Influence of peroperative lavage solutions on peritoneal defence mechanisms in vitro.

OBJECTIVE: To find out the in vitro reaction of mesothelial cells and polymorphonuclear leucocytes (PMN) to incubation with seven commonly-used lavage solutions. DESIGN: Experimental study. SETTING: Laboratories, The Netherlands. MATERIAL: Cultured human peritoneal mesothelial cells and isolated PMN. INTERVENTION: Incubation of cells with clinically used lavage solutions (sodium chloride, Hartmann's solution, povidone-iodine, Dakin's solution, taurolidine, chlorhexidine, and hydrogen peroxide). MAIN OUTCOME MEASURES: Activation of monolayers of mesothelial cells and PMN measured by release of oxygen free radicals (chemiluminescence) and interleukin (IL)-8 concentrations and toxic effects measured by morphology, release of lactate dehydrogenase, failure of the restriction of the passage of inulin, and incorporation of propidium iodide. RESULTS: All solutions activated and killed mesothelial cells and PMN to some extent; the more concentrated the solution the greater the effect on these cells. CONCLUSION: Lavage solutions both poison and stimulate mesothelial cells and neutrophils, and some solutions are more potent than others.

Piperacillin/tazobactam therapy for diabetic foot infection.

In this study, 29 patients were hospitalized with a diabetic foot infection and were treated with piperacillin/tazobactam. Of these 23 patients who were evaluated for efficacy of treatment, 22 patients improved or were clinically cured. In seven patients (30%), there was persistence of one of the baseline pathogens. Adverse events were reported in 15 patients (58%), three of which were serious. Piperacillin/tazobactam may be useful as monotherapy in diabetic foot infection giving an adequate clinical response and the level of side effects equivalent to those of other broad-spectrum antibiotics.

Peritoneal interleukin-8 in acute appendicitis.

Interleukin-8 (IL-8) is a chemoattractant that is highly selective for neutrophils. This study was designed to investigate the presence of IL-8 in peritoneal fluid of patients with acute appendicitis. The clinical circumstances underlying the secretion of IL-8 by mesothelium and its mechanism of activation have not been defined. In an in vitro model for bacterial peritonitis the role of bacteria in activating human mesothelial cells to secrete IL-8 was studied. Cultured human mesothelium was incubated with various species of pathogenic bacteria, isolated from peritoneal exudate fluids of patients with appendicitis. The amount of IL-8 secreted by the cultured mesothelial cells was determined in an IL-8 ELISA, as IL-8 was present in the original peritoneal fluid of these patients. Peritoneal fluids from patients with a perforated appendix were found to contain a significantly higher concentration of IL-8 compared to peritoneal fluids from patients with nonperforating appendicitis (121.6 (57.8) ng/ml versus 0.2 (0.07) ng/ml, respectively; mean (SEM), P < or = 0.01). Species of Bacteroïdes and Fusobacterium necrophorum induced IL-8 secretion from cultured mesothelial monolayers to levels comparable to those found in peritoneal fluids in vivo. Heat-killed bacteria and bacterial supernatant were also able to stimulate mesothelium to secrete IL-8. The results suggest that in the early phase of bacterial peritonitis the influx of PMN is regulated by bacteria-induced IL-8 secretion by the mesothelium lining the peritoneal cavity.

Neutrophil adherence to and migration across monolayers of human peritoneal mesothelial cells. The role of mesothelium in the influx of neutrophils during peritonitis.

Increased adherence to and subsequent migration of leukocytes across cultured human peritoneal mesothelial cell monolayers takes place after pretreatment of the mesothelial cells with interleukin-1beta. The contribution of the leukocyte beta2 integrins (CD11/CD18) and the mesothelial adhesion protein intercellular adhesion molecule-1 (ICAM-1) and the role of the cytokines interleukin-8, platelet-activating factor (PAF), and transforming growth factor-beta (TGF-beta) were studied in a three-dimensional model system for neutrophil-mesothelial monolayer interaction. Polymorphonuclear leukocytes (PMNs) showed minimal adherence to and migration across unactivated mesothelial monolayers, despite an extensive amount of ICAM-1 on the mesothelial membrane. Pretreatment of the monolayers with rIL-1beta induced enhanced PMN adherence to the mesothelial monolayer together with a further increase in ICAM-1 expression on the mesothelial membrane. PMN migration was observed across rIL-1beta-activated mesothelial cell (MC) monolayers whenever cytokines secreted by the MCs were present during migration. Monoclonal antibody (mAb) R6.5 against ICAM-1 and mAb CLB-LFA1/1 against CD18 both reduced the migration of PMNs across mesothelial monolayers with a predominant inhibitory effect of CLB-LFA1/1, indicating a significant role of the beta(2) integrins of PMNs in this process. Interleukin-8 was the major cytokine synthesized by the MCs to stimulate the migration of PMNs; both PAF and TGF-beta had a more modest role in our system. Adherence of PMNs to MC monolayers was not dependent on these latter cytokines. Neuraminidase did not have any effect, indicating that selectins were not involved in the adherence process. rIL-1beta-pretreated MCs induced a rapid increase in intracellular Ca2+ in PMNs; actinomycin D blocked this effect and was also able to prevent adhesion of neutrophils to activated MC monolayers. Neutrophil migration across activated cultured MCs is thus a cascade of events in which the MCs are actively involved.

Polarized secretion of interleukin-8 by human mesothelial cells: a role in neutrophil migration.

We investigated the role of human mesothelium in an in vitro model of peritonitis with emphasis on the secretion of the neutrophil chemoattractant interleukin-8 (IL-8) and the migration of polymorphonuclear leucocytes (PMN) across monolayers of peritoneal mesothelial cells. PMN showed minimal migration across non-activated mesothelial monolayers (< 2%). However, migration was induced after mesothelial cell activation by IL-1 beta (24%) and this induced migration was significantly blocked by antibodies against IL-8 (63% inhibition; P < or = 0.01). IL-1 beta-activated mesothelial monolayers were shown to secrete IL-8 in a polarized way, which was preferentially oriented towards the apical side of the monolayer. Our results indicate that the influx of PMN into the peritoneal cavity is, at least in part, controlled by the mesothelial cell layer of the peritoneal membrane.

CA 125 secretion by peritoneal mesothelial cells.

AIMS: To investigate the secretion of the tumour marker CA 125 by cultured human mesothelial cells; to determine if secretion of CA 125 could be observed by activating the mesothelial monolayers with different cytokines. METHODS: Mesothelial cells were isolated from human omentum and cultured to confluent monolayers on perforated polycarbonate membranes (pore size 0.4 micron). The mesothelial monolayers were activated and the apical and basolateral secretion of CA 125 compared. To investigate the influence of cytokines, mesothelial cells were cultured and activated with recombinant interleukin-1 beta (rIL-1 beta), tumour necrosis factor-alpha (TNF-alpha) or lipopolysaccharide from Escherichia coli. The secretion of CA 125 was tested using a microparticle enzyme immunoassay. RESULTS: Mesothelial monolayers secreted CA 125 in a polarised manner with preference for the apical side. Apical polarisation occurred irrespective of the side of the inducing stimulus (p < or = 0.05). Non-activated cultured mesothelial monolayers secreted significant quantities of CA 125, indicating constitutive production of this protein. However, CA 125 production was significantly enhanced if mesothelial cells were incubated with rIL-1 beta (p < or = 0.05), TNF-alpha (p < or = 0.05), and E coli LPS (p < or = 0.01). CONCLUSIONS: Human mesothelial monolayers secrete CA 125 preferentially from their apical surfaces. The secretion of CA 125 can be enhanced by the inflammatory cytokines Il-1 beta, TNF-alpha, and by E coli LPS.

Mesothelial cell adherence to vascular prostheses and their subsequent growth in vitro.

Cell seeding may decrease the thrombogenicity of implanted vascular grafts, but its application is hampered by the limited availability of autologous endothelial cells. Human peritoneal mesothelial cells have blood flow supporting qualities and are readily available. This study investigated the adherence of mesothelial cells to vascular prostheses and their subsequent growth in vitro. Circular pieces of various vascular prosthetic materials were seeded with 51Chromium-labeled mesothelial and endothelial cells and left for either 5, 15, 30, 60, and 120 minutes. The unattached cells were removed and the degree of cell attachment was measured. The number of mesothelial cells to Dacron increased during the first 60 min up to 35.2% of the seeded inoculum whereafter a plateau was reached. Scanning electron microscopy showed spread mesothelial cells adherent to the Dacron fibers. A significant increase in adherence was observed after preincubation of Dacron with 10 micrograms/mL fibronectin, but no improvement was found after preincubation with human serum albumin or gelatin. Mesothelial cells adhered better to Gel-coated than to Gel-sealed or plain Dacron. The adherence of mesothelial cells to ePTFE (Teflon) was significantly poorer. No significant differences in adherence were found between mesothelial and endothelial cells. Mesothelial cell growth on Dacron resulted in a modest increase in the number of viable cells during 27 days, which implies biocompatibility of Dacron and mesothelial cells in vitro.

A cobblestone cell isolated from the human omentum: the mesothelial cell; isolation, identification, and growth characteristics.

Normal human mesothelial cells (NHMC) were isolated from pieces of human omentum. The cell yield was approximately one million cells per square centimeter omentum. The mesothelial cells were identified by their positive staining with monoclonal antibodies against cytokeratins 6 and 18. Transmission electronmicroscopy of cultured NHMC revealed many microvilli on the apical surface and many mitochondria and pinocytotic vesicles in the cytoplasm, indicating active transmembrane transport. Growth of NHMC was directly related to the concentration of human serum or of fetal bovine serum in the growth medium. Addition of epidermal growth factor with or without hydrocortisone resulted in a significant increase of NHMC growth; when endothelial cell growth factor, insulin, or hydrocortisone were added no such increase was observed. Seeding NHMC at densities less than 3000/cm2 did not result in monolayer formation. The mesothelial cells were serially passed in growth medium M199 with added 10% fetal bovine serum up to 7 passages. However, after Passage 4 the cells changed into giant cells with an irregular pattern, and a lack of intracellular cytokeratin expression was observed for most of the cells.

Thrombogenicity and procoagulant activity of human mesothelial cells.

Cell seeding may decrease the thrombogenicity of implanted vascular grafts, but its application is hampered by the limited availability of autologous endothelial cells. We studied the interaction of alternate cells, human peritoneal mesothelial cells, with whole blood in a flow chamber. When citrated blood was perfused over mesothelial cells, platelet adhesion was seen on the intercellular matrix but not on the cells themselves. Perfusions with blood anticoagulated with low-molecular-weight heparin resulted in fibrin formation at the surface of mesothelial cells but not at the surface of human umbilical venous endothelial cells. At shear rates of 200 sec-1 fibrin deposition on the mesothelial cell surface increased during the first 5 minutes to 5.7 +/- 1.06 micrograms fibrin per square centimeter, whereafter these values stabilized. The procoagulant activity of cultured mesothelial cells was higher than that of peritoneal membrane studied ex vivo. However, cultured mesothelial cells incubated with polyclonal antibodies against tissue factor showed a significant decrease in procoagulant activity. We conclude that human peritoneal mesothelial cells may be used for cell seeding procedures, provided that their tissue factor expression can be controlled.

A haemodynamic comparison of epidural versus intrathecal sufentanil to supplement general anaesthesia for abdominal aortic surgery.

The present study was designed to evaluate the influence of epidural sufentanil (ES) and intrathecal sufentanil (IS) on the peri-operative haemodynamic responses during abdominal aortic surgery. Twenty-four ASA Grade II patients without clinical symptoms of coronary artery disease received, randomly, epidural (n = 12) or intrathecal (n = 12) sufentanil combined with light general anaesthesia for elective bifemoral grafting for aorto-iliac occlusive disease. The IS group contained significantly more hypertensive patients than the ES group. This resulted in a significantly higher systolic and mean blood pressure, which remained constant from the start to the end of the study. Following a single bolus injection of 150 micrograms of sufentanil epidurally or intrathecally, there was a significant decrease in heart rate (HR), systolic, mean and diastolic blood pressure, systemic vascular resistance (SVR) and coronary perfusion pressure in both groups. This suggests that IS and ES must be used with caution in patients with cardiovascular disease. The abdominal incision restored the haemodynamic changes produced by sufentanil administration, but these did not exceed pre-sufentanil values. There were no significant changes in filling pressure, cardiac index (CI) and left ventricular work after aortic cross-clamping in the two groups. Revascularization produced significant differences in HR, SVR and CI in both groups in comparison with the pre-declamping period. Notable was the maintenance of systemic blood pressure following revascularization due to preservation of sympathetic activity. It was concluded that both epidural and intrathecal sufentanil produce comparable and stable haemodynamics in this category of patients.

Locating and marking non-palpable, mammographically suspicious breast lesions with the aid of ultrasound.

Most non-palpable, mammographically suspicious breast lesions can be located by ultrasound (47 of 58 lesions in our series). We found that it was possible in 26 of 28 cases to mark these lesions prior to operation under ultrasound guidance thus simplifying the operating procedure.

The non-palpable, radiographically suspicious breast lesion: an analysis of 101 cases.

Analysis of 94 patients with 101 non-palpable, radiographically suspicious, breast lesions revealed 46 malignancies (46%). Comparison of the malignant lesions with 225 palpable carcinomas (221 patients) removed surgically during the same period, showed a higher incidence of carcinoma in situ. Contrary, lymph-node metastases and distant metastases were, at the time of operation, distinctly less than in the patients with non-palpable malignant lesions. A breast-conserving therapy was more often feasible in patients with nonpalpable malignant lesions.

[Therapeutic possibilities for the diabetic and ischemic foot]. / Possibilités thérapeutiques du pied diabétique et ischémique.

The prognosis and treatment of a diabetic foot depend on the extent of the lesions. If infected, these lesions are always mixed, secondary to aerobic and anaerobic bacteroides. The antibiotic treatment must be well adjusted and applied for a long enough time. It is absolutely necessary, for the treatment to be effective, to know the level of the ischemia in order to decide whether a lesion or an amputation scar will heal without revascularization. The author discusses various non-invasive examinations. Today, distal revascularization if possible due to new surgical techniques. But, one must emphasize that the patency of an adequate by-pass is definitely longer than the survival of a diabetic patient, in general. A accurate diagnosis and a multiudiscipline treatment will therefore improve the quality of the survival of the patient.

A study of valve incompetence that developed in an experimental model of venous hypertension.

Surgical treatment of venous valve incompetence should be aimed at restoring the underlying valve defect. For a better definition of the structural anatomical defects in acquired valve incompetence, an experimental study was performed with 40 rats. Venous valves were subjected to hemodynamic stress by creating femoral arteriovenous fistulas. The resulting valve incompetence was studied by using descending phlebography and a casting technique, which allowed for an accurate description of leaking valves when the scanning electron microscope was employed. A three-dimensional insight of the morphology of incompetent venous valves was obtained. A description of short- and long-term (two to four months) changes in valve architecture was also obtained; initially, there was retrograde passage of fluid through a separation of the cusps' free border. The cusps' bulgings were still clearly defined after two months, and, at this stage, commissures had started to widen. After a four-month period, commissures were lost and no recognizable valve sinus was present.