RSL3 | Micro rna2

Tetrahedral DNA Nanostructures Inhibit Ferroptosis and Apoptosis in Cisplatin-induced Renal Injury

Jiaying Li, Liwen Wei, Yuanqing Zhang,* and Minhao Wu*
This: ACS Appl. Bio Mater. 2021, 4, 5026−5032

ABSTRACT: Acute kidney injury (AKI) is the most serious adverse reaction during cisplatin chemotherapy, which limits the drug’s clinical eﬀects. Therefore, eﬀective strategies for protective therapy need to be developed. In the current study, we veriﬁed that tetrahedral DNA nanostructures (TDNs), promising DNA nano biomaterials, played protective roles against cisplatin-induced death of renal tubular cells. Herein, we observed that TDNs decreased the generation of lipid reactive oxygen species (ROS), restored the down-regulation of glutathione peroxidase 4 (GPX4), and hence inhibited ferroptosis induced by RSL3, a typical inducer of ferroptosis. In addition, we proved that TDNs attenuated cisplatin-induced ferroptosis by reversing the down-regulation of GPX4 and attenuated apoptosis induced by cisplatin via reducing the cleavage of poly(ADP-ribose) polymerase (PARP). Taking this all into account, our investigation suggested the potential of TDNs for cisplatin-induced AKI therapy.

KEYWORDS: TDNs, ferroptosis, apoptosis, cisplatin, renal injury, AKI

⦁ INTRODUCTION
Cisplatin was ﬁrst used clinically as a class of chemo-
therapeutic agents in 1978 in the United States and has now been widely used around the world.1,2 However, cisplatin siRNA,18 aptamers,19 and immunogenic molecules20,21 for pathogenic infections22 and cancer treatment.17 Moreover, TDNs also promote cell migration and tissue regeneration because of their antioxidant and antiapoptotic eﬀects. Zhang cytotoxicity remains a common side eﬀect.3 Acute kidney et al. conﬁrmed that TDNs inhibited LPS-induced inﬂam-injury (AKI) occurs in approximately 30% of patients with tumors after cisplatin chemotherapy treatment (referred to as cis-AKI in the following).4,5 AKI is closely related to cell death processes, including apoptosis and ferroptosis.6,7 PARP repairs the intracellular DNA damage. However, cisplatin induces the activation of apoptosis-associated caspases8,9 which execute the cleavage of PARP and thus trigger apoptosis.10,11 Additionally, recent reports have demonstrated that ferroptosis, a form of regulated cell death associated with lipid hydroperoxidation, is involved in cis-AKI via GSH mation and apoptosis in RAW264.7 cells.23 Zhang et al. reported that TDNs decreased oxidative damage and apoptosis in vitro myocardial ischemia-reperfusion injury model.24 In addition, TDNs are promising for application in wound healing.25,26 However, whether TDNs regulate the ferroptosis and cisplatin-induced death in renal tubular cells remains unknown. In addition, DNA origami nanostructures (DONs) inhibit ROS production and restore renal function in the AKI model in vitro and in vivo; however, the detailed mechanisms remain unclear.27
depletion along with the inactivation of the GPX familybmembers.12,13 Hence, targeting apoptosis and ferroptosis in cis-AKI could be a potential treatment.4Currently, DNA nanostructures, the programmable func- tionalized nano biomaterials, exhibit prospective roles in diﬀerent applications such as early disease diagnosis and target drug delivery.14,15 The advantages of DNA nanostruc- tures are their good stability, biocompatibility, biodegrad- ability, and easy synthesis. Through the appropriate design of DNA sequences, diﬀerent kinds of DNA self-assembly nanostructures such as Tetrahedral DNA nanostructures (TDNs), DNA origami nanostructures, and DNA nanotube nanostructures have been manufactured.16 TDNs are used to deliver bioactive molecules, including anticancer drugs,17

In this study, we veriﬁed that TDNs inhibited ferroptosis induced by RSL3 or cisplatin via regulating GPX4 expression in renal tubular cells. Additionally, we demonstrated that TDNs attenuated cisplatin-induced apoptosis by reducing the cleavage of PARP. To the best of our knowledge, this was the ﬁrst study proving that TDNs inhibited ferroptosis and
ImageReceived: March 10, 2021
Accepted: May 12, 2021
Published: May 25, 2021

■played protective roles against cisplatin-induced death of renal tubular cells. Beyond these, our study provided a potential targeted therapeutic strategy for cis-AKI.

MATERIALS AND METHODS
Materials. C11 BODIPY 581/591 was purchased from Invitrogen. RSL3, cisplatin, and ferrostatin-1 (Fer-1) were obtained
Transmission electron microscopy (TEM) and 3.5% agarose gel electrophoresis were performed to demonstrate the successful synthesis of TDNs as previously reported.26,28 The size of TDNs was measured by nanoparticle analyzers (Malvern Instrument Ltd., Malvern).
Cell Culture and Treatment. Human proximal tubular epithelial cells (HK-2 cells) were cultivated with Dulbecco’s modiﬁed Eagle’s medium/nutrient mixture F-12 (DMEM/F-12) from Selleck. Z-VAD-fmk was purchased from MCE. PI/AnnexinV apoptosis detection kit was obtained from BD.
Synthesis and Characterization of TDNs. Single strands of TDNs were obtained by Sangon Biotech. TDNs were assembled according to the method reported previously.21 Sequences of the four single strands of DNA of TDNs are shown in Table 1. CFX96 real-time PCR system (Bio-Rad). Sequences of primer pairs are shown in Table 2. Western Blot. Cells (1.5 × 105) were lysed by 60 μL of cell lysis buﬀer. An equal amount of total proteins were prepared and then divided according to their molecular weights by SDS-PAGE gel electrophoresis. The target proteins were transferred to polyvinyli- dene diﬂuoride membranes. After blocking in 5% BSA, the target membranes were incubated with primary antibodies as follows: β- actin (A1978, Sigma-Aldrich), GAPDH (G5262, Sigma-Aldrich),

Synthesis and characterizations of TDNs. (a) Assembly of TDNs displayed in the schematic. (b) Synthesis of TDNs analyzed by agarose gel electrophoresis. A, AB, and ABC are, respectively, single-stranded DNA, partially complemented double-stranded, and triple-stranded DNA. (c) TEM image of TDNs. (d) Nanoparticle analyzer performed to detect the size distribution of TDNs.
Cellular uptake and cytotoxicity of TDNs. (a) Confocal microscopy was used to observe the cellular uptake of Cy5-TDNs. Scale bars: 20 μm. (b, c) Flow cytometry was conducted to analyze the cellular uptake of Cy5-TDNs by HK-2 cells for 1, 2, and 3 h. ****p < 0.0001, ns, not signiﬁcant. (d) After treatment with TDNs (200 nM) for 24 h, viability was determined. ns, not signiﬁcant.

GPX4 (ab125066, Abcam), or cleaved PARP (5625S, CST). The washed membranes were incubated with secondary antibodies. The blots on membranes were exposed by chemoluminescence with New-SUPER ECL kit (KeyGEN).Cytotoxicity Analysis. Cytotoxicity was quantiﬁed using the
was 18.79 nm in diameter (Figure 1d). The results above indicated that we successfully assembled TDNs.Cellular Uptake and Cytotoxicity of TDNs. To explore the roles of TDNs in subsequent experiments, ﬁrst, we detected the cellular uptake by HK-2 cells and the nontoxic
CCK-8 assay (Glpbio) according to the explanatory memorandum.mStatistical Analysis. Data were displayed as mean ± SD of 3 independent experiments. The diﬀerence evaluation between the two groups was performed by Student’s t test and among multiple groups was performed by one-way ANOVA in Graphpad Prism.

⦁RESULTS AND DISCUSSION
Synthesis and Characterizations of TDNs. As the following schematic illustration displayed (Figure 1a), TDNs were assembled by four equal amounts of single-strand DNA.29 The result of 3.5% agarose gel demonstrated the successful synthesis of TDNs. A, AB, and ABC were, respectively, single-stranded DNA, partially complemented double-stranded DNA, and triple-stranded DNA (Figure 1b). The TEM image showed that the successful synthesized TDNs were in the shape of pyramids (Figure 1c). Moreover, based on particle analyzer, the average particle size of TDNsmnature of TDNs. We loaded Cy5 dyes in one of the ssDNA to track the cellular uptake of TDNs by HK-2 cells.21 As the confocal microscopy image showed, a mass of Cy5-loaded TDNs were observed in the cytoplasm (Figure 2a). Next, after incubation with Cy5-TDNs for 1, 2, and 3 h, the HK-2 cells were detected by ﬂow cytometry (Figure 2b, c). Our results demonstrated that TDNs rapidly entered HK-2 cells and reached a peak at 2 h post-treatment. Next, after treatment of TDNs for 24 h, cell viability was examined to conﬁrm the cytotoxic eﬀects of TDNs. And we found that TDNs exhibited no toxic eﬀects on HK-2 cells (Figure 2d). These results demonstrated that TDNs rapidly entered HK-2 cells and exerted good biocompatibility in HK-2 cells.

TDN-Attenuated Ferroptosis Induced by RSL3. Recent studies have shown that the apoptosis and ferroptosis of renal tubular cells are involved in cisplatin-induced AKI.12 To explore whether TDNs can alleviate cisplatin-induced TDN-attenuated ferroptosis induced by RSL3. After treatment of RSL3 (0.5 μM) and TDNs (200 nM) for 24 h, (a) cell viability was determined, (b) the mRNA level of GPX4 was determined by real-time PCR, (c) the protein level of GPX4 was determined by Western blot,(d) and the formation of lipid ROS was examined using ﬂow cytometry. *p < 0.05, **p < 0.01, ***p < 0.001.
apoptosis and ferroptosis, we introduced a series of experiments. GPX4 inhibits ferroptosis by directly reducing lipid peroxidation in cells and RSL3 works as a typical ferroptosis inducer by inhibiting GPX4.30 Therefore, we treated HK-2 cells with RSL3 and detected the cell viability with CCK-8 assay. We found that RSL3 caused the death of HK-2 cells, and TDNs remarkably reduced it (Figure 3a). We then detected the Gpx4 mRNA level in the HK2 cells after treatment of RSL3 and TDNs, and found that RSL3 reduced the mRNA level of Gpx4, whereas TDNs restored it (Figure 3b). Our Western blot data also showed that GPX4 expression was down-regulated by RSL3, but TDNs restored its expression (Figure 3c). Accumulation of lipid ROS is one of the main features of ferroptosis, which leads to lipid peroxidation.31 The formation of lipid ROS caused by RSL3 was also signiﬁcantly increased determined by ﬂow cytometry. However, TDNs remarkably inhibited lipid ROS generation (Figure 3d). Taken together, these results indicated that TDNs reversed the GPX4 down-regulation and reduced the production of lipid ROS thus relieved RSL3-induced ferroptosis. These results provided a basis for further investigation on the roles of TDNs in cis-AKI.

TDNs Attenuated Ferroptosis Induced by Cisplatin. Next, we conducted subsequent experiments to explore the potential eﬀects of TDNs in cisplatin-induced damage. First of all, HK-2 cells were treated with a serial concentration gradient TDNs followed by cisplatin. The results indicated that 200 nM TDNs exerted the best eﬀect to protect the renal cells against cisplatin-induced cell death (Figure 4a), which was consistent with a previous report showing that TDNs at around 200 nM concentration displayed prominent eﬀects for cell migration and wound healing.26 Accordingly, we used 200 nM TDNs in the following experiments. We also treated HK-2 cells with cisplatin and TDNs, single- stranded (A), partially folded double-stranded (AB), and triple-stranded DNA (ABC) of TDNs.

Only TDNs showed signiﬁcant protective roles against cisplatin-induced cell death (Figure 4b). Recent research has reported that ferroptosis and caspase-mediated apoptosis are both involved in cisplatin-induced injury.13 To investigate the potential protective mechanism of TDNs during cell injury, we conducted treatment of cisplatin and TDNs, the ferroptosis inhibitor Fer-1, or the broad-spectrum caspase inhibitor Z- VAD-fmk. (Figure 4c). Our data showed the application of Fer-1 or Z-VAD-fmk had a protective eﬀect on cisplatin- treated HK-2 cells. Co-treatment with these two drugs had a more protective outcome than that of either drug alone. The above results indicated that ferroptosis and apoptosis were jointly involved in damage mediated by cisplatin, which was consistent with a previous report.12 To sum up, the data above demonstrated that TDNs reversed a cisplatin-induced injury involved with ferroptosis and apoptosis concurrently.

We then investigated what eﬀect TDNs exerted in cisplatin-induced ferroptosis. The mRNA level of Gpx4 was down-regulated after cisplatin treatment, whereas TDNs restored it (Figure 4d). The Western blot results conﬁrmed this outcome (Figure 4e). These data collectively indicated that TDNs conferred protection against cisplatin-induced ferroptosis cell death through enhancing GPX4 expression. It has been reported that TDNs up-regulate the Nrf2-AKT-HO- 1 pathway to alleviate myocardial ischemia-reperfusion injury and increase the expression of Gpx family members.22,24 Nrf2 increases the transcription of Gpx4.32 It remains to be further .TDN-attenuated ferroptosis induced by cisplatin. Cell viability was detected after treatment of (a) diﬀerent dosages of TDNs and cisplatin (20 μg/mL), (b) diﬀerent geometry of DNA structures and cisplatin (20 μg/mL), (c) cisplatin (20 μg/mL) and Fer-1 (3 μM) or Z- VAD-fmk (10 mg/mL) for 24 h. After treatment of cisplatin (20 μg/mL) and TDNs, (d) the mRNA level of Gpx4 was examined by real-time PCR, and (e) the protein level of GPX4 was examined by Western blot. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, ns, not signiﬁcant.explored whether TDNs inhibit ferroptosis by up-regulating Gpx4 transcription via the Nrf2-AKT pathway.TDNs Attenuated Cisplatin-Induced Apoptosis. Be- cause of the antiapoptotic eﬀects of TDNs, we next explored the eﬀect of TDNs on cisplatin-induced apoptosis in HK-2 cells.33,34 Apoptotic cells were assessed using PI/AnnexinV staining followed by ﬂow cytometry. The results (Figure 5a, b) showed that treatment of TDNs remarkably decreased apoptosis induced by cisplatin. Consistently, Western blot results indicated that TDNs also signiﬁcantly reduced cleavage of PARP (Figure 5c), a common marker of cell apoptosis. Taken together, TDNs had the protective potential for cisplatin-induced apoptosis. Research has indicated that DNA nanostructures combine with platinum drugs,35 and it remains to be further explored whether TDNs present protective eﬀects by absorbing cisplatin in HK-2 cells.Taking all of this into account, TDNs inhibited ferroptosis and apoptosis and thus attenuated cisplatin-induced injury.

⦁CONCLUSIONS
In conclusion, our study demonstrated that TDNs had
protective eﬀects on kidney cells. Our results certiﬁed that TDNs inhibited ferroptosis induced by RSL3 via decreasing the generation of lipid ROS and restoring the down- regulation of GPX4. Additionally, we demonstrated that TDNs attenuated cisplatin-induced ferroptosis by reversing the down-regulation of GPX4 and attenuated cisplatin- induced apoptosis by reducing the cleavage of PARP (Figure 6). To the best of our knowledge, this work is the ﬁrst study

TDN-attenuated apoptosis induced by cisplatin. After treatment of TDNs (200 nM) and cisplatin (20 μg/mL) for 24 h, (a)
representative ﬂuorescent images and (b) semiquantitative analysis of apoptotic cells, (c) and protein level of cleaved PARP as detected by Western blot. **p < 0.01, ***p < 0.001, ****p < 0.0001. TDN-attenuated cisplatin-induced injury in HK-2 cells. Cisplatin induced apoptosis and ferroptosis of HK-2 cells by enhancing PARP cleavage and GPX4 expression, respectively. Pretreatment with TDNs relieved the cisplatin-induced cell death by suppressing PARP cleavage and GPX4 expression melucidating that TDNs relieved RSL3-induced renal tubular damage and established the treatment of cisplatin-induced tubular cell death using TDNs. These ﬁndings pave the way toward a potential targeted therapeutic strategy for cisplatin- induced AKI.

⦁AUTHOR INFORMATION
Corresponding Authors
Yuanqing Zhang − School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou 510006, China;
Imageorcid.org/0000-0002-8761-5328; Email: zhangyq65@ mail.sysu.edu.cn
Minhao Wu − Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou 510080, China;
Email: [email protected]
Authors
Jiaying Li − Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou 510080, China
Liwen Wei − Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou 510080, China
Complete contact information is available at: https://pubs.acs.org/10.1021/acsabm.1c00294

Author Contributions
Y.Z. and M.W. proposed and supervised the project. M.W. and J.L. designed the experiments. J.L. and W.W. carried out the experiments and the analysis of results. J.L. and W.W. wrote the manuscript.
Notes
The authors declare no competing ﬁnancial interest.
⦁ ACKNOWLEDGMENTS
This work was supported by the National Natural Science
Foundation of China (82072087, 31670880, and 31970893), the Guangdong Natural Science Fund for Distinguished Young Scholars (2017A030306016 and 2016A030306004),
and the Fundamental Research Funds for the Central Universities (19ykzd39).

⦁ REFERENCES
(1) Rottenberg, S.; Disler, C.; Perego, P. (1) The rediscovery of
platinum-based cancer therapy. Nat. Rev. Cancer 2021, 21, 37−50.
(2) Dasari, S.; Bernard Tchounwou, P. Cisplatin in cancer therapy: molecular mechanisms of action. Eur. J. Pharmacol. 2014, 740, 364− 378.
(3) Wu, Z.; Gong, Q.; Yu, Y.; Zhu, J.; Li, W. Knockdown of circ- ABCB10 promotes sensitivity of lung cancer cells to cisplatin via miR-556−3p/AK4 axis. BMC Pulm. Med. 2020, 20, 10.
(4) Holditch, S. J.; Brown, C. N.; Lombardi, A. M.; Nguyen, K. N.;
Edelstein, C. L. Recent Advances in Models, Mechanisms, Biomarkers, and Interventions in Cisplatin-Induced Acute Kidney Injury. Int. J. Mol. Sci. 2019, 20, 3011−3036.
(5) Chen, Q.; Ma, J.; Yang, X.; Li, Q.; Lin, Z.; Gong, F.
SIRT1Mediates Effects of FGF21 to Ameliorate Cisplatin-Induced Acute Kidney Injury. Front. Pharmacol. 2020, 11, 241.
(6) Oh, G.-S.; Kim, H.-J.; Shen, A.; Lee, S. B.; Khadka, D.; Pandit, A.; So, H.-S. Cisplatin-induced Kidney Dysfunction and Perspectives on Improving Treatment Strategies. Electrolyte Blood Pressure 2014, 12, 55−65.
(7) Deng, F.; Sharma, I.; Dai, Y.; Yang, M.; Kanwar, Y. S. Myo-
inositol oxygenase expression profile modulates pathogenic ferrop- tosis in the renal proximal tubule. J. Clin. Invest. 2019, 129, 5033− 5049.
(8) Sharp, C. N.; Doll, M. A.; Dupre, T. V.; Shah, P. P.; Subathra, M.; Siow, D.; Arteel, G. E.; Megyesi, J.; Beverly, L. J.; Siskind, L. J. Repeated administration of low-dose cisplatin in mice induces fibrosis. Am. J. Physiol. Renal Physiol. 2016, 310, 560−568.
(9) Basu, A.; Krishnamurthy, S. Cellular Responses to Cisplatin-
Induced DNA Damage. J. Nucleic Acids 2010, 2010, No. 201367.
(10) Zeng, X.; Wang, H.; He, D.; Jia, W.; Ma, R. LIMD1 Increases the Sensitivity of Lung Adenocarcinoma Cells to Cisplatin via the GADD45α/p38 MAPK Signaling Pathway. Front. Oncol. 2020, 10, 969.
(11) Yoon, S. Y.; Yoon, J. A.; Park, M.; Shin, E.-Y.; Jung, S.; Lee, J. E.; Eum, J. H.; Song, H.; Lee, D. R.; Lee, W. S.; Lyu, S. W. Recovery of ovarian function by human embryonic stem cell-derived mesenchymal stem cells in cisplatin-induced premature ovarian failure in mice. Stem Cell Res. Ther. 2020, 11, 255.
(12) Hu, Z.; Zhang, H.; Yi, B.; Yang, S.; Liu, J.; Hu, J.; Wang, J.; Cao, K.; Zhang, W. VDR activation attenuate cisplatin induced AKI by inhibiting ferroptosis. Cell Death Dis. 2020, 11, 1−11.
(13) Guo, J.; Xu, B.; Han, Q.; Zhou, H.; Xia, Y.; Gong, C.; Dai, X.;
Li, Z.; Wu, G. Ferroptosis: A Novel Anti-tumor Action for Cisplatin.
Cancer Res. Treat. 2018, 50, 445−460.
(14) Pinheiro, A. V.; Han, D.; Shih, W. M.; Yan, H. Challenges
and opportunities for structural DNA nanotechnology. Nat. Nanotechnol. 2011, 6, 763−72.
(15) Chang, M.; Yang, C. S.; Huang, D. M. Aptamer-conjugated
DNA icosahedral nanoparticles as a carrier of doxorubicin for cancer therapy. ACS Nano 2011, 5, 6156−63.
(16) Chiu, Y. T. E.; Li, H. Z.; Choi, C. H. J. Progress toward
Understanding the Interactions between DNA Nanostructures and the Cell. Small 2019, 15, 1805416.
(17) Kim, K. R.; Kim, D. R.; Lee, T.; Yhee, J. Y.; Kim, B. S.; Kwon, I. C.; Ahn, D. R. Drug delivery by a self-assembled DNA tetrahedron for overcoming drug resistance in breast cancer cells. Chem. Commun. 2013, 49, 2010−2012.
(18) Lee, J. B.; Hong, J.; Bonner, D. K.; Poon, Z.; Hammond, P.
T. Self-assembled RNA interference microsponges for efficient siRNA delivery. Nat. Mater. 2012, 11, 316−322.
(19) Li, Q.; Zhao, D.; Shao, X.; Lin, S.; Xie, X.; Liu, M.; Ma, W.;
Shi, S.; Lin, Y. Aptamer-modified tetrahedral DNA nanostructure for tumor-targeted drug delivery. ACS Appl. Mater. Interfaces 2017, 9, 36695−36701.
(20) Radovic-Moreno, A. F.; Chernyak, N.; Mader, C. C.;
Nallagatla, S.; Kang, R. S.; Hao, L.; Walker, D. A.; Halo, T. L.; Merkel, T. J.; Rische, C. H.; Anantatmula, S.; Burkhart, M.; Mirkin,

C. A.; Gryaznov, S. M. Immunomodulatory spherical nucleic acids.
Proc. Natl. Acad. Sci. U. S. A. 2015, 112, 3892−3897.
(21) Liang, L.; Li, J.; Li, Q.; Huang, Q.; Shi, J.; Yan, H.; Fan, C. Single-particle tracking and modulation of cell entry pathways of a tetrahedral DNA nanostructure in live cells. Angew. Chem., Int. Ed. 2014, 53, 7745−7750.
(22) Peng, Q.; Shao, X. R.; Xie, J.; Shi, S. R.; Wei, X.; Zhang, T.;
Cai, X.; Lin, Y. Understanding the Biomedical Effects of the Self- Assembled Tetrahedral DNA Nanostructure on Living Cells. ACS Appl. Mater. Interfaces 2016, 8, 12733−12739.
(23) Zhang, Q.; Lin, S.; Shi, S.; Zhang, T.; Ma, Q.; Tian, T.; Zhou,
T.; Cai, X.; Lin, Y. Anti-inflammatory and Antioxidative Effects of Tetrahedral DNA Nanostructures via the Modulation of Macro- phage Responses. ACS Appl. Mater. Interfaces 2018, 10, 3421−3430.
(24) Zhang, M.; Zhu, J.; Qin, X.; Zhou, M.; Zhang, X.; Gao, Y.;
Zhang, T.; Xiao, D.; Cui, W.; Cai, X. Cardioprotection of Tetrahedral DNA Nanostructures in Myocardial Ischemia-Reperfu- sion Injury. ACS Appl. Mater. Interfaces 2019, 11, 30631−30639.
(25) Lin, S.; Zhang, Q.; Li, S.; Zhang, T.; Wang, L.; Qin, X.;
Zhang, M.; Shi, S.; Cai, X. Antioxidative and Angiogenesis- Promoting Effects of Tetrahedral Framework Nucleic Acids in Diabetic wound healing with activation of the Akt/Nrf2/HO-1 pathway. ACS Appl. Mater. Interfaces 2020, 12, 11397−11408.
(26) Liu, N.; Zhang, X.; Li, N.; Zhou, M.; Zhang, T.; Li, S.; Cai,
X.; Ji, P.; Lin, Y. Tetrahedral Framework Nucleic Acids Promote Corneal Epithelial Wound Healing in Vitro and in Vivo. Small 2019, 15, 1901907.
(27) Jiang, D.; Ge, Z.; Im, H. J.; England, C. G.; Ni, D.; Hou, J.; Zhang, L.; Kutyreff, C. J.; Yan, Y.; Liu, Y.; Cho, S. Y.; Engle, J. W.; Shi, J.; Huang, P.; Fan, C.; Yan, H.; Cai, W. DNA origami nanostructures can exhibit preferential renal uptake and alleviate acute kidney injury. Nat. Biomed. Eng. 2018, 2, 865−877.
(28) Zhuang, J.; Tan, J.; Wu, C.; Zhang, J.; Liu, T.; Fan, C.; Li, J.;
Zhang, Y. Extracellular vesicles engineered with valency-controlled DNA nanostructures deliver CRISPR/Cas9 system for gene therapy. Nucleic Acids Res. 2020, 48, 8870−8882.
(29) Zhang, Y.; Ma, W.; Zhu, Y.; Shi, S.; Li, Q.; Mao, C.; Zhao,
D.; Zhan, Y.; Shi, J.; Li, W.; et al. Inhibiting Methicillin-Resistant Staphylococcus aureus by Tetrahedral DNA Nanostructure-enabled Antisense Peptide Nucleic Acid Delivery. Nano Lett. 2018, 18, 5652−5659.
(30) Fujiki, K.; Inamura, H.; Sugaya, T.; Matsuoka, M. Blockade of
ALK4/5 signaling suppresses cadmium- and erastin-induced cell death in renal proximal tubular epithelial cells via distinct signaling mechanisms. Cell Death Differ. 2019, 26, 2371−2385.
(31) Li, J.; Cao, F.; Yin, H.-l.; Huang, Z.-j.; Lin, Z.-t.; Mao, N.;
Sun, B.; Wang, G. Ferroptosis: past, present and future. Cell Death Dis. 2020, 11, 1−13.
(32) Abdalkader, M.; Lampinen, R.; Kanninen, K. M.; Malm, T.
M.; Liddell, J. R. Targeting Nrf2 to Suppress Ferroptosis and Mitochondrial Dysfunction in RSL3 Neurodegeneration. Front. Neurosci. 2018, 12, 466.
(33) Qin, X.; Li, N.; Zhang, M.; Lin, S.; Zhu, J.; Xiao, D.; Cui, W.; Zhang, T.; Lin, Y.; Cai, X. Tetrahedral framework nucleic acids prevent retina ischemia-reperfusion injury from oxidative stress via activating the Akt/Nrf2 pathway. Nanoscale 2019, 11, 20667−20675.
(34) Shao, X.; Cui, W.; Xie, X.; Ma, W.; Zhan, Y.; Lin, Y.
Treatment of Alzheimer’s disease with framework nucleic acids. Cell Proliferation 2020, 53, e12787.
(35) Ma, Y.; Wang, Z.; Ma, Y.; Han, Z.; Zhang, M.; Chen, H.; Gu,
Y. A telomerase-responsive DNA icosahedron for precise delivery of platinum nanodrugs to cisplatin-resistant cancer. Angew. Chem., Int. Ed. 2018, 57, 5389−5393.