Multiplexed imaging in live cells using pulsed interleaved excitation spectral FLIM.

Multiplexed fluorescence detection has become increasingly important in the fields of biosensing and bioimaging. Although a variety of excitation/detection optical designs and fluorescence unmixing schemes have been proposed to allow for multiplexed imaging, rapid and reliable differentiation and quantification of multiple fluorescent species at each imaging pixel is still challenging. Here we present a pulsed interleaved excitation spectral fluorescence lifetime microscopic (PIE-sFLIM) system that can simultaneously image six fluorescent tags in live cells in a single hyperspectral snapshot. Using an alternating pulsed laser excitation scheme at two different wavelengths and a synchronized 16-channel time-resolved spectral detector, our PIE-sFLIM system can effectively excite multiple fluorophores and collect their emission over a broad spectrum for analysis. Combining our system with the advanced live-cell labeling techniques and the lifetime/spectral phasor analysis, our PIE-sFLIM approach can well unmix the fluorescence of six fluorophores acquired in a single measurement, thus improving the imaging speed in live-specimen investigation.

A non-FRET DNA reporter that changes fluorescence colour upon nuclease digestion.

Fluorescence resonance energy transfer (FRET) reporters are commonly used in the final stages of nucleic acid amplification tests to indicate the presence of nucleic acid targets, where fluorescence is restored by nucleases that cleave the FRET reporters. However, the need for dual labelling and purification during manufacturing contributes to the high cost of FRET reporters. Here we demonstrate a low-cost silver nanocluster reporter that does not rely on FRET as the on/off switching mechanism, but rather on a cluster transformation process that leads to fluorescence color change upon nuclease digestion. Notably, a 90 nm red shift in emission is observed upon reporter cleavage, a result unattainable by a simple donor-quencher FRET reporter. Electrospray ionization-mass spectrometry results suggest that the stoichiometric change of the silver nanoclusters from Ag13 (in the intact DNA host) to Ag10 (in the fragments) is probably responsible for the emission colour change observed after reporter digestion. Our results demonstrate that DNA-templated silver nanocluster probes can be versatile reporters for detecting nuclease activities and provide insights into the interactions between nucleases and metallo-DNA nanomaterials.

Massively Parallel Selection of NanoCluster Beacons.

NanoCluster Beacons (NCBs) are multicolor silver nanocluster probes whose fluorescence can be activated or tuned by a proximal DNA strand called the activator. While a single-nucleotide difference in a pair of activators can lead to drastically different activation outcomes, termed polar opposite twins (POTs), it is difficult to discover new POT-NCBs using the conventional low-throughput characterization approaches. Here, a high-throughput selection method is reported that takes advantage of repurposed next-generation-sequencing chips to screen the activation fluorescence of ≈40 000 activator sequences. It is found that the nucleobases at positions 7-12 of the 18-nucleotide-long activator are critical to creating bright NCBs and positions 4-6 and 2-4 are hotspots to generate yellow-orange and red POTs, respectively. Based on these findings, a "zipper-bag" model is proposed that can explain how these hotspots facilitate the formation of distinct silver cluster chromophores and alter their chemical yields. Combining high-throughput screening with machine-learning algorithms, a pipeline is established to design bright and multicolor NCBs in silico.

Generative adversarial network enables rapid and robust fluorescence lifetime image analysis in live cells.

Fluorescence lifetime imaging microscopy (FLIM) is a powerful tool to quantify molecular compositions and study molecular states in complex cellular environment as the lifetime readings are not biased by fluorophore concentration or excitation power. However, the current methods to generate FLIM images are either computationally intensive or unreliable when the number of photons acquired at each pixel is low. Here we introduce a new deep learning-based method termed flimGANE (fluorescence lifetime imaging based on Generative Adversarial Network Estimation) that can rapidly generate accurate and high-quality FLIM images even in the photon-starved conditions. We demonstrated our model is up to 2,800 times faster than the gold standard time-domain maximum likelihood estimation (TD_MLE) and that flimGANE provides a more accurate analysis of low-photon-count histograms in barcode identification, cellular structure visualization, Förster resonance energy transfer characterization, and metabolic state analysis in live cells. With its advantages in speed and reliability, flimGANE is particularly useful in fundamental biological research and clinical applications, where high-speed analysis is critical.

Recent developments in the characterization of nucleic acid hybridization kinetics.

Hybridization of nucleic acids (NAs) is a fundamental molecular mechanism that drives many cellular processes and enables new biotechnologies as well as therapeutics. However, existing methods that measure hybridization kinetics of nucleic acids are either performed at the ensemble level or constrained to non-native physiological conditions. Recent advances in 3D single-molecule tracking techniques break these limitations by allowing multiple annealing and melting events to be observed on a single oligonucleotide freely diffusing inside a live mammalian cell. This review provides an overview of diverse approaches to measuring NA hybridization kinetics at the single-molecule level and in live cells, and concludes with a synopsis of unresolved challenges and opportunities in the live-cell hybridization kinetics measurements. Important discoveries made by NA kinetics measurements and biotechnologies that can be improved with a deeper understanding of hybridization kinetics are also described.

Deep learning-based classification of breast cancer cells using transmembrane receptor dynamics.

MOTIVATION: Motions of transmembrane receptors on cancer cell surfaces can reveal biophysical features of the cancer cells, thus providing a method for characterizing cancer cell phenotypes. While conventional analysis of receptor motions in the cell membrane mostly relies on the mean-squared displacement plots, much information is lost when producing these plots from the trajectories. Here we employ deep learning to classify breast cancer cell types based on the trajectories of epidermal growth factor receptor (EGFR). Our model is an artificial neural network trained on the EGFR motions acquired from six breast cancer cell lines of varying invasiveness and receptor status: MCF7 (hormone receptor positive), BT474 (HER2-positive), SKBR3 (HER2-positive), MDA-MB-468 (triple negative, TN), MDA-MB-231 (TN) and BT549 (TN). RESULTS: The model successfully classified the trajectories within individual cell lines with 83% accuracy and predicted receptor status with 85% accuracy. To further validate the method, epithelial-mesenchymal transition (EMT) was induced in benign MCF10A cells, noninvasive MCF7 cancer cells and highly invasive MDA-MB-231 cancer cells, and EGFR trajectories from these cells were tested. As expected, after EMT induction, both MCF10A and MCF7 cells showed higher rates of classification as TN cells, but not the MDA-MB-231 cells. Whereas deep learning-based cancer cell classifications are primarily based on the optical transmission images of cell morphology and the fluorescence images of cell organelles or cytoskeletal structures, here we demonstrated an alternative way to classify cancer cells using a dynamic, biophysical feature that is readily accessible. AVAILABILITY AND IMPLEMENTATION: A python implementation of deep learning-based classification can be found at https://github.com/soonwoohong/Deep-learning-for-EGFR-trajectory-classification. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Early-Stage Lung Cancer Diagnosis by Deep Learning-Based Spectroscopic Analysis of Circulating Exosomes.

Lung cancer has a high mortality rate, but an early diagnosis can contribute to a favorable prognosis. A liquid biopsy that captures and detects tumor-related biomarkers in body fluids has great potential for early-stage diagnosis. Exosomes, nanosized extracellular vesicles found in blood, have been proposed as promising biomarkers for liquid biopsy. Here, we demonstrate an accurate diagnosis of early-stage lung cancer, using deep learning-based surface-enhanced Raman spectroscopy (SERS) of the exosomes. Our approach was to explore the features of cell exosomes through deep learning and figure out the similarity in human plasma exosomes, without learning insufficient human data. The deep learning model was trained with SERS signals of exosomes derived from normal and lung cancer cell lines and could classify them with an accuracy of 95%. In 43 patients, including stage I and II cancer patients, the deep learning model predicted that plasma exosomes of 90.7% patients had higher similarity to lung cancer cell exosomes than the average of the healthy controls. Such similarity was proportional to the progression of cancer. Notably, the model predicted lung cancer with an area under the curve (AUC) of 0.912 for the whole cohort and stage I patients with an AUC of 0.910. These results suggest the great potential of the combination of exosome analysis and deep learning as a method for early-stage liquid biopsy of lung cancer.

Footprints of Nanoscale DNA-Silver Cluster Chromophores via Activated-Electron Photodetachment Mass Spectrometry.

DNA-templated silver clusters (AgC) are fluorescent probes and biosensors whose electronic spectra can be tuned by their DNA hosts. However, the underlying rules that relate DNA sequence and structure to DNA-AgC fluorescence and photophysics are largely empirical. Here, we employ 193 nm activated electron photodetachment (a-EPD) mass spectrometry as a hybrid MS3 approach to gain structural insight into these nanoscale chromophores. Two DNA-AgC systems are investigated with a 20 nt single-stranded DNA (ssDNA) and a 28 nt hybrid hairpin/single-stranded DNA (hpDNA). Both oligonucleotides template Ag10 clusters, but the two complexes are distinct chromophores: the former has a violet absorption at 400 nm with no observable emission, while the latter has a blue-green absorption at 490 nm with strong green emission at 550 nm. Via identification of both apo and holo (AgC-containing) sequence ions generated upon a-EPD and mapping areas of sequence dropout, specific DNA regions that encapsulate the AgC are assigned and attributed to the coordination with the DNA nucleobases. These a-EPD footprints are distinct for the two complexes. The ssDNA contacts the cluster via four nucleobases (CCTT) in the central region of the strand, whereas the hpDNA coordinates the cluster via 13 nucleobases (TTCCCGCCTTTTG) in the double-stranded region of the hairpin. This difference is consistent with prior X-ray scattering spectra and suggests that the clusters can adapt to different DNA hosts. More importantly, the a-EPD footprints directly identify the nucleobases that are in direct contact with the AgC. As these contacting nucleobases can tune the electronic structures of the Ag core and protect the AgC from collisional quenching in solution, understanding the DNA-silver contacts within these complexes will facilitate future biosensor designs.

Measuring DNA Hybridization Kinetics in Live Cells Using a Time-Resolved 3D Single-Molecule Tracking Method.

Single-molecule detection enables direct characterization of annealing/melting kinetics of nucleic acids without the need for synchronization of molecular states, but the current experiments are not carried out in a native cellular context. Here we describe an integrated 3D single-molecule tracking and lifetime measurement method that can follow individual DNA molecules diffusing inside a mammalian cell and observe multiple annealing and melting events on the same molecules. By comparing the hybridization kinetics of the same DNA strand in vitro, we found the association constants can be 13- to 163-fold higher in the molecular crowding cellular environment.

Aqueous synthesis of highly monodisperse sub-100 nm AgCl nanospheres/cubes and their plasmonic nanomesh replicas as visible-light photocatalysts and single SERS probes.

Despite the distinctive electrochemical and photocatalytic properties of nanostructured silver chloride (AgCl), the shape- and size-dependence of their properties have not been thoroughly investigated to date. The most substantial reason responsible for this incomplete study and the subsequent limited applications is the failure in controlling the structure of AgCl nanomaterials, mainly owing to the challenging synthetic conditions including organic phase and high reaction temperature. In this work, we reported a rapid one-pot room-temperature aqueous synthesis of highly monodisperse sub-100 nm AgCl nanomaterials with various shapes and sizes by controlling the precursor (Ag+ and AuCl4 -) ratios. The remaining unreacted metal precursors (Ag+ and AuCl4 -) used to produce AgClNC were subsequently reduced by ascorbic acid on the surface of the synthesized AgCl nanomaterials to form Ag/Au bimetallic nanomesh structures (AgClNC#AuAgCMs and SMs). After the removal of the AgCl nanotemplates, only nanomesh structures (AuAgCMs and SMs) were obtained. Importantly, we successfully decreased the size of the AgCl nanomaterials which were replicated into bimetallic spherical and cubic nanomesh structures that were small enough (∼100 nm) to show intense surface-plasmon-absorption bands. Based on these unique chemical and physical properties, we could take advantage of the plasmonic photocatalysis properties of the complex comprising semiconducting AgCl/metallic nanomesh replica for the complete removal of the environmentally harmful Cr6+ in the presence of sacrificial agents such as formic acid. Finally, the novel bimetallic nanomesh structures proved themselves to exhibit intense surface-enhanced Raman scattering properties in a single-particle enhancing the electromagnetic field.

Flexible and Stable Omniphobic Surfaces Based on Biomimetic Repulsive Air-Spring Structures.

In artificial biological circulation systems such as extracorporeal membrane oxygenation, surface wettability is a critical factor in blood clotting problems. Therefore, to prevent blood from clotting, omniphobic surfaces are required to repel both hydrophilic and oleophilic liquids and reduce surface friction. However, most omniphobic surfaces have been fabricated by combining chemical reagent coating and physical structures and/or using rigid materials such as silicon and metal. It is almost impossible for chemicals to be used in the omniphobic surface for biomedical devices due to durability and toxicity. Moreover, a flexible and stable omniphobic surface is difficult to be fabricated by using conventional rigid materials. This study demonstrates a flexible and stable omniphobic surface by mimicking the re-entrant structure of springtail's skin. Our surface consists of a thin nanohole membrane on supporting microstructures. This structure traps air under the membrane, which can repel the liquid on the surface like a spring and increase the contact angle regardless of liquid type. By theoretical wetting model and simulation, we confirm that the omniphobic property is derived from air trapped in the structure. Also, our surface well maintains the omniphobicity under a highly pressurized condition. As a proof of our concept and one of the real-life applications, blood experiments are performed with our flat and curved surfaces and the results including contact angle, advancing/receding angles, and residuals show significant omniphobicity. We hope that our omniphobic surface has a significant impact on blood-contacting biomedical applications.

Improving NanoCluster Beacon performance by blocking the unlabeled NC probes.

While NanoCluster Beacon (NCB) is a versatile molecular probe, it suffers from a low target-specific signal issue due to impurities. Here we show that adding a "blocker" strand to the reaction can effectively block the nonfunctional probes and enhance the target-specific signal by 14 fold at a 0.1 target/probe ratio.

Autoenhanced Raman Spectroscopy via Plasmonic Trapping for Molecular Sensing.

As a label-free and sensitive biosensor, surface-enhanced Raman spectroscopy (SERS) is a rapidly emerging technique. However, because SERS spectra are obtained in the area of light excitation and the enhancement effect can be varied depending on the position of a substrate, it is important to match the enhanced area with an illuminated spot. Here, in order to overcome such difficulty, we demonstrated a new technique combining SERS with plasmonic trapping. By plasmonic trapping, we can collect gold nanoparticles (GNPs) in the middle of initially fabricated nanobowtie structures where a laser is excited. As a result of trapping GNPs, hot-spots are formed at that area. Because SERS is measured in the area irradiated by a laser, hot-spot can be simultaneously coincided with a detection site for SERS. By using this, we detected Rhodamine 6G to 100 pM. To further verify and improve the reproducibility of our technique, we also calculated the electric field distribution, trapping force and trapping potential.

Photothermal inhibition of neural activity with near-infrared-sensitive nanotransducers.

A neural stimulation technique that can inhibit neural activity reversibly and directly without genetic modification is valuable for understating complex brain functions and treating brain diseases. Here, we propose a near-infrared (NIR)-activatable nanoplasmonic technique that can inhibit the electrical activity of neurons by utilizing gold nanorods (GNRs) as photothermal transducers on cellular membranes. The GNRs were bound onto the plasma membrane of neurons and irradiated with NIR light to induce GNR-mediated photothermal heating near the membrane. The electrical activity from the cultured neuronal networks pretreated with GNRs was immediately inhibited upon NIR irradiation, and fully restored when NIR light was removed. The degree of inhibition could be precisely modulated by tuning the laser intensity, thereby enabling restoration of firing of a hyperactive neuronal network with epileptiform activity. This nanotechnological approach to inhibit neural activity provides a powerful therapeutic platform to control cellular functions associated with disordered neural circuits.

Anterior dislodgement of a fusion cage after transforaminal lumbar interbody fusion for the treatment of isthmic spondylolisthesis.

Transforaminal lumbar interbody fusion (TLIF) is commonly used procedure for spinal fusion. However, there are no reports describing anterior cage dislodgement after surgery. This report is a rare case of anterior dislodgement of fusion cage after TLIF for the treatment of isthmic spondylolisthesis with lumbosacral transitional vertebra (LSTV). A 51-year-old man underwent TLIF at L4-5 with posterior instrumentation for the treatment of grade 1 isthmic spondylolisthesis with LSTV. At 7 weeks postoperatively, imaging studies demonstrated that banana-shaped cage migrated anteriorly and anterolisthesis recurred at the index level with pseudoarthrosis. The cage was removed and exchanged by new cage through anterior approach, and screws were replaced with larger size ones and cement augmentation was added. At postoperative 2 days of revision surgery, computed tomography (CT) showed fracture on lateral pedicle and body wall of L5 vertebra. He underwent surgery again for paraspinal decompression at L4-5 and extension of instrumentation to S1 vertebra. His back and leg pains improved significantly after final revision surgery and symptom relief was maintained during follow-up period. At 6 months follow-up, CT images showed solid fusion at L4-5 level. Careful cage selection for TLIF must be done for treatment of spondylolisthesis accompanied with deformed LSTV, especially when reduction will be attempted. Banana-shaped cage should be positioned anteriorly, but anterior dislodgement of cage and reduction failure may occur in case of a highly unstable spine. Revision surgery for the treatment of an anteriorly dislodged cage may be effectively performed using an anterior approach.

Comparison between the accuracy of percutaneous and open pedicle screw fixations in lumbosacral fusion.

BACKGROUND CONTEXT: In pedicle screw fixation, accurate insertion is essential to avoid neurological injury or weak stability. The percutaneous pedicle screw system was developed for minimally invasive spine surgery, and its safety has already been reported. However, the accuracy of percutaneous pedicle screw fixation (PPF) has not been compared with that of the open system to date. PURPOSE: To compare the accuracy of PPF with that of open pedicle screw fixation (open PF) and to investigate the risk factors associated with pedicle wall penetration. STUDY DESIGN/SETTING: A retrospective case series. PATIENT SAMPLE: The study group included 237 patients who underwent posterior pedicle screw fixation between January 2008 and October 2010 at a single institute with a total of 1,056 pedicle screw fixations completed. One hundred and twenty-six patients with 558 screws underwent open PF and 111 patients with 498 screws underwent PPF. OUTCOME MEASURES: Postoperative computerized tomography, including sagittal and coronal reformatted images. METHODS: Consecutive surgeries with either conventional open PF or PPF for anterior lumbar interbody fusion or transforaminal lumbar interbody fusion were performed. The open pedicle screw employed was from the WSH system (Winova, Seoul, Korea), and the two percutaneous pedicle screw systems were the Sextant (Medtronics, Minneapolis, MN, USA) and the Viper systems (DePuy Spine, Raynham, MA, USA). Computed tomography images were evaluated to determine pedicle wall penetration after operation. Severity was classified as mild (<3 mm), moderate (3-6 mm), and severe (≥6 mm), and the direction was assessed as medial, lateral, inferior, and superior. RESULTS: Pedicle wall penetration occurred in 75 patients (13.4%) in the open PF group and 71 patients (14.3%) in the PPF group and was not statistically different between the groups (p=.695). Assessment of the severity of the pedicle wall penetration revealed that minor penetration was the most common (open PF group, 9.7%; PPF group, 10.6%), although the distribution of the degree of severity was not statistically different between the groups (p=.863). A relatively higher incidence of lateral penetration was observed in the open PF group (66.7% vs. 43.7%), whereas medial, superior, and inferior penetrations were higher in the PPF group (p=.033). Other parameters such as age, sex, surgical method, and surgeon factors did not influence the penetration rate, but bone mineral densitometry negatively correlated with the penetration. CONCLUSIONS: Pedicle wall penetration during screw fixation was not different between the open PF and PPF groups. The lateral, paraspinal, muscle-splitting approach seems to lessen medial wall penetration, especially in the S1 vertebra. Distribution of the direction of penetration differs between the groups, with lateral wall penetration being more prominent in the open PF group. Careful placement of pedicle screws is necessary for a stronger construct because of the high incidence of penetration.

Factors affecting clinical outcomes in treating patients with grade 1 degenerative spondylolisthesis using interspinous soft stabilization with a tension band system: a minimum 5-year follow-up.

STUDY DESIGN: Retrospective clinical study. OBJECTIVE: To explore the factors influencing the clinical outcomes and motion-preserving stabilization after interspinous soft stabilization (ISS) with a tension band system for grade 1 degenerative spondylolisthesis (DS). SUMMARY OF BACKGROUND DATA: Despite increasing recognition of the benefits of dynamic stabilization systems for treating lumbar degenerative disorders, the factors affecting the clinical and radiological outcomes of these systems have rarely been identified. METHODS: Sixty-five patients (mean age, 60.3 years) who underwent ISS with a tension band system between 2002 and 2004 were analyzed. The mean follow-up period was 72.5 months. The patients were divided according to the postsurgical clinical improvements into the optimal (n = 44) and suboptimal groups (n = 21), and the radiological intergroup differences were analyzed. Multiple linear regression analysis was performed to determine the impact of the radiological factors on the clinical outcomes. RESULTS: Significant intergroup differences were observed on the follow-up clinical examination. Radiologically, total lumbar lordosis (TLL) and segmental lumbar lordosis (SLL) were significantly improved only in the optimal group, resulting in significant intergroup differences in TLL (P = 0.023), SLL (P = 0.001), and the L1 tilt (P = 0.002). All these measures were closely associated with postoperative segmental lumbar lordosis, which also was the most influential radiological variable for the clinical parameters. CONCLUSION: In the patients with grade 1 DS, the back pain relief and functional improvement following ISS were affected by the improvements in the sagittal spinal alignment through the achievement of segmental lumbar lordosis. ISS can be an alternative treatment to fusion surgery for grade 1 DS in patients who do not require fixation or reduction.

A comparison of unilateral and bilateral laminotomies for decompression of L4-L5 spinal stenosis.

STUDY DESIGN: A retrospective review of clinical and radiographic data was performed at a single institution. OBJECTIVES: To compare clinical and radiologic outcomes between unilateral and bilateral laminotomies for bilateral decompression in patients with L4­L5 spinal stenosis. SUMMARY OF BACKGROUND DATA: Laminotomy has been shown to be comparable with laminectomy with the advantage of potentially maintaining more stability by preserving more of the osseous structures. However, the comparison between unilateral and bilateral laminotomies is available only for short-term follow-up. METHODS: Fifty-three patients at one institution having decompressive surgery for L4­L5 spinal stenosis, including grade 1 degenerative spondylolisthesis without instability, were entered into this study with a minimum of 3-year follow-up. Clinical outcomes were assessed with visual analog scale for back and leg pain and the Oswestry disability index. Radiographic measurements were performed and included translational motion, angular motion, and epidural cross-sectional area. RESULTS: The average age of the patients was 62.4 years (range: 31­82). The mean follow-up period was 49.3 months (range: 40­61). Clinical outcomes and complication rates were similar in both groups. Intraoperative blood loss and operative time were less in the unilateral laminotomy group. Radiographically, the amount of increased translational motion was significantly increased in the bilateral laminotomy group (P 0.012), but the amount of increased angular motion was not significantly different (P 0.195) between the two groups. Postoperative radiographic instability was detected more frequently in bilateral laminotomy group than in the unilateral group, without statistical significance. CONCLUSIONS: Both unilateral and bilateral laminotomies provide sufficient decompression of spinal stenosis and excellent pain reduction. However, unilateral laminotomy can be performed with shorter operative times and less blood loss. Radiologically, the use of a unilateral laminotomy induces less translational motion increase after surgery; thus, it may reduce the risk of late instability when compared with a bilateral laminotomy.

Effect of lumbar angular motion on central canal diameter: positional MRI study in 491 cases.

BACKGROUND: Lumbar spinal stenosis is a common problem that is receiving attention with the advent of novel treatment procedures. Prior positional MRI studies demonstrated lumbar canal diameter changes with flexion and extension. There have not been any studies to examine the amount of spinal canal diameter change relative to the amount of angular motion. The purpose of this study was to evaluate the correlation between the lumbar canal diameter change and the angular motion quantitatively. METHODS: Positional MRI (pMRI) images for 491 patients, including 310 males and 181 females (16 years-85 years of age), were obtained with the subjects in sitting flexion 40 degree, upright, and with extension of 10 degrees within a 0.6 T Positional MRI scanner. Quantitative measurements of the canal diameter and segmental angle of each level in the sagittal midline plane were obtained for each position. Then the diameter change and angular motion were examined for correlation during flexion and extension with linear regression analysis. RESULTS: The lumbar segmental angles were lordotic in all positions except L1-2 in flexion. The changes of canal diameters were statistically correlated with the segmental angular motions during flexion and extension (P < 0.001). The amount of canal diameter change correlated with the amount of angular change and was expressed as a ratio. CONCLUSIONS: Positional MRI demonstrated the amount of spinal canal diameter change that was statistically correlated with the segmental angular motion of the spine during flexion and extension. These results may be used to predict the extent of canal diameter change when interspinous devices or positional changes are used to treat spinal stenosis and the amount of increased canal space may be predicted with the amount of angular or positional change of the spine. This may correlate with symptomatic relief and allow for improved success in the treatment of spinal stenosis.

Interspinous ligamentoplasty in the treatment of degenerative spondylolisthesis: midterm clinical results.

OBJECT Pseudarthrosis and adjacent-segment degeneration remain problems after fusion surgery. To overcome these complications, many dynamic stabilization methods have been developed. This study was conducted to elucidate the midterm results on the effectiveness of interspinous ligamentoplasty (ILP) to treat degenerative spondylolisthesis. METHODS Thirty-two consecutive surgeries involving decompression and ILP were performed by 2 surgeons at the authors' institution during 2001 and 2002. Nine patients were excluded from the study because of inadequate follow-up or radiological data, leaving a study population of 23 patients with a mean duration of follow-up of 64.6 months (range 60-77). All the patients had symptomatic spinal stenosis and Grade 1 spondylolisthesis at L4-5 level without foraminal stenosis and deformity. Clinical outcomes were evaluated by visual analog scale (VAS) for back and leg pain and the Oswestry Disability Index (ODI). Radiological measurements included segmental lordosis, total lumbar lordosis, posterior disc height, anterior slippage, angular motion, translational motion, and facet degeneration grade. Eighteen patients who had undergone bilateral laminotomy alone were included as a Control Group. RESULTS Twenty-two of the 23 patients who underwent ILP returned to their active daily lives. Symptomatic instability was less common in the ILP Group than in the Control Group (4.3% vs 27.8%). The mean postoperative VAS leg scores, VAS back scores, and ODI scores at final follow-up were significantly improved in both groups, in comparison to preoperative scores; however, the mean difference in ODI scores was significantly greater in the ILP group (29.3% vs 16.6%, p = 0.049). In radiological analysis, segmental and total lordoses were significantly increased in the ILP Group. In both groups slippage increased, disc height decreased, and angular motion was maintained, but translational motion decreased with statistical significance in the ILP Group, whereas it increased in the Control Group. Radiological instability was observed in 3 patients in the ILP Group, and 9 in the Control Group (significant between-groups difference, p = 0.016). CONCLUSIONS Interspinous ligamentoplasty is a good option treating patients with Grade 1 degenerative spondylolisthesis requiring surgery. It is less invasive and effectively stabilizes the unstable spine with a relatively small incidence of postoperative instability. Interspinous ligamentoplasty provides satisfactory clinical and radiological results at midterm follow-up.