Lentiviral vectors have been proven to be a powerful tool in gene therapies that includes the ability to perform long‐term gene editing in both dividing and non‐dividing cells. In order to meet the rising demand for clinical‐grade lentiviral vectors for future clinical trials and requirements by regulatory agencies, new methods and technologies were developed, including the rapid optimization of production and purification processes. However, gaps still exist in achieving ideal yields and recovery rates in large‐scale manufacturing process steps. The downstream purification process is a critical step required to obtain a sufficient quantity and high‐quality lentiviral vectors products, which is challenged by the low stability of the lentiviral vector particles and large production volumes associated with the manufacturing process. This review summarizes the most recent and promising technologies and enhancements used in the large‐scale purification process step of lentiviral vector manufacturing and aims to provide a significant contribution towards the achievement of providing sufficient quantity and quality of lentiviral vectors in scalable processes.
Objective To observe the effects of tissue factor pathway inhibitor(TFPI) on thrombosis formation in rabbit carotid arteries after ballon injury. Methods Fouty rabbits with the weight 2.5-3.0 kg were respectively divided into 4 groups, Ad-TFPI, Ad-LacZ, PBS and normal control groups. The normal control group was not given any treatment and other 3 groups were given 0.2 ml Ad-TFPI, Ad-LacZ or PBS reproduced by the Dispatch catheter respectively after the PTCA balloon iniury on the right carotid arteries. Ten days after gene transfer the repeated balloon injury was performed in the 3 groups, and the first balloon injury was performed in the normal control group by the same method. The carotid blood flow was recovered immediately after the injury. Thirty minutes later all the animals were sacrificed. The injured carotid arteries and one part of contralateral normal artery were cut down, scissored along the long axis, flattened and fixed in the 2% glutaral. The platelet aggregation and thrombosis formation on the luminal surfaces was observed under electron microscope. Results The electron microscope results showed that the vascular endothelial cell structure was integrated and lined up in order in the nomal artery which had no any injury. After the balloon injury in the normal control group, the structure of the endothelial cell was disintegrated, and there was some platelet aggregation but no fibrosis formation. A large amount of platelet aggregated but no fibrosis formed in Ad-TFPI group after the repeated balloon injury. A large amount of fibrosis formed and red cells piled up in the Ad-LacZ and PBS group. The positive rate of thrombosis formation among groups had siginificant differences(χ2=14.95, P 0.05). The positive rate in Ad-LacZ group(80%) was higher than in the normal control group(10%, χ2=9.90, P 0.05). The positive rate in PBS group(70%) was higher than that in the normal control group(10%, χ2=7.50, P< 0.01). Conclusions The repeated balloon injury method can cause a large amount of fibrosis formation in the rabbit carotid. TFPI gene inhibits thrombosis formation in balloon-injured rabbit carotid arteries.
Key words:
Tissue factor pathway inhibitor; Gene therapy; Carotid artery injuries; Thrombosis formation; Microscopy; electron; scanning
Hyperlipidemia is a medical condition characterized by elevated levels of blood lipids, especially triglycerides (TG). However, it remains unclear whether TG levels remain consistently elevated throughout the entire developmental stage of the high-lipid state. In our animal experiment, we found that TG levels were significantly higher in the early stage of the high-lipid model but significantly decreased at the 14th week of the late stage, reaching levels similar to those of the control group. This suggests that TG levels in the high-lipid model are not always higher than those of the control group. To determine the reason for this observation, we used in situ mass spectrometry imaging (MSI) to detect the distribution of metabolites in the liver of rats. The metabolite distribution of the control rats at different stages was significantly different from that of the model rats, and the high-lipid model differed significantly from the control rats. We identified nine functional metabolites that showed differences throughout the period, namely, PA(20:3-OH/i-21:0), PA(20:4-OH/22:6), PG(20:5-OH/i-16:0), PG(22:6-2OH/i-13:0), PG(O-18:0/20:4), PGP(18:3-OH/i-12:0), PGP(PGJ2/i-15:0), SM(d18:0/18:1-2OH), and TG(14:0/14:0/16:0), among which TG was most significantly correlated with hyperlipidemia and high lipid. This study is unique in that it used MSI to reveal the changes in metabolites in situ, showing the distribution of different metabolites or the same metabolite in liver tissue. The findings highlight the importance of considering the animal’s age when using TG as a biomarker for hyperlipidemia. Additionally, the MSI images of the liver in the high-lipid model clearly indicated the distribution and differences of more significant metabolites, providing valuable data for further research into new biomarkers and mechanisms of hyperlipidemia. This new pathway of in situ, visualized, and data-rich metabolomics research provides a more comprehensive understanding of the characteristics of high lipid and its implications for disease prevention and treatment.
The design of electron transport layers (ETLs) is crucial to the performance of optoelectronic devices. A composite ETL was constructed to overcome the poor carrier extraction issue in perovskite solar cells, resulting in a maximum PCE of 19.14% with reduced hysteresis. A similar enhancement phenomenon was observed in both devices based on TiO2 and SnO2 ETLs.
Objective To study the myocardium lesion indu ced by repetitive virus infection in mice and the function of transforming growt h factor-beta1(TGF-β 1) in the remodeling of myocardial matrix collagen.Methods The chronic virus injury model was established successfully by repetitive inoculated Coxsackie virus B3m (CVB3m) th ree times, with each infection the dose of virus was increased. The mice were ki lled in the 74th days after the first infection. The pathology injury of myocard ium and the change of collagen system were analysed by the means of pathology, h istochemistry and supermicroscope,and the collagen volume fraction was counted. Serum TGF-beta1 content was detected by the method of ELISA.Results The heart function of mice was decreased after repetitive virus infection. Myocardial collagen volume fraction was higher than that of the control group(0.05). The proliferation and remodeling of the m atrix collagen were significant, which was characterized as scar healing and int erstitial fibrosis. The serum TGF-β 1content(0.103 5±1.270 0)μg/L in the repetitive virus infection group was higher than that of the control group(P 0.05).Conclusions The main influences of repetitive virus inf ection are infected mice cardiac collagen system remodeling. Repetitive virus in fection and persistent elevation of TGF-β 1 are the key factors in the elvolu tion from myocarditis to dilated cardiomyopathy.
Accumulating evidence suggests that hypoxia-inducible factor 1α (HIF-1α) regulates numerous miRNAs and is crucial for cellular response to hypoxia. However, the relationship between HIF-1α and miR-21 in hypoxic cardiomyocytes is little known. We found that hypoxia induced HIF-1α and miR-21 expression. HIF-1α knockdown increased cell apoptosis and reduced miR-21 expression. Furthermore, we found that HIF-1α transcriptionally enhanced miR-21 promoter activity by binding to its promoter, which required the recruitment of CBP/p300. In addition, we found that miR-21 inhibition increased cell apoptosis and reduced HIF-1α expression, and modulated the PTEN/Akt pathway. Our results indicate that HIF-1α-miR-21 feedback contributes to the adaptation of cardiomyocytes to hypoxia, and has potential as therapeutic target for myocardial ischemia.
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