Diversity and function of mutations in p450 oxidoreductase in patients with Antley-Bixler syndrome and disordered steroidogenesis

P450 oxidoreductase (POR) is the obligatory flavoprotein intermediate that transfers electrons from reduced nicotinamide adenine dinucleotide phosphate (NADPH) to all microsomal cytochrome P450 enzymes. Although mouse Por gene ablation causes embryonic lethality, POR missense mutations cause disordered steroidogenesis, ambiguous genitalia, and Antley-Bixler syndrome (ABS), which has also been attributed to fibroblast growth factor receptor 2 (FGFR2) mutations. We sequenced the POR gene and FGFR2 exons 8 and 10 in 32 individuals with ABS and/or hormonal findings that suggested POR deficiency. POR and FGFR2 mutations segregated completely. Fifteen patients carried POR mutations on both alleles, 4 carried mutations on only one allele, 10 carried FGFR2 or FGFR3 mutations, and 3 patients carried no mutations. The 34 affected POR alleles included 10 with A287P (all from whites) and 7 with R457H (four Japanese, one African, two whites); 17 of the 34 alleles carried 16 "private" mutations, including 9 missense and 7 frameshift mutations. These 11 missense mutations, plus 10 others found in databases or reported elsewhere, were recreated by site-directed mutagenesis and were assessed by four assays: reduction of cytochrome c, oxidation of NADPH, support of 17alpha-hydroxylase activity, and support of 17,20 lyase using human P450c17. Assays that were based on cytochrome c, which is not a physiologic substrate for POR, correlated poorly with clinical phenotype, but assays that were based on POR's support of catalysis by P450c17--the enzyme most closely associated with the hormonal phenotype--provided an excellent genotype/phenotype correlation. Our large survey of patients with ABS shows that individuals with an ABS-like phenotype and normal steroidogenesis have FGFR mutations, whereas those with ambiguous genitalia and disordered steroidogenesis should be recognized as having a distinct new disease: POR deficiency.

Ankle dorsiflexion arthrodesis to salvage Chopart's amputation with anterior skin insufficiency

BACKGROUND In Chopart-level amputations the heel often deviates into equinus and varus when, due to the lack of healthy anterior soft tissue, rebalancing tendon transfers to the talar head are not possible. Consequently, anterior and lateral wound dehiscence and ulceration may occur requiring higher-level amputation to achieve wound closure, with considerable loss of function for the patients. METHODS Twenty-four consecutive patients (15 diabetes, 6 trauma, and 3 tumor) had Chopart's amputation and simultaneous or delayed additional ankle dorsiflexion arthrodesis to allow for tension-free wound closure or soft tissue reconstruction, or to treat secondary recurrent ulcerations. Percutaneous Achilles tendon lengthening and subtalar arthrodesis were added as needed. Wound healing problems, time to fusion and full weight-bearing in the prosthesis, complications in the prosthesis, and the ambulatory status were assessed. Satisfaction and function were evaluated by the AmpuPro score and the validated Prosthesis Evaluation Questionnaire scale. RESULTS Five patients had successful soft tissue healing and fusions but died of their underlying disease 2 to 46 months after the operation. Two diabetic patients required a transtibial amputation. The other 17 patients were followed for 27 months (range, 13-63). The average age of the 4 women and 13 men was 53.9 years (range, 16-87). Postoperative complications included minor wound healing problems in 8 patients, wound breakdown requiring revision in 4, phantom pain in 3, residual equinus in 1, and adjacent scar carcinoma in 1 patient. The time to full weight-bearing in the prosthesis ranged from 6 to 24 weeks (mean 10). The mean AmpuPro score was 107 points (of 120), and the mean Prosthesis Evaluation Questionnaire scale was 147 points (of 200). No complications occurred with the prosthesis. Twelve patients lost 1 to 2 mobility classes (mean 0.9). The arthrodeses all healed within 2.5 months (range, 1.5 to 5 months). CONCLUSION Adding an ankle arthrodesis to a Chopart's amputation either immediately or in a delayed fashion to treat anterior soft tissue complications was a successful salvage in most patients at this amputation level. It enabled the patients to preserve the advantages of a full-length limb with terminal weight-bearing. LEVEL OF EVIDENCE Level IV, retrospective case series.

Is there good simulation basic training for end-to-side vascular microanastomoses?

BACKGROUND Microvascular anastomosis is the cornerstone of free tissue transfers. Irrespective of the microsurgical technique that one seeks to integrate or improve, the time commitment in the laboratory is significant. After extensive previous training on several animal models, we sought to identify an animal model that circumvents the following issues: ethical rules, cost, time-consuming and expensive anesthesia, and surgical preparation of tissues required to access vessels before performing the microsurgical training, not to mention that laboratories are closed on weekends. METHODS Between January 2012 and April 2012, a total of 91 earthworms were used for 150 microsurgical training exercises to simulate vascular end-to-side microanastomosis. The training sessions were divided into ten periods of 7 days. Each training session included 15 simulations of end-to-side vascular microanastomoses: larger than 1.5 mm (n=5), between 1.0 and 1.5 mm (n=5), and smaller than 1.0 mm (n=5). A linear model with the main variables being the number of weeks (as a numerical covariate) and the size of the animal (as a factor) was used to determine the trend in time of anastomosis over subsequent weeks as well as the differences between the different size groups. RESULTS The linear model shows a significant trend (p<0.001) in time of anastomosis in the course of the training, as well as significant differences (p<0.001) between the groups of animals of different sizes. For microanastomoses larger than 1.5 mm, the mean anastomosis time decreased from 19.3±1.0 to 11.1±0.4 min between the first and last week of training (decrease of 42.5%). For training with smaller diameters, the results showed a decrease in execution time of 43.2% (diameter between 1.0 and 1.5 mm) and 40.9% (diameter<1.0 mm) between the first and last periods. The study demonstrates an improvement in the dexterity and speed of nodes execution. CONCLUSION The earthworm appears to be a reliable experimental model for microsurgical training of end-to-side microanastomoses. Its numerous advantages are discussed here and we predict training on earthworms will significantly grow and develop in the near future. LEVEL OF EVIDENCE III This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

The Critical Friends Group: An Innovative Way to Build Intercultural Competence Among Student and Faculty Groups

Introduction Few physicians involved in medical education are likely to have had formal training in teaching. One pedagogical method that can enhance relationships, thus improve teaching and learning is the Critical Friends Group (CFG). The CFG is a collegial support team that offers improved understanding of others. Unconditional high regard for team members frames the interactions in the CFG. These teams could be used to reduce bias and enhance intercultural competence among student CFGs and faculty CFGs. [See PDF for complete abstract]

Enterococcus faecalis PcfC, a spatially localized substrate receptor for type IV secretion of the pCF10 transfer intermediate.

Upon sensing of peptide pheromone, Enterococcus faecalis efficiently transfers plasmid pCF10 through a type IV secretion (T4S) system to recipient cells. The PcfF accessory factor and PcfG relaxase initiate transfer by catalyzing strand-specific nicking at the pCF10 origin of transfer sequence (oriT). Here, we present evidence that PcfF and PcfG spatially coordinate docking of the pCF10 transfer intermediate with PcfC, a membrane-bound putative ATPase related to the coupling proteins of gram-negative T4S machines. PcfC and PcfG fractionated with the membrane and PcfF with the cytoplasm, yet all three proteins formed several punctate foci at the peripheries of pheromone-induced cells as monitored by immunofluorescence microscopy. A PcfC Walker A nucleoside triphosphate (NTP) binding site mutant (K156T) fractionated with the E. faecalis membrane and also formed foci, whereas PcfC deleted of its N-terminal putative transmembrane domain (PcfCDelta N103) distributed uniformly throughout the cytoplasm. Native PcfC and mutant proteins PcfCK156T and PcfCDelta N103 bound pCF10 but not pcfG or Delta oriT mutant plasmids as shown by transfer DNA immunoprecipitation, indicating that PcfC binds only the processed form of pCF10 in vivo. Finally, purified PcfCDelta N103 bound DNA substrates and interacted with purified PcfF and PcfG in vitro. Our findings support a model in which (i) PcfF recruits PcfG to oriT to catalyze T-strand nicking, (ii) PcfF and PcfG spatially position the relaxosome at the cell membrane to stimulate substrate docking with PcfC, and (iii) PcfC initiates substrate transfer through the pCF10 T4S channel by an NTP-dependent mechanism.

The Family Preservation Philosophy and Therapy With Lesbian Clients

Family preservation is generally viewed in terms of a rather narrow practice definition. However, it's underlying philosophy offers a strong framework for building a positive, nonbiased helping alliance with lesbian clients in a therapeutic setting. The family preservation philosophy offers a unique heuristic for helping professionals to work with lesbians. Family preservation values teach that the therapist must start with the client's reality, recognize the particular needs of that client, and use the client's strengths in treatment. Also inherent in this perspective is respect and sensitivity to the lesbian client's "cultural context, experience, and history" (Family Preservation Institute, 1995). In other words, in the family preservation philosophy there is no assumption of heterosexuality in the therapeutic relationship; rather there is an assumption of unconditional positive regard. Further, clients are engaged in a dialogue and encouraged to name the challenges they encounter in their own words, from their own perspective. All of these principles will help empower lesbian clients. Lesbians may avoid traditional mental health services in times of need, preferring to depend on alternative services or friendship support networks. The choice not to seek help through mainstream agencies may be based on previous negative experience or on an assumption of the homophobic attitudes which are often inherent in such services. Traditional services are usually based on the medical model. Services based on the family preservation philosophy, however, have the capability of creating therapeutic relationships in which there is no assumption of heterosexuality, where the lesbian client is respected and viewed as a whole, healthy individual.

Expression, induction and regulation of the cytochrome P450 monooxygenase system in the rat glioma C6 cell line

The cytochrome P450 monooxygenase system consists of NADPH- cytochrome P450 reductase (P450 reductase) and cytochromes P450, which can catalyze the oxidation of a wide variety of endogenous and exogenous compounds, including steroid hormones, fatty acids, drugs, and pollutants. The functions of this system are as diverse as the substrates. P450 reductase transfers reducing equivalents from NADPH to P450, which in turn catalyzes metabolic reactions. This enzyme system has the highest level of activity in the liver. It is also present in other tissues, including brain. The functions of this enzyme system in brain seem to include: neurotransmission, neuroendocrinology, developmental and behavioral modulation, regulation of intracellular levels of cholesterol, and potential neurotoxicity.^ In this study, we have set up the rat glioma C6 cell line as an in vitro model system to examine the expression, induction, and tissue-specific regulation of P450s and P450 reductase. Rat glioma C6 cells were treated with P450 inducers phenobarbital (PB) or benzo(a)anthracene (BA). The presence of P450 reductase and of cytochrome P450 1A1, 1A2, 2A1, 2B1/2, 2C7, 2D1-5 and 2E1 was detected by reverse transcription followed by polymerase chain reaction (RT-PCR) and confirmed by restriction digestion. The induction of P450 1A1 and 2B1/2 and P450 reductase was quantified using competitive PCR. Ten- and five-fold inductions of P450 1A and 2B mRNA after BA or PB treatments, respectively, were detected. Western blot analysis of microsomal preparations of glioma C6 cells demonstrated the presence of P450 1A, 2B and P450 reductase at the protein level. ELISAs showed that BA and PB induce P450 1A and 2B proteins 7.3- and 13.5-fold, respectively. Microsomes prepared from rat glioma C6 cells showed cytochrome P450 CO difference spectra with absorption at or near 450 nm. Microsomes prepared from rat glioma C6 cells demonstrated much higher levels of ethoxyresorufin O-deethylase (EROD) and pentoxyresorufin O-dealkylase (PROD) activity, when treated with BA or PB, respectively. These experiments provide further evidence that the rat glioma C6 cell line contains an active cytochrome P450 monooxygenase system which can be induced by P450 inducers. The mRNAs of P450 1A1 and 2B1/2 can not bind to the oligo(dT) column efficiently, indicating they have very short poly(A) tails. This finding leads us to study the tissue specific regulation of P450s at post-transcriptional level. The half lives of P450 1A1 and 2B1/2 mRNA in glioma C6 cells are only 1/10 and 1/3 of that in liver. This may partly contribute to the low expression level of P450s in glial cells. The induction of P450s by BA or PB did not change their mRNA half lives, indicating the induction may be due to transcriptional regulation. In summary of this study, we believe the presence of the cytochrome P450 monooxygenase system in glial cells of the brain may be important in chemotherapy and carcinogenesis of brain tumors. ^

Hormonal risk factors and testicular cancer

The cause of testicular cancer is not known and recent hypotheses have suggested an altered hormonal milieu may increase the risk of testis cancer. This study examined modulation of testicular cancer risk by hormonal factors, specifically: environmental xenoestrogens (e.g. organochlorines), prenatal maternal estrogens, testosterone indices (age at puberty, severe adolescent acne, self-reported balding), sedentary lifestyle and dietary consumption of fats and phytoestrogens.^ A hospital based friend matched case-control study was conducted at the University of Texas M. D. Anderson Cancer Center in Houston, Texas, between January 1990 and October 1996. Cases had a first primary testis tumor diagnosed between age 18 to 50 years and resided in Texas, Louisiana, Oklahoma or Arkansas.^ Cases and friend controls completed a mail questionnaire and case/control mothers were contacted by phone regarding pregnancy related variables. The study population comprised 187 cases, 148 controls, 147 case mothers and 86 control mothers. Odds ratios were virtually identical whether the match was retained or dissolved, thus the analyses were conducted using unconditional logistic regression.^ Cryptorchidism was a strong risk factor for testis cancer with an age-adjusted odds ratio (OR) of 7.7 (95% confidence interval (CI): 2.3-26.3). In a final model (adjusted for age, education, and cryptorchidism), history of severe adolescent acne and self-reported balding were both significantly protective, as hypothesized. For acne (yes vs. no) the OR was 0.5 (CI: 0.3-1.0) and for balding (yes vs. no) the OR was 0.6 (CI: 0.3-1.0). Marijuana smoking was a risk factor among heavy, regular users (17 times/week, OR = 2.4; CI: 0.9-6.4) and higher saturated fat intake increased testis cancer risk (saturated fat intake $>$ 15.2 grams/day vs. $<$ 11.8 grams/day, OR = 3.3; CI: 1.5-7.1). Early puberty, xenoestrogen exposure, elevated maternal estrogen levels, sedentary lifestyle and dietary phytoestrogen intake were not associated with risk of testicular cancer.^ In conclusion, testicular cancer may be associated with endogenous androgen metabolism although environmental estrogen exposure can not be ruled out. Further research is needed to understand the underlying hormonal mechanisms and possible dietary influences. ^

Phototaxis signaling by the membrane complex of archaeal sensory rhodopsins and their transducers

Sensory rhodopsins I and II (SRI and SRII) are visual pigment-like phototaxis receptors in the archaeon Halobacterium salinarum. The receptor proteins each consist of a single polypeptide that folds into 7 $\alpha$-helical membrane-spanning segments forming an internal pocket where the chromophore retinal is bound. They transmit signals to their tightly bound transducer proteins, HtrI and HtrII, respectively, which in turn control a phosphotransfer pathway modulating the flagellar motors. SRI-HtrI mediates attractant responses to orange-light and repellent responses to UV light, while SRII-HtrII mediates repellent response to blue light. Experiments were designed to analyze the molecular processes in the SR-Htr complexes responsible for receptor activation, which previously had been shown by our laboratory to involve proton transfer reactions of the retinylidene Schiff base in the photoactive site, transfer of signals from receptor to transducer, and signaling specificity by the receptor-transducer complex.^ Site-directed mutagenesis and laser-flash kinetic spectroscopy revealed that His-166 in SRI (i) plays a role in the proton transfers both to and from the Schiffbase, either as a structurally critical residue or possibly as a direct participant, (ii) is involved in the modulation of SIU photoreaction kinetics by HtrI, and (iii) modulates the pKa of Asp-76, an important residue in the photoactive site, through a long-distance electrostatic interaction. Computerized cell tracking and motion analysis demonstrated that (iv) His-166 is crucial in phototaxis signaling: a spectrum of substitutions either eliminate signaling or greatly perturb the activation process that produces attractant and repellent signaling states of the receptor.^ The signaling states of SRI are communicated to HtrI, whose oligomeric structure and conformational changes were investigated by engineered sulfhydryl probes. It was found that signaling by the SRI-HtrI complex involves reversible conformational changes within a preexisting HtrI dimer, which is likely accomplished through a slight winding or unwinding of the two HtrT monomers via their loose coiled coil association. To elucidate which domains of the Htr dimers confer specificity for interaction with SRI or SRII, chimeras of HtrI and HtrII were constructed. The only determinant needed for functional and specific interaction with SRI or SRII was found to be the four transmembrane segments of the HtrI or HtrII dimers, respectively. The entire cytoplasmic parts of HtrI and HtrII, which include the functionally important signaling and adaptation domains, were interchangeable.^ These observations support a model in which SRI and SRII undergo conformational changes coupled to light-induced proton transfers in their photoactive sites, and that lateral helix-helix interactions with their cognate transducers' 4-helix bundle in the membrane relay these conformational changes into different states of the Htr proteins which regulate the down-stream phosphotransfer pathway. ^

Studies of subcellular localization and complex formation of the Agrobacterium tumefaciens transport ATPase VirB11

The VirB11 ATPase is an essential component of an Agrobacterium tumefaciens type IV bacterial secretion system that transfers oncogenic nucleoprotein complexes to susceptible plant cells. This dissertation investigates the subcellular localization and homo-oligomeric state of the VirB11 ATPase in order to provide insights about the assembly of the protein as a subunit of this membrane-associated transfer system. Subcellular fractionation studies and quantitative immunoblot analysis demonstrated that $\sim$30% of VirB11 partitioned as soluble protein and $\sim$70% was tightly associated with the bacterial cytoplasmic membrane. No differences were detected in VirB11 subcellular localization and membrane association in the presence or absence of other transport system components. Mutations in virB11 affecting protein function were mapped near the amino terminus, just upstream of a region encoding a Walker 'A' nucleotide-binding site, and within the Walker 'A' motif partitioned almost exclusively with the cytoplasmic membrane, suggesting that an activity associated with nucleotide binding could modulate the affinity of VirB11 for the cytoplasmic membrane. Merodiploid analysis of VirB11 mutant and truncation derivatives provided strong evidence that VirB11 functions as a homo- or heteromultimer and that the C-terminal half of VirB11 contains a protein interaction domain. A combination of biochemical and molecular genetic approaches suggested that VirB11 and the green fluorescence protein (GFP) formed a mixed multimer as demonstrated by immunoprecipitation experiments with anti-GFP antibodies. Second, a hybrid protein composed of VirB11 fused to the N-terminal DNA-binding domain of bacteriophage $\lambda$ cI repressor conferred immunity to $\lambda$ superinfection, demonstrating that VirB11 self-association promotes dimerization of the chimeric repressor. A conserved Walker 'A' motif, though required for VirB11 function in T-complex export, was not necessary for VirB11 self-association. Sequences in both the N- and the C-terminal halves of the protein were found to contribute to self-association of the full length protein. Chemical cross-linking experiments with His$\sb6$ tagged VirB11 suggested that VirB11 probably assembles into a higher order homo-oligomeric complex. ^

The role of menaquinone in the nitrate reductase complex

Membrane bound, respiratory nitrate reductase in Escherichia coli is composed of three subunits, αβγ. The active complex is anchored to the membrane by membrane-integrated γ subunit and can reduce nitrate to nitrite with membrane quinones, (ubiquinone or menaquinone) as physiological electron donors. The transfer of electrons through the complex is thought to involve the sequence: membrane quinols → b-type hemes (γ subunit) → Fe-S centers (β subunit) → molybdopterin (α subunit) → nitrate. The enzyme can be assayed with the artificial electron donor reduced methyl viologen (MVH) which transfers electrons directly to the molybdopterin cofactor. These studies have focused on the possible role of protein-bound menaquinone in the structure and function of this multisubunit complex. ^ Nitrate reductase was purified as two distinct forms; after solubilization of membrane proteins with detergents, purification rendered an αβγ complex (holoenzyme) which catalyzes nitrate reduction with MVH or the quinols analogs, menadiol and duroquinol, as electron donors. Alternatively, heat-treatment of the membranes in the absence of detergents and subsequent purification of the active enzyme produced an αβ complex, which reduces nitrate only with MVH as electron donor. The active αβ dimer was also separated from γ subunit by heat treatment of the holoenzyme. ^ Menaquinone-9 was isolated directly from the purified αβ complex, and identified by mass spectrometry. Based on the composition of the membrane quinone pool, it was concluded that menaquinone-9 is sequestered from the membrane pool in a specifically protein-bound form. ^ The role of the bound menaquinone in the structure-function of nitrate reductase was also investigated, along with its participation in UV-light inactivation of the enzyme. Menaquinone-depleted nitrate reductase from a menaquinone deficient mutant retained activity with all electron donors and it remained sensitive to UV inactivation. However, the MVH-nitrate reductase activity and the rate of UV inactivation of the enzyme were significantly reduced and the optical properties of the enzyme were modified by the absence of the bound menaquinone-9. ^ Menaquinone-9 is not absolutely required for electron transfer in nitrate reductase but it appears to be specifically-bound during assembly of the complex and to enhance the transfer of electrons through the complex. The possible plasticity of the functional electron transfer pathway in nitrate reductase is discussed. ^

Globale Akteure an den Randzonen von Souveränität und Legitimität

Der moderne Nationalstaat mit seinen Grenzlinien, welche die räumliche Reichweite des souveränen Gewaltmonopols fixierten und gleichsam einen Kulturraum mit homogenen sozialen und rechtlichen Normen abschlossen, war lange eine zentrale Orientierungsgröße für die Geschichtsschreibung. Die wirtschaftlichen, politischen und kulturellen Globalisierungsprozesse der letzten zwei Jahrzehnte haben jedoch das nationale Narrativ mit seinen eindeutigen Demarkationslinien in Frage gestellt. Stattdessen sind Randzonen und Zwischenräume in den Mittelpunkt des Interesses gerückt. Dieser Paradigmenwechsel in Methodik und Perspektive hat auch realhistorisch seine volle Berechtigung, denn solche Zwischenräume sind keine neuartigen Phänomene der Postmoderne mit ihren globalen Transfers und multikulturellen Gesellschaften. Sie hatten sich beispielsweise stets auch während des Epochen übergreifenden Prozesses der europäischen Expansion aufgetan, insbesondere an den fluiden imperialen Randzonen. Nach eurozentrischen Maßstäben handelte es sich dabei um rechtlich nicht regulierte und territorial undefinierte Räume. Aber selbst in der Hochphase des Imperialismus sowie auch in den postkolonialen Gesellschaften konnte das nationalstaatliche Modell nur partiell etabliert werden und konkurrierte stets mit „tribalen“ oder quasi-feudalen Formen sozio-politischer Organisation. Außerdem wurde selbst in den westlichen Nationalstaaten das staatliche Gewaltmonopol sowie die soziale, rechtliche und politische Ordnung bei innerstaatlichen Konflikten wie Revolutionen und Bürgerkriegen sowie internationalen Kriegen immer wieder in Frage gestellt oder sogar ausgehebelt. In solch gewaltbasierten Umfeldern des Ausnahmezustandes und der Insurrektion sowie in solch deregulierten Räumen, weitgehend frei von Hierarchien, bürokratischer Kontrolle und rechtlichen Einschränkungen, agierten stets auch besondere Akteurstypen. Sie hatten einerseits das Privileg einer geringen Kontrolle unterworfen zu sein oder völlig losgelöst davon zu handeln, waren aber andererseits überwiegend auf sich gestellt und konnten nur sehr beschränkt den Schutz des fernen Souveräns im Hintergrund beanspruchen oder hatten sich diesen durch ihre Verhaltensweise sogar zum Gegner gemacht. Diese Akteure an den Randzonen von Souveränität und Legitimität können grob in verschiedene Typen eingeteilt werden: Zu nennen sind die Avantgardisten der Expansion wie Entdecker und Eroberer, die klassischen „Men on the Spot“ an der Peripherie von Imperien wie Militärs und Administratoren, private Gewaltunternehmer und Söldner, Insurgenten und Revolutionäre, sowie Spione und Nachrichtenhändler. Anhand verschiedener Fallstudien soll diese Figur des Akteurs an den Randzonen in globaler Perspektive beleuchtet werden. Es stellt sich die Frage nach seiner Motivation für Sondermissionen, seiner ideellen Beweggründe und politischen Überzeugung sowie seiner Beziehung zum Auftraggeber oder Souverän im fernen Hintergrund. Insbesondere ist untersuchen, ob deren Interessen trotz beschränkter Kontrolle und Rückendeckung loyal verfolgt wurden oder ob für unsere Protagonisten vor allem materielle Interessen ausschlaggebend waren, womöglich sogar einfach das Streben nach einem Betätigungsfeld mit möglichst großer Handlungsfreiheit? Während entgrenzte Verhaltensweisen in der Regel auf rechtlich nur schwach regulierte Umfeldern zurückgeführt werden, ist auch zu untersuchen, ob sich gewisse Persönlichkeiten nicht gezielt Räume am Rande oder jenseits nationalstaatlicher Rechtsnormen als Agitationsfeld suchten oder ob sie sich sogar zur Verwirklichung ihrer persönlichen Bedürfnisse solche Räume durch Subversion staatlicher Ordnung gezielt schafften. Indem die Protagonisten mit den kulturellen, politischen und rechtlichen Ordnungen ihres Betätigungsfeldes in Beziehung gesetzt werden, kann auch eine Brücke von einer rein personenbezogenen Biographik zur Strukturgeschichte geschlagen werden. Die Grenzgänger wie auch die Zwischenräume, in denen sie agierten, sind wohl auch weniger als Randerscheinungen des Ausnahmezustandes zu verstehen, sondern vielmehr als die unmittelbar mit der sich intensivierenden Verrechtlichung und Homogenisierung der westeuropäischen Nationalstaaten einhergehende andere Seite dieses Prozesses: Einerseits brauchten Regierungen zuweilen für besondere Aufgaben Akteure, die nicht an die eigenen Rechtsnormen gebunden waren, andererseits gab es stets Persönlichkeiten, die sich in der engen Rechtswirklichkeit und sozialen Ordnung des Nationalstaates nicht zurechtfanden. Darüber hinaus sind diese Grenzgänger auch als Brückenköpfe politischer wie ökonomischer Machterweiterung sowie als Schlüsselfiguren von Interaktions-, Kooperations-, Vernetzungs- und Transformationsprozessen zu verstehen – nur in Extremfällen waren sie radikale Grenzüberschreiter sowie Träger der bloßen Zerstörung und Zersetzung. Da sie in einem kulturell fremden oder zerrütteten, krisengebeutelten Umfeld agierten, gilt ein besonderes Augenmerk ihrer Rolle als Makler von Interessen, Vermittler sowie Informationsbeschaffer. Schließlich soll auch die Außenwahrnehmung dieser Randfiguren in Betracht gezogen werden. Ihre Position am Rande der Gesellschaft und der Legalität löste in der Regel eine große Faszination beim „Normalbürger“ aus, die sich entweder in Bewunderung oder Abscheu wenden konnte. Teilweise veranlassten sie einen regelrechten Medienrummel, der den Staat wiederum in Zugzwang bringen konnte, oft mit beachtlichen Folgen.