[Diagnostics and surgical treatment of painful neuromas]. / Diagnostik und chirurgische Therapie schmerzhafter Neurome.

BACKGROUND: Painful neuromas that often develop after peripheral nerve injury require adequate diagnosis and treatment because of the suffering they cause. The scientific basis for the development of painful neuromas has not yet been sufficiently investigated. In addition to conservative procedures, a larger number of surgical techniques are available for treatment of painful neuromas. OBJECTIVE: A review of the basic principles, diagnostic and treatment options for painful neuromas. MATERIAL AND METHODS: Presentation of the scientific basis regarding the development of painful neuromas. Illustration and discussion of the most common diagnostic and treatment procedures. RESULTS: The scientific basis regarding the development of painful neuromas after peripheral nerve injury has not yet been adequately developed. In order to be able to make a correct diagnosis, the use of standardized diagnostic criteria and adequate imaging techniques are recommended. In the sense of a paradigm shift, the use of the formerly neuroma-bearing nerve for reinnervation of target organs is to be preferred over mere burying in adjacent tissue. CONCLUSION: In addition to standardized diagnostics the management of painful neuromas often requires a surgical intervention after all conservative therapeutic measures have been exhausted. As an alternative to restoring the continuity of the injured nerve, targeted reinnervation of electively denervated target organs by the formerly neuroma-bearing nerve is preferable over other techniques.

3D printed patient-specific fixation plates for the treatment of slipped capital femoral epiphysis: Topology optimization vs. conventional design.

Orthopedic plates are commonly used after osteotomies for temporary fixation of bones. Patient-specific plates have recently emerged as a promising fixation device. However, it is unclear how various strategies used for the design of such plates perform in comparison with each other. Here, we compare the biomechanical performance of 3D printed patient-specific bone plates designed using conventional computer-aided design (CAD) techniques with those designed with the help of topology optimization (TO) algorithms, focusing on cases involving slipped capital femoral epiphysis (SCFE). We established a biomechanical testing protocol to experimentally assess the performance of the designed plates while measuring the full-field strain using digital image correlation. We also created an experimentally validated finite element model to analyze the performance of the plates under physiologically relevant loading conditions. The results indicated that the TO construct exhibited higher ultimate load and biomechanical performance as compared to the CAD construct, suggesting that TO is a viable approach for the design of such patient-specific bone plates. The TO plate also distributed stress more evenly over the screws, likely resulting in more durable constructs and improved anatomical conformity while reducing the risk of screw and plate failure during cyclic loading. Although differences existed between finite element analysis and experimental testing, this study demonstrated that finite element modelling can be used as a reliable method for evaluating and optimizing plates for SCFE patients. In addition to enhancing the mechanical performance of patient-specific fixation plates, the utilization of TO in plate design may also improve the surgical outcome and decrease the recovery time by reducing the plate and incision sizes.

Design considerations for patient-specific bone fixation plates: a literature review.

In orthopedic surgery, patient-specific bone plates are used for fixation when conventional bone plates do not fit the specific anatomy of a patient. However, plate failure can occur due to a lack of properly established design parameters that support optimal biomechanical properties of the plate.This review provides an overview of design parameters and biomechanical properties of patient-specific bone plates, which can assist in the design of the optimal plate.A literature search was conducted through PubMed and Embase, resulting in the inclusion of 78 studies, comprising clinical studies using patient-specific bone plates for fracture fixation or experimental studies that evaluated biomechanical properties or design parameters of bone plates. Biomechanical properties of the plates, including elastic stiffness, yield strength, tensile strength, and Poisson's ratio are influenced by various factors, such as material properties, geometry, interface distance, fixation mechanism, screw pattern, working length and manufacturing techniques.Although variations within studies challenge direct translation of experimental results into clinical practice, this review serves as a useful reference guide to determine which parameters must be carefully considered during the design and manufacturing process to achieve the desired biomechanical properties of a plate for fixation of a specific type of fracture.

MicroRNA miR-371a-3p in serum of patients with germ cell tumours: evaluations for establishing a serum biomarker.

As only 60% of the patients with germ cell tumour (GCT) express the classical markers, new markers as for example microRNAs (miRNAs) are required. One promising candidate is miR-371a-3p, but data are sparse to date. We measured serum levels of miR-371a-3p in GCT patients, in controls, and in cases with other malignancies. We also assessed the expression in other body fluids and we looked to the decline of serum miR-371a-3p levels after treatment. miR-371a-3p levels were measured by quantitative polymerase chain reaction in serum samples of 25 GCT patients, 6 testicular intraepithelial neoplasia (TIN) patients, 20 healthy males and 24 non-testicular malignancies (NTMs). Testicular vein blood (TVB) was examined in five GCT patients and five controls. Five GCT patients had serial daily measurements after orchiectomy. Five seminal plasma samples, three urine specimens and one pleural effusion fluid were processed likewise. GCT patients had significantly higher miR-371a-3p serum levels than controls and NTMs. Serum levels of controls, TINs and NTMs were not significantly different. TVB samples of GCT patients had 65.4-fold higher serum levels than peripheral blood. Malignant pleural effusion fluid had extremely high levels of miR-371a-3p, seminal plasma had strongly elevated levels by comparison with serum levels of controls. In urine of GCT patients, no miR-371a-3p expression was detected. Daily measurements after orchiectomy in stage 1 patients revealed a decline by 95% within 24 h. Serum levels of miR-371a-3p appear to be a promising specific biomarker of GCTs as is suggested by high serum levels in GCT patients, the rapid return of elevated levels to normal range after treatment, the association of serum levels with tumour bulk, the non-expression in NTMs and the much higher levels of miR-371a-3p in TVB. This potential marker deserves further exploration in a large-scale clinical study.

Plasmidic versus insertional cloning of heterologous genes in Mycobacterium bovis BCG: impact on in vivo antigen persistence and immune responses.

Bivalent recombinant strains of Mycobacterium bovis BCG (rBCG) expressing the early regulatory nef and the structural gag(p26) genes from the simian immunodeficiency virus (SIV) SIVmac251 were engineered so that both genes were cotranscribed from a synthetic operon. The expression cassette was cloned into a multicopy-replicating vector, and the expression levels of both nef and gag in the bivalent rBCG(nef-gag) strain were found to be comparable to those of monovalent rBCG(nef) or rBCG(gag) strains. However, extrachromosomal cloning of the nef-gag operon into a replicative plasmid resulted in strains of low genetic stability that rapidly lost the plasmid in vivo. Thus, the nef-gag operon was inserted site specifically into the BCG chromosome by means of mycobacteriophage Ms6-derived vectors. The resulting integrative rBCG(nef-gag) strains showed very high genetic stability both in vitro and in vivo. The in vivo expression of the heterologous genes was much longer lived when the expression cassette was inserted into the BCG chromosome. In one of the strains obtained, integrative cloning did not reduce the expression levels of the genes even though a single copy was present. Accordingly, this strain induced cellular immune responses of the same magnitude as that of the replicative rBCG strain containing several copies of the genes.

Phosphorylation of Mycobacterium leprae heat-shock 70 protein at threonine 175 alters its substrate binding characteristics.

We have examined the functional properties including autophosphorylation of the Mycobacterium leprae Hsp70 homologue. Recombinant M. leprae Hsp70 had pH optima for its adenosine triphosphatase and autophosphorylating activities which were near pH 8 and 6, respectively. Both these activities were inhibited by reduced and alkylated bovine pancreatic trypsin inhibitor, but not other tested substrates. Autophosphorylation was augmented by up to 25 mM Ca2+. Using site-directed mutagenesis to construct two Thr-->Ala mutants at positions 175 and 193, and phosphoamino acid analysis, it was shown that Thr175 was the dominant threonine residue autophosphorylated in M. leprae Hsp70. Phosphorylation led to an increased affinity for a model polypeptide substrate, reduced and alkylated bovine albumin. These properties are compared with those of the DnaK protein of Escherichia coli.

The crystal structure of the liver fatty acid-binding protein. A complex with two bound oleates.

The crystal structure of the recombinant form of rat liver fatty acid-binding protein was completed to 2.3 A and refined to an R factor of 19.0%. The structural solution was obtained by molecular replacement using superimposed polyalanine coordinates of six intracellular lipid-binding proteins as a search probe. The entire amino acid sequence of rat liver fatty acid-binding protein along with an amino-terminal formyl-methionine was modeled in the crystal structure. In addition, the crystal was obtained in the presence of oleic acid, and the initial electron density clearly showed two fatty acid molecules bound within a central cavity. The carboxylate of one fatty acid molecule interacts with arginine 122 and is shielded from free solvent. It has an overall bent conformation. The more solvent-exposed carboxylate of the other oleate is located near the helix-turn-helix that caps one end of the beta-barrel, while the acyl chain lies in the interior. The cavity contains both polar and nonpolar residues but also shows extensive hydrophobic character around the nonpolar atoms of the ligands. The primary and secondary oleate binding sites appear to be totally interdependent, mainly because favorable hydrophobic interactions form between both aliphatic chains.

Recombinant Mycobacterium bovis BCG producing the N-terminal half of SIVmac251 Env antigen induces neutralizing antibodies and cytotoxic T lymphocyte responses in mice and guinea pigs.

Recombinant Mycobacterium bovis bacillus Calmette-Guérin (rBCG) represents a good candidate for the development of vaccines against AIDS. Several HIV or SIV genes including nef, gag, and env have already been expressed by rBCG strains and shown to induce strong humoral and cellular immune responses in experimental animals. Because a broad immune response directed to multiple HIV/SIV antigens is highly desirable in order to develop effective vaccines, we have also investigated the immune response induced by an rBCG strain expressing a large N-terminal portion of the SIVmac251 Env gp110-encoding gene. The rBCG(SIVmac251Env) strain obtained was able to induce strong CTL responses in mice as well as humoral immune responses in mice and guinea pigs immunized by parenteral routes. The anti-gp110 IgGs produced were able to neutralize in vitro growth of virulent SIVmac251 field isolates. Moreover, guinea pigs immunized by the oral route produced significant levels of anti-gp110 IgAs in the feces, demonstrating that rBCG is able to induce local humoral immunity in the intestinal mucosa. These data provide further evidence of the utility of BCG as a candidate vaccine vector against AIDS.

Urease activity does not contribute dramatically to persistence of Mycobacterium bovis bacillus Calmette-Guérin.

Multiplication of BCGure-, an isogenic urease-negative mutant of Mycobacterium bovis BCG constructed by allelic exchange (J. M. Reyrat, F. X. Berthet, and B. Gicquel, Proc. Natl. Acad. Sci. USA 92:8768-8772, 1995), was examined in human macrophages and mice. Although ureolytic activity was not essential to BCGure-growth, a slight decrease in the multiplication and persistence of the mutated strain compared with wild-type BCG was observed in lungs of infected mice.

Recombinant BCG strains expressing the SIVmac251nef gene induce proliferative and CTL responses against nef synthetic peptides in mice.

CTL responses are known to be important for the control of HIV and SIV infections. Such responses are targeted against various components of these viruses including regulatory proteins like Nef. The SIVmac251nef gene was cloned in Mycobacterium bovis BCG under the control of P(AN), a promoter from Mycobacterium paratuberculosis. Nef was expressed as a fused polypeptide with ORF2, an open reading frame adjacent to P(AN). Mice inoculated with rBCG(SIVmac251nef) exhibited proliferative and CD8+ cytotoxic T-cell (CTL) responses against several Nef synthetic peptides. A mapping of the epitopes recognized by CTLs revealed that the central region of Nef is mainly involved in responses. This region had already been demonstrated to induce CTLs in experimentally SIV-infected macaques as well as in HIV-infected individuals. These results demonstrate the feasibility of constructing BCG vaccine strains expressing nef for eliciting cytotoxic responses.

Mechanisms of persistence of mycobacteria.

Pathogenic mycobacteria use a variety of mechanisms to survive and replicate within mononuclear phagocytic cells, including avoidance of early direct activation of macrophages, interference with gamma-interferon-mediated activation and inhibition of bactericidal products. Developments in genetic manipulation should allow the genes involved in mycobacterial virulence and intracellular survival to be identified. Understanding these mechanisms may lead to more effective treatment and prevention of mycobacterial infections.

Regulation of the biosynthesis of subgroup C adenovirus protein IVa2.

The IVa2 gene is located between 16 and 11.3 map units on the left strand of the adenovirus type 5 (Ad5) genome. The coded RNA contains an intron of 277 nucleotides. To determine whether protein IVa2 is synthetized during productive infection and to obtain an immunological reagent to study its function, we prepared antibodies directed to 414 amino acids of protein IVa2 fused to the N-terminal domain of Staphylococcus aureus protein A. Western immunoblot analysis of viral proteins demonstrates that protein IVa2 is a minor component of mature viral particles and that it is also present in assembly intermediates and young virions. Thus, contrary to a previous report (H. Persson, B. Mathisen, L. Philipson, and U. Pettersson, Virology 93:198-208, 1979), protein IVa2 is not related to the 50-kDa polypeptide, a scaffolding protein present in assembly intermediates. The biosynthesis of protein IVa2 during productive infection was examined. Time course studies using immunofluorescence analysis with polyclonal antibodies targeted to protein IVa2 revealed that this protein is first synthesized at 12 h in a few cells exhibiting very striking fluorescence. Synthesis continues until at least 24 h postinfection. When hydroxyurea is added, protein IVa2 is not detected. In cells infected with mutant H5 ts125, blocked at the nonpermissive temperature (40 degrees C) in viral DNA replication, protein IVa2 is overexpressed. These results suggest that protein IVa2 synthesis requires cellular rather than viral DNA replication. RNase protection assay results indicate that hydroxyurea inhibits protein IVa2 synthesis at the transcriptional level. Thus, overexpression of protein IVa2 in H5 ts125-infected cells may be regulated at the translational level.

Cellular transformation by E1 genes of enteric adenoviruses.

The ability of Ad40 and Ad41 E1A plus E1B genes to transform BRK cells was considerably lower than that of Ad5 and Ad12 corresponding genes. However, as for Ad5, the E1A genes of enteric adenoviruses could cooperate with an activated ras oncogene for full cell transformation and the Ad41 E1B could be complemented by E1A gene of Ad5 or Ad12 for cell transformation. Complementation studies suggested that the conserved region 1 of Ad41 E1A was responsible for this inefficient transformation. The Ad40- and Ad41-transformed cell lines exhibited a low level of major histocompatibility complex (MHC) class I antigens correlated to the low level of Ad12-transformed cells. Class I MHC antigen amounts expressed at the surface of the cells transformed by the weakly oncogenic Ad3 were between the high level of Ad5- and the low level of Ad12-transformed cells.

Characterization of crystalline rat liver fatty acid binding protein produced in Escherichia coli.

The principal absorptive cell of the rat small intestinal epithelium contains two homologous cytosolic proteins that bind long chain fatty acids. These are known as intestinal and liver fatty acid binding proteins (FABP). While their precise physiological roles have not been defined, they are believed to represent a multifunctional cytosolic transport system that is involved in the trafficking of exogenous lipids to sites of metabolic processing. 13C NMR studies have revealed differences in their fatty acid binding stoichiometries, binding mechanisms, and the ionization properties of bound fatty acids. To understand the functional differences, liver FABP has been crystallized for eventual comparison with the known crystal structure of intestinal FABP. The lattice type is trigonal with unit cell dimensions of a = b = 84.1 A and c = 44.2 A. The space group as determined by examination of the Patterson symmetry is either P3(1)21 or P3(2)21.

Adenovirus transformed monkey cell lines permissive to enteric adenovirus type 40.

Plasmids containing the E1 regions of adenovirus serotypes 3 and 5 were transfected into primary Rhesus monkey kidney cells. The presence of viral DNA sequences was detected in transformed cell lines. All these cell lines expressed the E1A proteins. In addition, Ad5 transformed cells, have the E1B 21 kDa protein located in the nuclear membrane. These cell lines were permissive to the enteric adenovirus serotype 40 but not to serotype 41.

[Microscopically controlled excision of eyelid tumors (author's transl)]. / Mikroskopisch kontrollierte Exzision von Lidtumoren.

The completeness of excision is of decisive significance for surgical success in eyelid tumor treatment. Microscopically controlled excision is an approach that guarantees complete removal of the tumor while conserving as much healthy tissue as possible. In this way reconstruction can be simplified in many cases. The authors report on three cases.