In vitro endothelialization of bioprosthetic heart valves provides a cell monolayer with proliferative capacities and resistance to pulsatile flow.

OBJECTIVES: Degeneration of bioprosthetic heart valves has been suggested to be at least partly an immunogenic reaction toward the xenogeneic tissue. An autologous endothelial lining has been proposed to overcome this problem. We examined in vitro endothelialization of such tissue and retention of endothelial cells after exposure to flow resembling the in vivo situation. METHODS: Cultured human saphenous vein endothelial cells were used to in vitro endothelialize photo-oxidized bioprosthetic heart valves. The endothelialized valves were mounted in a specially designed flow device, creating a pulsatile flow through the valve. Maintenance of a confluent cell layer and deposition of basement membrane markers were determined with immunohistochemical labeling. RESULTS: Labeling of the main components of the basement membrane, laminin and collagen type IV, was verified within 6 hours after in vitro endothelialization. Under static conditions, 4-mm wide denudations were completely re-endothelialized in 4 days, which was similar to the growth rate on gelatin-coated cell culture plastic, which served as a control material. After exposure of endothelialized valves to pulsatile flows for 24 hours (80 beats/min, 3.4 L/min), there were minimal cell losses from the bioprostheses. The cell layer adapted to the pulsatile flow, as verified by rearrangement of morphology and intracellular stress fibers. CONCLUSIONS: This study shows the feasibility of in vitro endothelialization of photo-oxidized bioprosthetic heart valves. The cells are able to withstand a pulsatile flow in vitro, to develop basement membrane-like structures, and to re-endothelialize denuded areas. This technology may be used to enhance the performance of bioprosthetic heart valve prostheses.

A biodegradable bovine collagen membrane as a dermal template for human in vivo wound healing.

A bovine collagen membrane was used as a template for dermal regeneration in human full thickness wounds. Healing was allowed for 7, 21, or 42 days. The formation of neodermis, basement membrane, and terminal differentiation were assessed histologically and immunohistochemically. The collagen template was neovascularised within 7 days, and from day 21 small vessels were detected throughout the transplanted area. The procollagen content decreased whereas the number of fibroblasts increased with time. Collagen type IV was not detected after 7 days but was deposited with time from the wound edges and inwards over the transplanted area. Re-epithelialisation was complete at day 7 and terminal differentiation was similar to normal human skin from day 21. We have shown the time course of dermal and epidermal healing with the aid of a ready-to-use biodegradable collagen membrane. This material may be used as a true dermal template-because of the evidence of dermal regeneration and, in addition, its availability and ease of handling.

Time-course for in vitro development of basement membrane, gap junctions, and repair by adult endothelial cells seeded on precoated ePTFE.

OBJECTIVES: To establish some functional characteristics after in vitro endothelialisation of precoated ePTFE grafts. MATERIALS AND METHODS: Saphenous vein endothelial cells were confluently seeded on human serum-precoated ePTFE. Collagen I-precoated ePTFE served as control. Time-courses for development of laminin, collagen IV and connexin43 was followed-up with immunofluorescence. Endothelial migration and proliferation was studied with cresyl violet staining and tritiated thymidine-labelling. RESULTS: Six hours post-seeding, basement membrane components were visualised subendothelially as thin fibrils. After 24 h, there was a distinct fibril network, which did not seem to be further enhanced. Connexin43 was detected 6 h post-seeding, and the number of intercellular connections appeared to be similar up to day 7. The time-courses were similar on serum- and collagen I-precoated ePTFE. Endothelialisation of a 4 mm wide gap showed migrating fronts largely composed of proliferating cells. The process was completed within 10 days on collagen I-precoated ePTFE, and 15 days on serum-precoated ePTFE. CONCLUSIONS: These findings suggest human endothelial cells, seeded on precoated ePTFE, maintain some differentiated functions seen in vivo. This is of importance for cell retention, functionality and endothelial repair of in vitro endothelialised ePTFE-grafts after implantation in man.

Long-term alleviation of allodynia-like behaviors by intrathecal implantation of bovine chromaffin cells in rats with spinal cord injury.

Adrenal chromaffin cells produce analgesic substances, such as catecholamines and enkephalins, and intrathecal (i.t.) implantation of either allografted adrenal tissue or xenogenic chromaffin cells produce antinociception in animals. We evaluated the analgesic effect of bovine chromaffin cells in a model of central pain in which rats exhibit chronic allodynia-like behavior after photochemically induced ischemic spinal cord injury. Bovine chromaffin cells or endothelial cells were injected i.t. onto the lumbar spinal cord and their effects on mechanical and cold allodynia-like behaviors were studied for up to 8 weeks. The chronic allodynia-like behavior was stable for months without signs of remission and i.t. implantation of human endothelial cells did not alleviate the chronic allodynia-like behavior for the entire observation period. In contrast, 2 weeks after i.t. implantation of bovine chromaffin cells, the mechanical allodynia was abolished in the spinally injured rats, and the enhanced response to cold stimuli was significantly reduced. The overall effects were significant up to 8 weeks after i.t. implantation, although the anti-allodynic effect decreased towards the end of the observation period. No signs of side-effects were noted after i.t. implantation. The allodynia-like state was temporarily restored by naloxone (0.5 mg/kg) or phentolamine (0.3 mg/kg) injected intraperitoneally. Immunohistochemical examination revealed that tyrosine hydroxylase (TH)-positive chromaffin cells could be identified adjacent to the spinal cord up to 4 weeks after i.t. implantation, whereas at 8 weeks the TH-positive cells were sparse. It is concluded that bovine chromaffin cells stay viable in rat spinal cord for a considerable period of time after i.t. administration and alleviate chronic allodynia-like behavior in spinally injured rats, possibly through activation of opioid and alpha-adrenoceptors. The present results further document a new therapeutic approach for the treatment of chronic neuropathic pain.

Immunoisolating encapsulation of intrathecally implanted bovine chromaffin cells prolongs their survival and produces anti-allodynic effect in spinally injured rats.

We have previously reported that intrathecal (i.t.) implantation of bovine chromaffin cells has an anti-allodynic effect in a rat model of mechanical and cold allodynia-like neuropathic pain after spinal cord injury. The technique of encapsulation of the cells by a semipermeable membrane has been developed recently. The present study was undertaken to investigate the effects of encapsulated bovine chromaffin cells on the allodynia-like pain in the same model. Capsules with bovine chromaffin cells or control capsules were implanted in the spinal subarachnoidal space in rats. Their response in behavioural tests were recorded for 2 months. At termination, the capsules were explanted and examined morphologically with tyrosine hydroxylase immunohistochemistry. The mechanical allodynia was totally abolished from week 2 after implantation of the cells and throughout the 8-week test period. The abnormal cold response was also attenuated in about half of the animals. The threshold to acute nociceptive stimulation was not affected. Eight weeks after implantation, 60-80% of the encapsulated chromaffin cells were still tyrosine hydroxylase positive. No effects were observed with control capsules. The results indicate that spinal implantation of encapsulated xenogeneic chromaffin cells may be useful in treating some refractory painful states associated with spinal cord injury. Immunoisolation of chromaffin cells by a semipermeable membrane may inhibit immunorejection, prolong the survival of the cells and enhance their anti-allodynic effect. Copyright 1998 European Federation of Chapters of the International Association for the Study of Pain.

Effects of ropivacaine on eicosanoid release from human granulocytes and endothelial cells in vitro.

OBJECTIVE: To examine the effects of ropivacaine, currently being investigated for treatment of ulcerative colitis, on the release of arachidonic acid metabolites. MATERIAL: Human granulocytes and endothelial cells. TREATMENT: Ropivacaine, lidocaine, hydrocortisone, 5-aminosalicylic acid or acetylsalicylic acid (10-1000 microM). METHODS: Leukotriene B4, 5-hydroxyeicosatetraenoic acid, 6-keto PGF1 alpha and 15-hydroxyeicosatetraenoic acid were measured using immuno assays. Wilcoxon signed rank test was used for statistical calculations. RESULTS: Ropivacaine dose-dependently inhibited zymosan-induced release of leukotriene B4 and 5-hydroxyeicosatetraenoic acid whereas the release after ionophore stimulation was not affected. Ropivacaine was more potent than 5-aminosalicylic acid but less potent compared to hydrocortisone. Ropivacaine had only a weak inhibitory effect on the release of 15-hydroxyeicosatetraenoic acid from zymosan- or ionophore-stimulated cells. In contrast to hydrocortisone and 5-aminosalicylic acid, ropivacaine only weakly affected the release of 6-keto PGF1 alpha after stimulation with thrombin. CONCLUSIONS: The inhibited release of 5-lipoxygenase products may account for some of the anti-inflammatory effects of ropivacaine seen in the treatment of ulcerative colitis.

Characterization and partial purification of a keratinocyte-derived growth factor with wound-healing properties.

We have previously described the mitogenic and wound-healing properties of keratinocyte-conditioned medium (KCM). In this study we investigated the effect of KCM on the activation of second messenger systems and the expression of proto-oncogene in cultured human skin fibroblasts. We also present a partial purification of the factor responsible for the mitogenic and wound-healing effects of KCM. KCM was shown to increase the expression of the proto-oncogenes c-fos, c-myc and c-jun. The effect of KCM on three second messenger systems was investigated. The extracellular release of choline metabolites was increased by 40 per cent when cells were stimulated with KCM whereas the formation of cAMP and hydrolysis of phosphatidyl inositol (PI) was unaffected. KCM was purified by ion exchange chromatography and filtration. The biologically active fraction was eluted from an SP column and retained its activity after filtration through a 3-kDa filter. The fraction was inactivated by heat and acid, indicative of a peptide origin. Furthermore, the active fraction was shown to increase the extracellular release of choline metabolites and to stimulate re-epithelialization in wounds in human skin in vitro comparable to KCM. The study indicates that human keratinocytes produce a < 3 kDa peptide which may be partly responsible for the growth stimulatory and wound-healing properties of KCM.

In vitro endothelialisation of arteriovenous loop grafts for haemodialysis.

OBJECTIVES: To evaluate the feasibility in a pilot study of in vitro endothelialisation of PTFE grafts used as interposition arteriovenous fistulas in uraemic patients. METHODS: Autologous saphenous vein endothelial cells were harvested and cultured on PTFE grafts in seven patients undergoing maintenance haemodialysis. The patients had several previous failures of vascular access sites. The patients were followed with duplex ultrasound, clinical examination and in one case an explanted graft was examined. RESULTS: At the end of follow-up four of the seven patients had patent grafts. One patient occluded the graft immediately postoperatively and another after 3.5 months. The former patient received a second endothelialised graft. In two further patients revision of the outflow was performed. In two patients a functioning graft was excised, in one case because of bleeding of a venous aneurysm and in one case because of suspected infection. The former which was excised 5 weeks postoperatively revealed that 85% of the surface was covered by endothelial cells. CONCLUSIONS: This pilot study shows that in vitro endothelialisation of PTFE grafts used for haemodialysis is possible in uraemic patients. In this highly problematic patient group the results are promising with endothelial cell coverage after 5 weeks of implantation.

Cell adhesion and short-term patency in human endothelium preseeded 1.5-mm polytetrafluoroethylene vascular grafts: an experimental study.

It has been shown that endothelialization improves short-term patency of 1.5-mm expanded polytetrafluoroethylene vascular grafts. A model for endothelialization of 1.5-mm expanded polytetrafluoroethylene vascular grafts with human endothelial cells is described. In this model, the adherence of endothelial cells was increased significantly in grafts coated with serum proteins and collagen. By means of this protocol, graft patency was tested after implantation in two animal models: the rat aorta and the rabbit common carotid artery. Anastomosis was performed with a 3M Precise Microvascular Anastomotic System. In both animal models, no significant loss of endothelial cells in the precoated grafts (rat, n = 8) were noted 1 hour after blood flow restoration. All uncoated grafts showed significant endothelial cell loss. In the rabbit model, all nonendothelialized grafts (n = 8) clotted 5 to 25 minutes after flow restoration. Seven (n = 8) endothelialized grafts showed no clotting during 1 hour's observation: one clotted immediately for a patency rate of 87.5 percent. These results indicate that endothelialization of 1.5-mm grafts is practical. Furthermore, adhesion of endothelial cells to the graft walls is not affected by short-term, pulsatile, high-pressure blood flow.

Dialysis-induced serum factors inhibit adherence of monocytes and granulocytes to adult human endothelial cells.

In patients on hemodialysis, the sequences of events required for adhesion molecule-mediated adherence of monocytes and granulocytes to endothelial cells are pathophysiologically disrupted because activation of the cells with subsequent alterations in adhesion molecule phenotypes takes place in the extracorporeal circuit far away from the respective endothelial cell ligand. As a consequence, the host defense to infections may be affected. We analyzed the expression of CD62L and CD11b/CD18 on monocytes and granulocytes and the adherence of these cells, collected before, during, and after cuprophan hemodialysis, to resting and interleukin-1 beta (IL-1 beta)-stimulated adult human saphenous vein endothelial cells (HSVECs). Monocytes collected before dialysis adhered more avidly to HSVECs than did granulocytes. Adherence of both cell types increased to IL-1 beta-stimulated HSVECs (P < 0.05). Granulocytes obtained at 180 minutes adhered less to resting HSVECs than granulocytes harvested before hemodialysis (P < 0.05), despite an increase in CD11b/CD18 expression. To study if serum factors inhibiting adherence accumulate during hemodialysis, we incubated leukocytes harvested before hemodialysis in the same patients' serum collected before, during, and after dialysis. Serum collected at 180 minutes of cuprophan hemodialysis exerted inhibitory effects on both monocyte and granulocyte adhesion to HSVECs (P < 0.05). By contrast, serum collected during polysulfone hemodialysis of the same patients did not have any significant impact on the adherence of inflammatory cells to HSVECs. These findings contribute to an increased understanding of the factors involved in the impaired immune response observed in dialysis patients.

Characterization of a new in vitro model for studies of reepithelialization in human partial thickness wounds.

Reepithelialization of artificial partial thickness wounds made in biopsies of human skin was determined after 3, 5, or 7 d of incubation, submerged or elevated to the air-liquid interface. The biopsies were reepithelialized within 5-7 d, with a more complete epidermal healing in wounds exposed to air. Both types of wounds showed similar time-course in deposition of basement membrane components, as detected by immunofluorescence labeling. Laminin and collagen type VII were deposited underneath the migrating tips, whereas collagen type IV was detected after reepithelialization. Markers of terminal differentiation showed a pattern close to normal in the air-liquid incubated wounds after reepithelialization. Involucrin was detected in the suprabasal regions of the migrating epidermis and thereafter in the upper half of neo-epidermis in the air-liquid incubated wound. Filaggrin could not be detected in the submerged wounds at any time during healing, whereas wounds exposed to air showed a well-differentiated epidermis by Day 7. Tritiated thymidine-incorporation indicated proliferation of epidermal and dermal cells during reepithelialization and a maintained viability, as shown by cultivation of endothelial- and fibroblast-like cells obtained from the dermis 7 d after wounding. Reepithelialization in this human in vitro model is supported by a matrix close to normal with the possibility of extracellular influences and cell-cell interactions and, in addition, the technique is simple and reproducible. Therefore, we suggest this model for studies of regeneration in culture and as a complement to in vivo studies on epidermal healing.

Combinatorial expression patterns of the connexins 26, 32, and 43 during development, homeostasis, and regeneration of rat teeth.

Gap junctions permit the exchange of regulatory molecules between cells and play important roles during organogenesis. The expression pattern of the gap junction proteins connexin 26, 32, and 43 was studied by immunohistochemistry in the developing, adult, and injured rat teeth. Connexins 32 and 43, but not the connexin 26, were detected during the late stages of embryonic tooth development (bell stage). Expression of connexin 32 was predominant in epithelial cells, whereas connexin 43 was more widely distributed and found in both epithelial and mesenchymal cells. During cytodifferentiation (early postnatal stages), both connexin 32 and 43 were expressed in the epithelial-derived ameloblasts, synthesizing and secreting the enamel matrix proteins. In mesenchyme, connexin 32 was observed only in differentiating odontoblasts, while connexin 43 was expressed in both differentiating and functional odontoblasts, which secrete the dentin matrix. In adult rat teeth, connexin 26 and 43 were expressed in the odontoblastic layer at low and high levels, respectively, while connexin 32 was absent from odontoblasts. Electron microscopy showed that connexin 43 was distributed exclusively at sites of contacts between odontoblasts. However, double immunostaining combined with confocal microscopy suggested an occasional overlap between odontoblasts and calcitonin gene-related peptide-positive nerve fibers. Denervation experiments showed that the expression of connexins in dental pulp was independent of innervation, whereas in injured teeth connexin 43 was upregulated in pulpal fibroblasts. Finally, cultured dental epithelial cells expressed both connexin 32 and 43, and connexin 43 was detected in cultured pulp fibroblasts in vitro, thus mimicking the in vivo distribution pattern of connexins. These results demonstrate that connexins are involved in tooth development and suggest that a given connexin may have distinct roles during odontogenesis and tooth homeostasis.

Cultured human endothelial cells seeded on expanded polytetrafluoroethylene support thrombin-mediated activation of protein C.

PURPOSE: Reduction of the thrombogenicity of synthetic vascular grafts by endothelialization has been suggested. The purpose of this study was to examine some of the non-thrombogenic properties of cultured adult human great saphenous vein endothelial cells seeded on expanded polytetrafluoroethylene (ePTFE) grafts. METHODS: Endothelialized grafts, control grafts, and wells were incubated with thrombin. Assays of thrombin loss from solution, thrombin coagulant activity, and protein C activation on the surface were obtained. The presence of thrombomodulin was confirmed with immunohistochemistry. RESULTS: Significantly more thrombin was found on the ePTFE grafts or wells that underwent endothelialization, and larger amounts were lost from the thrombin solution compared with the control group. Thrombin enzyme activity on the endothelialization group was almost completely represented by activation of protein C and only to a minor extent by activity towards fibrinogen. CONCLUSIONS: It is concluded that endothelial cells seeded on ePTFE retain the possibility to inhibit thrombin coagulant activity and to activate protein C. These findings provide support for the clinical applicability of cultured autologous endothelium on ePTFE grafts.

Secretion of prostacyclin, tissue plasminogen activator and its inhibitor by cultured adult human endothelial cells grown on different matrices.

OBJECTIVES: The aim of this study was to investigate the secretion of prostacyclin (PGI2), tissue plasminogen activator (tPA) and plasminogen activator inhibitor type 1 (PAI- 1) produced by human great saphenous vein endothelial cells (HSVECs) in vitro when seeded to confluency on different matrices. METHODS: The matrices compared were: uncoated plastic, gelatin, collagen type-I gel, fibronectin, fibrin glue, de-endothelialized porcine aorta and ePTFE vascular grafts pre-coated with collagen type-I or human serum. RESULTS: There were no significant differences between basal production of PGI2 on the different matrices. The HSVECs responded with a significant increase in PGI2 secretion after stimulation with thrombin on all matrices, including the ePTFE grafts. A statistically significant higher secretion of tPA secretion was found in supernatants from cells cultured on collagen type-I gel. Interestingly, tPA secretion was lower by cells seeded on both collagen type-I and serum precoated ePTFE as compared to collagen type-1 gel. PAI-1 secretion however was significantly higher on collagen type-1 gel, gelatin, fibrin glue and porcine aorta but not on pre-coated ePTFE grafts. CONCLUSIONS: The findings emphasise the important effect of the matrix on endothelial secretion of PGI2, tPA and PAI-1. Differences in patency after implantation of in vitro endothelialized grafts, may be due to cellular function depending on the types of graft and matrix chosen.

Effects of in vitro hyperthermia on proliferative responses and lymphocyte activity.

Fever is induced by both exogenous products like endotoxin, and endogenous cytokines, most notably IL-1 and IL-6, and tumour necrosis factor (TNF). These mediators are believed to interact with the hypothalamus, to induce enhanced body temperature. However, little is known about the biological effects of fever on the function of the immune system. We here report that a 90-min pulse of mild hyperthermia (40 degrees C) induces enhanced proliferation of peripheral blood mononuclear cells (PBMC). This proliferative response was completely inhibited by antibodies to MHC class II, which had no effect on mitogen-induced proliferation of PBMC. The enzyme-linked immunospot (ELISPOT) assay is a sensitive method for detection of single cells secreting antibodies or cytokines. A 90-min pulse of mild hyperthermia (40 degrees C) induced a significantly enhanced immunoglobulin production in PBMC, as determined by ELISPOT, indicating B cell activation. The T cell cytokine pattern both with and without stimulation with hyperthermia differed between individuals. Enhanced interferon-gamma (IFN-gamma) secretion was noted at 39-41 degrees C. This IFN-gamma response was inhibited by antibodies to MHC class II and thus was MHC class II-restricted and dependent on antigen-presenting cells. None of the individuals tested showed IL-4 response after stimulation with hyperthermia. These findings favour the notion that fever may play an important role in immune responses, and it is possible that fever may act as a physiological adjuvant, with effects on the immune system both in infection and inflammation of other origins.

Effects of neuropeptides on growth of cultivated rat molar pulp fibroblasts.

The effect of the neuropeptides substance P (SP), neurokinin A (NKA), calcitonin gene-related peptide (CGRP), neuropeptide Y (NPY) and vasoactive intestinal polypeptide (VIP) on DNA synthesis of dental pulp cells was investigated in cells grown from molar tooth bud explants from 4-6 days old rat pups. A concentration response-assay of the proliferative response of pulpal cells was performed with SP, NPY, NKA, CGRP and VIP (0.01 to 1 nM) in the presence of EGF (10 ng/ml), hydrocortisone (0.4 microgram/ml) and 3% FCS, using [3H]thymidine incorporation. The results showed that SP, NKA and CGRP, but not NPY and VIP, increased the cell number in a concentration-dependent manner, with maxima at 10(-10)-10(-9) M (SP, NKA) and 10(-7) M (CGRP). No potentiating effect was noted when cells were simultaneously stimulated with SP and CGRP. The finding that SP, NKA and CGRP have growth regulatory properties on pulpal cells in vitro suggests that sensory neuropeptides may be involved during pulpal development or in wound healing after pulpal injury.

Effects of sera, basic fibroblast growth factor, heparin and cyclic AMP-stimulation on proliferation of human vascular endothelial cells.

The aim of this study was to find culture conditions that optimize the proliferation of human endothelial cells (ECs). The effects of different sera, growth factors and other additives on ECs derived from the adult human great saphenous or the umbilical vein were studied. Human serum (HS) at 10 and 40% was significantly more effective than fetal calf serum at 10 and 30%, respectively. The addition of basic fibroblast growth factor (bFGF) increased proliferation alone and in combination with heparin. Heparin alone increased EC growth only in medium containing 40% HS. The addition of cholera toxin (CT) and a phosphodiesterase inhibitor (IBMX) to raise cAMP-levels, stimulated proliferation of both cell types but was more pronounced on the ECs from the saphenous vein. The cAMP-levels were elevated equally in both cell types. However, db-cAMP stimulated proliferation only of the ECs from the saphenous vein. An additive stimulatory effect was observed when bFGF and CT/IBMX were combined. For saphenous vein ECs, a medium containing HS (40%), bFGF, heparin, CT and IBMX was found optimal for proliferation. We conclude that these compounds may be used to increase EC growth and that, even a limited number of donor ECs may be sufficient starting material for in vitro studies on the human endothelium.

Keratinocyte conditioned medium stimulates type IV collagenase synthesis in cultured human keratinocytes and fibroblasts.

We have previously shown that conditioned medium from cultured human keratinocytes stimulates proliferation of a variety of cell types involved in wound healing, as well as re-epithelialization of wounds in human skin in vitro. We now present evidence for an autocrine/paracrine control of the synthesis of type IV collagenases in human keratinocytes and fibroblasts. During wound healing, keratinocytes migrate over the wound bed, an activity coupled with lysis of basement membranes, and hence requiring the presence of collagenases. Collagenases are also needed for the production and remodelling of the granulation tissue. In order to study the autocrine/paracrine control of collagenase production in keratinocytes and fibroblasts, we stimulated these cells in culture with conditioned medium from cultured keratinocytes. Protease synthesis was determined by affinity labelling with 3H-diisopropylfluorophosphoridate (DFP) and by zymography. Keratinocyte-conditioned medium was found to increase the expression of 72 and 92 kDa type IV collagenase in human keratinocytes, and the 72 kDa collagenase in human fibroblasts, indicating that an autocrine/paracrine control mechanism is involved in collagenase production in these cell types during wound healing. This increased expression of collagenases could be partly responsible for the stimulated healing seen in wounds treated with sheets of cultured keratinocytes.

Reduction of monocyte adhesion to xenogenic tissue by endothelialization: an adhesion molecule and time-dependent mechanism.

Great interest has been shown for the seeding of autologous endothelial cells on prosthetic materials. We investigated the inflammatory and immunogenic properties of xenogeneic tissue before and after seeding with cultured human great saphenous vein endothelial cells in vitro. Adhesion of monocytes to xenogeneic tissue with or without endothelium and the endothelial cell expression of E-selectin, intercellular adhesion molecule 1, vascular adhesion molecule 1, and major histocompatibility complex class II antigens were investigated 1, 3, and 7 days after seeding. Both monocyte adhesion and endothelial adhesion molecule expression were relatively high 1 day after seeding and were significantly lowered after 3 to 7 days. There was no difference between monocyte adhesion and adhesion molecule expression on viable or nonviable xenogeneic tissue. Monocyte adhesion and adhesion molecule expression increased after interleukin-1 beta or interferon-gamma stimulation of the endothelial cells. The results suggest that human endothelial cells exhibit an early proinflammatory and immunogenic activity immediately after seeding. Three and 7 days after seeding, the endothelialized surface is less adhesive for monocytes as compared with nonendothelialized tissue. These findings have implications when cultured or intraoperatively recruited endothelial cells are used clinically.