Rapid Ped-2E9 cell-based cytotoxicity analysis and genotyping of Bacillus species

Bacillus species causing food-borne disease produce multiple toxins eliciting gastroenteritis. Toxin assays with mammalian cell cultures are reliable but may take 24 to 72 h to complete and also lack sensitivity. Here, a sensitive and rapid assay was developed using a murine hybridoma Ped-2E9 cell model. Bacillus culture supernatants containing toxins were added to a Ped-2E9 cell line and analyzed for cytotoxicity with an alkaline phosphatase release assay. Most Bacillus cereus strains produced positive cytotoxicity results within 1 h, and data were comparable to those obtained with the standard Chinese hamster ovary (CHO)-based cytotoxicity assay, which took about 72 h to complete. Moreover, the Ped-2E9 cell assay had 25- to 58-fold-higher sensitivity than the CHO assay. Enterotoxin-producing Bacillus thuringiensis also gave positive results with Ped-2E9 cells, while several other Bacillus species were negative. Eight isolates from food suspected of Bacillus contamination were also tested, and only one strain, which was later confirmed as B. cereus, gave a positive result. In comparison with two commercial diarrheal toxin assay kits (BDE-VIA and BCET-RPLA), the Ped-2E9 assay performed more reliably. Toxin fractions of >30 kDa showed the highest degree of cytotoxicity effects, and heat treatment significantly reduced the toxin activity, indicating the involvement of a heat-labile high-molecular-weight component in Ped-2E9 cytotoxicity. PCR results, in most cases, were in agreement with the cytotoxic potential of each strain. Ribotyping was used to identify cultures and indicated differences for several previously reported isolates. This Ped-2E9 cell assay could be used as a rapid (approximately 1-h) alternative to current methods for sensitive detection of enterotoxins from Bacillus species.     

Polycystin-1 induces resistance to apoptosis through the phosphatidylinositol 3-kinase/Akt signaling pathway

Polycystin-1 (PC-1), the PKD1 gene product, is a large receptor whose expression in renal epithelial cells results in resistance to apoptosis and tubulogenesis, a model consistent with the phenotype observed in patients. This study links PC-1 expression to a signaling pathway that is known to be both antiapoptotic and important for normal tubulogenesis. This study found that PC-1 expression results in phosphorylation of Akt and downstream effectors and that phosphatidylinositol 3-kinase (PI3-K) inhibitors prevent this process. In addition, it is shown that dominant negative Akt can revert PC-1-induced protection from apoptosis. Furthermore, it was observed that increased PI3-K beta activity in PC-1-expressing MDCK cells seems to be dependent on both tyrosine-kinase activity and heterotrimeric G proteins. It also was found that PC-1-induced tubulogenesis is inhibited by PI3-K inhibitors. Taken together, these data suggest that the PI3-K/Akt cascade may be a central modulator of PC-1 function and that its deregulation might be important in autosomal dominant polycystic kidney disease.     

Spontaneous infantile choledochal cyst perforation

Spontaneous perforation is a rare complication of infantile choledochal cysts. A 23-weeks female with acute biliary peritonitis is described and differentiation from a spontaneous perforation of the extrahepatic bile duct highlighted.     

A deflectable guiding catheter for real-time MRI-guided interventions

Purpose:

To design a deflectable guiding catheter that omits long metallic components yet preserves mechanical properties to facilitate therapeutic interventional MRI procedures.

Materials and Methods:

The catheter shaft incorporated Kevlar braiding. A 180° deflection was attained with a 5‐cm nitinol slotted tube, a nitinol spring, and a Kevlar pull string. We tested three designs: passive, passive incorporating an inductively coupled coil, and active receiver. We characterized mechanical properties, MRI properties, RF induced heating, and in vivo performance in swine.

Results:

Torque and tip deflection force were satisfactory. Representative procedures included hepatic and azygos vein access, laser cardiac septostomy, and atrial septal defect crossing. Visualization was best in the active configuration, delineating profile and tip orientation. The passive configuration could be used in tandem with an active guidewire to overcome its limited conspicuity. There was no RF‐induced heating in all configurations under expected use conditions in vitro and in vivo.

Conclusion:

Kevlar and short nitinol component substitutions preserved mechanical properties. The active design offered the best visibility and usability but reintroduced metal conductors. We describe versatile deflectable guiding catheters with a 0.057” lumen for interventional MRI catheterization. Implementations are feasible using active, inductive, and passive visualization strategies to suit application requirements. J. Magn. Reson. Imaging 2012;35:908–915. © 2011 Wiley Periodicals, Inc.     

Soft tissue knee injuries

Soft tissue injuries of the knee are some of the most common and clinically challenging musculoskeletal disorders in patients presenting to the emergency department. Establishing clear-cut diagnostic and therapeutic objectives for these injuries is critical. The aim of this article is to outline the pathophysiology and management principles of the most common of these injuries.     

Implications of acute left ventricular remodeling during squatting stress echocardiography

Background: We previously demonstrated that squatting induces left ventricular (LV) wall motion abnormalities (WMA) in areas subtended by stenotic coronary arteries. In addition, it was observed that some subjects developed acute changes in LV shape (acute left ventricular remodeling [ALVRM]) during squatting. Objective: This study tested the hypothesis that patients with ALVRM during squatting echocardiography have higher incidences of severe coronary artery disease (CAD). Methods: Echocardiography was performed in all standard views during standing and squatting. End‐systolic frames in the apical four‐chamber view were analyzed. Results: The subjects were divided into three groups. Group 1 consisted of 12 subjects who developed squatting‐induced ALVRM with apical and distal posterior septal akinesis, dilation of the apex and marked LV shape change at end‐systole. Group 2 consisted of 20 subjects with distal posterior septal and apical akinesis without ALVRM, during squatting. Group 3 consisted of 64 subjects who developed WMA in areas other than the apex (n = 49), or normal wall motion (n = 15) during squatting. Coronary angiography in group 1 revealed that 6 subjects had left main coronary artery stenosis (LMCAS ≥ 50%), two had severe three vessel disease (≥90% stenosis), and one had 100% left anterior descending coronary artery occlusion. Severe CAD was defined for purpose of this study as the presence of LMCAS, or severe three vessel disease (≥90% stenosis). Six subjects in group 2 had LMCAS and none had severe three vessel disease (P < 0.05 vs. group 1 for LMCAS and/or three vessel disease). In group 3, eight had LMCAS and none had severe three vessel disease (P < 0.0001 vs. group 1). Conclusion: Patients with ALVRM have severe CAD. Therefore, patients who develop ALVRM during squatting require urgent evaluation for revascularization therapy. (Echocardiography 2012;29:700‐705)     

Real-time cardiovascular magnetic resonance subxiphoid pericardial access and pericardiocentesis using off-the-shelf devices in swine

Background

Needle access or drainage of pericardial effusion, especially when small, entails risk of bystander tissue injury or operator uncertainty about proposed trajectories. Cardiovascular magnetic resonance (CMR) might allow enhanced imaging guidance.

Methods and results

We used real-time CMR to guide subxiphoid pericardial access in naïve swine using commercial 18G titanium puncture needles, which were exchanged for pericardial catheters. To test the value of CMR needle pericardiocentesis, we also created intentional pericardial effusions of a range of volumes, via a separate transvenous-transatrial catheter. We performed these procedures in 12 animals.Pericardiocentesis was performed in 2:47 ± 1:43 minutes; pericardial access was performed in 1:40 ± 4:34 minutes. The procedure was successful in all animals. Moderate and large effusions required only one needle pass. There were no complications, including pleural, hepatic or myocardial transit.

Conclusions

CMR guided pericardiocentesis is attractive because the large field of view and soft tissue imaging depict global anatomic context in arbitrary planes, and allow the operator to plan trajectories that limit inadvertent bystander tissue injury. More important, CMR provides continuous visualization of the needle and target throughout the procedure. Using even passive needle devices, CMR enabled rapid pericardial needle access and drainage. We believe this experience supports clinical testing of real-time CMR guided needle access or drainage of the pericardial space. We suspect this would be especially helpful in “difficult” pericardial access, for example, in distorted thoracic anatomy or loculated effusion.