Numerical and Structural Genomic Aberrations Are Reliably Detectable in Tissue Microarrays of Formalin-Fixed Paraffin-Embedded Tumor Samples by Fluorescence In-Situ Hybridization

Few data are available regarding the reliability of fluorescence in-situ hybridization (FISH), especially for chromosomal deletions, in high-throughput settings using tissue microarrays (TMAs). We performed a comprehensive FISH study for the detection of chromosomal translocations and deletions in formalin-fixed and paraffin-embedded (FFPE) tumor specimens arranged in TMA format. We analyzed 46 B-cell lymphoma (B-NHL) specimens with known karyotypes for translocations of IGH-, BCL2-, BCL6- and MYC-genes. Locus-specific DNA probes were used for the detection of deletions in chromosome bands 6q21 and 9p21 in 62 follicular lymphomas (FL) and six malignant mesothelioma (MM) samples, respectively. To test for aberrant signals generated by truncation of nuclei following sectioning of FFPE tissue samples, cell line dilutions with 9p21-deletions were embedded into paraffin blocks. The overall TMA hybridization efficiency was 94%. FISH results regarding translocations matched karyotyping data in 93%. As for chromosomal deletions, sectioning artefacts occurred in 17% to 25% of cells, suggesting that the proportion of cells showing deletions should exceed 25% to be reliably detectable. In conclusion, FISH represents a robust tool for the detection of structural as well as numerical aberrations in FFPE tissue samples in a TMA-based high-throughput setting, when rigorous cut-off values and appropriate controls are maintained, and, of note, was superior to quantitative PCR approaches.     

Effects of VRK2 (rs2312147) on White Matter Connectivity in Patients with Schizophrenia

Recent genome-wide association studies of schizophrenia reported a novel risk variant, rs2312147 at vaccinia-related kinase 2 gene (VRK2), in multiple Asian and European samples. However, its effect on the brain structure in schizophrenia is little known. We analyzed the brain structure of 36 schizophrenia patients and 18 healthy subjects with regard to rs2312147 genotype groups. Brain magnetic resonance scans for gray matter (GM) and white matter (WM) analysis, and genotype analysis for VRK2 rs2312147, were conducted. The Positive and Negative Syndrome Scale and the Digit Symbol Test were assessed for schizophrenia patients. There was no significant difference in either GM volume or WM connectivity with regard to rs2312147 genotype in healthy subjects. In contrast, we found significant differences in the WM connectivity between rs2312147 CC and CT/TT genotype groups of schizophrenia patients. The related brain areas included the splenium of corpus callosum, the left occipital lobe WM, the internal capsule (left anterior limb and right retrolenticular part), the bilateral temporal lobe WM, the left fornix/stria terminalis, the left cingulate gyrus WM, and the left parietal lobe WM. Voxelwise correlation analysis revealed that the Digit Symbol Test scores (age corrected) correlated with the fractional anisotropy in WM tracts that previously showed significant group differences between the CT/TT and CC genotypes in the rs2312147 CT/TT genotype group, while no significant correlation was found in the CC genotype group. Our data may provide evidence for the effect of VRK2 on WM connectivity in patients with schizophrenia.     

Multidimensional Measures of Impulsivity in Obsessive-Compulsive Disorder: Cannot Wait and Stop

Objective

Although the relationship between obsessive compulsive disorder (OCD) and impulsivity has long been debated, impulsivity has not been systematically examined in clinical samples of OCD. Meanwhile, recent findings suggest that impulsivity is multi-dimensional construct that can be examined through several constructs. Therefore, this study is aimed to evaluate multiple facets of impulsivity in OCD.

Method

The recruitment includes 80 OCD and 76 healthy control participants. Participants completed a test battery comprising three behavioral tasks of stop signal task (SST), delay discounting task (DDT) and balloon analog risk test (BART), and one self-report measure of the Barratt Impulsiveness scale (BIS-11).

Results

OCD subjects showed significantly lower stop signal reaction time of SST reflecting higher action impulsivity and higher delay discounting parameter of DDT suggesting increased choice impulsivity but significantly lower adjusted mean pump of BART implying lower risk taking propensity of BART than healthy control.

Conclusion

Increased Action and choice impulsivity, and decreased risk taking propensities were found in OCD. These findings seem to be consistent with clinical characteristics of OCD such as greater preference for or avoid risky situations (avoidance), inability to wait tension relief may provoke safety behaviors (compulsion) and inability to stop already started behaviors (repetition).     

抗病毒治疗乙型肝炎相关慢加急性肝衰竭患者的临床研究

目的:探讨抗病毒治疗乙型肝炎相关慢加急性肝衰竭(acute on chronic liver failure,ACLF)的临床意义。方法:回顾性分析2007年8月~2013年8月我院收治的乙型肝炎相关慢加急性肝衰竭的住院患者80例,按照患者有无接受抗病毒治疗分为抗病毒治疗组(A组)50例和未抗病毒治疗组(B组)30例,分析患者接受治疗后的近期与远期疗效、并发症及生存率。结果:1出院时A组好转率70%;B组好转率33.3%。两组比较差异有统计学意义(x2=10.243,P=0.0010.05)。2治疗14周后A组乙肝病毒DNA阴转率72%;B组阴转率30%,两组比较差异有统计学意义(x2=13.440,P=0.0000.05)。3A组出现细菌感染45例,电解质紊乱41例,消化道出血5例,肝性脑病10例,肝肾综合征10例,B组出现细菌感染30例,电解质紊乱27例,消化道出血6例,肝性脑病10例,肝肾综合征12例,两组比较差异无统计学意义(x2=2.755,P=0.0970.05)。4随访5年,A组存活36例,死亡14例,12、36和60个月累积生存率分别为78.5%、71.2%、71.2%,B组存活5例,死亡25例,12、36和60个月累积生存率分别为35.4%、27.5%、27.5%,两组比较差异有统计学意义(P0.05)。结论:对乙型肝炎相关慢加急性肝衰竭患者给予抗病毒治疗可明显改善预后,提高生存率。     

Imiquimod Induces Apoptosis of Squamous Cell Carcinoma (SCC) Cells via Regulation of A20

Imiquimod, a nucleoside analogue of the imidazoquinoline family, is being used to treat various cutaneous cancers including squamous cell carcinoma (SCC). Imiquimod activates anti-tumor immunity via Toll-like receptor 7 (TLR7) in macrophage and other immune cells. Imiquimod can also affect tumor cells directly, regardless of its impact on immune system. In this study, we demonstrated that imiquimod induced apoptosis of SCC cells (SCC12) and A20 was involved in this process. When A20 was overexpressed, imiquimod-induced apoptosis was markedly inhibited. Conversely, knockdown of A20 potentiated imiquimod-induced apoptosis. Interestingly, A20 counteracted activation of c-Jun N-terminal kinase (JNK), suggesting that A20-regulated JNK activity was possible mechanism underlying imiquimod-induced apoptosis of SCC12 cells. Finally, imiquimod-induced apoptosis of SCC12 cells was taken place in a TLR7-independent manner. Our data provide new insight into the mechanism underlying imiquimod effect in cutaneous cancer treatment.     

Advanced Radiology Utilization in a Tertiary Care Emergency Department from 2001 to 2010

Objective

To evaluate the utilization trends of advanced radiology, i.e. computed tomography (CT) and magnetic resonance imaging (MRI), examination in an emergency department (ED) of an academic medical center from 2001 to 2010.

Patients and Methods

We assessed the overall CT and MRI utilization, and the ED patient encounters. Each examination was evaluated according to the patient’s age and anatomically relevant regions.

Results

During the study period, 737,760 patient visited the ED, and 156,287 CT and 35,018 MRI examinations were performed. The number of annual ED patients increased from 63,770 in 2001 to 94,609 in 2010 (P = 0.018). The rate of CT utilization increased from 105.5 per 1000 patient visits in 2001 to 289.2 in 2010 (P<0.001), and the rate of MRI utilization increased from 8.1 per 1000 patient visits in 2001 to 74.6 in 2010 (P<0.001). In all of the patient age groups, the overall CT and MRI utilization increased. The greater the patient age, the more likely the use of advanced radiology [CT: 87.1 per 1000 patients in age <20 vs. 293.9 per 1000 in age>60 (P<0.001); MRI: 5.1 per 1000 patients in age <20 vs. 108.7 per 1000 in age>60 (P<0.001)]. Abdomen-pelvis (40.2%) and the head (35.7%) comprised the majority of CT scans, while the head (86.4%) comprised the majority of MRI examinations. The rates of advanced radiology use increased across all anatomical regions, with the highest increase being in chest CT (5.9 per 1000 to 49.2) and head MRI (7.2 per 1000 to 61.9).

Conclusion

We report a three-fold and nine-fold increase in the use of CT and MRI, respectively, during the study period. Additional studies will be required to understand the causes of this change and to determine the effect of advanced radiology utilization on the patient outcome.     

Factors Associated with the Occurrence of Cardiac Arrest after Emergency Tracheal Intubation in the Emergency Department

Objectives

Emergency tracheal intubation has achieved high success and low complication rates in the emergency department (ED). The objective of this study was to evaluate the incidence of post-intubation CA and determine the clinical factors associated with this complication.

Methods

A matched case-control study with a case to control ratio of 1∶3 was conducted at an urban tertiary care center between January 2007 and December 2011. Critically ill adult patients requiring emergency airway management in the ED were included. The primary endpoint was post-intubation CA, defined as CA within 10 minutes after tracheal intubation. Clinical variables were compared between patients with post-intubation CA and patients without CA who were individually matched based on age, sex, and pre-existing comorbidities.

Results

Of 2,403 patients who underwent emergency tracheal intubation, 41 patients (1.7%) had a post-intubation CA within 10 minutes of the procedure. The most common initial rhythm was pulseless electrical activity (78.1%). Patients experiencing CA had higher in-hospital mortality than patients without CA (61.0% vs. 30.1%; p<0.001). Systolic hypotension prior to intubation, defined as a systolic blood pressure ≤90 mmHg, was independently associated with post-intubation CA (OR, 3.67 [95% CI, 1.58–8.55], p = 0.01).

Conclusion

Early post-intubation CA occurred with an approximate 2% frequency in the ED. Systolic hypotension before intubation is associated with this complication, which has potentially significant implications for clinicians at the time of intubation.     

Functional modulation of lysophosphatidic acid type 2 G-protein coupled receptor facilitates alveolar bone formation

Lipid biosynthesis is recently studied its functions in a range of cellular physiology including differentiation and regeneration. However, it still remains to be elucidated in its precise function. To reveal this, we evaluated the roles of lysophosphatidic acid (LPA) signaling in alveolar bone formation using the LPA type 2 receptor (LPAR2) antagonist AMG-35 (Amgen Compound 35) using tooth loss without periodontal disease model which would be caused by trauma and usually requires a dental implant to restore masticatory function. In this study, in vitro cell culture experiments in osteoblasts and periodontal ligament fibroblasts revealed cell type-specific responses, with AMG-35 modulating osteogenic differentiation in osteoblasts in vitro. To confirm the in vivo results, we employed a mouse model of tooth loss without periodontal disease. Five to 10 days after tooth extraction, AMG-35 facilitated bone formation in the tooth root socket as measured by immunohistochemistry for differentiation markers KI67, Osteocalcin, Periostin, RUNX2, transforming growth factor beta 1 (TGF-β1) and SMAD2/3. The increased expression and the localization of these proteins suggest that AMG-35 elicits osteoblast differentiation through TGF-β1 and SMAD2/3 signaling. These results indicate that LPAR2/TGF-β1/SMAD2/3 represents a new signaling pathway in alveolar bone formation and that local application of AMG-35 in traumatic tooth loss can be used to facilitate bone regeneration and healing for further clinical treatment.     

β-Catenin regulates melanocyte dendricity through the modulation of PKCζ and PKCδ

The Wnt/β-catenin signaling pathway is involved in the melanocyte differentiation and melanoma development. However, the effect of β-catenin for dendrite formation has not been clearly elucidated yet in normal human epidermal melanocytes (NHEM). To investigate the effect of β-catenin, we transduced NHEM with recombinant adenovirus expressing β-catenin. Forced expression of β-catenin led to the dramatic morphological changes of NHEM, including the reduction of dendrite length and enlargement of cell body. Concomitantly with, the protein levels for dendrite formation-related molecules, such as Rac1 and Cdc42, were markedly decreased. In addition, phosphorylation of p38 MAPK was significantly reduced by β-catenin, potentiating its inhibitory role for dendrite formation. Interestingly, overexpression of β-catenin led to the increase of protein kinase C ζ (PKCζ) level, while protein kinase C δ (PKCδ) was decreased by β-catenin, suggesting that those PKCs were β-catenin-downstream modulators in NHMC. When PKCζ was overexpressed, dendrites were shortened, with the reduced protein levels for Rac1 and Cdc42. In contrast, PKCδ overexpression led to the elongation of dendrites, with the increased levels for Rac1 and Cdc42. These results suggest that β-catenin play an inhibitory role for dendrite formation through the modulation of PKCζ and PKCδ.     

Oral immunization of haemaggulutinin H5 expressed in plant endoplasmic reticulum with adjuvant saponin protects mice against highly pathogenic avian influenza A virus infection

Pandemics in poultry caused by the highly pathogenic avian influenza (HPAI) A virus occur too frequently globally, and there is growing concern about the HPAI A virus due to the possibility of a pandemic among humans. Thus, it is important to develop a vaccine against HPAI suitable for both humans and animals. Various approaches are underway to develop such vaccines. In particular, an edible vaccine would be a convenient way to vaccinate poultry because of the behaviour of the animals. However, an edible vaccine is still not available. In this study, we developed a strategy of effective vaccination of mice by the oral administration of transgenic Arabidopsis plants (HA‐TG) expressing haemagglutinin (HA) in the endoplasmic reticulum (ER). Expression of HA in the ER resulted in its high‐level accumulation, N‐glycosylation, protection from proteolytic degradation and long‐term stability. Oral administration of HA‐TG with saponin elicited high levels of HA‐specific systemic IgG and mucosal IgA responses in mice, which resulted in protection against a lethal influenza virus infection with attenuated inflammatory symptoms. Based on these results, we propose that oral administration of freeze‐dried leaf powders from transgenic plants expressing HA in the ER together with saponin is an attractive strategy for vaccination against influenza A virus.