Reduced expression of miR-503 is associated with poor prognosis in cervical cancer.

OBJECTIVE: We wished to evaluate the association between expression of microRNA (miR) -503 and prognosis in patients with cervical cancer. PATIENTS AND METHODS: 96 paired specimens of cervical cancer and adjacent normal cervical epithelial tissues were obtained. qPCR was used to evaluate expression levels of miR-503. We analyzed the associations between miR-503 expression levels and clinicopathological parameters, as well as recurrence-free and overall survival with Kaplan-Meier survival curves and proportional hazards model. RESULTS: miR-503 levels were significantly (p < 0.001) lower in cervical cancer tissue compared with adjacent normal cervical epithelial tissue. We further observed significant associations of expression of this miR and recurrence of cervical cancer, lymph node metastasization, and International Federation of Gynecology and Obstetrics (FIGO) stage. In addition, in multivariate analysis, miR-503 expression level was found to be an independent prognostic factor for both recurrence-free and overall survival. CONCLUSIONS: Reduced expression of miR-503 is an independent prognostic factor in cervical cancer indicating poor prognosis.

Immunohistochemistry of omega class glutathione S-transferase in human tissues.

Omega class glutathione transferase (GSTO) has been recently described in a number of mammalian species. We used immunohistochemistry to determine the cellular and tissue distribution of GSTO1-1 in humans. Expression of GSTO1-1 was abundant in a wide range of normal tissues, particularly liver, macrophages, glial cells, and endocrine cells. We also found nuclear staining in several types of cells, including glial cells, myoepithelial cells of the breast, neuroendocrine cells of colon, fetal myocytes, hepatocytes, biliary epithelium, ductal epithelium of the pancreas, Hoffbauer cells of the placenta, and follicular and C-cells of the thyroid. These observations and the known activity of GSTO1-1 suggest biological functions that are not shared with other GSTs.

Wash-in methodology and modeling to determine hepatocellular D-glucose transport in the perfused rat liver.

Wash-in experiments, which may be useful for the study of the disposition of substrates in the liver, have not been well described. To investigate physiological models for wash-in curves, we performed experiments on the perfused livers of male Wistar rats anesthetized with pentobarbital. Test perfusate contained (14)C-sucrose as the extracellular marker and (3)H-glucose. Liver perfusions were performed with background glucose concentrations of 5.7, 10.7, 51.2, or 108.5 mM. Outflow time-activity curves were analyzed with the use of four models. The V(max) and K(m) for the influx of glucose were 1.1 +/- 0.03 micromol/s/g and 41 +/- 3 mM with the Crone-Renkin early extraction model; 1.4 +/- 0.04 micromol/s/g and 36 +/- 3 mM with dispersion model analysis; 1.8 +/- 0.1 micromol/s/g and 25 +/- 4 mM with the Goresky distributed model to fit differentiated wash-in curves; and 2.7 +/- 0.6 micromol/s/g and 28 +/- 21 mM with compartmental analysis. There was reasonable agreement between the four models, and they yielded results similar to those reported for glucose uptake in other preparations.

Endovesical instillation of mitomycin-C in preventing recurrence of superficial bladder carcinoma.

It is now clear that MMC can be used as endovesical instillation after TUR, partial cystectomy or transurethral laser treatment and at the same time it has chemo-resection and chemoprophylactic efficacy. The success of treatment seems out of question on dosage but with the continuation of the instillation program. Recently, we have adopted 20 mg per instillation once every week for 40 times in the first year and once for 2-4 weeks for the second year, the overall recurrent rate was 17%. Such dosage used is more better than 2 mg, 10 mg or even 40 mg with acceptable side effects.

A nitric oxide donor (spermine-NONOate) prevents the formation of neointima in rabbit carotid artery.

1. In the present study we investigated the effect of spermine diazeniumdiolate (spermine-NONOate), a nitric oxide donor, on the early development of atheroma-like lesions induced by a peri-arterial collar in rabbits. 2. Spermine-NONOate was given locally by incorporating the compound (1 mg/mL) into a silastic collar, which was applied on one common carotid artery of rabbit while the other carotid artery had a placebo collar (without compound) applied. 3. Fourteen days postimplantation, both carotid arteries were dissected free for histological study (n = 6). 4. After 14 days with collars, treatment with spermine-NONOate had significantly reduced (by 74%) the thickness of the neointima in comparison with the contralateral collared artery without compound. Blood pressure did not change during treatment. Nitric oxide, detected as nitrite, was still released from spermine-NONOate silastic collars after 14 days implantation. 5. These results suggest that locally administered spermine-NONOate is effective in slowing the development of neointima in this model.

A platelet-activating factor antagonist (WEB 2170) preserves endothelium-dependent vasodilatation and prevents development of a neo-intima induced by a periarterial collar in rabbit carotid arteries.

Platelet-activating factor (PAF) may be involved in adhesion of leucocytes and migration of cells during vascular remodelling for it is expressed in leucocytes after cytokine priming and is required for cell adhesion. We studied the effects of WEB 2170, a potent PAF antagonist, on the development of an atheroma-like neo-intima induced by a peri-arterial collar in rabbits. Either WEB 2170 (3 mg/kg/day) or vehicle was given by subcutaneous injection once a day for 4 or 9 days, and on day 3 peri-arterial collars were applied to both carotid arteries in all animals. Two or 7 days after implanting the collars vasodilator responses to the endothelium-dependent vasodilator, acetylcholine and the endothelium-independent vasodilator, sodium nitroprusside were studied in isolated artery rings from both groups of rabbits. Neo-intima formation after 7 days (day 10 of treatment) was measured by light microscopy as the ratio of cross-sectional areas of intima and media, and expression of inducible nitric oxide synthase (iNOS) was studied by immunohistochemistry. PAF-induced platelet aggregation ex vivo was inhibited specifically in WEB 2170-treated rabbits. At day 5, acetylcholine-induced vasorelaxation in collared artery rings was markedly impaired as compared to control sections from both vehicle- and WEB 2170-treated rabbits. At day 10, acetylcholine-induced vasorelaxation in collared artery rings from vehicle rabbits was markedly less than in controls, but in WEB 2170-treated rabbits, the acetylcholine response in collared arteries was similar to control sections. Intimal thickening was much reduced in WEB 2170-treated rabbits, ratios of intima/media areas being vehicle: 0.21 +/- 0.02 (n = 5) and WEB 2170: 0.07 +/- 0.01 (n = 7; p < 0.01). Immunofluorescence showed expression of iNOS only in the neo-intima of vehicle-treated, collared arteries, but not in the residual neo-intima of WEB 2170-treated, collared arteries. These results suggest that WEB 2170 is effective in preserving endothelial function, prevents the development of neo-intima and blocks iNOS expression in the neo-intima in this model.

Induction of nitric oxide synthase in the neointima induced by a periarterial collar in rabbits.

A Silastic collar placed around the common carotid artery of rabbits causes the formation, within 7 days, of an atheroma-like neointima containing cells with the appearance of synthetic-phenotype smooth muscle cells. Using immunohisto-chemistry, we detected the appearance of the cytokine-inducible form of nitric oxide synthase (iNOS, or isoform II) in the neointima of rabbits that had the collar in place for 7 or 14 days. This iNOS immunofluorescence collocalized with anti-smooth muscle myosin in the intima, indicating that it is expressed in smooth muscle cells, and iNOS was also present in a few endothelial cells in collared sections. There was no evidence of iNOS expression in the arterial wall before the neointima was apparent, that is, after only 2 days with the collar. The expression of endothelial NOS (eNOS, or isoform III) immunofluorescence was confined to the endothelial cells in control sections, as it was in collared sections with neointima at 7 and 14 days. Specific immunofluorescence for neuronal NOS (nNOS, or isoform I) was not observed in any sections. Our results suggest that nitric oxide is produced by the inducible isoform of NOS in modified smooth muscle cells of the developing neointima. Activity of iNOS might deprive the endothelium of substrate for nitric oxide production and might explain the compromised endothelium-dependent vasodilatation observed both in this model of atherosclerosis and in human coronary artery disease.

Carbon monoxide disposition in the perfused rat liver.

A simple method for determining carbon monoxide (CO) disposition in the rat liver perfused with erythrocyte-free buffer was developed. Wash-in experiments were performed with buffer containing tracer quantities of [14C]sucrose and 3H2O and equilibrated with CO. Outflow samples were collected into tubes containing human erythrocytes, which avidly bind CO. Outflow curves were analyzed using compartmental models. Fractional recovery of CO was 1.07 +/- 0. 17, and the apparent volume of distribution was 1.37 +/- 0.30 ml/g of liver (n = 8). A flow-limited model fitted the data most effectively, although estimates of the permeability-to-surface area product were attempted using a barrier-limited model. This technique will facilitate investigation of the effects of disease on gaseous substrate disposition in perfused organs.

Nitric oxide in atherosclerosis: vascular protector or villain?

1. Nitric oxide (NO) has important roles in physiological vasodilatation, cytotoxicity and vascular disease. Nitric oxide and prostacyclin (PGI2), both released from the endothelium, act synergistically to inhibit platelet aggregation and adhesion. These autacoids also inhibit the adhesion and migration of leucocytes and, in some arteries, they synergize in terms of vasodilation. 2. The development of atherosclerosis and hyperlipaemia per se is accompanied by impairment of endothelium-dependent vasodilation. 3. Atherosclerosis is associated with marked changes in the activity of isoforms of NO synthase (NOS) in the artery wall, including increased expression of the NOS2 (inducible) isoform in complex human lesions as well as in the neointima of experimental animal models. 4. Failure of NO release from the endothelium with normal physiological stimuli, which has been attributed to a defect in the operation of the endothelial NOS (NOS3), provides conditions propitious for leucocyte adhesion, vasospasm, thrombosis and, in addition, may promote increased proliferation of intimal cells. 5. Nitric oxide and superoxide anions generated by inflammatory cells in atherosclerosis react to form cytodestructive peroxynitrite radicals, potentially causing injury to the endothelium and myocytes, and this may be a factor in apoptosis of cells leading to plaque rupture. 6. We have been able to reverse these NO defects with therapeutic agents, including angiotensin-converting enzyme inhibitors, antagonists of platelet-activating factor and NO donor compounds, all offering promise in protecting against some manifestations of vascular disease.

Pseudocapillarization and associated energy limitation in the aged rat liver.

Age-related impairment of drug metabolism by the liver is consistent with hepatocyte hypoxia, suggestive of the development of a diffusional barrier to oxygen supply. Because the effects of aging on the diffusional pathway (sinusoidal endothelium and space of Disse) have not been described, we performed comparative studies on the livers of Fischer F344 rats aged 4 to 7, 12 to 15, and 24 to 27 months. Light-microscopic examination revealed no evidence of fibrosis, cirrhosis, or other specific pathology. In contrast, scanning and transmission electron-microscopic examination revealed that aging is associated with pseudocapillarization of the sinusoidal endothelium, indicated by defenestration with reduced porosity, thickening of the endothelium, infrequent development of basal lamina, and only minor collagen deposits in the space of Disse. Furthermore, immunohistochemistry studies showed strong expression of collagen IV, moderate expression of factor VIII-related antigen, and weak expression of collagen I along the sinusoids of livers from old rats (P <.0001). In vitro (31)P magnetic resonance spectroscopy analysis showed that aging is associated with changes in high-energy phosphate and other metabolites, consistent with hepatocyte hypoxia. Aging in the liver is associated with changes in the sinusoidal endothelium and space of Disse that may restrict the availability of oxygen and other substrates.