Effect of physical cues of altered extract media from biodegradable magnesium implants on human gingival fibroblasts.

Volume stable barrier membranes made of magnesium are very promising in Guided Bone Regeneration (GBR) to treat periodontal bone defects in dentistry due to their excellent biocompatibility and biodegradability. During the degradation process the cells are exposed to the alteration of various parameters, so called physical cues, involving surface alterations due to the formed corrosion layer and medium alterations arising from the dissolved corrosion products. Cell migration of human gingival fibroblasts (HGF), as a crucial parameter for optimal healing process in GBR, has been investigated on magnesium membranes and revealed that medium alterations by dissolved corrosion products have a higher impact on cell migration than surface alterations. However, the effect of each altered medium parameter on cell migration has not been adequately studied, but their roles are crucial to explain the slower migration rate on magnesium surfaces compared to titanium and tissue culture plastic surfaces. Our study investigates the single effect of Mg2+, Ca2+, H2 and increased osmolality as well as the effect of magnesium extracts, which contain a dynamic mixture of previous parameters on cell migration, proliferation and viability of HGF. We showed that at 75 mM Mg2+ concentration and at 0 mM Ca2+, respectively, the cell migration rate is greatly reduced. In complex magnesium extract media, we found that a temporarily increased ratio of Mg2+ to Ca2+ conditioned a slow HGF migration rate. Based on these findings and the characterization of supernatants from HGF migration assays on Mg membranes, we propose, that the slower migration rate of HGF can be explained by the altered ratio of Mg2+ to Ca2+, caused by increasing concentrations of Mg2+ and decreasing concentrations of Ca2+ in the vicinity of the corroding Mg implant, combined with a constantly increased molecular hydrogen concentration in the supernatant. These results are cell type specific and should be checked carefully, if necessary, for Mg implant performance. STATEMENT OF SIGNIFICANCE: The study is providing a systematic approach to explain the main effects of extract medium parameters (physical cues) such as magnesium or calcium ion concentration, osmolality and dissolved molecular hydrogen and CO2 in cell culture media modified by co-incubating with corroding magnesium implants on the migration rate of human gingival fibroblasts (HGF). This study uncovers for the first time the combinatory effect of slightly increased molecular hydrogen and the change in Mg2+/Ca2+ ratio on HGF cell migration.

Design of a migration assay for human gingival fibroblasts on biodegradable magnesium surfaces.

A novel regenerative approach to Guided Bone Regeneration (GBR) in dental surgery is based on the development of biodegradable and volume stable barrier membranes made of metallic magnesium. Currently used volume stable barrier membranes are made of titanium-reinforced PTFE or titanium-reinforced collagen membranes, both, however, are accompanied by a high incidence of wound dehiscence resulting in membrane exposure, which leads to an increased infection risk. An exposed membrane could also occur directly after insertion due to insufficient soft tissue coverage of the membrane. In both cases, fast wound margin regeneration is required. As a first step of soft-tissue regeneration, gingival fibroblasts need to migrate over the barrier membrane and close the dehiscent wound. Based on this aim, this study investigated the migration behaviour of human gingival fibroblasts on a magnesium surface. Major experimental challenges such as formation of hydrogen bubbles due to initial magnesium corrosion and non-transparent material surfaces have been addressed to allow cell adhesion and to follow cell migration. The designed scratch-based cell migration assay involved vital fluorescent cell staining on a pre-corroded magnesium membrane to simulate invivo wound dehiscence. The assay has been used to compare cell migration on pre-corroded magnesium to titanium surfaces and tissue culture plastic as control substrates. First results of this assay showed that human gingival fibroblasts migrate slower on pre-corroded magnesium compared to plastic and titanium. However, the scratch was finally closed on all materials. Compared to titanium surfaces and tissue culture plastic, the surface roughness and the surface free energy (SFE) could not explain slower cell migration on magnesium surfaces. Immunohistological investigations of cellular structure revealed, that magnesium ions increased focal adhesion at concentration of additionally 75 mM MgCl2 in cell culture medium. The use of our designed cell migration assay has shown that ionic medium alterations due to magnesium corrosion has a higher impact on the cell migration rate than surface alterations. STATEMENT OF SIGNIFICANCE: The design of a migration assay on non-transparent magnesium surfaces will add the option to study cell response to surface modifications, coatings and the corrosion process itself under life view conditions.

Determining Tarsus Curvature in Relation To Direction of Gaze Using Non-contact Ultrasound Video Assistance.

PURPOSE: Plastic surgery on the eyelids for the purpose of aesthetic or functional correction requires precise knowledge of lid anatomy. Changes in the tarsal curvature of the upper eyelid relative to line of vision are important, particularly when a surgical correction of paralytic lagophthalmos is undertaken. We used a computer-based image-processing algorithm to establish a relationship between changes in the curvature of the tarsus relative to the line of vision. MATERIAL AND METHODS: A dynamic, ultrasound examination of the upper eyelids of 100 participants (100 eyes) was performed transpalpebrally using a 7.5 MHz scanner with the patient looking straight ahead, away from and towards the midline of the body. A computer-aided examination of the upper eyelid tarsus was then performed, followed by the calculation of the radius of curvature of the tarsus relative to the line of vision in each position. RESULTS: Using regression of a Taylor polynomial, the shape of the tarsus was mapped by a quadratic function, and the change in tarsal curvature relative to line of sight could be demonstrated. CONCLUSION: With objective evidence of change in the tarsal curvature relative to the line of sight, this may influence the treatment of pathological changes in the upper eyelid.

Secondary peritonitis: severity of disease and activation of peritoneal cells.

OBJECTIVE: To compare the degree of the inflammatory response of human peritoneum with the severity of peritonitis. DESIGN: Clinical laboratory study. SETTING: University hospital, Germany. SUBJECTS: 15 patients with diffuse secondary peritonitis and 5 having conventional cholecystectomy (controls) had peritoneal specimens taken from the site of incision. MAIN OUTCOME MEASURES: Correlation between presence of indicators of the inflammatory response: interleukin 1 (IL-1), interleukin 6 (IL-6), intercellular adhesion molecule-1 (ICAM-1), antibacterial protein (defensin 3 reflecting the activation of granulocytes), the antibody clone HAM 56 (for detection of local macrophages), and antibodies against macrophage migration inhibiting factor (MIF)-related proteins 8 and 14 (MRP 8 and 14), and clinical state evaluated by the Mannheim Peritonitis Index (MPI), the Peritonitis Index Altona II (PIA II) and the Acute Physiology Score (APS). C-reactive protein (CRP) concentrations were measured preoperatively in the serum. RESULTS: Expression of MRP 8 and 14, HAM 56, and defensin 3 was significantly higher in patients with peritonitis than in controls (p < 0.05). Expression of IL-1 and IL-6 was almost undetectable. ICAM-1 expression correlated significantly with phagocytic activation. There was no correlation between clinical scores, CRP, and immunohistochemically detectable variables. CONCLUSION: The pattern of peritoneal inflammatory reactions is relatively uniform and does not correlate with the clinical grading of severity.

Transfer of biological traces in cases of hanging and ligature strangulation.

In hanging and ligature strangulation, the noose mostly causes a mark or groove which is formed partly by compression of the skin and partly by abrasion with loss of the upper epidermal layers. The horny scales abraded from the neck may be transferred to the strangulation device or to the interposed textiles where they are sometimes visible at stereomicroscopic examination or even to the naked eye as silver-grey particles. The morphologic features of the epidermal transfer due to hanging and ligature strangulation is demonstrated by 14 case examples. The biological traces may be sufficient for comparative DNA typing by means of PCR-based methods. In 9 out of the 14 cases, genomic DNA typing was successful. Analysis of mtDNA succeeded in another two cases, although genomic DNA could not be detected. Beside the accumulation of solid epidermic particles the paper describes deposition of serous and fatty tissue fluid at the ligature (mainly adjacent to skin ridges).

Postmortem endoscopy of the ocular fundus. A valuable tool in forensic postmortem practice.

Pathological changes of the ocular fundus in acute severe head or extracranial injury are difficult to document. In vivo examination usually has to be carried out under intensive care conditions. Postmortem ophthalmoscopy is usually impeded by rapid blurring of the cornea. Therefore, endoscopic fundal photography has been applied by using a pars plana approach during forensic postmortem procedures. This technique provides fundoscopic pictures of high quality as a valuable supplement for subsequent macro- and histopathological preparations and sections. Different patterns of fundoscopic findings seem to correspond to different types of injuries such as craniocerebral trauma, traumatic asphyxia and decompression sickness, and may allow some insight into the respective pathogenetic mechanism.

Inflammatory consequences of the translocation of bacteria and endotoxin to mesenteric lymph nodes.

BACKGROUND: Translocation of intestinal bacteria to mesenteric lymph nodes (MLNs) has been documented in humans under a variety of circumstances, yet its clinical significance remains to be established. The aim of this study was to correlate detectable translocation to MLNs of bacteria and endotoxin with local and systemic signs of inflammation. METHODS: From each of 10 patients with carcinoma of the cecal region two MLNs were harvested prior to resection. The presence of bacteria and endotoxin in the lymphatic tissue and blood was determined by culture methods and DNA preparation (PCR) and by a Limulus assay, respectively. Inflammatory mediators were determined in plasma and in MLN homogenates. RESULTS: Viable bacteria were detected in MLNs of 7 patients and in 9 of 20 lymph nodes. PCR revealed traces of bacteria in 4 patients and in 6 of their MLNs. Combining both modalities, the translocation rate was 80% and 55% for patients and MLNs, respectively. There was no detectable bacteremia. Endotoxin was found in the plasma of 7 patients and in 9 MLNs from 5 patients. There was no correlation between culture findings and endotoxin concentrations. Moreover, bacteriological data did not correspond to local or systemic inflammation. The group of MLN with detectable endotoxin differed significantly from LPS-negative nodes with respect to interleukin-6, interleukin-10, and sCD14. Systemic concentrations of endotoxin and inflammatory parameters did not correspond to levels within MLNs. CONCLUSION: Translocation to MLNs occurs in patients with cecal carcinoma. This, however, seems not to be of major clinical significance if no additional physiologic insults are encountered. Irrespective of the presence of bacteria, there are variations in inflammatory reactions between lymph nodes from one and the same patient, probably reflecting fluctuating response mechanisms to low-grade translocation.

[Significance of mitochondrial DNA for forensic stain analysis, identification and forensic lineage determination]. / Zur Bedeutung der mitochondrialen DNA für die forensische Spurenkunde, Identifizierung und Abstammungsbegutachtung.

DNA testing using conventional STR systems may produce insufficient results, if the genomic DNA in the specimen is either highly degraded or the available quantity is very small (e.g. skin particles, hair shafts or ancient bones). In some of these cases the examination of mitochondrial DNA, which is present in considerably larger copy numbers in the cytoplasm, is more successful than that of nuclear DNA. Identification of unknown corpses by conventional DNA typing sometimes remains doubtful, if only samples from presumably distant relatives or putative brothers or sisters are available for comparison. Since mitochondrial DNA is generally transmitted in maternal lineages, its sequence pattern can be directly compared with those of other individuals and, in case of the same maternal lineage, corresponding sequence chromatograms are to be expected. In connection with nuclear DNA typing methods certain sequence motives may furnish clues to ethnic groups. The report presents three cases illustrating the application possibilities of mtDNA typing in forensic practice.

Local inflammatory peritoneal response to operative trauma: studies on cell activity, cytokine expression, and adhesion molecules.

OBJECTIVE: To test the hypothesis that different surgical procedures may lead to different degrees of activation of the human peritoneal response. DESIGN: Clinical laboratory study. SETTING: University Hospital, Germany. MATERIAL: Peritoneal specimens taken from the incision or parietal resection margins at the beginning and end of laparoscopic or open cholecystectomy, or other conventional open operations (n = 5 in each group). MAIN OUTCOME MEASURES: Detection of indicators of the inflammatory response: interleukin 1 (IL-1), interleukin 6 (IL-6), intercellular adhesion molecule- (ICAM-1), antibacterial protein (defensin 3 that reflects the activation of granulocytes), the antibody clone HAM 56 (for detection of local macrophages), and antibodies against macrophage inhibiting factor (MIF)-related proteins 8 and 14 (MRP 8 and 14). RESULTS: The rise between preoperative and postoperative evaluations was significant for each variable (p < 0.05). With one single exception (IL-6 between laparoscopic cholecystectomy and other operations), the one way analysis of variance (ANOVA) showed no significant differences among the three groups in the detectable increases in staining. Linear regression analysis showed no correlation between length of operation and increases in immunohistochemically detected inflammatory variables. CONCLUSION: Minimally invasive surgery does not necessarily mean minimal peritoneal damage. The immunohistochemical evaluation of the local cellular response may provide additional objective criteria for the grading of operative trauma.

Sudden cardiac death due to atypical isolated coronary arteritis?

A case of probable isolated primary arteritis of the coronary arteries (coronaritis) in a 68-year-old man who died suddenly and unexpectedly is presented. The histologic presentation of the disorder is discussed, especially the differential diagnosis of arteritis of the coronary arteries versus distinct coronary artery sclerosis with concomitant inflammatory change.

Systemic Fusarium oxysporum infection in an immunocompetent patient with an adult respiratory distress syndrome (ARDS) and extracorporal membrane oxygenation (ECMO).

We report a case of disseminated fusariosis in a 42-year-old patient with adult respiratory distress syndrome (ARDS) and extracorporal membrane oxygenation (ECMO), but without definite immunosuppression. Fusarium oxysporum was isolated from a bronchial lavage taken 6 days ante mortem. Despite antifungal treatment with amphotericin B and flucytosine the patient died in septic multiorgan failure. A post-mortem examination was performed. The patient's liver was found to contain fungus cells and F. oxysporum could be cultured from ascites.

Selenocysteine synthesis in mammalia: an identity switch from tRNA(Ser) to tRNA(Sec).

The mechanism of selenocysteine insertion into proteins is distinct from all other amino acids in all lines of descent in that it needs specific protein cofactors and a structurally unique tRNA(Sec). It is first aminoacylated with serine and further recognized among all other serylated serine isoacceptors by a selenocysteine synthase and is converted to selenocysteyl-tRNA(Sec). We present here the complete set of identity elements for selenylation of mammalian seryl-tRNA(Sec) and show that the transplantation of these elements into normal serine tRNA allows its selenylation. Four particular structural motifs differentiate eukaryotic tRNA(Sec) from normal tRNA(Ser): the orientation of the extra arm, the short 4 bp T psi C-stem, the extra long 9 bp acceptor-stem and the elongated 6 bp dihydrouridine-stem. Only the last two are essential and only together sufficient for selenocysteine synthesis, whereby the additional base-pairs of the acceptor-stem may be replaced by non-paired nucleotides. Each exchange of the first three structural motifs mentioned above between tRNA(Ser) and tRNA(Sec) resulted in a significant loss of serylation, indicating that the overall composition of particular structure elements is necessary to maintain normal functions of tRNA(Sec). Since we find that all seryl-tRNAs which are selenylated are also substrates for serine phosphorylation we propose that phosphoseryl-tRNA(Sec) is a storage form of seryl-tRNA(Sec).

[Differential diagnosis of hematotympanum in forensic autopsy]. / Zur Differentialdiagnose des Hämatotympanums bei forensischen Obduktionen.

In a living patient, a hemotympanum can be a sign of a temporal bone fracture. Postmortem endoscopy in forensic autopsy cases also revealed hemotympanum findings without any fracture. Thoracic compression or cranio-facial congestion were probably the pathogenetic mechanism in some of the cases. An engorgement of the internal and external jugular veins was found in all cases. Due to increased intravenous and intraarterial pressure, capillary hemorrhages were revealed in endoscopy and histological sections.

Editing does not exist for mammalian selenocysteine tRNAs.

It has been reported that selenocysteine tRNA from bovine liver is completely edited to two isoacceptor species, called tRNA([Ser]SecNCA) and tRNA([Ser]SecCmCa), which differ from the gene sequence. We used direct tRNA sequencing, mobility shift analyses, primer extension, restriction enzyme digestion and single strand conformational polymorphism (SSCP) analyses of products from reverse transcription coupled with polymerase chain reaction (RT/PCR), sequencing of RT/PCR products and HPLC-coupled mass spectrometry to reproduce this result and show here that editing of these tRNAs does not occur.

[Postmortem endoscopic diagnosis]. / Postmortale endoskopische Diagnostik.

The use of endoscopical methods augments the macroscopical diagnosis in autopsies. Cavities and sinuses which are hardly accessible may be inspected and photographically documented as well without damage of the anatomical context. In terms of forensic autopsies the endoscopical technique is mostly recommended for the inspection of the fundus of the eye, the external auditory meatus up to the tympanic membrane, the naso- and laryngopharynx. Pathological and traumatical changes can be valued at once in situ. Forensic evidence of postmortal endoscopical findings is exemplified.

Idiopathic chronic fatigue. A primary disorder.

Idiopathic chronic fatigue, an entity worthy of attention, understanding, and treatment, has disappeared from medical and psychiatric publications. Nonetheless, it is an important clinical phenomenon.

Identification by MRI.

A semi-skeletted water-lodged corpse was supposed to be an 82-year-old missed man. Fifteen years ago this person underwent an operation of a trimalleolar fracture of the left ankle joint. Conventional X-ray photograms of the left ankle joint of the water-lodged corpse showed no evidence of an operation. A T-1 weighted spin-echo sequence revealed the entire aspect of the osteosynthesis as documented in the previously performed post-operative X-ray photograms.

Immunohistological demonstration of lymphocyte surface antigens in postmortem lymphoid tissues.

The postmortem stability of cell antigens has hardly been studied. Using monoclonal antibodies (mabs) we examined the postmortem detectability of lymphocyte surface antigens in different lymphoid organs by comparing two sensitive, immunohistological staining procedures. To quantify the probable degree of autolysis of the tissues a score system was applied by taking into consideration the postmortem age as well as the core temperature of the corpses. The antigens examined generally proved to be very resistant to autolytic influences. Differences were found when comparing different mabs and with regard to the type of lymphoid tissue. The loss of immunohistological reactions was most extensive in the spleen whereas tonsils showed almost no qualitative alterations in staining patterns. Reactivity of mabs with postmortem tissues decreased in the following order: Dako CD22 and anti-Leu 4, anti-Leu 3a, anti-Leu 7, Dako T8. The mabs anti-Leu 7 and Dako-T8 frequently failed to demonstrate their respective antigens but no correlation between the loss of staining and the degree of autolytic decomposition (our score) could be detected. In general, postmortem tissues as well as tissues shock frozen after delay are suitable for qualitative immunohistology of those cells characterized by the mabs applied. The APAAP-method proved unequivocally to be the superior staining technique.

Treatment of autoimmune thrombocytopenic purpura with rhesus antibodies (anti-Rh0(D).

There is evidence that blockade of the reticuloendothelial system (RES) by sequestration of autologous red blood cells (RBC) leads to an elevation of platelet counts in immune thrombocytopenia. To substantiate this hypothesis, 10 Rh0(D)-positive adult patients (9 female, 1 male) with chronic autoimmune thrombocytopenic purpura (ITP) (1 to 21 years duration) were treated with low doses of intravenous IgG-anti-Rh0(D) (200 to 1,000 micrograms per dose; 300 to 3,600 micrograms per course; administration within 1 to 5 days). All patients improved clinically as indicated by cessation of bleeding. In eight out of ten patients there was a rise in platelet count. Platelet increments were excellent (greater than 100 X 10(9)/l) in one, good (50-100 X 10(9)/l) in three, fair (20-50 X 10(9)/1) in two and low (10-20 X 10(9)/1) in two patients. Splenectomized patients (N = 4) had a poorer response than non-splenectomized patients (N = 6) with mean increments of 16 X 10(9)/l (range 5-43 X 10(9)/l) versus 60 X 10(9)/l (range 10-110 X 10(9)/l). The increase in platelet counts persisted for seven to over 150 days. Transient and slight signs of haemolysis developed in seven out of ten patients (haemoglobin remained stable; increase of lactate dehydrogenase (greater than 250 IU/l) in four, decrease of haptoglobin (less than 60 mg/dl) in five patients). The direct antiglobulin test became positive in all cases due to IgG1 without complement fixation. We conclude that the interaction of antibody-coated RBC with macrophages (and, probably, other means of RBC alteration) is a feasible therapeutic approach in selected cases of ITP and related conditions.