Robotic assisted spinal surgery--from concept to clinical practice.

After several years of product development, animal trials and human cadaver testing, the SpineAssist--a miniature bone-mounted robotic system--has recently entered clinical use. To the best of the authors' knowledge, this is the only available image-based mechanical guidance system that enables pedicle screw insertion with an overall accuracy in the range of 1 mm in both open and minimally invasive procedures. In this paper, we describe the development and clinical trial process that has brought the SpineAssist to its current state, with an emphasis on the various difficulties encountered along the way and the corresponding solutions. All aspects of product development are discussed, including mechanical design, CT-to-fluoroscopy image registration, and surgical techniques. Finally, we describe a series of preclinical trials with human cadavers, as well as clinical use, which verify the system's accuracy and efficacy.

Crystal structure of colicin E3 immunity protein: an inhibitor of a ribosome-inactivating RNase.

BACKGROUND: Colicins are antibiotic-like proteins of Escherichia coli that kill related strains. Colicin E3 acts as an RNase that specifically cleaves 16S rRNA, thereby inactivating the ribosomes in the infected cell. The producing organism is protected against colicin E3 by a specific inhibitor, the immunity protein Im3, which forms a tight 1:1 complex with colicin E3 and renders it inactive. Crystallographic studies on colicin E3 and Im3 have been undertaken to unravel the structural basis for the ribonucleolytic activity and its inhibition. RESULTS: The crystal structure of Im3 has been determined to a resolution of 1.8 A. The structure consists of a four-standard antiparallel beta sheet flanked by three alpha helices on one side of the sheet. Thr7, Phe9, Phe16 and Phe74 form a hydrophobic cluster on the surface of the protein in the vicinity of Cys47. This cluster is part of a putative binding pocket which also includes nine polar residues. CONCLUSIONS: The putative binding pocket of Im3 is the probable site of interaction with colicin E3. The six acidic residues in the pocket may interact with some of the numerous basic residues of colicin E3. The involvement of hydrophobic moieties in the binding is consistent with the observation that the tight complex can only be dissociated by denaturation. The structure of Im3 resembles those of certain nucleic acid binding proteins, in particular domain II of topoisomerase I and RNA-binding proteins that contain the ribonucleoprotein (RNP) sequence motif. This observation suggests that Im3 has a nucleic acid binding function in addition to binding colicin E3.

Febotics - a marriage of fetal surgery and robotics.

For a fetus diagnosed with a severe congenital anomaly, surgery may offer an alternative to abortion, intra-uterine death, or a life with disability. Expertise is limited however, to a few treatment centers worldwide, and there are many technical hurdles to overcome, including requirements for miniaturized instrumentation, real-time high-resolution imaging, and harmless fetal access. This article highlights recent practices in prenatal intervention and various initiatives to integrate robotics into the fetal operating room. While the number of potential patients is low, research for implementation of robotics in the field of fetal surgery is justified by the morbidity rates of current procedures, proven favorable outcomes with intervention, and the educational value with potential for extension to other medical disciplines.

Image-guided system with miniature robot for precise positioning and targeting in keyhole neurosurgery.

This paper describes a novel image-guided system for precise automatic targeting in minimally invasive keyhole neurosurgery. The system consists of the MARS miniature robot fitted with a mechanical guide for needle, probe or catheter insertion. Intraoperatively, the robot is directly affixed to a head clamp or to the patient's skull. It automatically positions itself with respect to predefined targets in a preoperative CT/MRI image following an anatomical registration with an intraoperative 3D surface scan of the patient's facial features and registration jig. We present the system architecture, surgical protocol, custom hardware (targeting and registration jig), and software modules (preoperative planning, intraoperative execution, 3D surface scan processing, and three-way registration). We also describe a prototype implementation of the system and in vitro registration experiments. Our results indicate a system-wide target registration error of 1.7 mm (standard deviation = 0.7 mm), which is close to the required 1.0-1.5 mm clinical accuracy in many keyhole neurosurgical procedures.

The 6-hydroxymethyl group of a hexose is essential for the substrate-induced closure of the cleft in hexokinase.

Yeast hexokinase B (ATP:-hexose 6-phosphotransferase, EC 2.7.1.1) was crystallized in the presence of D-xylose and ADP, and its structure was determined at 7 A resolution. The enzyme is in the 'open' conformation which is characteristic of the enzyme crystallized in the absence of glucose, rather than in the 'closed' conformation that is observed with the glucose complex. That is, the binding of xylose into the large cleft that separates the molecule into two lobes does not cause the cleft to close. We conclude, then, that the glucose 6-hydroxymethyl group (which binds to an aspartic acid and a serine) is essential for the hexose-induced conformational change.

Crystal structure of an anticholera toxin peptide complex at 2.3 A.

Cholera toxin peptide 3 (CTP3) is a 15-residue peptide corresponding in sequence to an immunogenic loop on the surface of the B-subunits of both cholera toxin and the heat-labile toxin from Escherichia coli. TE33 is the Fab fragment of a monoclonal antibody elicited against CTP3. The crystal structure of the TE33-CTP3 complex at 2.3 A resolution reveals an antigen-binding pocket, 13 A deep and 13 A wide, which is lined with many aromatic residues. The N-terminal portion of the peptide antigen CTP3 forms a type II beta-turn that fits snugly into this pocket. At gln7 the peptide backbone of CTP3 forms a kink followed by an extended C-terminal chain that seals off the cleft and buries the beta-turn underneath it. All six complementarity-determining regions of TE33 contribute to the binding of CTP3. The antibody-peptide contacts include, in addition to van der Waals' interactions and hydrogen bonds, also one salt bridge and one water molecule, which mediates the interaction.

Crystallization and preliminary X-ray diffraction studies of colicin E3 immunity protein.

Immunity protein, an inhibitor of the ribonuclease activity of the protein antibiotic colicin E3, crystallizes in the orthorhombic space group C222 with cell dimensions a = 78.7 A, b = 54.1 A, c = 36.1 A and one molecule of Mr 9800 per asymmetric unit. The crystals are suitable for high resolution X-ray analysis.

Crystallization and preliminary X-ray diffraction studies of soybean agglutinin.

Soybean agglutinin crystallizes in the monoclinic space group C2 with unit cell dimensions a = 118.6 A, b = 88.9 A, c = 165.9 A, beta = 103.0 degrees and one tetramer of 120,000 Mr per asymmetric unit. The crystals are suitable for high-resolution work.

Crystallization and preliminary X-ray crystallographic studies of rusticyanin from Thiobacillus ferrooxidans.

Rusticyanin is a 16.5 kDa type I blue copper protein isolated from Thiobacillus ferrooxidans. This organism can grow on Fe2+ as its sole energy source. Rusticyanin is thought to be a principal component in the iron respiratory electron transport chain of T. ferrooxidans. As a component of the periplasmic space of an acidophilic bacterium, rusticyanin is remarkably stable at acidic pH. It is redox-active down to pH 0.2. Crystals of rusticyanin have been grown from solutions of PEG 8000 by the hanging-drop vapor diffusion method. The crystals are orthorhombic, space group P2(1)2(1)2(1), with unit cell dimensions a = 32.36 A, b = 60.37 A, c = 74.60 A. The crystals diffract to 2.0 A resolution and they are stable in the X-ray beam for at least two days.

Single crystals of large ribosomal particles from Halobacterium marismortui diffract to 6 A.

Large, well-ordered three-dimensional crystals of 50 S ribosomal subunits from Halobacterium marismortui have been obtained by seeding. The crystals have been characterized with synchrotron X-ray radiation as monoclinic, space group P2(1), with unit cell dimensions of a = 182(+/- 5) A, b = 584(+/- 10) A, c = 186(+/- 5) A, beta = 109 degrees. At 4 degrees C, the crystals (0.6 mm X 0.6 mm X 0.1 mm) diffract to 6 A resolution and are stable in the synchrotron beam for several hours. Compact packing is reflected from the crystallographic unit cell parameters and from electron micrographs of positively stained thin sections of embedded crystals.

Crystallization of halophilic malate dehydrogenase from Halobacterium marismortui.

Malate dehydrogenase from the extreme halophile Halobacterium marismortui crystallizes in highly concentrated phosphate solution in space group 12 with cell dimensions a = 113.8 A, b = 122.8 A, c = 126.7 A, beta = 98.1 degrees. The halophilic enzyme was found to be unstable at lower concentrations of phosphate. It associates with unusually large amounts of water and salt, and the combined particle volume shows a tight fit in the unit cell.

Multiple wavelength anomalous diffraction (MAD) crystal structure of rusticyanin: a highly oxidizing cupredoxin with extreme acid stability.

The X-ray crystal structure of the oxidized form of the extremely stable and highly oxidizing cupredoxin rusticyanin from Thiobacillus ferrooxidans has been determined by the method of multiwavelength anomalous diffraction (MAD) and refined to 1.9 A resolution. Like other cupredoxins, rusticyanin is a copper-containing metalloprotein, which is composed of a core beta-sandwich fold. In rusticyanin the beta-sandwich is composed of a six- and a seven-stranded beta-sheet. Also like other cupredoxins, the copper ion is coordinated by a cluster of four conserved residues (His85, Cys138, His143, Met148) arranged in a distorted tetrahedron. Rusticyanin has a redox potential of 680 mV, roughly twice that of any other cupredoxin, and it is optimally active at pH values < or = 2. By comparison with other cupredoxins, the three-dimensional structure of rusticyanin reveals several possible sources of the chemical differences, including more ordered secondary structure and more intersheet connectivity than other cupredoxins. The acid stability and redox potential of rusticyanin may also be enhanced over other cupredoxins by a more extensive internal hydrogen bonding network and by more extensive hydrophobic interactions surrounding the copper binding site. Finally, reduction in the number of charged residues surrounding the active site may also make a major contribution to acid stability. We propose that the resulting rigid copper binding site, which is constrained by the surrounding hydrophobic environment, structurally and electronically favours Cu(I). We propose that the two extreme chemical properties of rusticyanin are interrelated; the same unique structural features that enhance acid stability also lead to elevated redox potential.

Crystal parameters of an alcohol dehydrogenase from the extreme thermophile Thermoanaerobium brockii.

A bacterial thermophilic alcohol dehydrogenase which is stable and active at 85 degrees C, has been crystallized by vapor diffusion from solutions of polyethylene glycol. A monoclinic crystal form diffracts to 2.8 A resolution and belongs to space group C2 with unit cell dimensions a = 139.0 A, b = 137.4 A, c = 80.9 A and beta = 93.23 degrees. The asymmetric unit contains four molecules which exhibit 222 point symmetry. A second crystal form is orthohombic, space group P2(1)2(1)2 with unit cell dimensions a = 168.0 A, b = 123.0 A, c = 80.0 A, and it diffracts to 3.2 A resolution.

Structural diversity in a conserved cholera toxin epitope involved in ganglioside binding.

Cholera is a widespread disease for which there is no efficient vaccine. A better understanding of the conformational rearrangements at the epitope might be very helpful for the development of a good vaccine. Cholera toxin (CT) as well as the closely related heat-labile toxin from Escherichia coli (LT) are composed of two subunits, A and B, which form an oligomeric assembly AB5. Residues 50-64 on the surface of the B subunits comprise a conserved loop (CTP3), which is involved in saccharide binding to the receptor on epithelial cells. This loop exhibits remarkable conformational plasticity induced by environmental constraints. The crystal structure of this loop is compared in the free and receptor-bound toxins as well as in the crystal and solution structures of a complex with TE33, a monoclonal antibody elicited against CTP3. In the toxins this loop forms an irregular structure connecting a beta-strand to the central alpha-helix. Ser 55 and Gln 56 exhibit considerable conformational variability in the five subunits of the unliganded toxins. Saccharide binding induces a change primarily in Ser 55 and Gln 56 to a conformation identical in all five copies. Thus, saccharide binding confers rigidity upon the loop. The conformation of CTP3 in complex with TE33 is quite different. The amino-terminal part of CTP3 forms a beta-turn that fits snugly into a deep binding pocket on TE33, in both the crystal and NMR-derived solution structure. Only 8 and 12 residues out of 15 are seen in the NMR and crystal structures, respectively. Despite these conformational differences, TE33 is cross-reactive with intact CT, albeit with a thousandfold decrease in affinity. This suggests a different interaction of TE33 with intact CT.

Purification and characterization of ribosomal proteins from the 30 S subunit of the extreme halophile Halobacterium marismortui.

Ribosomal proteins were extracted from 30 S subunits of Halobacterium marismortui under native conditions.Their separation was based on gel filtration and hydrophobic chromatography, performed at a concentration of 3.2 M KC1 to avoid denaturation. A total of nine proteins were isolated, purified and identified by partial amino-terminal sequences and two-dimension a gel electrophoresis. There is a high degree of sequence homology with 30 S proteins from H. cutirubrum, and also some with 30 (S) proteins of eubacteria.Proton NMR data indicate unfolding of the proteins in low salt. One of the proteins, however, retains its secondary structure at a salt concentration as low as 0.1 M NaCl, and even in 8 M urea. One reason for this outstanding stability could be the high proportion (50%) of ß-structure in this protein as determined from circular dichroism measurements. In general, there is a higher ß-sheet content than for 30 S proteins from Escherichia coli. Measurements of Stokes radii indicate several of the proteins to have a rather elongated shape. One of these is a complex consisting of L3/L4 and L20, similar to the LI-complex from E. co&.The presence of this 50 S complex in the preparation of the small subunit suggests a location on the interface between the subunits.

Molecular pharmacology and modeling of vasopressin receptors.

AVP receptors represent a logical target for drug development. As a new class of therapeutic agents, orally active AVP analogs could be used to treat several human pathophysiological conditions including neurogenic diabetes insipidus, the syndrome of inappropriate secretion of AVP (SIADH), congestive heart failure, arterial hypertension, liver cirrhosis, nephrotic syndrome, dysmenorrhea, and ocular hypertension. By immunoprecipitation and immunoblotting, we elucidated the phosphorylation pattern of green fluorescent protein-tagged AVP receptors and showed interactions with the specific kinases PKC and GRK5 that are agonist-, time- and receptor subtype-dependent. The tyrosine residue of the NPWIY motif present in the 7th helix of AVP receptors is rapidly and transiently phosphorylated after agonist stimulation. This phosphorylation is instrumental in the genesis of the mitogenic cascade linked to the activation of this receptor, presumably by establishing key intramolecular contacts and by participating in the creation of a scaffold of proteins that produce the activation of downstream kinases. The random screening of chemical entities and optimization of lead compounds recently resulted in the development of orally active non-peptide AVP receptor agonists and antagonists. Furthermore, the identification of the molecular determinants of receptor-ligand interactions should facilitate the development of more potent and very selective orally active compounds via the approach of structure-based drug design. We developed three-dimensional molecular docking models of peptide and non-peptide ligands to the human V1 vascular, V2 renal and V3 pituitary AVP receptors. Docking of the peptide hormone AVP to the receptor ligand binding pockets reflects its dual polar and non-polar structure, but is receptor subtype-specific. The characteristics of non-peptide AVP analogs docking to the receptors are clearly distinct from those of peptide analogs docking. Molecular modeling of the results of site-directed mutagenesis experiments performed in CHO cells stably transfected with the human AVP receptor subtypes revealed that non-peptide antagonists establish key contacts with a few amino acid residues of the receptor subtypes that are different from those involved in agonist binding. Moreover, these interactions are species-specific. These findings provide further understanding of the signal transduction pathways of AVP receptors and new leads for elucidation of drug-receptor interactions and optimization of drug design. NOTE TO THE READER: The recent cloning and molecular characterization of AVP/OT receptor subtypes call for the revision of their nomenclature. For the sake of clarity and reference to their main site of expression, we call the V1a receptor the V1 vascular receptor, the V2 receptor the V2 renal receptor and the V1b or V3 receptor the V3 pituitary receptor in the present review.

Morphometric study of the human lumbar spine for operation-workspace specifications.

STUDY DESIGN: The anatomy of the lumbar vertebrae of 55 patients was measured by use of data provided by computed tomography. On the basis of these measurements, the location of puncture points and the orientation of the surgical instruments for pedicle, vertebral body, and disc entry points were calculated for open as well as percutaneous surgery. OBJECTIVE: Normal anatomic variations of the lumbar spine were investigated to define the workspace for several spinal procedures and to define the workspace of a robot designed to guide the physician during those procedures. SUMMARY OF BACKGROUND: Several comprehensive studies of vertebrae dimensions have been conducted in the past, but they lack several dimensions that are needed to determine the exact location of the entry point and orientation of the tool, in particular when a computerized guidance system is used. METHODS: Fifty-five spinal columns (L1-L5, total 250 vertebrae) were measured by computed tomography. These data provide geometric relations that determine entry points and tool orientations for different spinal interventions. RESULTS: The workspace for spinal operations was defined on the basis of anatomic data taken from computed tomography scans. The data included 15 measurements for each vertebra that defined its shape. The processed data provided puncture points for several spinal procedures in both open and percutaneous surgery. CONCLUSIONS: This study provides additional information on vertebral structure needed to calculate accurately the entry point and tool orientation in various spinal procedures. These statistical data are also valuable for model and implant designs and for workspace specifications for a robot-assisted surgery system.

A biomechanical model of index finger dynamics.

A dynamic model of the biomechanics of the index finger for flexion-extension and abduction-adduction motion is introduced. The model takes into account all the tendons in the finger and relates to their varying moment arms during motion. A new set of moment arm coefficients and elongation equations is derived based on experimental measurements of previous studies. Constraint equations using variable coefficients are introduced and an optimization approach used to obtain the tendon forces required for any given motion and external force. The model and optimization approach are tested with data from a rapid pinch experiment as well as a hypothetical disc rotation. Good correlation is obtained with respect to electromyographic data in the literature.

A surface-matching technique for robot-assisted registration.

Successful implementation of robot-assisted surgery (RAS) requires coherent integration of spatial image data with sensing and actuating devices, each having its own coordinate system. Hence, accurate estimation of the geometric relationships between relevant reference frames, known as registration, is a crucial procedure in all RAS applications. The purpose of this paper is to present a new registration scheme, along with the results of an experimental evaluation of a robot-assisted registration method for RAS applications in orthopedics. The accuracy of the proposed registration is appropriate for specified orthopedic surgical applications such as Total Knee Replacement. The registration method is based on a surface-matching algorithm that does not require marker implants, thereby reducing surgical invasiveness. Points on the bone surface are sampled by the robot, which in turn directs the surgical tool. This technique eliminates additional coordinate transformations to an external device (such as a digitizer), resulting in increased surgical accuracy. The registration technique was tested on an RSPR six-degrees-of-freedom parallel robot specifically designed for medical applications. A six-axis force sensor attached to the robot's moving platform enables fast and accurate acquisition of positions and surface normal directions at sampled points. Sampling with a robot probe was shown to be accurate, fast, and easy to perform. The whole procedure takes about 2 min, with the robot performing most of the registration procedures, leaving the surgeon's hands free. Robotic registration was shown to provide a flawless link between preoperative planning and robotic assistance during surgery.