Biomedical subjects

J Van Marle

Publications and source records attributed to J Van Marle.

At least 19 recordsLinked to original sources

PubMed2000

Coating of coverslips with glow-discharged carbon promotes cell attachment and spreading probably due to carboxylic groups.

BACKGROUND: For high-resolution microscopy, cells have to be analyzed through thin glass coverslips. Therefore, it is necessary to culture cells on coverslips for preservation of cell morphology. We found cell attachment and spreading to be relatively slow processes, even when cells were plated on coated coverslips. This slowness presents a problem, particularly when synchronized cell populations are used. METHODS: In this paper, we present a method that is based on glow-discharged carbon coating of coverslips which promotes rapid attachment and spreading of cells, enabling rapid analysis of cells after plating. Results obtained with carbon-coated coverslips were compared with those of other types of coating. Two fibroblast lines, an epithelial cell line, and a carcinoma cell line were tested. RESULTS AND CONCLUSIONS: All cell lines showed a rapid adhesion on carbon-coated coverslips. With fibroblasts we found the carbon coating to be superior to other coatings tested, mainly because the carbon did not influence cell morphology. Using synchronized or irradiated cells produced similar results. The superior performance of carbon coating is probably due to carboxylic groups on the glow-discharged carbon layer. The carbon layer does not interfere with microscopy or immunocytochemical staining procedures.

Animals↗

PubMed2000

Higher numbers of autologous fibroblasts in an artificial dermal substitute improve tissue regeneration and modulate scar tissue formation.

Cultured skin substitutes are increasingly important for the treatment of burns and chronic wounds. The role of fibroblast numbers present in a living-skin equivalent is at present unknown. The quality of dermal tissue regeneration was therefore investigated in relation to the number of autologous fibroblasts seeded in dermal substitutes, transplanted instantaneously or precultured for 10 days in the substitute. A full-thickness porcine wound model was used to compare acellular dermal substitutes (ADS) with dermal substitutes seeded with fibroblasts at two densities, 1x10(5) (0-DS10) and 5x10(5) cells/cm(2) (0-DS50), and with dermal substitutes seeded 10 days before operation at the same densities (10-DS10 and 10-DS50) (n=7 for each group, five pigs). After transplantation of the dermal substitutes, split-skin mesh grafts were applied on top. Wound healing was evaluated blind for 6 weeks. Cosmetic appearance was evaluated and wound contraction was measured by planimetry. The wound biopsies taken after 3 weeks were stained for myofibroblasts (alpha-smooth muscle actin), and after 6 weeks for scar tissue formation (collagen bundles organized in parallel and the absence of elastin staining). Collagen maturation was investigated with polarized light. For wound cosmetic parameters, the 10-DS50 and 0-DS50 treatments scored significantly better than the ADS treatment, as did the 10-DS50 treatment for wound contraction (p<0.05, paired t-test). Three weeks after wounding, the area with myofibroblasts in the granulation tissue, determined by image analysis, was significantly smaller for 0-DS50, 10-DS10, and 10-DS50 than for the ADS treatment (p<0.04, paired t-test). After 6 weeks, the wounds treated with 0-DS50, 0-DS10, and 10-DS50 had significantly less scar tissue and significantly more mature collagen bundles in the regenerated dermis. This improvement of wound healing was correlated with the higher numbers of fibroblasts present in the dermal substitute at the moment of transplantation. In conclusion, dermal regeneration of experimental full-skin defects was significantly improved by treatment with dermal substitutes containing high numbers of (precultured) autologous fibroblasts.

Animals↗

PubMed1998

Heterogeneous suppression of experimentally induced colon cancer metastasis in rat liver lobes by inhibition of extracellular cathepsin B.

Metastatic rat colon cancer cells but not normal rat hepatocytes showed activity of cathepsin B on their plasma membranes. Activity was visualized in living cells with a new fluorogenic substrate, [Z-Arg]2-cresyl violet, and confocal microscopy. When these cancer cells were injected into the portal vein of rats, the animals developed tumors in the liver in a heterogeneous fashion. Three- to four-fold more tumors were found in the small caudate lobe than in the other three large lobes of the liver. Oral treatment with a selective water-soluble inhibitor of extracellular cathepsin B, Mu-Phe-homoPhe-fluoromethylketone, resulted in 60% reduction of the number of tumors and 80% reduction of the volume of tumors in the three large lobes whereas tumor development was not affected in the small caudate lobe. This study supports the conclusions that (a) extracellular cathepsin B plays a crucial but complex role in liver colonisation by rat colon carcinoma cells in vivo, (b) its selective inhibition suppresses tumor growth heterogeneously in the liver and (c) the caudate lobe of the liver is a relatively large risk factor for tumor development.

Amino Acid Chloromethyl Ketones↗

PubMed1997

The contribution of quantitative confocal laser scanning microscopy in cartilage research: chondrocyte insulin-like growth factor-1 receptors in health and pathology.

The use of immunohistochemical detection techniques and fluorescent molecular probes in light and fluorescence microscopy allows accurate and specific analysis of a great variety of cell and tissue components. However, when staining yields only low intensity levels, serious problems may arise with discrimination of specific signals against background staining. This problem is often inherent with articular cartilage research. Application of confocal laser scanning microscopy (CLSM) can circumvent these problems. The CLSM collects images that are almost free of out-of-focus signals, which results in improved spatial resolution and discrimination as compared with conventional microscopy. Moreover, CLSM allows optical sectioning of specimens and three-dimensional reconstruction of the microscopical object. Quantitative evaluation of microscopical images is hampered by out-of-focus signals because they interfere with specific signals in the image. Interference of these nonspecific signals can be diminished by application of CLSM; in CLSM only one single point in microscopical objects is illuminated at any time and this point is then imaged into the pinhole at the entrance of the photo-detector and subsequently digitized. The present review is a discussion of the present state of the art in digital imaging with the use of CLSM in cartilage research. This discussion includes aspects such as sensitivity, specificity, spatial resolution and accuracy of quantitative analysis in microscopical immunofluorescent objects.

Animals↗

PubMed1997

Morphology and function of preserved microvascular arterial grafts: an experimental study in rats.

The aim of this study is to examine the morphology and function and small-caliber, arterial grafts after preservation in the University of Wisconsin solution (UW). Rat carotid arteries were stored in UW (n = 10) or in phosphate-buffered saline (PBS) (n = 10) for 1, 3, 7, and 14 days and were examined with light microscopy (LM) and scanning electron microscopy (SEM). Rat aortic preparations were stored in UW or PBS for 1 hour, 24 hours, 72 hours, 7 days, and 14 days and assessed for functional responses (stimulated contraction and endothelium-dependent relaxation). Segments (5 mm) of rat carotid arteries were stored in UW or PBS for 3 days, 7 days, and 14 days and orthotopically implanted as autografts and allografts. No immunosuppressive or anticoagulant agents were used. After 28 days of implantation, the grafts were assessed for patency and excised for LM and SEM. In UW, the endothelial layer remained intact up to 9 days of storage. In PBS, the endothelial layer showed deterioration after 1 day and was completely lost after 3 days. Functional responses were demonstrated to exist for as long as 7 days storage in UW. In PBS, no responses could be evoked after 24 hours storage. Autografts preserved in UW for 3 days (n = 6), 7 days (n = 6), and 14 days (n = 6) showed patency rates of 83.3%, 66.6%, and 66.6%, respectively, whereas patency rates of allografts were 66.6%, 33.3%, and 33.3%, respectively. Autografts stored in PBS for 3 days (n = 6), 7 days (n = 6), and 14 days (n = 6) showed patency rates of 33.3%, 33.3%, and 50%, respectively, whereas patency rates of allografts were 16.7%, 0%, and 33.3%, respectively. The UW preserved autografts showed normal morphology. All other groups showed vessel wall degeneration which in the allograft groups, were accompanied by lymphocellular infiltration. In conclusion, the endothelial layer and vessel wall of arteries are adequately preserved in UW. Functional responses are retained up to 14 days storage in UW, but, are lost after 24 hours storage in PBS. Autograft implantation studies accordingly show good performance of arterial segments preserved in UW, whereas allografts are subject to degradation as a result of rejection.

Adenosine↗

PubMed1997

Importance of the profunda femoral artery in distal limb revascularisation.

The profunda femoral artery (PFA) was recognised in the early 1960s as an essential artery to maintain perfusion in the lower limb. The aim of this retrospective study was to analyse the results obtained with aortobifemoral bypass surgery (ABF) where the distal perfusion was solely dependent on the PFA. An evaluation was made of 240 ABFs done between January 1988 and May 1995. This represented a combination of operations done by either the vascular unit at the HF Verwoerd teaching hospital or by Dr Van Marle privately. In 56 cases only the PFA was available for distal anastomoses in one or both limbs, giving a total of 80 PFA anastomoses. Evaluation was based on pre- and postoperative ankle brachial pulse indexes (ABPI). There were 3 deaths and 1 amputation in the early postoperative period. Over an average follow-up period of 32 months (1 month-63 months) 76 limbs were assessed. At initial presentation 30% of the patients had rest pain and 2 already had gangrenous changes. Mean pre-operative ABPI was 0.5. A further femoral-popliteal bypass was required within 1 year for 6 of the patients. The average postoperative ABPI was 0.86, giving a 73.2% average improvement. We therefore concluded that good results can be achieved when the PFA is used for distal anastomosis in ABF bypass surgery.

Adult↗

PubMed1996

Experimental arterial allografting under low and therapeutic dosages of cyclosporine for immunosuppression.

The aim of this study was to investigate performance of preserved arterial allografts under the protection of a high-dose and a low-dose immunosuppressive regimen, with cyclosporine (CsA). Dog carotid arteries were harvested and stored for 14 days at 4 degrees C in University of Wisconsin organ preservation solution. Segments (6 cm) of carotid artery were orthotopically and bilaterally implanted in mongrel dogs (n = 18). CsA was given in two dosage regimens: 25 mg/kg/day (group I, n = 7) and 10 mg/kg/day (group II, n = 7). The control group received no CsA (group III, n=4). After 3 months of implantation, patency was assessed by angiography. The grafts were excised for investigation of vessel wall and endothelial function and morphology. For assessment of function in vitro, slices of arterial segments were connected as ring preparations to an isometric force transducer and immersed in a 5 ml organ bath (37 degrees C) containing Tyrode's solution. The contractile response was examined by adding 40 mM KCl and phenylephrine (100 microM) to the organ bath; endothelium-dependent relaxation was examined by adding methacholine (100 microM). Morphology was assessed semiquantitatively. The functional responses to KCl, phenylephrine (Phe) and methacho- line (Met) after 14 days of storage in UW, were 30.2 +/- 1.2 mN, 26.9 +/- 1.0 and 45 +/- 1.2% (means +/- SEM, n=9), respectively. Patency after three months of implantation for group I was 100% (14/14), for group II 50% (7/14), and for group III 75% (6/8). In vitro functional responses of preserved arteries, after 3 months of implantation in group I were 58.5 +/- 10.6 mN (KCl), 36.5 +/- 5.8 mN (Phe), and 57.4 +/- 9.7% (Met), respectively. Functions in group II were 1.2 +/- 0.1 mN (KCl, 0.0 mN (Phe), and 0.0% (Met). Grafts in group III showed no function. Measurement of medial thickness showed significant thinning (P <0.05) in groups II and III. Patency and function of arterial allografts under a therapeutic dose of CsA were superior to grafts implanted under low-dose CsA or no immunosuppressive treatment.

Animals↗

PubMed1996

Three-point repositioning of axes: three-dimensional alignment procedure for electron microscope tomography using three markers.

A description is given of a new procedure to align series of tilted graphs, made with an electron microscope, for computer tomographic purposes. The procedure uses the coordinates of three projected markers to calculate parameters needed for the reconstruction. To that end the procedure computes the direction of the tilt-axis, the translation and rotation parameters, the tilt-angle of every micrograph, and the spatial coordinates of the individual markers with their centre of gravity as origin of the coordinate system. A searching technique, based on cross-correlation, is described to locate accurately the micrographs markers.

Animals↗

PubMed1996

Articular cartilage destruction in experimental inflammatory arthritis: insulin-like growth factor-1 regulation of proteoglycan metabolism in chondrocytes.

Rheumatoid arthritis, a disease of unknown aetiology, is characterized by joint inflammation and, in its later stages, cartilage destruction. Inflammatory mediators may exert not only suppression of matrix synthesis but also cartilage degradation, which eventually leads to severe cartilage depletion. Systemically and locally produced growth factors and hormones regulate cartilage metabolism. Alterations in levels of these factors or in their activity can influence the pathogenesis of articular cartilage destruction in arthritic joints. The main topic of the present review is the role of the anabolic factor insulin-like growth factor-1 in the regulation of chondrocyte metabolic functions in normal and in diseased cartilage. This is the most important growth factor that balances chondrocytes proteoglycan synthesis and catabolism to maintain a functional cartilage matrix. A brief overview of how chondrocytes keep the cartilage matrix intact, and how catabolic and anabolic factors are thought to be involved in pathological cartilage destruction precedes the review of the role of this growth factor in proteoglycan metabolism in cartilage.

Arthritis, Rheumatoid↗

PubMed1996

Histochemical analysis of insulin-like growth factor-1 binding sites in mouse normal and experimentally induced arthritic articular cartilage.

Insulin-like growth factor-1 (IGF-1) plays a key role in regulation of chondrocyte metabolism. We examined the localization of IGF-1 binding sites on chondrocytes in cartilage from normal and experimentally induced arthritic mouse knee joints. Cryostat sections from patellar cartilage were incubated either with IGF-1 receptor antibody or biotinylated IGF-1. Subsequently confocal laser scanning microscopy was applied to compare the two staining procedures qualitatively and quantitatively. This approach allowed detailed analysis of membrane-associated and intracellular staining. Using IGF-1 receptor antibody, IGF-1 receptors were found on the cell membrane of chondrocytes in the middle and deeper cartilage zones, whereas intracellular staining was highest in chondrocytes of superficial zones. After incubation with biotinylated IGF-1, distinct membrane staining was not present and fluorescence was localized homogeneously in the middle and deeper zones but not in superficial zones. In cartilage from inflamed knee joints staining with the use of IGF-1 receptor antibody did not change significantly, whereas a pronounced increase in staining was noted with biotinylated IGF-1 in chondrocytes of the middle and deeper zones of cartilage. It is concluded that the staining patterns obtained with the use of IGF-1 receptor antibody and biotinylated IGF-1 are remarkably different, suggesting that the latter also detects IGF-binding proteins. The results suggest that joint inflammation has no consistent effect on IGF-1 receptor expression but may induce a significant upregulation of IGF-binding proteins in chondrocytes of the middle and deeper zones of cartilage.

Animals↗

PubMed1996

Ultrastructural localization of activity of phosphatases by low temperature incubation of unfixed cryostat sections.

In the present study, we demonstrate the activity of several phosphatases ultrastructurally in long-term (up to 24 months) cold-stored (-80 degrees C) rat tissues. Phosphatase activity was histochemically studied with the use of unfixed cryostat sections in combination with low temperature (4 degrees C) incubation conditions in order to prevent inactivation of enzyme activity and to limit the loss of ultrastructure. 5'-Nucleotidase activity was observed at plasma membranes, mainly at bile canalicular membranes of hepatocytes in liver. Thiamine pyrophosphatase activity was detected not only in trans side cisternae but also in medial and cis side cisternae of Golgi complexes in the parotid gland. Glucose-6-phosphatase activity was localized in endoplasmic reticulum as well as at the outer membrane of the nuclear envelope. Acid phosphatase reaction product was found in lysosomes. Furthermore, the localization patterns of 5'-nucleotidase and thiamine pyrophosphatase activity were compared with those obtained after different fixation procedures such as immediate chemical fixation of tissues or fixation of tissues after freezing and thawing. The results showed similar localization patterns of these enzymes after the different pretreatments. However, with respect to the ultrastructural morphology, some damage was observed in unfixed material after incubation. It can be concluded that the procedure described here enables ultrastructural localization of activity of phosphatases in long-term cold-stored tissues. This procedure will be useful for a retrospective study on archival material when histochemical parameters are needed.

5'-Nucleotidase↗

PubMed1996

Disturbed structural interactions between microfilaments and tight junctions in rat hepatocytes during extrahepatic cholestasis induced by common bile duct ligation.

Microfilaments in epithelial cells are important for the structural and functional integrity of tight junctions. In the present study, we examined the relationship between microfilaments and tight junctions in hepatocytes of rat liver following common bile duct ligation (CBDL) for up to 2 weeks. Actin filaments and tight junctions were studied by fluorescence microscopy using 7-nitrobenzene-2-oxa-1,3-diazole phallacidin (NBD-ph) and an anti-ZO-1 antibody, respectively. Double-stained sections were examined with confocal laser scanning microscopy (CLSM). Electron microscopy was applied for the assessment of structural alterations in microfilaments and in tight junctions with detergent-extraction and freeze-fracture preparations. Our results showed that F-actin was present at the entire plasma membrane of hepatocytes in control liver, whereas CBDL increased the amount of F-actin mainly at the bile canalicular and lateral plasma membranes. Simultaneously, the immunofluorescence of ZO-1 underwent striking changes, i.e., from a uniform to an irregular staining pattern with various fluorescence intensities. CLSM demonstrated a colocalization of ZO-1 and F-actin in control liver and its deterioration in CBDL liver. Electron microscopy showed marked alterations of microfilaments and tight junctions due to CBDL. It is concluded that actin filaments are intimately associated with tight junctions in normal hepatocytes. CBDL impairs this association by progressively diminishing the structural interaction between F-actin and ZO-1, which may in turn lead to functional disturbances of tight junctions.

Actin Cytoskeleton↗

PubMed1996

Otoconial alterations after embryonic development in hypergravity.

The relation between prolonged hypergravity and structural adaptation of otoconia was studied in hamsters (n = 56). Three groups of hamsters (n = 27), were conceived and born in a centrifuge: group 1 (n = 10) 1 month under 2.5 G, group 2 (n = 9) 5 months under 2.5 G and 4 months under 1 G, group 3 (n = 8) 1 month under 2.5 G and 8 months under 1 G. Control hamsters (n = 29) were conceived and born under 1 G (1 month old, n = 7; 9 months old, n = 22). Histological study of the otoconial layers (energy dispersive x-ray element analysis and scanning electron microscopy) showed similar calcium content, size, and shape in utricular and saccular otoconia in all groups. Different were the utricular otoconial size classes, large, medium-sized, and small. The area with small otoconia increased in group 1 (p = 0.002). In group 2, the large otoconial area decreased (p = 0.001) and the medium-sized one increased (p < 0.001). In group 3, the large otoconial area decreased (p = 0.003) and the medium-sized one increased (p = 0.007). For age-related effects we found group 1 with an increased area of large otoconia (p = 0.001) and a decreased medium-sized one compared to groups 2 (p < 0.001) and 3 (p = 0.02). Hypergravity during formation of otoconia does not affect calcium content, size, or shape, but changes relative size of the areas with large, medium-sized, or small otoconia and the development of these areas. This resulted in a structural adaptation to hypergravity.

Animals↗

PubMed1996

Function of cryopreserved arterial allografts under immunosuppressive protection with cyclosporine A.

PURPOSE: Cryopreserved arterial allografts may be used for arterial reconstructive procedures. In this experimental study cryopreserved arteries were used as autografts and as allografts with or without immunosuppression with cyclosporine A. METHODS: In group A (three dogs, six bilateral grafts) cryopreserved carotid artery autografts were implanted. In groups B and C female mongrel dogs (three dogs and six bilateral grafts in each group) received cryopreserved male carotid artery allografts. Dogs in group C were treated with cyclosporine A (25 mg/kg/day). After 3 months of implantation patency was assessed by angiography. Contractile responses to KCl and phenylephrine (Phe) and the endothelium-dependent relaxation response to methacholine (Met) were examined in segments of the grafts after excision. Medial thickness was assessed semiquantitatively. The grafts were stained for sex chromatin analysis to determine the origin of cells in allografts. RESULTS: Patency: group A, 100% (6 of 6), group B, 66.6% (4 of 6), and group C, 100% (6 of 6). Functional responses: before implantation, after thawing, 2.7 +/- 0.5 mN (KCl), 4.8 +/- 1.0 mN (Phe), and 0.0% +/- 0.0% (Met), group A, 36.9 +/- 10.6 mN (KCl), 31.5 +/- 14.4 mN (Phe), and 59.3% +/- 20.4% (Met), group B, 0 for all agents used, group C, 34.0 +/- 7.5 mN (KCl), 28.8 +/- 7.0 mN (Phe), and 46.2% +/- 3.2% (Met). Morphologic characteristics: the media of grafts in group B showed significant thinning (p < 0.05). Smooth-muscle cells in vessel walls of grafts in group C were of female origin. CONCLUSION: Arteries showed no function and loss of endothelial integrity after cryopreservation and thawing. After 3 months of implantation cryopreserved arterial autografts and allografts under immunosuppressive treatment with cyclosporine A showed 100% patency and return of functional responses resulting from repopulation of grafts by host cells.

Animals↗

PubMed1996

Characterization of the cholesterol crystallization-promoting low-density particle isolated from human bile.

BACKGROUND & AIMS: Biliary concanavalin A-binding glycoprotein (CABG) contains cholesterol crystallization-promoting activity that is not accounted for by the pronucleators that have been characterized in this fraction. The aim of this study was to isolate and characterize the missing activity. METHODS: Biliary glycoprotein was isolated using concanavalin A-Sepharose. Promoting activity in CABG was purified using density gradient ultracentrifugation. RESULTS: Activity in CABG separated into two fractions at low (1.08) and high (1.29) density, which showed different crystallization kinetics in a model bile. The high-density fraction had a late onset time (49.2 +/- 17.8 hours) but a high crystal growth rate (13.4 +/- 5.2 micrograms. mL-1.h-1). The low-density fraction had a rapid onset time (33.9 +/- 20.9 hours) but a slower growth rate (6.5 +/- 3.8 micrograms.mL-1 .h-1). The high-density fraction was not further characterized in this study. The low-density fraction contained solid particles consisting of lipid and very little protein, and the activity was fully pronase resistant. Delipidation of the low-density fraction removed all activity. CONCLUSIONS: A potent pronase-resistant nucleation-promoting activity was activated from human bile and characterized. The low-density fraction may be responsible for the rapid nucleation in bile from typical patients with fast-nucleating gallstones.

Adult↗

PubMed1996

Redistribution of Ca2+, Mg2+-ATPase activity in relation to alterations of the cytoskeleton and tight junctions in hepatocytes of cholestatic rat liver.

Ca2+, Mg2+-ATPase is a membrane-bound enzyme localized at the bile canalicular membranes of hepatocytes. Cytoskeleton and tight junctions are important for maintenance of the polar distribution of plasma membrane proteins. In order to understand the mechanisms involved in the redistribution of Ca2+, Mg2+-ATPase due to cholestasis, the relationship between Ca2+, Mg2+-ATPase, microfilaments and tight junctions was examined. Cholestasis was induced in rat liver by common bile duct ligation (CBDL) for 2 weeks. Localization of Ca2+, Mg2+-ATPase activity was studied at the light and electron microscopic level. Double-staining of the enzyme and F-actin was performed using phase-contrast and fluorescence microscopy of 7-nitrobenzene-2-oxa-1,3-diazole phallacidin (NBD-ph), respectively. Immunofluorescence microscopy of ZO-1 was applied for the observation of tight junctions. Furthermore, cytoskeleton and junctional complexes were investigated electron microscopically in saponin-extracted tissues. The results showed that CBDL induced redistribution of Ca2+, Mg2+-ATPase activity from the apical to the entire plasma membrane of hepatocytes, which seemed to occur independently of F-actin. F-actin was present at all membrane domains of hepatocytes in control liver, whereas CBDL increased the amounts of F-actin mainly at the bile canalicular membranes. An inverse distribution pattern of Ca2+, Mg2+-ATPase activity and F-actin was found in epithelial cells of bile ducts in control and cholestatic livers. Marked alterations in microfilaments were observed at the bile canaliculi, which were defined as hypertrophy and atrophy and were in association with changes in tight junctions. Structural impairment of the tight junctions was proven by disordered immunofluorescence of ZO-1. It is concluded that changes in the distribution of Ca2+, Mg2+-ATPase and F-actin due to CBDL are independent of each other. CBDL-induced disorders of microfilaments are related to impairment of structural integrity of tight junctions that is suggested to be responsible for the redistribution of Ca2+, Mg2+-ATPase in hepatocytes.

Actins↗

PubMed1995

Morphology and function of dog arterial grafts preserved in UW-solution.

OBJECTIVES: To assess the function of arterial grafts after prolonged preservation in the University of Wisconsin solution (UW), in vitro and in vivo. METHODS: Carotid arteries were harvested from dogs and stored for 1-21 days at 4 degrees C in UW (n = 10) or in PBS (0.9% NaCl, pH 7.4), (PBS) (n = 10). Slices were examined by lightmicroscopy (LM) and scanning electron microscopy (SEM). For viability testing, specimens were connected to an isometric force transducer (2 x n = 9). Contractile and relaxation responses were examined by adding phenylephrine (200 microM) and metacholine (200 microM), respectively. For in vivo studies (n = 41), 2.5cm carotid artery segments were implanted or orthotopically, as autografts and allografts, after 14 days of storage in UW or in PBS. Autologous veins were used as controls. After 28 days or 56 days, arteriography was performed and the grafts were excised for LM and SEM. RESULTS: The arterial endothelial layer remained intact after up to 14 days of storage in UW. In PBS, the endothelium was lost after 3 days. The functional response after 14 days storage in UW was approximately 50% vs. 0% after 14 days in PBS. In the autografts, total patencies (28 days + 56 days) were 100% (8/8) and 63% (5/8) for UW and PBS stored grafts, respectively. In the allografts, the UW and PBS preserved grafts showed total patencies of 86% (12/14) and 83% (5/6), respectively. Microscopically, the allografts showed fibrotic degeneration. CONCLUSIONS: Arteries are well preserved in UW up to 14 days of storage. Arterial autografts preserved in UW showed good patency and better integrity of the vessel wall after implantation, than grafts stored in PBS or allografts (without immunosuppressive therapy).

Adenosine↗

PubMed1995

Reduced wound contraction and scar formation in punch biopsy wounds. Native collagen dermal substitutes. A clinical study.

In full-thickness skin wounds dermal regeneration usually fails, resulting in scar formation and wound contraction. We studied dermal regeneration by implantation of collagenous matrices in a human punch biopsy wound model. Matrices were made of native bovine collagen I fibres, and either hyaluronic acid, fibronectin, or elastin was added. Matrices were placed in 6-mm punch biopsy holes in seven patients (biopsies were used for the grafting of leg ulcers), and covered with a protective semi-permeable polyether urethane membrane. Histology, wound contraction and dermal architecture were studied. Dermal architecture was evaluated using a recently developed laser scatter technique. All collagen matrices showed a tendency to reduce wound contraction, compared with control wounds; elastin- and fibronectin-treated matrices showed significantly less contraction than control wounds. Only the addition of elastin had a clear beneficial effect on dermal architecture; collagen bundles were more randomly organized, compared with control wounds, and wounds treated with collagen matrices coated with fibronectin or hyaluronic acid, or without coating. We conclude that the punch biopsy wound model provides important information on dermal regeneration in humans. Native collagen matrices with elastin contributed to dermal regeneration and reduced wound contraction, in contrast with matrices coated with fibronectin or hyaluronic acid, or without coating. Future clinical studies of large-area, full-thickness wounds will be required to establish their clinical relevance for leg ulcer and burn treatment.

Aged↗