p53/Drp1-dependent mitochondrial fission mediates aldosterone-induced podocyte injury and mitochondrial dysfunction

Mitochondrial dysfunction is increasingly recognized as an important factor in glomerular diseases. Previous study has shown that mitochondrial fission contributed to mitochondrial dysfunction. However, the mechanism of mitochondrial fission on mitochondrial dysfunction in aldosterone-induced podocyte injury remains ambiguous. This study aimed to investigate the pathogenic effect of mitochondrial fission both in vivo and in vitro. In an animal model of aldosterone-induced nephropathy, inhibition of the mitochondrial fission protein dynamin-related protein 1 (Drp1) suppressed aldosterone-induced podocyte injury. In cultured podocytes, aldosterone dose dependently induced Drp1 expression. Knockdown of Drp1 inhibited aldosterone-induced mitochondrial fission, mitochondrial dysfunction, and podocyte apoptosis. Furthermore, aldosterone dose dependently induced p53 expression. Knockdown of p53 inhibited aldosterone-induced Drp1 expression, mitochondrial dysfunction, and podocyte apoptosis. These findings implicated that aldosterone induced mitochondrial dysfunction and podocyte injury mediated by p53/Drp1-dependent mitochondrial fission, which may provide opportunities for therapeutic intervention for podocyte injury.

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Mitochondrial dysfunction is an early event in aldosterone-induced podocyte injury

AJP Renal Physiology ◽

10.1152/ajprenal.00570.2012 ◽

2013 ◽

Vol 305 (4) ◽

pp. F520-F531 ◽

Cited By ~ 42

Author(s):

Min Su ◽

Asish-Roopchand Dhoopun ◽

Yanggang Yuan ◽

Songming Huang ◽

Chunhua Zhu ◽

...

Keyword(s):

Mitochondrial Dysfunction ◽

Early Event ◽

Mtdna Copy Number ◽

Glomerular Diseases ◽

Podocyte Injury ◽

Atp Production ◽

Podocyte Damage ◽

Mitochondrial Transcription Factor

We previously showed that mitochondrial dysfunction (MtD) is involved in an aldosterone (Aldo)-induced podocyte injury. Here, the potential role of MtD in the initiation of podocyte damage was investigated. We detected the dynamic changes of urinary protein, urinary F2-isoprostane and renal malondialdehyde levels, kidney ultrastructure morphology, mitochondrial DNA (mtDNA) copy number, mitochondrial membrane potential (ΔΨm), and nephrin and podocin expressions in Aldo-infused mice. Aldo infusion first induced renal oxidative stress, as evidenced by increased levels of urinary F2-isoprostane and renal malondialdehyde, and MtD, as demonstrated by reduced mtDNA, ΔΨm, and ATP production. Later, at 5 days after Aldo infusion, proteinuria and podocyte injury began to appear. In cultured podocytes, Aldo or hydrogen peroxide (H2O2) induced MtD after 2–8 h of treatment, whereas the podocyte damage, as shown by decreased nephrin and podocin expressions, occurred later after 12 h of treatment. Thus Aldo treatment both in vitro and in vivo indicated that MtD occurred before podocyte damage. Additionally, MtDNA depletion by ethidium bromide or mitochondrial transcription factor A (TFAM) RNAi induced MtD, further promoting podocyte damage. TFAM expression was found to be reduced in Aldo-infused mice and Aldo-treated podocytes. Adenoviral vector-mediated overexpression of TFAM prevented Aldo-induced MtD and protected against podocyte injury. Together, these findings support MtD as an early event in podocyte injury, and manipulation of TFAM may be a novel strategy for treatment of glomerular diseases such as podocytopathy.

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Increased long noncoding RNA maternally expressed gene 3 contributes to podocyte injury induced by high glucose through regulation of mitochondrial fission

Cell Death and Disease ◽

10.1038/s41419-020-03022-7 ◽

2020 ◽

Vol 11 (9) ◽

Author(s):

Qiongxia Deng ◽

Ruowei Wen ◽

Sirui Liu ◽

Xiaoqiu Chen ◽

Shicong Song ◽

...

Keyword(s):

High Glucose ◽

Noncoding Rna ◽

Diabetic Kidney Disease ◽

Mitochondrial Fission ◽

Diabetic Mice ◽

Podocyte Injury ◽

Mitochondrial Translocation ◽

Maternally Expressed Gene 3

Abstract Excessive mitochondrial fission plays a key role in podocyte injury in diabetic kidney disease (DKD), and long noncoding RNAs (lncRNAs) are important in the development and progression of DKD. However, lncRNA regulation of mitochondrial fission in podocytes is poorly understood. Here, we studied lncRNA maternally expressed gene 3 (Meg3) in mitochondrial fission in vivo and in vitro using human podocytes and Meg3 podocyte-specific knockdown mice. Expression of lncRNA Meg3 in STZ-induced diabetic mice was higher, and correlated with the number of podocytes. Excessive mitochondrial fission of podocytes and renal histopathological and physiological parameters were improved in podocyte-specific Meg3 knockdown diabetic mice. Elongated mitochondria with attenuated podocyte damage, as well as mitochondrial translocation of dynamin-related protein 1 (Drp1), were decreased in Meg3 knockout podocytes. By contrast, increased fragmented mitochondria, podocyte injury, and Drp1 expression and phosphorylation were observed in lncRNA Meg3-overexpressing podocytes. Treatment with Mdivi1 significantly blunted more fragmented mitochondria and reduced podocyte injury in lncRNA Meg3-overexpressing podocytes. Finally, fragmented mitochondria and Drp1 mitochondrial translocation induced by high glucose were reduced following treatment with Mdivi1. Our data show that expression of Meg3 in podocytes in both human cells and diabetic mice was higher, which regulates mitochondrial fission and contributes to podocyte injury through increased Drp1 and its translocation to mitochondria.

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Abstract 269: Cardiac Specific Disruption of Drp-1 Causes the Development of Cardiac Dysfunction via Inhibition of Autophagy and Induction of Apoptosis

Circulation Research ◽

10.1161/res.113.suppl_1.a269 ◽

2013 ◽

Vol 113 (suppl_1) ◽

Author(s):

Yoshiyuki Ikeda ◽

Junichi Sadoshima

Keyword(s):

Mitochondrial Dysfunction ◽

Systolic Function ◽

Mitochondrial Fission ◽

Mitochondrial Mass ◽

Reduced Ejection Fraction ◽

Tibia Length ◽

Cardiac Systolic Function

Fission and fusion affect mitochondrial turnover in part by modulating mitophagy. This study aimed to clarify the role of mitochondrial fission in regulating cardiac function and autophagy in the heart. Dynamin-related protein 1 (Drp-1) plays an essential role in mediating mitochondrial fission. Therefore, we generated cardiac specific Drp-1 KO mice and utilized cultured cardiomyocytes transduced with adenovirus harboring short hairpin Drp-1 (Ad-shDrp-1) to test the effect of Drp-1 disruption both in vivo and in vitro. In Drp-1 KO hearts we observed a significantly greater mitochondrial mass ratio compared to control, as assessed by electron microscopy (Drp-1 KO: 3.57 ± 1.38, control: 1.18 ± 0.31, P<0.05). Mitochondrial ATP content was significantly lower (0.70 ± 0.07 vs 1.03 ± 0.10, P<0.05), while mitochondrial swelling was significantly greater (% decrease in absorbance; 8.01 ± 1.99 vs 2.01 ± 0.58, P<0.05) in Drp-1 KO hearts versus control. Mitochondrial membrane potential, assessed by JC-1 staining, was significantly reduced in myocytes with knockdown of Drp-1. Taken together, these results suggest that inhibition of fission causes mitochondrial dysfunction. We also examined the effect of Drp-1 depletion on autophagy. We found that the amount of LC-3 II was significantly less (0.47 ± 0.16 vs 1.32 ±0.34, P<0.05), whereas p62 expression was significantly greater (1.14 ± 0.16 vs 0.16 ± 0.06, P<0.01) in Drp-1 KO hearts compared to control. The number of LC3 dots in Ad-shDrp-1 transduced myocytes was lower than that of sh-scramble treatment. We investigated apoptosis and found that the amount of cleaved caspase-3 (0.62 ± 0.24 vs 0.18 ± 0.04, P<0.05) and the number of TUNEL positive cells (0.22 ± 0.12 vs 0.03 ± 0.06%, P<0.01) were higher in Drp-1 KO versus control hearts. Cardiac systolic function was reduced (ejection fraction; 44.5 ± 6.3 vs 85.4 ± 5.7%, P<0.01) and LVW/tibia length was greater (4.48 ± 0.38 vs 3.84 ± 0.58, P<0.05) in Drp-1 KO mice compared to control. Finally, we observed that the survival rate of Drp-1 KO mice was significantly reduced compared to control mice. Our results demonstrate that inhibition of mitochondrial fission via disruption of Drp-1 inhibits autophagy and causes mitochondrial dysfunction, thereby promoting cardiomyopathy.

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GFP fluorescence tagging alters dynamin-related protein 1 oligomerization dynamics and creates disassembly-refractory puncta to mediate mitochondrial fission

Scientific Reports ◽

10.1038/s41598-020-71655-x ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Felipe Montecinos-Franjola ◽

Brianna L. Bauer ◽

Jason A. Mears ◽

Rajesh Ramachandran

Keyword(s):

Fluorescent Protein ◽

Imaging Techniques ◽

Unit Length ◽

Mitochondrial Fission ◽

Related Protein ◽

Gfp Fluorescence ◽

Green Fluorescent ◽

Quantitative Fluorescence

Abstract Green fluorescent protein (GFP)-tagging is the prevalent strategy to monitor protein dynamics in living cells. However, the consequences of appending the bulky GFP moiety to the protein of interest are rarely investigated. Here, using a powerful combination of quantitative fluorescence spectroscopic and imaging techniques, we have examined the oligomerization dynamics of the GFP-tagged mitochondrial fission GTPase dynamin-related protein 1 (Drp1) both in vitro and in vivo. We find that GFP-tagged Drp1 exhibits impaired oligomerization equilibria in solution that corresponds to a greatly diminished cooperative GTPase activity in comparison to native Drp1. Consequently, GFP-tagged Drp1 constitutes aberrantly stable, GTP-resistant supramolecular assemblies both in vitro and in vivo, neither of which reflects a more dynamic native Drp1 oligomerization state. Indeed, GFP-tagged Drp1 is detected more frequently per unit length over mitochondria in Drp1-null mouse embryonic fibroblasts (MEFs) compared to wild-type (wt) MEFs, indicating that the drastically reduced GTP turnover restricts oligomer disassembly from the mitochondrial surface relative to mixed oligomers comprising native and GFP-tagged Drp1. Yet, GFP-tagged Drp1 retains the capacity to mediate membrane constriction in vitro and mitochondrial division in vivo. These findings suggest that instead of robust assembly-disassembly dynamics, persistent Drp1 higher-order oligomerization over membranes is sufficient for mitochondrial fission.

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SIRT1 Alleviates Aldosterone-Induced Podocyte Injury by Suppressing Mitochondrial Dysfunction and NLRP3 Inflammasome Activation

Kidney Diseases ◽

10.1159/000513884 ◽

2021 ◽

pp. 1-13

Author(s):

Mingzhu Jiang ◽

Min Zhao ◽

Mi Bai ◽

Juan Lei ◽

Yanggang Yuan ◽

...

Keyword(s):

Mitochondrial Dysfunction ◽

Knockout Mice ◽

Nlrp3 Inflammasome ◽

Inflammasome Activation ◽

Glomerular Injury ◽

Glomerular Diseases ◽

Podocyte Injury ◽

Pyrin Domain ◽

Nlrp3 Inflammasome Activation

Background: Podocyte injury contributes to progressive glomerulosclerosis. Previously, we demonstrated the important role of the NLR family pyrin domain containing 3 (NLRP3) inflammasome in mediating the podocyte injury induced by aldosterone. Silent mating type information regulation 2 homolog 1 (SIRT1) is an NAD+-dependent deacetylase that is associated with the regulation of cellular inflammation. However, whether the activation of the NLRP3 inflammasome in podocytes is regulated by SIRT1, and the mechanism involved, remains unknown. Methods: In this study, we detected SIRT1 expression in patients with podocyte disease and performed an aldosterone infusion model in podocyte-specific Sirt1 knockout mice. In cultured podocytes, we used plasmids to overexpress SIRT1 and treated the podocyte with aldosterone. Results: SIRT1 was significantly decreased in the glomeruli of patients with podocyte disease. Sirt1-deficient mice showed significant urinary albumin excretion after aldosterone infusion, and the severity of the glomerular injury was significantly greater in podocyte-specific Sirt1 knockout mice than in the wild-type mice. Moreover, podocyte conditional Sirt1 knockout aggravated NLRP3 inflammasome activation and mitochondrial dysfunction (MtD). In vitro, overexpression of SIRT1 inhibited NLRP3 activation, protected against MtD and podocyte injury. Conclusion: Taken together, these findings revealed a novel regulatory mechanism of the NLRP3 inflammasome by SIRT1 by promoting mitochondrial function, which provides some potential targets for the treatment of glomerular diseases.

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Luteolin Prevents Cardiac Dysfunction and Improves the Chemotherapeutic Efficacy of Doxorubicin in Breast Cancer

Frontiers in Cardiovascular Medicine ◽

10.3389/fcvm.2021.750186 ◽

2021 ◽

Vol 8 ◽

Author(s):

Youyang Shi ◽

Feifei Li ◽

Man Shen ◽

Chenpin Sun ◽

Wei Hao ◽

...

Keyword(s):

Breast Cancer ◽

Mitochondrial Fission ◽

Mitochondrial Morphology ◽

Protective Effects ◽

Myocardial Cells ◽

Related Protein ◽

Zebrafish Model ◽

Protein Levels

Background: Doxorubicin (Dox) is one of the most effective chemotherapy agents used in the treatment of solid tumors and hematological malignancies. However, it causes dose-related cardiotoxicity that may lead to heart failure in patients. Luteolin (Lut) is a common flavonoid that exists in many types of plants. It has been studied for treating various diseases such as hypertension, inflammatory disorders, and cancer. In this study, we evaluated the cardioprotective and anticancer effects of Lut on Dox-induced cardiomyopathy in vitro and in vivo to explore related mechanisms in alleviating dynamin-related protein (Drp1)-mediated mitochondrial apoptosis.Methods: MTT and LDH assay were used to determine the viability and toxicity of cardiomyocytes treated with Dox and Lut. Flow cytometry was used to examine ROS levels, and electron and confocal microscopy was employed to assess the mitochondrial morphology. The level of apoptosis was examined by Hoechst 33258 staining. The protein levels of myocardial fission protein and apoptosis-related protein were examined using Western blot. Transcriptome analysis of the protective effect of Lut against Dox-induced cardiac toxicity in myocardial cells was performed using RNA sequencing technology. The protective effects of Lut against cardiotoxicity mediated by Dox in zebrafish were quantified. The effect of Lut increase the antitumor activity of Dox in breast cancer both in vitro and in vivo were further employed.Results: Lut ameliorated Dox-induced toxicity in H9c2 and AC16 cells. The level of oxidative stress was downregulated by Lut after Dox treatment of myocardial cells. Lut effectively reduced the increased mitochondrial fission post Dox stimulation in cardiomyocytes. Apoptosis, fission protein Drp1, and Ser616 phosphorylation were also increased post Dox and reduced by Lut. In the zebrafish model, Lut significantly preserved the ventricular function of zebrafish after Dox treatment. Moreover, in the mouse model, Lut prevented Dox-induced cardiotoxicity and enhanced the cytotoxicity in triple-negative breast cancer by inhibiting proliferation and metastasis and inducing apoptosis.

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Squamosamide Derivative FLZ Diminishes Aberrant Mitochondrial Fission by Inhibiting Dynamin-Related Protein 1

Frontiers in Pharmacology ◽

10.3389/fphar.2021.588003 ◽

2021 ◽

Vol 12 ◽

Author(s):

Hanyu Yang ◽

Lu Wang ◽

Caixia Zang ◽

Xu Yang ◽

Xiuqi Bao ◽

...

Keyword(s):

Mitochondrial Dysfunction ◽

Dopaminergic Neurons ◽

Motor Coordination ◽

Mitochondrial Dynamics ◽

Mitochondrial Fission ◽

Mitochondrial Morphology ◽

Protective Effects ◽

Related Protein ◽

Mitochondrial Fragmentation

Mitochondrial dysfunction is involved in the pathogenesis of Parkinson’s disease (PD). Mitochondrial morphology is dynamic and precisely regulated by mitochondrial fission and fusion machinery. Aberrant mitochondrial fragmentation, which can result in cell death, is controlled by the mitochondrial fission protein, dynamin-related protein 1 (Drp1). Our previous results demonstrated that FLZ could correct mitochondrial dysfunction, but the effect of FLZ on mitochondrial dynamics remain uncharacterized. In this study, we investigated the effect of FLZ and the role of Drp1 on 1-methyl-4-phenylpyridinium (MPP+)–induced mitochondrial fission in neurons. We observed that FLZ blocked Drp1, inhibited Drp1 enzyme activity, and reduced excessive mitochondrial fission in cultured neurons. Furthermore, by inhibiting mitochondrial fission and ROS production, FLZ improved mitochondrial integrity and membrane potential, resulting in neuroprotection. FLZ curtailed the reduction of synaptic branches of primary cultured dopaminergic neurons caused by MPP+ exposure, reduced abnormal fission, restored normal mitochondrial distribution in neurons, and exhibited protective effects on dopaminergic neurons. The in vitro research results were validated using an MPTP-induced PD mouse model. The in vivo results revealed that FLZ significantly reduced the mitochondrial translocation of Drp1 in the midbrain of PD mice, which, in turn, reduced the mitochondrial fragmentation in mouse substantia nigra neurons. FLZ also protected dopaminergic neurons in PD mice and increased the dopamine content in the striatum, which improved the motor coordination ability of the mice. These findings elucidate this newly discovered mechanism through which FLZ produces neuroprotection in PD.

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mPGES-1-derived PGE2 contributes to adriamycin-induced podocyte injury

AJP Renal Physiology ◽

10.1152/ajprenal.00499.2015 ◽

2016 ◽

Vol 310 (6) ◽

pp. F492-F498 ◽

Cited By ~ 6

Author(s):

Jing Yu ◽

Wei Gong ◽

Yimei Wu ◽

Shuzhen Li ◽

Yiyun Cui ◽

...

Keyword(s):

Underlying Mechanism ◽

Pathological Feature ◽

Glomerular Diseases ◽

Podocyte Injury ◽

Phenotypic Alteration ◽

Tnf Α ◽

Progression Of Kidney Disease

Podocyte damage is a common pathological feature in many types of glomerular diseases and is involved in the occurrence and progression of kidney disease. However, the pathogenic mechanisms leading to podocyte injury are still uncertain. The present study was undertaken to investigate the role of microsomal PGE synthase (mPGES)-1 in adriamycin (ADR)-induced podocyte injury as well as the underlying mechanism. In both mouse kidneys and in vitro podocytes, application of ADR remarkably enhanced mPGES-1 expression in line with a stimulation of cyclooxygenase-2. Interestingly, inhibition of mPGES-1 with a small interfering RNA approach significantly attenuated ADR-induced downregualtion of podocin and nephrin. Moreover, ADR-induced podocyte apoptosis was also markedly blocked in parallel with blunted caspase-3 induction. In agreement with the improvement of cell phenotypic alteration and apoptosis, the enhanced inflammatory markers of IL-1β and TNF-α were also significantly suppressed by mPGES-1 silencing. More importantly, in mPGES-1-deficient mice, albuminuria induced by ADR showed a remarkable attenuation in line with decreased urinary output of PGE2 and TNF-α, highly suggesting an in vivo role of mPGES-1 in mediating podocyte injury. In summary, findings from the present study offered the first evidence demonstrating a pathogenic role of mPGES-1 in mediating ADR-induced podocyte injury possibly via triggering an inflammatory response.

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Huaier Cream Protects against Adriamycin-Induced Nephropathy by Restoring Mitochondrial Function via PGC-1αUpregulation

PPAR Research ◽

10.1155/2015/720383 ◽

2015 ◽

Vol 2015 ◽

pp. 1-11 ◽

Cited By ~ 7

Author(s):

Ruochen Che ◽

Chunhua Zhu ◽

Guixia Ding ◽

Min Zhao ◽

Mi Bai ◽

...

Keyword(s):

Mitochondrial Dysfunction ◽

Mitochondrial Function ◽

Cell Injury ◽

Therapeutic Drug ◽

Mtdna Copy Number ◽

Glomerular Diseases ◽

Mitochondrial Transcription Factor ◽

Transmission Electron

The mechanism by which Huaier, a Chinese traditional medicine, protects podocytes remains unclear. We designed the present study to examine whether mitochondrial function restored by PGC-1αserves as the major target of Huaier cream in protecting ADR nephropathy. After ADR administration, the podocytes exhibited remarkable cell injury and mitochondrial dysfunction. Additionally, ADR also reduced PGC-1αbothin vivoandin vitro. Following the Huaier treatment, the notable downregulation of PGC-1αand its downstream molecule mitochondrial transcription factor A (TFAM) were almost entirely blocked. Correspondingly, Huaier markedly ameliorated ADR-induced podocyte injury and mitochondrial dysfunction in both rat kidneys and incubated cells as it inhibited the decrease of nephrin and podocin expression, mtDNA copy number, MMP, and ATP content. Transmission electron microscopy result also showed that Huaier protected mitochondria against ADR-induced severe mitophagy and abnormal changes of ultrastructural morphology. In conclusion, Huaier can protect podocytes against ADR-induced cytotoxicity possibly by reversing the dysfunction of mitochondria via PGC-1αoverexpression, which may be a novel therapeutic drug target in glomerular diseases.

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Dynamin-related protein 1 inhibition reduces hepatic PCSK9 secretion

Cardiovascular Research ◽

10.1093/cvr/cvab034 ◽

2021 ◽

Cited By ~ 1

Author(s):

Maximillian A Rogers ◽

Joshua D Hutcheson ◽

Takehito Okui ◽

Claudia Goettsch ◽

Sasha A Singh ◽

...

Keyword(s):

Disease Risk ◽

Mitochondrial Fission ◽

Low Density Lipoprotein ◽

Density Lipoprotein ◽

Mrna Levels ◽

Related Protein ◽

Pcsk9 Inhibitor ◽

Density Lipoprotein Receptor

Abstract Aims Proteostasis maintains protein homeostasis and participates in regulating critical cardiometabolic disease risk factors including proprotein convertase subtilisin/kexin type 9 (PCSK9). Endoplasmic reticulum (ER) remodeling through release and incorporation of trafficking vesicles mediates protein secretion and degradation. We hypothesized that ER remodeling that drives mitochondrial fission participates in cardiometabolic proteostasis. Methods and results We used in vitro and in vivo hepatocyte inhibition of a protein involved in mitochondrial fission, dynamin-related protein 1 (DRP1). Here, we show that DRP1 promotes remodeling of select ER microdomains by tethering vesicles at ER. A DRP1 inhibitor, mitochondrial division inhibitor 1 (mdivi-1) reduced ER localization of a DRP1 receptor, mitochondrial fission factor, suppressing ER remodeling-driven mitochondrial fission, autophagy, and increased mitochondrial calcium buffering and PCSK9 proteasomal degradation. DRP1 inhibition by CRISPR/Cas9 deletion or mdivi-1 alone or in combination with statin incubation in human hepatocytes and hepatocyte-specific Drp1-deficiency in mice reduced PCSK9 secretion (−78.5%). In HepG2 cells, mdivi-1 increased low-density lipoprotein receptor via c-Jun transcription and reduced PCSK9 mRNA levels via suppressed sterol regulatory binding protein-1c. Additionally, mdivi-1 reduced macrophage burden, oxidative stress, and advanced calcified atherosclerotic plaque in aortic roots of diabetic Apoe-deficient mice and inflammatory cytokine production in human macrophages. Conclusions We propose a novel tethering function of DRP1 beyond its established fission function, with DRP1-mediated ER remodeling likely contributing to ER constriction of mitochondria that drives mitochondrial fission. We report that DRP1-driven remodeling of select ER micro-domains may critically regulate hepatic proteostasis and identify mdivi-1 as a novel small molecule PCSK9 inhibitor.

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