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Kyphotic deformity after cervical laminoplasty (CLP) often leads to unfavorable neurological recovery due to insufficient indirect decompression of the spinal cord. Existing literature has described that segmental cervical instability is a contraindication for CLP because it is a potential risk factor for kyphotic changes after surgery; however, this has never been confirmed in any clinical studies.To confirm whether segmental cervical instability was an independent risk factor for postoperative kyphotic change and to examine whether segmental cervical instability led to poor neurological outcomes after CLP for cervical spondylotic myelopathy (CSM).A retrospective study PATIENT SAMPLE: Patients who underwent CLP for CSM between January 2013 and January 2021 with a follow-up period of ≥1 year were enrolled.Cervical radiographic measurements including C2-C7 lordosis (C2-7 angle), cervical sagittal vertical axis, C7 slope, flexion range of motion (fROM) and extension ROM (eROM) were assessed using neutral and flexion-extension views. Segmental cervical instability was classified into anterolisthesis (AL) of ≥2 mm displacement, retrolisthesis (RL) of ≥2 mm displacement, and translational instability (TI) of ≥3 mm translational motion. The amount of C2-7 angle loss at the follow-up period compared to the preoperative measurements was defined as cervical lordosis loss (CLL). Neurological outcomes were assessed using the recovery rate of the Japanese Orthopedic Association score (JOA-RR).CLL was compared among patients with and without segmental cervical instability. Further, multiple linear regression model for CLL was built for the evaluation with adjustment of the reported risks, including cervical sagittal vertical axis, C7 slope, fROM, eROM, and patient age together with AL, RL, and TI, as independent variables. The JOA-RR was also compared between patients with and without segmental cervical instability.A total of 138 patients (mean age, 68.7 years; 65.9% male) were included in the analysis. AL, RL, and TI were found in 12 (8.7%), 33 (23.9%), and 16 (11.6%) patients, respectively. Comparisons among the groups showed that AL led to greater CLL; however, RL and TI did not. Multiple linear regression analysis revealed that greater CLL is significantly associated with greater fROM and smaller eROM (regression coefficient [β]=0.328, 95% confidence interval: 0.178 to 0.478, p.001; β=-0.372, 95% confidence interval: -0.591 to -0.153, p=.001, respectively). However, there were no significant statistical associations in the AL, RL, and TI. Whereas, patients with AL tended to exhibit lower JOA-RR than those without AL (37.8% vs. 52.0%, p=.108).Segmental cervical instability is not the definitive driver for loss of cervical lordosis after CLP in patients with CSM; thus, is not a contraindication in and of itself. However, it is necessary to consider the indications for CLP, according to individual cases of patients with AL on baseline radiograph, which is a sign of poor neurological recovery.

Serum/glucocorticoid-regulated kinase 1 (SGK1) is predominantly expressed in endothelial cells of mouse embryos, and Sgk1 null mice show embryonic lethality due to impaired vascular formation. However, how the SGK1 expression is controlled in developing vasculature remains unknown. In this study, we first identified a proximal endothelial enhancer through lacZ reporter mouse analyses. The mouse Sgk1 proximal enhancer was narrowed down to the 5' region of the major transcription initiation site, while a human corresponding region possessed relatively weak activity. We then searched for distal enhancer candidates using in silico analyses of publicly available databases for DNase accessibility, RNA polymerase association and chromatin modification. A region approximately 500 kb distant from the human SGK1 gene was conserved in the mouse, and the mouse and human genomic fragments drove transcription restricted to embryonic endothelial cells. Minimal fragments of both proximal and distal enhancers had consensus binding elements for the ETS transcription factors, which were essential for the responsiveness to ERG, FLI1 and ETS1 proteins in luciferase assays and the endothelial lacZ reporter expression in mouse embryos. These results suggest that endothelial SGK1 expression in embryonic vasculature is maintained through at least two ETS-regulated enhancers located in the proximal and distal regions.

Although the spontaneous regression of pseudotumors after posterior fusion has been reported, the predictive factors remain unclear. We examined the radiological features that predict for the regression of retro-odontoid pseudotumors after posterior fusion, with a specific focus on cyst formation.We included 28 patients with a diagnosis of retro-odontoid pseudotumor using preoperative magnetic resonance imaging. The radiographic parameters and pseudotumor thickness were measured pre- and postoperatively. The regression rate for each pseudotumor was calculated. The presence of a cyst around the retro-odontoid pseudotumor was investigated. If present, the cyst thickness was measured. To elucidate the predictors for the postoperative regression of pseudotumors, the patients were divided into 2 cohorts: the regression group with a regression rate40% and the no-regression group with a regression rate of40%. Multivariate logistic regression analysis, including the demographic data and preoperative radiographic parameters as independent variables, was performed.The mean pseudotumor size had decreased significantly from 8.8 ± 3.6 mm preoperatively to 5.3 ± 2.0 mm postoperatively (P0.0001). The mean regression rate was 35.9% during a magnetic resonance imaging follow-up period of 8.6 months (range, 6-12 months). Cystic lesions were noted in 10 patients (35.7%) preoperatively. The mean cyst size was 4.7 ± 1.9 mm. All cysts were located dorsal to the pseudotumors and were involved at the maximum spinal compression levels. Nevertheless, all the cysts had disappeared postoperatively. Multivariate logistic regression analysis revealed that the pseudotumor regression group had had a significantly greater proportion of cysts (57.1% vs. 14.3%; odds ratio, 11.7; P = 0.013).The presence of cystic lesions protruding from retro-odontoid pseudotumors might serve as a predictive factor for the spontaneous regression of pseudotumors after posterior fusion.

Study design A retrospective study. Objective To investigate the effects of surgery on the subjective perception of bodily unsteadiness and the objective measurements of postural instability in patients with cervical compressive myelopathy (CCM). Summary of background data Several studies have demonstrated that CCM patients have impaired postural stability and investigated its surgical outcomes. However, these studies have only objectively measured postural stability by using a stabilometer or three-dimensional motion capture system. There have been no studies examining the subjective perception of postural instability in CCM patients. Materials and methods We retrospectively reviewed patients who underwent decompressive surgery for CCM. The Fall Efficacy Scale-International (FES-I) and a self-prepared questionnaire were used to evaluate subjective perception of bodily unsteadiness. To objectively assess postural instability, a stabilometric analysis was performed with the following parameters: sway area (SwA, cm2), sway velocity (SwV, cm/s), and sway density (SwD, /cm). The evaluations were performed preoperatively, during the early postoperative period (3-6 mo postoperatively), and at 1-year postoperatively in patients with CCM. The evaluation results were compared with age-matched, sex-matched, and body mass index-matched healthy subjects. Results We included 70 CCM patients and 36 healthy subjects in this study. In both the FES-I and self-prepared questionnaire, CCM patients reported significantly milder postoperative bodily unsteadiness. The stabilometric parameters were significantly improved during the postoperative period when compared with preoperative values. Nevertheless, neither the self-reported outcome measures nor stabilometric parameters of CCM patients reached the levels of those in healthy controls in the postoperative period. Conclusion This was the first study to examine CCM surgical outcomes in terms of both subjective perception and objective postural instability. While both objective postural stability and subjective perception improved following decompressive surgery, they did not reach the levels seen in healthy participants. Level of evidence Level III.

STUDY DESIGN This was a retrospective study. OBJECTIVE The objective of this study was to investigate the diagnostic utility of percutaneous ultrasonography (PUS) for postoperative epidural hematoma (EH) as a postoperative complication. SUMMARY OF BACKGROUND DATA We investigated the usefulness of PUS for determining the need of surgical evacuation of postoperative EH by comparing the postoperative magnetic resonance imaging (MRI) and PUS of the spinal cord. MATERIALS AND METHODS This study included patients who underwent cervical laminoplasty using suture anchors. Regular MRI and PUS were performed 1 week postoperatively. Whenever the patients exhibited neurological deterioration, MRI and PUS were performed. The spinal cord decompression status was classified into 3 grades using MRI and PUS. The existence of spinal pulsation was determined by PUS. RESULTS One hundred thirty-one patients were investigated. The decompression status by MRI and PUS, and the pulsation status by PUS showed a correlation with neurological deterioration (P

Funder: GlaxoSmithKline; doi: http://dx.doi.org/10.13039/100004330, BACKGROUND AND OBJECTIVES: GSK2982772 is an oral small-molecule RIPK1 inhibitor with potential therapeutic efficacy in immune-mediated inflammatory diseases (IMIDs). An inter-ethnic comparison of GSK2982772 pharmacokinetics was conducted based on data from Western (Study 1) and Japanese subjects (Study 2). METHODS: Both studies were single-centre, randomised, double-blind, placebo-controlled studies with objectives to assess the safety and characterise the pharmacokinetics of GSK2982772. Western subjects in Study 1 (NCT03305419), Part A (N = 15), were randomly assigned to receive 120 mg three times daily (TID), 240 mg TID, or 360 mg twice daily (BID) doses of GSK2982772, or placebo (TID or BID) for 1 day. Part B subjects (N = 47) received GSK2982772 120 mg TID, 240 mg TID, or placebo TID for 14 days. Japanese subjects in Study 2 (N = 13) (NCT03590613) were randomly assigned to receive TID doses of GSK2982772 60, 120, 240 mg TID or placebo TID for 1 day. RESULTS: GSK2982772 was well tolerated and adverse events were generally mild. Maximum observed plasma drug concentration (Cmax), time to reach Cmax (Tmax), area under the plasma drug concentration versus time curve after the first GSK2982772 dose (AUC(0-7)) of 120 and 240 mg, and (AUC(0-24)) values for the 120 and 240 mg TID doses over a single day were similar in Japanese and Western subjects. CONCLUSIONS: The pharmacokinetics and tolerability of GSK2982772 were similar between Western and Japanese subjects, justifying inclusion of Japanese subjects in future global clinical studies to assess the therapeutic potential of RIPK1 inhibition for the treatment of IMIDs. Clinical Trials: NCT03305419 and NCT03590613 available from http://www.clinicaltrials.gov .

Kyphotic deformity is a rare but serious complication after cervical laminoplasty (CLP), and several studies have investigated its predictors. In these studies, a kyphotic Cobb angle of 0°-5° between C2 and C7 at a certain postoperative time-point was often used to detect kyphotic deformity. However, studies considering the amount of cervical lordosis loss compared to the preoperative measurement are scarce. This study aimed to elucidate risk factors for kyphotic change after CLP by comparing patients with and without marked loss of cervical lordosis postoperatively. The study population was divided into seven patients with and 92 patients without a loss of10° of the C2-7 angle during the follow-up period compared to the preoperative measurements [cervical lordosis loss (CLL) group and no CLL (NCLL) group, respectively]; demographic characteristics, surgical information, preoperative radiographic sagittal parameters of the cervical spine, and posterior paravertebral muscle morphology evaluated by magnetic resonance imaging were compared between two groups. A univariate analysis revealed that the CLL group had significantly greater flexion range of motion (fROM) than the NCLL group (43.0° vs. 25.8°, P 0.001); however, no statistical significance was identified for other parameters. The fROM had a high capacity to discriminate between the CLL and NCLL groups (area under the receiver-operating characteristic curve, 0.880; P 0.001; 95% confidence interval, 0.589-0.974) with an optimal cutoff point of 37°. This study suggests that greater fROM is a risk factor for the development of kyphotic changes after CLP. For patients with preoperative fROM exceeding 40°, CLP should be carefully indicated.

Development of multi-chambered heart is associated with spatio-temporal regulation of gene expression. A basic helix-loop-helix transcription factor Hey2 is specifically expressed in the embryonic mouse ventricles and is indispensable for ventricular myocyte differentiation, compartment identity and morphogenesis of the heart. However, how Hey2 transcription is precisely regulated in the heart remains unclear. In this study, we identified a distal Hey2 enhancer conserved in the mouse and human to possess specific transcriptional activity in ventricular free wall myocytes at the looping stage of cardiac development. Deletion of the enhancer significantly decreased endogenous Hey2 expression in the ventricular myocardium but not in other tissues of mouse embryos. Mutation/deletion of the conserved binding sites for T-box and Gata proteins, but not NK-2 proteins, abolished the enhancer activity, and Tbx20 null mice completely lost the enhancer activity in the embryonic ventricles. Luciferase reporter analysis suggested that the ventricular enhancer activity was controlled by Tbx20 through its DNA binding and cooperative function with cardiac Gata proteins. These results delineate a regulatory mechanism of ventricular Hey2 expression and help fully understand molecular cascades in myocardial cell differentiation and cardiac morphogenesis during embryonic development.

Levocetirizine is classified as a second‐generation antihistamine. Levocetirizine is available for the treatment of allergic disorders such as allergic rhinitis and chronic idiopathic urticaria. This was a single‐center, single‐dose, open‐label, randomized, 2‐way crossover study in healthy Japanese male subjects consisting of 2 parts. Part 1 compared the bioavailability of levocetirizine oral disintegrating tablet (ODT) and levocetirizine immediate‐release tablet (IRT) taken with water in the fasted state in 24 subjects; all subjects completed this part of the trial. In part 2, the bioavailability of levocetirizine ODT without water was compared with that of levocetirizine IRT with water in the fasted state in 48 subjects; 47 subjects completed this part of the trial. Bioequivalence was demonstrated between levocetirizine IRT 5 mg and ODT 5 mg. The safety profiles were generally similar between levocetirizine ODT and levocetirizine IRT, with no serious adverse events, deaths, or adverse events leading to withdrawal reported during the study.

A retrospective multicenter study. Body mass index (BMI) is recognized as an important determinant of osteoporosis and spinal postoperative outcomes; however, the specific impact of BMI on surgery for osteoporotic vertebral fractures (OVFs) remains inconclusive. This retrospective multicenter study investigated the impact of BMI on clinical outcomes following fusion surgery for OVFs. 237 OVF patients (mean age, 74.3 years; 48 men and 189 women) with neurological symptoms who underwent spinal fusion were included in this study. Patients were grouped by World Health Organization BMI categories: low BMI (18.5 kg/m2), normal BMI (≥18.5 and 25 kg/m2), and high BMI (≥25 kg/m2). Patients' backgrounds, surgical method, radiological findings, pain measurements, activities of daily living (ADL), and postoperative complications were compared after a mean follow-up period of 4 years. As results, the proportion of patients able to walk independently was significantly smaller in the low BMI group (75.0%) compared with the normal BMI group (89.9%; P = .01) and the high BMI group (94.3%; P = .04). Improvement in the visual analogue scale for leg pain was significantly less in the low BMI group than the high BMI group (26.7 vs 42.8 mm; P = .046). Radiological evaluation, the Frankel classification, and postoperative complications were not significantly different among all 3 groups. Improvement of pain intensity and ADL in the high BMI group was equivalent or non-significantly better for some outcome measures compared with the normal BMI group. Leg pain and independent walking ability after fusion surgery for patients with OVFs improved less in the low versus the high BMI group. Surgeons may want to carefully evaluate at risk low BMI patients before fusion surgery for OVF because poor clinical results may occur.

Abstract Among neonatal cardiomyopathies, primary endocardial fibroelastosis (pEFE) remains a mysterious disease of the endomyocardium that is poorly genetically characterized, affecting 1/5000 live births and accounting for 25% of the entire pediatric dilated cardiomyopathy (DCM) with a devastating course and grave prognosis. To investigate the potential genetic contribution to pEFE, we performed integrative genomic analysis, using whole exome sequencing (WES) and RNA-seq in a female infant with confirmed pathological diagnosis of pEFE. Within regions of homozygosity in the proband genome, WES analysis revealed novel parent-transmitted homozygous mutations affecting three genes with known roles in cilia assembly or function. Among them, a novel homozygous variant [c.1943delA] of uncertain significance in ALMS1 was prioritized for functional genomic and mechanistic analysis. Loss of function mutations of ALMS1 have been implicated in Alstrom syndrome (AS) [OMIM 203800], a rare recessive ciliopathy that has been associated with cardiomyopathy. The variant of interest results in a frameshift introducing a premature stop codon. RNA-seq of the proband’s dermal fibroblasts confirmed the impact of the novel ALMS1 variant on RNA-seq reads and revealed dysregulated cellular signaling and function, including the induction of epithelial mesenchymal transition (EMT) and activation of TGFβ signaling. ALMS1 loss enhanced cellular migration in patient fibroblasts as well as neonatal cardiac fibroblasts, while ALMS1-depleted cardiomyocytes exhibited enhanced proliferation activity. Herein, we present the unique pathological features of pEFE compared to DCM and utilize integrated genomic analysis to elucidate the molecular impact of a novel mutation in ALMS1 gene in an AS case. Our report provides insights into pEFE etiology and suggests, for the first time to our knowledge, ciliopathy as a potential underlying mechanism for this poorly understood and incurable form of neonatal cardiomyopathy. Key message Primary endocardial fibroelastosis (pEFE) is a rare form of neonatal cardiomyopathy that occurs in 1/5000 live births with significant consequences but unknown etiology. Integrated genomics analysis (whole exome sequencing and RNA sequencing) elucidates novel genetic contribution to pEFE etiology. In this case, the cardiac manifestation in Alstrom syndrome is pEFE. To our knowledge, this report provides the first evidence linking ciliopathy to pEFE etiology. Infants with pEFE should be examined for syndromic features of Alstrom syndrome. Our findings lead to a better understanding of the molecular mechanisms of pEFE, paving the way to potential diagnostic and therapeutic applications.

Cells display distinct metabolic characteristics depending on its differentiation stage. The fuel type of the cells serves not only as a source of energy but also as a driver of differentiation. Glucose, the primary nutrient to the cells, is a critical regulator of rapidly growing embryos. This metabolic change is a consequence as well as a cause of changes in genetic program. Disturbance of fetal glucose metabolism such as in diabetic pregnancy is associated with congenital heart disease. In utero hyperglycemia impacts the left-right axis establishment, migration of cardiac neural crest cells, conotruncal formation and mesenchymal formation of the cardiac cushion during early embryogenesis and causes cardiac hypertrophy in late fetal stages. In this review, we focus on the role of glucose in cardiogenesis and the molecular mechanisms underlying heart diseases associated with hyperglycemia.

Background Controversy exists regarding surgical treatment of high-grade spondylolisthesis (HGS) in terms of decompression with surgical reduction or in situ fusion. In situ fusion has the advantage of being less technically demanding. However, the residual bone graft area is extremely limited, and posterolateral bone grafting is complex as the transverse process of the slipped vertebrae is located anterior to the sacral ala, which correlates with high rates of pseudoarthrosis. Methods Four cases of L5/S1 HGS complaining of low back pain were treated using our new bone graft technique with an exposed osseous end plate. Transdiskal screw holes were made from the S1 pedicle, and bone chips were packed into the L5/S disk space through the screw holes. The slipped L5 vertebra was fixed with a combination of L5/S1 transdiskal and L5 pedicle screws. Results All cases exhibited good bony fusion, and the low back pain disappeared in all patients up to the 12-month follow-up. This technique involved packing the bone into the closed space, relieving any concerns regarding bone graft migration. Conclusions Transpedicular bone graft via transdiskal screw holes is an easy and practical technique for raising the fusion rate in surgical treatment of HGS.

Diabetes mellitus (DM) can cause left ventricular (LV) diastolic dysfunction, leading to heart failure with preserved ejection fraction (HFpEF). Dipeptidyl peptidase IV (DPP-IV) inhibitors have failed to reduce hospitalization due to HF in type 2 DM (T2D) patients in a large-scale clinical trial, despite their cardiovascular protective effects. Therefore, it is important to investigate whether DPP-IV inhibitors can improve LV diastolic dysfunction in T2D patients. The aim of the study was to evaluate whether teneligliptin, the strongest of the DPP-IV inhibitors, improves LV dysfunction or prevents the worsening of LV diastolic function in T2D patients. The TOPLEVEL study is designed as an open-labeled, marker-stratified randomized, parallel-group comparison, standard treatment-controlled multicenter study. TOPLEVEL includes two marker-defined subgroups to give treatment recommendations for T2D patients with normal (E/e′

Background The effect of indirect decompression after lateral lumbar interbody fusion (LLIF) is widely acknowledged; however, its details remain unclear. This study aimed to evaluate the immediate effects of indirect decompression just after LLIF cage placement but just before posterior instrumentation, using intraoperative computed tomography myelogram (iCTM). Methods Fifty-three levels from 28 patients undergoing LLIF with iCTM, were included in this retrospective study. Radiographic parameters were obtained from preoperative computed tomography myelogram and iCTM. Segmental correction, cross-sectional areas of the spinal canal, and bilateral foramen were compared preoperatively and intraoperatively to assess the neural decompression just after LLIF cage placement. Canal stenosis status during axial computed tomography myelogram was classified into 3 grades according to modified Schizas's grading to determine the necessity of additional posterior decompression procedures. The Oswestry Disability Index was obtained before and 3 months after the operation. Results Significant improvements in all radiological parameters of segmental correction, cross-sectional areas of the spinal canal, and bilateral foramen were observed just after LLIF cage placement. However, 11 (21%) levels had insufficient neural decompression status with iCTM grade (10 central canal and 1 lateral recess stenosis), requiring further direct posterior decompression. The difference in the improvement of Oswestry Disability Index between the decompression and nondecompression group was not significant, suggesting the validity of our decision. Conclusions Detailed evaluation with iCTM revealed that adequate indirect decompression with LLIF was not always obtained, validating the intraoperative decision of further posterior decompression. This procedure, LLIF with iCTM, may reduce the risk of unnecessary direct decompression and reoperation after insufficient indirect decompression.

Many neurological disorders can present similar symptomatology to degenerative cervical myelopathy (DCM) or myeloradiculopathy (DCMR). Therefore, to avoid misdiagnosis, it is important to recognise the differential diagnosis, which has been well described in previous literature. Additionally, DCM or DCMR can also coexist with other diseases that overlap some of its clinical manifestations, which may be overlooked before cervical surgery. Nevertheless, few studies have addressed this clinical situation. In clinical practice, the diagnosis of coexisting disease with DCM or DCMR would be typically made when some symptoms persist without improvement after cervical surgery. To inform the patients of this possibility preoperatively and arrive at the early diagnosis during the postoperative period, some knowledge of the possible coexisting diseases would be necessary. In this report, we reviewed 230 patients who underwent surgery for DCM or DCMR in an academic centre to examine the prevalence and kind of underlying disease that was overlooked preoperatively. The coexisting diseases relevant to their baseline symptoms were diagnosed only after cervical surgery in three patients (1.3%) and included amyotrophic lateral sclerosis, lung cancer and polymyalgia rheumatica. The overlapping symptoms were gait difficulty, scapular pain and neck pain, respectively. Surgeons should recognise that the coexisting disease with DCM or DCMR may be overlooked before cervical surgery because of overlapping symptomatology, although its prevalence is not certainly high. Further, when the specific symptom persisted without improvement after surgery for DCM or DCMR, the patient should be comprehensively examined, considering diverse pathological conditions, not only neurological disorders.

STUDY DESIGN Retrospective cohort study. OBJECTIVE To clarify the poor patient satisfaction after lumbar spinal surgery in elderly patients. SUMMARY OF BACKGROUND DATA As the global population continues to age, it is important to consider the surgical outcome and patient satisfaction in the elderly. No studies have assessed patient satisfaction in elderly patients undergoing surgical treatment and risk factors for poor satisfaction in elderly patients after lumbar spinal surgery. MATERIALS AND METHODS A retrospective multicenter survey was performed in 169 patients aged above 80 years who underwent lumbar spinal surgery. Patients were followed up for at least 1 year after surgery. We assessed patient satisfaction from the results of surgery by using a newly developed patient questionnaire. Patients were assessed by demographic data, surgical procedures, complications, reoperation rate, pain improvement, and risk factors for poor patient satisfaction with surgery for lumbar spinal disease. RESULTS In total, 131 patients (77.5%, G-group) were satisfied and 38 patients (22.5%, P-group) were dissatisfied with surgery. The 2 groups did not differ significantly in baseline characteristics and surgical data. Postoperative visual analog scale score for low back pain and leg pain were significantly higher in the P-group than in the G-group (low back pain: G-group, 1.7±1.9 vs. P-group, 5.2±2.5, P

Background Vertebroplasty with posterior spinal fusion (VP + PSF) is one of the most widely accepted surgical techniques for treating osteoporotic vertebral collapse (OVC). Nevertheless, the effect of the extent of fusion on surgical outcomes remains to be established. This study aimed to evaluate the surgical outcomes of short- versus long-segment VP + PSF for OVC with neurological impairment in thoracolumbar spine. Methods We retrospectively collected data from 133 patients (median age, 77 years; 42 men and 91 women) from 27 university hospitals and their affiliated hospitals. We divided patients into two groups: a short-segment fusion group (S group) with 2- or 3-segment fusion (87 patients) and a long-segment fusion group (L group) with 4- through 6-segment fusion (46 patients). Surgical invasion, clinical outcomes, local kyphosis angle (LKA), and complications were evaluated. Results No significant differences between the two groups were observed in terms of neurological recovery, pain scale scores, and complications. Surgical time was shorter and blood loss was less in the S group, whereas LKA at the final follow-up and correction loss were superior in the L group. Conclusion Although less invasiveness and validity of pain and neurological relief are secured by short-segment VP + PSF, surgeons should be cautious regarding correction loss.

Background The optimal treatment of osteoporosis after reconstruction surgery for osteoporotic vertebral fractures (OVF) remains unclear. In this multicentre retrospective study, we investigated the effects of typically used agents for osteoporosis, namely, bisphosphonates (BP) and teriparatide (TP), on surgical results in patients with osteoporotic vertebral fractures. Methods Retrospectively registered data were collected from 27 universities and affiliated hospitals in Japan. We compared the effects of BP vs TP on postoperative mechanical complication rates, implant-related reoperation rates, and clinical outcomes in patients who underwent posterior instrumented fusion for OVF. Data were analysed according to whether the osteoporosis was primary or glucocorticoid-induced. Results A total of 159 patients who underwent posterior instrumented fusion for OVF were included. The overall mechanical complication rate was significantly lower in the TP group than in the BP group (BP vs TP: 73.1% vs 58.2%, p = 0.045). The screw backout rate was significantly lower and the rates of new vertebral fractures and pseudoarthrosis tended to be lower in the TP group than in the BP group. However, there were no significant differences in lumbar functional scores and visual analogue scale pain scores or in implant-related reoperation rates between the two groups. The incidence of pseudoarthrosis was significantly higher in patients with glucocorticoid-induced osteoporosis (GIOP) than in those with primary osteoporosis; however, the pseudoarthrosis rate was reduced by using TP. The use of TP also tended to reduce the overall mechanical complication rate in both primary osteoporosis and GIOP. Conclusions The overall mechanical complication rate was lower in patients who received TP than in those who received a BP postoperatively, regardless of type of osteoporosis. The incidence of pseudoarthrosis was significantly higher in patients with GIOP, but the use of TP reduced the rate of pseudoarthrosis in GIOP patients. The use of TP was effective to reduce postoperative complications for OVF patients treated with posterior fusion.

Thoracic great vessels such as the aorta and subclavian arteries are formed through dynamic remodeling of embryonic pharyngeal arch arteries (PAAs). Previous work has shown that loss of a basic helix-loop-helix transcription factor Hey1 in mice causes abnormal fourth PAA development and lethal great vessel anomalies resembling congenital malformations in humans. However, how Hey1 mediates vascular formation remains unclear. In this study, we revealed that Hey1 in vascular endothelial cells, but not in smooth muscle cells, played essential roles for PAA development and great vessel morphogenesis in mouse embryos. Tek-Cre–mediated Hey1 deletion in endothelial cells affected endothelial tube formation and smooth muscle differentiation in embryonic fourth PAAs and resulted in interruption of the aortic arch and other great vessel malformations. Cell specificity and signal responsiveness of Hey1 expression were controlled through multiple cis-regulatory regions. We found two distal genomic regions that had enhancer activity in endothelial cells and in the pharyngeal epithelium and somites, respectively. The novel endothelial enhancer was conserved across species and was specific to large-caliber arteries. Its transcriptional activity was regulated by Notch signaling in vitro and in vivo, but not by ALK1 signaling and other transcription factors implicated in endothelial cell specificity. The distal endothelial enhancer was not essential for basal Hey1 expression in mouse embryos but may likely serve for Notch-dependent transcriptional control in endothelial cells together with the proximal regulatory region. These findings help in understanding the significance and regulation of endothelial Hey1 as a mediator of multiple signaling pathways in embryonic vascular formation.

Fil: Anderson, Allison. University of Minnesota. Department of Pharmacology; Estados Unidos Fil: Masuho, Ikuo. The Scripps Research Institute, Department of Neuroscience; Estados Unidos Fil: Marrón Fernández de Velasco, Ezequiel. University of Minnesota. Department of Pharmacology; Estados Unidos Fil: Nakano, Atsushi. University of California. Department of Molecular, Cell, and Developmental Biology; Estados Unidos Fil: Birnbaumer, Lutz. National Institute of Environmental Health Sciences. Neurobiology Laboratory; Estados Unidos Fil: Birnbaumer, Lutz. Pontificia Universidad Católica Argentina. Facultad de Ciencias Médicas. Instituto de Investigaciones Biomédicas; Argentina Fil: Martemyanov, Kirill A. The Scripps Research Institute, Department of Neuroscience; Estados Unidos Fil: Wickman, Kevin. University of Minnesota. Department of Pharmacology; Estados Unidos Abstract: How G protein-coupled receptors (GPCRs) evoke specific biological outcomes while utilizing a limited array of G proteins and effectors is poorly understood, particularly in native cell systems. Here, we examined signaling evoked by muscarinic (M2R) and adenosine (A1R) receptor activation in the mouse sinoatrial node (SAN), the cardiac pacemaker. M2R and A1R activate a shared pool of cardiac G protein-gated inwardly rectifying K+ (GIRK) channels in SAN cells from adult mice, but A1R-GIRK responses are smaller and slower than M2R-GIRK responses. Recordings from mice lacking Regulator of G protein Signaling 6 (RGS6) revealed that RGS6 exerts a GPCRdependent influence on GIRK-dependent signaling in SAN cells, suppressing M2R-GIRK coupling efficiency and kinetics and A1R-GIRK signaling amplitude. Fast kinetic bioluminescence resonance energy transfer assays in transfected HEK cells showed that RGS6 prefers Gαo over Gαi as a substrate for its catalytic activity and that M2R signals preferentially via Gαo, while A1R does not discriminate between inhibitory G protein isoforms. The impact of atrial/SAN-selective ablation of Gαo or Gαi2 was consistent with these findings. Gαi2 ablation hadminimal impact onM2R-GIRK and A1R-GIRK signaling in SAN cells. In contrast, Gαo ablation decreased the amplitude and slowed the kinetics of M2R-GIRK responses, while enhancing the sensitivity and prolonging the deactivation rate of A1R-GIRK signaling. Collectively, our data show that differences in GPCR-G protein coupling preferences, and the Gαo substrate preference of RGS6, shape A1R- and M2R-GIRK signaling dynamics in mouse SAN cells.

Summary Transition from primed to naive pluripotency is associated with dynamic changes in transposable element (TE) expression and demethylation of imprinting control regions (ICRs). In mouse, ICR methylation and TE expression are each regulated by TRIM28; however, the role of TRIM28 in humans is less clear. Here, we show that a null mutation in TRIM28 causes significant alterations in TE expression in both the naive and primed states of human pluripotency, and phenotypically this has limited effects on self-renewal, instead causing a loss of germline competency. Furthermore, we discovered that TRIM28 regulates paternal ICR methylation and chromatin accessibility in the primed state, with no effects on maternal ICRs. Taken together, our study shows that abnormal TE expression is tolerated by self-renewing human pluripotent cells, whereas germline competency is not., Graphical Abstract, Highlights • Primed and naive hESC self-renewal does not depend on TRIM28 • Human germ cell formation in vitro requires TRIM28 • TRIM28 differentially regulates transposons in primed and naive hESCs • DNA methylation of H19 and MEG3 requires TRIM28, In this article, Clark and colleagues show that both primed and naive hESCs are compatible with a TRIM28 deletion, despite massive deregulation of transposable elements. However, human germline competency and paternal imprint methylation are compromised upon loss of TRIM28.

Background The optimal treatment of neurological deficits following osteoporotic vertebral collapse (OVC) is controversial, owing to complications that result from fragile bone quality. In the present study, we assessed surgical results following posterior spinal fusion without decompression. We achieved stable fusion over a short segment of the spinal column using pedicle screws and spinous process plates, maximizing the use of the preserved posterior elements. Methods We reviewed surgical data, perioperative complications, clinical outcomes, and radiographic data of 20 consecutively recruited patients with delayed neurological deficits following OVC, who experienced posterior short fusion without neural decompression. The average follow-up period was 24.3 months. The spine was typically stabilized with pedicle screws and spinous process plates from one level above to one level below the collapsed vertebrae, without using neural decompression or considerable correction of kyphosis. Results All patients experienced relief from back pain and neurological improvements of at least one modified Frankel grade. Bone unions of the collapsed vertebrae were achieved in all patients, and spinal fusions of the instrumented segments were achieved in all but one patient. The mean loss of correction was 5.9°, and the average spinal canal compromise by bone fragments was 32.4% before surgery as against 26.0% at the final follow-up time point. Fractures in adjacent or upper instrumented vertebrae were observed in four cases (20%). Conclusions Rigid augmentation by spinous process plates and an enough bed for the bone grafts were available in patients with severe osteoporosis, without neural decompression. All patients had satisfactory neurological recovery regardless of the extent of spinal canal remodeling, demonstrating that dynamic factors are the primary contributor to neurological deficits following OVC.

Background This study aimed to clarify the prognostic impact of partial pressure of end-tidal carbon dioxide (PETCO 2 ) in patients with advanced chronic heart failure (HF). Methods Forty-eight patients (mean age 43.1±11.9years, 32 males) with chronic HF (44 with non-ischemic and 4 with ischemic cardiomyopathy) were prospectively enrolled. Echocardiography, blood tests, pulmonary function testing, and PETCO 2 measurements were performed as noninvasive tests, whereas right heart catheterization and arterial blood gas analysis were conducted as invasive tests. The primary end point of this study was left ventricular assist device (LVAD) implantation or cardiac death. Results Eighteen patients underwent LVAD implantation at the Interagency Registry for Mechanically Circulatory Support (INTERMACS) profile 3 during the follow-up period, and no patient died. PETCO 2 was significantly lower in a stepwise manner with New York Heart Association functional class (class I or II, 34.2±9.3mmHg vs. class III or IV, 27.7±2.5mmHg; p 2 ≤31mmHg is an independent noninvasive predictor of LVAD implantation. Univariable and multivariable linear regression analyses showed that pulmonary arterial pressure was independently and highly correlated with PETCO 2 ( r 2 =−0.512, p Conclusions Among various noninvasive clinical parameters investigated, PETCO 2 was the independent predictor of LVAD implantation at the INTERMACS profile 3 in patients with chronic HF. Pulmonary congestion may significantly contribute to decreases in PETCO 2 in patients with HF.

Background Few studies have examined long-term outcomes after posterior spinal fusion using Cotrel-Dubousset instrumentation (CDI) for adolescent idiopathic scoliosis (AIS). Most patients with AIS are female, and their main concern is how spinal fusion will affect their future life. This study aimed to investigate the long-term health-related quality of life (HRQOL), including marital and reproductive status, of middle-aged Japanese women who underwent posterior spinal fusion using CDI for AIS in its earliest days in Japan. Methods Japanese women who were younger than 20 years of age at the time of surgery using CDI, between 1985 and 1995, were targeted. Roland-Morris Disability Questionnaire, Oswestry Disability Index, Scoliosis Research Society-22 questionnaire, and 36-Item Short-Form Survey (SF-36) were used to evaluate HRQOL. Marital and reproductive status were also investigated. These results were compared to those of healthy women controls and Japanese national data for 2015. Results Of 87 female patients, 29 (33.3%) were included, with 71 healthy women as controls. The average age of the patient group was 42.7 years (range 37–48 years), and the average follow-up period was 27.5 years (range 22–32 years). HRQOL scores in the patient group were generally lower than that in the healthy control group, although there was no significant difference between the two groups in the role component summary score (RCS) of SF-36. Marital and reproductive status were not significantly different between patient and control groups, and results for the patient group were similar to Japanese national data. Conclusions This is the first study of HRQOL in middle-aged patients who underwent posterior spinal fusion using CDI for AIS in Japan. Although HRQOL scores expect RCS of the patient group were lower than those of the healthy control group, the effects of posterior spinal fusion using CDI on women's social life and marital and reproductive statuses were minimal.

Study design Retrospective radiological analysis OBJECTIVE.: The aim of this study was to identify the effects of posterior cervical muscle swelling on C5 palsy (C5P) by evaluating early postoperative magnetic resonance (MR) images. Summary of background data Cervical laminoplasty is an established technique, but the risk of C5P after surgery has not been fully resolved. Studies have reported that lateral stretching and postoperative swelling of the multifidus muscle may cause stretching of the medial branches and cervical nerves. Methods A total of 214 C5 nerves of 107 consecutive patients who underwent laminoplasty were examined. We reviewed their demographic and surgical data and radiographic and MR images as parameters, including the axial cross-sectional area (CSA) of the posterior muscles. The patients and C5 nerves were divided into C5P and non-C5P groups. The parameters and changes were compared between the two groups to examine correlations with C5P. Results In demographic data, age, sex, history of smoking, diabetes mellitus, and preoperative Japanese Orthopedic Association scores were not significantly different between the groups. Only body mass index (BMI) was significantly higher in the C5P group. Regarding the surgical and imaging data, the number of laminoplasty, operative time, decompression trough width, cervical sagittal alignment, preoperative spinal cord rotation, and posterior shift of the spinal cord were not significantly different, but the multifidus CSA change ratio was significantly higher in the C5P group. Multiple logistic regression analyses revealed that the multifidus CSA change ratio and BMI were significant independent factors. Conclusion Multifidus swelling is associated with C5P, possibly through traction of the C5 nerve via the medial branches of the cervical dorsal rami. The medial branch is the shortest of the dorsal rami and may have the largest effect by traction force. Therefore, a gentle maneuver of the deep posterior muscles during surgery is a potential countermeasure to prevent C5P. Level of evidence 3.

There have been few reports on the incidence and risk factors of the complications after spinal fixation surgery for osteoporotic vertebral collapse (OVC) with neurological deficits. This study aimed to identify the incidence and risk factors of the complications after OVC surgery.In this retrospective multicenter study, a total of 403 patients (314 women and 89 men; mean age 73.8 years) who underwent spinal fixation surgery for OVC with neurological deficits between 2005 and 2014 were enrolled. Data on patient demographics were collected, including age, sex, body mass index, smoking, steroid use, medical comorbidities, and surgical procedures. All postoperative complications that occurred within 6 weeks were recorded. Patients were classified into two groups, namely, complication group and no complication group, and risk factors for postoperative complications were investigated by univariate and multivariate analyses.Postoperative complications occurred in 57 patients (14.1%), and the most common complication was delirium (5.7%). In the univariate analysis, the complication group was found to be older (p = 0.039) and predominantly male (p = 0.049), with higher occurrence rate of liver disease (p = 0.001) and Parkinson's disease (p = 0.039) compared with the no-complication group. In the multivariate analysis, the significant independent risk factors were age (p = 0.021; odds ratio [OR] 1.051, 95% confidence interval [CI] 1.007-1.097), liver disease (p 0.001; OR 8.993, 95% CI 2.882-28.065), and Parkinson's disease (p = 0.009; OR 3.636, 95% CI 1.378-9.599).Complications after spinal fixation surgery for OVC with neurological deficits occurred in 14.1%. Age, liver disease, and Parkinson's disease were demonstrated to be independent risk factors for postoperative complications.

Background To date, there have been little published data on surgical outcomes for patients with PD with thoracolumbar OVF. We conducted a retrospective multicenter study of registry data to investigate the outcomes of fusion surgery for patients with Parkinson’s disease (PD) with osteoporotic vertebral fracture (OVF) in the thoracolumbar junction. Methods Retrospectively registered data were collected from 27 universities and their affiliated hospitals in Japan. In total, 26 patients with PD (mean age, 76 years; 3 men and 23 women) with thoracolumbar OVF who underwent spinal fusion with a minimum of 2 years of follow-up were included (PD group). Surgical invasion, perioperative complications, radiographic sagittal alignment, mechanical failure (MF) related to instrumentation, and clinical outcomes were evaluated. A control group of 296 non-PD patients (non-PD group) matched for age, sex, distribution of surgical procedures, number of fused segments, and follow-up period were used for comparison. Results The PD group showed higher rates of perioperative complications (p

Osteoporotic vertebral fracture (OVF) with nonunion or neurological deficit may be a candidate for surgical treatment. However, some patients do not show improvement as expected. Therefore, we conducted a nationwide multicenter study to determine the predictors for postoperative poor activity of daily living (ADL) in patients with OVF. We retrospectively reviewed the case histories of 309 patients with OVF who underwent surgery. To determine the factors predicting postoperative poor ADL, uni- and multivariate statistical analyses were performed. The frequency of poor ADL at final follow-up period was 9.1%. In univariate analysis, preoperative neurological deficit (OR, 4.1; 95% CI, 1.8–10.3; P

Endocardial development involves a complex orchestration of cell fate decisions that coordinate with endoderm formation and other mesodermal cell lineages. Historically, investigations into the contribution of endocardium in the developing embryo was constrained to the heart where these cells give rise to the inner lining of the myocardium and are a major contributor to valve formation. In recent years, studies have continued to elucidate the complexities of endocardial fate commitment revealing a much broader scope of lineage potential from developing endocardium. These studies cover a wide range of species and model systems and show direct contribution or fate potential of endocardium giving rise to cardiac vasculature, blood, fibroblast, and cardiomyocyte lineages. This review focuses on the marked expansion of knowledge in the area of endocardial fate potential. This article is part of a Special Issue entitled: Cardiomyocyte Biology: Integration of Developmental and Environmental Cues in the Heart edited by Marcus Schaub and Hughes Abriel.

We aimed to investigate the clinical performance of the constructs of double-door cervical laminoplasty with suture anchors by examining bony fusion at the hinges and lamina closure. We retrospectively analysed computed tomography (CT) scans obtained preoperatively, immediately after the operation, and at follow-up of patients who underwent cervical laminoplasty using suture anchors. Hinge fracture, bony healing at the hinges, suture anchor failure, and the lamina angle (LA) were evaluated using CT. Lamina closure was defined as an LA

Mutations in DMD disrupt the reading frame, prevent dystrophin translation, and cause Duchenne muscular dystrophy (DMD). Here we describe a CRISPR/Cas9 platform applicable to 60% of DMD patient mutations. We applied the platform to DMD-derived hiPSCs where successful deletion and non-homologous end joining of up to 725 kb reframed the DMD gene. This is the largest CRISPR/Cas9-mediated deletion shown to date in DMD. Use of hiPSCs allowed evaluation of dystrophin in disease-relevant cell types. Cardiomyocytes and skeletal muscle myotubes derived from reframed hiPSC clonal lines had restored dystrophin protein. The internally deleted dystrophin was functional as demonstrated by improved membrane integrity and restoration of the dystrophin glycoprotein complex in vitro and in vivo. Furthermore, miR31 was reduced upon reframing, similar to observations in Becker muscular dystrophy. This work demonstrates the feasibility of using a single CRISPR pair to correct the reading frame for the majority of DMD patients.

The heart switches its energy substrate from glucose to fatty acids at birth, and maternal hyperglycemia is associated with congenital heart disease. However, little is known about how blood glucose impacts heart formation. Using a chemically defined human pluripotent stem-cell-derived cardiomyocyte differentiation system, we found that high glucose inhibits the maturation of cardiomyocytes at genetic, structural, metabolic, electrophysiological, and biomechanical levels by promoting nucleotide biosynthesis through the pentose phosphate pathway. Blood glucose level in embryos is stable in utero during normal pregnancy, but glucose uptake by fetal cardiac tissue is drastically reduced in late gestational stages. In a murine model of diabetic pregnancy, fetal hearts showed cardiomyopathy with increased mitotic activity and decreased maturity. These data suggest that high glucose suppresses cardiac maturation, providing a possible mechanistic basis for congenital heart disease in diabetic pregnancy., eLife digest Congenital heart disease is the most common type of birth defect, affecting nearly 1 in 100 children born. It can involve a weak heart, narrowed arteries, narrowed heart valves, or the main arteries of the heart switching places. These conditions can be fatal if untreated and often need surgery to correct. The mother’s blood sugar levels during pregnancy can have a large effect on how likely the baby is to have congenital heart disease. If a pregnant woman has poorly controlled diabetes with rapidly fluctuating sugar levels, she may be at a higher risk of having a child with the condition. High sugar levels in the mother’s blood make the baby up to five times more likely to have congenital heart disease. It has been difficult to find out exactly how sugar levels interfere with heart development because diabetes can affect the fetus in many ways. Nakano et al. used stem cells and experiments in pregnant mice with diabetes to hone in on how high sugar levels affect the fetus’s heart development. First, heart cells were grown from human stem cells, and exposed to high levels of glucose in a dish. This revealed a new mechanism for how high sugar levels affect heart formation: the cells created too many nucleotides, the building blocks of molecules such as DNA. It turns out that high glucose levels boosted a chemical process in the cell known as the pentose phosphate pathway. Some of the products of this pathway are nucleotides. This made the cells divide rapidly, but did not allow them to mature well compared with cells exposed to normal levels of sugar. In another experiment, Nakano et al. found similar results in pregnant diabetic mice. The heart cells in mouse fetuses also divided quickly but matured slowly when exposed to high sugar levels. An estimated 60 million women at an age to have children have diabetes. These new findings help us to understand why and how these women are more likely to have children with congenital heart disease, and further study will hopefully lead to a better way to prevent this condition.

The angiotensin II type 1 receptor (AT1) is a 7-transmembrane domain GPCR that when activated by its ligand angiotensin II, generates signaling events promoting vascular dysfunction and the development of cardiovascular disease. Here, we show that the single-transmembrane oxidized LDL (oxLDL) receptor (LOX-1) resides in proximity to AT1 on cell-surface membranes and that binding of oxLDL to LOX-1 can allosterically activate AT1-dependent signaling events. oxLDL-induced signaling events in human vascular endothelial cells were abolished by knockdown of AT1 and inhibited by AT1 blockade (ARB). oxLDL increased cytosolic G protein by 350% in Chinese hamster ovary (CHO) cells with genetically induced expression of AT1 and LOX-1, whereas little increase was observed in CHO cells expressing only LOX-1. Immunoprecipitation and in situ proximity ligation assay (PLA) assays in CHO cells revealed the presence of cell-surface complexes involving LOX-1 and AT1. Chimeric analysis showed that oxLDL-induced AT1 signaling events are mediated via interactions between the intracellular domain of LOX-1 and AT1 that activate AT1. oxLDL-induced impairment of endothelium-dependent vascular relaxation of vascular ring from mouse thoracic aorta was abolished by ARB or genetic deletion of AT1. These findings reveal a novel pathway for AT1 activation and suggest a new mechanism whereby oxLDL may be promoting risk for cardiovascular disease.

Cardiovascular diseases, which lead to cardiovascular events including death, progress with many deleterious pathophysiological sequels. If a cause-and-effect relationship follows a one-to-one relation, we can focus on a cause to treat an effect, but such a relation cannot be applied in cardiovascular diseases. To identify novel drugs in the cardiovascular field, we generally adopt two different strategies: induction and deduction. In the cardiovascular field, it is difficult to use deduction because cardiovascular diseases are caused by many factors, leading us to use induction. In this method, we consider all clinical data, such as medical records or genetic data, and identify a few candidates. Recent computational and mathematical advances enable us to use data-mining methods to uncover hidden relationships between many parameters and clinical outcomes. However, because these candidates are not identified as promoting or inhibiting factors, or as causal or consequent factors of cardiovascular diseases, we need to test them in basic research, and bring them back to the clinical field to test their efficacy in clinical trials. With such a "back-and-forth loop" between clinical observation and basic research, data-mining methods may provide novel strategies leading to new tools for clinicians, basic findings for researchers, and better outcomes for patients.

Kim et al. identify a novel shear stress–induced pathway involving protein kinase A, CREB, and bone morphogenetic protein that regulates hematopoietic stem cell generation in the embryonic aorta., Fluid shear stress promotes the emergence of hematopoietic stem cells (HSCs) in the aorta–gonad–mesonephros (AGM) of the developing mouse embryo. We determined that the AGM is enriched for expression of targets of protein kinase A (PKA)–cAMP response element-binding protein (CREB), a pathway activated by fluid shear stress. By analyzing CREB genomic occupancy from chromatin-immunoprecipitation sequencing (ChIP-seq) data, we identified the bone morphogenetic protein (BMP) pathway as a potential regulator of CREB. By chemical modulation of the PKA–CREB and BMP pathways in isolated AGM VE-cadherin+ cells from mid-gestation embryos, we demonstrate that PKA–CREB regulates hematopoietic engraftment and clonogenicity of hematopoietic progenitors, and is dependent on secreted BMP ligands through the type I BMP receptor. Finally, we observed blunting of this signaling axis using Ncx1-null embryos, which lack a heartbeat and intravascular flow. Collectively, we have identified a novel PKA–CREB–BMP signaling pathway downstream of shear stress that regulates HSC emergence in the AGM via the endothelial-to-hematopoietic transition.

In the previous investigations, we showed that intravenous immunoglobulin (IVIG) prevented cytokine release in procalcitonin (PCT)-stimulated monocytic cells. The aim of the present study was to investigate the underlying mechanisms of inhibition of IVIG on cytokine production in PCT-stimulated THP-1 cells. THP-1 cells treated with phorbol myristate acetate were stimulated with PCT. The protein levels of pro-inflammatory cytokines [tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and high-mobility group box 1 (HMGB1)] in the culture supernatants were determined using enzyme-linked immunosorbent assay kits. The mRNA level of TNF-α was determined by reverse transcription-polymerase chain reaction. The phosphorylations of nuclear factor kappa B (NFκB) and the mitogen-activated protein kinases (MAPKs) were determined by Western blotting. IVIG reduced mRNA expression and protein production of TNF-α in PCT-stimulated THP-1 cells. Not only IVIG but also both the Fc fragment and the F(ab’)2 fragment inhibited PCT-induced TNF-α, IL-6, and HMGB1 production. Furthermore, IVIG and its fragments suppressed PCT-induced phosphorylations of NFκB, p38 MAPK, and c-Jun N-terminal kinase. Our results indicate that IVIG prevents PCT-induced cytokine production mediated by not only the Fab region but also the Fc region. The activity of IVIG and its fragments might be regulated by inhibiting NFκB and MAPKs pathways in THP-1 cells.

The oxidative phosphorylation (OXPHOS) system generates most of the ATP in respiring cells. ATP-depleting conditions, such as hypoxia, trigger responses that promote ATP production. However, how OXPHOS is regulated during hypoxia has yet to be elucidated. In this study, selective measurement of intramitochondrial ATP levels identified the hypoxia-inducible protein G0/G1 switch gene 2 (G0s2) as a positive regulator of OXPHOS. A mitochondria-targeted, FRET-based ATP biosensor enabled us to assess OXPHOS activity in living cells. Mitochondria-targeted, FRET-based ATP biosensor and ATP production assay in a semiintact cell system revealed that G0s2 increases mitochondrial ATP production. The expression of G0s2 was rapidly and transiently induced by hypoxic stimuli, and G0s2 interacts with OXPHOS complex V (FoF1-ATP synthase). Furthermore, physiological enhancement of G0s2 expression prevented cells from ATP depletion and induced a cellular tolerance for hypoxic stress. These results show that G0s2 positively regulates OXPHOS activity by interacting with FoF1-ATP synthase, which causes an increase in ATP production in response to hypoxic stress and protects cells from a critical energy crisis. These findings contribute to the understanding of a unique stress response to energy depletion. Additionally, this study shows the importance of assessing intramitochondrial ATP levels to evaluate OXPHOS activity in living cells.

Objectives: In the present study, we aimed to evaluate the diagnostic accuracy and suitability of the ‘Tibial Nerve Compression Test (TNCT)’ as a screening tool for lumbar spinal canal stenosis (LSS). Methods: A total of 108 consecutive patients admitted to our hospital for surgical treatment or diagnosis of LSS were included in this study. Fifty healthy volunteers were examined as a control group. The severity of tenderness was scored (tenderness score) and measured on a visual analogue scale (P-VAS score). These scores were compared between the LSS and control groups. Moreover, they were compared before and after the operation among operated patients. Results: The positive tenderness rate was significantly higher (92.6% [100/108]) in the LSS group than in the control group (30% [15/50]). The sensitivity and specificity of TNCT (95% confidence interval) were 0.93 (0.88–0.96) and 0.70 (0.61–0.77), respectively. Positive tenderness rates and P-VAS scores were significantly higher in the LSS group (p

Tissue clearing methods promise to provide exquisite three-dimensional imaging information; however, there is a need for simplified methods for lower resource settings and for non-fluorescence based phenotyping to enable light microscopic imaging modalities. Here we describe the simplified CLARITY method (SCM) for tissue clearing that preserves epitopes of interest. We imaged the resulting tissues using light sheet microscopy to generate rapid 3D reconstructions of entire tissues and organs. In addition, to enable clearing and 3D tissue imaging with light microscopy methods, we developed a colorimetric, non-fluorescent method for specifically labeling cleared tissues based on horseradish peroxidase conversion of diaminobenzidine to a colored insoluble product. The methods we describe here are portable and can be accomplished at low cost and can allow light microscopic imaging of cleared tissues, thus enabling tissue clearing and imaging in a wide variety of settings.

Stem cell-derived cardiomyocytes provide a promising tool for human developmental biology, regenerative therapies, disease modeling, and drug discovery. As human pluripotent stem cell-derived cardiomyocytes remain functionally fetal-type, close monitoring of electrophysiological maturation is critical for their further application to biology and translation. However, to date, electrophysiological analyses of stem cell-derived cardiomyocytes has largely been limited by biologically undefined factors including 3D nature of embryoid body, sera from animals, and the feeder cells isolated from mouse. Large variability in the aforementioned systems leads to uncontrollable and irreproducible results, making conclusive studies difficult. In this report, a chemically-defined differentiation regimen and a monolayer cell culture technique was combined with multielectrode arrays for accurate, real-time, and flexible measurement of electrophysiological parameters in translation-ready human cardiomyocytes. Consistent with their natural counterpart, amplitude and dV/dtmax of field potential progressively increased during the course of maturation. Monolayer culture allowed for the identification of pacemaking cells using the multielectrode array platform and thereby the estimation of conduction velocity, which gradually increased during the differentiation of cardiomyocytes. Thus, the electrophysiological maturation of the human pluripotent stem cell-derived cardiomyocytes in our system recapitulates in vivo development. This system provides a versatile biological tool to analyze human heart development, disease mechanisms, and the efficacy/toxicity of chemicals.

A cadaveric study.To investigate the accuracy of pedicle screw placement using a robotic guidance system (RGS).RGS is a unique surgery assistance-apparatus. Although several clinical studies have demonstrated that RGS provides accurate pedicle screw placement, very few studies have validated its accuracy.A total of 216 trajectories performed with the assistance of the RGS in eight cadavers were evaluated. The RGS was used, with different mounting platforms, to drill pilot holes in the thoracic and lumbosacral spine, using 3-mm diameter fiducial wires as trajectory markers. Deviation between the preoperative plan and executed trajectories was measured at the entry points to the vertebrae and at a depth of 30 mm along the wire. Both the deviation from the preoperative plan and the wire position were evaluated in the axial and sagittal planes using computed tomography (CT).The average deviation from the planned wire placement was 0.64 ± 0.59 mm at the entry point and 0.63 ± 0.57 mm at a depth of 30 mm in the axial plane, and 0.77 ± 0.62 mm and 0.80 ± 0.66 mm, respectively, in the sagittal plane. The magnitude of deviation was not affected by the vertebral level or the platform used. The use of an open approach achieved greater screw placement accuracy at a depth of 30 mm in the sagittal plane, compared with the percutaneous approach. The fiducials were placed completely within the pedicle in 93.9% of trajectories in the axial plane (n = 164 pedicles with a width ≥5 mm) and 98.6% in the sagittal plane (n = 216).In this cadaveric study, RGS supported execution of accurate trajectories that were equal or slightly superior to reports of CT-based navigation systems.N/A.

Three mammalian isoforms of heterochromatin protein 1 (HP1), α, β, and γ, play diverse roles in gene regulation. Despite their structural similarity, the diverse functions of these isoforms imply that they are additionally regulated by post-translational modifications. Here, we have identified intermolecular disulfide bond formation of HP1 cysteines in an isoform-specific manner. Cysteine 133 in HP1α and cysteine 177 in HP1γ were involved in intermolecular homodimerization. Although both HP1α and HP1γ contain reactive cysteine residues, only HP1γ readily and reversibly formed disulfide homodimers under oxidative conditions. Oxidatively dimerized HP1γ strongly and transiently interacted with TIF1β, a universal transcriptional co-repressor. Under oxidative conditions, HP1γ dimerized and held TIF1β in a chromatin component and inhibited its repression ability. Our results highlight a novel, isoform-specific role for HP1 as a sensor of the cellular redox state.

Oxidized LDL (OxLDL) is implicated in endothelial dysfunction as well as the formation and progression of atherosclerosis. It has become evident that the atherogenic properties induced by OxLDL are mainly mediated via lectin-like OxLDL receptor-1 (LOX-1). Over the past decade, much research has been performed to investigate lipid metabolism and atherogenesis using genetically engineered mice. To understand the significance of OxLDL, methods to measure the levels of OxLDL in these experimental animals should be established. Utilizing a chicken monoclonal antibody technique, here, we generated anti-human ApoB antibodies that are able to recognize mouse VLDL/LDL. These antibodies were selected from single chain fragment of variable region (scFv) phage library constructed from chickens immunized with human LDL. One of these antibodies, HUC20, was reconstructed into IgY form. Immunohistochemical analysis revealed that this novel antibody specifically stains atherosclerotic lesions of ApoE-deficient mice, associated with Oil red O positive and macrophage-antigen-positive regions. Furthermore, in combination with recombinant LOX-1, a sandwich enzyme immunoassay was developed to measure the levels of LOX-1 ligands in mouse plasma. The sandwich enzyme immunoassay revealed a dramatic increase in the level of LOX-1 ligands in the plasma of ApoE-deficient mice fed high-fat diet, suggesting a link between the level of LOX-1-ligands and the progression of atherosclerosis in mice. Hence, the chicken anti-ApoB monoclonal antibody HUC20 developed here, could be a useful tool to analyze the role of ApoB-containing lipoprotein in atherogenesis in mice. © 2008 Elsevier Ireland Ltd. All rights reserved.

Chemotherapeutic agents to induce DNA damage have been limited to use due to severe side effects of cardiotoxicity. ATM (Ataxia-telangiectasia mutated) is an essential protein kinase in triggering DNA damage responses. However, it is unclear how the ATM-mediated DNA damage responses are involved in the cardiac cell damage. To elucidate these functions in heart, we searched for specific substrates of ATM from mouse heart homogenate. Combining an in vitro phosphorylation following anion-exchange chromatography with purification by reverse-phase high-performance liquid chromatography (HPLC), we successfully identified p32, an ASF/SF2-associated protein, as a novel substrate for ATM. An in vitro kinase assay using recombinant p32 revealed that ATM directly phosphorylated p32. Furthermore, we determined Ser 148 of p32 as an ATM phosphorylation site. Since p32 is known to regulate mRNA splicing and transcription, p32 phosphorylation by ATM might be a new transcriptional regulatory pathway for specific DNA damage responses in heart.

TNFalpha has multiple important cellular functions both in normal cells and in tumor cells. To explore the role of TNFalpha, we identified NUAK family, SNF1-like kinase 2 (NUAK2), as a TNFalpha-induced kinase by gene chip analysis. NUAK2 is known to be induced by various cellular stresses and involved in cell mortality, however, its substrate has never been identified. We developed original protocol of de novo screening for kinase substrates using an in vitro kinase assay and high performance liquid chromatography (HPLC). Using this procedure, we identified myosin phosphatase target subunit 1 (MYPT1) as a specific substrate for NUAK2. MYPT1 was phosphorylated at another site(s) by NUAK2, other than known Rho-kinase phosphorylation sites (Thr696 or Thr853) responsible for inhibition of myosin phosphatase activity. These data suggests different phosphorylation and regulation of MYPT1 activity by NUAK2.

Osteoporotic vertebral compression fractures occur frequently in the elderly, and in most cases, they can be managed well with conservative treatment [1,2]. However, in some cases, the fracture site fails to unite, resulting in vertebral collapse and instability. Surgical treatment is required when delayed neurological deficits occur, which are caused by spinal cord compression resulting from retropulsed bone fragments in the spinal canal, progression of kyphosis, and spinal instability at the fracture site [3,4]. However, there is no consensus on surgical indication when the patients complained of only pain in the back or lower extremities [5]. In the present study, we report our experience of surgery in a patient with osteoporotic vertebral collapse and instability. The patient presented with no motor deficits, and had a long history of conservative treatments. However, the surgery revealed L2 spinal nerve root transection caused by spinal instability. The present case shows the importance of surgical indication and timing in patients with pain as the only symptom of osteoporotic vertebral compression fracture. Informed consent for the publication of this case was obtained from the patient.

Changes in the toxicity in chlorinated water after chlorine addition were examined. For toxicity evaluation, the chromosomal aberration test and the transformation test were conducted as indexes of initiation activity and promotion activity, respectively, in the carcinogenesis process. Activity inducing chromosomal aberrations in chlorinated Lake Biwa water gradually decreased over time after chlorination. In contrast, activity inducing transformations determined by the two-stage assay gradually increased. Thus, toxicity that decreases or increases is present in chlorinated water. Furthermore, activity inducing transformations determined by the non-two-stage assay gradually decreased over time. This direction of change is opposite to that of activity inducing transformations determined by the two-stage assay and is consistent with that of activity inducing chromosomal aberrations. The drastic decrease in initiation activity detected as activity inducing chromosomal aberrations could be the main cause for the decrease in activity inducing transformations determined by the non-two-stage assay (an index of the sum of initiation and promotion activity). MX change was quantitatively consistent with those of activity inducing chromosomal aberrations and transformations determined by the non-two-stage assay. On the other hand, directions of changes in concentrations of typical by-products such as chloroform were consistent only with that of activity inducing transformations determined by the two-stage assay. Findings of this study suggest that MX is appropriate as an index for comparing the carcinogenicity of tap water near and far from a water purification plant.