Up-regulated microRNA-185-3p inhibits the development of hyperlipidemia in rats

Objective MicroRNA (miR)-185-3p roles have been probed in multiple cancers, while the underlying function of miR-185-3p in hyperlipidemia remained obscure. This research was conducted to unravel miR-185-3p function in hyperlipidemia development via modulating mastermind-like 1 (MAML1). Methods The hyperlipidemia rat model was constructed. MiR-185-3p and MAML1 levels in hyperlipidemia rats were detected. Adenoviral vectors altering miR-185-3p and MAML1 levels were injected into hyperlipidemia rats to examine the levels of biochemical indices, inflammatory factors, oxidative stress, lipid accumulation and cellular morphology in liver tissues of hyperlipidemia rats. The targeting relation between miR-185-3p and MAML1 was manifested. Results MiR-185-3p levels were depleted while MAML1 expression was elevated in HH rats. MiR-185-3p overexpression or MAML1 silencing reduced levels of inflammatory factors in serum, mitigated oxidative stress and biochemical response, relieved lipid accumulation and cellular morphology in liver tissues; while up-regulated MAML1 reversed the effects of augmented MAML1 in hyperlipidemia rats. MiR-185-3p targeted MAML1. Conclusion Up-regulated miR-185-3p represses hyperlipidemia development via modulating MAML1. This research provides novel therapeutic candidates for the treatment of hyperlipidemia.

Macrophage Depletion via Clodronate Pretreatment Reduces Transgene Expression from AAV Vectors In Vivo

Adeno-associated virus is a popular gene delivery vehicle for gene therapy studies. A potential roadblock to widespread clinical adoption is the high vector doses required for efficient transduction in vivo, and the potential for subsequent immune responses that may limit prolonged transgene expression. We hypothesized that the depletion of macrophages via systemic delivery of liposome-encapsulated clodronate would improve transgene expression if given prior to systemic AAV vector administration, as has been shown to be the case with adenoviral vectors. Contrary to our expectations, clodronate liposome pretreatment resulted in significantly reduced transgene expression in the liver and heart, but permitted moderate transduction of the white pulp of the spleen. There was a remarkable localization of transgene expression from the red pulp to the center of the white pulp in clodronate-treated mice compared to untreated mice. Similarly, a greater proportion of transgene expression could be observed in the medulla located in the center of the lymph node in mice treated with clodronate-containing liposomes as compared to untreated mice where transgene expression was localized primarily to the cortex. These results underscore the highly significant role that the immune system plays in influencing the distribution and relative numbers of transduced cells in the context of AAV-mediated gene delivery.

COVID-19 Vaccines: Adenoviral Vectors

The worldwide pandemic of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has led to the unprecedented pace of development of multiple vaccines. This review evaluates how adenovirus (Ad) vector platforms have been leveraged in response to this pandemic. Ad vectors have been used in the past for vaccines against other viruses, most notably HIV and Ebola, but they never have been produced, distributed, or administered to humans at such a large scale. Several different serotypes of Ads encoding SARS-CoV-2 Spike have been tested and found to be efficacious against COVID-19. As vaccine rollouts continue and the number of people receiving these vaccines increases, we will continue to learn about this vaccine platform for COVID-19 prevention and control. Expected final online publication date for the Annual Review of Medicine, Volume 73 is January 2022. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

Concepts in Oncolytic Adenovirus Therapy

Oncolytic adenovirus therapy is gaining importance as a novel treatment option for the management of various cancers. Different concepts of modification within the adenovirus vector have been identified that define the mode of action against and the interaction with the tumour. Adenoviral vectors allow for genetic manipulations that restrict tumour specificity and also the expression of specific transgenes in order to support the anti-tumour effect. Additionally, replication of the virus and reinfection of neighbouring tumour cells amplify the therapeutic effect. Another important aspect in oncolytic adenovirus therapy is the virus induced cell death which is a process that activates the immune system against the tumour. This review describes which elements in adenovirus vectors have been identified for modification not only to utilize oncolytic adenovirus vectors into conditionally replicating adenoviruses (CRAds) that allow replication specifically in tumour cells but also to confer specific characteristics to these viruses. These advances in development resulted in clinical trials that are summarized based on the conceptual design.

Human Adenovirus Serotypes Efficiently Transducing HEK293 Cells: An In Vitro Propagation of HAdv

Generally, a gene which is inserted directly into a cell does not operate independently. Instead, the transmission of the gene is genetically modified by a biological messenger called a vector, consists of a transgene and a large DNA sequence as a backbone. Since they can deliver the new gene by infecting the cell, such viruses are also used as vectors. The adenovirus is a non-enveloped virus that can be tailored to transfer DNA to target cells, and it has sparked a lot of interest in the field, particularly in clinical trial therapy techniques. For the new age production of COVID-19 vaccine, development of different mammalian cell lines like HEK293 (most reliable growth and prosperity for transfection) and recombinant adenoviral vectors have become the first priority for biopharmaceutical giants and globally approved vaccine manufacturers to scale up their vaccine production. Adenoviruses have an icosahedral shape, with a protein coat encasing the viral double-stranded DNA genome. Because the adenovirus genome is relatively small, it's a good candidate for insertion of foreign DNA. The adenovirus E1A gene is deleted, and the virus loses its capacity to replicate. This ability can be restored during cell culture propagation by employing cells that produce the E1A protein, for example. Hence, in this mini research, I have shared an overview of the propagation of adenoviral vectors, i.e. recombinant adenovirus SARS CoV-2 vector in HEK-293 cell suspension culture.

Human AdV-20-42-42, a promising novel adenoviral vector for gene therapy and vaccine product development

Pre-existing immune responses towards adenoviral vector limit the use of a vector based on particular serotypes and its clinical applicability for gene therapy and/or vaccination. Therefore, there is a significant interest to vectorize novel adenoviral types that have low seroprevalence in the human population. Here, we describe the discovery and vectorization of a chimeric human adenovirus, which we call HAdV-20-42-42. Full genome sequencing revealed that this virus is closely related to human serotype 42, except for the penton-base which is derived from serotype 20. The HAdV-20-42-42 vector could be propagated stably to high titers on existing E1-complementing packaging cell lines. Receptor binding studies revealed that the vector utilized both CAR and CD46 as receptors for cell entry. Furthermore, the HAdV-20-42-42 vector was potent in transducing human and murine cardiovascular cells and tissues, irrespective of the presence of blood coagulation factor X. In vivo characterizations demonstrate that when delivered intravenously (i.v.) in mice, HAdV-20-42-42 mainly targeted the lungs, liver and spleen and triggered robust inflammatory immune response. Finally, we demonstrate that potent T-cell responses against vector-delivered antigens could be induced upon intramuscular vaccination in mice. In summary, from the data obtained we conclude that HAdV-20-42-42 provides a valuable addition to the portfolio of adenoviral vectors available to develop efficacious products in the fields of gene therapy and vaccination. IMPORTANCE Adenoviral vectors are currently under investigation for a broad range of therapeutic indications in diverse fields, such as oncology and gene therapy, as well as for vaccination both for human and veterinary use. A wealth of data shows that pre-existing immune responses may limit the use of a vector. Particularly in the current climate of global pandemic, there is a need to expand the toolbox with novel adenoviral vectors for vaccine development. Our data demonstrates that we have successfully vectorized a novel adenovirus type candidate with low seroprevalence. The cell transduction data and antigen-specific immune responses induced in vivo demonstrate that this vector is highly promising for the development of gene therapy and vaccine products.

Inhibition of Arginyl-tRNA Synthetase Promotes the Protection of Neuronal Ischemic Tolerance in Vitro Author Imformation

BackgroundsStudies have shown that the metabolic rate of mammals decreases during hibernation, suggesting that effective reduction of the energy consumption of ischemic cells may be the basis of the protective effect of ischemic tolerance. Anderson et al. reported after inhibition of the gene encoding arginyl-tRNA synthetase, the protein translation and mortality in C. elegans under anoxic conditions decreased significantly.PurposesWhether inhibition of arginyl-tRNA synthetase (RARS), in addition to combating hypoxic injury in C. elegans, protects rat neurons from ischemic damage remains unknown. The aim of this study is to determine whether knockdown of arginyl-tRNA synthetase improves the tolerance of primary cultured rat neurons to ischemic anoxia. Methods For the primary neuronal cutlure, cerebral cortex tissues were collected from newborn 24 hours Sprague-Dawley rats. Different viral vectors were transfected into cultured primary neurons, and the optimal viral vector and time points for gene silencing were determined by detecting the expression of RARS gene and protein. The adenovirus vector expressing shRNA-RARS with the highest silencing efficiency was transfected into rat primary cultured cortical neurons. The time point at which the viral vector exhibits optimal gene silencing efficiency was selected as the detection time point for subsequent experiments. The neurons after OGD treatment were divided into 2 groups: the experimental group and the control group. Each group was divided into 3 subgroups: the normal group, the control shRNA group and the shRNA-RARS group. After 3 hours of OGD treatment, cell survival, ATP levels and RARS protein expression were evaluated , and the data was analyzed. ResultsPrimary cultured neurons were identified and purified. The adenoviral vector expressing RARS-RNAi (27394-1) exhibited the highest silencing efficiency of the three adenoviral vectors, and 3 days after transfection of the adenoviral vector was selected as the optimal time point for gene silencing. Transfection of adenoviral vectors expressing shRNA-RARS reduced RARS protein expression, ATP consumption and neuronal death, and increased CCK8 activity in neurons after OGD insult.Conclusions Our work revealed neurons transfected with adenoviral vectors expressing shRNA-RARS exhibited stronger tolerance to ischemia and hypoxia, which was due to the inhibition of RARS activity andreduced cell energy metabolism rate. These results suggested that RARS inhibition reduced protein translation and energy consumption, and played a protective role in ischemic tolerance.