Chemical generation of nitric oxide in the mouth from the enterosalivary circulation of dietary nitrate.

High concentrations of nitrite present in saliva (derived from dietary nitrate) may, upon acidification, generate nitrogen oxides in the stomach in sufficient amounts to provide protection from swallowed pathogens. We now show that, in the rat, reduction of nitrate to nitrite is confined to a specialized area on the posterior surface of the tongue, which is heavily colonized by bacteria, and that nitrate reduction is absent in germ-free rats. We also show that in humans increased salivary nitrite production resulting from nitrate intake enhances oral nitric oxide production. We propose that the salivary generation of nitrite is accomplished by a symbiotic relationship involving nitrate-reducing bacteria on the tongue surface, which is designed to provide host defence against microbial pathogens in the mouth and lower gut. These results provide further evidence for beneficial effects of dietary nitrate.

Sustained inhibition of intimal thickening. In vitro and in vivo effects of polymeric beta-cyclodextrin sulfate.

Intimal thickening after vascular injury may be modulated in part by heparin binding growth factors. We hypothesized that placement of a therapeutic polymer in the periadventitial space capable of tightly binding growth factors might alter the vascular response to injury. We first demonstrated that incubation of rat aortic smooth muscle cells with an insoluble, sulfated polymer of beta-cyclodextrin (P-CDS) was associated with a dose-dependent inhibition of proliferation induced by fetal calf serum, fibroblast growth factor-2 (FGF-2), platelet-derived growth factor BB, or epidermal growth factor. Preincubation studies of P-CDS with FGF-2 revealed a very rapid removal of mitogenic activity. Using radiolabeled FGF-2 (0.25 microg/ml), we observed a very rapid association rate (0.34 +/- 0.07 min-1, n=4) and a very slow dissociation rate (3.3 +/- 0.2 X 10(-7) min-1) at 37 degrees C, suggesting a high affinity interaction. Using both Transwell and linear under-agarose assays, we demonstrated a significant inhibition of random migration (chemokinesis) by P-CDS. Unsulfated polymeric beta-cyclodextrin (P-CD) had little if any of these effects, suggesting that the high negative charge density of P-CDS was important for the effects. Finally, rats undergoing carotid artery balloon injury were randomized to treatment with periadventitial P-CDS or no treatment, and were killed at 4 (n=20), 14 (n=59), and 88 d (n=14). Morphometric analysis demonstrated significant and sustained inhibition of intimal thickening in P-CDS-treated rats at 14 (P < 0.01) and 88 d (P < 0.05) using absolute intimal area or intima/media area ratios. No inhibition was seen in a group of rats treated with P-CD. In P-CDS-treated rats, bromodeoxyuridine labeling studies revealed fewer labeled smooth muscle cells in the intima at 14 d (P=0.01), while staining with Evans blue revealed enhanced late endothelial cell regrowth. Thus, periadventitially applied sulfated beta-cyclodextrin polymer, which can tightly bind heparin binding growth factors, inhibits intimal thickening in vivo in a sustained fashion without using an additional delivery system. These studies suggest that cellular processes mediated by heparin binding growth factors may be modulated by P-CDS.

Perigraft air, fever, and leukocytosis after endovascular repair of abdominal aortic aneurysms.

BACKGROUND: The postimplantation syndrome of fever and leukocytosis after endovascular repair of infrarenal aortic aneurysms has not been previously characterized and its etiology is not known. METHODS: We studied the first 12 patients who underwent successful endovascular repair of infrarenal aortic aneurysms with Dacron-covered stent-grafts, as part of an ongoing phase II clinical trial. Sepsis syndrome evaluations (physical examination, urinalysis, chest radiograph, urine cultures, and blood cultures) were performed for all patients with postoperative temperature (T) greater than 101.4 degrees F. Computed tomography scans of the abdomen were performed, as part of the clinical protocol, on postoperative days 2 and 30. RESULTS: Fever (T > 101.4 degrees F) was seen in 8 of 12 (67%) patients (P < 05). An additional 2 of 12 (17%) patients had low-grade fevers (100.3 degrees F, 100.6 degrees F). Only 2 of 12 (17%) patients remained afebrile postoperatively. Leukocytosis with counts over 11,000 white blood cells (WBC)/dL was observed in 7 of 12 (58%) patients (P < 05). Sepsis evaluations failed to identify any source of infection in 11 of 12 (97%) patients. Computed tomography scan evidence of perigraft air was noted in 8 of 12 (67%) patients. All patients were afebrile, had normal white blood cell counts, and were discharged within 1 week postoperatively. There has been no evidence of graft infection after 1 to 6 months of follow-up. CONCLUSIONS: Fever and leukocytosis after stent-graft repair of aortic aneurysms does not represent evidence of systemic or graft infection and is not clearly related to nonspecific causes of postoperative fever and leukocytosis. Moreover, the finding of early postoperative perigraft air is not necessarily an indication of graft infection even when concurrently present with fever and leukocytosis.

Dietary nitrate in man: friend or foe?

Based on the premise that dietary nitrate is detrimental to human health, increasingly stringent regulations are being instituted to lower nitrate levels in food and water. Not only does this pose a financial challenge to water boards and a threat to vegetable production in Northern Europe, but also may be eliminating an important non-immune mechanism for host defence. Until recently nitrate was perceived as a purely harmful dietary component which causes infantile methaemoglobinaemia, carcinogenesis and possibly even teratogenesis. Epidemiological studies have failed to substantiate this. It has been shown that dietary nitrate undergoes enterosalivary circulation. It is recirculated in the blood, concentrated by the salivary glands, secreted in the saliva and reduced to nitrite by facultative Gram-positive anaerobes (Staphylococcus sciuri and S. intermedius) on the tongue. Salivary nitrite is swallowed into the acidic stomach where it is reduced to large quantities of NO and other oxides of N and, conceivably, also contributes to the formation of systemic S-nitrosothiols. NO and solutions of acidified nitrite, mimicking gastric conditions, have been shown to have antimicrobial activity against a wide range of organisms. In particular, acidified nitrite is bactericidal for a variety of gastrointestinal pathogens such as Yersinia and Salmonella. NO is known to have vasodilator properties and to modulate platelet function, as are S-nitrosothiols. Thus, nitrate in the diet, which determines reactive nitrogen oxide species production in the stomach (McKnight et al. 1997), is emerging as an effective host defence against gastrointestinal pathogens, as a modulator of platelet activity and possibly even of gastrointestinal motility and microcirculation. Therefore dietary nitrate may have an important therapeutic role to play, not least in the immunocompromised and in refugees who are at particular risk of contracting gastroenteritides.

Growth factor gene expression by intimal cells in healing polytetrafluoroethylene grafts.

Vascular smooth muscle cell proliferation resulting in intimal hyperplasia is a major cause of late graft failure. In baboons, healing 60 microns internodal distance polytetrafluoroethylene vascular grafts form an intima composed of proliferating smooth muscle cells with a luminal lining of endothelium. The presence of intimal smooth muscle cell proliferation underneath an intact endothelium, without platelet adherence, suggests that intimal cells rather than platelets may provide the growth factors regulating the smooth muscle cell proliferation. This idea is supported by the observation that, when segments of graft and artery are excised and perfused ex vivo, there is greater mitogenic activity present in the graft perfusate compared to artery perfusate. Two factors expressed by vascular wall cells and known to influence smooth muscle cell growth in vitro are platelet-derived growth factor and transforming growth factor-beta 1. The expression of these growth factors was measured by Northern blot analysis of total ribonucleic acid extracted from thoracic aorta and the intima of 6-week thoracoabdominal polytetrafluoroethylene grafts, and from smooth muscle cell cultured from the aorta and polytetrafluoroethylene graft. Growth of the cultured smooth muscle cell was arrested in serum-free conditions for 3 days and then stimulated with 10% fetal calf serum. Twenty-four hours later, the smooth muscle cells were harvested. Probing the blots for platelet-derived growth factor-A, platelet-derived growth factor-B, and transforming growth factor-beta 1 messenger ribonucleic acid revealed that in vivo, the graft intima expressed more platelet-derived growth factor-A than the aorta.(ABSTRACT TRUNCATED AT 250 WORDS)

Healing of polytetrafluoroethylene arterial grafts is influenced by graft porosity.

The importance of porosity in synthetic arterial graft healing has not been adequately defined. To determine the effect of porosity on graft healing, we measured the extent of cellular response at late times in 4 mm internal diameter polytetrafluoroethylene grafts of varying porosity (between 10 and 90 microns internodal distance) inserted into the arterial system of baboons. After 1 and 3 months the grafts were retrieved and examined for endothelial coverage, intimal thickening, and endothelial cell and smooth muscle cell proliferation. The pattern of intimal healing with endothelial cells and smooth muscle cells was only related to porosity in the sense that there was an abrupt switch in the pattern of healing as porosity was increased from 30 to 60 microns. In low porosity grafts (10 and 30 microns internodal distances) endothelial coverage of the luminal surface was incomplete and, along with intimal thickening, was limited to graft near the anastomosis. In high porosity grafts (60 and 90 microns internodal distances) luminal endothelial coverage was complete, and intimal thickening was uniformly distributed throughout the graft. The highest porosity graft studied (90 microns) developed areas of focal loss of endothelial cells at late time periods. In this limited series there does appear to be an optimal porosity for polytetrafluoroethylene grafts near 60 microns, since 10 and 30 micron grafts fail to achieve luminal endothelial cell coverage, and 90 micron grafts exhibit instability of the intima with focal endothelial cell loss.

Chemical synthesis of nitric oxide in the stomach from dietary nitrate in humans.

BACKGROUND/AIMS: It has been suggested that dietary nitrate, after concentration in the saliva and reduction to nitrite by tongue surface bacteria, is chemically reduced to nitric oxide (NO) in the acidic conditions of the stomach. This study aimed to quantify this in humans. METHODS: Ten healthy fasting volunteers were studied twice, after oral administration of 2 mmol of potassium nitrate or potassium chloride. Plasma, salivary and gastric nitrate, salivary and gastric nitrite, and gastric headspace NO concentrations were measured over six hours. RESULTS: On the control day the parameters measured varied little from basal values. Gastric nitrate concentration was 105.3 (13) mumol/l (mean (SEM), plasma nitrate concentration was 17.9 (2.4) mumol/l, salivary nitrate concentration 92.6 (31.6) mumol/l, and nitrite concentration 53.9 (22.8) mumol/l. Gastric nitrite concentrations were minimal (< 1 mumol/l). Gastric headspace gas NO concentration was 16.4 (5.8) parts per million (ppm). After nitrate ingestion, gastric nitrate peaked at 20 minutes at 3430 (832) mumol/l, plasma nitrate at 134 (7.2) mumol/l, salivary nitrate at 1516.7 (280.5) mumol/l, and salivary nitrite at 761.5 (187.7) mumol/l after 20-40 minutes. Gastric nitrite concentrations tended to be low, variable, and any rise was non-sustained. Gastric NO concentrations rose considerably from 14.8 (3.1) ppm to 89.4 (28.6) ppm (p < 0.0001) after 60 minutes. All parameters remained increased significantly for the duration of the study. CONCLUSIONS: A very large and sustained increase in chemically derived gastric NO concentrations after an oral nitrate load was shown, which may be important both in host defence against swallowed pathogens and in gastric physiology.

Failure of endovascular abdominal aortic aneurysm graft limbs.

OBJECTIVE: Endovascular abdominal aortic aneurysm (AAA) grafts are subject to subsequent failure of endograft limbs. We sought to determine what device-related factors could be identified that might contribute to limb failure. METHODS: We reviewed the records of patients who had undergone endovascular AAA repair and femorofemoral bypass grafting at a single institution. RESULTS: Endovascular AAA repair was performed in 173 patients. There were 137 bifurcated endografts and 36 aortomonoiliac grafts combined with femorofemoral bypass grafts, yielding a total population of 310 aortic graft limbs and 36 femorofemoral grafts. Thirty-nine additional patients underwent femorofemoral bypass grafting for occlusive disease. The cumulative primary patency of all endografts performed for AAA was 92% at 21 months. Secondary patency was achieved for all failed endograft limbs. There were 24 aortic graft limb "failures" that required intervention: seven limbs underwent thrombosis requiring revision; kinked limbs requiring stenting either at the time of graft placement (17) or subsequently (7) were identified. Fully supported endograft limbs had better primary patency (97% at 18 months) than unsupported limbs (69% at 18 months, P <.001). The aortomonoiliac grafts with femorofemoral bypass grafts tended to have better patency (97% at 18 months) than bifurcated endografts (90% at 18 months), but this did not reach statistical significance (P =.28, not significant). Femorofemoral grafts performed for occlusive disease were found to have somewhat lower patency than those performed for AAA (83% vs 92% at 18 months of follow-up, P =.37, not significant). CONCLUSIONS: Fully supported AAA endografts provide superior endograft limb patency compared with unsupported designs. Consideration should be given to routine stenting of all unsupported endograft limbs. Aortomonoiliac grafts and bifurcated grafts provide similar results for endograft limb patency. Femorofemoral bypass grafts performed in conjunction with aortomonoiliac grafts for AAA disease provide excellent short-term patency.

Protection against oral and gastrointestinal diseases: importance of dietary nitrate intake, oral nitrate reduction and enterosalivary nitrate circulation.

Over the last 20 years, dietary nitrate has been implicated in the formation of methemoglobin and carcinogenic nitrosamines in humans. This has led to restrictions of nitrate and nitrite levels in food and drinking water. However, there is no epidemiological evidence for an increased risk of gastric and intestinal cancer in population groups with high dietary vegetable or nitrate intake. A reevaluation of our currently very negative perception of dietary nitrates comes from recent research into the metabolism and enterosalivary circulation of nitrate in mammals. These studies showed that nitrate is converted to nitrite in the oral cavity that then "fuels" an important mammalian resistance mechanism against infectious diseases. Moreover, there is now evidence that the conversion of nitrate into oxides of nitrogen prevents the formation carcinogenic nitrosamines.