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Biomedical subjects

R Bischoff

Publications and source records attributed to R Bischoff.

At least 55 records · Page 3Linked to original sources
PubMed1994

Clinical results of tension band fixation of avulsion fractures of the hand.

We report 100 cases in which a tension-band technique was used to secure fixation of avulsion fractures within the digits. Included are 51 bony mallet fractures that were subluxed or irreducible, 38 displaced bony gamekeeper's fractures of the thumb, 8 fractures of the lateral phalangeal base, and 3 avulsion fractures that involved the base of the dorsal aspect of the middle phalanx. We evaluated results on the basis of clinical and x-ray film criteria. Excellent or satisfactory results were found in all 38 bony gamekeeper's injuries. All but one gamekeeper's fracture healed; the one nonunion was asymptomatic. All eight patients with lateral avulsion fractures had an excellent or satisfactory result on clinical examination. Seven of the eight had an excellent result on x-ray film evaluation; the one malnuion was asymptomatic. Two of the three fractures with dorsal avulsions of the base of the middle phalanx had a satisfactory result on clinical and x-ray film examination; the one poor result was due to the severity of the comminution of the fracture. However, 21 of the 51 bony mallet injuries had a poor result both clinically and radiographically. We noted numerous postoperative complications including dorsal skin breakdown, superficial and deep infection, and secondary displacement of the fragment. Tension-band fixation is an excellent method for treating various avulsion fractures of the hand such as bony gamekeeper's thumb, lateral avlusion injuries, and dorsal avulsions of the middle phalanx. However, the treatment of bony mallet fingers with tension band fixation is less predictable and should be used with caution.

Adult↗
PubMed1994

Heterotopic ossification following uncemented total hip arthroplasty. Effect of the operative approach.

A consecutive series of 112 primary uncemented total hip arthroplasties using the same type of prosthesis was reviewed for the incidence and severity of heterotopic ossification (HO). Sixty-three of these patients underwent a posterior approach (group 1) and 49 underwent an anterolateral approach (group 2). The two groups were virtually identical when risk factors, such as age, sex, estimated blood loss, length of surgery, and diagnosis were compared. Patients in group 1 had a significantly lower incidence and severity of HO than those in group 2. The overall incidence of class III or IV HO was very low (8%), indicating that uncemented total hip arthroplasty in itself is not predisposed to moderate or severe degrees of HO. Patients undergoing uncemented total hip arthroplasty through an anterolateral approach had a higher incidence of HO than those undergoing a posterior approach in this series.

Adult↗
PubMed1994

A new extracellular protein of Pseudomonas aeruginosa PA103 regulated by regA.

The expression of exotoxin A (ExoA) from Pseudomonas aeruginosa is influenced by iron and is under the control of the regulatory gene regA. To test whether regA plays a role in the expression of other iron-regulated proteins a RegA- mutant was constructed by insertional mutagenesis. The polypeptide pattern of this mutant (PA103R) was compared with the parental strain (PA103) and a trans-complemented strain PA103R(pREX18) after growth of the strains in conditions containing low or high concentrations of iron. An iron-regulated 42 kDa protein (RRP) was identified and purified from the culture supernatant of PA103 and PA103R(pREX18) which was missing in PA103R. Database analysis of the N-terminal sequence of this regA-regulated protein (RRP) revealed no similarity to other proteins. Preliminary investigations into the function of RRP revealed that it has no proteolytic or cytotoxic activity. Using two-dimensional electrophoretic analysis of whole cells, a technique which allowed separation of over 600 polypeptides, we were unable to identify any other iron-regulated protein whose expression was regulated by regA.

ADP Ribose Transferases↗
PubMed1994

Alpha 1-antitrypsin Pittsburgh (Met358-->Arg) inhibits the contact pathway of intrinsic coagulation and alters the release of human neutrophil elastase during simulated extracorporeal circulation.

Cardiopulmonary bypass prolongs bleeding time, increases postoperative blood loss, and triggers activation of plasma proteolytic enzyme systems and blood cells referred to as the "whole body inflammatory response". Contact of blood with synthetic surfaces leads to qualitative and quantitative alterations in platelets, neutrophils, contact and complement systems. Contact and complement pathway proteins both induce neutrophil activation. alpha 1-antitrypsin Pittsburgh (Met358-->Arg), a mutant of alpha 1-antitrypsin, is a potent inhibitor of plasma kallikrein and thrombin. We investigated whether this recombinant mutant protein inhibited platelet activation, as well as contact and/or complement-induced neutrophil activation during simulated extracorporeal circulation. Arg358 alpha 1-antitrypsin did not prevent the 34% drop in platelet count at 5 min of recirculation, did not block the 50% decrease in ADP-induced platelet aggregation at 120 min of recirculation, nor inhibit the release of 6.06 +/- 1.07 micrograms/ml beta-thromboglobulin at 120 min of recirculation suggesting that the inhibitor had little effect on platelet activation. However, Arg358 alpha 1-antitrypsin totally blocked kallikrein-C1-inhibitor complex formation but not C1-C1-inhibitor complex formation. Most importantly, Arg358 alpha 1-antitrypsin decreased the release of 1.11 +/- 0.16 micrograms/ml human neutrophil elastase by 43%. The attenuation of neutrophil activation in the absence of an effect on complement activation via the classical pathway, supports the concept that kallikrein is a major mediator of neutrophil degranulation during cardiopulmonary bypass.

Arginine↗
PubMed1993

Sequence-specific deamidation: isolation and biochemical characterization of succinimide intermediates of recombinant hirudin.

Natural hirudin variant 2 with a lysine residue in position 47 (rHV2-Lys47) was produced in a genetically engineered strain of Saccharomyces cerevisiae as a secreted protein of 65 amino acids and purified to greater than 99% homogeneity. Only reversed-phase high-performance liquid chromatography (RP-HPLC) using very shallow acetonitrile gradients indicated the presence of a component in the final product (approximately 1% of total protein) with a slightly increased retention time. Using successive RP-HPLC purification steps, this hydrophobic impurity was isolated and separated into two constituents defined as components A1 and A2 which differed from the parent molecule by mass reductions of 17.2 Da (A1) and 17.6 Da (A2), respectively, as determined by electrospray mass spectrometry (ESMS). Proteolytic digestion with endoprotease Glu-C from Staphylococcus aureus (V8 protease) and analysis of the peptide mixture by ESMS showed that the mass difference between rHV2-Lys47 and component A1 was due to a modification between amino acids 1 and 43, while the corresponding mass difference with component A2 was the result of a modification within the peptide fragment comprising residues 50-61. Further analyses using amino acid sequencing and ESMS in combination with collision-activated dissociation (CAD) detected modifications at residues Asn33-Gly34 in component A1 and at Asn53-Gly54 in component A2. Both of these sites were previously shown to be susceptible to spontaneous deamidation under slightly basic pH conditions. Thus, the mass reductions of approximately 17 Da and the fact that both asparagines, Asn33 in component A1 and Asn53 in component A2, proved to be resistant to Edman degradation provided strong support for them being stable succinimide intermediates of the corresponding deamidation reactions. Both intermediates were shown to have inhibition constants for human alpha-thrombin on the order of 1 pM, identical to that of rHV2-Lys47. The isoelectric point of component A2 was determined to be within 0.01 pH unit of that of the parent molecule by isoelectric focusing in an immobilized pH gradient.

Amides↗
PubMed1993

The use of Texas Scottish-Rite instrumentation in idiopathic scoliosis. A preliminary report.

The Texas Scottish-Rite instrumentation (TSRH) is a new system for the correction of spinal deformities that uses smooth rods and hooks stabilized to the rod with a three-point interference fit. This obviates the need for the break-off hex bolts and should make for easier removal and/or revision. Twenty-three patients with TSRH instrumentation were followed for an average of 18 months after surgery. The major scoliotic curve was corrected an average of 49% with the compensatory curve averaging 37% correction. Loss of thoracic kyphosis was less than 9 degrees although it increased on average 8 degrees in those patients where kyphosis was improved. There were no acute complications, but late complications included four cases of hook pull-out. Results are comparable to those found with the CD instrumentation. The TSRH instrumentation is an attractive alternative in scoliosis surgery, particularly because of the relative ease in which revision surgery may be performed.

Adolescent↗
PubMed1993

Hydrophobic cluster analysis of G protein-coupled receptors: a powerful tool to derive structural and functional information from 2D-representation of protein sequences.

Current methods for comparative analyses of protein sequences are 1D-alignments of amino acid sequences based on the maximization of amino acid identity (homology) and the prediction of secondary structure elements. This method has a major drawback once the amino acid identity drops below 20-25%, since maximization of a homology score does not take into account any structural information. A new technique called Hydrophobic Cluster Analysis (HCA) has been developed by Lemesle-Varloot et al. (Biochimie 72, 555-574), 1990). This consists of comparing several sequences simultaneously and combining homology detection with secondary structure analysis. HCA is primarily based on the detection and comparison of structural segments constituting the hydrophobic core of globular protein domains, with or without transmembrane domains. We have applied HCA to the analysis of different families of G-protein coupled receptors, such as catecholamine receptors as well as peptide hormone receptors. Utilizing HCA the thrombin receptor, a new and as yet unique member of the family of G-protein coupled receptors, can be clearly classified as being closely related to the family of neuropeptide receptors rather than to the catecholamine receptors for which the shape of the hydrophobic clusters and the length of their third cytoplasmic loop are very different. Furthermore, the potential of HCA to predict relationships between new putative and already characterized members of this family of receptors will be presented.

Amino Acid Sequence↗
PubMed1993

The Arg-4 mutant factor IX Strasbourg 2 shows a delayed activation by factor XIa.

We have characterized at the DNA and protein levels a mutant factor IX, factor IX Strasbourg 2, which is responsible for a severe form (< 0.01 U/ml) of haemophilia B. Factor IX Strasbourg 2 has a higher molecular weight than normal factor IX. A mutation G-->A at position 6365 of the gene was demonstrated by DNA sequencing and confirmed by restriction mapping which showed absence of a Hae III site. This leads to the substitution of glutamine for arginine at position -4 of the propeptide. Factor IX Strasbourg 2 was purified from plasma by DEAE Sepharose chromatography and immunoaffinity and relative to normal factor IX, binding of calcium to the mutant protein was clearly reduced in calcium lactate agarose gel. Quantification of gamma-carboxyglutamic acid residues gave about 50% carboxylation as compared to normal factor IX. Microsequencing of the NH2-terminal part of factor IX Strasbourg 2 confirmed the attachment of the propeptide and the mutation Arg-->Gln. Activation of factor IX Strasbourg 2 by purified factor XIa was found to be retarded as compared to normal factor IX, but after activation the mutant factor IXa was able to activate factor X. In conclusion, factor IX Strasbourg 2 circulates with the attached propeptide and shows reduced gamma-carboxylation and delayed activation by factor XIa but a normal capacity to activate factor X after total cleavage by factor XIa.

1-Carboxyglutamic Acid↗
PubMed1992

A novel insect defensin mediates the inducible antibacterial activity in larvae of the dragonfly Aeschna cyanea (Paleoptera, Odonata).

The injection of low doses of bacteria into the aquatic larvae of dragonflies (Aeschna cyanea, Odonata, Paleoptera) induces the appearance in their hemolymph of a potent antibacterial activity. We have isolated a 38-residue peptide from this hemolymph which is strongly active against Gram-positive bacteria and also shows activity against one of the Gram-negative bacteria which was tested. The peptide is a novel member of the insect defensin family of inducible antibacterial peptides, which had so far only been reported from the higher insect orders believed to have evolved 100 million years after the Paleoptera. Aeschna defensin is more potent than defensin from the dipteran Phormia, from which its structure differs in several interesting aspects, which are discussed in the paper.

Amino Acid Sequence↗
PubMed1992

A 17.6 kbp region located upstream of the rabbit WAP gene directs high level expression of a functional human protein variant in transgenic mouse milk.

We have investigated whether DNA regions present in the rabbit whey acidic protein (WAP) promoter/5' flanking sequence could potentially confer, in vivo, high level expression of reporter genes. Transgenic mice were generated expressing a variant of human alpha 1-antitrypsin, which has inhibitory activity against plasma kallikrein under the control of a 17.6 kbp DNA fragment located upstream of the rabbit WAP gene. Up to 10 mg/ml of active and correctly processed recombinant protein were detected in mouse milk, thus suggesting that the far upstream DNA sequences from the rabbit WAP gene might be useful for engineering efficient protein production in the mammary glands of transgenic animals.

Animals↗
PubMed1992

Analysis of recombinant proteins by isoelectric focusing in immobilized pH gradients.

Isoelectric focusing in immobilized pH gradients (IEF-IPG) was used to analyze three different recombinant proteins. Recombinant leech hirudin (65 amino acids, three disulfide bonds) expressed in Saccharomyces cerevisiae as a secreted protein and purified by anion-exchange and reversed-phase chromatography proved to be homogeneous with regard to its isoelectric point (pI). In addition, the theoretical pI, calculated on the basis of the primary structure, corresponded precisely to the measured pI of 4.30. IEF-IPG was further employed to follow the stability of recombinant hirudin at pH 9, indicating that deamidation occurred under these conditions. A variant of recombinant human alpha 1-antitrypsin (AAT) (389 amino acids, one cysteine residue) expressed in Escherichia coli and purified by anion-exchange, metal chelate and hydrophobic-interaction chromatography appeared to be homogeneous by polyacrylamide gel electrophoresis under reducing and denaturing conditions as well as by various high performance liquid chromatography methods. However, some heterogeneity was detected by IEF-IPG between pH 5-6. The measured pI values of 5.43-5.58 were slightly lower than the calculated pI based on the primary structure (5.72). This indicated deamidations of Asn or Gln residues. A recombinant Schistosoma mansoni parasite antigen, p28 (210 amino acids, one cysteine residue) obtained after intracellular expression in Saccharomyces cerevisiae and affinity purification on glutathione agarose was analyzed by IEF-IPG in a pH 7.3-8.3 gradient. It appeared to be heterogeneous with regard to its pI, with the major component having a pI of 7.81 compared to the calculated value of 7.17.(ABSTRACT TRUNCATED AT 250 WORDS)

Antigens, Helminth↗
PubMed1992

Topical application of 13-cis-retinoic acid in the treatment of chronic plaque psoriasis.

Topical retinoids are of potential value in the treatment of psoriasis. The aim of the present study was to find out whether topical application of 13-cis-retinoic acid (13-cis-RA) has an antipsoriatic effect. Nine patients participated in the investigation. In each patient, two comparable psoriatic lesions (5 x 5 cm or more) were selected for treatment with either 13-cis-RA in a 0.1% cream base or with the vehicle only (placebo), using a double-blind approach. The investigation was a left-right within-subject comparison. The lesions were recorded for clinical scores 4 weeks before and after the investigation. Punch biopsies were taken from eight patients before and after treatment and examined using immunohistochemical methods to assess epidermal proliferation and keratinization, and to assess inflammation. Thirteen-cis-RA treatment resulted in a mild decrease of scaling and induration. Erythema however increased. No statistically significant difference in biological effects was achieved between 13-cis-RA and placebo treated lesions and no changes in expression of the immunohistochemical markers were seen.

Administration, Topical↗
PubMed1991

Inhibitory activity and conformational transition of alpha 1-proteinase inhibitor variants.

Several variants of alpha 1-proteinase inhibitor (alpha 1-PI) were investigated by spectroscopic methods and characterized according to their inhibitory activity. Replacement of Thr345 (P14) with Arg in alpha 1-PI containing an Arg residue in position 358 (yielding [Thr345----Arg, Met358----Arg]alpha 1-PI) results in complete loss of its inhibitory activity against human alpha-thrombin; whereas an exchange of residue Met351 (P8) by Glu [( Met351----Glu, Met358----Arg]alpha 1-PI) does not alter activity. [Thr345----Arg, Met358----Arg]alpha 1-PI is rapidly cleaved by thrombin, while [Met358----Arg]alpha 1-PI and [Met351----Glu, Met358----Arg]alpha 1-PI form stable proteinase-inhibitor complexes. The stability of [Thr345----Arg, Met358----Arg]alpha 1-PI against guanidinium chloride denaturation is significantly enhanced compared to wild-type alpha 1-PI, and does not change after cleavage, resembling ovalbumin, a serpin with no inhibitory activity, from which the Thr345----Arg amino acid exchange had been derived. [Met351----Glu, Met358----Arg]alpha 1-PI and [Met358----Arg]alpha 1-PI resemble the wild-type protein in this respect. The CD spectra of intact and cleaved alpha 1-PI variants do not compare well with the wild-type protein, probably reflecting local structural differences. Insertion of a synthetic peptide, which corresponds to residues Thr345----Met358 of human alpha 1-PI, leads to the formation of binary complexes with all variants having the characteristic features of the binary complex between peptide and wild-type protein.

Amino Acid Sequence↗
PubMed1991

Purification and biochemical characterization of recombinant alpha 1-antitrypsin variants expressed in Escherichia coli.

Site-directed variants of alpha 1-antitrypsin (alpha 1AT) expressed in a recombinant strain of Escherichia coli have been isolated with an overall process yield of 50% following tangential flow ultrafiltration, anion-exchange, immobilized metal affinity, and hydrophobic interaction chromatography. The primary structure of the purified variants including the integrity of the N- and C-termini has been verified by electrospray mass spectrometry of the intact molecules (44 kDa) for two of the variants (alpha 1AT Leu-358 and alpha 1AT Ala-357, Arg-358). Complementary classical peptide mapping and automated amino acid sequencing have verified 75% of the primary sequence of alpha 1AT Ala-357, Arg-358. Isoelectric focusing in an immobilized pH gradient revealed some microheterogeneity which proved to be reproducible from one purification batch to another. The isolated variants of alpha 1AT did not show any signs of proteolytic degradation during the purification process and proved to be fully active against their target proteases. The described process also allowed the complete removal of endotoxins from the preparations, opening the possibility to evaluate these novel protease inhibitors for their in vivo efficacy in different animal models of human disease.

Amino Acid Sequence↗
PubMed1991

Expression of active recombinant human alpha 1-antitrypsin in transgenic rabbits.

A DNA construct containing the human alpha 1-antitrypsin gene including 1.5 and 4 kb of 5' and 3' flanking sequences, was microinjected into the pronucleus of rabbit embryos. The recombinant human protein was (a) expressed in the blood circulation of F0 and F1 transgenic rabbits at an average concentration of 1 mg ml-1, (b) shown to be fully active and (c) shown to be separable from its rabbit counterpart. Transgenic rabbits might represent a novel source of human proteins of therapeutic interest.

Animals↗
PubMed1990

Sulphation of hirudin in BHK cells.

Hirudin, a thrombin inhibitor of the leech, was expressed in BHK cells; the alpha 1-antitrypsin signal peptide was used to direct secretion into the culture medium. The recombinant hirudin so produced inhibited thrombin and was shown by labelling experiments with [35S]sulphate to have been posttranslationally modified.

Amino Acid Sequence↗