A perfusable microfluidic device with on-chip total internal reflection fluorescence microscopy (TIRFM) for in situ and real-time monitoring of live cells

A microfluidic device integrated with a Total Internal Reflection (TIR)-based chip for cell observation and analysis was developed. This integrated device enables in situ Total Internal Reflection Fluorescence Microscopy (TIRFM) on adherent cells cultured under continuous medium perfusion. This TIR-based chip, allows TIRFM to be easily performed on cells without the assembly of complicated optical components and cell culture chambers. The integrated device was evaluated by tracking the movement of fluorescent beads and monitoring the location of insulin granules in mouse pancreatic β-cells. This system offers higher signal-to-noise (S/N) ratio than epi-fluorescence microscopy (EPIFM), and comparable image quality to commercial TIRFM systems when imaging insulin granules. We also detected repetitive changes in intracellular Ca(2+) concentration in MIN6-m9 cells stimulated with KCl, which demonstrates quick perfusion for cell analysis while maintaining high S/N ratio.     

Stem cell-mediated transfer of a human artificial chromosome ameliorates muscular dystrophy

In contrast to conventional gene therapy vectors, human artificial chromosomes (HACs) are episomal vectors that can carry large regions of the genome containing regulatory elements. So far, HACs have not been used as vectors in gene therapy for treating genetic disorders. Here, we report the amelioration of the dystrophic phenotype in the mdx mouse model of Duchenne muscular dystrophy (DMD) using a combination of HAC-mediated gene replacement and transplantation with blood vessel-associated stem cells (mesoangioblasts). We first genetically corrected mesoangioblasts from dystrophic mdx mice with a HAC vector containing the entire (2.4 Mb) human dystrophin genetic locus. Genetically corrected mesoangioblasts engrafted robustly and gave rise to many dystrophin-positive muscle fibers and muscle satellite cells in dystrophic mice, leading to morphological and functional amelioration of the phenotype that lasted for up to 8 months after transplantation. Thus, HAC-mediated gene transfer shows efficacy in a preclinical model of DMD and offers potential for future clinical translation.     

Clinical study of blood purification therapy in critical care in Japan: results from the survey research of the Japan Society for Blood Purification in Critical Care in 2013

To clarify the clinical status of blood purification therapy (BPT) in critical care in Japan, we conducted a cohort study using data from a nationwide registry of the Japan Society for Blood Purification in Critical Care in 2013. We enrolled 2227 patients treated with BPT (female, 39.1%; mean age, 65.5 ± 12.1 years) in the intensive care units of 43 facilities. Patient characteristics, modes of BPT, and survival rate for each disease were investigated. In total, BPT was performed 3053 times. Continuous renal replacement therapy (CRRT) (57.9%) was the most common mode of BPT, followed by intermittent renal replacement therapy (20.2%) and direct hemoperfusion with the polymyxin B-immobilized fiber column (PMX-DHP) (11.5%). Nafamostat mesilate (84.9%) was most frequently used as the anticoagulant. The 28-day survival rate was 56.8% in all patients. The most common mode for acute kidney injury (AKI) and multiple organ failure was CRRT, while PMX-DHP and CRRT were most common for sepsis. There was no significant difference in survival rates among AKI stages 1–3. Survival rate (38.3%) was significantly lower in patients with acute lung injury (ALI) than in those with multiple organ failure (41.8%) and those with sepsis (46.6%). Multivariate regression analysis revealed that the APACHE II score and the presence of acute ALI and acute hepatic failure were significantly associated with death. This large-scale cohort study showed the clinical status of BPT in Japan. Further investigations are required to clarify the efficacy of BPT for critically ill patients.     

A practical prediction model for early hematoma expansion in spontaneous deep ganglionic intracerebral hemorrhage

Objective

Early hematoma expansion is a known cause of morbidity and mortality in patients with intracerebral hemorrhage (ICH). The goal of this study was to identify clinical predictors of ICH growth in the acute stage.

Materials and methods

We studied 201 patients with acute (<6 h) deep ganglionic ICH. Patients underwent CT scan at baseline and hematoma expansion (>33% or >12.5 ml increase) was determined on the second scan performed within 24 h. Fourteen clinical and neuroimaging variables (age, gender, GCS at admission, hypertension, diabetes mellitus, kidney disease, stroke, hemorrhagic, antiplatelet use, anticoagulant use, hematoma density heterogeneity, hematoma shape irregularity, hematoma volume and presence of IVH) were registered. Additionally, blood pressure was registered at initial systolic BP (i-SBP) and systolic BP 1.5 h after admission (1.5 h-SBP). The discriminant value of the hematoma volume and 1.5 h-SBP for hematoma expansion were determined by the receiver operating characteristic (ROC) curves. Factors associated with hematoma expansion were analyzed with multiple logistic regression.

Results

Early hematoma expansion occurred in 15 patients (7.0%). The cut-off value of hematoma volume and 1.5 h-SBP were determined to be 16 ml and 160 mmHg, respectively. Hematoma volume above 16 ml (HV > 16) ([OR] = 5.05, 95% CI 1.32–21.36, p = 0.018), hematoma heterogeneity (HH) ([OR] = 7.81, 95% CI 1.91–40.23, p = 0.004) and 1.5 h-SBP above 160 mmHg (1.5 h-SBP > 160) ([OR] = 8.77, 95% CI 2.33–44.56, p = 0.001) independently predicted ICH expansion. If those three factors were present, the probability was estimated to be 59%.

Conclusions

The presented model (HV > 16, HH, 1.5 h-SBP > 160) can be a practical tool for prediction of ICH growth in the acute stage. Further prospective studies are warranted to validate the ability of this model to predict clinical outcome.     

Preoperative scoring system for predicting early adjacent vertebral fractures after Balloon Kyphoplasty

BackgroundAdjacent vertebral fracture (AVF) is a major complication following Balloon Kyphoplasty (BKP). There is no scoring system for predicting AVF using only preoperative elements. The purposes of this study were to develop a scoring system for predicting early AVF after BKP based on preoperative factors and to investigate the appropriate surgical indication for BKP.MethodsOf 220 patients who underwent BKP at a single institution since 2011, 65 patients over the age of 60 who had undergone a standing whole spine X-ray preoperatively were enrolled. Factors affecting the occurrence of early AVF were examined. A scoring system was created consisting of the factors exhibiting significant differences, and the correlation between the total score and the incidence of early AVF was investigated.ResultsTwenty of the 65 patients (30.8%) had early AVF. In a univariate analysis, age, previous vertebral fracture, pelvic tilt, and Local kyphosis significantly influenced early AVF. In a multivariate logistic regression analysis, age had an odds ratio of 1.136 (95% CI 1.001–1.289), previous vertebral fractures 4.181 (1.01–17.309), and Local kyphosis 1.103 (1.021–1.191). The scoring system was set as follows: ①Age (<75 years: 0 points(P), 75years≦: 1P), ②The number of previous vertebral fractures (0: 0 P, 1: 1P, 2: 2P, 3 or more: 3P), and ③Local kyphosis (<10°: 0P, 10°≦: 1P). There was a correlation between the total score and the incidence of early AVF (r = 0.812, 1P = 0.05). The incidence of early AVF was 6.4% (2 cases/31 cases) for a score of ≦1P and 54.5% (18 cases/33 cases) for a score of ≧2P.ConclusionsThere was a correlation between the total score and the incidence of early AVF. A score of 1 point or less may represent the appropriate surgical indication for BKP.     

Hip range‐of‐motion (ROM) is less than normal after rotational acetabular osteotomy for developmental dysplasia of the hip: A simulated ROM analysis

The optimal reorientation of the acetabulum for developmental dysplasia of the hip (DDH) is unknown in terms of hip range‐of‐motion (ROM). The simulated ROMs of 52 DDHs after rotational acetabular osteotomy (RAO) with several patterns of femoral head coverage and those of 73 normal hips were analyzed using computer models reconstructed from CT images. After RAO with a lateral center edge angle (LCEA) of 30° and an anterior center edge angle (ACEA) of 55° producing coverage similar to that of normal hips, the maximal flexion and maximal internal rotation at 110° flexion with 20° adduction were significantly smaller than those of the normal group. To achieve ROMs after RAO similar to those of the normal group, an LCEA of 30° with an ACEA of 45°, an LCEA of 25° with an ACEA of 45° to 50°, and an LCEA of 20° with an ACEA of 50° could be preferred angles to target, even though they provided smaller coverage than that of normal hips. After RAO producing femoral head coverage similar to that of normal hips, the maximal flexion and the maximal internal rotation at 110° flexion with 20° adduction were significantly smaller than those of the normal group. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:217–223, 2016.     

Patient-specific radiotherapy quality assurance for estimating actual treatment dose

This study aimed to evaluate the optimal method for planning computed tomography (CT) for prostate cancer radiotherapy to avoid a dose difference of ≥3% between the actual and planned treatments using multiple acquisition planning CT (MPCT). We calculated the 3-dimensional (3D) displacement error between the pelvic bone and matching fiducial marker on MPCT and cone-beam CT scans of 25 patients who underwent prostate volumetric-modulated arc therapy for prostate cancer. The correlation of the 3D displacement error and the dose difference between planned and actual treatments was calculated using least squares second-order polynomial model. The 3D displacement error showed a moderate correlation with differences between planned and accumulated treatment doses (r = 0.587, p < 0.0001). Moreover, the improvement rate of the minimum 3D displacement error showed a strong correlation with the relative error between each MPCT image (r = 0.793, p < 0.0001). Significant differences were observed between planned and actual treatment doses (p < 0.0001) in the relative 3D displacement errors of <1 mm, 1 to 3 mm, and >3 mm. The 3D displacement error on MPCT (as the selection estimation index for optimal planning CT) is useful for monitoring patient-specific intensity-modulated radiation therapy quality assurance. This new method allows to estimate dose differences from the planned dose before commencing treatment, thereby ensuring high-quality therapy. As radiotherapy quality is critical for patient outcome, these findings may contribute to better management of prostate cancer.     

Psychopathology and working memory-induced activation of the prefrontal cortex in schizophrenia-like psychosis of epilepsy: Evidence from magnetoencephalography

Aim: The aim of this study was to investigate whether magnetoencephalographic oscillations underlying working memory dysfunction in the dorsolateral prefrontal cortex (DLPFC) are related to psychopathological disturbance in patients with schizophrenia‐like psychosis of epilepsy (SLPE). Methods: Twelve patients with SLPE and 14 non‐psychotic epilepsy controls participated in this study. Magnetoencephalography was recorded while patients performed a visual working memory (WM) task. Psychopathology was assessed using a four‐factor structure of the Brief Psychiatric Rating Scale, and regression analyses were carried out to examine the relative impact of severity of psychopathology on WM‐induced activation of the DLPFC. Results: We found that activation of the WM‐compromising DLPFC, as indicated by increased alpha desynchronization in patients with SLPE compared with their non‐psychotic counterparts, showed a positive linear correlation with disorganization symptom scores. This association remained significant after controlling for confounding factors, including age, task performance, IQ, and duration of psychosis. Conclusion: Our results indicate that abnormal activation in prefrontal areas engaged during working memory may be critical to domains of psychopathology, in particular disorganized thought‐processing in patients with SLPE.