Effects of Cardiac Glycoside Digoxin on Dendritic Spines and Motor Learning Performance in Mice.

Synapse formation following the generation of postsynaptic dendritic spines is essential for motor learning and functional recovery after brain injury. The C-terminal fragment of agrin cleaved by neurotrypsin induces dendritic spine formation in the adult hippocampus. Since the α3 subunit of sodium-potassium ATPase (Na/K ATPase) is a neuronal receptor for agrin in the central nervous system, cardiac glycosides might facilitate dendritic spine formation and subsequent improvements in learning. This study investigated the effects of cardiac glycoside digoxin on dendritic spine turnover and learning performance in mice. Golgi-Cox staining revealed that intraperitoneal injection of digoxin less than its IC50 in the brain significantly increased the density of long spines (≥2 µm) in the cerebral cortex in wild-type mice and neurotrypsin-knockout (NT-KO) mice showing impairment of activity-dependent spine formation. Although the motor learning performance of NT-KO mice was significantly lower than control wild-type mice under the control condition, low doses of digoxin enhanced performance to a similar degree in both strains. In NT-KO mice, lower digoxin doses equivalent to clinical doses also significantly improved motor learning performance. These data suggest that lower doses of digoxin could modify dendritic spine formation or recycling and facilitate motor learning in compensation for the disruption of neurotrypsin-agrin pathway.

Effects of arm and leg positions on lumbar multifidus muscle activity while on hands and knees or while standing.

BACKGROUND: The lumbar multifidus (LMF) muscle, which is involved in the mechanical stability of the lumbar spine, reportedly undergoes atrophy in patients with low back pain. Preventing or mitigating low back pain requires strengthening the LMF muscle; however, methods for triggering selective and significant contraction of this muscle have not been fully studied. This study aims to clarify how, in the hands-and-knees or standing position, the position of the arm and leg on one side affects the activity of the lumbar erector spinae (LES) and LMF muscles. METHODS: We recruited nine adult men with no prior history of low back pain. Measurements were taken in four different postures under varying conditions (that is, one arm and one leg were lifted in either the hands-and-knees or standing position,) as follows: (1) shoulder joint flexion and hip joint extension in the hands-and-knees position; (2) 90° shoulder joint abduction and hip joint abduction in the hands-and-knees position; (3) shoulder joint flexion and hip joint extension in the standing position; and (4) 90° shoulder joint abduction and hip joint abduction in the standing position. The 90° shoulder joint abduction involved simultaneous horizontal abduction, while the hip joint abduction involved simultaneous extension. Muscle activity of the LES and LMF in each posture was measured using a surface electromyograph. RESULTS: Muscle activity of the LMF was significantly higher in 90° shoulder joint abduction and hip joint abduction than in shoulder joint flexion and hip joint extension in both the hands-and-knees and standing positions. The LES muscle showed no significant differences in activity between each posture. CONCLUSIONS: The results suggest that unilateral 90° shoulder joint abduction and contralateral hip joint abduction in the hands-and-knees and standing positions may produce selective and significant contraction of the LMF muscle.

The nitric oxide-cyclic guanosine monophosphate pathway inhibits the bladder ATP release in response to a physiological or pathological stimulus.

The release of ATP from the epithelium of the urinary bladder (urothelium) in response to mechanical/chemical stimuli contributes to the visceral sensation in the micturition reflex. The nitric oxide (NO)-mediated induction of cyclic guanosine monophosphate (cGMP) has been detected in urothelial cells and may inhibit the micturition reflex. However, the function of the NO-cGMP pathway in the regulation of urothelial ATP release remains poorly understood in contrast to its effects on smooth muscles or primary afferent nerves. Therefore, we investigated the relevance of the NO-cGMP pathway to ATP release on the mucosal side in the present study. The administration of l-arginine (NO precursor) or NOC 12 (NO donor) significantly reduced ATP release to the mucosal side at a physiologically normal urine storage pressure (5 cmH2 O). L-NAME (NO synthase inhibitor) significantly increased the distention-induced release of ATP. The phosphodiesterase-5 inhibitor, sildenafil, which increases cGMP levels, inhibited distention-induced ATP release. Furthermore, sildenafil significantly reduced ATP release in response to the administration of lipopolysaccharide. These results suggest that the NO-cGMP pathway inhibited urothelial ATP release during the storage phase under both physiological and pathological conditions.

Serum tenascin-C predicts resistance to steroid combination therapy in high-risk Kawasaki disease: a multicenter prospective cohort study.

BACKGROUND: Tenascin-C (TN-C) is an extracellular matrix glycoprotein related to tissue inflammation. Our previous retrospective study conducted in 2016 revealed that the serum tenascin-C level was higher in patients with Kawasaki disease (KD) who were resistant to intravenous immunoglobulin (IVIG) and developed coronary artery lesions (CALs). The present study is a prospective cohort study to assess if the serum level of tenascin-C could be used as a novel biomarker to predict the risk of resistance to initial treatment for high-risk patients. METHODS: A total of 380 KD patients were registered and provided serum samples for tenascin-C measurement before commencing their initial treatment. Patients who did not meet the inclusion criteria were excluded from analysis; of the 181 remaining subjects, there were 144 low-risk patients (Kobayashi score: ≤4 points) and 37 high-risk patients (Kobayashi score: ≥5 points). The initial treatments for low-risk patients and high-risk patients were conventional therapy (IVIG with aspirin) and prednisolone combination therapy, respectively. The patient clinical and laboratory data, including the serum tenascin-C level, were compared between initial treatment responders and non-responders. RESULTS: In the low-risk patients, there was no significant difference in the median levels of serum tenascin-C between the initial therapy responders and non-responders. However, in the high-risk patients, the median serum tenascin-C level in initial therapy non-responders was significantly higher than that in initial therapy responders (175.8 ng/ml vs 117.6 ng/ml). CONCLUSIONS: Serum tenascin-C could be a biomarker for predicting the risk of high-risk patients being non-responsive to steroid combination therapy. TRIAL REGISTRATION: This study was a prospective cohort study. It was approved by the ethics committee of each institute and performed in accordance with the Declaration of Helsinki.

Proposal of a New Index Based on Signal-to-Noise Ratio for Low-contrast Detectability in Computed Tomographic Imaging.

The low-contrast detectability of computed tomography (CT) images is commonly evaluated by the contrast-to-noise ratio (CNR) because of its convenience to measure. However, the correlation between CNR and visual detectability is poor because the CNR is a simple index determined by both the contrast of the object and the standard deviation of the image noise. On the other hand, the signal-to-noise ratio (SNR), especially SNR based on the statistical decision theory model (SNRS, D) and SNR based on the matched-filter model (SNRM) are considered superior to CNR. In this study, we investigated a new physical image quality index for evaluating low-contrast detectability (SNRA), which is approximately derived from SNRS, D and SNRM. The new index, which was calculated using the object size, contrast of the object and the noise power spectrum, provided good approximations when the diameter of the rod object was equal and >5 mm. The diameter dependency of the SNRA was also found to provide better sensitivity than the sensitivities of CNR and object-specific CNR, similar to SNRS, D and SNRM. The results suggested that the proposed convenient index should be useful for evaluating the low-contrast detectability of CT images.

The Bone Regeneration Using Bone Marrow Stromal Cells with Moderate Concentration Platelet-Rich Plasma in Femoral Segmental Defect of Rats.

BACKGROUND: Platelet-rich plasma (PRP) can provide an assortment of growth factors, but how PRP effects bone regeneration is still unknown. The aim of the study was to explore an optimal method of using PRP and bone marrow stromal cells (BMSCs). METHODS: An in vitro experiment was first conducted to determine an appropriate quantity of PRP. BMSCs were cultured with PRP of different concentrations to assess cell proliferation and osteogenic differentiation. Following the in vitro study, a rat femoral segmental defect model was used. Five collagen mixtures consisting of different concentrations of PRP and BMSCs were prepared as follows, i) BMSCs and PRP (platelet 20 x 104/µl), ii) BMSCs and PRP (platelet 100 x 104/µl), iii) BMSCs and PRP (platelet 500 x 104/µl), iv) BMSCs, and v) PRP group (platelet 100 x 104/µl), were used to fill defect. New bone formation was evaluated by soft X-ray and histologic analyses were performed at 2, 4, 6 and 8 weeks postoperatively. RESULTS: The cell proliferation increased PRP concentration-dependently. Cellular alkaline phosphatase activity was higher in moderate concentration than high or low concentration group's in vitro study. In vivo study, the bone fill percentage of newly formed bone in BMSCs and PRP (platelet 100 x 104/µl) was 46.9% at 8 weeks and increased significantly compared with other groups. CONCLUSION: BMSCs with moderate level of PRP significantly enhanced bone formation in comparison with BMSCs or PRP transplant in a rat femoral defect model.

JNK is critical for the development of Candida albicans-induced vascular lesions in a mouse model of Kawasaki disease.

BACKGROUND: Kawasaki disease (KD) is the most common systemic vasculitis of unknown etiology in children, and can cause the life-threatening complication of coronary artery aneurysm. Although a novel treatment strategy for patients with KD-caused vascular lesions is eagerly awaited, their molecular pathogenesis remains largely unknown. c-Jun N-terminal kinase (JNK) is a signaling molecule known to have roles in inflammation and tissue remodeling. The aim of this study was to elucidate significant involvement of JNK in the development of vascular lesions in a mouse model of KD. METHODS AND RESULTS: We injected Candida albicans cell wall extract (CAWE) into 4-week-old C57BL/6 mice. Macroscopically, we found that CAWE caused the development of bulging lesions at coronary artery, carotid artery, celiac artery, iliac artery and abdominal aorta. Histological examination of coronary artery and abdominal aorta in CAWE-treated mice showed marked inflammatory cell infiltration, destruction of elastic lamellae, loss of medial smooth muscle cells and intimal thickening, which are similar to histological features of vascular lesions of patients with KD. To find the role of JNK in lesion formation, we evaluated the effects of JNK inhibitor, SP600125, on abdominal aortic lesions induced by CAWE. Interestingly, treatment with SP600125 significantly decreased the incidence of lesions and also protected against vascular inflammation and tissue destruction histologically, compared with the placebo treatment. CONCLUSIONS: Our findings suggest that JNK is crucial for the development of CAWE-induced vascular lesions in mice, and potentially represents a novel therapeutic target for KD.

Effect of platelet-rich plasma with fibrin matrix on healing of intrasynovial flexor tendons.

PURPOSE: To investigate the effects of platelet-rich plasma (PRP) with fibrin matrix on the healing of intrasynovial flexor tendons in a rabbit model in vivo. METHODS: We transected and repaired 156 toe flexors of 73 rabbits using the technique of Tsuge et al and a simple running epitendinous suture. We randomly assigned Repaired tendons to groups that recieved no additional treatment (control) or to which we applied PRP, fibrin (F), or PRP with fibrin matrix (PRP-F) at the repair site. We scored edema and adhesion at 2, 3, and 6 weeks after surgery, and linearly tested repaired tendons for load to failure. We also histologically evaluated tendons at 2 and 3 weeks. RESULTS: Edema scores and adhesion scores did not significantly differ among the 4 groups at any time point. Mean load to failure in the PRP-F group (14.7 N) was the highest among the 4 groups at 2 weeks after surgery, and was significantly higher than in the control group (10.0 N). Median histological scores in the PRP-F group (3.3 points) were significantly higher than in the control group (1.0 point). Mean load to failure in the PRP-F group (16.1 N) was highest, and median histological scores in the PRP-F group (3.5 points) were higher than in the control group (2.4 points) at 3 weeks, although there were no significant differences at 3 or 6 weeks. CONCLUSIONS: In a rabbit model of cut flexor tendons, PRP with fibrin matrix significantly increased healing strength within 2 weeks after surgery. Side effects such as increases in toe edema or adhesions around the tendons did not arise. CLINICAL RELEVANCE: Platelet-rich plasma with fibrin matrix might help reduce the risk of repeated rupture after flexor tendon surgery, and lead to early rehabilitation.

Feasibility study of two schedules of sunitinib in combination with pemetrexed in patients with advanced solid tumors.

BACKGROUND: Sunitinib is an oral multitargeted tyrosine kinase inhibitor of vascular endothelial growth factor and platelet-derived growth factor receptors, as well as of other receptor types. We have performed a feasibility study to investigate the safety of sunitinib in combination with pemetrexed for treatment of advanced refractory solid tumors. METHODS: Sunitinib was administered once daily on a continuous daily dosing (CDD) schedule (37.5 mg/day) or a 2-weeks-on, 1-week-off treatment schedule (50 mg/day, Schedule 2/1) in combination with pemetrexed at 500 mg/m(2) on day 1 of repeated 21-day cycles. RESULTS: Twelve patients were enrolled in the study: six on the CDD schedule and six on Schedule 2/1. None of the treated patients experienced a dose-limiting toxicity. Toxicities were manageable and similar in type to those observed in monotherapy studies of sunitinib and pemetrexed. Pharmacokinetic analysis did not reveal any substantial drug-drug interaction. One patient with squamous cell lung cancer showed a partial response and five patients had stable disease. CONCLUSIONS: Combination therapy with sunitinib administered on Schedule 2/1 (50 mg/day) or a CDD schedule (37.5 mg/day) together with standard-dose pemetrexed (500 mg/m(2)) was well tolerated in previously treated patients with advanced solid tumors.

Effect of axitinib (AG-013736) on fatigue, thyroid-stimulating hormone, and biomarkers: a phase I study in Japanese patients.

Axitinib is an oral, potent, and selective inhibitor of vascular endothelial growth factor receptor (VEGFR) 1, 2, and 3. This phase I study evaluated the safety, pharmacokinetics, pharmacodynamics, antitumor activity, and recommended starting dose of axitinib in patients with advanced solid tumors. Twelve patients received single-dose axitinib 5 mg and were monitored for > or =48 h. Continuous 5 mg twice-daily dosing was then initiated. One patient had dose-limiting toxicity (grade 3 proteinuria and fatigue). Common treatment-related adverse events were anorexia, fatigue, and diarrhea. Grade 3 treatment-related adverse events were fatigue and hypertension. Maximum axitinib plasma concentration occurred 1-4 h after steady-state dosing. Eleven patients experienced thyroid-stimulating hormone elevation; time-course change and fatigue onset appeared to be related in some patients. Significant correlation was observed between thyroid-stimulating hormone change and area under the plasma concentration-time curve (AUC; r = 0.80, P = 0.005). Axitinib decreased plasma soluble vascular endothelial growth factor receptor 2 (s-VEGFR2), with significant correlation between change in s-VEGFR2 and AUC (r = -0.92, P < 0.0001). Fluorodeoxyglucose positron emission tomography revealed a substantial decrease in tumor metabolic activity associated with axitinib. Tumor size decreased in nine patients. The time-course of thyroid-stimulating hormone change appeared correlated with fatigue. There were significant correlations between thyroid-stimulating hormone or s-VEGFR2 and axitinib exposure. Axitinib 5 mg twice-daily is the recommended starting dose for Japanese patients. This trial is registered with ClinicalTrials.gov, identifier NCT00447005.

Phase I study of TLR9 agonist PF-3512676 in combination with carboplatin and paclitaxel in patients with advanced non-small-cell lung cancer.

This phase I, open-label study investigated the Toll-like receptor 9 agonist, PF-3512676, in combination with carboplatin and paclitaxel in Japanese patients with advanced, non-small-cell lung cancer (NSCLC). Patients (n = 12) with treatment-naive stage IIIB or IV NSCLC received single-agent PF-3512676 subcutaneously once during the first 7 days (monotherapy phase) in three escalating dose levels (0.1, 0.2, and 0.4 mg/kg) followed by a combination phase during which patients received 0.1 or 0.2 mg/kg PF-3512676 subcutaneously on days 8 and 15 of each 3-week cycle of carboplatin (area under the curve, 6 mg x min/mL) and paclitaxel (200 mg/m(2)). Safety and pharmacokinetics of PF-3512676 were assessed during monotherapy and combination therapy phases. PF-3512676 was tolerable as monotherapy or in combination with chemotherapy in patients with NSCLC. Most common treatment-related, non-hematologic adverse events (AEs) throughout the study were injection-site reactions (n = 12, 100%) and flu-like symptoms (n = 11, 91.7%) that were each grade 1 or 2 in all but one patient. All patients experienced neutropenia and leukopenia (>or=grade 3 in 11 [91.7%] and seven [58.3%] patients, respectively). One patient in dose level 2 had a dose-limiting toxicity: grade 3 rash and grade 3 increase in gamma-glutamyltransferase during combination therapy. Mean PF-3512676 half-life ranged from 4.8 to 21.6 h (longer with higher doses). Four (33%) patients had objective responses (one complete response, three partial responses), and seven (58%) patients achieved stable disease. PF-3512676 as monotherapy and in combination with chemotherapy had an acceptable safety profile in Japanese patients with treatment-naive NSCLC.

Absence of biglycan accelerates the degenerative process in mouse intervertebral disc.

STUDY DESIGN: A study of the histologic changes of the intervertebral discs (IVDs) in biglycan (Bgn)-deficient mice. OBJECTIVE: In this study, we investigate whether the absence of Bgn accelerates the degenerative process in mouse intervertebral disc (IVD). SUMMARY OF BACKGROUND DATA: Proteoglycans and collagen fibrils are major components in the extracellular matrix (ECM) composition of IVD. The ECM of IVD contains several members of the small leucine repeat proteoglycans (SLRPs) family. Bgn is one member of SLRPs family, and showed a unique expression with age and degeneration in the human IVD. To date, there have been no in vivo studies to see whether SLRPs have a role in maintaining the structural integrity of IVD. To explore the functions of Bgn in the IVD, we examined discs in Bgn-deficient mice. METHODS: A total of 30 spine specimens were harvested from wild-type (WT) and Bgn-deficient mice. Five specimens for each genotype at 4-, 6-, and 9-month old were examined in the experiments. Histologic analysis of the IVD was performed. Histologic gradings were performed separately on nucleus pulposus, anulus fibrosus, and endplate according to the classification system proposed by Boos et al. RESULTS: We found that Bgn-deficient mice developed an early onset of disc degeneration compared with WT mice. The degenerative scores of Bgn-deficient mice were significantly higher than those of WT mice at 4- and 9-month-old. High scores for nucleus pulposus and anulus fibrosus in Bgn-deficient mice significantly affected the difference in total degenerative scores at 9 months of age. CONCLUSION: Bgn deficiency significantly accelerated disc degeneration.

Epimerization of diastereomeric alpha-amino nitriles to single stereoisomers in the solid state.

[reaction: see text] A diastereomeric mixture of the alpha-amino nitrile prepared by the Strecker reaction of benzaldehyde, (1S,2R)-1-aminoindan-2-ol, and cyanotrimethylsilane thermally epimerizes in the solid state to give a single diastereomer with an (S)-configuration at the alpha position to the nitrile moiety. This shows a sharp contrast to the reaction conducted in DMSO at room temperature, which gives a 1:1 mixture of (S)- and (R)-isomers. Several other alpha-amino nitriles also epimerize in the solid-state toward single diastereomers.

Engineered allogeneic mesenchymal stem cells repair femoral segmental defect in rats.

Bone marrow derived mesenchymal stem cells (MSC) have been shown to be progenitor cells for mesenchymal tissues. These cells may also provide a potential therapy for bone repair. Our previous studies showed that MSC engineered with the gene for bone morphogenetic protein 2 (BMP-2), a growth factor for bone cells, were capable of differentiating into osteoblast lineage and inducing autologous bone formation in several animal models. Culturing individual MSC for autologous implantation, however, remains problematic. The number of human MSC with osteogenic potential decreases with age, and, in certain diseases, the patient's marrow may be damaged or the healthy cells reduced in number. In this study, we used rats with a femoral segmental defect to investigate whether allogeneic BMP-2 engineered MSC would facilitate bone healing. We show that BMP-2 engineered allogeneic MSC can repair critical bone defects to the same degree as rats treated with BMP-2 engineered autologous MSC, if the allogeneic group receives short-term treatment with immunosuppressant FK506. We also show that allogeneic gene transferred MSC are directly involved in bone repair, in addition to acting as gene deliverers.