[Types of evidence syntheses]. / Formen der Evidenzsynthese.

This manuscript outlines various types of review articles as forms of evidence synthesis with special regard to their strengths and limitations. Review articles not only present summarised data, but also offer an evaluation of the quality of the individual studies included in it. The validity and the reliability of outcomes of reviews is strongly dependent on the quality of the data included. For this reason, a comprehensive literature selection process is paramount. Fundamental knowledge of bias and literature assessment is also necessary when reading reviews. This article presents selected tools for evidence appraisal and evaluation of bias risk.

Hypoglycin A Content in Blood and Urine Discriminates Horses with Atypical Myopathy from Clinically Normal Horses Grazing on the Same Pasture.

Hypoglycin A (HGA) in seeds of Acer spp. is suspected to cause seasonal pasture myopathy in North America and equine atypical myopathy (AM) in Europe, fatal diseases in horses on pasture. In previous studies, this suspicion was substantiated by the correlation of seed HGA content with the concentrations of toxic metabolites in urine and serum (MCPA-conjugates) of affected horses. However, seed sampling was conducted after rather than during an outbreak of the disease. The aim of this study was to further confirm the causality between HGA occurrence and disease outbreak by seed sampling during an outbreak and the determination of i) HGA in seeds and of ii) HGA and MCPA-conjugates in urine and serum of diseased horses. Furthermore, cograzing healthy horses, which were present on AM affected pastures, were also investigated. AM-pastures in Germany were visited to identify seeds of Acer pseudoplatanus and serum (n = 8) as well as urine (n = 6) from a total of 16 diseased horses were analyzed for amino acid composition by LC-ESI-MS/MS, with a special focus on the content of HGA. Additionally, the content of its toxic metabolite was measured in its conjugated form in body fluids (UPLC-MS/MS). The seeds contained 1.7-319.8 µg HGA/g seed. The content of HGA in serum of affected horses ranged from 387.8-8493.8 µg/L (controls < 10 µg/L), and in urine from 143.8-926.4 µg/L (controls < 10 µg/L), respectively. Healthy cograzing horses on AM-pastures showed higher serum (108.8 ± 83.76 µg/L) and urine concentrations (26.9 ± 7.39 µg/L) compared to control horses, but lower concentrations compared to diseased horses. The range of MCPA-carnitine and creatinine concentrations found in diseased horses in serum and urine were 0.17-0.65 mmol/L (controls < 0.01), and 0.34-2.05 µmol/mmoL (controls < 0.001), respectively. MCPA-glycine levels in urine of cograzing horses were higher compared to controls. Thus, the causal link between HGA intoxication and disease outbreak could be further substantiated, and the early detection of HGA in cograzing horses, which are clinically normal, might be a promising step in prophylaxis.

Morphometric analysis of the retina from horses infected with the Borna disease virus.

Borna disease (BD) is a fatal disorder of horses, often characterized by blindness. Although degeneration of retinal neurons has been demonstrated in a rat model, there are controversial data concerning whether a similar degeneration occurs in the retina of infected horses. To investigate whether BD may cause degeneration of photoreceptors and possibly of other neuronal cells at least at later stages of the disease, we performed a detailed quantitative morphologic study of retinal tissue from Borna-diseased horses. BD was diagnosed by detection of pathognomonic Joest-Degen inclusion bodies in the postmortem brains. Paraffin sections of paraformaldehyde-fixed retinae were used for histologic and immunohistochemical stainings. Numbers of neurons and Müller glial cells were counted, and neuron-to-Müller cell ratios were calculated. Among tissues from 9 horses with BD, we found retinae with strongly altered histologic appearance as well as retinae with only minor changes. The neuron-to-Müller cell ratio for the whole retina was significantly smaller in diseased animals (8.5 +/- 0.4; P < .01) as compared with controls (17.6 +/- 0.8). It can be concluded that BD in horses causes alterations of the retinal histology of a variable degree. The study provides new data about the pathogenesis of BD concerning the retina and demonstrates that a loss of photoreceptors may explain the observed blindness in infected horses.

Myoblasts for survive 16 weeks after intracardiac transfer and start differentiation.

INTRODUCTION: Skeletal myoblasts have been shown to survive transfer into myocardial scar tissue. The purpose of this study was to investigate differentiation after intramyocardial transplantation. Additionally, we evaluated an intravital fluorescence dye. MATERIALS AND METHODS: Skeletal myoblasts were harvested from 8 male inbred Lewis rats and expanded in culture. For each transplantation planned, 10(6) cells were trypsinized and incubated for three minutes with 2 ml of buffered PKH-26 solution (Sigma, St. Louis). This dye was integrated into the cell membrane. After washing three times, the cells were plated again for two more days. The cells were then transplanted into the hearts of 60 female Lewis-rats, mean weight 380 g (260 - 450 g). At implantation, 10(6) cells suspended in 100 microliters were injected into the apex region of the left ventricle. 12 animals served as the control group with only cell medium injected. Animals were sacrificed after 1, 2, 6, 8, 12, and 16 weeks (n = 10 each). The hearts were explanted and serial frozen sections of the hearts were prepared for detecting labeled cells. Sections with labeled cells were stained immunohistochemically for Myo D1 (myogenic origin), n-cam (early myotubes), desmin (muscular filament), myosin light chain (muscular contractile protein), and connexin 43 (tight junction). RESULTS: Cell labeling was successful in all cases. After two days, the myoblasts had recovered from the staining procedure. The fluorescing dye, however, was only rarely transmitted by cell division. Marked cells were found in the intercellular spaces between the cardiac myofibers in at least 8 animals from each group. No fibrotic reaction or inflammation was seen surrounding the transplanted cells. Up to 6 weeks after implantation, the cells stained positive for n-cam and Myo D1, and particularly for desmin. More n-cam positive cells were found than labeled cells, indicating cell division after the cell transfer. Two animals suffered sudden death after a follow-up time of 8 and 10 weeks, which was possibly due to cardiac arrhythmia. After 8 weeks, the cells formed conglomerates and stained positive for desmin, myosin light chain, and connexin 43. The cells were not structurally integrated into the recipient myocardial tissue, however. CONCLUSIONS: Myoblasts divided further after transplantation into rat myocardium. Positive staining for desmin demonstrated the development of myofibers. Starting at 8 weeks after transplantation, the cells started differentiation without reaching structural integration during follow-up. Labeling the cells with PKH-26 proved to be a reliable method to detect the cells.

Five years of experience with mitral valve homografts.

INTRODUCTION: The main advantages of mitral homografts are preservation of the subvalvular apparatus and avoidance of life-long anticoagulation. In this communication, we will present our five-year experience using mitral homografts in mitral valve surgery. PATIENTS AND METHODS: Since 1996, 14 patients (mean age 46 +/- 8 years, range 27 - 65 years have had mitral homografts implanted. Thirteen patients had mitral valve replacement; the septal leaflet of the tricuspid valve was replaced in one case. The indications were mitral (n = 6) or tricuspid endocarditis (n = 1), mitral valve stenosis (n = 3), and combined mitral valve disease (n = 4). Complete mitral homografts were implanted in eight patients; partial homografts were used in six cases. Preoperatively, the dimensions of the left ventricle and the mitral valve were measured by transoesophageal echocardiography (TOE). The mean left ventricular ejection fraction was 56 +/- 9%, the mean end-diastolic diameter 58 +/- 6 mm. The technique described by Acar/Carpentier was adapted for implantation; a Carpentier ring was implanted in all cases for annular stabilization. The patients had anticoagulative therapy which was discontinued when stable sinus rhythm was present after three months postoperatively. Follow-up included clinical examination, ECG, and echocardiography, and was initiated six months postoperatively and continued on a yearly basis. The following parameters were determined by echocardiography--left atrial size, left ventricular end-diastolic and end-systolic diameter, pressure gradient across the mitral valve (c/w Doppler, Bernoulli's equation), and mitral regurgitation. RESULTS: All patients survived surgery; the mean operation-time was 281 +/- 37 minutes. Intraoperative TOE revealed a first degree insufficiency in 7 patients. Follow-up was completed in all patients, with a mean period of 30 months (6 - 60 months). Two patients had an acute endocarditis two years postoperatively, requiring repeat valve replacement with a mechanical prosthesis. An additional patient had to be reoperated due to chordal rupture three years postoperatively. All three patients had mitral valve stenosis as the initial indication for surgery and had received a complete homograft. In the remaining eleven patients, the morphological and functional state of the implanted grafts remained unchanged during follow-up. The freedom from valve-related events was 93% after one year, 86% after two years, and 79% after three years. At six-month follow-up, ECG and echocardiography revealed sinus rhythm and sufficient atrial contractions in 13 cases. At the last follow-up, the pressure gradients were 3.4 +/- 0.6 mmHg for complete homografts and 2.8 +/- 0.6 mmHg for partial homografts. In five cases, a mild insufficiency was documented, while six patients presented with competent grafts. CONCLUSIONS: Mitral homografts can be used with acceptable mid-term results in selected cases with good left ventricular function and only slightly dilated left ventricles. Partial mitral homografts represent an additional technique, especially for mitral valve repair in patients with acute endocarditis. The susceptibility to bacterial infections of a homograft makes strict prophylaxis against endocarditis mandatory.

Homografts in patients with combined disease of the aortic valve and the ascending aorta: an alternative to the classical Bentall procedure.

BACKGROUND AND AIM OF THE STUDY: In patients with aneurysms or dissections of the ascending aorta and additional aortic valve disease, valve-containing composite grafts are used in clinical routine. The study aim was to present our experience with homografts for aortic valve replacement extended by a vascular prosthesis as an alternative to the classical Bentall procedure. METHODS: Thirty consecutive patients (mean age 46+/-14 years) were included in this study. Indications for valve replacement were aortic stenosis (n = 15), aortic insufficiency (n = 6), combined aortic valve disease (n = 6), endocarditis of the native valve (n = 1), and endocarditis of a previously placed bioprosthesis (n = 2). The mean diameter of the ascending aorta was 5.6+/-0.5 cm; one patient had an acute dissection (diameter 4.4 cm). For valve replacement, cryopreserved homografts (mean size 24+/-2 mm) were used in a mini-root technique, and the ascending aorta was replaced by collagen-coated vascular prostheses (mean diameter 28+/-3 mm). The size of the vascular prosthesis was adjusted to the diameter of the sinutubular junction of the implanted homograft. Follow up included annual clinical examinations, transthoracic echocardiography and ultrafast computed tomography (CT) scans. RESULTS: All patients survived surgery, and no deaths occurred during follow up. None of the patients had postoperative anticoagulation, and no thromboembolic events were noted. Follow up was complete, with an average 48 months (range: 6 to 84 months). Doppler echocardiography revealed trivial to mild aortic regurgitation in nine patients postoperatively, with no deterioration during follow up. No pathologic pressure gradients over the aortic valves were measured at Doppler echocardiography; the mean valvular orifice area was 2.5+/-0.3 cm2. At ultrafast CT, normal homograft anatomy including the sinotubular junction, no calcifications, and no signs of annular dilatation were seen. In the patient who had surgery for acute endocarditis of the native valve, ultrafast CT revealed a small pseudoaneurysm below the left coronary artery, without need for reoperation. CONCLUSION: Short- and mid-term results show that cryopreserved homografts extended by small-sized vascular prostheses can be used safely for Bentall procedures in selected cases where the diameter of the aortic valve annulus is moderately dilated.

Pseudoaneurysm complicating homograft aortic valve replacement--two different--courses and strategies.

The courses of two patients with pseudoaneurysm formation following aortic valve replacement using homografts are reported. Both patients had aortic valve replacement due to acute infective endocarditis with paravalvular abscess formation. The first case had an uneventful postoperative course; but on routine echocardiography, a pseudoaneurysm located at the left coronary commissure was found at one year follow-up. As the pseudoaneurysm was completely asymptomatic, the patient was followed up carefully at six months intervals. The second patient also had pseudoaneurysm formation postoperatively, but he presented one year after surgery with fever and elevated leucocyte count. He had surgical revision of the pseudoaneurysm because no other reason for an infection could be found. Four months later he presented again with infection signs and pseudoaneurysm formation. This time, the homograft was completely excised and another homograft was implanted. One year after the final operation, he is now free from re-infection. The two presented courses show that pseudoaneurysms complicating aortic valve replacement should be managed according to attendant circumstances and symptoms.

Mitral valve surgery utilizing homografts: early results.

BACKGROUND AND AIM OF THE STUDY: Mitral valve repair is superior to prosthetic valve replacement due to preservation of the subvalvular apparatus. We used cryopreserved homografts for mitral valve replacement in selected cases, in whom valve repair would not have been successful. METHODS: Cryopreserved homografts were used in 10 patients (four males, six females; mean age 47 +/- 8 years; range: 27-65 years), for either complete (n = 7) or partial (n = 3) mitral valve replacement. Surgery was indicated due to acute endocarditis (n = 3), stenosis (n = 4) or combined mitral valve disease (n = 3). Transesophageal echocardiography (TEE) was performed to determine correct homograft size; these values were compared with intraoperative measurements of valvular dimensions. The function of the implanted grafts was examined intraoperatively by TEE. Follow up included clinical examination, electrocardiography and echocardiography. RESULTS: All patients survived surgery. Intraoperative TEE revealed mild insufficiency (grade I) in six cases and no insufficiency in four. The papillary muscle-mitral annulus distance was the most reliable preoperative echocardiographic parameter to determine the required homograft. Mean pressure gradients were 3.2 +/- 0.7 mmHg for complete and 2.4 +/- 0.5 mmHg for partial homograft replacement. Sinus rhythm was present in all cases at discharge. Mean follow up was 24 months (range: 6-36 months). At six-month and one-year follow up, the pressure gradients had risen slightly to 3.4 +/- 0.6 mmHg (complete) and 2.8 +/- 0.6 mmHg (partial). To date, four patients had competent grafts, and six presented with an insufficiency (grade I). All patients had normal left ventricular function (EF 65 +/- 6%) at their last follow up; there were no signs of endocarditis during the follow up period. CONCLUSION: Mitral homografts for valve replacement or repair are recommended in selected cases in whom conservative reconstruction techniques are not possible. The avoidance of long-term anticoagulation therapy and preservation of left ventricular geometry are clear advantages to other mitral valve prostheses.

[Homografts for mitral valve replacement in florid endocarditis--an alternative to prosthetic replacement]. / Homografts für den Mitralklappenersatz bei florider Endokarditis--eine Alternative zum prothetischen Ersatz.

INTRODUCTION: Homografts for valve replacement are indicated in acute valve endocarditis. It is assumed that they possess anti-infective properties. Homografts are an established indication in aortic valve replacement. We present our early results with homografts for mitral valve replacement in acute endocarditis. PATIENTS AND METHODS: Between July 1996 and March 1998 we used cryopreserved homografts for mitral valve replacement in seven patients. In three cases (age 24, 42, and 34 years) the indication was an acute endocarditis with subsequent severe mitral valve insufficiency. The size of the required homograft was measured preoperatively using transesophageal echocardiography. For implantation the technique described by A. Carpentier was used; for stabilization of the mitral anulus a valvular ring (Physio) was implanted. Follow-up was done every six months including clinical and echocardiographical examinations. After the first postoperative year an Ultrafast-CT was done in addition. RESULTS: One patient had complete mitral valve replacement, in the other two cases the diseased parts of the valve were completely excised and the valve was repaired using a partial homograft. There were no perioperative deaths. In the follow-ups, up to 24 months of uneventful homograft function was documented by echocardiography; no insufficiency > degree I was seen on color Doppler echocardiography. At the last follow-up (mean follow-up 16 months, range 12 to 24 months) the average mitral valve orifice was 2.5 +/- 0.5 cm2, the mean pressure gradient 2.8 +/- 0.8 mm Hg. In Ultrafast-CT no morphological abnormalities of the mitral valves and no dilatation of the left ventricle were seen. There were no signs of a recurrence of the endocarditis in any patient during the follow-up period. CONCLUSION: Homografts for mitral valve replacement are an interesting alternative to prosthetic valve replacement, especially in younger patients. In cases with acute endocarditis, in which mechanical prosthesis should not be used, a reconstruction or repair of the mitral valve with preservation of the ventricular geometry is possible even if large parts of the mitral valve are infected.

Miniaturized ion-selective chip electrode for sensor application.

The performance of miniaturized potentiometric cells, with multilayer, planar ion-selective sensors in aqueous electrolyte solutions, human serum, urine, and whole blood, is presented. The basic steps of the fabrication with silicon technology are summarized. The effect of the contact surface between the internal reference system and the ion-sensitive membrane on the analytical characteristics of potassium- and calcium-sensitive sensors is studied. Silicone rubber-, high molecular weight PVC-, carboxylated PVC and aliphatic polyurethane (Tecoflex)-based solvent polymeric membranes were dispensed into anisotropically etched wells on silicon wafers, and the resulted planar sensors were tested in terms of their ion sensitivity (slopes of the cell voltage-pK or pCa calibration curves), long-term stability, and reproducibility. For the assay of potassium in whole blood, the miniaturized potentiometric cell was built in a flow-through manifold. To achieve the required precision, the flow conditions were optimized and the sensors calibrated periodically. The results prove the feasibility of the new sensor design and satisfy the particularly difficult requirements for the analysis of biological samples.

Microbiosensors using electrodes made in Si-technology.

The combination of electrochemical transducers made in silicon technology with chemical and biochemical components has been used to manufacture miniaturized sensor structures. Three different types of sensors have been developed and optimized for practical use; (i) an ion-selective sensor, (ii) a glucose enzyme sensor, (iii) a redox-amplifying sensor for immunosensing. The immunodetection based on the redox recycling of mediator molecules is shown for low and high molecular weight analytes. The sensors have been integrated with miniaturized fluidic components and combined with sensor-related electronics and a common microcontroller.

Immunologic reaction and viability of cryopreserved homografts.

Homograft cell viability after cryopreservation was investigated and cytoimmunologic monitoring was performed during the early postoperative course to research possible immunologic reactions after allograft aortic valve replacement. After cryopreservation, morphologic observations were made, a nonradioactive cell proliferation assay was used, and prostaglandin I2 secretion of the remaining endothelial cells was determined. Cytoimmunologic monitoring was performed daily within the first 3 weeks postoperatively. An increase of the activation index greater than 1 was rated as an immunologic reaction. Maintained metabolic activity of graft endothelial cells after cryopreservation was confirmed by prostaglandin I2 release (9.24 +/- 3.48 ng/cm2 basic release and 20.1 +/- 5.76 ng/cm2 when stimulated with 25 mumol/L Na arachidonic acid). Cell proliferation was indicated after graft incubation with the nonradioactive viability kit (0.27 +/- 0.9 at 450 nm). Cytoimmunologic examinations (n = 861) after homograft implantation showed a more intense activation in patients with ABO-incompatible grafts (activation index 2.1 +/- 1.6, n = 16) than in those with ABO-compatible grafts (activation index 1.3 +/- 0.8, n = 17). In these groups, the duration of activation by cytoimmunologic monitoring was 2.8 +/- 1.5 days and 1.3 +/- 0.6 days, respectively (p < 0.041). No activation was observed in 8 patients after xenograft valve replacement (p < 0.01). Our data indicate that cryopreservation of homograft valves represents a cell- and tissue-protective preservation method. Postoperatively, all homograft valves caused immunologic reactions, which were reversible without immunosuppression treatment.

Integrity and viability of homograft valves.

Since it has been suggested that the leaflet tissue viability influences durability after homologous valve replacement, we compared different harvest and preservation techniques in order to examine the quality and smoothness of homograft conservation. We analyzed human aortic and pulmonary valve leaflets obtained from 'heart-beating donors' (HBD) during heart transplantation and from 'non-heart-beating donors' (NHBD) during coroner's autopsies. Valves were either cryopreserved in liquid nitrogen or stored at 4 degrees C in nutrient medium similar to the procedure reported in our protocols of the homograft bank system. All grafts from NHBD had been antibiotically sterilized for 3 days beforehand. Morphological observations were made using light and electron microscopy and, in order to characterize the endothelial cell viability, a non-radioactive cell proliferation assay was used. The PGI2 secretion of the remaining endothelium was defined as the 6-keto-prostaglandin F1 alpha metabolite by an enzyme immunoassay. Observations in the scanning electron microscope revealed that, after cryopreservation, homografts show an almost confluent endothelium when processed within 24 h after harvest from HBD, but lack endothelial cells when obtained from NHBD. Cryopreserved grafts from NHBD exhibited an altered tissue structure with edema and vacuolization within the spongiosa of the leaflets as well as irreversible cell damage when examined under the light and transmission electron microscope. That the metabolic activity of HBD homografts was maintained was confirmed by proven PGI2 secretion (6150 +/- 1200 pg/3 ml M199 after cryopreservation), whereas specimens from NHBD showed a reduced (1950 +/- 730 pg/3 ml M199) and, after cryopreservation, almost no release (P < 0.0001).(ABSTRACT TRUNCATED AT 250 WORDS)

Cytoimmunological monitoring after homograft valve replacement.

Examination results concerning immunological reactions in cryopreserved allograft valves during the early postoperative course are so far not available. Cytoimmunological monitoring (CIM) is a well established method to prove rejection reaction after allograft transplantation and was used in this study. Allograft valves were harvested from patients who underwent heart transplantation, and did not require sterilizing in antibiotic solutions. The valves were dissected, conserved and subsequently frozen to -40 degrees C and stored in a freezing system at -196 degrees C. During the first 3 weeks following allograft implantation, CIM was performed daily. An activation index (AI) was determined from the cytological evaluation of the mononuclear concentrate in the peripheral blood. An increase of the AI > 1 was defined as an immunological reaction. As control we performed 98 CIM examinations in eight patients who underwent bioprosthetic valve replacement in the aortic position. Echocardiography (TTE and TEE) was used postoperatively as function control. Out of 16 patients who underwent cryopreserved aortic valve implantation in the aortic position, 336 CIM-results were obtained. An immunological reaction could be detected in all patients, starting on the 5th day on average. Comparing ABO-compatible (group I, n = 9) with ABO-incompatible (group II, n = 7) allografts, the AI-maximum in group I was 1.4 with a mean duration of 1.5 days. Group II was characterized by more intensive immunoreactions (mean = 2.3) which proved to be even more prolonged (mean = 3.3 days, P < 0.05). Nevertheless it became apparent that all observed immunological events were completely reversible without the need for immunosuppressive drugs.(ABSTRACT TRUNCATED AT 250 WORDS)

[Clinical forms of enzootic pneumonia in calves and their effects on respiration]. / Klinische Formen der enzootischen Pneumonie bei Kälbern und ihre Auswirkungen auf die Atmung.

Enzootic pneumonia was diagnosed in 160 of 225 bullocks in a livestock fattening unit. Within the first 10 weeks, 51 of those affected contracted the disease a second time after the symptoms had vanished. Acute catarrhal bronchopneumonia were most common (49.3%) during the first outbreak, whereas acute catarrhal purulent bronchopneumonia (39.2%) and chronic pneumonia (29.4%) were most common during the second. Arterial blood gas analysis was undertaken for 33 clinically healthy bullocks and 100 bullocks with clinically diagnosed acute bronchitis, acute catarrhal bronchopneumonia, acute catarrhal purulent bronchopneumonia and chronic pneumonia. The pO2, SATO2 and A-aDO2 levels found in blood from the abdominal aorta of all diseased animals differed significantly from those of healthy animals. The values for animals with distinct catarrhal bronchopneumonia and catarrhal purulent bronchopneumonia showed the greatest differences.