Prospective Memory and Default Mode Network Functional Connectivity in Normal and Pathological Aging.

Background: Prospective memory (PM), the ability to execute a previously formed intention given the proper circumstance, has been proven to be vulnerable to Alzheimer's disease. Previous studies have indicated the involvement of the frontoparietal networks; however, it is proposed that PM may also be associated with other neural substrates that support stimulus-dependent spontaneous cognition.

Objective: The present study aimed to examine the hypothesis that PM deficit in Alzheimer's disease is related to altered functional connectivity (FC) within the default mode network (DMN).

In vitro evaluation of the hyaluronic acid/alginate composite powder for topical haemostasis and wound healing.

The use of haemostatic agents can provide life-saving treatment for patients who suffer from massive bleeding in both prehospital and intraoperative conditions. However, there are still urgent demands for novel haemostatic materials that exhibit better haemostatic activity, biocompatibility, and biodegradability than existing products. In the present study, we aim to evaluate the feasibility of new wound dressing, RapidClot, for treating uncontrolled haemorrhage through a series of in vitro assessments to determine the swelling ratio, clotting time, enzymatic degradation, haemolytic activity, cytotoxicity, cell proliferation, and migration.

Enhancing Protein Production Yield from Chinese Hamster Ovary Cells by CRISPR Interference.

Chinese hamster ovary (CHO) cells are an important host for biopharmaceutical production. Generation of stable CHO cells typically requires cointegration of dhfr and a foreign gene into chromosomes and subsequent methotrexate (MTX) selection for coamplification of dhfr and foreign gene. CRISPR interference (CRISPRi) is an emerging system that effectively suppresses gene transcription through the coordination of dCas9 protein and guide RNA (gRNA).

Enhanced enterovirus 71 virus-like particle yield from a new baculovirus design.

Enterovirus 71 (EV71) is responsible for the outbreaks of hand-foot-and-mouth disease in the Asia-Pacific region. To produce the virus-like particle (VLP) vaccine, we previously constructed recombinant baculoviruses to co-express EV71 P1 polypeptide and 3CD protease using the Bac-to-Bac(®) vector system. The recombinant baculoviruses resulted in P1 cleavage by 3CD and subsequent VLP assembly in infected insect cells, but caused either low VLP yield or excessive VLP degradation.

Development of EV71 virus-like particle purification processes.

Enterovirus 71 (EV71) causes the outbreaks of hand-foot-and-mouth disease and results in deaths of hundreds of young children. EV71 virus-like particles (VLPs) are empty capsids consisting of viral structural proteins and can elicit potent immune responses, thus holding promise as an EV71 vaccine candidate. However, an efficient, scalable production and purification scheme is missing.

Enterovirus-71 virus-like particles induce the activation and maturation of human monocyte-derived dendritic cells through TLR4 signaling.

Enterovirus 71 (EV71) causes seasonal epidemics of hand-foot-and-mouth disease and has a high mortality rate among young children. We recently demonstrated potent induction of the humoral and cell-mediated immune response in monkeys immunized with EV71 virus-like particles (VLPs), with a morphology resembling that of infectious EV71 virions but not containing a viral genome, which could potentially be safe as a vaccine for EV71. To elucidate the mechanisms through which EV71 VLPs induce cell-mediated immunity, we studied the immunomodulatory effects of EV71 VLPs on human monocyte-derived dendritic cells (DCs), which bind to and incorporate EV71 VLPs.

Update on baculovirus as an expression and/or delivery vehicle for vaccine antigens.

After three decades of development, the baculovirus/insect cell expression system is now recognized as a powerful platform for recombinant protein production. With a number of distinct advantages, the baculovirus/insect cell expression system has been extensively used for the production of various vaccine candidates, and several human and veterinary vaccine products have been commercially available. In addition to insect cells, baculovirus is capable of entering a broad range of mammalian cells, lending itself to a promising gene delivery vehicle for antigen expression and display in vivo.

Efficient gene delivery into cell lines and stem cells using baculovirus.

Baculovirus is a promising vector for transducing numerous types of mammalian cells. We have developed hybrid baculovirus vectors and protocols for the efficient transduction of a variety of cell lines, primary cells and stem cells, including bone marrow-derived mesenchymal stem cells (BMSCs) and adipose-derived stem cells (ASCs). The hybrid vector enables intracellular minicircle formation and prolongs transgene expression.

Long-term tracking of segmental bone healing mediated by genetically engineered adipose-derived stem cells: focuses on bone remodeling and potential side effects.

We previously showed that transplantation of adipose-derived stem cells (ASCs) engineered with hybrid baculovirus (BV) persistently expressing bone morphogenetic protein 2 (BMP2)/vascular endothelial growth factor (VEGF) into segmental defects in New Zealand White (NZW) rabbits led to successful defect reunion. By using microcomputed tomography and histology, here we further demonstrated that transplanting the hybrid BV-engineered ASCs into the massive defects (10 mm in length) at the femoral diaphysis of NZW rabbits resulted in trabecular bone formation in the interior via endochondral ossification and bone remodeling at 3 months post-transplantation. The progression of bone remodeling gave rise to the resorption of trabecular bone and conspicuous reconstruction of medullary cavity and cortical bone with lamellar structure at 8 months post-transplantation, hence conferring mechanical properties that were comparable to those of nonoperated femora.

Evaluation of the stability of enterovirus 71 virus-like particle.

Enterovirus 71 (EV71) is responsible for the outbreaks of hand-foot-and-mouth disease that caused significant mortality in children, but no vaccine is available yet. EV71 virus-like particle (VLP) is the empty capsid consisting of viral structural proteins but can elicit potent immune responses, rendering VLP a promising EV71 vaccine candidate. To evaluate whether VLP remains stable after long-term storage, which is crucial for advancing the VLP vaccine to the clinical setting, we evaluated the effects of NaCl concentration, buffers and temperatures on the VLP stability.

Recent progresses in gene delivery-based bone tissue engineering.

Gene therapy has converged with bone engineering over the past decade, by which a variety of therapeutic genes have been delivered to stimulate bone repair. These genes can be administered via in vivo or ex vivo approach using either viral or nonviral vectors. This article reviews the fundamental aspects and recent progresses in the gene therapy-based bone engineering, with emphasis on the new genes, viral vectors and gene delivery approaches.

Regenerating cartilages by engineered ASCs: prolonged TGF-β3/BMP-6 expression improved articular cartilage formation and restored zonal structure.

Adipose-derived stem cells (ASCs) hold promise for cartilage regeneration but their chondrogenesis potential is inferior. Here, we used a baculovirus (BV) system that exploited FLPo/Frt-mediated transgene recombination and episomal minicircle formation to genetically engineer rabbit ASCs (rASCs). The BV system conferred prolonged and robust TGF-β3/BMP-6 expression in rASCs cultured in porous scaffolds, which critically augmented rASCs chondrogenesis and suppressed osteogenesis/hypertrophy, leading to the formation of cartilaginous constructs with improved maturity and mechanical properties in 2-week culture.

Enhanced and prolonged baculovirus-mediated expression by incorporating recombinase system and in cis elements: a comparative study.

Baculovirus (BV) is a promising gene vector but mediates transient expression. To prolong the expression, we developed a binary system whereby the transgene in the substrate BV was excised by the recombinase (ΦC31o, Cre or FLPo) expressed by a second BV and recombined into smaller minicircle. The recombination efficiency was lower by ΦC31o (≈40-75%), but approached ≈90-95% by Cre and FLPo in various cell lines and stem cells [e.

Adaptive immune responses elicited by baculovirus and impacts on subsequent transgene expression in vivo.

Baculovirus (BV) is a promising gene therapy vector and typically requires readministration because BV mediates transient expression. However, how the prime-boost regimen triggers BV-specific adaptive responses and their impacts on BV readministration, transgene expression, and therapeutic/vaccine efficacy remain unknown. Here we unraveled that BV injection into BALB/c mice induced the production of BV-specific antibodies, including IgG1 and IgG2a, which could neutralize BV by antagonizing the envelope protein gp64 and impede BV-mediated transgene expression.

Baculovirus vector as an avian influenza vaccine: hemagglutinin expression and presentation augment the vaccine immunogenicity.

Baculovirus simultaneously displaying and expressing the avian influenza virus (AIV) hemagglutinin (HA) protein can induce potent anti-HA humoral and cellular immune responses. Based on the hypothesis that improving the antigen expression and presentation can further boost the AIV vaccine efficacies, we first constructed a baculoviral vector (Bac-HAW) with HA gene fused with the woodchuck hepatitis virus post-transcriptional regulatory element (WPRE) near its 3' end and expressed under the control of the hybrid CAG promoter. The WPRE fusion improved the HA expression and augmented the humoral and Th1 cellular immune responses after intramuscular administration into BALB/c mice.

Recent patents on the baculovirus systems.

Baculovirus has been widely utilized as a protein production tool in insect cells for nearly 3 decades and has captured growing interests as a vector for gene delivery into mammalian cells and animals over the past decade. This review summarizes important patents pertaining to the use of baculovirus for insect cell infection and mammalian cell transduction, with special emphasis on the vector development, new applications and downstream processing.

Enterovirus 71 virus-like particle vaccine: improved production conditions for enhanced yield.

To develop the enterovirus 71 (EV71) vaccine, we previously constructed a recombinant baculovirus (Bac-P1-3CD) co-expressing EV71 P1 (under polyhedrin promoter) and 3CD (under p10 promoter) proteins, which caused P1 cleavage by 3CD protease and self-assembly of virus-like particles (VLPs) in Sf-9 cells. Assuming that reducing the 3CD expression can alleviate the competition with P1 expression and elevate the VLPs yield, hereby we constructed Bac-P1-C3CD and Bac-P1-I3CD expressing 3CD under weaker CMV and IE-1 promoters, respectively. Western blot and ELISA analyses revealed that Bac-P1-C3CD and Bac-P1-I3CD led to the VLPs release into the supernatant and enhanced the extracellular VLPs yield in Sf-9 cells, but gave poor VLPs production in High Five™ (Hi-5) cells.

ASPM is a novel marker for vascular invasion, early recurrence, and poor prognosis of hepatocellular carcinoma.

Purpose: Abnormal spindle-like microcephaly associated (ASPM) plays an important role in neurogenesis and cell proliferation. This study is to elucidate its role in hepatocellular carcinoma (HCC), particularly early tumor recurrence (ETR) and prognosis.

Experimental Design: We used reverse transcription-PCR assays to measure the ASPM mRNA levels in 247 HCC and correlated with clinicopathologic and molecular features.