Matrix vesicles in the fibrous cap of atherosclerotic plaque: possible contribution to plaque rupture.

Plaque rupture is the most common type of plaque complication and leads to acute ischaemic events such as myocardial infarction and stroke. Calcification has been suggested as a possible indicator of plaque instability. Although the role of matrix vesicles in the initial stages of arterial calcification has been recognized, no studies have yet been carried out to examine a possible role of matrix vesicles in plaque destabilization. Tissue specimens selected for the present study represented carotid specimens obtained from patients undergoing carotid endarterectomy. Serial frozen cross-sections of the tissue specimens were cut and mounted on glass slides. The thickness of the fibrous cap (FCT) in each advanced atherosclerotic lesion, containing a well developed lipid/necrotic core, was measured at its narrowest sites in sets of serial sections. According to established criteria, atherosclerotic plaque specimens were histologically subdivided into two groups: vulnerable plaques with thin fibrous caps (FCT <100 microm) and presumably stable plaques, in which fibrous caps were thicker than 100 microm. Twenty-four carotid plaques (12 vulnerable and 12 presumably stable plaques) were collected for the present analysis of matrix vesicles in fibrous caps. In order to provide a sufficient number of representative areas from each plaque, laser capture microdissection (LCM) was carried out. The quantification of matrix vesicles in ultrathin sections of vulnerable and stable plaques revealed that the numbers of matrix vesicles were significantly higher in fibrous caps of vulnerable plaques than those in stable plaques (8.908+0.544 versus 6.208+0.467 matrix vesicles per 1.92 microm2 standard area; P= 0.0002). Electron microscopy combined with X-ray elemental microanalysis showed that some matrix vesicles in atherosclerotic plaques were undergoing calcification and were characterized by a high content of calcium and phosphorus. The percentage of calcified matrix vesicles/microcalcifications was significantly higher in fibrous caps in vulnerable plaques compared with that in stable plaques (6.705+/-0.436 versus 5.322+/-0494; P= 0.0474). The findings reinforce a view that the texture of the extracellular matrix in the thinning fibrous cap of atherosclerotic plaque is altered and this might contribute to plaque destabilization.

Stent-graft treatment for bleeding from a presumed aortoenteric fistula.

PURPOSE: To describe a technique for the endovascular treatment of aortoenteric fistula. METHODS AND RESULTS: A 67-year-old man who had undergone aortobi-iliac grafting for aneurysmal disease 8 years previously presented with life-threatening upper gastrointestinal hemorrhage. Endoscopy after resuscitation did not identify the source of the bleeding. Computed tomographic (CT) scanning and angiography revealed pseudoaneurysm formation at the upper anastomosis 1 cm below the renal arteries. Measurements were taken for endovascular repair. Uncomplicated emergency aortic endografting for exclusion of the pseudoaneurysm was performed using a 28-mm x 3.75-cm AneuRx device. Gastrointestinal hemorrhage ceased. CT scanning at 6 months confirmed the absence of a pseudoaneurysm, and the patient remains symptom free at 18 months. CONCLUSIONS: Endovascular treatment of aortoenteric fistula may represent a technique for treating gastrointestinal hemorrhage and for lessening the morbidity and mortality of open repair.

The diagnosis and management of mixed arterial/venous leg ulcers in community-based clinics.

OBJECTIVES: To assess a management protocol for mixed arterial/venous leg ulcers in a community service. DESIGN: Two-year prospective study of outcome with intention of assessing limbs with mixed arterial/venous ulcers when managed by a new protocol. METHOD: Limbs were assessed for venous reflux by duplex and arterial insufficiency by ankle-brachial pressure index (ABPI) and defined into three categories: ABPI > 0.85, 0.5 > ABPI < or = 0.85 (moderate), ABPI < or = 0.5 (severe). Four-layer compression was applied to limbs with normal arteries. Modified compression was applied to limbs with venous and moderate arterial disease with treatment failure triggering arterial imaging and revascularisation. Limbs with venous and severe arterial disease were investigated for revascularisation. RESULTS: Of 267 consecutive limbs, 221 had pure chronic venous ulcers and 46 had mixed arterial/venous ulcers with 33 having moderate and 13 having severe arterial disease. Thirty-six week healing rates for chronic venous, moderate arterial/venous and severe arterial/venous ulcers were 70%, 64% and 23%, respectively. CONCLUSION: Limbs with mixed moderate arterial/venous ulcers achieved rates comparable with venous ulcers with this protocol although nurse-led surveillance was required. Limbs with mixed severe arterial/venous ulcers healed slowly despite an aggressive approach to correct arterial disease.

Influence of a specialised leg ulcer service and venous surgery on the outcome of venous leg ulcers.

OBJECTIVES: To assess the influence of a vascular-led community service on the outcome of chronic leg ulcers. DESIGN: Before and after study. METHOD: Healing and recurrence were compared between ulcerated limbs (n = 149) from a random sample of 200 patients treated in the community and consecutive limbs (n = 200) from 180 patients treated in specialised clinics. In these clinics, vascular disease was routinely identified with venous duplex and ankle-brachial pressure index. Surgery was offered if superficial vein reflux alone was detected. Compression bandaging was applied to limbs with ABPI > 0.85. Healed limbs were treated with compression hosiery. RESULTS: After the clinics were introduced, the 12 and 24-week healing rates increased from 12 and 29 per cent to 53 and 68 per cent respectively (p < 0.01), and the 6 and 12 month recurrence rates decreased from 43 and 54 per cent to 21 and 23 per cent respectively (p < 0.01). Superficial venous surgery reduced recurrence at 1 year to 9 per cent. CONCLUSION: Outcome of leg ulcers is improved in a vascular-led community service. Routine surgical correction, in cases of reflux limited to the superficial system, may further reduce the chance of recurrence.

Colour duplex ultrasonography in the rational management of chronic venous leg ulcers.

Colour duplex ultrasonographic imaging has largely replaced venography in the assessment of lower-limb venous disorders. This is a study of the use of duplex in the management of patients with chronic venous ulceration in community ulcer clinics. Patients with chronic leg ulceration and an ankle: brachial pressure index of 0.85 or greater were studied. Assessment of venous competence in both the deep and superficial systems of the affected and unaffected legs was performed using colour venous duplex imaging. Reflux was defined as reverse flow for greater than 1 s after manual calf compression. One hundred consecutive patients were assessed over 15 months. Of 111 ulcerated legs, 96 had active ulceration, while 15 had been ulcerated within the previous 6 months. Fifty-seven (51 per cent) of the 111 ulcerated legs had superficial incompetence alone (88 per cent long saphenous system or its perforators, 12 per cent short saphenous system). Six legs (5 per cent) had isolated deep venous incompetence. Forty-two legs had mixed superficial and deep venous reflux; 22 of these had undergone previous venous surgery. Colour venous duplex assessment demonstrated superficial venous disease in approximately half of limbs with chronic leg ulceration. Venous dysfunction in these patients is potentially curable by surgery.

High-performance liquid chromatographic assay for the measurement of melphalan and its hydrolysis products in perfusate and plasma and melphalan in tissues from human and rat isolated limb perfusions.

A sensitive, specific and rapid reversed-phase high-performance liquid chromatographic (HPLC) assay was developed for the quantitation of melphalan and its hydrolysis products in samples from the isolated perfusion of human and rat limbs. Samples of perfusate, plasma and tissue were analysed, following methanol precipitation, using a phenyl column and fluorescence detection. Dansyl-arginine (38 micrograms ml-1) was employed as the internal standard. Good resolution was observed allowing quantitation of melphalan, monohydroxymelphalan (MOH) and dihydroxymelphalan (DOH) in perfusate and plasma were all 100 +/- 10%. The recovery of melphalan in tissue was 93.5%. A linear response was demonstrated for melphalan in the concentration range 1.8 - 56.8 micrograms ml-1, for DOH in the concentration range 0.5 - 30.0 micrograms ml-1 and for MOH in the range 1.4-25.1 micrograms ml-1, in perfusate and plasma. The lower limits of quantitation of melphalan, MOH and DOH in perfusate and plasma were 1.4, 2.4 and 1.2 ng on column, respectively, and 7.2 ng of melphalan on column in tissue. Intra-assay coefficients of variation (C.V.) for melphalan, MOH and DOH, at low and high concentrations were all less than 5% and the inter-assay C.V.s were less than 9%. An ultra-filtration study to determine the protein binding of melphalan and the hydrolysis products showed that the unbound fractions (fu) of melphalan in buffer containing dextran and bovine serum albumin were 0.873 and 0.521, respectively. The assay was used to quantitate melphalan and its hydrolysis products in samples from isolated perfusions in the human limb and rat hindlimb.

Melphalan uptake, hyperthermic synergism and drug resistance in a human cell culture model for the isolated limb perfusion of melanoma.

Isolated limb perfusion with melphalan is a long-standing treatment for melanoma but the clinical conditions have not been subjected to a systematic evaluation. In order to establish optimal conditions for perfusion, three human melanoma cell lines were cultured with melphalan in vitro under conditions comparable to in vivo therapy. The most important findings were that: (a) 41.5 degrees C was synergistic for melphalan killing of three human melanoma cell lines; (b) prolonging the treatment time beyond 1 h had little additional toxicity; and (c) varying the initial pH of the culture medium had no effect. After 1 h of treatment, cells accumulated more melphalan at 41.5 degrees C than at 37 degrees C, relative to the extracellular concentration. A cell line (MM418) derived from a primary tumour was the most resistant of the three lines; pigmented or non-pigmented sublines were equally resistant. The A2058 line showed the lowest level of synergism with hyperthermia, and displayed a marked plateau at 10% of controls in the dose-response for survival, yet no melphalan-resistant subpopulation could be isolated. The implications of this work are that (a) enhanced cellular uptake of melphalan may account for hyperthermic synergism of melphalan; (b) varying conditions other than treatment time will be necessary to deal with the variation in resistance between tumours; and (c) repeated cycles of treatment may be needed for phenotypes such as A2058 where melphalan resistance appears to be based on an epigenetic mechanism.