G. Uhlschmid

Development of degradable polyesterurethanes for medical applications: In vitro and in vivo evaluations

To evaluate the biocompatibility of a newly developed degradable class of polyesterurethanes and their possible use as biomaterials, we investigated the cell and tissue interactions with these polymers using a small number of chemical base entities. The polymers were prepared by chain extension with diisocyanates of PHB/HV-diol and either PCL-diol or Diorez, another aliphatic polyester-diol. Regardless of the chemical composition of the four tested polyesterurethanes used as substrates, no morphological difference was observed either in the macrophages (macrophage cell line J774) or in the fibroblasts (fibroblast cell line 3T3) cultured on the polymers. In contrast, however, cell adhesion and growth of macrophages and fibroblasts were affected by the polymer properties. Compared to macrophages cultured on tissue culture polystyrene (TCPS), cells cultured on the test polymers exhibited levels of cell adhesion that varied from 65-100% of TCPS, and the doubling time was 25-43% higher on the polymers than on TCPS. Likewise, fibroblasts adhered to the polymers at lower rates (50-85% of TCPS) and grew at higher doubling times (125-140% of TCPS). Furthermore, cells cultured on the test polymers preserved their phenotypes: fibroblasts produced high amounts (up to 280% of control cells) of collagens Type I and Type IV and fibronectin; and macrophages produced nitric oxide (NO) and tumor necrosis factor alpha (TNF-alpha) in the same concentrations as control cells and responded to lipopolysaccharide treatment by the elevation of the production of NO and TNF-alpha, indicating that the cell-to-polymer interactions allow fibroblasts and macrophages to maintain their phenotypes. In vivo investigations showed that all four test polymers exhibit favorable tissue compatibility. The formed capsule was 60-250 microns thick. In addition, the polymers are degradable. After one years subcutaneous implantation in rats, the molecular weight of the test polymers were reduced to about 50%, depending on the composition. Taken collectively, the present data demonstrate that the newly developed polyesterurethanes are cell and tissue compatible and biodegradable.

New versatile, elastomeric, degradable polymeric materials for medicine.

The present investigation was focused on the cell compatibility of recently developed biodegradable polyesterurethane-foam (DegraPol-foam) to chondrocytes and osteoblasts. Both chondrocytes and osteoblasts, isolated from adult male rats, exhibited relatively high cell adhesion on DegraPol-foam. Scanning electron microscopy (SEM) showed that cells grew on the surface and into the open cell pores of the foam. Morphologically, cells found on the surface of the foam exhibited a flat cell appearance and built a confluent cell multilayer. In contrast, inside the foams cell showed rounded morphology building cell aggregates and cell islets. In addition, chondrocytes and osteoblasts proliferated on the DegraPol-foam and preserved their phenotype for up to 2 weeks. During degradation of these polymers, small crystalline particles of short-chain poly[(R)-3-hydroxybutyric acid] (Mn approximately 2300) (PHB-P) and lysine methyl ester are released. Therefore, lysine methyl ester and PHB-P, as possible degradation products of the polymer, are investigated here for their effects on macrophages and osteoblasts. Results obtained in the present study clearly indicate that macrophages and, to a lesser degree, osteoblasts have the ability to take up (phagocytose) PHB-P. At low concentrations, particles of PHB failed to induce cytotoxic effects or to activate macrophages. Osteoblasts showed only limited PHB-P phagocytosis and no signs of any cellular damage. At high concentrations of PHB-P, the cell viability of macrophages and to a lesser extent of osteoblasts was affected.

Degradable and highly porous polyesterurethane foam as biomaterial: Effects and phagocytosis of degradation products in osteoblasts

Recently, a new class of biodegradable PHB-based polyesterurethane (DegraPol/btc) has been prepared and found to exhibit favorable cell and tissue compatibility. The present study has been designed to evaluate the response of primary isolated rat tibia osteoblasts to small crystalline particles of short-chain poly[(R)-3-hydroxybutyric acid] (PHB-P diameter: 2-20 microm), of fluorescent-labeled analogs (DPHP-P), and of lysine methyl ester as possible degradation products of DegraPol/btc. Observations made using confocal microscopy clearly indicate that osteoblasts have the capability of taking up PHB-P particles. Although in single-cell analysis the number of DPHB-P-positive osteoblasts gradually increased up to 16 days, the fluorescence intensity per osteoblast increased only during the first 4 h after DPHB-P incubation, and then it retained the 4 h level up to 16 days. No significant change in the production levels of collagen type I and osteocalcin was detectable after treatment with low concentrations of PHB-P for up to 32 days. In contrast, a time- and dose-dependent alteration of the alkaline phosphatase (ALP) activity was found. Maximal activity was measured after 4 days of treatment with 2 microg of PHB-P/mL (170% of control cells). Rat peritoneal macrophages co-cultured with osteoblasts in a transwell culture system mimicked the observed PHB-P induced ALP elevation. Therefore, the PHB-P-induced ALP increase could be the result of direct or indirect stimulation of osteoblasts, possibly via soluble factors produced by contaminating osteoclasts. Taken collectively, the data demonstrate that osteoblasts are capable of phagocytosing PHB-P and that this process is accompanied at low PHB-P concentrations by dose- and time-dependent alteration of alkaline phosphatase activity but not of collagen type I or osteocalcin.

Chondrocyte-biocompatibility of DegraPol®-foam: In vitro evaluations

Histological and biochemical investigations were carried out in order to evaluate the chondrocyte compatibility of a recently developed biodegradable polyesterurethane-foam (DegraPol-foam). Therefore, cell adhesion, cell growth, and the preservation of chondrocyte phenotype was measured in rat xyphoid chondrocytes seeded on DegraPol-foam. Chondrocytes, isolated from xyphoids of adult male rats, exhibited relatively high cell adhesion on DegraPol-foam (about 60% of that found on TCPS). Scanning electron microscopy (SEM) showed that chondrocytes grew on the surface and into the open cell pores of the foam. Morphologically, cells found on the surface of the foam exhibited a flat cell appearance and built a confluent cell multilayer. In contrast, the interior of the foam cells showed rounded morphology in cell aggregates and cell islets. In addition, chondrocytes proliferated on the DegraPol-foam (doubling-time of about 12.5 days) and preserved their phenotype for up to 14 days. Compared to freshly isolated chondrocytes, cells seeded on the foam produced high concentrations of collagen type II for up to 2 weeks: the ratio of type II/I collagen was 1.2-1.4 fold higher than the ratio found in freshly isolated cells. No significant difference was observed in chondroitin sulfate levels produced by freshly isolated cells and cells cultured on DegraPol-foam for up to 14 days. To sum up, our results indicate that DegraPol-foam is a compatible substrate for chondrocytes.

Hepatic artery embolisation with a novel radiopaque polymer causes extended liver necrosis in pigs due to occlusion of the concomitant portal vein.

BACKGROUND/AIM In an attempt to overcome some of the problems encountered with the materials available for liver embolisation, we investigated a novel radiopaque polymer of the polyurethane family (Degra-Bloc). METHODS Hepatic artery embolisation of one liver lobe using polyurethane was performed in 19 healthy pigs. Microcirculatory changes were assessed by laser Doppler flowmetry. Radiological and pathological examinations of the livers, hearts and lungs removed provided information about the extent and effect of the embolisation. RESULTS None of the pigs died due to hepatic failure or toxicity of polyurethane. Microcirculation of embolised liver lobes significantly decreased from 106 (+/-15) perfusion units (PU) to 45 (+/-6) PU immediately after embolisation and further to 28 (+/-7) PU before euthanasia. At this time conventional and angiographic X-ray controls demonstrated the radiopaque casts extending up to the peripheral arteries with signs of degradation over time but without formation of collateral vessels. The main pathological findings consisted of destruction of the portal tract structures and also of large areas of liver necrosis. Polyurethane was encountered in arterioles as small as 10-20 microm, but not in liver sinusoids, hearts or lungs. CONCLUSIONS The novel polymer called DegraBloc is a biocompatible, slowly degradable, radiopaque embolic agent. The occlusion of the arterial tree up to the smallest arteriolar diameter combined with concomitant portal vein occlusion leads to sharp segmental necrosis in pig livers without formation of significant collaterals and without systemic embolism. In the treatment of liver tumours polyurethane might provide a promising alternative to conventional embolic materials, provided that it is used with care in patients with advanced liver cirrhosis.

Magnetic resonance-guided laparoscopic interstitial laser therapy of the liver

BACKGROUND There is a necessity for an imaging method during laparoscopy to get a three-dimensional access to the target. In this study we evaluated laparoscopic interstitial laser therapy of the liver under magnetic resonance imaging guidance. METHODS Five domestic pigs underwent laparoscopy in an open-configuration magnetic resonance system. Under simultaneous real-time magnetic resonance imaging interstitial laser therapy was applied to the liver. Magnetic resonance images, macroscopic aspects of the lesions, and light microscopic findings were compared. RESULTS The interventions could be safely performed. There was no image artifact caused by instruments or by the carbon dioxide. Dynamic gadolinium-enhanced imaging proved to significantly predict the macroscopic volume of the laser lesions. CONCLUSIONS Magnetic resonance-guided laparoscopic interstitial laser therapy of the liver combines the advantages of minimal invasive surgery and magnetic resonance imaging. Further development should focus on laparoscopic instruments and temperature sensitive sequences.

Morphological and Functional Changes in Canine Kidneys following Extracorporeal Shock-Wave Treatment

Extracorporeal shock-wave lithotripsy (ESWL) has rapidly become established worldwide as a routine method for treatment of nephro- and ureterolithiasis. Although initial studies showed no tissue-damaging effect by the shock waves, we found, in an animal experiment using canine kidneys, that the ESWL-induced damage to the renal parenchyma is more marked than originally assumed. The damage is limited to the area that was focused on, and heals relatively rapidly by connective tissue encapsulation with final cicatrisation without any further residual effects being observed up to the present. This parenchymal damage is probably also the cause of the macrohematuria that is always observed during therapy. The resulting tissue damage is not extensive enough to cause a demonstrable reduction of function as measured by the usual methods (serum creatinine, creatinine clearance, isotopy renography, i.v. urography). In serum we observed a transient decrease of calcium, an immediate increase of lactate-dehydrogenase, transaminases (SGOT and SGPT) and a delayed increase of alkaline phosphates. Creatinine, blood urea nitrogen, sodium, potassium and amylase remained within normal limits. In urine, a decrease of creatinine and an increase of glucose excretion were noted. We believe that these changes represent a relatively mild and transient damage of renal cells and do not reflect the occasionally heavy morphological changes observed after shock-wave exposure. The main clinical complication is the large subcapsular hematoma which, according to the present knowledge, could well result from a lesion of the larger peripheral vessels. Damage to other organs such as subserous colonic and small bowel hematomata are to be expected although they do not lead to clinical symptoms.

Phagocytosis and biodegradation of short-chain poly [(R)-3-hydroxybutyric acid] particles in macrophage cell line

The biocompatibility and the degradation mechanisms of block co-polyurethanes containing crystallizable telechelic poly [(R)-3-hydroxybutyric acid] (PHB) segments have found recently growing interest for the possible biomedical applications of these new materials. The random hydrolytic cleavage of the amorphous part of these polymers might result, in vivo, in the production of small crystalline particles of low molecular weight PHB that could then undergo phagocytosis and biodegradation inside phagosomes. To test this possibility, a fluorescent-labelled PHB segment was synthesized, precipitated in the form of crystalline particles, and used for an in vitro investigation of its interaction with macrophage cell line. Light and fluorescence microscopy performed in the present study clearly show that the fluorescent particles are well internalized in phagosomes already after 1 h of incubation. The number of phagocytized particles decreases notably after 8 days of incubation.A quantitative determination of the time dependence of the phagocytosis was obtained through laser cytometry and fluorescence-activated cell sorting (FACS). Fluorescence spectroscopy and high-performance liquid chromatography (HPLC) analysis of extracts of cell supernatants revealed the presence of supposed degradation products of PHB after 8 days of incubation, suggesting that macrophages could degrade low molecular weight PHB.

Urologische Komplikationen nach Nierentransplantation

Between December 1964 and April 1974, 216 renal transplants were performed on 200 recipients at the University Clinic of the Cantonal Hospital at Zurich. The ureteroneocystostomy was performed in most

In vivo effects of angiotensin antagonists on plasma aldosterone in the dog.

The effects of infusions of equimolar doses of Sar1-Ile8-angiotensin II and of Sar1-Ala8-angiotensin II on plasma aldosterone, plasma renin activity and arterial blood pressure were compared in normal dogs. Plasma aldosterone increased significantly after Sar1-Ile8-angiotensin II whereas it was unaffected by Sar1-Ala8-angiotensin II. Changes in blood pressure and renin activity were small without marked differences between both groups of animals. The experiments demonstrate a clear steroidogenic potency of Sar1-Ile8-angiotensin II. Therefore, Sar1-Ala8-angiotensin II should be preferred as antagonist of the action of angiotensin II in the adrenal gland.