Pontages veineux in situ. Indications, technique et résultats. A propos d'une étude prospective de 26 cas consécutifs [In situ venous grafts. Indications, technique and results. Prospective study of 26 consecutive cases].

The in situ saphenous vein bypass has been introduced in our department since 1989. A total of 26 bypasses in 22 patients have been followed prospectively. Indications for revascularisation have been severe arterial insufficiency in 73% of the cases (stage III or IV). With the exception of one postoperative death (myocardial infarction), all the patients have recovered uneventfully, with a regression to stage I. No amputation has been necessary. Morbidity has been 30%, with mainly minor local complications. The primary patency rate is 83% at one year and 78% after 2 and 3 years, whereas the secondary patency rate is 91% at one year, and remains constant thereafter up to 3 years. Considering our results and those from the literature, we believe that the in situ technique is very valuable, especially for below-knee vascular reconstruction. Technical difficulties of the method are analysed.

Local moderate magnetically induced hyperthermia using an implant formed in situ in a mouse tumor model.

Purpose: We investigate a new heat delivery technique for the local treatment of solid tumors. The technique involves injecting a formulation that solidifies to form an implant in situ. This implant entraps superparamagnetic iron oxide nanoparticles (SPIONs) embedded in silica microbeads for magnetically induced moderate hyperthermia. Particle entrapment prevents phagocytosis and distant migration of SPIONs. The implant can be repeatedly heated by magnetic induction. Methods: We evaluated heating and treatment efficacies by means of thermometry and survival studies in nude mice carrying subcutaneous human colocarcinomas. At day 1, we injected the formulation into the tumor. At day 2, a single 20-min hyperthermia treatment was delivered by 141-kHz magnetic induction using field strengths of 9 to 12 mT under thermometry. Results: SPIONs embedded in silica microbeads were effectively confined within the implant at the injection site. Heat-induced necro-apoptosis was assessed by histology on day 3. On average, 12 mT resulted in tumor temperature of 47.8 degrees C, and over 70% tumor necrosis that correlated to the heat dose (AUC = 282 degrees C.min). In contrast, a 9-mT field strength induced tumoral temperature of 40 degrees C (AUC = 131 degrees C.min) without morphologically identifiable necrosis. Survival after treatment with 10.5 or 12 mT fields was significantly improved compared to non-implanted and implanted controls. Median survival times were 27 and 37 days versus 12 and 21 days respectively. Conclusion: Five of eleven mice (45%) of the 12 mT group survived one year without any tumor recurrence, holding promise for tumor therapy using magnetically induced moderate hyperthermia through injectable implants.

Fingermark detection based on the in situ growth of luminescent nanoparticles: towards a new generation of multimetal deposition

The in situ deposition of zinc oxide on gold nanoparticles in aqueous solution has been here successfully applied in the field of fingermark detection on various non-porous surfaces. In this article, we present the improvement of the multimetal deposition, an existing technique limited up to now to non-luminescent results, by obtaining luminescent fingermarks with very good contrast and details. This is seen as a major improvement in the field in terms of selectivity and sensitivity of detection, especially on black surfaces.

Applications of a transportable Raman spectrometer for the in situ detection of controlled substances at border controls

A transportable Raman spectrometer was tested for the detection of illicit drugs seized during border controls. In a first step, the analysis methodology was optimized using reference substances such as diacetylmorphine (heroin), cocaine and amphetamine (as powder or liquid forms). Adequate focalisation distance and times of analysis, influence of daylight and artificial light sources, repeatability and limits of detection were studied. In a second step the applications and limitations of the technique to detect the illicit substances in different mixtures and containers was evaluated. Transportable Raman spectroscopy was found to be adequate for a rapid screen of liquids and powders for the detection and identification of controlled substances. Additionally, it had the advantage over other portable techniques, such as ion mobility spectrometry, of being non-destructive and capable of rapid analysis of large quantities of substances through containers such as plastic bags and glass bottles.

In situ introducer sheath dilatation for complex aortic access.

Aortic access problems due to diseased or small peripheral vessels are a major issue in endovascular aneurysm repair (EVAR). In the emergency setting, like aortic rupture after blunt trauma, or in patients with a hostile abdomen, a more proximal access to the aorta is not a pleasant perspective. We developed in situ introducer sheath dilatation as a bail-out technique for patients with difficult aortic access under various circumstances including EVAR, intra-aortic balloon pump insertion and cannulation for perfusion. The method described allows to increase the access vessel diameter by 50% (from 6 to 9 mm) or the luminal circumference from 18 to 27 F. We have used this technique in five patients without complication, very much in contrast to the traditionally practiced 'forced device insertion'.

Diagnostic and identification in situ of fungal pathogens in superficial mycosis

Fungi are divided in 3 groups in the field of medical mycology. The dermatophytes are filamentous fungi able to grow on keratinized tissues from human or animals. They are the main cause of superficial and cutaneous mycoses of the skin and its appendix (hair and nail). The yeasts, or dimorphic fungi, can be responsible of diverse types of infections (superficial to deep mycoses). The moulds include all Non-dermatophyte Filamentous Fungi (NDF). In medical mycology, the most representative moulds are Aspergillus spp., Fusarium spp. and Mucor spp. Diagnosis of mycosis is currently based on direct mycological examination of biological samples, as well as macroscopic and microscopic identification of the infectious fungus in culture assay. However, culture assays were found to remain sterile in roughly 40% of cases otherwise positive by direct mycological examinations. Additionally, results from culture assays are often difficult to interpret as various NDF are sometimes isolated. This thesis work is composed of three projects focusing on the development of new assays for direct in situ identification of fungi from dermatological samples. Part 1. A Polymerase Chain Reaction - Terminal Restriction Fragment Length Polymorphism assay (PCR-TRFLP) targeting the 28S rDNA was developed to identify dermatophytes and NDF in nails with suspected onychomycosis. This method is faster and more efficient than culture. It further enables the distinction of more than one agent in case of mixed infection. A fast and reliable assay for the identification of dermatophytes and NDF in onychomycosis was found to be highly relevant since onychomycosis with Fusarium spp. or other NDF are weakly responsive or unresponsive to standard onychomycosis treatments with oral terbinafine and itraconazole. Part 2. A nested PCR-sequencing assay targeting the 28S rDNA was developed to identify dermatophyte species in skin and hair samples. This method is especially suitable for tinea capitis where dermatophytes identification is critical for subsequently prescribing the adequate treatment. The challenge presented when performing direct PCR fungi identification in skin and hair differs from that seen in onychomycosis as small amount of material is generally collected, few fungal elements are present in the clinical sample and one dermatophyte among a dozen species must be identified. Part 3. Fusarium spp. is currently isolated from nails with a frequency of 15% of that of dermatophytes in the laboratory of Mycology of the CHUV (2005-2012). The aim of this work was to examine if the intensive use of terbinafine and itraconazole could be a cause of the high incidence of Fusarium nail infections. For that purpose, two different methods, specific PCR and TRFLP, were used to detect both Fusarium spp. and Trichophyton spp. in nails of previously treated or untreated patients. TRFLP assay was found to be less sensitive than classical PCR assays specifically detecting Fusarium spp. or Trichophyton spp. Independently of the detection method used, the prevalence of Fusarium spp. appears not to be higher in patients previously treated by oral standard treatment with terbinafine and azoles which are highly effective to fight Trichophyton spp. in nails. In many cases Fusarium sp. was detected in samples of patients not previously subjected to antifungal therapy. Therefore, these treatments do not appear to favor the establishment of Fusarium spp. after elimination of a dermatophyte in nail infection. - En mycologie médicale, les champignons sont classés en 3 groupes. Les dermatophytes sont des champignons filamenteux capables de se développer dans les tissus kératinisés des hommes et des animaux, ils représentent la principale cause des mycoses superficielles et cutanées de la peau et de ses appendices (ongles et cheveux). Les levures, ou champignons dimorphiques, peuvent être responsables de divers types d'infections (superficielles à profondes). Les moisissures incluent tous les champignons filamenteux non-dermatophytes (NDF), les Aspergillus spp., les Fusarium spp. et les Mucor spp. sont les principales espèces rencontrées. Le diagnostic d'une mycose est basé sur un examen mycologique direct des prélèvements biologiques ainsi que sur l'identification macroscopique et microscopique du champignon infectieux isolé en culture. Cependant, dans environ 40% des cas, l'identification de l'agent pathogène est impossible par cette méthode car la culture reste stérile, bien que l'examen direct soit positif. De plus, la croissance de moisissures et/ou autres contaminants peut rendre l'interprétation de l'examen difficile. Ce travail de thèse est composé de trois projets focalisés sur le développement de nouvelles méthodes d'identification des champignons directement à partir d'échantillons dermatologiques. Projet 1. Une méthode de Réaction en chaîne de polymérase couplée à du polymorphisme de longueur des fragments de restriction terminaux (PCR-TRFLP), en ciblant l'ADN ribosomal 28S, a été développée pour l'identification des dermatophytes et moisissures dans les ongles avec suspicion d'onychomycoses. Cette technique s'est avérée plus rapide et plus efficace que la culture, permettant l'identification de plusieurs champignons en même temps. Posséder une méthode d'identification rapide et fiable des dermatophytes et des NDF dans les onychomycoses a été jugée nécessaire du fait que les Fusarium et d'autres NDF sont peu ou pas sensibles aux traitements oraux standards à la terbinafine et à Γ itraconazole. Projet 2. Une PCR nichée couplée au séquençage d'un fragment de l'ADN ribosomal 28S a été développée afin de différencier les dermatophytes dans la peau et les cheveux. Cette méthode est particulièrement adaptée au cas de tinea capitis, où l'identification du dermatophyte est essentielle afin de prescrire le traitement adéquat. Le problème de l'identification du pathogène fongique dans les cheveux et la peau diffère des onychomycoses car de petites quantités sont prélevées chez les patients, peu d'éléments fongiques sont présents et il faut discriminer un dermatophyte parmi une douzaine d'espèces potentielles. Projet 3. Au laboratoire de Mycologie du CHUV, les Fusarium ont été isolé dans les ongles à une fréquence de 15% pour la période 2005-2012. Le but de ce travail était d'examiner si l'utilisation intensive de terbinafine et d'itraconazole pouvait être une des causes de la forte incidence des infections des ongles par Fusarium. A cet effet, deux méthodes ont été utilisées pour détecter à la fois Fusarium spp. et Trichophyton spp., la PCR spécifique et le TRFLP. Indépendamment de la méthode choisie, il en résulte que la prévalence des Fusarium η'apparaît pas liée à un traitement au préalable des patients avec de la terbinafine ou des azoles, thérapies très efficaces contre les Trichophyton spp. dans les ongles. De plus, il existe de nombreux cas où Fusarium était détecté chez des patients non traités.

The Discrimination of Colored Acrylic, Cotton, and Wool Textile Fibers Using Micro-Raman Spectroscopy. Part 1 : In Situ Detection and Characterization of Dyes

Raman spectroscopy has been applied to characterize fiber dyes and determine the discriminating ability of the method. Black, blue, and red acrylic, cotton, and wool samples were analyzed. Four excitation sources were used to obtain complementary responses in the case of fluorescent samples. Fibers that did not provide informative spectra using a given laser were usually detected using another wavelength. For any colored acrylic, the 633-nm laser did not provide Raman information. The 514-nm laser provided the highest discrimination for blue and black cotton, but half of the blue cottons produced noninformative spectra. The 830-nm laser exhibited the highest discrimination for red cotton. Both visible lasers provided the highest discrimination for black and blue wool, and NIR lasers produced remarkable separation for red and black wool. This study shows that the discriminating ability of Raman spectroscopy depends on the fiber type, color, and the laser wavelength.

Modeling of in-situ crystallization processes in the Permian mafic layered intrusion of Mont Collon (Dent Blanche nappe, western Alps)

The Mont Collon mafic complex is one of the best preserved examples of the Early Permian magmatism in the Central Alps, related to the intra-continental collapse of the Variscan belt. It mostly consists (> 95 vol.%) of ol+hy-nonnative plagioclase-wehrlites, olivine- and cpx-gabbros with cumulitic structures, crosscut by acid dikes. Pegmatitic gabbros, troctolites and anorthosites outcrop locally. A well-preserved cumulative, sequence is exposed in the Dents de Bertol area (center of intrusion). PT-calculations indicate that this layered magma chamber emplaced at mid-crustal levels at about 0.5 GPa and 1100 degrees C. The Mont Collon cumulitic rocks record little magmatic differentiation, as illustrated by the restricted range of clinopyroxene mg-number (Mg#(cpx)=83-89). Whole-rock incompatible trace-element contents (e.g. Nb, Zr, Ba) vary largely and without correlation with major-element composition. These features are characteristic of an in-situ crystallization process with variable amounts of interstitial liquid L trapped between the cumulus mineral phases. LA-ICPMS measurements show that trace-element distribution in the latter is homogeneous, pointing to subsolidus re-equilibration between crystals and interstitial melts. A quantitative modeling based on Langmuir's in-situ crystallization equation successfully duplicated the REE concentrations in cumulitic minerals of all rock facies of the intrusion. The calculated amounts of interstitial liquid L vary between 0 and 35% for degrees of differentiation F of 0 to 20%, relative to the least evolved facies of the intrusion. L values are well correlated with the modal proportions of interstitial amphibole and whole-rock incompatible trace-element concentrations (e.g. Zr, Nb) of the tested samples. However, the in-situ crystallization model reaches its limitations with rock containing high modal content of REE-bearing minerals (i.e. zircon), such as pegmatitic gabbros. Dikes of anorthositic composition, locally crosscutting the layered lithologies, evidence that the Mont Collon rocks evolved in open system with mixing of intercumulus liquids of different origins and possibly contrasting compositions. The proposed model is not able to resolve these complex open systems, but migrating liquids could be partly responsible for the observed dispersion of points in some correlation diagrams. Absence of significant differentiation with recurrent lithologies in the cumulitic pile of Dents de Bertol points to an efficiently convective magma chamber, with possible periodic replenishment, (c) 2005 Elsevier B.V. All rights reserved.

In situ evaluation of single-cell lysis by cytosol extraction observation through fluorescence decay and dielectrophoretic trapping time

We present a new method for lysis of single cells in continuous flow, where cells are sequentially trapped, lysed and released in an automatic process. Using optimized frequencies, dielectrophoretic trapping allows exposing cells in a reproducible way to high electrical fields for long durations, thereby giving good control on the lysis parameters. In situ evaluation of cytosol extraction on single cells has been studied for Chinese hamster ovary (CHO) cells through out-diffusion of fluorescent molecules for different voltage amplitudes. A diffusion model is proposed to correlate this out-diffusion to the total area of the created pores, which is dependent on the potential drop across the cell membrane and enables evaluation of the total pore area in the membrane. The dielectrophoretic trapping is no longer effective after lysis because of the reduced conductivity inside the cells, leading to cell release. The trapping time is linked to the time required for cytosol extraction and can thus provide additional validation of the effective cytosol extraction for non-fluorescent cells. Furthermore, the application of one single voltage for both trapping and lysis provides a fully automatic process including cell trapping, lysis, and release, allowing operating the device in continuous flow without human intervention.

High-magnification vascular imaging to reject false-positive sites in situ during Hexvix® fluorescence cystoscopy.

Fluorescence imaging for detection of non-muscle-invasive bladder cancer is based on the selective production and accumulation of fluorescing porphyrins-mainly, protoporphyrin IX-in cancerous tissues after the instillation of Hexvix®. Although the sensitivity of this procedure is very good, its specificity is somewhat limited due to fluorescence false-positive sites. Consequently, magnification cystoscopy has been investigated in order to discriminate false from true fluorescence positive findings. Both white-light and fluorescence modes are possible with the magnification cystoscope, allowing observation of the bladder wall with magnification ranging between 30× for standard observation and 650×. The optical zooming setup allows adjusting the magnification continuously in situ. In the high-magnification (HM) regime, the smallest diameter of the field of view is 600 microns and the resolution is 2.5 microns when in contact with the bladder wall. With this cystoscope, we characterized the superficial vascularization of the fluorescing sites in order to discriminate cancerous from noncancerous tissues. This procedure allowed us to establish a classification based on observed vascular patterns. Seventy-two patients subject to Hexvix® fluorescence cystoscopy were included in the study. Comparison of HM cystoscopy classification with histopathology results confirmed 32?33 (97%) cancerous biopsies and rejected 17?20 (85%) noncancerous lesions.

Fas ligand expression is restricted to nonlymphoid thymic components in situ.

The cell surface receptor Fas (Apo-1/CD95) and its ligand (FasL) are mediators of apoptosis that have been shown to be implicated in activation-induced death of mature T cells and in killing mediated by cytolytic T cells. The role of the Fas pathway in apoptosis associated with thymic selection events is, however, controversial. Although Fas and FasL are known to be expressed in the thymus, the nature and in vivo localization of FasL-expressing cells have not been determined. Using recently developed anti-FasL Abs in combination with in situ hybridization on tissue sections, we show in this work that FasL-expressing cells are present in the thymus, particularly within the medulla. FasL mRNA was detected readily in thymic stromal cell extracts, but not in isolated thymocytes. Moreover, immunohistochemical analysis of serial tissue sections stained with Abs against FasL in conjunction with epithelial and dendritic cell markers indicated that both thymic epithelial and dendritic cells express FasL in situ. The coexistence of FasL-expressing stromal cells and Fas-expressing thymocytes may have important implications for the role of the Fas pathway in apoptosis associated with thymic selection events.

The in vivo performance of magnetic particle-loaded injectable, in situ gelling, carriers for the delivery of local hyperthermia.

We investigated the use of in situ implant formation that incorporates superparamagnetic iron oxide nanoparticles (SPIONs) as a form of minimally invasive treatment of cancer lesions by magnetically induced local hyperthermia. We developed injectable formulations that form gels entrapping magnetic particles into a tumor. We used SPIONs embedded in silica microparticles to favor syringeability and incorporated the highest proportion possible to allow large heating capacities. Hydrogel, single-solvent organogel and cosolvent (low-toxicity hydrophilic solvent) organogel formulations were injected into human cancer tumors xenografted in mice. The thermoreversible hydrogels (poloxamer, chitosan), which accommodated 20% w/v of the magnetic microparticles, proved to be inadequate. Alginate hydrogels, however, incorporated 10% w/v of the magnetic microparticles, and the external gelation led to strong implants localizing to the tumor periphery, whereas internal gelation failed in situ. The organogel formulations, which consisted of precipitating polymers dissolved in single organic solvents, displayed various microstructures. A 8% poly(ethylene-vinyl alcohol) in DMSO containing 40% w/v of magnetic microparticles formed the most suitable implants in terms of tumor casting and heat delivery. Importantly, it is of great clinical interest to develop cosolvent formulations with up to 20% w/v of magnetic microparticles that show reduced toxicity and centered tumor implantation.

In-situ revascularisation for secondary aorto-enteric fistulae: the success of silver-coated Dacron is closely linked to a suitable bowel repair.

OBJECTIVES: The purpose of this study was to assess short- and mid-term results of in-situ revascularisation (ISR) using silver-coated Dacron prostheses and bowel repair for management of secondary aorto-enteric fistulae (SAEF). DESIGN: Single-centre retrospective chart review. MATERIAL AND METHODS: This study includes all the patients treated by ISR using silver-coated Dacron for SAEF between 2006 and 2010. Primary end points were mortality and survival rates. Secondary end points were reinfection-free survival and secondary patency rates. RESULTS: Eighteen male patients with SAEF with a median age of 64 years were operated by ISR using silver-coated Dacron during the study period without operative death. The 30-day mortality was 22% and the in-hospital mortality rate was 39%. Indeed, during hospitalisation, a duodenal leak was observed in four patients including three who died. Four others patients died due to multi-system organ failure. Median follow-up was 16 months (range 1-66). The survival rate at 12 months was 55%. One duodenal leak was observed leading to death. The reinfection-free survival and the secondary patency rates at 12 months were 60% and 89%, respectively. CONCLUSION: In-situ revascularisation with silver-coated Dacron provides acceptable results in terms of mortality. This treatment may be useful for simple vascular reconstruction and allow greater attention to bowel repair that is a determinant in short- and mid-term survival.

Carcinomes canalaires in situ: données anatomo-pathologiques actuelles [Ductal carcinoma in situ: current anatomo-pathologic data]

Ductal carcinoma in situ (DCIS), accounting for 15-25% of all breast cancers, is frequently diagnosed by mammographic examination. This heterogeneous disease requires a rigorous local treatment based, in about two-third of cases, on conservative surgery and radiotherapy. DCIS are currently classified on the basis of nuclear grade. Most lesions, and especially high nuclear grade DCIS, are limited to one quadrant. Micropapillary DCIS are likely to be of larger size/extent and thus a conservative approach is often difficult. A careful pathological examination of an oriented excisional biopsy is a pre-requisite for optimal therapy.

Differential distribution of two microtubule-associated proteins, MAP2 and MAP5, during chick dorsal root ganglion development in situ and in culture.

Microtubule-associated proteins (MAPs) are essential components necessary for the early growth process of axons and dendrites, and for the structural organization within cells. Both MAP2 and MAP5 are involved in these events, MAP2 occupying a role predominantly in dendrites, and MAP5 being involved in both axonal and dendritic growth. In the chick dorsal root ganglia, pseudo-unipolar sensory neurons have a T-shaped axon and are devoid of any dendrites. Therefore, they offer an ideal model to study the differential expression of MAPs during DRG development, specifically during axonal growth. In this study we have analyzed the expression and localization of MAP2 and MAP5 isoforms during chick dorsal root ganglia development in vivo, and in cell culture. In DRG, both MAPs appeared as early as E5. MAP2 consists of the 3 isoforms MAP2a, b and c. On blots, no MAP2a could be found at any stage. MAP2b increased between E6 and E10 and thereafter diminished slowly in concentration, while MAP2c was found between stages E6 and E10 in DRG. By immunocytochemistry, MAP2 isoforms were mainly located in the neuronal perikarya and in the proximal portion of axons, but could not be localized to distal axonal segments, nor in sciatic nerve at any developmental stage. On blots, MAP5 was present in two isoforms, MAP5a and MAP5b. The concentration of MAP5a was highest at E6 and then decreased to a low level at E18. In contrast, MAP5b increased between E6 and E10, and rapidly decreased after E14. Only MAP5a was present in sciatic nerve up to E14. Immunocytochemistry revealed that MAP5 was localized mainly in axons, although neuronal perikarya exhibited a faint immunostaining. Strong staining of axons was observed between E10 and E14, at a time coincidental to a period of intense axonal outgrowth. After E14 immunolabeling of MAP5 decreased abruptly. In DRG culture, MAP2 was found exclusively in the neuronal perikarya and the most proximal neurite segment. In contrast, MAP5 was detected in the neuronal cell bodies and all along their neurites. In conclusion, MAP2 seems involved in the early establishment of the cytoarchitecture of cell bodies and the proximal axon segment of somatosensory neurons, while MAP5 is clearly related to axonal growth.