Christian Mittermayer

Topography of the inferior rectal artery: A possible cause of chronic, primary anal fissure

The authors believed that it might be possible to explain the local frequency of the anal fissure at the posterior commissure by an anatomic relationship, and examined the blood supply of the anus. The inferior rectal artery is demonstrated by postmortem angiography and by manual preparations (N=4) and histologic study after angiography of the vessels (N=10). The blood supply at the different sites of the anal canal are demonstrated by a morphometric study (N=20). The inferior rectal artery presents two variants in the postmortem angiographies, type 1 (85. 4 percent) and type 2 (14.6 percent). In type 1, the posterior commissure is less perfused than the other sections of the anal canal. In addition, the blood supply may be more compromised by contusion of the vessels passing vertically through the muscle fibers of the sphincter ani internus muscle during increased sphincter tone. The role of topography in the pathogenesis of the primary anal fissure is illustrated in a model

Successful long-term implantation of electrically inactive epiretinal microelectrode arrays in rabbits.

BACKGROUND In the ongoing discussion concerning the realization of an epiretinal prosthesis for electric stimulation of retinal ganglion cells, long-term fixation of such a device is a crucial question. We evaluated surgical techniques for implantation and fixation of electrically inactive microelectrode arrays (MA) into the retinas of rabbits and secondary tissue reactions to the implant. METHODS Vitrectomy and laser coagulation of the prospective fixation area were performed in rabbits. Implantation of MAs was performed 3 weeks later in 10 animals. The MA was fixated using retinal tacks. The follow-up included ophthalmoscopy and electrophysiology. At the end of the follow-up, the enucleated eyes were processed for light microscopy using standard procedures and grinding techniques. RESULTS Nine of 10 rabbits were implanted without serious complications. Clinical and electrophysiologic data through 6 months of follow-up did not indicate any adverse effect of the surgery, the implant, or the tack itself. No change in retinal architecture underneath the implant was found by light microscopy. In these cases, the implant was stable at its original fixation area. In three cases, mild cataract formation was observed, and in one case, a total retinal detachment was found. CONCLUSION Tack fixation of electrode arrays for electric stimulation of the inner retinal surface seems to be a useful approach in long-term implantation of an epiretinal prosthesis.

Theoretical and practical aspects of testing potential biomaterialsin vitro

In vitro methods provide a necessary and useful adjunct toin vivo studies in testing potential biomaterials. One of the most important functions is the screening for toxic effects of the biomaterials. The spectrum of changes elicited ranges from cell death to alterations of cell adhesion, proliferation and biosynthetic activity. Such test systems may involve the direct contact of cells with the biomaterials or the use of soluble extracts of the latter. The rapid, cost-effective and highly sensitivein vitro methods have to be weighed against the problem of validity of extrapolation to thein vivo situation. The first step in testing potential biomaterials may be termed “general” biocompatibility testing and usually involves the use of various cell lines (i.e. transformed cells) which can be easily cultivated and passaged in the laboratory for long periods. Although the latter is convenient and highly sensitive for recognizing and excluding toxic materials at an early stage in the process of toxicity testing, this method cannot be regarded as exhaustive. It is proposed that such screening methods be followed by a secondin vitro phase, in which primary and early passage cells of a type relevant to the proposed application of the medical device are used. This “specific” biocompatibility testing is an attempt to simulate thein vivo situation as closely as possible. A further component of such a testing scheme involves the use of relevant biological parameters, such as cell adhesion or the production of specific biosynthetic products by the relevant cell type. It is thus possible to construct a spectrum ofin vitro changes, ranging from marked inhibition of growth with frank cell death (“not biocompatible”) to marked stimulation of relevant growth and other cell biological parameters (“biocompatible and bioactive”). An expansion ofin vitro testing methods can offer a method to “tailor” biomaterials for specificin vivo applications. In conclusion, it must be stressed that allin vitro experimentation, whether “general” of “specific”, cannot replace the subsequentin vivo testing. Both components are necessary in an adequate scheme for testing potential biomaterials.

Elevated soluble adhesion molecules in women with pre-eclampsia: Do cytokines like tumour necrosis factor-α and interleukin-1β cause endothelial activation?

OBJECTIVE The purpose of the present study was to evaluate the clinical significance of tumour necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) for endothelial cell activation in pre-eclampsia. Therefore, we determined and compared the correlations between these cytokines and circulating adhesion molecules in the sera of pre-eclamptic pregnant women, normotensive pregnant women and nonpregnant women. METHODS The soluble adhesion molecules VCAM-1, ICAM-1, E-selectin, and P-selectin were determined in the serum of 38 pre-eclamptic pregnant women and 40 normotensive pregnant and nonpregnant controls using ELISA-techniques. We correlated these serum concentrations with the serum levels of TNF-alpha and IL-1beta, respectively, also determined by ELISA. RESULTS Elevated serum levels of VCAM-1 and E-selectin could be detected in pre-eclamptic patients, with and without HELLP-syndrome. In contrast, no increased serum concentration of ICAM-1, P-selectin, TNF-alpha and IL-1beta were found in these patients. While significant correlation between VCAM-1 and E-selectin could be determined (r=0.604; p<0.001) no unambiguous correlations, however, were found between TNF-alpha or between IL-1beta and the examined adhesion molecules or the selectins. CONCLUSIONS In contrast to in vitro investigations on cultured umbilical vein endothelium, our experimental results indicate that the cytokines TNF-alpha and IL-1beta can not explain endothelial cell activation, and that their measurement in serum is not useful for the detection of pre-eclampsia.

Induction of heat shock protein 70 by zinc-bis-(DL-hydrogenaspartate) reduces cytokine liberation, apoptosis, and mortality rate in a rat model of LD100 endotoxemia.

A prospective, randomized model of LD100/24 h endotoxemia was performed in male Wistar rats (n = 26; 250–300 g). The animals were divided into four groups: Group I (n = 5; saline treatment only), Group II (n = 5; Zn2+ treatment only), Group III (n = 8; saline pretreatment, lipdpolysaccharide (LPS) treatment), and Group IV (n = 8; Zn2+ pretreatment, LPS treatment). Zn2+ pretreatment was carried out by intraperitoneal injection of 50 mg/kg zinc-bis-(DL-hydrogenaspartate) (10 mg/kg Zn2+). LD100/24 h endotoxemia was induced by intraperitoneal administration of 20 mg/kg LPS of the Escherichia coli strain WO111:B4. Tumor necrosis factor α, interleukin-1β, and interleukin-6 were detected by enzyme-linked immunosorbent assay (ELISA). HSP70 expression in the lungs, the liver, and the kidneys was determined by immunohistochemistry, Western blotting, and an HSP70 ELISA. Apoptosis was also detected by an in situ apoptosis detection kit (TUNEL) and a cell death detection ELISA, respectively. This rat model of endotoxemia proves the close relationship between HSP70 expression, cytokine liberation, and development of apoptosis. The data demonstrate that: 1) Zn2+ is a potent inducer of HSP70 expression; 2) the application of Zn2+ leads to slightly increased cytokine plasma levels; and 3) the manipulation of the heat shock response by Zn2+ significantly increases the survival rate after LD100 endotoxemia. Enhanced survival rate in animals pretreated with Zn2+ may be explained by increased tissue levels of HSP70, a subsequent significantly decreased liberation of the proinflammatory cytokines after LPS challenge, and a significantly decreased rate of apoptosis.

Sclerosis of the internal anal sphincter--a process of aging.

Functional and organic alterations such as sclerosis of the internal anal sphincter muscle are often discussed as a cause of proctologic diseases. Systematic histopathologic examinations of this muscle are rare, although the internal anal sphincter is of primary significance for the continence of the anal canal and its functional efficiency. The authors believe that the degree of formation of connective tissue in this unstriated muscle depends on the age of the examined specimen. The authors can demonstrate that sclerosis is a physiologic process of aging.

The role of soluble adhesion molecules in evaluating endothelial cell activation in preeclampsia.

OBJECTIVE Adhesion molecules, such as vascular cell adhesion molecule 1, are known to be increased in the serum of patients with preeclampsia, indicating that these molecules are possible markers of endothelial cell activation. We investigated the influence of serum from women with preeclampsia on the expression of adhesion molecules by cultured endothelial cells. STUDY DESIGN Human umbilical vein endothelial cells were cultured in Ham/Iscove modified Dulbeccos medium containing 20% pooled human serum, l -glutamine (200 mmol/L), penicillin, and streptomycin. We stimulated these cells for 24 hours with sera from patients with preeclampsia and then determined the levels of vascular cell adhesion molecule 1, intercellular cell adhesion molecule 1, E-selectin, and P-selectin in the supernatant and in the maternal serum by means of enzyme-linked immunosorbent assay. These results were compared with those of sera obtained from normotensive pregnant and nonpregnant women. In addition, the expressions of these adhesion molecules on the endothelial surface were determined by immunofluo-rescence microscopy. RESULTS Only for vascular cell adhesion molecule 1 and E-selectin were elevated plasma levels found in hypertensive patients, whereas intercellular cell adhesion molecule 1 and P-selectin showed similar plasma levels in all the patients. No differences in the levels of the adhesion molecules were found between the supernatants of endothelial cell cultures after stimulation with sera from patients with preeclampsia and those after stimulation with normotensive control sera. In contrast, with immunofluorescence microscopy we could detect higher amounts of vascular cell adhesion molecule 1, intercellular cell adhesion molecule 1, and E-selectin on the endothelial surface after stimulation with sera from women with preeclampsia. CONCLUSION Although vascular cell adhesion molecule 1 and E-selectin were elevated in maternal serum samples from women with preeclampsia and on the endothelial surface after stimulation with such sera, there were no detectable increases in the levels of both of these adhesion molecules in the supernatant of cultured endothelial cells. We therefore assume that sera from women with preeclampsia may cause endothelial cell activation. Because we could not detect elevated concentrations of any of the investigated adhesion molecules in the supernatant of endothelial cells, we believe that factors other than sera from women with preeclampsia seem to play a major role in the release of soluble forms of adhesion molecules from the endothelial membrane.

Atherosclerosis and cancer: Common pathways on the vascular endothelium

This article reviews experimental and clinical data on atherosclerosis and cancer showing common pathogenic mechanisms. It is suggested that common pathways follow dysfunction of the vascular endothelium. The activation of the haemostatic system and the overexpression of cytokines and adhesion molecules by the endothelial cells represent important features of this dysfunction. These mechanisms can be responsible for progression of both diseases and explain the higher incidence of thromboembolic events in cancer patients, the occurrence of similar laboratory findings and the effect of many drugs on the course of the two diseases. Our article confirms that atherosclerosis and cancer share common mechanisms, and we hope it will stimulate further clinical trials on the use of drugs active on the haemostatic system in cancer patients.

Skeletal muscle oedema and muscle fibre necrosis during septic shock. Observations with a porcine septic shock model

In domestic pigs, intermitted application of Escherichia coli-endotoxin was used to create an animal model for a prolonged hypo- and hyperdynamic septic shock-like state and to investigate mechanisms of multiple organ failure. Here, we describe the changes in skeletal muscle after 18 h (2 animals) and 48 h (6 animals) of septic shock. Two pigs for each observation period that received physiologic saline solutions instead of endotoxin served as controls. The earliest lesions were endothelial cell damage with endomysial oedema and swelling of mitochondria in muscle fibres. With increasing degree of endothelial cell damage, pericytes showed degenerative changes with cytoplasmic fragmentation and karyolysis. After 48 h of shock, endomysial oedema was increased with fibrinogen present. Muscle fibre diameters were increased and swollen mitochondria and segmental necrosis of muscle fibres were frequently observed. However, phagocytic reaction or regenerative changes were not detected. In this respect, skeletal muscle lesions in septic shock differ from ischemic damage, which is characterized by early phagocytosis. Tumour necrosis factor alpha (TNFα) was increased greatly and significantly in the serum of the pigs that received endotoxin. The lesions described may be the result of both direct damage to muscle fibres by the endotoxin and/or the increased levels of TNFα and indirect damage because of the increased diffusion distance, due to the endomysial oedema. The loss of blood proteins into the endomysium may also play a role in generating hypoproteinemia in patients with septic shock.

Tumour-cell-endothelial interactions: free radicals are mediators of melanoma-induced endothelial cell damage

Damage to vascular endothelium may play an important role during metastasis. We used a three-dimensional model of tumour cell extravasation to test the hypothesis that certain types of tumour cells are able to induce vascular endothelial cell injury. Multicellular tumour spheroids (MCTS) of 14 human cancer cell lines and spheroids from two benign cell lines were transferred onto confluent monolayers of human endothelial cells (EC). MCTS from 4 of 7 melanoma cell lines induced damage of the endothelium which was closely associated with tumour cell attachment. Endothelial cell injury became evident morphologically by loss of cell membrane integrity and sensitivity to shear stress. Similar results were obtained with EC derived from human umbilical veins, umbilical arteries and saphenous veins. Addition of the oxygen radical scavenger catalase showed a dose- and time-dependent inhibition (up to 48 h) of EC damage in the case of the melanoma cell lines ST-ML-11, ST-ML-14 and SK-MEL-28. The scavenging enzyme superoxide dismutase proved to be protective (up to 12 h) in ST-ML-12 MCTS. In contrast, allopurinol, deferoxamine mesylate, ibuprofen, nor-dihydroguaretic acid, soybean trypsin inhibitor or aprotinin had no protective effect. None of the non-melanoma cancer cell lines or benign cells induced endothelial cell damage. Endothelial injury has been shown to enhance the process of metastasis. Our results suggest that free-radical-mediated endothelial cell damage may be one of the mechanisms contributing to the devastating metastatic potential of melanoma.