Ultrasound-guided intermediate cervical plexus and additional peripheral facial nerve block for carotid endarterectomy : A prospective pilot study.

BACKGROUND AND OBJECTIVES: Ultrasound-guided intermediate cervical plexus block with perivascular local anesthetic infiltration is an established anesthetic procedure for carotid endarterectomy. In this prospective pilot study an additional subplatysmal block of the superficial ansa cervicalis is presented for the first time. The target structures are the anastomoses between the facial nerve (cervical and marginal mandibular branches) and cervical plexus. METHODS: An ultrasound-guided intermediate cervical plexus block (20 ml of ropivacaine 0.75%) was performed (n = 28). Then, depending on the individual sonoanatomy, 5 ml of prilocaine 1% was injected into the carotid sheath (group 1: no perivascular infiltration, n = 14, group 2: perivascular infiltration, n = 14). The third step was subplatysmal injection of 5 ml of prilocaine 1% between the medial edge of the sternocleidomastoid muscle and the submandibular gland (n = 28). The investigated parameters included the need for supplementation and block-related side effects. RESULTS: The requirement for supplemental local anesthetic infiltration in the skin incision area was minimal at mean (M) 1.1 ml (standard deviation (SD) ±2.4 ml). Perivascular infiltration in group 2 significantly decreased the total amount of local anesthetic supplemented: group 1 M = 4.2 ml (SD = ±3.1 ml), group 2 M = 1.7 ml (SD = ±2.0 ml) (p = 0.018). The incidence of block-related side effects was not significantly different between the two groups. CONCLUSION: This study presents an ultrasound-guided subplatysmal block of the superficial ansa cervicalis for the first time, with the aim of optimizing anesthesia quality during surgical interventions in the carotid triangle.

Evaluation of a panel of spermatological methods for assessing reprotoxic compounds in multilayer semen plastic bags.

The increase of fertility performance in sows is one of the biggest achievements in pig production over the last 30 years. Nevertheless, pig farms using artificial insemination (AI) repeatedly experienced in recent year's fertility problems with dramatic consequences due to toxic compounds from plastic semen bags. In particular, bisphenol A diglycidyl-ether (BADGE) present in multilayer plastic bags can leach into the semen and could affect the functionality of the spermatozoa. Former studies could not find any alterations in spermatozoa based on the exposure to BADGE. The aim of the study was to evaluate effects of BADGE on boar spermatozoa using an extended panel of spermatological methods. In spring 2019, a large drop in farrowing rates from 92.6 ± 2.3% to 63.7 ± 11.1% in four sow farms in Croatia was detected. In migration studies, BADGE could be identified as a causal toxic compound and leached into the extended semen in concentration of 0.37 ± 0.05 mg/L. Detailed spermatological studies showed that significant predictors for effects on spermatozoa were different levels of motility and kinematic data after a prolonged storage time, thermo-resistance test (prolonged incubation time), mitochondrial activity, membrane integrity and fluidity. No serious effects were observed for sperm morphology and DNA fragmentation. These results provide new insights into the development of a new quality assurance concept for a detailed spermatological examination during testing of plastic materials for boar semen preservation. It could be shown that boar spermatozoa are an excellent biosensor to detect potential toxicity and fertility-relevant compounds.

Depolymerization of lignosulfonates by submerged cultures of the basidiomycete Irpex consors and cloning of a putative versatile peroxidase.

Lignosulfonates are abundantly available byproducts of the paper and pulping industry, and they therefore represent a promising feedstock for new sustainable processes. For industrial applications of lignosulfonates, their molecular weight distribution is a critical factor. In order to decrease the average molecular weight of lignosulfonates, Seventeen basidiomycetes were screened for their capability to depolymerize lignosulfonates from spent sulfite liquor (SSL) in surface and liquid cultures. Five basidiomycetes polymerized the lignosulfonates under the selected conditions. Only Irpex consors was found to efficiently degrade calcium lignosulfonates when SSL (0.5%, w/w) was used as the sole carbon and nitrogen source. The average molecular weight of the lignosulfonates was reduced from ∼26 to ∼4 kDa as determined by size exclusion chromatography (SEC) within two weeks. Various extracellular enzyme activities of I. consors were determined over the culture period. High peroxidase activities were correlating with a high degradation rate and the culture was harvested at the day of highest peroxidase activity. A putative versatile peroxidase was isolated by fast protein liquid chromatography (FPLC) and its encoding cDNA was cloned.

Plasma levels of bactericidal/permeability-increasing protein (BPI) and lipopolysaccharide-binding protein (LBP) during hemodialysis.

Several proteins modify the biological response to lipopolysaccharide (LPS). Both bactericidal/permeability-increasing factor (BPI), a protein stored in neutrophils, and the acute phase protein LPS-binding protein (LBP) bind to LPS; however, BPI inhibits while LBP enhances binding of LPS to leukocytes and subsequent induction of cytokines. We investigated plasma levels of BPI, LBP, elastase and C5a before, during and after hemodialysis (HD). Six patients were dialysed with Cuprophane (Cup) and polysulfone (PS) low-flux dialyzers on two consecutive HD sessions. There was a significant, 10.9 +/- 2.8-fold increase in BPI after 4-hour HD compared to predialysis and a 4.4 +/- 1.6-fold increase in elastase after 4-hour HD using Cup. Plasma levels of BPI and elastase decreased rapidly after the dialysis session. HD with PS resulted in a smaller, but still significant rise in BPI (3.7 +/- 1.6-fold at 4 hours) and elastase (1.69 +/- 0.2-fold at 4 hours). Levels for BPI and elastase were similar in the arterial and venous blood lines of the dialyzer. Plasma levels of LBP did not change during or after the HD session. These data indicate that BPI, but not LBP is released during HD with Cup and to a lesser extent with PS. Activation of neutrophils and release of BPI during HD may influence the biological response to bacterial products possibly introduced during HD.

A laboratory investigation to assess the influence of cement augmentation of screw and plate fixation in a simulation of distal femoral fracture of osteoporotic and non-osteoporotic bone.

The augmentation of fixation with bone cement is increasingly being used in the treatment of severe osteoporotic fractures. We investigated the influence of bone quality on the mechanics of augmentation of plate fixation in a distal femoral fracture model (AO 33 A3 type). Eight osteoporotic and eight non-osteoporotic femoral models were randomly assigned to either an augmented or a non-augmented group. Fixation was performed using a locking compression plate. In the augmented group additionally 1 ml of bone cement was injected into the screw hole before insertion of the screw. Biomechanical testing was performed in axial sinusoidal loading. Augmentation significantly reduced the cut-out distance in the osteoporotic models by about 67% (non-augmented mean 0.30 mm (sd 0.08) vs augmented 0.13 mm (sd 0.06); p = 0.017). There was no statistical reduction in this distance following augmentation in the non-osteoporotic models (non-augmented mean 0.15 mm (sd 0.02) vs augmented 0.15 mm (sd 0.07); p = 0.915). In the osteoporotic models, augmentation significantly increased stability (p = 0.017).

The potential of implant augmentation in the treatment of osteoporotic distal femur fractures: a biomechanical study.

PURPOSE: Osteoporotic fractures of the distal femur are an underestimated and increasing problem in trauma and orthopaedic surgery. Therefore, this study investigates the biomechanical potential of implant augmentation in the treatment of these fractures. METHODS: Twelve osteoporotic surrogate distal femora were randomly assigned to the augmented or non-augmented group. All specimens were fixed using the LCP DF. In the augmented group additionally 1ml Vertecem V+ was injected in each screw hole before screw positioning. The construct represents an AO 33 A3 fracture. Biomechanical testing was performed as sinusoidal axial loading between 50 and 500N with 2Hz for 45,000 cycles, followed by loading between 50 and 750N until failure. RESULTS: The augmented group showed significant higher axial stiffness (36%). Additionally the displacement after 45,000 cycles was 3.4 times lower for the augmented group (0.68±0.2mm vs. 2.28±0.2mm). Failure occurred after 45,130 cycles (SD 99) in all of the non-augmented specimens and in two specimens of the augmented group after 69,675 cycles (SD 1729). Four of the augmented specimens showed no failure. The failure mode of all specimens in both groups was a medial cut-out. CONCLUSIONS: This study shows a promising potential of implant augmentation in the treatment of osteoporotic distal femur fractures.

A sensitive ELISA for the quantitation of human C5a in blood plasma using a monoclonal antibody.

An ELISA for the quantitation of the C5a anaphylatoxin of complement in human plasma is presented which is based on the use of both a monoclonal antibody and polyclonal antibodies to C5a. Its detection limit is 1 ng C5a/ml plasma. It detects no C5a in fresh EDTA-plasma. Complete removal of C5 by an optimized precipitation step prior to the assay procedure is an essential feature of the method.

Bioartificial cartilage.

Cartilage is a highly differentiated tissue. Its three-dimensional composition of cells and matrix is able to resist intensive mechanical loads. The capacity of cartilage tissue for regeneration is limited. Chondrocytes are responsible for matrix production of cartilage tissue. Enzymatic isolation and expansion of chondrocytes with cell culture techniques has been improved in the last years. These cells can be cultured on different three-dimensional culture systems suitable for transplantation to repair localized cartilage defects. Two types of bioresorbable polymer fleece matrices (PLLA and a composite fleece of polydioxanone and polyglactin) and lyophilized dura as a biological carrier are tested. Phenotypic and morphological appearance of the cultured articular rabbit chondrocytes is preserved on all three types of transport media. Production of glycosaminoglycans has been shown by Alcian blue staining, production of collagen by azan staining. Chondroitin 4- and 6-sulfate are detected immunohistochemically in the created constructs. The different carriers have specific characteristics regarding their suitability for the creation of bioartificial cartilage. This tissue is transplantable into articular cartilage defects and could, therefore, improve the minor intrinsic healing capacity of cartilage tissue.

Synthesis of articular cartilage-like tissue in vitro.

Defects in mature articular cartilage do not heal without residues, and therefore they remain a challenging problem in orthopaedic surgery. Modern tissue culture techniques facilitate the synthesis of cartilage-like tissue. A requirement of retaining the phenotypic characteristics of chondrocytes in vitro is the use of three-dimensional culture techniques. Articular chondrocytes of adult rabbits were isolated and cultured on different transplantable media for several weeks. A resorbable fleece, a non-absorbable net and lyophilized dura were used. Viability was tested by immunohistochemical techniques. Deposition of extracellular matrix could be observed by electron microscopy. The phenotypical and morphological appearance of cultured chondrocytes was preserved on the resorbable polymer fleece and the lyophilized dura. Cells cultured on the non-absorbable net had a more fibroblastic appearance. The resorbable fleece is apparently most suitable in terms of viability of the cultured chondrocytes and biocompatibility. The cultured three-dimensional artificial cartilage constructs reveal a future possibility for autologous cartilage transplantation into mature cartilage defects.

Agglutinating and precipitating capacity of rabbit anti-Salmonella typhosa gamma G and gamma M-antibodies during prolonged immunization.

Pike, Robert M. (University of Texas Southwestern Medical School, Dallas), Mary L. Schulze, and Cleo H. Chandler. Agglutinating and precipitating capacity of rabbit anti-Salmonella typhosa gammaG and gammaM antibodies during prolonged immunization. J. Bacteriol. 92:880-886. 1966.-Antibody produced in rabbits immunized with acetone-dried typhoid bacilli was followed over a period of 445 days by agglutination and by quantitative precipitation. Repeated injections of vaccine resulted in suppression of antibody titers. Both gammaG and gammaM antibodies were rapidly increased by booster injections after rest periods during which titers had decreased to low levels. The O agglutinin titers and the amount of antibody protein, as determined by precipitation with endotoxin, generally were parallel, except in serum specimens in which unusually large proportions of the agglutinating activity were found in the gammaG fraction. These exceptions were explained by the greater agglutinating capacity of the gammaM. Endotoxin precipitated about 10 times as much antibody from gammaG preparations as it did from gammaM fractions of equivalent agglutinating strength. A much higher proportion of the serological activity, therefore, was found in the gammaG fractions when antibody was measured by precipitation than when agglutination was used as the measure of activity.

L-fucose residues on cellulose-based dialysis membranes: quantification of membrane-associated L-fucose and analysis of specific lectin binding.

Contact of mononuclear human leukocytes with cellulose dialysis membranes may result in complement-independent cell activation, i.e. enhanced synthesis of cytokines, prostaglandins and an increase in beta 2-micro-globulin synthesis. Cellular contact activation is specifically inhibited by the monosaccharide L-fucose suggesting that dialysis membrane associated L-fucose residues are involved in leukocyte activation. In this study we have detected and quantitated L-fucose on commercially-available cellulose dialysis membranes using two approaches. A sensitive enzymatic fluorescence assay detected L-fucose after acid hydrolysis of flat sheet membranes. Values ranged from 79.3 +/- 3.6 to 90.2 +/- 5.0 pmol cm-2 for Hemophan or Cuprophan respectively. Enzymatic cleavage of terminal alpha-L-fucopyranoses with alpha-L-fucosidase yielded 7.7 +/- 3.3 pmol L-fucose per cm2 for Cuprophan. Enzymatic hydrolysis of the synthetic polymer membranes AN-69 and PC-PE did not yield detectable amounts of L-fucose. In a second approach, binding of the fucose specific lectins of Lotus tetragonolobus and Ulex europaeus (UEAI) demonstrated the presence of biologically accessible L-fucose on the surface of cellulose membranes. Specific binding was observed with Cuprophan, and up to 2.6 +/- 0.3 pmol L-fucose per cm2 was calculated to be present from Langmuir-type adsorption isotherms. The data presented are in line with the hypothesis that surface-associated L-fucose residues on cellulose dialysis membranes participate in leukocyte contact activation.

Comparative study of C5a plasma levels with different hemodialysis membranes using an enzyme-linked immunosorbent assay.

Using a specific and sensitive ELISA for C5a, the present study shows that predialysis levels of C5a in end-stage renal disease patients are not elevated and that all membranes studied (Cuprophan, Hemophan, Gambrane and hydrophilic polysulfone) cause significant increases of C5a plasma levels albeit to different degrees. Higher increases of C5a were accompanied by larger decreases of circulating granulocytes and monocytes. Dialyzers with a newly introduced modified regenerated cellulose membrane, Hemophan, showed lower C5a plasma levels during hemodialysis than Cuprophan. With Hemophan C5a plasma levels were comparable to those with polysulfone membranes.

Glycosylphosphatidylinositol-specific phospholipase D of human serum--activity modulation by naturally occurring amphiphiles.

The enzymatic properties of glycosylphosphatidylinositol-specific phospholipase D (EC 3.1.4.50) were characterized using a 6,000-fold purified enzyme. This was obtained in 100 microg amounts from human serum with a recovery of 35%. Pure alkaline phosphatase containing one anchor moiety per molecule was used as substrate. The enzyme is stimulated by n-butanol, but in contrast to other phospholipases this activation is not produced by a transphosphatidylation reaction. The previously reported non-linearity of the specific activity with respect to phospholipase concentration in the test was no longer observed upon purification, indicating inhibitor removal. The serum inhibitor(s) co-chromatograph with serum proteins and lipoproteins. The main part of the inhibitory activity was found in the lipid fraction after protein denaturation and can be subfractionated into acid phospholipids, cholesteryl esters and triacylglycerides. Added phosphatidyl-serine, phosphatidylinositol, phosphatidylglycerol, gangliosides, cholesteryl esters, and sphingomyelins turned out to be strong inhibitors, as well as phosphatidic acid. Phosphatidylethanolamine and various monoacylglycerols were found to be activators. The low glycosylphosphatidylinositol-specific phospholipase activity found in native serum did not increase significantly upon 90% removal of phospholipids by n-butanol. High serum concentrations of strongly inhibiting compounds, complex kinetic interactions among aggregates of these substances, and compartmentalization effects are discussed as possible reasons for the observed inactivity.

Gene expression of interleukin-1 beta during hemodialysis.

It is still controversial whether the hemodialysis (HD) procedure is an inflammatory process in vivo. Therefore, we studied the gene expression of interleukin-1 beta (IL-1 beta) as a marker of inflammation in peripheral blood mononuclear cells (PBMC) of patients during HD by Northern blotting and polymerase chain reaction. Compared to PBMC separated pre-HD (1.0 densitometric units), the amount of IL-1 beta mRNA was increased in PBMC leaving the dialyzer (12.2 +/- 2 densitometric units, P < 0.01), but was not increased in PBMC re-entering the dialyzer from the systemic circulation (0.6 +/- 0.1 densitometric units) in all 12 patients studied. The maximal amount of IL-1 beta mRNA in PBMC was seen at five minutes after start of HD. There was a significant correlation between the increase in IL-1 beta mRNA and the increase in activated complement C5a (r = 0.71, P < 0.01). HD using less complement-activating membranes (hemophan, polysulfone, polyamide or polyacrylonitrile) resulted in no detectable IL-1 beta mRNA. Furthermore, a monoclonal antibody against human C5a reduced the increase in IL-1 beta mRNA by 83% (P < 0.05), indicating that C5a plays a major role for induction of IL-1 beta mRNA during HD. This study demonstrates that during HD with regenerated cellulose, gene expression for IL-1 beta takes place in PBMC.

Permeability of dialyzer membranes to TNF alpha-inducing substances derived from water bacteria.

Pro-inflammatory cytokine-inducing substances derived from cultured E. coli have previously been shown to pass across low-flux regenerated cellulosic dialyzer membranes. In the present study, a sterile filtrate of Pseudomonas maltophilia grown from standard bicarbonate dialysis fluid was used to test the permeability of various dialyzer membranes (regenerated cellulose, cellulose triacetate, polyacrylonitrile, polysulfone and polyamide) to TNF alpha-inducing bacterial substances. Pyrogen-free tissue culture medium (MEM) was recirculated for 60 minutes in the dialysate compartment of a closed-loop dialysis system, then P. maltophilia filtrate was added and recirculation was continued for a further hour. Samples from the dialysate (MEM) and the blood side (containing 10% human plasma in MEM) were incubated with donor mononuclear cells (MNC) for 18 hours and TNF alpha release was measured in MNC supernatants by radioimmunoassay. Five minutes after the addition of P. maltophilia filtrate, mean TNF alpha-inducing activity in the dialysate increased from (mean +/- SEM) 0.10 +/- 0.02 to 18.2 +2- 1.5 (ng/2.5 x 10(6) MNC/18 hr). TNF alpha-inducing activity in the blood side increased with regenerated cellulose from 0.10 +/- 0.01 to 4.57 +/- 1.55 (N = 8; P less than 0.001); with cellulose triacetate from 0.20 +/- 0.05 to 0.44 +/- 0.10 (N = 5; P less than 0.05), and with polyacrylonitrile from 0.10 +/- 0.02 to 1.16 +/- 0.45 (N = 5; P less than 0.03). No increased TNF alpha-inducing activity was observed in the blood side of polysulfone (N = 5) or polyamide dialyzers (N = 5).(ABSTRACT TRUNCATED AT 250 WORDS)