Large molecules as anti-adhesive compounds against pathogens.

Anti-adhesive compounds are potential prophylactic tools in alternative treatment regimes against bacterial infection, as bacterial adhesion is commonly mediated by carbohydrate-protein interactions between surface adhesions of microorganisms and the host cell. The use of exogenous polyvalent, high-molecular carbohydrates and tannin-like plant-derived compounds should antagonize the adhesive interaction. A range of carbohydrates and carbohydrate- and proanthocyanidin-enriched plant extracts were screened for potential anti-adhesive effects against Helicobacter pylori, Campylobacter jejuni, Porphyromonas gingivalis and Candida albicans in different in-situ assays on primary tissue. The adhesion of H. pylori on human stomach tissue was effectively blocked by glucuronic acid-enriched polysaccharides from immature okra fruits (Abelmoschus esculentus). These compounds also had strong in-vitro effects against C. jejuni (inhibition up to 80%), but were ineffective in an in-vivo study in infected chicken broilers due to metabolism in the gastrointestinal system. Polysaccharides from Glycyrrhizia glabra, also enriched with glucuronic acid, showed strong anti-adhesive properties against H. pylori and P. gingivalis (inhibition 60-70%). Pelargonium sidoides extract, containing mainly polymeric proanthocyanidins, was effective against H. pylori in a dose-dependent manner. Due to the multifunctional adhesive strategy of C. albicans, no effective compounds were detected against this yeast. Structure-activity relationships are presented and the potential in-vivo use of carbohydrate-based anti-adhesives is discussed.

Reaction of rat pulp tissue to Carisolv 'new gel'--a histocytological evaluation.

BACKGROUND: This study was an histological examination of pulp tissue exposed to Carisolv 'new gel' after 1 to 28 days. METHODS: An occlusal cavity was prepared in 64 caries-free molar teeth of 16 Wistar rats. The roofs of the pulp chambers were perforated and Carisolv 'new gel' solution was placed onto the exposed pulps of 32 molar teeth for 20 minutes. Thirty-two contralateral molar teeth served as controls and were coated with an inert liquid containing isotonic saline solution and carmellose for 20 minutes as well. The pulps of all teeth were capped with Ca(OH)2 and the cavities were filled with a flowable composite in combination with a self-etching dentine adhesive. The animals were sacrificed after 1, 3, 7 and 28 days. Eight teeth per group and the time period were histologically examined, scored, and statistically evaluated (Wilcoxon-test). RESULTS: The results showed no statistically significant differences between the Carisolv group and the control group (p > 0.05). The observed pulp reaction was essentially the same as those reported in the past being typical for the effect of calcium hydroxide as a direct pulp capping agent. CONCLUSION: Compared to Ca(OH)2, Carisolv 'new gel' did not cause any different or additional pulp reaction in healthy teeth.

Electrical stimulation influences mineral formation of osteoblast-like cells in vitro.

The aim of the present study was to assess the structure of newly formed mineral crystals after electrical stimulation of osteoblast-like cells in vitro. Pulsed electrical stimulation was coupled capacitively or semi-capacitively to primary osteoblast-like cells derived from bovine metacarpals. Computer calculations revealed that the chosen input signal (saw-tooth, 100 V, 63 ms width, 16 Hz repetition rate) generated a short pulsed voltage drop of 100 microV (capacitive coupled mode) and of 350 microV (semi-capacitive coupled mode) across the cell-matrix layer. Stimulated cultures showed an enhanced mineral formation compared to the non stimulated controls. In cultures exposed to capacitively coupled electric fields and in control cultures nodules and mineralized globules were found. Nodules with a diameter of less than 200 nm covered the cell surface, whereas mineral globules with a diameter of up to 700 nm formed characteristic mineral deposits in the vicinity of the cells similar to biomineral formations occurring in mineralizing tissues. In contrast, large rod-shaped crystals were found in cultures stimulated by semi-capacitive coupled electric fields, indicating a non-physiological precipitation process. In conclusion, osteoblasts in culture are sensitive to electrical stimulation resulting in an enhancement of the biomineralization process.

Magnesium in newly formed dentin mineral of rat incisor.

Small amounts of magnesium are always detectable in addition to calcium and phosphorus in mineralized tissues such as dentin or bone. Magnesium has been considered to influence the mineralization process, especially crystal growth. The present study reports on the location and enrichment of magnesium in the newly mineralized dentin by using the high lateral resolution of energy dispersive X-ray microanalysis combined with scanning transmission electron microscopy. To this end, we have used the continuously growing rat incisor as a model for a collagenous mineralizing system. Dental tissue was dissected free and cryofixed in liquid nitrogen-cooled propane. The distribution of elements was measured in freeze-dried ultrathin cryosections. The magnesium distribution of the newly formed dentin area near the predentin area was found to be inhomogeneous. In certain small dentin areas, characteristical magnesium enrichments were observed. Further, high magnesium-to-phosphate molar ratios were found in these areas, and these were correlated with low calcium-to-phosphate molar ratios. Our results support the theory that magnesium is involved in the process of biological apatite crystal formation.

Ultrastructural characterization of the implant/bone interface of immediately loaded dental implants.

Primary stability and an optimized load transfer are assumed to account for an undisturbed osseointegration process of implants. Immediate loaded newly designed titanium dental implants inserted in the mandible of minipigs were used for the characterization of the interfacial area between the implant surface and the surrounding bone tissue during the early healing phase. Histological and electron microscopical studies were performed from implant containing bone specimens. Two different load regimens were applied to investigate the load related tissue reaction. Histological and electron microscopical analysis revealed a direct bone apposition on the implant surfaces, as well as the attachment of cells and matrix proteins in the early loading phase. A striking finding of the ultrastructural immunocytochemical investigations was the synthesis and deposition of bone related proteins (osteonectin, fibronectin, fibronectin receptor) by osteoblasts from day one of bone/biomaterial interaction. Calcium-phosphate needle-like crystallites were newly synthesized in a time-related manner directly at the titanium surface. No difference in the ultrastructural appearance of the interface was found between the two loading groups. Our experimental data suggest that loading of specially designed implants can be performed immediately after insertion without disturbing the biological osseointegration process.

Ultrastructural findings after intraarticular application of hyaluronan in a canine model of arthropathy.

We investigated the effect of intraarticularly applied hyaluronic acid (hyaluronan) on the cartilaginous structure of experimentally induced chondromalacia patellae in dogs. For the induction of chondromalacia, we used the Pond-Nuki technique, which involved severance and resection of the anterior cruciate ligament, as a canine model of arthropathy in 27 foxhounds (three groups of nine animals each). In a pilot study, we evaluated the effect of resection of the anterior cruciate ligament with no therapy. Patellar specimens were retrieved at 3, 6, and 12 weeks postoperatively. Subsequently, we compared a treatment group that received intraarticular injections of hyaluronan with a placebo group that received saline solution. The groups were compared at 3, 6, and 12 weeks postoperatively. Three animals from the treatment and placebo groups received five injections of hyaluronan during one of the 4-week intervals (weeks 3-6, 6-9, or 12-15). Specimens were retrieved 5 weeks after the last injection. In both groups, the uninvolved contralateral knee served as a control. The specimens were taken from the medial and lateral patellar poles. Histological analysis included light microscopy and transmission electron microscopy. The structural and ultrastructural changes were assessed qualitatively and were quantified with use of a modified Mankin score. Our results indicate that chondromalacia patellae may be induced with the Pond-Nuki technique. We found a significant reduction (p < 0.01) of cartilaginous lesions in the hyaluronan group compared with the placebo group. Our results suggest that intraarticularly applied hyaluronan is effective in delaying the degenerative process of cartilage degradation. Therefore, we conclude that the use of hyaluronan may be indicated during the early stages of chondromalacia.

Achilles tendon healing: long-term biomechanical effects of postoperative mobilization and immobilization in a new mouse model.

The aim of the study was to investigate the long-term effects of postoperative immobilization as opposed to mobilization on the biomechanical attributes of healing Achilles tendons in a new experimental mouse model. In 114 Balb-C-mice the left Achilles tendon was transected and sutured by the Kirchmayr-Kessler technique. The tendons healed either under postoperative immobilization effected by fixing the upper ankle joint in equinus position or under mobilization through a limited range of movement. The contralateral Achilles tendons served as internal control. All tendons were tested biomechanically at short intervals up to the 112th postoperative day in terms of load to failure [N], tendon deflection [mm] and tendon stiffness [N/mm], and were evaluated histologically after 8 and 112 days. Postoperative mobilization resulted in a continuous and significantly more rapid restoration of load to failure in comparison to the immobilization group. Tendon deflection was decreased by postoperative mobilization, whereas under immobilization it paradoxically increased still further in the later course. After 112 days the tendons of the mobilization group had regained their original tendon stiffness, whereas the tendons after immobilization reached only about half the values seen in the control tendons. Histologically, postoperative mobilization led to increased immigration of inflammatory cells in the early phase. In the late phase, as compared to immobilization, tendon structure was more mature, with fibre bundles arranged in parallel and interposed tendocytes. Tensile loading of the healing tendon by postoperative mobilization leads to fundamental changes in the biological process of tendon healing resulting in accelerated restoration of load to failure and reduced tendon deflection.

The mineralization of mantle dentine and of circumpulpal dentine in the rat: an ultrastructural and element-analytical study.

The purpose of this study was to compare the biomineralization of circumpulpal dentine with that of mantle dentine by ultrastructural and element-analytical techniques. Forty upper second molar germs of 10-day-old albino rats were cryofixed in liquid nitrogen-cooled propane and embedded in resin after freeze drying. Semithin dry sections were cut for analyzing the calcium and phosphorous concentration in initial mantle dentine, at the mineralization front of circumpulpal dentine, in the middle region of circumpulpal dentine and in mantle dentine peripheral to circumpulpal dentine. For the morphological evaluation of mineral deposits we compared ultrathin and unstained sections of cryofixed molars with chemically fixed molars. For both dentine types it was found that they develop via identical steps of mineral formation at collagen fibrils and non-collagenous matrix molecules. In circumpulpal dentine no globular mineral protrusions along the mineralization front (i.e. calcospherites) and no indications of interglobular dentine at the transition from circumpulpal dentine to mantle dentine were present. Two von Korff fibres were not only visible in mantle dentine but also in circumpulpal dentine. Matrix vesicles were present only during the formation of an initial coherent layer of mantle dentine and could not be observed during successive formation of mantle dentine and circumpulpal dentine. The element-analytical data did not demonstrate any difference in the mineral content between the two dentine types. Therefore, we conclude that mantle dentine and circumpulpal dentine in the rat molar possess a high degree of structural and chemical similarity and that only the extent of terminal branching of the odontoblast processes gives an approximate estimation of the thickness of mantle dentine.

Development and evaluation of an experimental model for investigating the pathogenesis and therapeutic strategies of acute liver failure.

Because of the various etiologies of acute liver failure (ALF) a clinically relevant model must fulfill four criteria--reversibility, reproducibility, ALF-induced death, and a sufficient time interval for diagnosis and therapy between induction and death. In this study an experimental model was evaluated for these criteria. A total of 49 rats were randomized into seven groups: First, a pilot study was performed regarding the survival rate after different treatments: In group I, animals underwent a 70% liver resection. In group II, 70% liver resection was combined with ascending doses of postoperative endotoxin administration up to 400 microg/kg (group IIc). In group III, animals only underwent liver mobilization. In group IV, ALF was induced according to the protocol of group IIc, but with additional treatment of an endothelin-A-receptor (ETAR) antagonist. Animals in group V received only 400 microg endotoxin. After induction of ALF, all animals died within the first day, showing significantly elevated bilirubin and ammonium levels and severe damage to hepatocellular integrity. Application of the ETAR antagonist resulted in the survival of 6/7 animals until the 14th day; the biochemical and histomorphological changes were reversible. All other animals survived to the 14th day. A clinically relevant model of ALF in rats can be created by the combination of 70% liver resection and endotoxin application to produce an inflammatory component.

Strain-related bone remodeling in distraction osteogenesis of the mandible.

Distraction osteogenesis has become a mainstay in craniofacial surgery. However, there are several unresolved problems concerning the biology of bone regeneration. We investigated the biomechanical effects of mandibular lengthening in 32 rabbits on a cellular and histologic level. The mandible was subjected to a corticotomy, held in a neutral position for 4 days, and then lengthened at various strain rates and frequencies for 10 days. Radiographic, histologic, and electron microscopic examinations showed a strain-related bone regeneration. Application of physiologic strain rates (2000 microstrains or 0.2 percent) led to a bridging of the artificial fracture exhibiting woven ossification, whereas at 20,000 microstrains trabecular bone formation was demonstrated. In contrast, hyperphysiologic strain magnitudes (200,000 microstrains and 300,000 microstrains) showed a fibrous tissue formation. Multiple strain applications (10 cycles/day versus 1 cycle/day) increased the width of the distraction gap without changing the stage of bone regeneration. The gradual distraction of bone in physiologic magnitudes at higher frequencies seems to be desirable for a bony differentiation and may help to improve clinical applications.

Histocytological evaluation of the reaction of rat pulp tissue to Carisolv.

OBJECTIVES: Carisolv is a viscous substance for the chemo-mechanical removal of caries. The aim of this study was to examine the tissue reactions of pulp and dentin tissue to Carisolv in vivo. METHODS: In 48 caries-free molars (24 Wistar rats, age: 3 months) an occlusal cavity was prepared. The roofs of the pulp chambers were perforated with a sharp probe and Carisolv solution was placed onto the exposed pulp of 24 molars for 1, 10 and 20 min. Twenty-four contralateral molars served as controls and were coated with an inert liquid containing isotonic saline solution, carmellose and erythrosin. RESULTS: After a contact period of 10 and 20 min in the experimental group, pulpal destruction of intercellular substance and cells including odontoblasts was observed up to a depth of 150 microm, in addition to the mechanical damage by perforation (e.g. dentin particles). The pulpal and predentin fibrils as well as the dentin fibrils appeared to be intact and did not differ from the controls. After a contact time of 1 min a weaker damage of pulp cells and odontoblasts, as well as intact fibrils in pulp, predentin and dentin could be observed. CONCLUSIONS: Radicals of Carisolv (OH- and OCI-) cause alkaline hydrolysis of cellular components but do not decompose collagenous tissue components.

Mechanical tension in distraction osteogenesis regulates chondrocytic differentiation.

Differentiation of chondrocytes to cells of osteoblastic phenotype occurs during an interim period of bone development, fracture repair and distraction osteogenesis. To study the relationship between tension-stress and chondrogenesis, uniaxial strains (0 microstrains, 2000 microstrains, 20000 microstrains, 200000 microstrains, 300000 microstrains) were applied in a rabbit model of mandibular distraction osteogenesis. The results demonstrated that cell differentiation, apoptosis and tissue development in the newly formed gap tissue showed a correlation to the applied strain magnitudes. Only strains of 20000 microstrains resulted in a statistically significant (P<0.05) formation of cartilage struts with embedded chondrocyte-like cells. However, chondrocyte-like cells were rarely detected in samples distracted at lower or higher strain magnitudes. Osteoblasts appeared to replace cartilaginous matrix by mineralized bone matrix. The phenotypic change from chondrocytes to osteoblasts was accompanied by a decreased proteoglycan synthesis. a change in the expression from type II collagen towards type I and involved asymmetric cell divisions and apoptotic cell death. Therefore, we suggest that mechanical strain is an external stimulus responsible for phenotypic cell alterations.

The rat esophagus: ultrastructure and radiological aspects of tissue response after 1320 nm Nd:YAG laser irradiation.

Morphological tissue response towards laser treatment was investigated in the esophagi of adult Wistar rats by light- and transmission electron microscopy. The specimens were fixed by perfusion immediately, 2 days and 14 days after laser treatment in order to assess different stages of the healing process. The epithelium of the lasercentre was completely destroyed in the immediate group. The connective tissue showed damaged cells, fused collagenous fibres and occluded blood vessels. Smooth muscle cells presented a vacuolated sarcoplasm and pycnotic nuclei. The cross striation of skeletal muscle cells had disappeared and their nuclei were karyolytic. In a distance of 4 mm from the lasercentre all wall tissues had an almost normal appearance. After 2 days the morphological feature of the lasercentre was the same as in the immediate group. In a distance of 2 mm some layers of flat and intact epithelial cells were observed below the necrotic epithelium. The adjacent connective tissue was infiltrated by inflammatory cells. After 14 days the formation of granulation tissue had caused an occlusion of the lumen in the lasercentre. In a distance of 2 mm the lumen was patent and the wall tissues had been partly restored. As the rat esophagus serves as a model for esophagotracheal fistulae in newborn children we assume the 1320 nm Nd:YAG laser to be a possible application in occlusion of these fistulae.

[The cell response of rat esophagus after intraluminal irradiation with Nd-YAG-laser (1064 nm) after various postoperative times: a light and electron microscopic study]. / Die Gewebeantwort des Rattenösophagus nach intraluminaler Bestrahlung mit Nd-YAG-Laser (1064 nm) zu unterschiedlichen postoperativen Zeitpunkten: Eine licht- und elektronenmikroskopische Studie.

The esophagi of Wistar rats were irradiated by a Nd-YAG-laser and studied by light and transmission electron microscopy immediately, 2 days and 14 days after the operation. In the immediate group the lasercentre showed a destruction of the stratified epithelium. On the contrary the cells and fibres of the underlying connective tissue were hardly affected. In the lasercentre we have found occlusion of microvascular lumina. Both the layer of smooth muscular tissue and the layer of skeletal muscular tissue showed defects in their myofilaments and altered nuclei. The damage of the skeletal muscle fibres extended up to 4 mm distant from the lasercentre. After 2 days a migrating epithelial sheet was present below the necrotic epithelium and an inflammatory reaction was found in the connective tissue. After 14 days a new regenerated epithelium and an underlying granulation tissue had caused a stenosis of the esophageal lumen. The smooth and striated muscle fibres also showed signs of regeneration. We assume a high regenerative capacity of all involved tissues.

A new device for long-term continuous enteral nutrition of rats by elementary diet via gastrostomy, following extensive oesophageal or lower gastrointestinal surgery.

A device for intragastric nutrition of unsedated and minimally restrained rats under complete alimentary abstinence has been developed. The cannulation system consists of an infusion pump, modified glass syringe as flow swivel, rat-harness and a silicone-tube-gastrostomy. The animals were kept in special cages and coprophagy was prevented by an own model of faecal collection cup. Methionine and Ca-glycerophosphate had to be added to a commercial elementary diet. The rats were allowed to move freely during intragastric infusion, which was applied for 14 to 28 days in 118 Wistar-rats (350-400 g). They maintained weight on an energy supply of 86.4 kcal/day.

Tracheoesophageal anastomosis, continent gastrostomy and oesophagostomy--a new experimental model in minipiglets.

Tracheoesophageal fistulae endanger newborn children by aspiration pneumonia. An animal model of this dangerous malformation was thought to be impossible until now. By tracheoesophageal anastomosis, occlusion of the oesophagogastral junction, proximal cervical oesophagostomy and continent gastrostomy in 25 minipiglets a reliable model of tracheoesophageal fistula has been developed. After establishing the model in a pilot study the mortality following the establishment of this procedure was 5.6% and the minipiglets survived in excellent condition for 2-10 week periods under complete alimentary restriction and gastrostomy feeding. Therefore the method seems to be suitable for long-term investigations on major oesophagotracheal surgery.

The scope of minimally invasive pediatric surgery (MIPS) and its influence on reduced length of hospitalization.

Minimally invasive pediatric surgery (MIPS) is a new dimension in pediatric surgery. The scope of methods replacing large operations in children by smaller endo- or laparoscopic interventions is presented.Electromagnetic double-segmental elongation has achieved auto anastomosis in 16 out of 22 children with long gap esophageal atresias since 1975. A new radial laser-applicator with entirely lateral radiation has led to a method for endoscopic occlusion of esophagotracheal and other fistulas with a 100% success rate in a minipig-model. Our experience with 54 laparoscopic interventions including large bowel resections, hiatus hernia, and a new technique for pyloromyotomy documents that laparoscopy halves hospitalisation-periods and may well become the most rapidly growing branch of pediatric surgery.

Laparoscopic and thoracoscopic surgery in infancy and childhood, the Münster/Gent experience.

Laparoscopic and thoracoscopic surgery have had a tremendous impact on adult surgery, but are still rarely used in children. In the past 3 years 168 children have been treated endoscopically in the three cooperating institutions. The course of all patients was documented prospectively in order to determine the value and prove the safety of endoscopic surgery in children. The operations performed endoscopically comprised appendectomy (n = 39), diagnostic procedures (24), adhesiolysis (n = 22), cholecystectomy (n = 13), bowel resection (n = 8), pyloromyotomy (n = 6), ovarial resection (n = 6), hiatus hernia repair (n = 22), splenectomy (n = 6), first stage of Fowler-Stephens operation for kryptorchidism (n = 3) and interruption of persistent botallic ducts (n = 6) in preterm infants. Apart from four wound infections (2.4%) following perforated appendicitis, one of which required laparotomy for perityphlitic abscess there were no serious complications. No patient died, and overall morbidity related to the procedure was 1.8% (i.e. incidence of uneventful minor intraoperative complications). Furthermore endoscopic surgery provides a better diagnostic survey of the abdomen or thorax so that missing of secondary pathology is unlikely. Pain and reflectory impairment of bowel-function or respiration were diminished, the aesthetic results were excellent and hospitalization could be reduced to a minimum. It is concluded that laparoscopic and thoracoscopic surgery are safe and reliable procedures for children and yield very encouraging results.

[Improved model of a fecal collection device for the prevention of coprophagia in the rat]. / Verbessertes Modell einer Kotfalle zur Unterbindung der Koprophagie bei dei Ratte.

Previous fecal collection cups with tape fixation frequently led to serious necroses of the rat tail due to strangulations, and did not allow for growth of the tail. A recently developed new model gives space for rapidly growing tails and is securely fixed by a cannula perforating both fecal collection cup and tail. It is transparent, cheap to be made, and easy to empty and was tested in 118 animals for two and four weeks. In no case the cup did loosen or had to be removed due to infections, strangulations or any local problems.

[No opiates against cannabis hyperemesis syndrome]. / Keine Opiate gegen das Cannabis-Hyperemesis-Syndrom.

HISTORY AND CLINICAL FINDINGS: An otherwise healthy 25-year-old patient with heavy cannabis-abuse suffered from an undiagnosed cannabis hyperemesis syndrome (CHS) over years, which characteristically was resistant to usual antiemetics. In an apparently last attempt at healing, opiates (morphine, methadone) were administrated and improved the CHS, however, this led to an at least as equally distressing and painful opiate withdrawal syndrome. TREATMENT AND COURSE: In the controlled cannabis abstinence during the 2-week inpatient treatment of opiate addiction syndrome the CHS has not recurred. CONCLUSION: Opiates are not suited for the treatment of CHS because they are addictive and lead to respiratory depression in overdose.