Investigation of the cerebral hemodynamic response function in single blood vessels by functional photoacoustic microscopy

The specificity of the hemodynamic response function (HRF) is determined spatially by the vascular architecture and temporally by the evolution of hemodynamic changes. Here, we used functional photoacoustic microscopy (fPAM) to investigate single cerebral blood vessels of rats after left forepaw stimulation. In this system, we analyzed the spatiotemporal evolution of the HRFs of the total hemoglobin concentration (HbT), cerebral blood volume (CBV), and hemoglobin oxygen saturation (SO2). Changes in specific cerebral vessels corresponding to various electrical stimulation intensities and durations were bilaterally imaged with 36 × 65-μm2 spatial resolution. Stimulation intensities of 1, 2, 6, and 10 mA were applied for periods of 5 or 15 s. Our results show that the relative functional changes in HbT, CBV, and SO2 are highly dependent not only on the intensity of the stimulation, but also on its duration. Additionally, the duration of the stimulation has a strong influence on the spatiotemporal characteristics of the HRF as shorter stimuli elicit responses only in the local vasculature (smaller arterioles), whereas longer stimuli lead to greater vascular supply and drainage. This study suggests that the current fPAM system is reliable for studying relative cerebral hemodynamic changes, as well as for offering new insights into the dynamics of functional cerebral hemodynamic changes in small animals. © 2012 Society of Photo-Optical Instrumentation Engineers (SPIE)

Recombinant lipidated dengue-4 envelope protein domain III elicits protective immunity

AbstractThe combination of recombinant protein antigens with an immunostimulator has the potential to greatly increase the immunogenicity of recombinant protein antigens. In the present study, we selected the dengue-4 envelope protein domain III as a dengue vaccine candidate and expressed the protein in lipidated form using an Escherichia coli-based system. The recombinant lipidated dengue-4 envelope protein domain III folded into the proper conformation and competed with the dengue-4 virus for cellular binding sites. Mice immunized with lipidated dengue-4 envelope protein domain III without exogenous adjuvant had higher frequencies of dengue-4 envelope protein domain III-specific B cells secreting antibodies than mice immunized with the nonlipidated form. Importantly, lipidated dengue-4 envelope protein domain III-immunized mice demonstrated a durable neutralizing antibody response and had reduced viremia levels after challenge. The study demonstrates that lipidated dengue-4 envelope protein domain III is immunogenic and may be a potential dengue vaccine candidate. Furthermore, the lipidation strategy can be applied to other serotypes of dengue virus

Including Total EGFR Staining in Scoring Improves EGFR Mutations Detection by Mutation-Specific Antibodies and EGFR TKIs Response Prediction

Epidermal growth factor receptor (EGFR) is a novel target for therapy in subsets of non-small cell lung cancer, especially adenocarcinoma. Tumors with EGFR mutations showed good response to EGFR tyrosine kinase inhibitors (TKIs). We aimed to identify the discriminating capacity of immunohistochemical (IHC) scoring to detect L858R and E746-A750 deletion mutation in lung adenocarcinoma patients and predict EGFR TKIs response. Patients with surgically resected lung adenocarcinoma were enrolled. EGFR mutation status was genotyped by PCR and direct sequencing. Mutation-specific antibodies for L858R and E746-A750 deletion were used for IHC staining. Receiver operating characteristic (ROC) curves were used to determine the capacity of IHC, including intensity and/or quickscore (Q score), in differentiating L858R and E746-A750 deletion. We enrolled 143 patients during September 2000 to May 2009. Logistic-regression-model-based scoring containing both L858R Q score and total EGFR expression Q score was able to obtain a maximal area under the curve (AUC: 0.891) to differentiate the patients with L858R. Predictive model based on IHC Q score of E746-A750 deletion and IHC intensity of total EGFR expression reached an AUC of 0.969. The predictive model of L858R had a significantly higher AUC than L858R intensity only (p = 0.036). Of the six patients harboring complex EGFR mutations with classical mutation patterns, five had positive IHC staining. For EGFR TKI treated cancer recurrence patients, those with positive mutation-specific antibody IHC staining had better EGFR TKI response (p = 0.008) and longer progression-free survival (p = 0.012) than those without. In conclusion, total EGFR expression should be included in the IHC interpretation of L858R. After adjusting for total EGFR expression, the scoring method decreased the false positive rate and increased diagnostic power. According to the scoring method, the IHC method is useful to predict the clinical outcome and refine personalized therapy

Intra- and interreader reproducibility of PI-RADSv2: A multireader study

Background: The Prostate Imaging Reporting and Data System version 2 (PI-RADSv2) has been in use since 2015; while interreader reproducibility has been studied, there has been a paucity of studies investigating the intrareader reproducibility of PI-RADSv2. Purpose: To evaluate both intra- and interreader reproducibility of PI-RADSv2 in the assessment of intraprostatic lesions using multiparametric magnetic resonance imaging (mpMRI). Study Type: Retrospective. Population/Subjects: In all, 102 consecutive biopsy-naïve patients who underwent prostate MRI and subsequent MR/transrectal ultrasonography (MR/TRUS)-guided biopsy. Field Strength/Sequences: Prostate mpMRI at 3T using endorectal with phased array surface coils (TW MRI, DW MRI with ADC maps and b2000 DW MRI, DCE MRI). Assessment: Previously detected and biopsied lesions were scored by four readers from four different institutions using PI-RADSv2. Readers scored lesions during two readout rounds with a 4-week washout period. Statistical Tests: Kappa (κ) statistics and specific agreement (Po) were calculated to quantify intra- and interreader reproducibility of PI-RADSv2 scoring. Lesion measurement agreement was calculated using the intraclass correlation coefficient (ICC). Results: Overall intrareader reproducibility was moderate to substantial (κ = 0.43–0.67, Po = 0.60–0.77), while overall interreader reproducibility was poor to moderate (κ = 0.24, Po = 46). Readers with more experience showed greater interreader reproducibility than readers with intermediate experience in the whole prostate (P = 0.026) and peripheral zone (P = 0.002). Sequence-specific interreader agreement for all readers was similar to the overall PI-RADSv2 score, with κ = 0.24, 0.24, and 0.23 and Po = 0.47, 0.44, and 0.54 in T2-weighted, diffusion-weighted imaging (DWI), and dynamic contrast-enhanced (DCE), respectively. Overall intrareader and interreader ICC for lesion measurement was 0.82 and 0.71, respectively. Data Conclusion: PI-RADSv2 provides moderate intrareader reproducibility, poor interreader reproducibility, and moderate interreader lesion measurement reproducibility. These findings suggest a need for more standardized reader training in prostate MRI. Level of Evidence: 2. Technical Efficacy: Stage 2

A Novel Single-Dose Dengue Subunit Vaccine Induces Memory Immune Responses

To protect against dengue viral infection, a novel lipidated dengue subunit vaccine was rationally designed to contain the consensus amino acid sequences derived from four serotypes of dengue viruses. We found that the lipidated consensus dengue virus envelope protein domain III (LcED III) is capable of activating antigen-presenting cells and enhancing cellular and humoral immune responses. A single-dose of LcED III immunization in mice without extra adjuvant formulation is sufficient to elicit neutralizing antibodies against all four serotypes of dengue viruses. In addition, strong memory responses were elicited in mice immunized with a single-dose of LcED III. Quick, anamnestic neutralizing antibody responses to a live dengue virus challenge were elicited at week 28 post-immunization. These results demonstrate the promising possibility of a future successful tetravalent vaccine against dengue viral infections that utilizes one-dose vaccination with LcED III

Phosphorylation of LCRMP-1 by GSK3β Promotes Filopoda Formation, Migration and Invasion Abilities in Lung Cancer Cells

LCRMP-1, a novel isoform of CRMP-1, can promote cancer cell migration, invasion and associate with poor clinical outcome in patients with non-small-cell lung cancer (NSCLC). However, the underlying regulatory mechanisms of LCRMP-1 in cancer cell invasiveness still remain obscure. Here, we report that GSK3β can phosphorylate LCRMP-1 at Thr-628 in consensus sequences and this phosphorylation is crucial for function of LCRMP-1 to promote filopodia formation, migration and invasion in cancer cells. Impediment of Thr-628 phosphorylation attenuates the stimulatory effects of LCRMP-1 on filopodia forming, migration and invasion abilities in cancer cells; simultaneously, kinase-dead GSK3β diminishes regulation of LCRMP-1 on cancer cell invasion. Furthermore, we also found that patients with low-level Ser-9-phosphorylated GSK3β expression and high-level LCRMP-1 expression have worse overall survival than those with high-level inactive GSK3β expressions and low-level LCRMP-1 expressions (P<0.0001). Collectively, these results demonstrate that GSK3β-dependent phosphorylation of LCRMP-1 provides an important mechanism for regulation of LCRMP-1 on cancer cell invasiveness and clinical outcome

Soft Spectrum in Yukawa-Gauge Mediation

We introduce a model independent parametrization for a subclass of gauge mediated theories, which we refer to as Yukawa-gauge mediation. Within this formalism we study the resulting soft masses in the visible spectrum. We find general expressions for the gaugino and scalar masses. Under generic conditions, the gaugino mass is screened, vanishing at first order in the SUSY breaking scale.Comment: 22 pages, 4 figures; v2: minor corrections, published versio

SLC37A1 and SLC37A2 Are Phosphate-Linked, Glucose-6-Phosphate Antiporters

Blood glucose homeostasis between meals depends upon production of glucose within the endoplasmic reticulum (ER) of the liver and kidney by hydrolysis of glucose-6-phosphate (G6P) into glucose and phosphate (Pi). This reaction depends on coupling the G6P transporter (G6PT) with glucose-6-phosphatase-α (G6Pase-α). Only one G6PT, also known as SLC37A4, has been characterized, and it acts as a Pi-linked G6P antiporter. The other three SLC37 family members, predicted to be sugar-phosphate:Pi exchangers, have not been characterized functionally. Using reconstituted proteoliposomes, we examine the antiporter activity of the other SLC37 members along with their ability to couple with G6Pase-α. G6PT- and mock-proteoliposomes are used as positive and negative controls, respectively. We show that SLC37A1 and SLC37A2 are ER-associated, Pi-linked antiporters, that can transport G6P. Unlike G6PT, neither is sensitive to chlorogenic acid, a competitive inhibitor of physiological ER G6P transport, and neither couples to G6Pase-α. We conclude that three of the four SLC37 family members are functional sugar-phosphate antiporters. However, only G6PT/SLC37A4 matches the characteristics of the physiological ER G6P transporter, suggesting the other SLC37 proteins have roles independent of blood glucose homeostasis

Enterovirus 71 3C Protease Cleaves a Novel Target CstF-64 and Inhibits Cellular Polyadenylation

Identification of novel cellular proteins as substrates to viral proteases would provide a new insight into the mechanism of cell–virus interplay. Eight nuclear proteins as potential targets for enterovirus 71 (EV71) 3C protease (3Cpro) cleavages were identified by 2D electrophoresis and MALDI-TOF analysis. Of these proteins, CstF-64, which is a critical factor for 3′ pre-mRNA processing in a cell nucleus, was selected for further study. A time-course study to monitor the expression levels of CstF-64 in EV71-infected cells also revealed that the reduction of CstF-64 during virus infection was correlated with the production of viral 3Cpro. CstF-64 was cleaved in vitro by 3Cpro but neither by mutant 3Cpro (in which the catalytic site was inactivated) nor by another EV71 protease 2Apro. Serial mutagenesis was performed in CstF-64, revealing that the 3Cpro cleavage sites are located at position 251 in the N-terminal P/G-rich domain and at multiple positions close to the C-terminus of CstF-64 (around position 500). An accumulation of unprocessed pre-mRNA and the depression of mature mRNA were observed in EV71-infected cells. An in vitro assay revealed the inhibition of the 3′-end pre-mRNA processing and polyadenylation in 3Cpro-treated nuclear extract, and this impairment was rescued by adding purified recombinant CstF-64 protein. In summing up the above results, we suggest that 3Cpro cleavage inactivates CstF-64 and impairs the host cell polyadenylation in vitro, as well as in virus-infected cells. This finding is, to our knowledge, the first to demonstrate that a picornavirus protein affects the polyadenylation of host mRNA