Gelatin-poly(lactic-co-glycolic acid) scaffolds with oriented pore channel architecture - From in vitro to in vivo testing.

A gelatin-poly(lactic-co-glycolic acid), PLGA, composite scaffold, featuring a highly oriented pore channel structure, was developed as a template for articular cartilage regeneration. As a design principle the composite scaffold was optimized to contain only medical grade educts and accordingly no chemical cross linking agents or other toxicological relevant substances or methods were used. Scaffolds were synthesized using a freeze structuring method combined with an electrochemical process followed by freeze-drying. Finally, cross linking was performed using dehydrothermal treatment, which was simultaneously used for sterilization purposes. These composite scaffolds were analyzed in regard to structural and biomechanical properties, and to their degradation behavior. Furthermore, cell culture performance was tested using chondrocytes originated from joint articular cartilage tissue from 6 to 10 months old domestic pigs. Finally, the scaffolds were tested for tissue biocompatibility and their ability for tissue integration in a rat model. The scaffolds showed both excellent functional performance and high biocompatibility in vitro and in vivo. We expect that these gelatin-PLGA scaffolds can effectively support chondrogenesis in vivo demonstrating great potential for the use in cartilage defect treatment.

Effect of a silver device in preventing catheter-related infections in peritoneal dialysis patients: silver ring prophylaxis at the catheter exit study.

Catheter-related infections remain a significant cause of method failure in chronic peritoneal dialysis (PD) therapy. Given the increasing antibiotic resistance, such nonpharmacological strategies as local silver devices attract more interest. To establish whether a silver ring device (designed by Grosse-Siestrup in 1992) mounted onto the PD catheter and placed at the exit site at skin level is effective in preventing exit-site and other catheter-related infections, a prospective 12-month, multicenter, controlled study stratified by diabetes status was conducted. The study subjects were assessed by an extensive structured inventory, including a broad spectrum of control variables, such as age, body mass index (BMI), Staphylococcus aureus carrier status, catheter features, mode and quality of PD therapy, comorbidity, and psychosocial rehabilitation. Ten experienced German outpatient dialysis centers (seven adult, three pediatric) participated in the trial. All eligible patients (n=195) from the study area without catheter-related infections during the ascertainment period were included (incidental subjects undergoing PD therapy for at least 3 months). The main outcome measures were the occurrence of first exit-site infections (primary study end point), sinus tract/tunnel infection, and peritonitis. Ninety-seven patients were assigned to the silver ring and 98 patients to the control group. Baseline characteristics of age, sex, proportion of pediatric and incidental patients, S aureus carrier status, and other variables were similar in both groups. The incidence of infections in the silver ring group versus the control group was as follows: 23 of 97 versus 16 of 98 patients had exit-site infections, 12 of 97 versus 12 of 98 patients had sinus tract/tunnel infections, 16 of 97 versus 18 of 98 patients had peritonitis, respectively. Kaplan-Meier analysis for the probability of an infection-free interval showed no statistical difference (log-rank test) between the two groups. Displacement of the silver ring contributed to study termination in 6% of the study group patients, including two patients with catheter loss. Univariate analysis and multiple logistic regression identified younger age (<50 years), low serum albumin level (<35 g/L), number of previously placed PD catheters, short cuff-exit distance (<2 cm), and S aureus nasal carriage as risk factors for the development of exit-site infections. In conclusion, our study does not show any benefit of the silver ring in preventing catheter-related infections in PD patients. Thus, prevention of infection-related method failure in PD still has to rely on conventional antibiotic treatment strategies and less so on alternative methods.

Histologic evaluation of a natural permanent percutaneous structure and clinical percutaneous devices.

The longevity of percutaneous devices (PD) is often hampered by exit-site infection. The babyrussa tusk, the only permanent natural percutaneous structure, was histologically studied and the implant-tissue interface of 11 continuous ambulatory peritoneal dialysis (CAPD) catheters that had been implanted in humans was evaluated histologically. Attachment of the epidermis to the tusk surface was observed. All the CAPD catheters showed varying degrees of epidermal downgrowth. These results demonstrate that the epidermis is capable of adhering closely to a percutaneous structure. Regarding the CAPD catheters, no steady state at the epidermis-implant interface was achieved, so the risk of infection still remains.

Biocompatibility of silver-coated polyurethane catheters and silver-coated Dacron material.

The local effects of silver-coated polyurethane catheters and Dacron material were compared to uncoated polyurethane catheters and Dacron material in a long-term implantation test using rabbits. The tissue-implant interaction was analysed by investigating the type and number of inflammatory cells, capillaries, fat tissue, the extent of fibrosis, thickness of the fibrous capsule, number and distribution of silver particles, and the size of giant cells. Silver-coated and uncoated materials displayed comparable signs of inflammation and tissue reaction.

The isolated perfused liver. a new model using autologous blood and porcine slaughterhouse organs.

INTRODUCTION: Current models of isolated and perfused livers are limited by nonphysiologic perfusates or the need for the use of high numbers of laboratory animals. The present study was performed in order to rectify these difficulties. METHODS: To establish a new isolated perfused liver model, a perfusion circuit was developed using normothermic, autologous hemoperfusion and organs obtained from a slaughterhouse. RESULTS: Stable organ function was maintained over 220 min. The organs displayed physiologic values for measured variables, including oxygen consumption which varied from 5.2+/-1.5 ml/min at 40 min to 5.2+/-2.4 ml/min at 220 min, and bile production (0.15-0.31 ml/min, respectively). DISCUSSION: The present studies demonstrate a new approach for experimental liver perfusion by combining the optimal perfusion medium of autologous blood and slaughterhouse organs as source material.

Totally implantable catheter system in the dog.

Four totally implantable catheter systems for arterial and venous access were implanted for a maximum of 17 months. Hematological and bacteriological tests were performed. The platelets remained normal. Occasionally observed bacteremias were successfully treated. The morphological and SEM results show that in comparison to the standard externalized catheter the totally implantable catheter system has a longer functional lifetime and less problems with sepsis while the well-being of the experimental animal is improved.

A new method to evaluate the CAPD-catheter-exit and other percutaneous devices.

A method that has proved to be the most sensitive approach for the early diagnosis of infection of the oral mucosa around dental implants has been used to monitor the exit-sites of percutaneous devices. For three months the state of the skin-catheter interface of 13 CAPD-patients was examined every four weeks. This examination included the measurement of pocket depth with the aid of a periodontal probe, the measurement of SFFR (sulcus-fluid-flow-rate) using standardized filter paper strips and a bacteriological swab of the exit-site. The results obtained demonstrated that SFFR can be measured around percutaneous implants. The measurement of SFFR appeared to be more useful in the monitoring of exit-sites of percutaneous devices than the measurement of pocket depth or the use of bacteriological swabs. Further studies for prolonged periods are necessary to determine whether measurement of SFFR can reliably predict onset of exit-site infection before clinical signs become evident.

Isolated hemoperfused heart model of slaughterhouse pigs.

A model of hemoperfused slaughterhouse pighearts is described providing a wide range of applications which leads to a reduction in animal experiments. The size of a pigheart, heart rate, coronary perfusion, metabolism, etc. are more comparable to conditions in patients than those in hearts of small laboratory animals. Global heart function can be assessed either by measuring stroke volume, ejection fraction, Emax etc. in the working model or by measuring intraventricular pressure with balloon catheters in the isovolumetric model. Regional cardiac function can be measured by sonomicrometry and ischemic and non-ischemic areas can be compared. Local metabolic changes are measurable as well with microdialysis. Cardiac function can be kept on any given functional level by infusion of norepinephrine in spite of the fact that functional parameters are lower without adrenergic drive in vitro than in vivo. Stable heart function can be maintained for several hours with only 500 to 1000 ml of blood because the blood is permanently regenerated by a special dialysis system. This model can be applied in many research projects dealing with reperfusion injuries, inotropic, antiarrhythmic or arrhythmogenic effects of certain drugs, immunological rejection, evaluation of imaging systems (NMR, echocardiography etc.) or cardiac assist devices.

Influence of differences in body weight and volume management on experimental results in porcine models.

BACKGROUND: In contrast to humans, young pigs naturally have a low COPpl (12-16 mmHg versus 22-26 mmHg in young humans). Thus, behavior occurring when volume management is performed similar to human medicine might be different. Potentially underestimated intra- to extravascular fluid and solute shifts could influence time course of variables investigated. That is why we studied whether differences in the basic protocol for infusion therapy and different levels of COP according to age or body weight, respectively - impair reproducibility and reliability of experimental results. METHODS: Group A [n=6, lower body weight (LBW < 22 kg)] was treated with unrestricted infusion rates (UIR) adjusted to maintain a constant blood pressure; group B [n=6, higher body weight (HBW > 28 kg)] was treated with a restricted continuous infusion rate for fluid balance (RIR); group C (n=6) combined HBW and UIR protocol. Blood pressure, plasma solutes, diuresis, and peritoneal fluid were analysed. Statistical analysis was performed using Kruskal-Wallis test and Wilcoxon test. RESULTS: UIR-treated pigs with LBW (group A) developed ascites and demonstrated time dependent decreases of plasma solute concentrations whereas in pigs of group C mainly diuresis was increased and subcutaneous edema occurred. None of the protocols enabled constant blood pressure. DISCUSSION: In young pigs (LBW), an adaptive volume substitution using crystalloids to standardize blood pressure may induce fluid extravasation; in turn data may not show statistical significance, stable hemodynamics may not be achieved and changes in plasma solute concentrations may lead to false interpretations. RIR-strategy provided reproducible, plausible results and thus should be recommended in combination with pigs >29 kg BW for the use of porcine models.

Validation of bone conversion in osteoconductive and osteoinductive bone substitutes.

Bone reconstruction can be performed with an autogeneic graft from various donor regions. Osteoconductive and osteoinductive bone substitutes originate from substances of diverse chemical and morphological types and can have a synthetic or a biological derivation. Alongside autogeneic bone transplants and allogenic and xenogeneic bone implants, alloplastic bone replacements of synthetic or semi-synthetic origin are being used for defect reconstruction. In an animal model in rabbits five bone substitutes and one autogeneic graft were surgically incorporated into identical bone defects (10times 10 mm in size) in six anatomically defined regions of the skull. With scintigraphic and histological methods, the metabolic dynamics of the bone is examined as it reacts to the transplantation of autogeneic bone or to implanted bone replacement material.The different autogeneic, xenogeneic and alloplastic bone replacement materials can be differentiated according to the functional quality of the new tissue and the dynamics of the bone conversion thus induced. In the comparison of mineralized, osteoconductive bone subsitutes (TCP, HA, calcium carbonate ceramics) with demineralized, osteoinductive implants (DBM new, DBM old) and autogeneic bone grafts, the bone inducing matrices show the largest quantity of new bone formation, making possible a volume-constant reconstruction.

[A new method of ex vivo whole blood perfusion of isolated mammalian organs, exemplified by the kidney of swine]. / Eine neue Methode zur Ex-vivo-Vollblut-Perfusion isolierter Warmblüterorgane, dargestellt an der Niere von Schweinen.

A new method for the ex vivo perfusion of organs from large mammals is described. Gas exchange and dialysis are carried out simultaneously with a low-flux polysulfon dialysis module. The dialysate (e.g. Tyrode solution) is aerated with a mixture of oxygen and carbon dioxide to ensure gas exchange with the blood. Dialysis is carried out in a closed thermostatically controlled system. Monitoring of ultrafiltration is maintained by continuously weighing the blood reservoir and adjusting an afferent and efferent blood pump. Initial results obtained with isolated pig kidneys demonstrate the suitability of the new method for use as a model for the replacement of animal experiments. Theoretically, clinical application in the area of in vivo regional organ perfusion may also be possible.

Dependence of renal blood flow on renal artery stenosis measured using CT angiography.

PURPOSE: The present study investigates the suitability of computed tomography angiography (CTA) depicting the degree of renal artery stenosis for estimating renal blood flow (RBF) in a kidney. MATERIALS AND METHODS: We investigated renal artery stenosis assessment by CTA in eight adult female hybrid pigs with an ultrasound probe implanted at the renal vein for RBF measurement. An inflatable metal-free cuff was placed around the renal artery to control the RBF. The RBF was then reduced in four steps. For each reduced RBF value and baseline RBF, CTA with a reconstructed slice thickness of 0.625 mm was performed in the arterial phase following injection of 80 ml of nonionic intravenous contrast medium. The radius of the stenotic and non-stenotic renal artery segment was measured in the reconstructed images. RESULTS: A significant linear correlation (p < 0.0001) was found between the relative apparent stenosis (calculated as the ratio of the radii of the actual stenotic segment and a non-stenotic renal artery segment) and RBF. The linear regression yielded a slope of 0.57 and a y-axis of 24.1 %. A significant linear correlation (p < 0.0001) was also found between the relative true stenosis (the ratio of the radii of the actual stenotic segment and a non-stenotic renal artery segment at baseline) and the RBF. The linear regression yielded a slope of 0.67 and a y-axis of 13.8 %. CONCLUSION: The results show that the relative stenosis apparent on CTA differs from the true degree of renal artery stenosis. Nevertheless, the degree of renal artery stenosis determined by CTA provides a reliable estimate of the resulting RBF reduction.

Thermography as potential real-time technique to assess changes in flow distribution in hemofiltration.

Flow distributions are critical determinants in the function of hemofilters. Despite their importance, however, flow distributions cannot currently be measured in filters during experimental or clinical applications. Here, we demonstrate that the thermal conduction properties of extracorporeal circuits may provide a tool to overcome this limitation. More specifically, we show that thermography provides an indirect approach to visualize differences in regional perfusion rates through temperature profiles on the filter surface. Thermograms were recorded using a TVS700 system (Ca. Goratec) during recirculating in vitro hemofiltration of porcine blood. Different test protocols were executed to characterize the contribution of thermal conduction and convection to the measurable changes in the temperature at the surface of the filter housing. For comparison and validation, these experiments were supplemented by computer tomography (CT) of filters after dye injection. Thermography enabled real-time visualization of the flow distributions in a hemofilter. Moreover, 'point' trends taken from different regions of the filter provided quantitative information about changes of flow distributions in response to changing experimental conditions. Our preliminary data suggest that thermography is a promising new approach for assessing the principles and time-related changes in flow distributions in hemofiltration. As expected, resolution is lower than that in CT measurements and further studies will be necessary to determine the smallest temperature gradient that still identifies differences in regional perfusion rates. Given its potential to develop into an inexpensive tool for the 'bedside' level monitoring of flow distributions during clinical studies, further investigation of thermography is highly desirable.

Design criteria for percutaneous devices.

A theoretical model of a percutaneous device is described. The model is split into several components, each with its own function. Special structures such as horns, hair, feathers, fingernails, hoofs , teeth, and antlers are taken as examples where nature has solved the problems of "percutaneous devices." These structures have been regarded in relation to dimensional and structural differences of epidermis, dermis, and subcutis. Theoretical guidelines are described for the design of a prosthetic percutaneous device.

A computer-assisted in-vitro biomaterial test for percutaneous devices using human keratinocyte cultures.

The aim of this study was to assess the suitability of a computer-assisted in vitro test system to evaluate candidate biomaterials used for percutaneous devices. Silicone rubber (Silastic) and five different polymers from the Eurobiomat concerted action, polyetherurethane, polyvinylchloride with plasticizer, di-ethylhexylphthalate (PVC-DEHP), polyvinylchloride with plasticizer, tri-ethylhexyltrimellitate (PVC-TEHTM), polyethylene and polypropylene were examined with respect to their qualities to facilitate keratinocyte attachment. HaCaT-cells, a spontaneously transformed non-tumorigenic human keratinocyte cell line, were cultured on the different materials for 3 days. Cellular behaviour was examined morphologically by phase-contrast and scanning electron microscopy throughout the 72 h incubation period. For the computer-assisted measurement of the cell-covered substrate surface and subsequent statistical analysis the cells were fixed after 3 days of incubation, stained, photographed and the images then digitally transformed. Of the different polymers examined silicone rubber showed the most favourable results with respect to the quantitative analysis of the cell-covered substrate surface as well as concerning cytomorphological findings. The results of this study indicate that the in vitro test system described is a sensitive screening method for evaluating candidate biomaterials used for percutaneous devices in a fast and reproducible manner, thus reducing number of animal experiments.