CCR1 acts downstream of NFAT2 in osteoclastogenesis and enhances cell migration.

We found that a chemokine receptor gene, CCR1, acts downstream of NFAT2 in RANKL-stimulated RAW264 and bone marrow cells. The upstream regulatory region of CCR1 showed RANKL-dependent and CsA-suppressible promoter activity. Downregulation of the expression and function of CCR1 suppressed cell migration. INTRODUCTION: We previously reported that the expression of NFAT2 induced by RANKL is a key process for progression to multinucleated cells in an in vitro osteoclastogenesis system. Identifying the target genes of NFAT2 would thus be informative about the differentiation process. We focused here on chemokine and chemokine receptor genes that act downstream of NFAT2 in RAW264 cells as well as osteoclast precursors prepared from bone marrow cells. MATERIALS AND METHODS: RAW264 mouse monocyte/macrophage line cells were cultured with or without cyclosporin A (CsA) in the presence of RANKL or glutathione S-transferase (GST). Osteoclast precursors were prepared from bone marrow cells. RANKL-inducible and CsA-suppressible genes were searched for by microarray analysis, and expression was confirmed by quantitative RT-PCR. Promoter activity was measured by luciferase gene reporter assay. Short interfering (si)RNA for CCR1 was introduced in RAW264 cells. Cell migration activity was examined using a Boyden chamber assay. RESULTS AND CONCLUSIONS: We identified the chemokine receptor gene CCR1 as a gene showing significant differential expression profiles in osteoclastogenesis in the presence versus the absence of CsA, an inhibitor of NFAT. This property was unique to CCR1 among the chemokine and chemokine receptor genes examined in both RAW264 and bone marrow cells. The upstream regulatory region was isolated from CCR1, and its RANKL-dependent and CsA-suppressible promoter activity was confirmed. The functional significance of CCR1 was assessed by monitoring the migration of cells in a transwell migration assay, and this activity was abolished when either CsA- or CCR1 siRNA-treated cells were used. Moreover, treatment with a Galpha inhibitor pertussis toxin (PTX) or methiolynated-regulated on activation, normal T cells expressed and secreted (Met-RANTES), an antagonist of CCR1, suppressed multinucleated cell formation in the bone marrow cell system. Together, these results suggest that the CCR1 signaling cascade is under the control of NFAT2 and seems to enhance the migration of differentiating osteoclasts.     

Imaging of a large collection of human embryo using a super-parallel MR microscope.

Using 4 and 8-channel super-parallel magnetic resonance (MR) microscopes with a horizontal bore 2.34T superconducting magnet developed for 3-dimensional MR microscopy of the large Kyoto Collection of Human Embryos, we acquired T(1)-weighted 3D images of 1204 embryos at a spatial resolution of (40 microm)(3) to (150 microm)(3) in about 2 years. Similarity of image contrast between the T(1)-weighted images and stained anatomical sections indicated that T(1)-weighted 3D images could be used for an anatomical 3D image database for human embryology.     

Pharmacologic preconditioning effects: Prostaglandin E1 induces heat-shock proteins immediately after ischemia/reperfusion of the mouse liver

Prostaglandin E1 (PGE1) has several potential therapeutic effects, including cytoprotection, vasodilation, and inhibition of platelet aggregation. This study investigates the protective action of PGE1 against hepatic ischemia/reperfusion injury in vivo using a complementary DNA microarray. PGE1 or saline was continuously administered intravenously to mice in which the left lobe of the liver was made ischemic for 30 minutes and then reperfused. Livers were harvested 0, 10, and 30 minutes postreperfusion. Messenger RNA was extracted, and the samples were labeled with two different fluorescent dyes and hybridized to the RIKEN set of 18,816 full-length enriched mouse complementary DNA microarrays. Serum alanine aminotransferase and aspartate aminotransferase levels at 180 minutes postreperfusion were significantly lower in the PGE1-treated group than in the saline-treated group. The cDNA microarray analysis revealed that the genes encoding heat-shock protein (HSP) 70, glucose-regulated protein 78, HSP86, and glutathione S-transferase were upregulated at the end of the ischemic period (0 minutes postreperfusion) in the PGE1 group. Our results suggested that PGE1 induces HSPs immediately after ischemia reperfusion. HSPs might therefore play an important role in the protective effects of PGE1 against ischemia/reperfusion injury of the liver.     

Subtle computed tomography abnormalities in cerebral deep sinus thrombosis.

A patient with cerebral deep sinus thrombosis, which was not diagnosed on the first examination, is reported. A 46-year-old woman presented with headache and vomiting. Neurological examination and a brain computed tomography (CT) scan showed no obvious abnormal findings. The patient suffered disturbed consciousness on the day after the examination, and was admitted to our emergency centre. A CT scan and magnetic resonance imaging revealed an ischaemic lesion in the left basal ganglia, suggesting deep sinus occlusion. Anticoagulant therapy was administered. One day after admission, a CT scan showed a haematoma and severe brain swelling in the same region. Cerebral angiography demonstrated a straight sinus occlusion. Intracranial pressure was not controlled with hypothermia, and the patient died 25 days after admission. Review of the initial CT scan revealed subtle, early findings of deep venous thrombosis that were missed on first examination.     

Congenital esophageal atresia successfully treated by early ligation of a tracheoesophageal fistula and delayed repair of the esophagus in a premature infant: Report of a case

We report herein the case of a premature infant with esophageal atresia (EA) and a tracheoesophageal fistula (TEF) associated with cardiac anomalies who was successfully treated by an early ligation of the TEF following gastrostomy, and delayed repair of the esophagus. A 1212-g male was born prematurely at 31 gestational weeks, at which time he was diagnosed as having EA with TEF and patent ductus arteriosus (PDA), ventricular septal defect (VSD), and atrial septal defect (ASD). A gastrostomy was initially performed but following extubation he gradually became tachypneic. A chest roentogenogram revealed atelectasis and ground-glass appearance, and reintubation was required. Ligation of the TEF was performed 53h after his birth. Following the improvement of his respiratory condition through ventilatory support and the intratracheal administration of pulmonary surfactant, he underwent repair of the esophagus on the 6th day of life. Postoperatively, he suffered from heart failure, but was treated with peritoneal dialysis and pharmacological closure of the PDA. Weaning the infant from the ventilator proved difficult, but it was finally achieved when he had reached a weight of 2268g at 3 months of age by enteral feeding. Our experience of this case demonstrates that early ligation of TEF should be performed for a premature infant with EA and TEF before respiratory distress syndrome (RDS) has developed. If a gastrostomy is required to prevent gastric distention, it should be followed by simultaneous or immediate ligation of the TEF.     

A Comparison of Two Simple Measures to Evaluate the Health Status of Asthmatics: The Asthma Bother Profile and the Airways Questionnaire 20

Simple and concise measures for health status are desirable in clinical practice. The Asthma Bother Profile (ABP), which consists of 23 items, has been developed to assess how much asthma bothers patients. The Airways Questionnaire 20 (AQ20) is a simple instrument which consists of 20 items. The purpose of this study was to investigate how the ABP and AQ20 evaluate the health status of patients with asthma. A total of 166 patients with chronic asthma (age: 48 ± 16 yr, 77 males) completed pulmonary function testing, measurement of airway hyperresponsiveness, dyspnea rating, assessments of their anxiety and depression (HADS; Hospital Anxiety and Depression Scale), and assessments of their health status. The health status was assessed using the ABP, AQ20, the short-form 36 health survey questionnaire (SF-36), the Living With Asthma Questionnaire (LWAQ) and the Asthma Quality of Life Questionnaire (AQLQ). The Japanese version of the ABP included only 15 'bother' items out of the original 23 items due to cultural differences. The scores on the ABP were widely distributed, whereas the scores on the AQ20 were skewed towards the milder end of the scale. The ABP had a strong correlation with the Avoidance and Distress constructs on the LWAQ, and Anxiety and Depression on the HADS (Rs = 0.56 ∼ 0.79), and its strongest correlation with the General Health (Rs = - 0.64) scale among the 8 subscales on the SF-36. The AQ20 had a less significant correlation with the LWAQ, AQLQ, and SF-36 than the ABP. The ABP and AQ20 were short and simple to complete, and both measures could easily be used in clinical practice. The ABP can evaluate patients more specifically with respect to distress and bother than the AQ20.