Harvesting local cylinder autograft from adjacent vertebral body for anterior lumbar interbody fusion: surgical technique, operative feasibility and preliminary clinical results

Autogenous iliac crest has long served as the gold standard for anterior lumbar arthrodesis although added morbidity results from the bone graft harvest. Therefore, femoral ring allograft, or cages, have been used to decrease the morbidity of iliac crest bone harvesting. More recently, an experimental study in the animal showed that harvesting local bone from the anterior vertebral body and replacing the void by a radio-opaque beta-tricalcium phosphate plug was a valid concept. However, such a concept precludes theoretically the use of posterior pedicle screw fixation. At one institution a consecutive series of 21 patients underwent single- or multiple-level circumferential lumbar fusion with anterior cages and posterior pedicle screws. All cages were filled with cancellous bone harvested from the adjacent vertebral body, and the vertebral body defect was filled with a beta-tricalcium phosphate plug. The indications for surgery were failed conservative treatment of a lumbar degenerative disc disease or spondylolisthesis. The purpose of this study, therefore, was to report on the surgical technique, operative feasibility, safety, benefits, and drawbacks of this technique with our primary clinical experience. An independent researcher reviewed all data that had been collected prospectively from the onset of the study. The average age of the patients was 39.9 (26-57) years. Bone grafts were successfully harvested from 28 vertebral bodies in all but one patient whose anterior procedure was aborted due to difficulty in freeing the left common iliac vein. This case was converted to a transforaminal interbody fusion (TLIF). There was no major vascular injury. Blood loss of the anterior procedure averaged 250 ml (50-350 ml). One tricalcium phosphate bone plug was broken during its insertion, and one endplate was broken because of wrong surgical technique, which did not affect the final outcome. One patient had a right lumbar plexopathy that was not related to this special technique. There was no retrograde ejaculation, infection or pseudoarthrosis. One patient experienced a deep venous thrombosis. At the last follow up (mean 28 months) all patients had a solid lumbar spine fusion. At the 6-month follow up, the pain as assessed on the visual analog scale (VAS) decreased from 6.9 to 4.5 (33% decrease), and the Oswestry disability index (ODI) reduced from 48.0 to 31.7 with a 34% reduction. However, at 2 years follow up there was a trend for increase in the ODI (35) and VAS (5). The data in this study suggest that harvesting a cylinder of autograft from the adjacent vertebral body is safe and efficient. Filling of the void defect with a beta-tricalcium phosphate plug does not preclude the use of posterior pedicle screw stabilization.

Synergistic inhibition in cell-cell fusion mediated by the matrix and nucleocapsid protein of canine distemper virus

Canine distemper virus (CDV) causes a chronic, demyelinating, progressive or relapsing neurological disease in dogs, because CDV persists in the CNS. Persistence of virulent CDV, such as the A75/17 strain has been reproduced in cell cultures where it is associated with a non-cytolytic infection with very limited cell-cell fusion. This is in sharp contrast to attenuated CDV infection in cell cultures, such as the Onderstepoort (OP) CDV strain, which produces extensive fusion activity and cytolysis. Fusion efficiency may be determined by the structure of the viral fusion protein per se but also by its interaction with other structural proteins of CDV. This was studied by combining genes derived from persistent and non-persistent CDV strains in transient transfection experiments. It was found that fusion efficiency was markedly attenuated by the structure of the fusion protein of the neurovirulent A75/17-CDV. Moreover, we showed that the interaction of the surface glycoproteins with the M protein of the persistent strain greatly influenced fusion activity. Site directed mutagenesis showed that the c-terminus of the M protein is of particular importance in this respect. Interestingly, although the nucleocapsid protein alone did not affect F/H-induced cell-cell fusion, maximal inhibition occurred when the latter was added to combined glycoproteins with matrix protein. Thus, the present study suggests that very limited fusogenicity in virulent CDV infection, which favours persistence by limiting cell destruction involves complex interactions between all viral structural proteins.

Signal peptide and helical bundle domains of virulent canine distemper virus fusion protein restrict fusogenicity

Persistence in canine distemper virus (CDV) infection is correlated with very limited cell-cell fusion and lack of cytolysis induced by the neurovirulent A75/17-CDV compared to that of the cytolytic Onderstepoort vaccine strain. We have previously shown that this difference was at least in part due to the amino acid sequence of the fusion (F) protein (P. Plattet, J. P. Rivals, B. Zuber, J. M. Brunner, A. Zurbriggen, and R. Wittek, Virology 337:312-326, 2005). Here, we investigated the molecular mechanisms of the neurovirulent CDV F protein underlying limited membrane fusion activity. By exchanging the signal peptide between both F CDV strains or replacing it with an exogenous signal peptide, we demonstrated that this domain controlled intracellular and consequently cell surface protein expression, thus indirectly modulating fusogenicity. In addition, by serially passaging a poorly fusogenic virus and selecting a syncytium-forming variant, we identified the mutation L372W as being responsible for this change of phenotype. Intriguingly, residue L372 potentially is located in the helical bundle domain of the F(1) subunit. We showed that this mutation drastically increased fusion activity of F proteins of both CDV strains in a signal peptide-independent manner. Due to its unique structure even among morbilliviruses, our findings with respect to the signal peptide are likely to be specifically relevant to CDV, whereas the results related to the helical bundle add new insights to our growing understanding of this class of F proteins. We conclude that different mechanisms involving multiple domains of the neurovirulent A75/17-CDV F protein act in concert to limit fusion activity, preventing lysis of infected cells, which ultimately may favor viral persistence.

Cardiac image fusion from stand-alone SPECT and CT: clinical experience

Myocardial perfusion imaging with SPECT (SPECT-MPI) and 64-slice CT angiography (CTA) are both established techniques for the noninvasive evaluation of coronary artery disease (CAD). Three-dimensional (3D) SPECT/CT image fusion may offer an incremental diagnostic value by integrating both sets of information. We report our first clinical experiences with fused 3D SPECT/CT in CAD patients. METHODS: Thirty-eight consecutive patients with at least 1 perfusion defect on SPECT-MPI (1-d adenosine stress/rest SPECT with (99m)Tc-tetrofosmin) and 64-slice CTA were included. 3D volume-rendered fused SPECT/CT images were generated and compared with the findings from the side-by-side analysis with regard to coronary lesion interpretation by assigning the perfusion defects to their corresponding coronary lesion. RESULTS: The fused SPECT/CT images added information on pathophysiologic lesion severity in 27 coronary stenoses (22%) of 12 patients (29%) (P<0.001). Among 40 equivocal lesions on side-by-side analysis, the fused interpretation confirmed hemodynamic significance in 14 lesions and excluded functional relevance in 10 lesions. In 3 lesions, assignment of perfusion defect and coronary lesion appeared to be reliable on side-by-side analysis but proved to be inaccurate on fused interpretation. Added diagnostic information by SPECT/CT was more commonly found in patients with stenoses of small vessels (P=0.004) and involvement of diagonal branches (P=0.01). CONCLUSION: In addition to being intuitively convincing, 3D SPECT/CT fusion images in CAD may provide added diagnostic information on the functional relevance of coronary artery lesions.

Fluorescent annexin A1 reveals dynamics of ceramide platforms in living cells

Upon its genesis during apoptosis, ceramide promotes gross reorganization of the plasma membrane structure involving clustering of signalling molecules and an amplification of vesicle formation, fusion and trafficking. The annexins are a family of proteins, which in the presence of Ca(2+), bind to membranes containing negatively charged phospholipids. Here, we show that ceramide increases affinity of annexin A1-membrane interaction. In the physiologically relevant range of Ca(2+) concentrations, this leads to an increase in the Ca(2+)sensitivity of annexin A1-membrane interaction. In fixed cells, using a ceramide-specific antibody, we establish a direct interaction of annexin A1 with areas of the plasma membrane enriched in ceramide (ceramide platforms). In living cells, the intracellular dynamics of annexin A1 match those of plasmalemmal ceramide. Among proteins of the annexin family, the interaction with ceramide platforms is restricted to annexin A1 and is conveyed by its unique N-terminal domain. We demonstrate that intracellular Ca(2+)overload occurring at the conditions of cellular stress induces ceramide production. Using fluorescently tagged annexin A1 as a reporter for ceramide platforms and annexin A6 as a non-selective membrane marker, we visualize ceramide platforms for the first time in living cells and provide evidence for a ceramide-driven segregation and internalization of membrane-associated proteins.

The role of 3D fusion imaging in sentinel lymphadenectomy for vulvar cancer

OBJECTIVES: To evaluate the feasibility of fusion imaging compound tomography (FICT) of CT/MRI and single photon emission tomography (SPECT) versus planar scintigraphy only (plSc) in pre-surgical staging for vulvar cancer. MATERIALS AND METHODS: Analysis of consecutive patients with vulvar cancer who preoperatively underwent sentinel scintigraphy (planar and 3D-SPECT imaging) and CT or MRI. Body markers were used for exact anatomical co-registration and fusion datasets were reconstructed using SPECT and CT/MRI. The number and localisation of all intraoperatively identified and resected sentinel lymph nodes (SLN) were compared between planar and 3D fusion imaging. RESULTS: Twenty six SLN were localized on planar scintigraphy. Twelve additional SLN were identified after SPECT and CT/MRI reconstruction, all of them were confirmed intraoperatively. In seven cases where single foci were identified at plSc, fusion imaging revealed grouped individual nodes and five additional localisations were discovered at fusion imaging. In seven patients both methods identified SLN contra lateral to the primary tumor site, but only fusion imaging allowed to localise iliac SLN in four patients. All SLN predicted on fusion imaging could be localised and resected during surgery. CONCLUSIONS: Fusion imaging using SPECT and CT/MRI can detect SLN in vulvar cancer more precisely than planar imaging regarding number and anatomical localisation. FICT revealed additional information in seven out of ten cases (70%).

Evaluation of early tissue reactions after lumbar intertransverse process fusion using CT in a rabbit

OBJECTIVE: The objective of the study was to evaluate tissue reactions such as bone genesis, cartilage genesis and graft materials in the early phase of lumbar intertransverse process fusion in a rabbit model using computed tomography (CT) imaging with CT intensity (Hounsfield units) measurement, and to compare these data with histological results. MATERIALS AND METHODS: Lumbar intertransverse process fusion was performed on 18 rabbits. Four graft materials were used: autograft bone (n = 3); collagen membrane soaked with recombinant human bone morphogenetic protein-2 (rhBMP-2) (n = 5); granular calcium phosphate (n = 5); and granular calcium phosphate coated with rhBMP-2 (n = 5). All rabbits were euthanized 3 weeks post-operatively and lumbar spines were removed for CT imaging and histological examination. RESULTS: Computed tomography imaging demonstrated that each fusion mass component had the appropriate CT intensity range. CT also showed the different distributions and intensities of bone genesis in the fusion masses between the groups. Each component of tissue reactions was identified successfully on CT images using the CT intensity difference. Using CT color mapping, these observations could be easily visualized, and the results correlated well with histological findings. CONCLUSIONS: The use of CT intensity is an effective approach for observing and comparing early tissue reactions such as newly synthesized bone, newly synthesized cartilage, and graft materials after lumbar intertransverse process fusion in a rabbit model.