High time resolution fluorescence imaging with a CCD camera

We have built a high speed, sensitive camera system capable of capturing sequences of low-light level images synchronized with recordings of membrane potential. The camera system is based on a cooled, scientific grade CCD camera controlled by a PC/AT computer. It can take 100 frames/sec of 18 X 18 element images and 40 frames/sec of 50 X 50 element images with no lag in response to step changes in light intensity. High accuracy and dynamic range of the measurements result from the fact that light levels of the picture elements are digitized with 12 bit accuracy with intrinsic camera noise levels typically less than 1/10,000 of the maximum detectable light level. We have used this system to record calcium dependent fura-2 fluorescence transients in the dendrites of cerebellar Purkinje cells and from different regions of leech neurons in segmental ganglia or isolated in culture.     

Isolation and molecular characterization of the CaPHO85 gene: a negative regulator of phosphate metabolism (PHO system) in Candida albicans]

The PHO system is an ingenious mechanism by which the yeast Saccharomyces cerevisiae regulates the expression of a set of genes involved in phosphate metabolism in response to the change of phosphate concentrations in the environment. A key factor in this mechanism is the Pho85 kinase, which has been discovered as a negative regulator of the PHO system. One of the genes isolated in our laboratory in screening the protein kinase genes from Candida albicans was identified as a homologue (CaPHO85) of the PHO85 of S. cerevisiae, based on the following results. a) Pho85 is the polypeptide with the highest homology to CaPho85 (62% identity) among the S. cerevisiae genome sequence. b) The position of insertion of the intron is quite similar between CaPHO85 (in the 7th codon of the N-terminal MTGSSSQ) and S. cerevisiae PHO85 (in the 6th codon of the N-terminal MSSSSQ). c) The nucleotide sequences in the intron possess the consensus sequences for yeast intron: the 5-splice-site, internal, and the 3-splice-site sequence. d) CaPHO85 complemented the S. cerevisiae pho85 mutation. e) CaPho85 contains all of the consensus sequences for the ATP-binding domain and for the kinase domain found in S. cerevisiae Pho85.     

Intrapancreatic gastric duplication cyst mimicking pancreatic cystic tumor

Cystic lesions of the pancreas are sometimes difficult to diagnose. We report a case of a gastric duplication cyst (GDC) of the pancreas in an adult. A 45-year-old woman was admitted to our department for the investigation of anemia. Abdominal ultrasonography revealed a large cystic lesion, measuring about 40 × 70 mm with calcification in the tail of the pancreas. Contrast-enhanced computed tomography of the abdomen revealed a non-enhanced cystic lesion with non-enhanced wall. Endoscopic retrograde pancreatography revealed a mild extended main pancreatic duct and an aberrant pancreatic duct, but there was no communication with the cyst. We could not deny the malignancy, so distal pancreatectomy was performed. The pathological examination revealed that the inner wall of the cyst consisted of columnar epithelium and smooth muscle layer. Immunohistochemical analysis revealed the columnar epithelium to be immunopositive for cytokeratin 7 (CK7) and immunonegative for cytokeratin 20 (CK20) and the glands to be immunonegative for CK 7 and immunopositive for CK 20. Therefore, a diagnosis of GDC of the pancreas was made. On imaging, a GDC usually mimics a pancreatic pseudocyst or a cystic neoplasm of the pancreas. Therefore, even though it is rare, a GDC should be considered in the differential diagnosis of cystic tumors of the pancreas.     

Clinical features of pancreaticobiliary maljunction: update analysis of 2nd Japan-nationwide survey

Introduction

Pancreaticobiliary maljunction (PBM) is a congenital anomaly, which can be defined as a union of the pancreatic and biliary ducts located outside off the duodenal wall. We herein investigate clinical features of PBM including as the 2nd report of a Japanese nationwide survey.

Patients and methods

During a period of 18 years (from 1990 to 2007), 2,561 patients with PBM were registered at 141 medical institutions in Japan. Among them, eligible patients (n = 2,529) were divided into two groups: adult (n = 1,511) and pediatric patients (n = 1,018). Comparisons of clinical features including associated biliary cancers were performed according to the biliary dilatation (BD), age factor, and time era.

Results

Only one case in pediatric patients with BD combined with a bile duct cancer (0.1 %). In adult patients, the bile duct cancer and the gallbladder cancer was seen in 6.9 and 13.4 % patients with BD and in 3.1 and 37.4 % patients without BD, respectively. In adult patients with BD, the occurrence rates of biliary cancers were increased in latter period (00’–07’) compared with former period (90’–99’). The ratio of biliary cancer localization was changed between former and latter period, and the bile duct cancer was increased in latter period (from 5.5 to 9.3 %).

Conclusions

The largest series of PBM were evaluated to clarify the clinical features including the associated biliary cancer in this Japan-nationwide survey. This report could be widely used in the future as a reference data for diagnosis and treatment of PBM.     

Porphyromonas gingivalis induces murine macrophage foam cell formation

Atherosclerosis is a complex pathologic process initialed by the formation of cholesterol-rich plaque. Macrophages play a central role in the development of atherosclerosis, specifically in the initial accumulation of cholesterol in the arterial wall. It has been suggested that infection and chronic inflammatory conditions such as periodontitis may influence the atherosclerosis process. Porphyromonas gingivalis, one of the major pathogens involved in periodontitis, has been detected in human atheromas, suggesting that P. gingivalis infection may be associated with atherosclerosis. However, a causal relationship between this pathogen and the disease process has not yet been established. The purpose of the present investigation was to determine whether P. gingivalis could induce macrophages to form foam cells using the murine macrophage cell line (J774) as a model system. For inocula smaller than one bacterium per ten cells, P. gingivalis 381, as well as its lipopolysaccharide (LPS), induced foam cell formation of macrophages when cultured in the presence of human low-density lipoprotein (LDL). Infection of macrophages with increasing doses of P. gingivalis resulted in higher levels of foam cell formation. More than 70% of the cultured macrophages form cholesterol ester droplet-rich cells in the presence of 100 mug/ml of LDL when the inocula was more than 10 bacteria per cell. Low concentrations of P. gingivalis outer membrane vesicles also induced foam cell formation in the presence of LDL. In addition, it was demonstrated that P. gingivalis LPS alone was able to induce macrophage foam cell formation. P. gingivalis and its vesicles not only promoted LDL binding to macrophages but also induced macrophages to modify native LDL, which plays an important role in foam cell formation and the pathogenesis of atherosclerosis. Therefore, P. gingivalis cells or its vesicles released from periodontal lesions into the circulation may deliver virulence factor(s) such as LPS to the arterial wall to initiate or promote foam cell formation in macrophages and contribute to atheroma development.     

Effect of polycyclic aromatic hydrocarbons on generation and efflux of glutathione conjugates in primary cultured alveolar epithelial cells

This study was initiated to functionally characterize multidrug resistance associated protein (MRP)-mediated transport across the lung epithelium. Alveolar type II cells were isolated from rabbit lung and cultured on Transwell until forming a tight monolayers. After the cell monolayer was preloaded with monochlorobimane (mBCl) that is metabolized to a fluorescent glutathione conjugate (mBCl-SG), amount of mBCl-SG exported to apical and basal compartments were measured periodically. mBCl-SG was more preferentially exported in the apical direction than in the basolateral direction. Efflux of mBCl-SG from alveolar epithelial cells was significantly inhibited by a MRP inhibitor MK-571. Pharmacokinetic analysis of efflux profiles revealed that increased efflux of mBCl-SG by B[a]P is not due to enhanced MRP activity but simply due to an elevated level of mBCl-SG in the cells. Elevation of the intracellular level of mBCl-SG corresponded well to that of reduced GSH caused by B[a]P pretreatment.     

The hemorrhage caused by sporadic‐type cerebral amyloid angiopathy occurs primarily in the cerebral sulci

We examined a solitary hematoma in a patient with sporadic cerebral amyloid angiopathy (CAA). The hematoma affected the middle frontal sulcus, cerebral cortex (CC) and subcortical frontal white matter (sfWM). We embedded the hematoma in four paraffin blocks, each of which was cut serially into 6‐µm‐thick sections. The first section and every 18th section from each block were subjected to Elastica‐Goldner (E‐G) staining, and the distribution and diameter of the ruptured blood vessels (rBVs) were examined. The rBVs were then marked on diagrams representing each E‐G‐stained section. The present study yielded the following important findings: (i), early‐ and recently ruptured Aβ‐positive arteries were present mainly in the intrasulcal hematoma (ISH), rather than in the CC; (ii) many early‐ruptured arteries in the ISH were larger in diameter than those in the CC; and (iii) ruptures of the cortical arteries, even near the cortical surface, did not occur so frequently and the ruptured vessels were small in size. We concluded that in patients with subcortical hematoma caused by sporadic‐type CAA, successive rupturse of the meningeal vessels, mainly arteries, occur in the cerebral sulcus initially, followed by formation of an ISH and development of a fresh hemorrhagic or anemic infarct in the CC surrounding the ISH, the latter in most cases then extending into the brain parenchyma through the necrotic CC at the depth of the sulcus, finally creating a secondary hematoma in the subcortical white matter.     

Clinical practice guidance for juvenile idiopathic arthritis (JIA) 2018

Juvenile idiopathic arthritis (JIA) is the most common disease in pediatric rheumatism. There is no specific symptom or examination finding for JIA, and the diagnosis is made by exclusion and differentiation. Because non-pediatric rheumatologists are sometimes involved in medical care, ‘proposal for JIA guidance on diagnosis and treatment for primary care pediatricians and non-pediatric rheumatologists’ was first published in 2007. In these 10 years, a number of new findings on pathophysiology and treatment of JIA have been published; therefore, we propose this guidance of 2018th edition aiming at updating and standardization of JIA medical care in Japan. This edition included the management of uveitis, macrophage activation syndrome, infectious diseases before and during treatment. Moreover, details of biologics are also described. Although this guidance is tailored to adaptation of examinations and drugs, we do not purpose to limit the physicians' discretion in clinical practice. This guidance should be viewed as recommendations and be individualized according to the condition of the patient. We hope that medical care for JIA will advance and more patients will get benefit based on this guidance. Then, further revisions are needed due to changes in future conditions.     

Juvenile fibromyalgia: Guidance for management

Juvenile fibromyalgia (JFM) is a disease in which patients complain of acute and chronic severe pain, an overt primary cause for which cannot be found or surmised. Although patients with JFM mainly complain of systemic pain or allodynia in the medical interview and physical examination, the concept of the disease is the total sum of painful illness, chronic fatigue, hypothermia and many other autonomic symptoms and signs. Many issues are interacting including individual traits (personality, temperament, sensitivity, memory of pain; age: early adolescence), individual states (self‐esteem, anxiety, developmental level), and external stressors (parent especially mother, school environment). JFM is diagnosed on the combination of disease history, physical examination to determine the 18 tender points and allodynia, pain from gently touching their hair, and negative results of blood tests (inflammatory markers, thyroid function, myogenic enzymes). The goals of treatment are the following: restoration of function and relief of pain. Psychological support is advocated. Although the exact number of patients with JFM is still to be elucidated, it seems to be growing because pediatric rheumatologists in Japan encounter children with a wide variety of musculoskeletal pains. This guideline describes how to diagnose JFM in children and how to treat them appropriately.     

Optimal image reconstruction using multidetector‐row computed tomography to facilitate cardiac resynchronization therapy

Preprocedural recognition of the segment of latest mechanical contraction along with the anatomy of the coronary venous system is important for successful and effective cardiac resynchronization therapy. We present a case of ischemic cardiomyopathy who underwent implantation of a cardiac resynchronization therapy device with a defibrillator, which was facilitated by preprocedural computed tomographic images reconstructed to visualize the left ventricular slab and the coronary venous system simultaneously on the cardiac contour. The present reconstruction method using computed tomography is optimal and feasible method to incorporate the echocardiographic findings into the procedure performed under fluoroscopy appropriately.