Elastic modulus of muscle and tendon with shear wave ultrasound elastography: variations with different technical settings

Standardization on Shear wave ultrasound elastography (SWUE) technical settings will not only ensure that the results are accurate, but also detect any differences over time that may be attributed to true physiological changes. The present study evaluated the variations of elastic modulus of muscle and tendon using SWUE when different technical aspects were altered. The results of this study indicated that variations of elastic modulus of muscle and tendon were found when different transducer's pressure and region of interest (ROI)'s size were applied. No significant differences in elastic modulus of the rectus femoris muscle and patellar tendon were found with different acquisition times of the SWUE sonogram. The SWUE on the muscle and tendon should be performed with the lightest transducer's pressure, a shorter acquisition time for the SWUE sonogram, while measuring the mean elastic modulus regardless the ROI's size.     

Paradoxical Impact of Two Folate Receptors,FRα and RFC,in Ovarian Cancer: Effect on Cell Proliferation,Invasion and Clinical Outcome

Despite being an essential vitamin, folate has been implicated to enhance tumor growth, as evidenced by reports on overexpression of folate receptor alpha (FRα) in carcinomas. The role of another folate transporter, reduced folate carrier (RFC), is largely unknown. This study investigated the roles of folate, FRα and RFC in ovarian cancers. We demonstrated FRα mRNA and protein overexpression and reduced RFC expression in association with FRα gene amplification and RFC promoter hypermethylation, respectively. FRα overexpression was associated with tumor progression while RFC expression incurred a favorable clinical outcome. Such reciprocal expression pattern was also observed in ovarian cancer cell lines. Folate was shown to promote cancer cell proliferation, migration and invasion in vitro, and down-regulate E-cadherin expression. This effect was blocked after either stable knockdown of FRα or ectopic overexpression of RFC. This hitherto unreported phenomenon suggests that, RFC can serve as a balancing partner of FRα and confer a protective effect in patients with high FRα-expressing ovarian carcinomas, as evidenced by their prolonged overall and disease-free survivals. In conclusion, we report on the paradoxical impact of FRα (putative oncogenic) and RFC (putative tumor suppressive) in human malignancies. FRα and RFC may potentially be explored as therapeutic target or prognostic marker respectively. We recommend caution and additional research on folate supplements in cancer patients.     

Evaluation of Tyrosine Kinase Inhibitor Combinations for Glioblastoma Therapy

Glioblastoma multiforme (GBM) is the most common intracranial cancer but despite recent advances in therapy the overall survival remains about 20 months. Whole genome exon sequencing studies implicate mutations in the receptor tyrosine kinase pathways (RTK) for driving tumor growth in over 80% of GBMs. In spite of various RTKs being mutated or altered in the majority of GBMs, clinical studies have not been able to demonstrate efficacy of molecular targeted therapies using tyrosine kinase inhibitors in GBMs. Activation of multiple downstream signaling pathways has been implicated as a possible means by which inhibition of a single RTK has been ineffective in GBM. In this study, we sought a combination of approved drugs that would inhibit in vitro and in vivo growth of GBM oncospheres. A combination consisting of gefitinib and sunitinib acted synergistically in inhibiting growth of GBM oncospheres in vitro. Sunitinib was the only RTK inhibitor that could induce apoptosis in GBM cells. However, the in vivo efficacy testing of the gefitinib and sunitinib combination in an EGFR amplified/ PTEN wild type GBM xenograft model revealed that gefitinib alone could significantly improve survival in animals whereas sunitinib did not show any survival benefit. Subsequent testing of the same drug combination in a different syngeneic glioma model that lacked EGFR amplification but was more susceptible to sunitinib in vitro demonstrated no survival benefit when treated with gefitinib or sunitinib or the gefitinib and sunitinib combination. Although a modest survival benefit was obtained in one of two animal models with EGFR amplification due to gefitinib alone, the addition of sunitinib, to test our best in vitro combination therapy, did not translate to any additional in vivo benefit. Improved targeted therapies, with drug properties favorable to intracranial tumors, are likely required to form effective drug combinations for GBM.     

Phenotyping of tianma-stimulated differentiated rat neuronal b104 cells by quantitative proteomics

Gastrodia elata blume (tianma) is a traditional Chinese herb often used in the treatment of convulsions, headaches, and hypertension. Although interest in neuronal-related actions of tianma is increasing, minimal studies have been conducted to determine its specific effects on neuronal cells. This study was designed to examine the effects of tianma on the metabolism in differentiated neuroblastoma cells using the isobaric tag for relative and absolute quantitation (iTRAQ) technology. Stimulation of these cells with tianma caused changes in the expression of 38 proteins that were subsequently classified according to their physiological functions and association with neurodegenerative diseases. We identified six proteins with altered functional activities in neurodegenerative disease states that were modulated by tianma: triosephosphate isomerase (Tpi1), peptidyl-prolyl cis-trans isomerase A (Ppia), neural cell adhesion molecule 1 (Ncam1), ubiquitin carboxyl-terminal hydrolase isozyme L1 (Uchl1), septin-2 (Sept2) and heat shock protein 90 (Hsp90aa1). We postulate that tianma mediates its neuroprotective effects via upregulation of Ncam1, Hsp90aa1, Tpi1 and Ppia while downregulating Sept2 and Uchl1. These changes in protein expression aid in the restoration of the intracellular environment to a metabolically balanced state, promoting cell survival. Based on these observed data, we conclude that tianma has therapeutic potential, especially for neurodegenerative diseases.     

Seroepidemiology of <Emphasis Type="Italic">Klebsiella pneumoniae</Emphasis> colonizing the intestinal tract of healthy chinese and overseas chinese adults in Asian countries

Background  

Capsular serotypes K1 and K2 of Klebsiella pneumoniae are thought to the major virulence determinants responsible for liver abscess. The intestine is one of the major reservoirs of K. pneumoniae, and epidemiological studies have suggested that the majority of K. pneumoniae infections are preceded by colonization of the gastrointestinal tract. The possibility of fecal-oral transmission in liver abscess has been raised on the basis of molecular typing of isolates. Data on the serotype distribution of K. pneumoniae in stool samples from healthy individuals has not been previously reported. This study investigated the seroepidemiology of K. pneumoniae isolates from the intestinal tract of healthy Chinese in Asian countries. Stool specimens from healthy adult Chinese residents of Taiwan, Japan, Hong Kong, China, Thailand, Malaysia, Singapore, and Vietnam were collected from August 2004 to August 2010 for analysis.     

Testing microstructural adaptation in the earliest dental tools

Conodont elements are the earliest vertebrate dental structures. The dental tools on elements responsible for food fracture—cusps and denticles—are usually composed of lamellar crown tissue (a putative enamel homologue) and the enigmatic tissue known as ‘white matter’. White matter is unique to conodonts and has been hypothesized to be a functional adaptation for the use of elements as teeth. We test this quantitatively using finite-element analysis. Our results indicate that white matter allowed cusps and denticles to withstand greater tensile stresses than do cusps comprised solely of lamellar crown tissue. Microstructural variation is demonstrably associated with dietary and loading differences in teeth, so secondary loss of white matter through conodont phylogeny may reflect changes in diet and element occlusal kinematics. The presence, development and distribution of white matter could thus provide constraints on function in the first vertebrate dental structures.     

Testosterone Plus Low-Intensity Physical Training in Late Life Improves Functional Performance,Skeletal Muscle Mitochondrial Biogenesis,and Mitochondrial Quality Control in Male Mice

Testosterone supplementation increases muscle mass in older men but has not been shown to consistently improve physical function and activity. It has been hypothesized that physical exercise is required to induce the adaptations necessary for translation of testosterone-induced muscle mass gain into functional improvements. However, the effects of testosterone plus low intensity physical exercise training (T/PT) on functional performance and bioenergetics are unknown. In this pilot study, we tested the hypothesis that combined administration of T/PT would improve functional performance and bioenergetics in male mice late in life more than low-intensity physical training alone. 28-month old male mice were randomized to receive T/PT or vehicle plus physical training (V/PT) for 2 months. Compare to V/PT control, administration of T/PT was associated with improvements in muscle mass, grip strength, spontaneous physical movements, and respiratory activity. These changes were correlated with increased mitochondrial DNA copy number and expression of markers for mitochondrial biogenesis. Mice receiving T/PT also displayed increased expression of key elements for mitochondrial quality control, including markers for mitochondrial fission-and-fusion and mitophagy. Concurrently, mice receiving T/PT also displayed increased expression of markers for reduced tissue oxidative damage and improved muscle quality. Conclusion: Testosterone administered with low-intensity physical training improves grip strength, spontaneous movements, and respiratory activity. These functional improvements were associated with increased muscle mitochondrial biogenesis and improved mitochondrial quality control.     

Ezrin Interacts with the SARS Coronavirus Spike Protein and Restrains Infection at the Entry Stage

Background

Entry of Severe Acute Respiratory Syndrome coronavirus (SARS-CoV) and its envelope fusion with host cell membrane are controlled by a series of complex molecular mechanisms, largely dependent on the viral envelope glycoprotein Spike (S). There are still many unknowns on the implication of cellular factors that regulate the entry process.

Methodology/Principal Findings

We performed a yeast two-hybrid screen using as bait the carboxy-terminal endodomain of S, which faces the cytosol during and after opening of the fusion pore at early stages of the virus life cycle. Here we show that the ezrin membrane-actin linker interacts with S endodomain through the F1 lobe of its FERM domain and that both the eight carboxy-terminal amino-acids and a membrane-proximal cysteine cluster of S endodomain are important for this interaction in vitro. Interestingly, we found that ezrin is present at the site of entry of S-pseudotyped lentiviral particles in Vero E6 cells. Targeting ezrin function by small interfering RNA increased S-mediated entry of pseudotyped particles in epithelial cells. Furthermore, deletion of the eight carboxy-terminal amino acids of S enhanced S-pseudotyped particles infection. Expression of the ezrin dominant negative FERM domain enhanced cell susceptibility to infection by SARS-CoV and S-pseudotyped particles and potentiated S-dependent membrane fusion.

Conclusions/Significance

Ezrin interacts with SARS-CoV S endodomain and limits virus entry and fusion. Our data present a novel mechanism involving a cellular factor in the regulation of S-dependent early events of infection.     

The M, E, and N structural proteins of the severe acute respiratory syndrome coronavirus are required for efficient assembly, trafficking, and release of virus-like particles

The production of virus-like particles (VLPs) constitutes a relevant and safe model to study molecular determinants of virion egress. The minimal requirement for the assembly of VLPs for the coronavirus responsible for severe acute respiratory syndrome in humans (SARS-CoV) is still controversial. Recent studies have shown that SARS-CoV VLP formation depends on either M and E proteins or M and N proteins. Here we show that both E and N proteins must be coexpressed with M protein for the efficient production and release of VLPs by transfected Vero E6 cells. This suggests that the mechanism of SARS-CoV assembly differs from that of other studied coronaviruses, which only require M and E proteins for VLP formation. When coexpressed, the native envelope trimeric S glycoprotein is incorporated onto VLPs. Interestingly, when a fluorescent protein tag is added to the C-terminal end of N or S protein, but not M protein, the chimeric viral proteins can be assembled within VLPs and allow visualization of VLP production and trafficking in living cells by state-of-the-art imaging technologies. Fluorescent VLPs will be used further to investigate the role of cellular machineries during SARS-CoV egress.