New PCR-Based Open Reading Frame Typing Method for Easy,Rapid, and Reliable Identification of Acinetobacter baumannii International Epidemic Clones without Performing Multilocus Sequence Typing

Antimicrobial resistance issues have become a global health concern. The rapid identification of multidrug-resistant microbes, which depends on microbial genomic information, is essential for overcoming growing antimicrobial resistance challenges. However, genotyping methods, such as multilocus sequence typing (MLST), for identifying international epidemic clones of Acinetobacter baumannii are not easily performed as routine tests in ordinary clinical laboratories. In this study, we aimed to develop a novel genotyping method that can be performed in ordinary microbiology laboratories. Several open reading frames (ORFs) specific to certain bacterial genetic lineages or species, together with their unique distribution patterns on the chromosomes showing a good correlation with the results of MLST, were selected in A. baumannii and other Acinetobacter spp. by comparing their genomic data. The distribution patterns of the ORFs were visualized by agarose gel electrophoresis after multiplex PCR amplification and digitized. A. baumannii sequence types (STs) corresponding to international clones I and II were successfully discriminated from other STs and Acinetobacter species by detecting the distribution patterns of their ORFs using the multiplex PCR developed here. Since bacterial STs can be easily expressed as digitized numeric data with plus (+) expressed as 1 and minus (−) expressed as 0, the results of the method can be easily compared with those obtained by different tests or laboratories. This PCR-based ORF typing (POT) method can easily and rapidly identify international epidemic clones of A. baumannii and differentiate this microbe from other Acinetobacter spp. Since this POT method is easy enough to be performed even in ordinary clinical laboratories, it would also contribute to daily infection control measures and surveillance.     

Negative regulation of cell motile and invasive activities by lysophosphatidic acid receptor-3 in colon cancer HCT116 cells

Lysophosphatidic acid (LPA) mediates a wide range of biological responses with G protein-coupled transmembrane receptors (LPA receptors). So far, at least six types of LPA receptors (LPA receptor-1 (LPA₁) to LPA₆) have been identified. Recently, it has been reported that LPA₃ indicates opposite effects on cellular functions of cancer cells. In the present study, to assess a biological role of LPA₃ on cell migration ability of colon cancer cells, we generated LPA receptor-3 (LPAR3) knockdown (HCT-sh3-3) cells from HCT116 and measured cell motile and invasion activities. In motility assay with a cell culture insert, HCT-sh3-3 cells showed significantly high cell motile activity, compared with control cells. For invasion assay, the filter was coated with Matrigel. The invasive activity of HCT-sh3-3 cells was significantly higher than that of control cells. Furthermore, we also examined the effects of LPAR3 knockdown on the interaction between colon cancer cells and endothelial F-2 cells. When F-2 cells were cultured with serum-free DMEM containing a supernatant from HCT-sh3-3 cells, the cell growth rate and migration activity of F-2 cells were significantly stimulated, associating with the elevated expressions of vascular endothelial growth factor (VEGF)-A and VEGF-C genes in HCT-sh3-3 cells. These results suggest that LPA₃ may act as a negative regulator on cell motile and invasive abilities of colon cancer HCT116 cells.     

Adduction-Abduction Asymmetry in Saccades During Video-Oculographic Monocular Recording: A Word of Caution

In eye movement examination, video-oculographic monocular recording has become more popular than electro-oculographic binocular recording. The aim of this study was to examine the characteristics of monocular movements recorded using video-oculography. In 66 healthy subjects, the horizontal saccades and smooth pursuit eye movements of the right eye within a range of 30º were evaluated using a video-oculographic eye movement recording system. Saccade latency, velocity, accuracy, and smooth pursuit gain were measured and analysed by age and direction. Saccade parameters (latency, velocity, and amplitude) and smooth pursuit gain deteriorated with age in healthy subjects. Saccade velocity and accuracy were significantly larger during adduction than during abduction. The smooth pursuit gain did not differ between adduction and abduction. In conclusion, unlike smooth pursuit eye movements, saccadic eye movements have adduction-abduction asymmetry. In video-oculographic monocular recording of saccades, it is necessary to recognise the possibility of the existence of adduction-abduction asymmetry.     

Localized dissection of the sinus of valsalva without coronary artery involvement during percutaneous coronary intervention

Dissection of the sinus of Valsalva is an extremely rare accident during percutaneous coronary intervention (PCI), but it can lead to serious complications such as dissection of the ascending aorta. We experienced a localized dissection of the right coronary cusp without coronary artery involvement that was induced by a guiding catheter during PCI in a patient with acute myocardial infarction. The localized dissection showed pooling of the contrast medium in the acute phase, but it subsided spontaneously after 12 days without any sequelae. Manipulation of the guiding catheter should be performed with great caution not only in the coronary artery but also in the sinus of Valsalva.     

A case of factor X (FX) deficiency due to novel mutation V196M,FX Hofu

The patient, a 20-year-old male, was found to have a slightly prolonged prothrombin time (PT). No episodes of bleeding were noted. The measurement of coagulation factors revealed that the level of factor X (FX) activity was solely deficient, 51% (normal range: 70–130% ), and that of FX antigen was 100%. Analysis of the entire FX gene revealed the novel missense mutation of GTG to ATG, resulting in the substitution of the 196th amino acid valine → methionine. The mother and younger brother had a normal PT time and expressed no episode of bleeding. The mother exhibited a normal level of FX activity and antigen; however the younger brother showed a slight decrease in both the parameters. This mutation was not observed in the mother and younger brother. Polymorphism is not observed at this point in healthy persons. The present novel FX mutation was named FX Hofu.     

Increased macrophage colony-stimulating factor levels in patients with Graves’ disease

Previous studies have found markedly elevated serum concentrations of proinflammatory cytokines in patients with Graves’ disease (GD). We investigated the role of macrophage colony-stimulating factor (M-CSF) in GD. We assayed concentrations of M-CSF in sera from 32 patients with GD (25 untreated; 7 receiving thiamazole therapy). We also studied 32 age-matched healthy subjects as controls. Relationships between serum M-CSF and both thyroid state and serum lipids were examined. Moreover, to examine the effect of thyroid hormone alone on serum M-CSF, T3 was administered orally to normal subjects. Serum concentrations of M-CSF in GD patients who were hyperthyroid were significantly increased compared with GD patients who were euthyroid (P < 0.05) and control subjects (P < 0.0001). Serum M-CSF concentrations correlated closely with T3 levels in patients (r = 0.51, P < 0.005). Serial measurement of five individual patients revealed that serum concentrations of M-CSF were significantly decreased (P < 0.05), reaching normal control values upon attainment of euthyroidism. Furthermore, oral T3 administered to 15 volunteers for 7 days produced significant increases in serum levels of M-CSF (P < 0.05). The close correlation between serum M-CSF and serum thyroid hormone levels suggests that high circulating levels of thyroid hormones may directly or indirectly potentiate the production of M-CSF in patients with GD.     

Pioglitazone reduces urinary podocyte excretion in type 2 diabetes patients with microalbuminuria

In various renal diseases, including diabetic nephropathy, detection of podocytes in the urine indicates severe injury to podocytes in the glomeruli. Pioglitazone is a newly developed antidiabetic agent that attenuates insulin resistance. The aim of the present study was to determine whether pioglitazone affects urinary albumin excretion (UAE) or the number of urinary podocytes or both in type 2 diabetes patients with microalbuminuria. Twenty-eight patients with normotensive type 2 diabetes and microalbuminuria (18 men and 10 women; mean age, 52.5 years) and 30 age-matched normotensive controls (20 men and 10 women; mean age, 51.5 years) were included in the study. Urinary podocytes were detected by immunofluorescence with a monoclonal antibody against podocalyxin. Patients were randomly assigned to 2 groups: a pioglitazone-treatment group (30 mg/day, n = 14) and a placebo group (n = 14). Treatment was continued for 6 months. Podocytes were absent in the urine of healthy controls, but detected in 17 of 28 diabetic patients (60.7%). UAE was reduced from 96.7 +/- 50.5 microg/min to 39.7 +/- 22.9 microg/min (P <.01) in the pioglitazone-treatment group, and the number of urinary podocytes was reduced from 0.9 +/- 1.0 cells/mL to 0.1 +/- 0.2 cells/mL (P <.001). Neither UAE nor the number of urinary podocytes was affected in the placebo group. These data indicate that pioglitazone is effective for reducing UAE and podocyte injury in early-stage diabetic nephropathy.