Fast and automatic periacetabular osteotomy fragment pose estimation using intraoperatively implanted fiducials and single-view fluoroscopy.

Accurate and consistent mental interpretation of fluoroscopy to determine the position and orientation of acetabular bone fragments in 3D space is difficult. We propose a computer assisted approach that uses a single fluoroscopic view and quickly reports the pose of an acetabular fragment without any user input or initialization. Intraoperatively, but prior to any osteotomies, two constellations of metallic ball-bearings (BBs) are injected into the wing of a patient's ilium and lateral superior pubic ramus. One constellation is located on the expected acetabular fragment, and the other is located on the remaining, larger, pelvis fragment. The 3D locations of each BB are reconstructed using three fluoroscopic views and 2D/3D registrations to a preoperative CT scan of the pelvis. The relative pose of the fragment is established by estimating the movement of the two BB constellations using a single fluoroscopic view taken after osteotomy and fragment relocation. BB detection and inter-view correspondences are automatically computed throughout the processing pipeline. The proposed method was evaluated on a multitude of fluoroscopic images collected from six cadaveric surgeries performed bilaterally on three specimens. Mean fragment rotation error was 2.4 ± 1.0 degrees, mean translation error was 2.1 ± 0.6 mm, and mean 3D lateral center edge angle error was 1.0 ± 0.5 degrees. The average runtime of the single-view pose estimation was 0.7 ± 0.2 s. The proposed method demonstrates accuracy similar to other state of the art systems which require optical tracking systems or multiple-view 2D/3D registrations with manual input. The errors reported on fragment poses and lateral center edge angles are within the margins required for accurate intraoperative evaluation of femoral head coverage.

CSB cooperates with SMARCAL1 to maintain telomere stability in ALT cells.

Elevated replication stress is evident at telomeres of about 10-15% of cancer cells, which maintain their telomeres via a homologous recombination (HR)-based mechanism, referred to as alternative lengthening of telomeres (ALT). How ALT cells resolve replication stress to support their growth remains incompletely characterized. Here, we report that CSB (also known as ERCC6) promotes recruitment of HR repair proteins (MRN, BRCA1, BLM and RPA32) and POLD3 to ALT telomeres, a process that requires the ATPase activity of CSB and is controlled by ATM- and CDK2-dependent phosphorylation. Loss of CSB stimulates telomeric recruitment of MUS81 and SLX4, components of the structure-specific MUS81-EME1-SLX1-SLX4 (MUS-SLX) endonuclease complex, suggesting that CSB restricts MUS-SLX-mediated processing of stalled forks at ALT telomeres. Loss of CSB coupled with depletion of SMARCAL1, a chromatin remodeler implicated in catalyzing regression of stalled forks, synergistically promotes not only telomeric recruitment of MUS81 but also the formation of fragile telomeres, the latter of which is reported to arise from fork stalling. These results altogether suggest that CSB-mediated HR repair and SMARCAL1-mediated fork regression cooperate to prevent stalled forks from being processed into fragile telomeres in ALT cells.

The SUV4-20 inhibitor A-196 verifies a role for epigenetics in genomic integrity.

Protein lysine methyltransferases (PKMTs) regulate diverse physiological processes including transcription and the maintenance of genomic integrity. Genetic studies suggest that the PKMTs SUV420H1 and SUV420H2 facilitate proficient nonhomologous end-joining (NHEJ)-directed DNA repair by catalyzing the di- and trimethylation (me2 and me3, respectively) of lysine 20 on histone 4 (H4K20). Here we report the identification of A-196, a potent and selective inhibitor of SUV420H1 and SUV420H2. Biochemical and co-crystallization analyses demonstrate that A-196 is a substrate-competitive inhibitor of both SUV4-20 enzymes. In cells, A-196 induced a global decrease in H4K20me2 and H4K20me3 and a concomitant increase in H4K20me1. A-196 inhibited 53BP1 foci formation upon ionizing radiation and reduced NHEJ-mediated DNA-break repair but did not affect homology-directed repair. These results demonstrate the role of SUV4-20 enzymatic activity in H4K20 methylation and DNA repair. A-196 represents a first-in-class chemical probe of SUV4-20 to investigate the role of histone methyltransferases in genomic integrity.

TRF1 phosphorylation on T271 modulates telomerase-dependent telomere length maintenance as well as the formation of ALT-associated PML bodies.

TRF1, a component of the shelterin complex, plays a key role in both telomerase-dependent telomere maintenance and alternative lengthening of telomeres, the latter also known as ALT. Characteristics of ALT cells include C-circles and ALT-associated PML bodies, referred to as APBs. The function of TRF1 is tightly regulated by post-translational modification including phosphorylation, however TRF1 phosphorylation sites have yet to be fully characterized. Here we report a novel TRF1 phosphorylation site threonine 271. We show that a nonphosphorylatable mutation of T271A impairs TRF1 binding to telomeric DNA in vivo and renders TRF1 defective in inhibiting telomerase-dependent telomere elongation. On the other hand, TRF1 carrying a phosphomimic mutation of T271D is competent in not only binding to telomeric DNA but also inhibiting telomerase-mediated telomere lengthening. These results suggest that TRF1 phosphorylation on T271 negatively regulates telomerase-mediated telomere maintenance. We find that in telomerase-negative ALT cells, TRF1 carrying either a T271A or T271D mutation is able to promote C-circle production but fails to support APB formation. These results suggest that TRF1 phosphorylation on T271 is necessary for APB formation but dispensable for C-circle production. These results further imply that APB formation can be mechanistically separated from C-circle production.

Endoscopic-CT: Learning-Based Photometric Reconstruction for Endoscopic Sinus Surgery.

In this work we present a method for dense reconstruction of anatomical structures using white light endoscopic imagery based on a learning process that estimates a mapping between light reflectance and surface geometry. Our method is unique in that few unrealistic assumptions are considered (i.e., we do not assume a Lambertian reflectance model nor do we assume a point light source) and we learn a model on a per-patient basis, thus increasing the accuracy and extensibility to different endoscopic sequences. The proposed method assumes accurate video-CT registration through a combination of Structure-from-Motion (SfM) and Trimmed-ICP, and then uses the registered 3D structure and motion to generate training data with which to learn a multivariate regression of observed pixel values to known 3D surface geometry. We demonstrate with a non-linear regression technique using a neural network towards estimating depth images and surface normal maps, resulting in high-resolution spatial 3D reconstructions to an average error of 0.53mm (on the low side, when anatomy matches the CT precisely) to 1.12mm (on the high side, when the presence of liquids causes scene geometry that is not present in the CT for evaluation). Our results are exhibited on patient data and validated with associated CT scans. In total, we processed 206 total endoscopic images from patient data, where each image yields approximately 1 million reconstructed 3D points per image.

Rendering-Based Video-CT Registration with Physical Constraints for Image-Guided Endoscopic Sinus Surgery.

We present a system for registering the coordinate frame of an endoscope to pre- or intra- operatively acquired CT data based on optimizing the similarity metric between an endoscopic image and an image predicted via rendering of CT. Our method is robust and semi-automatic because it takes account of physical constraints, specifically, collisions between the endoscope and the anatomy, to initialize and constrain the search. The proposed optimization method is based on a stochastic optimization algorithm that evaluates a large number of similarity metric functions in parallel on a graphics processing unit. Images from a cadaver and a patient were used for evaluation. The registration error was 0.83 mm and 1.97 mm for cadaver and patient images respectively. The average registration time for 60 trials was 4.4 seconds. The patient study demonstrated robustness of the proposed algorithm against a moderate anatomical deformation.

Methylated TRF2 associates with the nuclear matrix and serves as a potential biomarker for cellular senescence.

Methylation of N-terminal arginines of the shelterin component TRF2 is important for cellular proliferation. While TRF2 is found at telomeres, where it plays an essential role in maintaining telomere integrity, little is known about the cellular localization of methylated TRF2. Here we report that the majority of methylated TRF2 is resistant to extraction by high salt buffer and DNase I treatment, indicating that methylated TRF2 is tightly associated with the nuclear matrix. We show that methylated TRF2 drastically alters its nuclear staining as normal human primary fibroblast cells approach and enter replicative senescence. This altered nuclear staining, which is found to be overwhelmingly associated with misshapen nuclei and abnormal nuclear matrix folds, can be suppressed by hTERT and it is barely detectable in transformed and cancer cell lines. We find that dysfunctional telomeres and DNA damage, both of which are potent inducers of cellular senescence, promote the altered nuclear staining of methylated TRF2, which is dependent upon the ATM-mediated DNA damage response. Collectively, these results suggest that the altered nuclear staining of methylated TRF2 may represent ATM-mediated nuclear structural alteration associated with cellular senescence. Our data further imply that methylated TRF2 can serve as a potential biomarker for cellular senescence.

Phosphorylated (pT371)TRF1 is recruited to sites of DNA damage to facilitate homologous recombination and checkpoint activation.

TRF1, a duplex telomeric DNA-binding protein, plays an important role in telomere metabolism. We have previously reported that a fraction of endogenous TRF1 can stably exist free of telomere chromatin when it is phosphorylated at T371 by Cdk1; however, the role of this telomere-free (pT371)TRF1 has yet to be fully characterized. Here we show that phosphorylated (pT371)TRF1 is recruited to sites of DNA damage, forming damage-induced foci in response to ionizing radiation (IR), etoposide and camptothecin. We find that IR-induced (pT371)TRF1 foci formation is dependent on the ATM- and Mre11/Rad50/Nbs1-mediated DNA damage response. While loss of functional BRCA1 impairs the formation of IR-induced (pT371)TRF1 foci, depletion of either 53BP1 or Rif1 stimulates IR-induced (pT371)TRF1 foci formation. In addition, we show that TRF1 depletion or the lack of its phosphorylation at T371 impairs DNA end resection and repair of nontelomeric DNA double-strand breaks by homologous recombination. The lack of TRF1 phosphorylation at T371 also hampers the activation of the G2/M checkpoint and sensitizes cells to PARP inhibition, IR and camptothecin. Collectively, these results reveal a novel but important function of phosphorylated (pT371)TRF1 in facilitating DNA double-strand break repair and the maintenance of genome integrity.

Deformable image registration for cone-beam CT guided transoral robotic base-of-tongue surgery.

Transoral robotic surgery (TORS) offers a minimally invasive approach to resection of base-of-tongue tumors. However, precise localization of the surgical target and adjacent critical structures can be challenged by the highly deformed intraoperative setup. We propose a deformable registration method using intraoperative cone-beam computed tomography (CBCT) to accurately align preoperative CT or MR images with the intraoperative scene. The registration method combines a Gaussian mixture (GM) model followed by a variation of the Demons algorithm. First, following segmentation of the volume of interest (i.e. volume of the tongue extending to the hyoid), a GM model is applied to surface point clouds for rigid initialization (GM rigid) followed by nonrigid deformation (GM nonrigid). Second, the registration is refined using the Demons algorithm applied to distance map transforms of the (GM-registered) preoperative image and intraoperative CBCT. Performance was evaluated in repeat cadaver studies (25 image pairs) in terms of target registration error (TRE), entropy correlation coefficient (ECC) and normalized pointwise mutual information (NPMI). Retraction of the tongue in the TORS operative setup induced gross deformation >30 mm. The mean TRE following the GM rigid, GM nonrigid and Demons steps was 4.6, 2.1 and 1.7 mm, respectively. The respective ECC was 0.57, 0.70 and 0.73, and NPMI was 0.46, 0.57 and 0.60. Registration accuracy was best across the superior aspect of the tongue and in proximity to the hyoid (by virtue of GM registration of surface points on these structures). The Demons step refined registration primarily in deeper portions of the tongue further from the surface and hyoid bone. Since the method does not use image intensities directly, it is suitable to multi-modality registration of preoperative CT or MR with intraoperative CBCT. Extending the 3D image registration to the fusion of image and planning data in stereo-endoscopic video is anticipated to support safer, high-precision base-of-tongue robotic surgery.

Conditions associated with Clostridium sporogenes growth as a surrogate for Clostridium botulinum in nonthermally processed canned butter.

The objective of this study was to better understand the effect of butter composition and emulsion structure on growth and survival of Clostridium sporogenes, used as a surrogate for C. botulinum in canned butter. The lack of a thermal process step in commercially available canned butter raises questions of potential safety, because it is hermetically sealed and generally exhibits anaerobic growth conditions, which are optimal for Clostridium botulinum growth. Without thermal processing, low-acid canned foods must have inhibitory factors present to prevent C. botulinum growth. Some potential intrinsic inhibitory factors, or hurdles, within butter include: reduced water activity, acidity in cultured products, elevated salt content, and the micro-droplet nature of the aqueous phase in the butter emulsion. It was hypothesized that a normal, intact butter emulsion would have sufficient hurdles to prevent C. botulinum growth, whereas a broken butter emulsion would result in a coalesced aqueous phase that would allow for C. botulinum growth. Batch-churned butter was inoculated with C. sporogenes; butter samples with varying salt contents (0, 0.8, 1.6, and 2.4% wt/wt NaCl) were prepared and stored in coated steel cans for varying times (1 or 2 wk) and temperatures (22 or 41°C) to determine temperature and emulsion structure effects on C. sporogenes growth. Samples stored at 41°C showed a significant increase in C. sporogenes growth compared with those stored at 22°C. Furthermore, NaCl addition was found to have a significant effect on C. sporogenes growth, with 0.8% NaCl promoting more growth than 0%, but with decreases in growth observed at 1.6 and 2.4%. Uninoculated control plates were also found to have bacterial growth; this growth was attributed to other anaerobic bacteria present within the cream. It was concluded that removal of the hurdle created by the micro-droplet size of the emulsion aqueous phase could result in C. botulinum growth even at elevated salt levels and, therefore, home preparation of canned butter is not advisable. It is also possible that commercially canned butter, if heat abused, could potentially allow for C. botulinum growth and, therefore, consumption is not recommended.

Retinal haemorrhages and related findings in abusive and non-abusive head trauma: a systematic review.

AIM: To report the retinal signs that distinguish abusive head trauma (AHT) from non-abusive head trauma (nAHT). METHODS: A systematic review of literature, 1950-2009, was conducted with standardised critical appraisal. Inclusion criteria were a strict confirmation of the aetiology, children aged <11 years and details of an examination conducted by an ophthalmologist. Post mortem data, organic disease of eye, and inadequate examinations were excluded. A multivariate logistic regression analysis was conducted to determine odds ratios (OR) and probabilities for AHT. RESULTS: Of the 62 included studies, 13 provided prevalence data (998 children, 504 AHT). Overall, retinal haemorrhages (RH) were found in 78% of AHT vs 5% of nAHT. In a child with head trauma and RH, the OR that this is AHT is 14.7 (95% confidence intervals 6.39, 33.62) and the probability of abuse is 91%. Where recorded, RH were bilateral in 83% of AHT compared with 8.3% in nAHT. RH were numerous in AHT, and few in nAHT located in the posterior pole, with only 10% extending to periphery. True prevalence of additional features, for example, retinal folds, could not be determined. CONCLUSIONS: Our systematic review confirms that although certain patterns of RH were far commoner in AHT, namely large numbers of RH in both the eyes, present in all layers of the retina, and extension into the periphery, there was no retinal sign that was unique to abusive injury. RH are rare in accidental trauma and, when present, are predominantly unilateral, few in number and in the posterior pole.

Cockayne Syndrome group B protein interacts with TRF2 and regulates telomere length and stability.

The majority of Cockayne syndrome (CS) patients carry a mutation in Cockayne Syndrome group B (CSB), a large nuclear protein implicated in DNA repair, transcription and chromatin remodeling. However, whether CSB may play a role in telomere metabolism has not yet been characterized. Here, we report that CSB physically interacts with TRF2, a duplex telomeric DNA binding protein essential for telomere protection. We find that CSB localizes at a small subset of human telomeres and that it is required for preventing the formation of telomere dysfunction-induced foci (TIF) in CS cells. We find that CS cells or CSB knockdown cells accumulate telomere doublets, the suppression of which requires CSB. We find that overexpression of CSB in CS cells promotes telomerase-dependent telomere lengthening, a phenotype that is associated with a decrease in the amount of telomere-bound TRF1, a negative mediator of telomere length maintenance. Furthermore, we show that CS cells or CSB knockdown cells exhibit misregulation of TERRA, a large non-coding telomere repeat-containing RNA important for telomere maintenance. Taken together, these results suggest that CSB is required for maintaining the homeostatic level of TERRA, telomere length and integrity. These results further imply that CS patients carrying CSB mutations may be defective in telomere maintenance.

An on-board surgical tracking and video augmentation system for C-arm image guidance.

PURPOSE: Conventional tracker configurations for surgical navigation carry a variety of limitations, including limited geometric accuracy, line-of-sight obstruction, and mismatch of the view angle with the surgeon's-eye view. This paper presents the development and characterization of a novel tracker configuration (referred to as "Tracker-on-C") intended to address such limitations by incorporating the tracker directly on the gantry of a mobile C-arm for fluoroscopy and cone-beam CT (CBCT). METHODS: A video-based tracker (MicronTracker, Claron Technology Inc., Toronto, ON, Canada) was mounted on the gantry of a prototype mobile isocentric C-arm next to the flat-panel detector. To maintain registration within a dynamically moving reference frame (due to rotation of the C-arm), a reference marker consisting of 6 faces (referred to as a "hex-face marker") was developed to give visibility across the full range of C-arm rotation. Three primary functionalities were investigated: surgical tracking, generation of digitally reconstructed radiographs (DRRs) from the perspective of a tracked tool or the current C-arm angle, and augmentation of the tracker video scene with image, DRR, and planning data. Target registration error (TRE) was measured in comparison with the same tracker implemented in a conventional in-room configuration. Graphics processing unit (GPU)-accelerated DRRs were generated in real time as an assistant to C-arm positioning (i.e., positioning the C-arm such that target anatomy is in the field-of-view (FOV)), radiographic search (i.e., a virtual X-ray projection preview of target anatomy without X-ray exposure), and localization (i.e., visualizing the location of the surgical target or planning data). Video augmentation included superimposing tracker data, the X-ray FOV, DRRs, planning data, preoperative images, and/or intraoperative CBCT onto the video scene. Geometric accuracy was quantitatively evaluated in each case, and qualitative assessment of clinical feasibility was analyzed by an experienced and fellowship-trained orthopedic spine surgeon within a clinically realistic surgical setup of the Tracker-on-C. RESULTS: The Tracker-on-C configuration demonstrated improved TRE (0.87 ± 0.25) mm in comparison with a conventional in-room tracker setup (1.92 ± 0.71) mm (p < 0.0001) attributed primarily to improved depth resolution of the stereoscopic camera placed closer to the surgical field. The hex-face reference marker maintained registration across the 180° C-arm orbit (TRE = 0.70 ± 0.32 mm). DRRs generated from the perspective of the C-arm X-ray detector demonstrated sub- mm accuracy (0.37 ± 0.20 mm) in correspondence with the real X-ray image. Planning data and DRRs overlaid on the video scene exhibited accuracy of (0.59 ± 0.38) pixels and (0.66 ± 0.36) pixels, respectively. Preclinical assessment suggested potential utility of the Tracker-on-C in a spectrum of interventions, including improved line of sight, an assistant to C-arm positioning, and faster target localization, while reducing X-ray exposure time. CONCLUSIONS: The proposed tracker configuration demonstrated sub- mm TRE from the dynamic reference frame of a rotational C-arm through the use of the multi-face reference marker. Real-time DRRs and video augmentation from a natural perspective over the operating table assisted C-arm setup, simplified radiographic search and localization, and reduced fluoroscopy time. Incorporation of the proposed tracker configuration with C-arm CBCT guidance has the potential to simplify intraoperative registration, improve geometric accuracy, enhance visualization, and reduce radiation exposure.

An in vivo comparison of hip structure analysis (HSA) with measurements obtained by QCT.

SUMMARY: In a population of elderly women, bone cross-sectional area (CSA), cross-sectional moment of inertia (CSMI), section modulus (Z), femoral neck axis length (FNAL), and width measured with hip structure analysis (HSA) on dual-energy x-ray absorptiometry (DXA) images in the femoral neck and trochanteric regions are highly correlated to quantitative computed tomography (QCT) measurements. INTRODUCTION: HSA is a method of obtaining measurements of proximal femur structure using 2D DXA technology. This study was designed to examine the correlations between HSA measurements and 3D QCT. METHODS: Forty-one women (mean age, 82.8 ± 2.5 years) were measured using DXA and a 64-slice CT scanner (1 mm slice thickness, 0.29 mm in plane resolution). HSA parameters were calculated at the narrow neck (NN) and trochanteric (IT) regions on the DXA image. These regions were then translated to anatomically equivalent regions on the QCT dataset by co-registering the DXA image and QCT dataset using four DXA images acquired at different angles. RESULTS: At the NN and IT regions, high linear correlations were measured between HSA and QCT for CSA r = 0.95 and 0.93, CSMI r = 0.94 and 0.93, and Z r = 0.93 and 0.89, respectively. All correlations were highly significant (p < 0.001), but there were differences in slope and offset between the two techniques, at least in part due to differences in calibration between the two techniques. FNAL and width of the bone at the NN and IT regions, physical measurements independent of the calibration, were highly correlated (r = 0.90-0.95, p < 0.001) and had slopes close to 1.0 (range, 0.978 to 1.003). CONCLUSION: CSA, CSMI, Z, FNAL, and width measured by HSA correlate highly to high-resolution QCT.

TREK: an integrated system architecture for intraoperative cone-beam CT-guided surgery.

PURPOSE: A system architecture has been developed for integration of intraoperative 3D imaging [viz., mobile C-arm cone-beam CT (CBCT)] with surgical navigation (e.g., trackers, endoscopy, and preoperative image and planning data). The goal of this paper is to describe the architecture and its handling of a broad variety of data sources in modular tool development for streamlined use of CBCT guidance in application-specific surgical scenarios. METHODS: The architecture builds on two proven open-source software packages, namely the cisst package (Johns Hopkins University, Baltimore, MD) and 3D Slicer (Brigham and Women's Hospital, Boston, MA), and combines data sources common to image-guided procedures with intraoperative 3D imaging. Integration at the software component level is achieved through language bindings to a scripting language (Python) and an object-oriented approach to abstract and simplify the use of devices with varying characteristics. The platform aims to minimize offline data processing and to expose quantitative tools that analyze and communicate factors of geometric precision online. Modular tools are defined to accomplish specific surgical tasks, demonstrated in three clinical scenarios (temporal bone, skull base, and spine surgery) that involve a progressively increased level of complexity in toolset requirements. RESULTS: The resulting architecture (referred to as "TREK") hosts a collection of modules developed according to application-specific surgical tasks, emphasizing streamlined integration with intraoperative CBCT. These include multi-modality image display; 3D-3D rigid and deformable registration to bring preoperative image and planning data to the most up-to-date CBCT; 3D-2D registration of planning and image data to real-time fluoroscopy; infrared, electromagnetic, and video-based trackers used individually or in hybrid arrangements; augmented overlay of image and planning data in endoscopic or in-room video; and real-time "virtual fluoroscopy" computed from GPU-accelerated digitally reconstructed radiographs (DRRs). Application in three preclinical scenarios (temporal bone, skull base, and spine surgery) demonstrates the utility of the modular, task-specific approach in progressively complex tasks. CONCLUSIONS: The design and development of a system architecture for image-guided surgery has been reported, demonstrating enhanced utilization of intraoperative CBCT in surgical applications with vastly different requirements. The system integrates C-arm CBCT with a broad variety of data sources in a modular fashion that streamlines the interface to application-specific tools, accommodates distinct workflow scenarios, and accelerates testing and translation of novel toolsets to clinical use. The modular architecture was shown to adapt to and satisfy the requirements of distinct surgical scenarios from a common code-base, leveraging software components arising from over a decade of effort within the imaging and computer-assisted interventions community.

Calculating the error in refractive error.

AIMS AND PURPOSE: To demonstrate a quick way of calculating the optical difference between two refractions using vector analysis, and to express this as a score for examination purposes. METHODS: An existing formula is applied, converted to a defocus equivalent, and then converted to a score. The formula is set out in an Excel spreadsheet. RESULTS: The spreadsheet enabled rapid assessment of the difference between two refractive errors. Examples are demonstrated. CONCLUSION: The spreadsheet was successful in enabling a comparison of any two refractions, expressing the difference either as a correcting (third) refraction, a defocus equivalent or a one-figure score.

Arginine methylation regulates telomere length and stability.

TRF2, a component of the shelterin complex, functions to protect telomeres. TRF2 contains an N-terminal basic domain rich in glycines and arginines, similar to the GAR motif that is methylated by protein arginine methyltransferases. However, whether arginine methylation regulates TRF2 function has not been determined. Here we report that amino acid substitutions of arginines with lysines in the basic domain of TRF2 induce telomere dysfunction-induced focus formation, leading to induction of cellular senescence. We have demonstrated that cells overexpressing TRF2 lysine mutants accumulate telomere doublets, indicative of telomere instability. We uncovered that TRF2 interacts with PRMT1, and its arginines in the basic domain undergo PRMT1-mediated methylation both in vitro and in vivo. We have shown that loss of PRMT1 induces growth arrest in normal human cells but has no effect on cell proliferation in cancer cells, suggesting that PRMT1 may control cell proliferation in a cell type-specific manner. We found that depletion of PRMT1 in normal human cells results in accumulation of telomere doublets, indistinguishable from overexpression of TRF2 lysine mutants. PRMT1 knockdown in cancer cells upregulates TRF2 association with telomeres, promoting telomere shortening. Taken together, these results suggest that PRMT1 may control telomere length and stability in part through TRF2 methylation.

Human XPF controls TRF2 and telomere length maintenance through distinctive mechanisms.

XPF-ERCC1, a structure-specific endonuclease, is involved in nucleotide excision repair, crosslink repair and homologous recombination. XPF-ERCC1 is also found to interact with TRF2, a duplex telomeric DNA binding protein. We have previously shown that XPF-ERCC1 is required for TRF2-promoted telomere shortening. However, whether XPF-ERCC1 by itself has a role in telomere length maintenance has not been determined. Here we report that overexpression of XPF induces telomere shortening in XPF-proficient cells whereas XPF complementation suppresses telomere lengthening in XPF-deficient cells. These results suggest that XPF-ERCC1 can function as a negative mediator of telomere length maintenance. In addition, we find that introduction of wild type XPF into XPF-deficient cells leads to over 40% reduction in TRF2 association with telomeric DNA, indicating that XPF-ERCC1 negatively regulates TRF2 binding to telomeric DNA. Furthermore, we show that XPF carrying mutations in the conserved nuclease domain fails to control TRF2 association with telomeric DNA but it is competent for modulating telomere length maintenance. These results imply that XPF-ERCC1 controls TRF2 and telomere length maintenance through two distinctive mechanisms, with the former requiring its nuclease activity. Our results further imply that TRF2 association with telomeres may be deregulated in cells derived from XPF patients.

Precision placement of instruments for minimally invasive procedures using a "needle driver" robot.

Medical practice continues to move toward less invasive procedures. Many of these procedures require the precision placement of a needle in the anatomy. Over the past several years, our research team has been investigating the use of a robotic needle driver to assist the physician in this task. This paper summarizes our work in this area. The robotic system is briefly described, followed by a description of a clinical trial in spinal nerve blockade. The robot was used under joystick control to place a 22 gauge needle in the spines of 10 patients using fluoroscopic imaging. The results were equivalent to the current manual procedure. We next describe our follow-up clinical application in lung biopsy for lung cancer screening under CT fluoroscopy. The system concept is discussed and the results of a phantom study are presented. A start-up company named ImageGuide has recently been formed to commercialize the robot. Their revised robot design is presented, along with plans to install a ceiling-mounted version of the robot in the CT fluoroscopy suite at Georgetown University.