OBJECTIVETo observe the distribution and expression changes of caveolin-1 under shear stress in a turbulence flow channel model mimicking susceptible sites of atherosclerosis in vitro.

METHODSHuman umbilical vein endothelial cells (HUVECs) were inoculated to flow channel and then exposed to disturbed shear stress for 6, 12, 24 and 48 hours, respectively. HUVECs inoculated to flow channel without shear stress served as controls. Immunofluorescence staining was performed with SABC-FITC to detect the caveolin-1 distribution and RT-PCR was used to determine the mRNA expression of caveolin-1 gene in these cells.

RESULTSConfocal microcopy revealed ellipse-like shaped HUVECs under disturbed shear stress and caveolin-1 didn't move toward to the upstream edge of HUVECs but moved toward to the edge of HUVECs, and expression of caveolin-1 gene in HUVECs was significantly downregulated after exposure to shear stress for 48 hours compared to HUVECs without shear stress stimulation.

CONCLUSIONDisturbed shear stress affected disposition and expression of caveolin-1 which might be responsible for shear stress induced endothelial cell pathological atherosclerotic changes.

Caveolin 1 ; Cells, Cultured ; Endothelium, Vascular ; metabolism ; Humans ; Interleukin-8 ; Stress, Mechanical ; Umbilical Veins

OBJECTIVETo study the relationship between the dynamic changes of caveolin-1 with the degrees of liver fibrosis and portal venous pressure (PVP) in the process of rat liver cirrhosis formation induced by dimethylnitrosamine (DMN); also to investigate the mechanisms of caveolin-1 in the regulation of endothelial nitric oxide synthase (eNOS).

METHODSLiver cirrhosis was induced in rats by DMN. The degrees of liver fibrosis and PVP were measured. NOS activity was assessed by citrulline generation. Protein expressions of caveolin-1, eNOS and caveolin-1-eNOS interactions were examined by Western blot and immunoprecipitation, respectively.

RESULTSFour weeks after DMN administration, liver fibrosis was at its peak and then decreased gradually. Immunoprecipitation and Western blot demonstrated that there was enhanced binding of caveolin-1 with eNOS in the process of rat liver cirrhosis. An increase in caveolin-1 expression was detected but the expression of eNOS was lower in cirrhotic tissues than in normal liver tissues. Caveolin-1 protein levels were positively correlated with the degrees of liver fibrosis and the levels of PVP (r=0.967, P < 0.01; r=0.922, P < 0.01, respectively), while NOS catalytic activity was negatively correlated with the degrees of liver fibrosis and levels of PVP (r= 0.973, P < 0.01; r=-0.947, P < 0.01) respectively.

CONCLUSIONSCaveolin-1 upregulation is associated with the development of portal hypertension in liver cirrhosis. Over-expression of caveolin-1 in perisinusoidal cells may promote caveolin-1-eNOS binding and reduce the activity of eNOS.

Animals ; Caveolin 1 ; metabolism ; Hypertension, Portal ; metabolism ; Liver Cirrhosis, Experimental ; metabolism ; Nitric Oxide Synthase Type III ; metabolism ; Rats ; Rats, Wistar

Caveolae are specialized lipid rafts that form flask-shaped invaginations of the plasma membrane. Many researches show that caveolae are involved in cell signaling and transport. Caveolin-1 is the major coat protein essential for the formation of caveolae. Recently, several reports indicated that the other caveolae-associated proteins, Cavins, are required for caveola formation and organization. It's worth noting that Cavin-1 could cooperate with Caveolin-1 to accommodate the structural integrity and function of caveolae. Here, we reviewed that the relationship between Cavins and Caveolins and explore the role of them in regulating caveolae.
Animals ; Caveolae ; physiology ; Caveolin 1 ; metabolism ; physiology ; Caveolins ; metabolism ; physiology ; Humans ; Membrane Proteins ; metabolism ; physiology ; RNA-Binding Proteins ; metabolism ; physiology

Caveolin 1 ; genetics ; metabolism ; Cells, Cultured ; Homocysteine ; physiology ; Human Umbilical Vein Endothelial Cells ; cytology ; metabolism ; Humans ; RNA, Messenger ; genetics ; metabolism

OBJECTIVETo construct a reverse transcriptase-polymerase chain reaction (RT-PCR)approach that can improve the specificity of primers while dropping down the nonspecific amplification.

METHODSIn the recent study we reported a new RT-PCR assay which improved markedly the specificity. However its efficiency of regressing nonspecific amplification remains to be accurately checked and further documented. In primer design, we looked over again some sequences that showed differences at 5' or 3' ends between human CAV1 and mouse Cav1 genes. cDNAs and the diluted plasmids which harbored the sequence of human CAV1 or mouse Cav1 gene were chosen as the templates. The ordinary PCR compared with one, of which primers modified by phosphorothioate and combined with proofreading polymerase, for their efficiencies of nonspecific amplification inhibited.

RESULTSTaq DNA polymerase without proofreading activity could efficiently catalyze the extension of primers with a single or multiple mismatched base pairs at the 3' terminus, but the kind of primer extension can be effectively blocked by phosphorothioate modified primers combined with proofreading polymerase. Compared with ordinary PCR reaction, this new PCR method can effectively regress the primer mismatched amplification of 50 ng DNA almost equaling to 2 x 10(4) unmatched template copies in a final volume of 50 microL.

CONCLUSIONCompared with the first generation of polymerases with or without proofreading activities mediating RT-PCR reaction, the introduction of nuclease-resistant 3' modified primers (3' phosphorothioate primer extension) can offer more simplicity, accuracy, and also decrease cost.

Animals ; Caveolin 1 ; genetics ; Deoxyribonucleases ; metabolism ; Gene Amplification ; Humans ; Mice ; Reverse Transcriptase Polymerase Chain Reaction ; methods ; Thionucleotides ; metabolism

BACKGROUND/AIMS: Hepatocellular carcinoma (HCC) is becoming one of the common malignant tumors worldwide, and it is characterized by its high vascularity. Caveolin is the major structural protein in caveolae, which are small omega-shaped invaginations within the plasma membrane. Caveolin has been implicated in mitogenic signaling, oncogenesis and angiogenesis. The expression of caveolin-1 and -2 in HCC and its potential relationship with angiogenesis has not been examined. METHODS: Paraffin sections of 35 HCC specimens were immunostained with caveolin-1, caveolin-2, alpha-smooth muscle actin, and CD34 antibodies. In addition, the expression of caveolin-1 and -2 mRNA in HCC was examined. The relationship between the radiological findings and the number of unpaired arteries and microvessel density (MVD) was also investigated. RESULTS: Caveolin-1 and -2 were expressed in the sinusoidal endothelial cells in 20 out of 35, and 18 out of 35 HCC specimens, respectively. Caveolin-1 and -2 were also expressed in the smooth muscle cells of the unpaired arteries in 26 out of 35, and 18 out of 35 HCC specimens, respectively. Increased expression of caveolin-1 and -2 mRNA was detected in 26.7% and 33.3% of the tumor specimens, respectively, compared with the corresponding non-tumorous adjacent liver tissues. There was a significant correlation between expression of caveolin-1, -2 in the smooth muscle cells of unpaired arteries and the number of unpaired arteries. The number of unpaired arteries in HCCs was found to be associated with the degree of contrast enhancement in the arterial phase imaging. However, it did not correlate with the degree of MVD. CONCLUSIONS: These findings suggest that the expression of caveolin-1, -2 is associated with the formation of unpaired arteries in HCC. In addition, there is a correlation between the degree of contrast enhancement of the HCC in the arterial phase image and the number of unpaired arteries.
Adult ; Aged ; Carcinoma, Hepatocellular/*blood supply/metabolism/radiography ; Caveolin 1/genetics/*metabolism ; Caveolin 2/genetics/*metabolism ; Female ; Hepatic Artery/pathology/radiography ; Humans ; Liver Neoplasms/*blood supply/metabolism/radiography ; Male ; Middle Aged ; Neoplasm Staging ; Neovascularization, Pathologic/etiology/*metabolism/radiography ; Retrospective Studies

This study was purposed to explore the effect of lentivirus-mediated CAV1 overexpression on proliferation and apoptosis in HL-60 cells. Recombinant lentiviral expression vector pcDNA-EF1-CAV1 was constructed, and cotransfected the 293TN cells with a mixture of pPACK packaging plasmids. Then collecting virus suspension infects the HL-60 cells, which make CAV1 gene stable transfection and high expression in the cells. The CAV1 protein expression status in HL-60 cells transfected was evaluated through Western blot method. Proliferative activity and apoptosis of HL-60 cells before and after transfection were detected by CCK-8 method and flow cytometry, respectively. The results showed that the PCR-positive clone screening and results of nucleotide sequencing confirmed that the CAV1 gene inserted into the expression vector pcDNA-EF1-GFP correctly, recombinant lentiviral particles Lv-CAV1 transfected HL-60 cells successfully and with transfection rate up to 90%. The result of Western blot showed that CAV1 protein expression in HL-60 cells significantly increased at 48 hours after transfection. CCK-8 result indicated that cell proliferation activity increased at 48 h after transfection (P < 0.05), flow cytometry testing results displayed that apoptosis rate of HL-60 cells obviously decreased after transfection (P < 0.01). It is concluded that the overexpression of CAV1 in HL-60 cells can inhibit cell proliferation activity and promote cell apoptosis.
Apoptosis ; Caveolin 1 ; genetics ; metabolism ; Cell Proliferation ; Genetic Vectors ; HL-60 Cells ; Humans ; Lentivirus ; genetics ; Plasmids ; Transfection

BACKGROUND: Recent studies indicate that light-emitting diodes (LED) may represent a novel and effective anti-aging light source for the skin. Among many candidate molecules known to control collagens, caveolin-1 (Cav-1) is known to play an inhibitory role in cutaneous collagen metabolism. OBJECTIVE: This study aimed to evaluate the effects of LED irradiation on the expression levels of Cav-1 and procollagens (proCOLs) in human dermal fibroblasts (HDFs). METHODS: Cultured HDFs were irradiated with 630 nm LED at different doses, and the mRNA and protein expression levels of Cav-1 and proCOLs I/III were analyzed. RESULTS: In LED-irradiated HDFs, mRNA and protein levels of Cav-1 were found to be down-regulated, whereas those of proCOLs I/III were up-regulated in a dose-dependent manner. A negative correlation between Cav-1 and proCOLs was verified in Cav-1 siRNA transfected HDFs. LED was moreover found to result in up-regulation of transforming growth factor (TGF)-beta1 and its receptors (TbetaRI, TbetaRII), SMAD1, and SMAD2 mRNA levels, indicating that LED may activate the TGF-1/TbetaR/SMAD pathway in HDFs. CONCLUSION: The anti-aging effects of 630 nm LED on human skin are likely mediated by up-regulation of proCOLs I/III and inhibition of Cav-1 expression levels in HDFs.
Caveolin 1* ; Collagen ; Fibroblasts* ; Humans ; Metabolism ; Procollagen* ; RNA, Messenger ; RNA, Small Interfering ; Skin ; Transforming Growth Factors ; Up-Regulation

Animals ; Autophagy ; Biomarkers, Tumor ; metabolism ; Caveolin 1 ; deficiency ; metabolism ; Drug Delivery Systems ; Humans ; Myofibroblasts ; pathology ; Neoplasms ; metabolism ; pathology ; Prognosis ; Stromal Cells ; metabolism ; Tumor Microenvironment

OBJECTIVETo observe the therapeutic effect and mechanism of penehyclidine hydrochloride on paraquat-induced acute lung injury.

METHODS80 healthy adult male Wistar rats were randomly assigned into control groups (10 rats), 100 mg/kg PQ group (10 rats), 100 mg/kg PQ plus 33 µg/kg penehyclidine hydrochloride treatment group (30 rats), 100 mg/kg PQ plus 66 µg/kg penehyclidine hydrochloride treatment group (30 rats). The two treatment groups were executed respectively at 36 h, 72 h and 7 d. Lung tissues were used to assess histopathological change by HE staining. The level of MMP-2, caveolin-1 and HYP were detected in the lung homogenate. The serum and BALF contents of ET were measured.

RESULTSPathology inspection confirmed that the model of acute rat pulmonary injury were duplicated successfully. The level of MMP-2, HYP in lung tissues and the serum and BALF ET contents in PQ group were (1.77 ± 0.40) µg/g, (2.91 ± 0.79) µg/g, (505.23 ± 124.69) µg/ml, (640.38 ± 136.60) µg/ml. The level of those was higher than that in control group [(0.95 ± 0.66) µg/g, (1.48 ± 0.69) µg/g, (95.48 ± 46.01) µg/ml, (200.40 ± 88.39) µg/ml, P < 0.05]; The above-mentioned index in two treatment groups was lower than that in PQ group (P < 0.05). The caveolin-1 content [(1.77 ± 0.82) µg/g] in PQ group was lower than that in control group [(5.39 ± 1.68) µg/g, P < 0.05]. The level of caveolin-1 in two treatment groups was higher than that in PQ group (P < 0.05).

CONCLUSIONPenehyclidine hydrochloride can decrease the level of MMP-2, HYP in lung tissues and the ET in serum and BALF, increase that of caveolin-1 and lessen the damage induced by paraquat.

Acute Lung Injury ; chemically induced ; drug therapy ; Animals ; Caveolin 1 ; metabolism ; Endothelins ; metabolism ; Hydroxyproline ; metabolism ; Male ; Matrix Metalloproteinase 2 ; metabolism ; Paraquat ; toxicity ; Quinuclidines ; therapeutic use ; Rats ; Rats, Wistar