Evaluating target silencing by short hairpin RNA mediated by the group I intron in cultured mammalian cells

<p>Abstract</p> <p>Background</p> <p>The group I intron, a ribozyme that catalyzes its own splicing reactions in the absence of proteins <it>in vitro</it>, is a potential target for rational engineering and attracted our interest due to its potential utility in gene repair using trans-splicing. However, the ribozyme activity of a group I intron appears to be facilitated by RNA chaperones <it>in vivo</it>; therefore, the efficiency of self-splicing could be dependent on the structure around the insert site or the length of the sequence to be inserted. To better understand how ribozyme activity could be modulated in cultured mammalian cells, a group I intron was inserted into a short hairpin RNA (shRNA), and silencing of a reporter gene by the shRNA was estimated to reflect self-splicing activity <it>in vivo</it>. In addition, we appended a theophylline-binding aptamer to the ribozyme to investigate any potential effects caused by a trans-effector.</p> <p>Results</p> <p>shRNA-expression vectors in which the loop region of the shRNA was interrupted by an intron were constructed to target firefly luciferase mRNA. There was no remarkable toxicity of the shRNA-expression vectors in Cos cells, and the decrease in luciferase activity was measured as an index of the ribozyme splicing activity. In contrast, the expression of the shRNA through intron splicing was completely abolished in 293T cells, although the silencing induced by the shRNA-expressing vector alone was no different from that in the Cos cells. The splicing efficiency of the aptamer-appended intron also had implications for the potential of trans-factors to differentially promote self-splicing among cultured mammalian cells.</p> <p>Conclusions</p> <p>Silencing by shRNAs interrupted by a group I intron could be used to monitor self-splicing activity in cultured mammalian cells, and the efficiency of self-splicing appears to be affected by cell-type specific factors, demonstrating the potential effectiveness of a trans-effector.</p

Lateral Lumbar Interbody Fusion for Ossification of the Yellow Ligament in the Lumbar Spine: First Reported Case

When ossification of the yellow ligament (OYL) occurs in the lumbar spine and extends to the lateral wall of the spinal canal, facetectomy is required to remove all of the ossified lesion and achieve decompression. Subsequent posterior fixation with interbody fusion will then be necessary to prevent postoperative progression of the ossification and intervertebral instability. The technique of lateral lumbar interbody fusion (LLIF) has recently been introduced. Using this procedure, surgeons can avoid excess blood loss from the extradural venous plexus and detachment of the ossified lesion and the ventral dura mater is avoidable. We present a 55-year-old male patient with OYL at L3/4 and anterior spondylolisthesis of L4 vertebra, with concomitant ossification of the posterior longitudinal ligament, who presented with a severe gait disturbance. He underwent a 2-stage operation without complications: LLIF for L3/4 and L4/5 was performed at the initial surgery, and posterior decompression fixation using pedicle screws from L3 to L5 was performed at the second surgery. His postoperative progress was favorable, and his interbody fusion was deemed successful. Here, we present the first reported case of LLIF for OYL of the lumbar spine. This procedure can be a good option for OYL of the lumbar spine

Survey of Period Variations of Superhumps in SU UMa-Type Dwarf Novae. II: The Second Year (2009-2010)

As an extension of the project in Kato et al. (2009, arXiv:0905.1757), we collected times of superhump maxima for 61 SU UMa-type dwarf novae mainly observed during the 2009-2010 season. The newly obtained data confirmed the basic findings reported in Kato et al. (2009): the presence of stages A-C, as well as the predominance of positive period derivatives during stage B in systems with superhump periods shorter than 0.07 d. There was a systematic difference in period derivatives for systems with superhump periods longer than 0.075 d between this study and Kato et al. (2009). We suggest that this difference is possibly caused by the relative lack of frequently outbursting SU UMa-type dwarf novae in this period regime in the present study. We recorded a strong beat phenomenon during the 2009 superoutburst of IY UMa. The close correlation between the beat period and superhump period suggests that the changing angular velocity of the apsidal motion of the elliptical disk is responsible for the variation of superhump periods. We also described three new WZ Sge-type objects with established early superhumps and one with likely early superhumps. We also suggest that two systems, VX For and EL UMa, are WZ Sge-type dwarf novae with multiple rebrightenings. The O-C variation in OT J213806.6+261957 suggests that the frequent absence of rebrightenings in very short-Porb objects can be a result of sustained superoutburst plateau at the epoch when usual SU UMa-type dwarf novae return to quiescence preceding a rebrightening. We also present a formulation for a variety of Bayesian extension to traditional period analyses.Comment: 63 pages, 77 figures, 1 appendix, Accepted for publication in PASJ, data correctio

A case of thoracic disc herniation characterized by marked posture-related dynamic changes in neurological symptoms

Thoracic disc herniation is less common than lumbar and cervical disc herniations. It is usually accompanied by severe myelopathy, which often leads to surgery. Because the thoracic spine is less mobile, thoracic disc herniation is considered to be minimally affected by dynamic spine factors in cases with myelopathy. We experienced a case of thoracic disc herniation (T4/5 and T6/7) characterized by posture-related dynamic changes in neurological symptoms; that is, numbness extending from the trunk to the entire lower limbs was deteriorated in the standing and sitting positions, was relieved in the supine position, and disappeared in the prone position. In addition, the patient reported dysuria with a delay when attempting to urinate in the standing position. Computed tomographic myelography revealed diffuse idiopathic skeletal hyperostosis extending from T3 to T11, and the kyphosis angles at T1 to T11 levels were 68 degrees in the half-sitting position and 58 degrees in the prone position, showing posture-related changes. The patient underwent the posterior fusion in the prone position, by which symptoms disappeared, without undergoing disc herniotomy or laminectomy, and favorable outcomes were achieved. Thoracic disc herniation with marked posture-related neurological symptoms is extremely rare. Here we report a case presentation and literature review of pathophysiology observed in our patient

Initial hospitalization with rigorous bed rest followed by bracing and rehabilitation as an option of conservative treatment for osteoporotic vertebral fractures in elderly patients: a pilot one arm safety and feasibility study

SummaryWe assessed the safety and feasibility of a unified conservative treatment protocol for osteoporotic vertebral fractures in the elderly patients with a 24-week follow-up. Our results showed that initial hospitalization with rigorous bed rest followed by a rehabilitation program using a Jewett brace was safe and feasible in managing patients.PurposeThe purpose of this study was to prove the safety and feasibility of a unified conservative treatment protocol, which included initial hospitalization with rigorous bed rest followed by a rehabilitation program with Jewett brace for osteoporotic vertebral fractures (OVFs) in the elderly patients with a 24-week follow-up.MethodsBetween April 2012 and Mach 2015, one hundred fifty-four patients met the eligibility for this study. Radiological findings at the 3-week, 6~8-week, 24-week assessment were evaluated. Among these, 11 patients underwent early surgery within the first 2 weeks after admission and 19 patients lost follow-up. Therefore, 124 patients were assessed at the final follow-up visit.ResultsThe average vertebral instability in all the present series was 4.9 ± 4.8° at 3-week, 2.9 ± 3.5° at 6~8-week, and 1.8 ± 3.0° at 24-week follow-up visit. Delayed union was observed in 16 patients on the 24-week follow-up visit. Therefore, the present conservative treatment protocol resulted in bony union in 98 out of 124 patients (79.0%, per protocol set analysis) and 98 out of 154 patients including drop-out (63.6%, intention-to-treat analysis). There was no severe adverse event related to initial bed rest. The vertebral instability at 3-week assessment was significantly higher in the delayed union group when compared with that in the union group. Univariate analyses followed by multivariate logistic regression analysis revealed that T2-weighted image of confined high intensity on MRI and having more than 5° of vertebral instability on dynamic X-ray at 3-week assessment are the independent risk factors for delayed union of conservative treatment in the present series.ConclusionsOur results showed that initial hospitalization with rigorous bed rest followed by a rehabilitation program using a Jewett brace was safe and feasible. Therefore, the present conservative treatment protocol can be one of the acceptable treatment options in managing OVF patients

Visualization of walking speed variation-induced synchronized dynamic changes in lower limb joint angles and activity of trunk and lower limb muscles with a newly developed gait analysis system

Purpose:To evaluate a newly developed system for dynamic analysis of gait kinematics and muscle activity.Methods:We recruited 10 healthy men into this study. Analyses of three-dimensional motion and wireless surface electromyogram (EMG) were integrated to achieve synchronous measurement. The participants walked continuously for 10 min under two conditions: comfortable and quick pace. Outcome measures were joint angles of the lower limbs determined from reflective markers and myoelectric activity of trunk and lower limbs determined from EMG sensors, comparing comfortable and quick gait pace.Results:Lower limb joint angle was significantly greater at the quick pace (maximum flexion of the hip joint: 4.1°, maximum extension of hip joint: 2.3°, and maximum flexion of the knee joint while standing: 7.4°). The period of maximum flexion of the ankle joint during a walking cycle was 2.5% longer at a quick pace. EMG amplitudes of all trunk muscles significantly increased during the period of support by two legs (cervical paraspinal: 55.1%, latissimus dorsi: 31.3%, and erector spinae: 32.6%). EMG amplitudes of quadriceps, femoral biceps, and tibialis anterior increased significantly by 223%, 60.9%, and 67.4%, respectively, between the periods of heel contact and loading response. EMG amplitude of the gastrocnemius significantly increased by 102% during the heel-off period.Conclusion:Our gait analysis synchronizing three-dimensional motion and wireless surface EMG successfully visualized dynamic changes in lower limb joint angles and activity of trunk and lower limb muscles induced by various walking speeds

Lateral Lumbar Interbody Fusion for Ossification of the Yellow Ligament in the Lumbar Spine: First Reported Case

When ossification of the yellow ligament (OYL) occurs in the lumbar spine and extends to the lateral wall of the spinal canal, facetectomy is required to remove all of the ossified lesion and achieve decompression. Subsequent posterior fixation with interbody fusion will then be necessary to prevent postoperative progression of the ossification and intervertebral instability. The technique of lateral lumbar interbody fusion (LLIF) has recently been introduced. Using this procedure, surgeons can avoid excess blood loss from the extradural venous plexus and detachment of the ossified lesion and the ventral dura mater is avoidable. We present a 55-year-old male patient with OYL at L3/4 and anterior spondylolisthesis of L4 vertebra, with concomitant ossification of the posterior longitudinal ligament, who presented with a severe gait disturbance. He underwent a 2-stage operation without complications: LLIF for L3/4 and L4/5 was performed at the initial surgery, and posterior decompression fixation using pedicle screws from L3 to L5 was performed at the second surgery. His postoperative progress was favorable, and his interbody fusion was deemed successful. Here, we present the first reported case of LLIF for OYL of the lumbar spine. This procedure can be a good option for OYL of the lumbar spine

Spina Bifida Occulta with Bilateral Spondylolysis at the Thoracolumbar Junction Presenting Cauda Equina Syndrome

Several reports have described the coexistence of spina bifida occulta (SBO) and spondylolysis, but the majority of defects occur at L5. No report has described the coexistence of SBO and spondylolysis at the thoracolumbar junction. We report a case of SBO with spondylolysis at L1, presenting cauda equine syndrome. A 37-year-old man presented with a gait disorder as a result of bilateral motor weakness of the lower extremities. A plain radiograph showed local kyphosis at L1-2 as a result of severe degenerative change and wedging of the vertebral body at L1. Magnetic resonance imaging (MRI) revealed degenerative disc changes and severe canal stenosis at L1-2. Computed tomography (CT) revealed SBO and spondylolysis at L1. He was diagnosed with cauda equina syndrome related to SBO and spondylolysis at L1. Posterior interbody fusion and decompression at L1-2 were performed. After surgery, his muscle power recovered to normal strength. The possible mechanisms in this case are the strain on anterior elements as a result of disruption of the posterior elements due to SBO and spondylolysis. The coexistence of SBO and spondylolysis at the thoracolumbar junction might induce at-risk status of increased strain to the anterior elements that may cause cauda equina syndrome

A case of thoracic disc herniation characterized by marked posture-related dynamic changes in neurological symptoms

Thoracic disc herniation is less common than lumbar and cervical disc herniations. It is usually accompanied by severe myelopathy, which often leads to surgery. Because the thoracic spine is less mobile, thoracic disc herniation is considered to be minimally affected by dynamic spine factors in cases with myelopathy. We experienced a case of thoracic disc herniation (T4/5 and T6/7) characterized by posture-related dynamic changes in neurological symptoms; that is, numbness extending from the trunk to the entire lower limbs was deteriorated in the standing and sitting positions, was relieved in the supine position, and disappeared in the prone position. In addition, the patient reported dysuria with a delay when attempting to urinate in the standing position. Computed tomographic myelography revealed diffuse idiopathic skeletal hyperostosis extending from T3 to T11, and the kyphosis angles at T1 to T11 levels were 68 degrees in the half-sitting position and 58 degrees in the prone position, showing posture-related changes. The patient underwent the posterior fusion in the prone position, by which symptoms disappeared, without undergoing disc herniotomy or laminectomy, and favorable outcomes were achieved. Thoracic disc herniation with marked posture-related neurological symptoms is extremely rare. Here we report a case presentation and literature review of pathophysiology observed in our patient. Keywords: Thoracic disc herniation, Posture-related symptom, Posterior fusion, Flexion myelopath