoglobin, blood urea nitrogen, serum collagen IV, blood urea nitrogen, serum creatinine, urinary albumin excretion rate of the 24-hour, and clearance rate of creatinine collagen IV protein were determined by immunohistochemistoche analysis as well as type the gene expression of collagen IV alpha 1 mRNA were determined by in situ hybridization analysis in the kidney tissue of different groups.

RESULTS(1) Diabetes mellitus and renal function lesion occurred in the four groups. (2) Vitamin C and Puerarin could improve the general conditions of diabetic Rats, decrease blood urea nitrogen [(8.68 +/- 0.43), (7.98 +/- 0.47) and (5.76 +/- 0.82) micromol/L, serum creatinine [(74.68 +/- 8.20), (75.52 +/- 7.98) and (58.66 +/- 6.65) mmol/L], and urinary albumin excretion rate of the 24-hour [(18.40 +/- 0.37), (17.24 +/- 0.30) and (9.97 +/- 1.27) mg/24 h x 10(-3)]; increase clearance rate of creatinine [(0.59 +/- 0.21), (0.61 +/- 0.14) and (0.69 +/- 0.32) ml/min], the expression of collage IV absorbance [(111.56 +/- 14.61), (110.78 +/- 9.69) and (95.44 +/- 9.97) ] in the diabetic Rats were significantly inhibited at the same time.

CONCLUSIONThe compound Puerarin might have some functions on preventing ren by inhibiting expression of type IV collagen.

Animals ; Collagen Type IV ; antagonists & inhibitors ; biosynthesis ; Diabetes Mellitus, Experimental ; drug therapy ; metabolism ; Diabetic Nephropathies ; drug therapy ; metabolism ; Isoflavones ; pharmacology ; therapeutic use ; Male ; Phytotherapy ; Rats ; Rats, Sprague-Dawley

Whether tranilast had antagonistic effect on proliferation inhibition and collagen synthesis promotion induced by TGF-beta2 in cultured human trabecular meshwork cells was investigated. Suspension of 1 x 10(4) cultured human trabecular meshwork cells of 3-5 passage was distributed in each well of a 96-well disk and divided into control group and experimental group. After 24 h, 0 microg/ml (control), 12.5 microg/ml, 25 microg/ml, 50 microg/ml tranilast with 3.2 ng/ml TGF-beta2 were added into the incubation medium. Another 24 h later, proliferation and collagen synthesis in cultured human trabecular meshwork cells were examined respectively by using tetrazolium-based semiautomated colormetric (MTT) assay and 3H-proline incorporation with liquid scintillation technique. The results showed absorbance (A) values of the experimental groups were 0.9036 +/- 0.3017, 1.1361 +/-0.1352, 1.2457 +/- 0.1524 according to the different concentrations of tranilast, and 0.8956 +/-0.1903 of the control group. In comparison with the control group, 25 microg/ml (q'= 3.23, P< 0.05), 50 microg/ml (q'=4.70, P<0.01) tranilast significantly antagonized the decrease of the A values induced by TGF-beta2 in the cultured human trabecular meshwork cells. In comparison with the control group [817.37+/-124.21 cpm/10(4) cells], 12.5 microg/ml (620.33+/-80.46 cpm/10(4) cells, q'= 4.26, P<0.05), 25 microg/ml (594.58+/-88.13 cpm/10(4) cells, q'=4.81, P<0.01), 50 microg/ml (418.64+/-67.90 cpm/10(4) cells, q'=8.62, P<0.01) tranilast significantly inhibited the incorporation of 3H-proline into the cultured human trabecular meshwork cells promoted by TGF-beta2 in a dose-dependent manner. It was concluded that tranilast had the antagonistic effect on the proliferation inhibition and collagen synthesis promotion induced by TGF-alpha2 in the cultured human trabecular meshwork cells.
Cell Proliferation ; drug effects ; Cells, Cultured ; Collagen ; biosynthesis ; Humans ; Trabecular Meshwork ; cytology ; metabolism ; Transforming Growth Factor beta ; antagonists & inhibitors ; Transforming Growth Factor beta2 ; ortho-Aminobenzoates ; pharmacology

Acetaldehyde ; pharmacology ; Animals ; Anthracenes ; pharmacology ; Apoptosis ; drug effects ; Cell Line ; Cell Proliferation ; drug effects ; Collagen ; biosynthesis ; Collagen Type I ; biosynthesis ; Collagen Type III ; biosynthesis ; Flow Cytometry ; Hepatic Stellate Cells ; drug effects ; metabolism ; JNK Mitogen-Activated Protein Kinases ; antagonists & inhibitors ; Liver Cirrhosis, Alcoholic ; pathology ; prevention & control ; Rats ; Signal Transduction ; drug effects

OBJECTIVETo study the therapeutic effects to block the TGF-beta1 (transforming growth factor beta1) signal transduction by antisense Smad4 gene on experimental fibrotic liver.

METHODSUsing the rat model of liver fibrosis induced by Carbon Tetrachloride (CCl4)/ethanol, we transfected antisense Smad4 gene mediated by adenovirus via portal vein infusion into the liver, and observed the expression of Smad4 by Retro-Polymerase Chain Reaction (RT-PCR) and Western Blot. We also investigated the pathologic features and collagen expression.

RESULTSIn the non-therapeutic cirrhotic liver, the expression of Smad4 mRNA was significantly increased than normal liver, and so was the collagen I. After antisense Smad4 gene being transfected, the expression of Smad4 mRNA and that of collagen I in the therapeutic liver was significantly decreased, compared with the non-therapeutic cirrhotic liver. The fibrous degree of therapeutic liver was also reduced compared with the non-therapeutic fibrous liver.

CONCLUSIONThese results indicate that because antisense Smad4 gene could block TGF-beta1 signal transduction by reducing the expression of Smad4, so it could inhibit the production of extracellular matrix (ECM) and improve hepatic fibrosis.

Adenoviridae ; genetics ; Animals ; Antisense Elements (Genetics) ; therapeutic use ; Collagen Type I ; analysis ; DNA-Binding Proteins ; antagonists & inhibitors ; genetics ; Liver ; pathology ; Liver Cirrhosis, Experimental ; metabolism ; pathology ; therapy ; Male ; Rats ; Rats, Wistar ; Signal Transduction ; Smad4 Protein ; Trans-Activators ; antagonists & inhibitors ; genetics ; Transforming Growth Factor beta ; antagonists & inhibitors ; Transforming Growth Factor beta1

Rapamycin, a macrocyclic lactone, is effective in reducing the incidence of acute rejection after renal transplantation. The inhibitory effects of rapamycin on lymphocyte proliferation and the molecular mechanisms that were involved have been described. However, its effects on glomerular mesangial cells have not been clearly understood, and here, we examined the effect of rapamycin on platelet-derived growth factor (PDGF) - induced extracellular matrix synthesis as well as cell proliferation in mesangial cells. Rat mesangial cells were isolated from the glomeruli of Sprague-Dawley rats and cultured with Dulbecco's modified Eagles medium containing 20% fetal bovine serum. Different concentrations of rapamycin were administered 1 hour before the addition of 10 ng/ml of PDGF into growth arrested and synchronized cells. Cell proliferation was assessed by [3H]thymidine incorporation, total collagen synthesis by [3H]proline incorporation, and fibronectin secretion into the medium by Western blot analysis. In the mesangial cells, PDGF increased cell proliferation by 4.6-fold, total collagen synthesis by 1.8-fold, and fibronectin secretion by 3.2-fold. Rapamycin above 10 nM significantly inhibited PDGF-induced proliferation and collagen synthesis, but the treatment of rapamycin up to 1micrometer did not show any significant effects on PDGF-induced fibronectin secretion. These inhibitory effects of rapamycin on PDGF-induced mesangial cell proliferation and collagen synthesis reflect the potential value of rapamycin in the prevention and treatment of glomerulosclerosis in patients with chronic allograft nephropathy.
Animals ; Cells, Cultured ; Collagen/*antagonists & inhibitors/biosynthesis ; Fibronectins/*biosynthesis ; Glomerular Mesangium/cytology/drug effects/*metabolism ; Immunosuppressive Agents/*pharmacology ; Male ; Platelet-Derived Growth Factor/*pharmacology ; Rats ; Rats, Sprague-Dawley ; Research Support, Non-U.S. Gov't ; Sirolimus/*pharmacology

OBJECTIVETo study the role of connective tissue growth factor (CTGF) in the pathogenesis of human keloid.

METHODSHuman keloid fibroblasts (HKF) were isolated from human keloid and cultured in vitro. The cells were then divided into 3 groups according to different processing, i.e. ASODN treatment (AT), in which phosphorothioate CTGF antisense oligonucleotides (ASODN) labeled by fluorescent isothiocyananate were transfected into the HKFs by liposome; liposome control (LC, with liposome only); control groups (without liposome or ASODN). The distribution of CTGF ASODN in all groups of cells was observed under fluorescent microscope. The CTGF mRNA index (RI) of HKF was assessed by reverse transcription polymerase chain reaction method (RT-PCR). The collagen synthesis of HKF was assessed by (3)H-proline incorporation method.

RESULTSA large amount of fluorescence could be observed in the cytoplasm of HKFs in AT 12 hours after transfection, but not in LC and C groups. The CTGF mRNA index of HKF in AT group 48 hours after transfection was significantly lower than that in LC and C groups (0.12 +/- 0.62 vs 0.51 +/- 0.18 vs 0.54 +/- 0.35, P < 0.01). The (3)H-proline incorporation rate in AT group (108.96 +/- 79.05) was lower than that in LC and C groups (P < 0.01).

CONCLUSIONThe expression of CTGF gene and collagen synthesis of the cultured HKF could be inhibited by CTGF ASODN, implying that CTGF played a role in the development of excessive fibrosis of human keloid.

Collagen ; biosynthesis ; Connective Tissue Growth Factor ; Fibroblasts ; metabolism ; Humans ; Immediate-Early Proteins ; antagonists & inhibitors ; genetics ; physiology ; Intercellular Signaling Peptides and Proteins ; genetics ; physiology ; Keloid ; etiology ; metabolism ; Oligonucleotides, Antisense ; pharmacology ; RNA, Messenger ; analysis ; Transfection

BACKGROUND: Aldosterone antagonists are reported to have beneficial effects on diabetic nephropathy by effective blocking of the renin-angiotensin-aldosterone system. We investigated the renoprotective effect of the selective aldosterone receptor blocker eplerenone, the angiotensin converting enzyme inhibitor lisinopril, and combined eplerenone and lisinopril treatment in type 2 diabetic rats. METHODS: Animals were divided into six groups as follows: Otsuka Long-Evans Tokushima Fatty (OLETF) rat control, OLETF rats treated with a low dose of eplerenone (50 mg/kg/day), OLETF rats treated with a high dose of eplerenone (200 mg/kg/day), OLETF rats treated with lisinopril (10 mg/kg/day), OLETF rats treated with a combination of both drugs (eplerenone 200 mg/kg/day and lisinopril 10 mg/kg/day), and obese non-diabetic Long-Evans Tokushima Otsuka rats for 26 weeks. RESULTS: Urinary albumin excretion was significantly lower in the lisinopril group, but not in the eplerenone group. Urinary albumin excretion was decreased in the combination group than in the lisinopril group. Glomerulosclerosis and renal expression of type I and type IV collagen, plasminogen activator inhibitor-1, transforming growth factor-beta1, connective tissue growth factor, and fibronectin mRNA were markedly decreased in the lisinopril, eplerenone, and combination groups. CONCLUSION: Eplerenone and lisinopril combination showed additional benefits on type 2 diabetic nephropathy compared to monotherapy of each drug.
Aldosterone ; Animals ; Collagen Type IV ; Connective Tissue Growth Factor ; Diabetic Nephropathies ; Fibronectins ; Lisinopril ; Mineralocorticoid Receptor Antagonists ; Peptidyl-Dipeptidase A ; Plasminogen Activators ; Rats ; Rats, Inbred OLETF ; Receptors, Mineralocorticoid ; Renin-Angiotensin System ; RNA, Messenger ; Spironolactone