AIM AND METHODSTo determine the relationship between the nuclear envelope nucleoside triphosphatase (EC 3. 6. 1. 15, NTPase) activity and the phenotypic modulation of vascular smooth muscle cell (VSMC), the NTPase activity was detected during restenosis after de-endothelialization in vascular wall. The activities of three enzymes involved in carbohydrate and nucleic acid metabolism were also investigated by spectrophotometry.

RESULTSThe activity of NTPase increased continuously and associated with the process of intimal thickening. Western blotting showed that expression of SMalpha-actin, as the marker of contractile phenotype of VSMC, decreased continuously. Osteopontin (OPN), the marker of synthetic phenotype of VSMC, was up-regulated during the process. These suggested that intimal injury induced phenotypic modulation of VSMC. The activities of 5'-nucleotidase, adenosine deaminase and succinate dehydrogenase increased and reached their peaks on 7 days after de-endothelialization. The changes of three enzymes were associated with proliferation in VSMC.

CONCLUSIONThe efflux of mRNA and the changes of enzyme activity involved in carbohydrate or nucleic acid metabolism may be the biochemical basis in the development and progression of restenosis.

Animals ; Constriction, Pathologic ; Endothelium, Vascular ; pathology ; Female ; Hyperplasia ; enzymology ; pathology ; Male ; Muscle, Smooth, Vascular ; pathology ; Nucleoside-Triphosphatase ; metabolism ; Rats ; Rats, Sprague-Dawley ; Tunica Intima ; enzymology ; pathology

OBJECTIVETo investigate whether extracellular signal-regulated kinase (ERK1/2) was involved in changes of vascular smooth muscle cell (VSMC) under hypertension.

METHODSTwo-kidney one clip Wistar hypertensive rats (WHR) were sacrificed and their right kidneys were harvested 4 weeks after surgery. The spontaneously hypertensive rats (SHR) were divided into 4, 8, and 16 weeks old groups (SHR4w, SHR8w, and SHR16w), respectively. The control group were sham operated age-matched Wistar rats. Immunohistochemical technique and Western blotting were applied to study ERK1/2 protein expression in VSMC of the renal vascular trees in WHR, SHR, and control rats.

RESULTSBlood pressure in two-kidney one clip WHR obviously increased at one week after surgery, and reached to 198. 00 +/- 33. 00 mm Hg at the end of experiment, significantly higher than that in the control rats (P < 0.01). Blood pressure in SHR4w (108.00 +/- 11.25 mm Hg) was similar to that in the controls. However, it rose to 122.25 +/- 21.75 mm Hg in SHR8w, and even up to 201.75 +/- 18.00 mm Hg in SHR16w, which were significantly higher than that of both the SHR4w and the controls (P < 0.01). The rate and degree of glomerular fibrosis in WHR were significantly higher than controls (P < 0.05). Hyaline degeneration of the afferent arterioles was found in WHR. In contrast, either fibrosis of glomerulus or hyaline degeneration of the arterioles or protein casts was not observed in SHR4w, SHR8w, and SHR16w. Immunohistochemical staining results showed expression of ERK1 was similar to that of ERK2. The positive rates of ERK2 staining in VSMC of afferent arterioles, interlobular, interlobar, and arcuate arteries in two-kidney one clip WHR were significantly higher (7.09% +/- 1.75%, 14.57% +/- 4.58%, 29.44% +/- 7.35%, and 13.63% +/- 3. 85%, respectively) than that of the controls(P < 0.01). The positive rates of ERK2 staining in VSMC at afferent arterioles, interlobular, interlobar, and arcuate arteries in SHR16w were significantly higher (12.09% +/- 1.40%, 24.17% +/- 6.92%, 32.44% +/- 4.05%, and 18.61% +/- 3.35%, respectively) than that of the controls (P < 0.01), too. The expression of ERK1/2 protein of kidney in WHR and SHR16w was significantly higher than that in the controls by Western blotting assay (P < 0.01).

CONCLUSIONExtracellular signal transduction system are highly expressed in kidney VSMC of two-kidney one clip WHR and SHR. Phospho-ERKI/2 may play an important role in VSMC hypertrophy and hyperplasia under hypertension.

Animals ; Arterioles ; enzymology ; Fibrosis ; Hypertension ; metabolism ; pathology ; Kidney Glomerulus ; blood supply ; pathology ; Male ; Mitogen-Activated Protein Kinase 1 ; metabolism ; Mitogen-Activated Protein Kinase 3 ; metabolism ; Muscle, Smooth, Vascular ; cytology ; Myocytes, Smooth Muscle ; enzymology ; Rats ; Rats, Inbred SHR ; Rats, Wistar

OBJECTIVESTo detect the expression of Cathepsin B (CatB) in the intracranial aneurysm wall and its effect to the apoptosis of vascular smooth muscle cells, aimed at clarifying the pathological formation mechanism of intracranial aneurysm.

METHODSFrom November 2006 to February 2009, 20 intracranial aneurysm samples were collected as the experimental group, and 6 cases of normal pallium artery samples were collected as the control group. Immunohistochemical technique was used to evaluate the expressions of CatB, alpha-smooth muscle actin (alpha-SMA) and Caspase-3. The expression of CatB mRNA was evaluated by real-time PCR. The ultrastructure of intracranial aneurysms were observed by using the transmission electronic microscope.

RESULTSCompared with the normal pallium artery specimens, the expression of CatB and Caspase-3 both significantly increased in the intracranial aneurysm walls where alpha-SMA decreased (P < 0.05). The mean expression of CatB mRNA in intracranial aneurysm samples was about 3.8-folds than that in control group (P < 0.01). There were excessive apoptotic vascular smooth muscle cells (VSMCs) in the tunica median, and typical apoptotic body were observed in some aneurysm walls.

CONCLUSIONCathepsin B may be involved in the formation and the progression of intracranial aneurysm.

Adolescent ; Adult ; Aged ; Apoptosis ; Case-Control Studies ; Cathepsin B ; metabolism ; Female ; Humans ; Intracranial Aneurysm ; enzymology ; pathology ; Male ; Middle Aged ; Muscle, Smooth, Vascular ; pathology ; Young Adult

The effect of genistein on aortic atherosclerosis was studied by immunohistochemistry with RAM-11 and HHF-35 antibodies and western blotting for matrix metalloproteinase-3 (MMP-3) in New Zealand White rabbits. After provocation of atherosclerosis with hyperlipidemic diet, the rabbits were divided as hyperlipidemic diet group (HD), normal diet group (ND) and hyperlipidemic plus genistein diet group (HD+genistein) for 4 and half months. The average cross sectional area of atherosclerotic lesion was 0.269 mm2 after provocation. The lesion was progressed by continuous hyperlipidemic diet (10.06 mm2) but was increased mildly by genistein (0.997 mm2), and decreased by normal diet (0.228 mm2). The ratio of macrophages to smooth muscle cells in the lesion was not changed by genistein supplementation. The western blotting showed reduction of MMP-3 expression in HD+genistein and ND groups than HD group. The inhibition of atherogenesis by genistein was might be due to improve the endothelial dysfunction rather than direct action on macrophages and/or smooth muscle cells in the lesion, since endothelial dysfunction by lipid peroxidation was the main atherogenic factor in the hypercholesterolemicrabbits. The genistein supplementation also suggests that it helps the stabilization of the atherosclerotic lesion by inhibition of MMP-3 expression.
Animals ; Aorta/pathology ; Arteriosclerosis/*drug therapy/pathology/*prevention & control ; Blotting, Western ; Diet, Atherogenic ; Genistein/*pharmacology ; Growth Inhibitors/*pharmacology ; Hypercholesterolemia/*drug therapy/pathology ; Macrophages/pathology ; Male ; Muscle, Smooth, Vascular/enzymology/pathology ; Rabbits ; Research Support, Non-U.S. Gov't ; Stromelysin 1/metabolism

The heme oxygenase-1 (HO-1), a rate-limiting enzyme in heme metabolism, has been recently defined as a novel stress-stimulated protein, since the intracellular expression of HO-1 in response to various stimuli as oxidation, ischemia and endotoxin injury has been proved to be able to protect the cells from damage. In this study, a retroviral vector containing human HO-1 gene was constructed and transfected to rat vascular smooth muscle cells (VSMCs). Using Southern and Northern blot analyses, the integration and mRNA expression of HO-1 gene in the transfected cells were confirmed. The profound protein expression of HO-1 as well as HO enzyme activity in the transfected cells increased by 1.8-fold and 2.0-fold respectively as compared with the non-transfected cells. It was found that the HO-1 transfected-VSMCs presented dominant resistance to toxicity produced by exposure to H2O2, as a significant protective effect of HO-1 marked by cell survival and LDH leakage was observed when 200, 400 and 600 micromol/L of H2O2 were used. The protection of HO-1 rapidly declined after the transfected-VSMCs were pretreated 24 h with an HO-1 specific inhibitor (ZnPP-IX). The results of this investigation suggest that the functional expression of HO-1 gene within VSMCs raises an alternative ability to protect the vascular cells against active oxygen injury.
Animals ; Cells, Cultured ; Gene Expression ; Genetic Vectors ; Heme Oxygenase (Decyclizing) ; biosynthesis ; genetics ; Heme Oxygenase-1 ; Hydrogen Peroxide ; toxicity ; Muscle, Smooth, Vascular ; enzymology ; pathology ; physiology ; Oxidants ; toxicity ; Rats ; Rats, Inbred WKY ; Retroviridae ; genetics ; Transfection