Double transduction of a Cre/LoxP lentiviral vector: a simple method to generate kidney cell-specific knockdown mice

Nam, Bo Young; Kim, Yon Su; Park, Sun-Hee; Kang, Hye Young; Paeng, Jisun; Kim, Seong-Hun; Park, Jimin; Um, Jae Eun; Oh, Hyung Jung; Han, Seung Hyeok; Yoo, Tae‐Hyun; Kang, Shin Wook

doi:10.1152/ajprenal.00251.2015

Cited by 13 publications

(8 citation statements)

References 43 publications

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“…Short-hairpin RNA (shRNA) expression for Nrf2 or the scrambled sequence was produced according to published protocols (Naito and Ui-Tei, 2013). Lentivirus samples were set up at a titer of 8 × 10 8 transfection units (TU)/mL and stored at −80°C for the following experiments (Nam et al, 2015). To knock down Nrf2 in vivo , C57 mice were randomly divided into four groups and injected with corresponding lentivirus shRNA or scrambled sequences via tail vein injection at the age of 6 weeks.…”

Section: Methodsmentioning

confidence: 99%

“…To knock down Nrf2 in vivo , C57 mice were randomly divided into four groups and injected with corresponding lentivirus shRNA or scrambled sequences via tail vein injection at the age of 6 weeks. Lentivirus was injected at 3-day intervals for three consecutive injections (Nam et al, 2015). To maintain effective expression of shRNA, the injections were repeated every 4 weeks.…”

Section: Methodsmentioning

confidence: 99%

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Myricetin Attenuated Diabetes-Associated Kidney Injuries and Dysfunction via Regulating Nuclear Factor (Erythroid Derived 2)-Like 2 and Nuclear Factor-κB Signaling

Yang

Wang

et al. 2019

Front. Pharmacol.

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Background/Aims: Previous studies have suggested that myricetin (Myr) could promote the expression and nuclear translocation of nuclear factor (erythroid-derived 2)-like (Nrf2). This study aimed to investigate whether Myr could attenuate diabetes-associated kidney injuries and dysfunction in wild-type (WT) and Nrf2 knockdown (Nrf2-KD) mice. Methods: Lentivirus-mediated Nrf2-KD and WT mice were used to establish type 1 diabetes mellitus (DM) by streptozotocin (STZ) injection. WT and Nrf2-KD mice were then randomly allocated into four groups: control (CON), Myr, STZ, and STZ + Myr. Myr (100 mg/kg/day) or vehicle was administered for 6 months. Kidneys were harvested and weighed at the end of the experiment. Hematoxylin and eosin staining and Masson’s trichrome staining were used to assess the morphology and fibrosis of the kidneys, respectively. Urinary albumin-to-creatinine ratio was used to test renal function. Western blotting was performed to determine oxidative-stress- or inflammation-associated signaling pathways. Real-time polymerase chain reaction (RT-PCR) was performed to detect the expression of fibrosis or inflammatory cytokines at the message Ribonucleic Acid (mRNA) level. Results: In WT mice, Myr alleviated DM-induced renal dysfunction, fibrosis, and oxidative damage and enhanced the expression of Nrf2 and its downstream genes. After knockdown of Nrf2, Myr treatment partially but significantly mitigated DM-induced renal dysfunction and fibrosis, which might be associated with inhibition of the I-kappa-B (IκB)/nuclear factor-κB (NF-κB) (P65) signaling pathway. Conclusions: This study showed that Myr prevented DM-associated decreased expression of Nrf2 and inhibited IκB/NF-κB (P65) signaling pathway. Moreover, inhibition of IκB/NF-κB (P65) signaling pathway is independent of the regulation of Nrf2. Thus, Myr could be a potential treatment for preventing the development and progression of DM-associated kidney injuries and dysfunction.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Myricetin Attenuated Diabetes-Associated Kidney Injuries and Dysfunction via Regulating Nuclear Factor (Erythroid Derived 2)-Like 2 and Nuclear Factor-κB Signaling

Yang

Wang

et al. 2019

Front. Pharmacol.

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“…After the reaction of PCR, the temperature was increased to 95°C at a rate of 2°C/minute to make a melting curve. 27 The ratio of mtDNA/nuclear DNA was calculated as 2 ΔCT , where ΔCT is CT LDL − CT ND1 . Intra-assay and inter-assay coefficients of variation of mtDNA copy number were 4.7% (range 1.8–6.7) and 5.9% (range 3.7–9.1), respectively.…”

Section: Methodsmentioning

confidence: 99%

Low Mitochondrial DNA Copy Number is Associated With Adverse Clinical Outcomes in Peritoneal Dialysis Patients

et al. 2016

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“…To downregulate expression of the IL4 receptor α-chain (IL4Rα) in hippocampal microglia, a lentivirus encoding a short hairpin RNA (shRNA) against the receptor was constructed as described 29 . The lentivirus contained the cassette u6-LoxP-stop-LoxP-shRNA-IL4Rα.…”

Section: Expression Systems and Stereotactic Injectionmentioning

confidence: 99%

IL4-driven microglia modulate stress resilience through BDNF-dependent neurogenesis

Zhang

Rong

Zhang

et al. 2020

Preprint

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Running title: IL4-driven microglia modulate neurogenesis and stress resilience In Brief:Zhang et al. show that IL4-induced Arg1 + microglia restore hippocampal neurogenesis and promote resilience against stress in mice by increasing BDNF levels. Targeting microglia with immunomodulatory factors may be a strategy for treating mood disorders.2 Highlights:1. Vulnerability to stress in mice is associated with reduced IL4 signaling in the hippocampus 2. Brain-derived IL4 promotes adult hippocampal neurogenesis and stress resistance by driving Arg1 + microglia 3. IL4-driven Arg1 + microglia enhance hippocampal neurogenesis via a BDNF-dependent pathway 3 Abstract Adult neurogenesis in the dentate gyrus of the hippocampus is regulated by specific groups of microglia and is functionally implicated in behavioral responses to stress. However, the role of microglia in modulating hippocampal neurogenesis in stress responses remains poorly understood. Here we investigated the effects of IL4-driven Arg1 + microglia in the restoration of hippocampal neurogenesis and conferment of stress resilience. We found that low IL4 levels in the hippocampus of mice was associated with greater stress vulnerability and, conversely, overexpression of IL4 in the hippocampus induced a large number of Arg1 + microglia and ameliorated stress-induced depressive-like behaviors. Knockdown of microglial IL4 receptors in the hippocampus of mice exacerbated the stress-induced inflammatory response and abolished the antidepressant effects of IL4 overexpression. Enhancement or inhibition of IL4 signaling in hippocampal microglia modulated neurogenesis, and blockade of neurogenesis abolished the resilience to stress-induced depression. We further show that IL4-activated microglia is associated with upregulation of BDNF levels and neurogenesis. Taken together, our findings suggest that IL4-driven microglia in the hippocampus trigger BDNF-dependent neurogenesis in response to chronic stress, helping protect against depressive-like symptoms. These findings identify the modulation of a specific microglial phenotype as a treatment strategy for mood disorders.

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Double transduction of a Cre/LoxP lentiviral vector: a simple method to generate kidney cell-specific knockdown mice

Cited by 13 publications

References 43 publications

Myricetin Attenuated Diabetes-Associated Kidney Injuries and Dysfunction via Regulating Nuclear Factor (Erythroid Derived 2)-Like 2 and Nuclear Factor-κB Signaling

Myricetin Attenuated Diabetes-Associated Kidney Injuries and Dysfunction via Regulating Nuclear Factor (Erythroid Derived 2)-Like 2 and Nuclear Factor-κB Signaling

Low Mitochondrial DNA Copy Number is Associated With Adverse Clinical Outcomes in Peritoneal Dialysis Patients

IL4-driven microglia modulate stress resilience through BDNF-dependent neurogenesis

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