Netrin-1 improves functional recovery through autophagy regulation by activating the AKT/mTOR signaling pathway in high-glucose-induced renal proximal tubule injury

doi:10.21203/rs.3.rs-1597367/v1

Download PDF

Research Article

Netrin-1 improves functional recovery through autophagy regulation by activating the AKT/mTOR signaling pathway in high-glucose-induced renal proximal tubule injury

https://doi.org/10.21203/rs.3.rs-1597367/v1

This work is licensed under a CC BY 4.0 License

Version 1

posted

You are reading this latest preprint version

Netrin-1, a survival factor, is highly expressed in the kidney and reported to participate in some types of acute or chronic kidney diseases. Nevertheless, the causal link beneath the effect on renal proximal tubule cells of Netrin-1 under pathological conditions in diabetic nephropathy (DN) hasn't been elucidated. Our team explored the effects of Netrin-1 on the etiopathogenesis of DN. In vitro, high blood glucose caused the downregulation of Netrin-1, which induced the stimulation of AKT/mTOR signal transmission with the elevated programmed cell death of renal proximal tubule cells and increased actin cytoskeleton derangement. The supplementation of recombinant rat Netrin-1 reversed these pathological changes and it was tightly related to the recovery of autophagy flux. In addition, those pathologic variations were abolished by transfection with a lentivirus expressing short hairpin RNA against Netrin-1 or a mTOR inhibitor in vitro. The discoveries of the present research reveal that Netrin-1 is responsible for the mediation of high glucose-triggered loss of renal proximal tubule cells by regulating their detachment and programmed cell death in vitro. The present paper facilitates our comprehension regarding the causal links of renal proximal tubule cell injury in DN and reveals that targeting Netrin-1-associated signal transduction has treatment significance for DN.

AKT/mTOR signaling

autophagy

Netrin-1

renal proximal tubule injury

The fact that high glucose (HG) causes glomerular cell damage in DN has aroused extensive interest among nephrologists; nevertheless, mounting researches have revealed that the renal tubule, particularly the proximal tubule, is a site of damage as well in DN [1]. The elevation of the programmed cell death of renal proximal tubules has been discovered in diabetic sufferers and HG milieu [2].

Netrin-1 is a secreted glycosidoprotein pertaining to the Netrins protein family, which has a strong chemotropic function for axonal guidance. From the perspective of biofunction, it's associated with the etiopathogenesis of cellular proliferation, differentiation adhesion and antiapoptosis events [3]. As a pivotal signal transduction protein regulating several molecule pathways, the injury repair effect of Netrin-1 in other organs has been corroborated. More and more researches have revealed that abnormal Netrin-1 stimulation in kidneys participates in renal fibrogenesis as well [4]. Our recent study has demonstrated that Netrin-1 expression and activation are reduced in diabetic proximal tubule epitheliums, whereas the potential molecule-level causal links of Netrin-1 stimulation and its effects on diabetes-related renal damage are still elusive.

HG stimulates the phosphoinositide 3-kinase (PI3K)/AKT signaling regulating cellular proliferative activity, cellular growth and death, and such pathway broadly participates in the course of ischemia-related illnesses [5]. The activation of PI3K-AKT signal paths triggers the phosphonation of mTOR, causing the eventual inhibition of autophagy. Recently, researches have unveiled an effect of Netrin-1 on modulating autophagic activities via the regulation of lysosomal functions [6]. Intriguingly, another study has revealed that exogenous Netrin-1 reduces autophagosomal formation and eventually suppresses autophagic activities[7]. Nevertheless, if elevated expressing level of Netrin-1 in DN is under the mediation of AKT/mTOR signal transmission and if autophagy further affects the cell skeleton and function of proximal tubule epitheliums are still elusive.

Our team explored the function and the potential causal link of Netrin-1 in the etiopathogenesis of DN via completing cell and molecule analysis. The discoveries of the present paper reveal that Netrin-1 can regulate apoptosis and affect the renal proximal tubule cytoskeleton via the interplay with and the stimulation of AKT signaling, hence promoting autophagic flux.

Cell culture

HK-2 is a human proximal tubule cell line from the kidney, provided by Hong Kong Biotechnology Limited. Shanghai FuHeng (Shanghai FH0228, China). After a while, cells were subcultured using RPMI - 1640 (SH30809.01 B; Hyclone) medium including 10% FBS (10270106; Gibco). Cell differentiation was induced by 1% dual antibiotics without interferon γ in a 5% CO2 incubator incubated under 37°C for 10-14 days. The medium was substituted daily or every other day depending on the growth of the cells. In addition, HK-2 cells were given 5.5 mM glucose (termed NG group) or 30 mM glucose (termed HG group) for 24 h when HG was administered. The mTOR inhibitor rapamycin (100nmol/L) was purchased from Selleck (S1039) treated cells. Recombinant rat Netrin-1 was obtained from R&D Systems (6419-N1-025). HK-2 cells were treated with the specified concentration of Netrin-1 (0 or 20ng/ml) for 24 h.

Western blotting

Cell proteins were extracted with RIPA lysis buffer (Solarbio, P0013B) and electrophoretized with 10% SDS-Page gel. After the membrane was transferred to PVDF, anti-Netrin-1 (20235-1-AP; proteintech), anti-cleavage Caspase 3(#9664T; cst), anti-Bax (#14796; cst), anti-Bcl-2 (ab1722; millipore), anti-LC3 (#12741S; cst), anti-p62 (#5114S; cst), anti-Beclin-1 (#3738S; cst), anti-p-AKT (#4060S; cst), anti-AKT (#4691; cst), anti-p-mTOR (ab109268; abcam), anti-mTOR (ab134903; abcam), anti-mouse IgG (HRP,#7076, cst), anti-rabbit IgG(HRP, #7074; cst) and anti-GAPDH (#5174; cst). After PBS washing and incubation with secondary antibody, the membrane was visualized using intesive chemiluminescence (ECL) detection system (Santa Cruz Biotechnology).

Cell apoptosis analysis

Apoptosis rate was detected by flow cytometry (40302ES20; YEASENusing) Annexin V-FITC/PI staining. In simple terms, different groups of cells are treated with trypsin. HK-2 was originally digested with 0.25% trypsin (without EDTA), centrifuged for 5 min, and then suspended with PBS. The cells were incubated with 300 μL binding buffer and stained with Annexin V-FITC (5 μL) and PI (10 μL).

RNA extraction and real-time quantitative PCR (RT-qPCR)

Total RNA was extracted with Trizol reagent (Invitrogen, CA) and reversed transcribed into complementary DNA (cDNA) using Prime Script RT Reagent Kit (Takara). SYBR Premix Ex Taq II (Takara Bio, Otsu, Japan) was used to detect gene fold change by RT-qPCR. The mRNA levels of target genes were normalized to GAPDH. Primer sequences are as follows:

Netrin-1: 5’-TGCAAGCCCTTCCACTACG-3′ (F) and 5’- TGTTGTGGCGACAGTTGAGG-3′ (R); GAPDH: 5’- GGAGCGAGATCCCTCCAAAAT-3′ (F) and 5’- GGCTGTTGTCATACTTCTCATGG-3′ (R).

GFP-mRFP-LC3 adenoviral transfection

HK-2 cells were placed in a glass substrate petri dish and infected with an adenovirus vector containing GFP-MRFP-LC3 (Asia-Vector Biotechnology, Shanghai Co.LTD). Fresh medium was used instead of medium, and HK-2 was incubated for one day.

Then, confocal laser scanning microscope (OLYMPUS; FV1000) was used to observe HK-2 in order to monitor autophagy flux and assess the number of yellow and green dots.

Actin cytoskeletal disorder score

F-actin in HK-2 cells was stained with TRITC Phalloidin (G1401, purchased from Servicebio, China) and the images were observed under a Zeiss (Zeiss, Jena, Germany) fluorescence microscope.The cells were inoculated into a 24-well plate with a glass lid and fixed in PBS with 4% paraformaldehyde. 0.5% Triton X-100 in PBS can pass through the cell membrane. DAPI (G1012, Servicebio) and TRITC Phalloidin (100 nM; G1401, Servicebio, Wuhan, China) for cell structure staining. For the semi-quantitative analysis of actin cytoskeleton disorder, we conducted a scoring study according to the above method [8]. The disorganized regions of F-actin stained by Phalloidin were defined as disordered regions.

Statistical Analysis.

The data are expressed as mean ± SEMs. One way ANOVA followed by Bonferroni correction was used to analyze means between more than two groups. Unpaired t-test was used to analyze data when two groups were present. GraphPad Prism software was used for statistical analyses. All experiments were repeated at least 3 times, and representative experiments are shown. P<0.05 was considered as statistically significant.

1. Netrin ameliorated hyperglycemia induced apoptosis and actin cytoskeleton derangement (Fig. 1)

The dysfunction of renal proximal tubules is pivotal for the progression of DN. Our team identified the role of Netrin-1 in the programmed cell death and actin cell skeleton of HK-2 cells under HG status. Western blott (WB) analysis revealed that Netrin-1 level was remarkably decreased under HG status vs the controls (Fig. 1A-B). Flow cytometry analysis revealed that the quantity of apoptosis-related HK-2 cells was remarkably elevated under HG vs the controls. Furthermore, the HG-triggered programmed cell death of HK-2 cells was weakened by recombinant Netrin-1 (Fig. 1E). The results in Fig.1C and Fig.1D displayed that the expression of proapoptosis protein (Cleaved caspase-3 and Bax) was remarkably reinforced in HK-2 cells under HG, whereas their contents were down-regulated posterior to the exposure to recombinant Netrin-1 vs the controls. Bcl-2, as a vital antiapoptosis protein, displayed opposite results.

Netrin-1 suppresses endothelium adhesive and migratory abilities, and alters their actin cell skeleton [9]. For the sake of identifying if Netrin-1 affected cytoskeleton reorganisation in renal proximal tubule cells, we overexpressed Netrin-1 in mankind foot cells via Netrin-1 recombinant protein. The treatment with HG remarkably reduced the quantity of F-actin fibers and considerably elevated the quantity of disorganised. HK-2 cells incubated under HG and Netrin-1 recombination protein showed reserve effects (Fig. 1F). Hence, HG-triggered cytoskeleton disorder in HK-2 cells might be owing to a decrease in the expressing level of Netrin-1. Those results reveal that Netrin-1 is vital for renal proximal tubule function disorder under HG status.

2. Netrin-1 mediated AKT/mTOR pathway and induced autophagy in high glucose-treated HK-2 cells (Fig 2.)

Subsequently, our team determined the causal link by which Netrin-1 triggered the programmed cell death of HK-2 cells and actin cytoskeleton disorder under HG status. Our team revealed that Netrin-1 decreased AKT and mTOR phosphorylation under HG conditions (Fig. 2A-B). Those results reveal that the AKT/mTOR signal path exerts an effect on the decreased Netrin-1-triggered programmed cell death of HK-2 cells under HG status.

The expressing levels of Beclin-1 and LC3, 2 biological markers of autophagic activity, were identified in HG-treated HK-2 via WB assay to explore the regulatory roles of Netrin-1 in autophagy. Our team subsequently identified autophagy flux via exposing HK-2 to GFP-mRFP-LC3 adenovirus. As a result, the exposure to HG reduced the red and green puncta vs controls, and this phenomenon was reversed by the addition of Netrin-1 in HK-2 cells (Fig. 2C), which suggested that Netrin-1 recovered autophagy flux repressed by HG in HK-2. suggesting that Netrin-1 restored autophagic flux that was inhibited by HG in HK-2. In addition, our team substantiated that the addition of Netrin-1 abolished the reduction of LC-3II/LC-3I and Beclin-1 contents and the elevation of p62 contents in HG-exposed HK-2 cells (Fig. 2A-B). The number of autophagosomes was increased via the observation with an electron microscope under HG conditions (Fig. 2D). Collectively, those outcomes revealed that Netrin-1 weakened programmed cell death, caused autophagic activity and repressed the AKT/mTOR signal path in HG-exposed HK-2 cells.

3. Effects of autophagy on apoptosis and cytoskeletal derangement in HK-2 cells (Fig 3.)

We hypothesize that Netrin-1 binds to and activates mTOR on the HK-2 cells, hence causing programmed cell death and actin cytoskeleton disorder in an autocrine or paracrine manner. For the sake of testing such assumption, our team knocked down Netrin-1 in differentiated human renal proximal tubules by lentivirus-mediated shRNA interference. Real-time quantitative PCR analysis substantiated the validity of knockout by the Netrin-1 shRNA (Fig. 3A). Subsequently, HK-2 cells were cultivated under HG status (30 mM), or exposed to rapamycin (a mTOR inhibitor) and lentivirus expressing shRNA against Netrin-1 (shNetrin-1) for 24 h. The silencing of Netrin-1 significantly increased the expression of phosphorylation proteins (phospho-AKT and phospho-mTOR ) and caused programmed cell death, as displayed by the reduced content of Bcl-2 and the elevated contents of cleaved caspase-3 and Bax (Fig. 3B-E). Nevertheless, the phenomenon that Netrin-1 silencing triggered programmed cell death under HG status was remarkably weakened by rapamycin, so it was with the phenomenon that Netrin-1 silencing triggered cytoskeleton disorder (Fig. 3F-G). LC-3 fluorescence intensity was induced by Netrin-1 silencing or the treatment of rapamycin. Autophagy flux was caused by silencing Netrin-1 in HG-exposed HK-2 cells, and this phenomenon was offset by the treatment of rapamycin. The decrease of LC-3II/LC-3I and Beclin-1 and the increase of p62 induced by Netrin-1 silencing were terminated via the inhibition of mTOR. Increased autophagosomes by silencing Netrin-1 in HG-exposed HK-2 cells was reversed by the treatment of rapamycin(Fig. 3H-I). In conclusion, it was suggested that Netrin-1 attenuated programmed cell death and promoted autophagic activity in HG-exposed HK-2 cells via the AKT/mTOR signal path.

DN is a chronic disease characterized by aberrant glomerulus perfusion and saccharometabolism, which thickens glomerulus basement membrane, exocellular matrix deposition, proliferation variations and atrophy of kidney tubules inducing interstitial fibrosis and glomerular sclerosis. Researches have displayed that HG exert deleterious influence on renal tubular cells (RTCs) [10]. Tubular hypertrophy and tubulointerstial fibrosis are identified as important steps accompanied by progressive renal dysfunction.The murine renal injury models established by reveal Reeves et al. [11] revealed that the primary source of Netrin-1 in acute renal damage was proximal tubule epitheliums. Moreover, the expression of Netrin-1 is minimal in tubule epitheliums of healthy kidneys, whereas its expression is extensive in impaired kidneys. Another study found elevated Netrin-1 in transplant recipients after surgery, and the levels were reduced with the amelioration of graft function [12]. Coherent with this paper, a recently finished research used a diverse method to reveal the roles of Netrin-1 in DN, finding that transgene mice with tubule-specific Netrin-1 over expression were protected in DN. Herein, our team intended to explore the roles as well as the mechanism of Netrin-1 in RTC injury.

Our team discovered that the expressing level of Netrin-1 in HK-2 cells from the kidney cortex was remarkably decreased under high glucose condition. In addition, it caused the programmed cell death of HK-2 cells and actin cell skeleton disorganisation. Furthermore, those effects were weakened via the blockade of Netrin-1 or using a mTOR inhibitor in vitro. Recent studies have provided preliminary evidence pertaining to the crosstalk between proximal tubular epithelial cells and determined podocytes [13]. Disordered actin cytoskeleton complexes have been discovered to be pivotal for foot process effacement. After podocyte impairment, cytoskeleton reorganisation induces foot process effacement. The remarkably organised parallel contractile actin filament bundles in the foot process become disorganised, shortened, and branched under pathologic circumstances. We demonstrated that Netrin-1 altered renal tubular cell morphology by activating the AKT/mTOR pathway under HG conditions.

The suppression of autophagic activity through the AKT/mTOR signal path is identified RTC damage. Previously, researches revealed that the AKT/mTOR signal transmission was stimulated via HG and facilitated RTC impairment and autophagic activity suppression in DN. Those discoveries have unveiled that AKT/mTOR is a vital signal transduction process for the regulation of autophagic activity and RTC impairment, and that targeting such signal transmission is a prospective method to mitigate DN. Intriguingly, a recently finished research unraveled that Netrin-1 could stimulate the AKT/mTOR signal path in neuron growth cones [14]. As is known, mTOR is a serine/threonine protein kinase regulating cellular metabolic, proliferative, death-related, and survival-related processes; such protein is associated with certain physiologic processes as well, like transcriptional process, mRNA turnover and translational process, ribosome genesis, vesicular transport, autophagic activity and cytoskeleton organisation [15–16]. Previously, researches revealed that the suppression of the mTOR signal path had neuroprotection effect via activating autophagic activity in the central nervous system [17]. Based on these findings, our team explored the roles of Netrin-1 down-expression in this progress. Interference technology was used to knock down Netrin-1 under HG conditions. Importantly, our data suggested that slience Netrin-1 significantly increased the expressing levels p-AKT and p-mTOR and caused programmed cell death, as displayed by the reduced content of Bcl-2 and the elevated contents of cleaved caspase-3 and Bax. As per WB and immune fluorescence dyeing outcomes for Beclin-1 and LC3, our team displayed that Netrin-1 could activate autophagic activity in damaged renal proximal tubules posterior to HG treatment. The roles of Netrin-1 sliencing in modulating autophagic activity and inducing apoptosis were partially terminated by rapamycin, so it was with the fact that Netrin-1 silencing induced cytoskeletal derangement, which suggested that the decreased expression of Netrin-1 exacerbated the severity of renal proximal tubule injury and that knocking down Netrin-1 could directly reverse this phenomenon by improving autophagy signal transduction. Those results might reveal that Netrin-1 activated autophagic activity via stimulating the AKT/mTOR signal path. In contrast to rapamycin alone, the combination usage of Netrin-1 silencing and rapamycin could remarkably reduce the ratio of p-AKT/AKT and p-mTOR/mTOR and decline the expressing level of Beclin-1 and ratio of LC3B-II/LC3B-I. Such outcome verified that Netrin-1 activated autophagic activity via stimulating the AKT/mTOR signal path posterior to DN. Nevertheless, our team failed to rule out that Netrin-1 could activate autophagic activity via other underlying signal paths posterior to DN. Thereby, indepth researches are required to explore other underlying molecule-level causal links of Netrin-1 in autophagic activity activation posterior to DN.

Hence, one of the deficiencies of this paper is that the potential causal links, by which Netrin-1 acceptor therapy modulated autophagosomal formation and autophagosomal-lysosomal fusion, weren't revealed. In addition, insufficient proofs from in vivo assays is another flaw. Those deficiencies can offer guidance for future researches.

Collectively, the decreased expressing level of Netrin-1 under HG status induced renal proximal tubule impairment. Netrin-1 silencing induced the programmed cell death of renal proximal tubule cells and actin cytoskeleton disorder through the AKT/mTOR signal path under HG status. Functionally, Netrin-1 triggered autophagy in HG-exposed HK-2 cells.The slience Netrin-1-triggered apoptosis of renal proximal tubule cells was attenuated by rapamycin. Therefore, the pivotal role of Netrin-1 in DN reveals that manipulating Netrin-1 associated signal paths is probably valid.

Funding This work was funded by Suzhou Science and Education Project (No. KJXW2019037) .

Conflict of interest

The authors declare that there is no conflict of interest.

Ethical approval

This article contains no original data using human or animal subjects.

Informed consent

For this type of study formal consent is not required.

Authors Contributions

CXL and MW designed experiments, conducted literature review and wrote the manuscript.YZ and AJH helped with the acquisition and interpretation of data and with manuscript revisions. XNH, JQC and TL prepared figure 1-3. ML, MF and RX helped to polish the language. HH L guaranteed this work and provide academic guidance. All authors read and approved the final manuscript. The authors declare that all data were generated in-house and that no paper mill was used.

Consent to publish

Not applicable.

Availability of data and materials

Our data will not be shared due to it was involved in individual privacy and needed further study.

Chevalier RL.The proximal tubule is the primary target of injury and progression of kidney disease: role of the glomerulo tubular junction. Am J Physiol Renal Physiol 311:F145–F161. http://doi.org/10.1152/ajprenal.00164.2016 (2016) .
Su SS, Li BP, Li CL, Xiu FR, Wang DY, Zhang FR. Downregulation of MiR-218 can alleviate high-glucose-induced renal proximal tubule injury by targeting GPRC5A. Biosci Biotechnol Biochem 84(6): 1123–1130. http:// doi.org/ 10.1080/ 09168451. 2020. 1717330 (2020).
Ke XJ, Liu CX, Wang Y,Ma JH, Mao XM, Li Q. Netrin-1 promotes mesenchymal stem cell revascularization of limb ischaemia. Diabetes & Vascular Disease Research 13(2):145–56. http:// doi.org/ 10.1177/1479164115611594 (2016).
Jiao XJ, Zhang D, Hong Q, Yan L, Han QX, Shao FM, Cai GY, Chen XM, Zhu. HY. Netrin-1 works with UNC5B to regulate angiogenesis in diabetic kidney disease. Front Med 14(3): 293–304. http:// doi.org/ 10.1007/s11684-019-0715-7 (2020).
Xu ZX, Jia KQ, Wang H, Gao F, Zhao S, Li F, Hao J. METTL14-regulated PI3K/AKT signaling pathway via PTEN affects HDAC5-mediated epithelial-mesenchymal transition of renal tubular cells in diabetic kidney disease. Cell Death Dis 12(1):32. http:// doi.org/10.1038/s41419-020-03312-0 (2021).
Bai LJ, Mei XF, Shen ZL, Bi YL, Yuan YJ, Guo ZP, Wang HY, Zhao HS, Zhou ZP, Wang C, Zhu KM, Li G, Lv G. Netrin-1 Improves Functional Recovery through Autophagy Regulation by Activating the AMPK/mTOR Signaling Pathway in Rats with Spinal Cord Injury. Scientific Reports 7: 42288. http:// doi.org/ 10.1038/ srep42288 (2017).
Tang TC, Gao DK, Yang XS, Hua XM, Li ST, Sun H. Exogenous Netrin-1 Inhibits Autophagy of Ischemic Brain Tissues and Hypoxic Neurons via PI3K/mTOR Pathway in Ischemic Stroke. J Stroke Cerebrovasc Dis May 28(5): 1338–1345. http:// doi.org/ 10.1016/j.jstrokecerebrovasdis (2019).
Guo KF, Pan P, Wu M, Ma YW, Lu JX, Chen HB. Hyposialylated angiopoietin-like-4 induces apoptosis of podocytes via β1 Integrin/FAK signaling in diabetic nephropathy. Molecular and Cellular Endocrinology 505: 110730. http:// doi.org/ 10.1016/j.mce.2020.110730 (2020).
Philippe MD, Nathalie LV. The calcium-activated protease calpain regulates netrin-1 receptor deleted in colorectal cancer-induced axon outgrowth in cortical neurons. J Neurochem 152(3): 315–332. http:// doi.org/ 10.1111/jnc.14837 (2020).
Lee WC, Chau YY, Ng HY, Chen CH, Wang PW, Liou CW, Lin TK, Chen JB. Empag7liflozin Protects HK-2 Cells from High Glucose-Mediated Injuries via a Mitochondrial Mechanism. Cells 8(9): 1085. http:// doi.org/ 10.3390/ cells8091085 (2019).
Reeves WB, Kwon O, Ramesh G. Netrin-1 and kidney injury. II. Netrin-1 is an early biomarker of acute kidney injury. American journal of physiology. Renal physiology 294(4):F731-738. http:// doi.org/ 10.1152/ajprenal.00507.2007 (2008).
Wang WW, Reeves WB, Ramesh G. Netrin-1 and kidney injury. I. Netrin-1 protects against ischemia-reperfusion injury of the kidney, American journal of physiology. Renal physiology 294(4): F739-747. http:// doi.org/ 10.1152/ ajprenal. 00508.2007 (2008).
Xu CM, Zhou XJ, Xie TY, Zhou Y, Zhang Q, Jiang S, Zhang R, Liao L, Dong JJ. Renal tubular Bim mediates the tubule-podocyte crosstalk via NFAT2 to induce podocyte cytoskeletal dysfunction.Theranostics 10(15): 6806–6824. http:// doi.org/ 10.7150/thno.43145 (2020).
Piper M, Lee AC, Horck FP, McNeilly H, Lu TB, Harris WA, Holt CE. Differential requirement of F-actin and microtubule cytoskeleton in cue-induced local protein synthesis in axonal growth cones. Neural Development 10:3. http:// doi.org/ 10.1186/s13064-015-0031-0.
Yoon MS. mTOR as a Key Regulator in Maintaining Skeletal Muscle Mass. Front Physiol 8:788. doi.org/ 10.3389/fphys.2017.0078 (2017)8 .
Murugan AK. mTOR: Role in cancer, metastasis and drug resistance. Semin Cancer Biol 59:92–111. doi.org/ 10.1016/j.semcancer.2019.07.003 (2019).
Nerea DS, Alejo E. The mTOR-Autophagy Axis and the Control of Metabolism. Front Cell Dev Biol 9: 655731. doi.org/ 10.3389/fcell.2021.655731 (2021).

No competing interests reported.

Download PDF

Version 1

posted

You are reading this latest preprint version

Netrin-1 improves functional recovery through autophagy regulation by activating the AKT/mTOR signaling pathway in high-glucose-induced renal proximal tubule injury

Status:

Version 1

Abstract

Figures

Introduction

Materials And Methods

Results

Discussion

Declarations

References

Additional Declarations

Status:

Version 1