BIOSCIENCE BIOTECHNOLOGY AND BIOCHEMISTRY
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2024
·
IF 1.4
Wu Tao,Wu Ji,Liu Li,Song Hongping
Lipopolysaccharide (LPS) causes inflammatory cholestasis in sepsis. We investigated the role of PDZK1 in the repression of ABC transporters in LPS-induced cholestasis. Lentiviral gene transfer of PDZK1 to rats was conducted to explore its influence on cholestasis induced by LPS. And the effect of lentivirus-mediated shRNA targeting PDZK1 on ABC transporters in rat liver BRL-3A cells was evaluated. Lentiviral vector encoding rat PDZK1 was administered to rats by tail intravenous injection. Obviously elevated serum total bile acid (TBA) level and liver biochemical markers in cholestatic rats were decreased by the Lv-PDZK1 delivery. Also Lv-PDZK1 delivery stimulated the suppressed hepatic ABC transporters expression. In vitro, after the lentiviral vector LV3/PDZK1 shRNA transfection, no expression of PDZK1 and mild expression of ABC transporters were detected in BRL-3A cells by Western blotting. Our results indicate that the lentiviral-mediated hepatocyte PDZK1 expression ameliorates LPS-induced cholestasis in rats by rescuing hepatic ABC transporters expression.
研究领域:
分子生物学
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肝病学
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基因治疗
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产品关联
DOI
PubMed
高影响因子
CANCER RESEARCH
·
2025
·
IF 16.6
Shang Yan,Li Shuaihu,Xu Tian,Sun Yihua,Li Fei
The CRISPR/Cas9 lentiviral system has emerged as a powerful tool for gene knockout in cancer immunology research. However, the persistent expression of exogenous elements, such as Cas9 and resistance markers, often triggers excessive tumor immune rejection. This can lead to prolonged experimental timelines, increased data variability, biased outcomes, and even experimental failures. To address this challenge, several strategies have been investigated that offer partial solutions, but they have failed so far to comprehensively resolve the issue. In response, we developed the VL-AdCre system, a strategy that enables efficient excision of exogenous expression elements following gene knockout. The VL-AdCre system effectively reduced tumor immune rejection in allograft models, streamlined experimental workflows, and improved the reliability of research outcomes. Overall, this optimized CRISPR/Cas9 lentiviral gene knockout system offers a robust and practical solution for studying gene functions in vivo and advancing immunotherapeutic strategies.
研究领域:
免疫学
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肿瘤学
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分子生物学
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基因编辑
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转化医学
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产品关联
DOI
PubMed
高影响因子
INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
·
2021
·
IF 6.953
Yundi Duan,Nan Jiang,Jing Chen,Jianhua Chen
A high serum uric acid (SUA) concentration is associated with hyperuricemia (HUA) and gout. In order to obtain long-acting therapeutic effect, correction of purine metabolism at genetic level is advantageous. For this purpose, we expressed three “human-like” urate oxidases in human hepatocytes (HL-7702) by lentivirus-mediated transduction. Enzymatic assay revealed that the recombinant urate oxidases expressed in HL-7702 cells were functionally active. Electron microscopy study showed that the recombinant enzymes were localized to peroxisome and formed distinct crystalloid core structures as in other mammal cells. Although similar rate of uric acid degradation was observed for all recombinant urate oxidases, HL-7702-pLVX-UOX 83 cells and HL-7702-pLVX-UOX 214/217 cells retained more cell viability compared with HL-7702-pLVX-UOX PBC at high uric acid level. This study provides a new direction for the treatment of gout and hyperuricemia.
研究领域:
遗传学
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代谢性疾病
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分子生物学
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治疗学
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产品关联
DOI
PubMed
高影响因子
EXPERIMENTAL ANIMALS
·
2022
·
IF 1.798
Ning HAN,Ying SU,Mi GUO,Li YU
Diabetic retinopathy (DR) is the prevalent microvascular complication of diabetes mellitus (DM), and it may lead to permanent blindness. The previous publication has indicated that both inflammatory response and oxidative stress are critical factors involved in DR progression, however, the accurate regulatory mechanism remains to be revealed. Src homology region 2 (SH2)-containing protein tyrosine phosphatase 2 (SHP2), a member of the protein tyrosine phosphatase family, was reported to play a role in diabetic nephropathy, whereas its function in DR was unknown and required further exploration. The level of phosphorylated, not the total, SHP2 increased in the retinas of rats with streptozotocin injection-induced DM. Further, the intravitreal injection of SHP2 shRNA lentivirus alleviated retinal pathological changes, and inhibited inflammatory response and oxidative stress, which were accompanied with Yes-associated protein 1 (YAP1) deactivation in DR rats. Additional co-immunoprecipitation results confirmed the interaction of SHP2 and YAP1. Collectively, our data preliminarily show that DR amelioration-induced by SHP2 inhibition in rats may attribute to the deactivation of YAP1 pathway.
研究领域:
眼科学
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分子生物学
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糖尿病研究
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细胞信号传导
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产品关联
DOI
PubMed
高影响因子
BIOCHEMICAL PHARMACOLOGY
·
2021
·
IF 5.858
Xiaodan Tan,Wenjia Guo,Zhe Peng,Chao Gu,Pu Xiang,Yujun Tu,Huizhi Fei,Xia Liu,Yi Lu,Miaomiao Li,Hong Wang,Ying Luo,Junqing Yang
Based on the previous studies, this study was carried out to explore the interaction of LncRNA-Malat1/miR-211-5p in cerebral ischemia reperfusion injury (CIRI). Firstly, the expression changes of LncRNA-MALAT1 and miR-211-5p in ischemia patients’ blood were determined, and the binding sites of them were predicted by bioinformatics. Furthermore, middle cerebral artery occlusion/reperfusion (MCAO/R) injury model was established in adult male SD rats , and primary neuronal oxygen-glucose deprivation/reoxygen-glucose reoxygenation (OGD/R) was established in vitro. The results showed that LncRNA-MALAT1 was significantly up-regulated and miR-211-5p down-regulated in the peripheral blood of patients with ischemic stroke, and the expression changes were negatively correlated. Bioinformatics prediction results showed that LncRNA-MALAT1 had a binding site with miR-211-5p. We also found that LncRNA-Malat1 was significantly up-regulated while miR-211-5p down-regulated in rat cortex tissue and primary neurons treated with OGD/R. In addition, lentivirus interfered with LV-Malat1-RNAi decreased the expression of LncRNA-Malat1 and promoted the up-regulation of miR-211-5p. Combination of LV-Malat1-RNAi and miR-211-5p inhibitor significantly reversed the protective effect of down-regulation of LncRNA-Malat1. Inhibition of LncRNA-Malat1 expression alleviated the neurological deficit score after MCAO/R, improved histopathological damage, and reduced the size of cerebral infarction . Combined administration of LV-Malat1-RNAi + Antagomir-211-5p reversed these effects above. In short, our data suggest that LncRNA-Malat is involved in the occurrence and development of cerebral ischemia reperfusion injury by acting on miR-211-5p and is then regulating the expression of COX-2.
研究领域:
神经科学
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分子生物学
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医学遗传学
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DOI
PubMed
高影响因子
BIOTECHNOLOGY LETTERS
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2024
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IF 2
Yao Yongchang,Chen Ke,Pan Qian,Gao Hui,Su Weixian,Zheng Shicong,Dong Weiqiang,Qian Dongyang
Objectives We genetically modified dedifferentiated chondrocytes (DCs) using lentiviral vectors and adenoviral vectors encoding TGF-β3 (referred to as transgenic groups below) and encapsulated these DCs in the microcavitary hydrogel and investigated the combinational effect on redifferentiation of the genetically manipulated DCs. Results The Cell Counting Kit-8 data indicated that both transgenic groups exhibited significantly higher cell viability in the first week but inferior cell viability in the subsequent timepoints compared with those of the control group. Real-time polymerase chain reaction and western blot analysis results demonstrated that both transgenic groups had a better effect on redifferentiation to some extent, as evidenced by higher expression levels of chondrogenic genes, suggesting the validity of combination with transgenic DCs and the microcavitary hydrogel on redifferentiation. Although transgenic DCs with adenoviral vectors presented a superior extent of redifferentiation, they also expressed greater levels of the hypertrophic gene type X collagen. It is still worth further exploring how to deliver TGF-β3 more efficiently and optimizing the appropriate parameters, including concentration and duration. Conclusions The results demonstrated the better redifferentiation effect of DCs with the combinational use of transgenic TGF-β3 and a microcavitary alginate hydrogel and implied that DCs would be alternative seed cells for cartilage tissue engineering due to their easily achieved sufficient cell amounts through multiple passages and great potential to redifferentiate to produce cartilaginous extracellular matrix.
研究领域:
生物医学工程
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组织工程
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细胞生物学
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分子生物学
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产品关联
DOI
PubMed
高影响因子
Frontiers in Immunology
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2023
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IF 7.3
Xi Zhang,Tiaoxia Wang,Xiaona Zhu,Yong Lu,Mingpeng Li,Zhihong Huang,Deping Han,Longzhen Zhang,Yang Wu,Liantao Li,Frank Klawonn,Renata Stripecke
Introduction Epstein-Barr virus (EBV) is a widely spread pathogen associated with lymphoproliferative diseases, B/ T/ NK cell lymphomas, nasopharyngeal carcinoma (NPC) and gastric carcinoma (GC). EBV lytic reactivations contribute to the genomic instability, inflammation and tumorigenesis of NPC, promoting cancer progression. Patients with NPC refractory to standard therapies show dismal survival. EBV gp350 is an envelope protein detectable in NPC specimens intracellularly and on the cell membrane of malignant cells, and is a potential viral antigen for T cell-directed immunotherapies. The potency of T cells engineered with a chimeric antigen receptor (CAR) targeting gp350 against EBV + lymphoproliferative disease was previously shown. Methods Here, we advanced towards preclinical and non-clinical developments of this virus-specific CAR-T cell immunotherapy against NPC. Different gp350CAR designs were inserted into a lentiviral vector (LV) backbone. Results A construct expressing the scFv 7A1-anti−gp350 incorporating the CD8 transmembrane and CD28.CD3ζ signaling domain (ZT002) was selected. High titer ZT002 (~1x10 8 TU/ml) was manufactured in HEK 293T/17 suspension cells in serum free media as large-scale production under good manufacturing practices (GMP). A LV multiplicity of infection (MOI) of 1 resulted in high frequencies of functional gp350CAR + T cells (>70%) at a low (<2) vector copy numbers in the genome. ZT002 was therefore used to establish gp350CAR-T batch run production methods. GMP upscaling and validation of T cell transduction and expansion in several runs resulted in average 3x10 9 gp350CAR-T cells per batch. >80% CD3 + gp350CAR-T cells bound to purified gp350 protein. In vitro cytotoxicity and cytokine secretion assays (IFN-γ and TNF-α) confirmed the specificity of gp350CAR-T cells against gp350 + NPC, GC and lymphoma cell targets. Immunocompromised B-NDG mice (NOD.CB17- PrkdcscidIl2rgtm1 /Bcgen) were challenged s.c. with a EBV + NPC C666.1 cell line expressing gp350 and then treated with escalating doses of gp350CAR-T cells or with non-transduced T cells. gp350CAR-T cells promoted antitumor responses, bio-distributed in several tissues, infiltrated in tumors and rejected gp350 + tumor cells. Discussion These results support the use of gp350CAR-T cells generated with ZT002 as an Innovative New Drug to treat patients with solid and liquid EBV-associated malignancies.
研究领域:
免疫治疗
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肿瘤学
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分子生物学
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病毒学
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临床前研究
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产品关联
DOI
PubMed
高影响因子
EXPERIMENTAL CELL RESEARCH
·
2024
·
IF 3.3
Yanhua Jin,Huan Wang,Yu Xin,Yanning Zhang
Mitochondrial dysfunction is a pivotal factor in the onset of podocyte damage, which is a central component in the pathogenesis of nephrotic syndrome (NS). However, the precise mechanisms underlying the changes in podocyte mitochondria remain elusive. Our study aims to clarify the potential mechanisms involved in the role of F-box protein 11 (Fbxo11) in NS, specifically concentrating on its impact on mitochondrial function. A mouse model was established by tail vein injection of adriamycin (ADR, 10 mg/kg) and was infected with lentivirus overexpressing Fbxo11. Mouse podocytes (MPC-5) were infected with lenti-Fbxo11-OE, followed by treatment with 0.4 μg/mL of ADR. We identified the decreased expression of Fbxo11 in mouse kidney tissues and MPC-5 cells induced by ADR. Lenti-Fbxo11-OE intervention relieved ADR-induced glomerular lesion, podocyte injury, and mitochondrial dysfunction. In vitro, overexpression of Fbxo11 in mouse podocytes improved mitochondrial function and reduced podocyte damage, thereby inhibiting podocyte apoptosis. Mechanistically, Fbxo11 decreased the protein expression of Fosl2 through ubiquitin-dependent proteasomal degradation. Rescue experiments revealed that overexpression of Fosl2 abolished the protective effects of Fbxo11 overexpression on mitochondrial damage and podocyte injury. Importantly, the regulatory effects of the Fbxo11/Fosl2 axis were reversed when treated with the mitochondrial fission inhibitor mdivi-1. Taken together, our results demonstrated that Fbxo11-mediated ubiquitin degradation of Fosl2 is critical for maintaining mitochondrial function and preventing podocyte injury during NS.
研究领域:
肾病学
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分子生物学
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细胞生物学
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DOI
PubMed
高影响因子
Frontiers in Aging Neuroscience
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2020
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IF 4.364
Feng Chenxi,Bao Xiaoming,Shan Ling,Ling Yunxiang,Ding Yanfei,Wang Jia,Cao Yanzi,Wang Qinwen,Cui Wei,Xu Shujun
Calcium-sensing receptor (CaSR) is a G protein-coupled receptor (GPCRs). Soluble β-amyloid peptide (Aβ) is one of the orthosteric modulators of CaSR, while, the role and underlying mechanism of CaSR in cognitive decline in Alzheimer’s disease (AD) is unclear. In this study, molecular technology such as live-cell imaging combined with behavioral tests were used to explore the role and the underlying mechanism of CaSR in the cognitive deficits in AD mice. The expression levels of CaSR were increased both in AD mice and Aβ1–42 (β-amyloid protein)-treated primary cultured neurons. Pharmacological inhibition of CaSR ameliorated recognitive and spatial memory deficits of Aβ1–42-oligomer-treated mice in a dose-dependent manner. Pharmacological inhibition of CaSR or down-regulation of the expression of CaSR by CaSR-shRNA-lentivirus prevented the impairment of filopodia, and the synapse induced by oligomeric Aβ1–42. The contents of cytosolic phospholipase A2 (cPLA2) and prostaglandin E2 (PGE2) in hippocampal neurons and tissue were increased after treatment with Aβ1–42 oligomers. Inhibition or down-regulation of CaSR mediates Aβ-induced synapse formation and cognitive deficits partially, through the activation of the cPLA2/PGE2 pathway. This study provides novel insights on CaSR, which is a promising therapeutic target for AD.
研究领域:
神经科学
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药理学
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分子生物学
·
神经退行性疾病
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DOI
PubMed
高影响因子
Molecular Medicine Reports
·
2017
·Vol. 15
·期 5
·3048-3054
·
IF 1.692
Wenting Zhao,Zhen Wang,Zewei Sun,Shuai Wang,Mingjie Wu,Liangrong Zheng
Elevated blood glucose levels contribute to a series of complications in patients with diabetes mellitus, including chronic ulcers and accelerated atherosclerosis. Dysregulated endothelial migration induced by high glucose is important in vascular‑associated complications. In the present study, cluster of differentiation (CD)97/adhesion G protein‑coupled receptor 5 (ADGRE5), a member of the G protein‑coupled receptor protein family, stimulated angiogenesis was investigated to determine its role in cell models of diabetes mellitus using lentivirus‑mediated overexpression and siRNA transfection. The results revealed reduced expression in high glucose‑treated human umbilical vein endothelial cells and in the endothelium of diabetic mice. Among the three CD97 isoforms, the majority of the expression of CD97 (EGF1,2,5) in the endothelial cells was regulated by high levels of glucose. Using stable lentivirus‑mediated transfection and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR‑associated protein 9 (Cas9) technology, the present study constructed CD97‑overexpression and CD97‑knockout endothelial cell lines. Migration assays showed that the lentivirus‑mediated overexpression of CD97/ADGRE5 improved the inhibition of high glucose‑induced endothelial cell migration. In addition, using cytoskeleton staining, it was found that CD97 promoted membrane ruffling and lamellipodia formation. Cell division cycle 42, a small GTP‑binding protein, and its downstream factor, actin‑related protein 2, were involved in CD97‑induced actin reorganization in endothelial cells. Additionally, the use of transcription factor filter plate assays revealed that the nuclear translocation of signal transducer and activator of transcription 1 stimulated by high glucose contributed to the inhibited transcription of CD97. In conclusion, the present study established that the overexpression of CD97 improved high glucose‑induced dysfunction of endothelial cell migration. These findings provide insight to assist in identifying therapeutic targets with potential to ameliorate certain vascular complications of diabetes.
研究领域:
分子生物学
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细胞生物学
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糖尿病研究
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血管生物学
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产品关联
DOI
PubMed
高影响因子
ARCHIVES OF ORAL BIOLOGY
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2021
·
IF 2.635
Yujia Cui,Mingru Bai,Daimo Guo,Yueyi Yang,Haoran Chen,Jianxun Sun,Jing Xie,Xuedong Zhou
Objective Insulin-like growth factor 1 (IGF1) is one of the vital factors in regenerative endodontics . Previous studies have focused on the role of IGF1 in the mineralization of dental tissues. However, the role of IGF1 in the neural differentiation of dental stem cells was little discussed. Design IGF1 was overexpressed in human stem cells from the apical papilla (hSCAPs) by lentivirus and knocked down in hSCAPs by small interfering RNA . The neural differentiation level of hSCAPs was investigated histologically by HE staining and Nissl staining after neural induction for 3 days. The expression of proteins was examined by western blot and immunofluorescence. Results IGF1 promoted neural differentiation of hSCAPs, more cell processes and Nissl-positive body stained cells. IGF1 overexpression could both promote glial differentiation in hSCAPs, characterized by the increase of S100β and GFAP proteins, and neuronal differentiation , characterized by the increase of βIII-tubulin and functional GAD67/vGLUT1 proteins. Conversely, IGF1 knockdown suppressed both glial and neuronal differentiation. IGF1 activated AKT to regulate the early neural differentiation of hSCAPs. Conclusions The results indicate IGF1 could promote neural differentiation of hSCAPs by activating AKT signaling and provide a cue for the candidate of induced neural seeding cells in regenerative endodontics.
研究领域:
牙科学
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干细胞生物学
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神经科学
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再生医学
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产品关联
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）