Analysis of the biologic functions of H- and L-ferritins in HeLa cells by transfection with siRNAs and cDNAs: evidence for a proliferative role of L-ferritin
Open accessAbstract
We describe the use of small interfering RNAs (siRNAs) to down-regulate H- and L-ferritin levels in HeLa cells. siRNAs repressed H- and L-ferritin expression to about 20% to 25% of the background level in both stable and transient transfections. HeLa cells transfected with H- and L-ferritin cDNAs were analyzed in parallel to compare the effects of ferritin up- and down-regulation. We found that large modifications of L-ferritin levels did not affect iron availability in HeLa cells but positively affected cell proliferation rate in an iron-independent manner. The transient down-regulation of H-ferritin modified cellular iron availability and resistance to oxidative damage, as expected. In contrast, the stable suppression of H-ferritin in HeLa cell clones transfected with siRNAs did not increase cellular iron availability but made cells less resistant to iron supplementation and chelation. The results indicate that L-ferritin has no direct effects on cellular iron homeostasis in HeLa cells, while it has new, iron-unrelated functions. In addition, they suggest that H-ferritin function is to act as an iron buffer.
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HeLa cells expressing mutant ferritin L chains exhibit a phenotype of iron loading.
“In addition, HeLa cells that express mutant ferritin L chains exhibited a phenotype of iron loading (61).”
From Intracellular iron transport and storage: from molecular mechanisms to health implications (2008) · cites this paper as [61]
Recombinant mutant ferritin L can assemble into 24-subunit ferritin shells, but efficiency of assembly and capacity to incorporate iron are decreased in vitro.
“Studies with recombinant mutant ferritin L showed that, although it was able to assemble into 24-subunit ferritin shells, the efficiency of assembly and capacity to incorporate iron were both decreased in vitro (61).”
From Intracellular iron transport and storage: from molecular mechanisms to health implications (2008) · cites this paper as [61]
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A novel view of ferritin in cancer
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Wu, T., Li, Y., Liu, B., Zhang, S., Wu, L., Zhu, X., & Chen, Q. (2016) · PLoS ONE · doi:10.1371/journal.pone.0149361 · Cited by 71 · see citing works
“FTL levels were previously reported to positively modulate the rate of HeLa cell proliferation [7].”
BackgroundH-Ferritin is essential for macrophages’ capacity to store or detoxify exogenously added iron
Mesquita, G., Silva, T., Gomes, A. C., Oliveira, P. F., Alves, M. G., Fernandes, R., Almeida, A. A., Moreira, A. C., & Gomes, M. S. (2020) · Scientific Reports · doi:10.1038/s41598-020-59898-0 · Cited by 70 · see citing works
Dissecting plant iron homeostasis under short and long-term iron fluctuations
Darbani, B., Briat, J., Holm, P. B., Husted, S., Noeparvar, S., & Borg, S. (2013) · Biotechnology Advances · doi:10.1016/j.biotechadv.2013.05.003 · Cited by 68 · see citing works
Wu, M., Zhao, Y., Zhang, C., & Pu, K. (2024) · ACS Nano · doi:10.1021/acsnano.4c09800 · Cited by 65 · see citing works
Oxidative stress caused by lead (Pb) induces iron deficiency in Drosophila melanogaster
Liu, Z.-H., Shang, J., Yan, L., Tian, W., Xiang, L., Wang, H.-L., Cheng, J., & Xiao, G. (2019) · Chemosphere · doi:10.1016/j.chemosphere.2019.125428 · Cited by 64 · see citing works
Ferritin heavy chain mediates the protective effect of heme oxygenase-1 against oxidative stress
Cheng, H., Yen, C., Chang, C.-C., Huang, K.-T., Chen, K.-H., Zhang, R.-Y., Lee, P.-Y., Miaw, S., Huang, J., Chiang, C., et al. (2015) · Biochimica et Biophysica Acta (BBA) - General Subjects · doi:10.1016/j.bbagen.2015.09.018 · Cited by 62 · see citing works
Neurodegeneration with Brain Iron Accumulation
Schneider, S. A. (2016) · Current Neurology and Neuroscience Reports · doi:10.1007/s11910-015-0608-3 · Cited by 61 · see citing works
Haldar, S., Bevers, L. E., Tosha, T., & Theil, E. C. (2011) · Journal of Biological Chemistry · doi:10.1074/jbc.m110.205278 · Cited by 60 · see citing works
Ferritin binds and activates p53 under oxidative stress
Lee, J., Jang, H., Cho, E., & Youn, H. (2009) · Biochemical and Biophysical Research Communications · doi:10.1016/j.bbrc.2009.08.125 · Cited by 58 · see citing works
Paired Bacillus anthracis Dps (Mini-ferritin) Have Different Reactivities with Peroxide
Liu, X., Kim, K., Leighton, T., & Theil, E. C. (2006) · Journal of Biological Chemistry · doi:10.1074/jbc.m601398200 · Cited by 58 · see citing works
Zheng, W., Hu, Y., & Sun, L. (2010) · Comparative Biochemistry and Physiology Part B Biochemistry and Molecular Biology · doi:10.1016/j.cbpb.2010.03.012 · Cited by 56 · see citing works
Lopez, M., Sánchez‐Más, J., Pascual-Figal, D. A., Abenza, S., Pérez‐Martínez, M. T., Valdés, M., & Lax, A. (2012) · Free Radical Biology and Medicine · doi:10.1016/j.freeradbiomed.2012.09.009 · Cited by 55 · see citing works
Luscieti, S., Santambrogio, P., Langlois d'Estaintot, B., Granier, T., Cozzi, A., Poli, M., Gallois, B., Finazzi, D., Cattaneo, A., Levi, S., et al. (2010) · Journal of Biological Chemistry · doi:10.1074/jbc.m109.096404 · Cited by 51 · see citing works
Self-citationRole and regulation of ferritin H in rotenone-mediated mitochondrial oxidative stress
Mackenzie, E. L., Ray, P. D., & Tsuji, Y. (2008) · Free Radical Biology and Medicine · doi:10.1016/j.freeradbiomed.2008.01.031 · Cited by 53 · see citing works
Cozzi, A., Santambrogio, P., Privitera, D., Broccoli, V., Rotundo, L. I., Garavaglia, B., Benz, R., Altamura, S., Goede, J. S., Muckenthaler, M. U., et al. (2013) · Journal of Experimental Medicine · doi:10.1084/jem.20130315 · Cited by 47 · see citing works
“, 2000) and RNA interference studies in HeLa cells (Cozzi et al., 2004), indicated that alterations of FtL levels do not affect cellular iron homeostasis or the sensitivity to oxidative stress.”
BackgroundSelf-citationLudwiczek, S., Theurl, I., Bahram, S., Schümann, K., & Weiß, G. (2005) · Journal of Cellular Physiology · doi:10.1002/jcp.20315 · Cited by 50 · see citing works
“(Cozzi et al., 2004).”
Cytoprotective effects of ferritin on doxorubicin-induced breast cancer cell death
Buranrat, B., & Connor, J. R. (2015) · Oncology Reports · doi:10.3892/or.2015.4250 · Cited by 48 · see citing works
“It has recently been shown that the repression of ferritin by siRNA increases the chemosensitivity of HeLa cells (11) and human glioma (19).”
BackgroundInfluentialLi, Z., Liu, J., Chen, H., Zhang, Y., Shi, H., Huang, L., Tao, J., Shen, R., & Wang, T. (2019) · Carcinogenesis · doi:10.1093/carcin/bgz181 · Cited by 48 · see citing works
α‐Synuclein in blood cells differentiates Parkinson’s disease from healthy controls
Elhadi, S. A., Grigoletto, J., Poli, M., Arosio, P., Arkadir, D., & Sharon, R. (2019) · Annals of Clinical and Translational Neurology · doi:10.1002/acn3.50944 · Cited by 46 · see citing works
“H-ferritin: determined by sandwich-ELISA, with a monoclonal antibody rH02 as described previously.(26) A standard curve consisting of a recombinant homo-polymer H ferritin was used for reference amount.”
MethodSelf-citationLoss of the von Hippel Lindau Tumor Suppressor Disrupts Iron Homeostasis in Renal Carcinoma Cells
Alberghini, A., Recalcati, S., Tacchini, L., Santambrogio, P., Campanella, A., & Cairo, G. (2005) · Journal of Biological Chemistry · doi:10.1074/jbc.m500971200 · Cited by 41 · see citing works
Self-citationMcCullough, K., & Bolisetty, S. (2020) · Seminars in Nephrology · doi:10.1016/j.semnephrol.2020.01.007 · Cited by 40 · see citing works
Cozzi, A., Santambrogio, P., Corsi, B., Campanella, A., Arosio, P., & Levi, S. (2006) · Neurobiology of Disease · doi:10.1016/j.nbd.2006.05.004 · Cited by 38 · see citing works
Self-citationZiv, K., Meir, G., Harmelin, A., Shimoni, E., Klein, E., & Neeman, M. (2010) · NMR in Biomedicine · doi:10.1002/nbm.1491 · Cited by 38 · see citing works
“Indeed previous studies showed varied response to elevated expression of h-ferritin ranging from protection of cells from reactive oxygen species (18,19), to regulation of free iron (20) and increased susceptibility to neurodegeneration in aging mice (21).”
BackgroundAl-Qenaei, A., Yiakouvaki, A., Reelfs, O., Santambrogio, P., Levi, S., Hall, N. D., Tyrrell, R. M., & Pourzand, C. (2013) · Free Radical Biology and Medicine · doi:10.1016/j.freeradbiomed.2013.12.006 · Cited by 37 · see citing works
“Accordingly FtL knockdown did not have a significant impact on intracellular iron availability [63].”
BackgroundSelf-citationRen, C., Chen, T., Jiang, X., Wang, Y., & Hu, C. (2014) · Fish & Shellfish Immunology · doi:10.1016/j.fsi.2014.03.022 · Cited by 37 · see citing works
Repression of ferritin light chain translation by human eIF3
Pulos-Holmes, M. C., Srole, D. N., Lee, A. S. Y., Juarez, M. G., McSwiggen, D. T., Ingolia, N. T., & Cate, J. H. D. (2019) · eLife · doi:10.7554/elife.48193 · Cited by 34 · see citing works
“Primers for the qPCR were as follows: FTL forward: 5’ -ACCTCTCTCTGGGCTTCTAT-3’, FTL reverse: 5’ -AGCTGGC TTCTTGATGTCCT-3’ (Cozzi et al., 2004), ACTB forward: 5’-CTCTTCCAGCCTTCCTTCCT-3’, ACTB reverse: 5’-AGCACTGTGTTGGCGTACAG-3’ (Chen et al., 2008), PSMB6 forward: 5’-GGACTCCAGAACAACCACTG-3’, PSMB6 reverse: 5’-CAGCTGAGCC…”
MethodNeurons lacking iron regulatory protein-2 are highly resistant to the toxicity of hemoglobin
Regan, R. F., Chen, M., Li, Z., Zhang, X., Benvenisti‐Zarom, L., & Chen‐Roetling, J. (2008) · Neurobiology of Disease · doi:10.1016/j.nbd.2008.04.008 · Cited by 33 · see citing works
“H-ferritin knockdown markedly increased HeLa cell vulnerability to hydrogen peroxide, while upregulation was protective; modulation of L-ferritin levels had no significant effect (Cozzi et al., 2004).”
Gallì, A., Bergamaschi, G., Recalde, H. R., Biasiotto, G., Santambrogio, P., Boggi, S., Levi, S., Arosio, P., & Cazzola, M. (2004) · British Journal of Haematology · doi:10.1111/j.1365-2141.2004.05238.x · Cited by 32 · see citing works
“Recent studies support the hypothesis that ferroportin plays an important role in macrophage recycling of iron from senescent red blood cells (Yang et al, 2002; Knutson et al, 2003; Ludwiczek et al, 2003; Cozzi et al, 2004).”
BackgroundResultSelf-citationFerritin L is the Sole Serum Ferritin Constituent and a Positive Hepatic Acute-Phase Protein
Naz, N., Moriconi, F., Ahmad, S., Amanzada, A., Khan, S., Mihm, S., Ramadori, G., & Malik, I. A. (2013) · Shock · doi:10.1097/shk.0b013e31829266b9 · Cited by 32 · see citing works
“tissue-specific regulations, whereas multiple conditions activate FTH gene transcription (7, 8), including cell differentiation, changes in the cell proliferation status (9), cytokines, and heme.”
BackgroundIRP1-independent alterations of cardiac iron metabolism in doxorubicin-treated mice
Corna, G., Galy, B., Hentze, M. W., & Cairo, G. (2006) · Journal of Molecular Medicine · doi:10.1007/s00109-006-0068-y · Cited by 31 · see citing works
“Further evidence for this protective role of ferritin stems from the demonstration of higher H2O2 toxicity in HeLa cells depleted of ferritin H-chain by small interfering RNAs [38].”
BackgroundH ferritin silencing induces protein misfolding in K562 cells: A Raman analysis
Zolea, F., Biamonte, F., Candeloro, P., Sanzo, M. D., Cozzi, A., Vito, A. D., Quaresima, B., Lobello, N., Trecroci, F., Fabrizio, E. M. D., et al. (2015) · Free Radical Biology and Medicine · doi:10.1016/j.freeradbiomed.2015.07.161 · Cited by 30 · see citing works
“In this work, we confirmed this ferritin primary role and uncovered an unexpected and interesting effect on protein folding in K562 cells caused by the silencing of the H-ferritin chain.”
ResultInfluentialSelf-citationPicard, É., Ranchon‐Cole, I., Jonet, L., Beaumont, C., Béhar‐Cohen, F., Courtois, Y., & Jeanny, J. (2010) · Investigative Ophthalmology & Visual Science · doi:10.1167/iovs.10-5705 · Cited by 30 · see citing works
Keogh, M. J., Morris, C. M., & Chinnery, P. F. (2013) · International review of neurobiology · doi:10.1016/b978-0-12-410502-7.00006-5 · Cited by 28 · see citing works
Ahmad, S., Moriconi, F., Naz, N., Sultan, S., Sheikh, N., Ramadori, G. P., & Malik, I. A. (2013) · Europe PMC (PubMed Central) · Cited by 28 · see citing works
Silva-Gomes, S., Bouton, C., Silva, T., Santambrogio, P., Rodrigues, P., Appelberg, R., & Gomes, M. S. (2013) · PLoS ONE · doi:10.1371/journal.pone.0082874 · Cited by 26 · see citing works
“On the other hand, H-ferritin has also been implicated in protection against oxidative stress [32,33] and apoptosis [34].”
BackgroundSelf-citationKe, S., Wang, C., Su, Z., Lin, S., & Wu, G. (2022) · Frontiers in Genetics · doi:10.3389/fgene.2022.897683 · Cited by 26 · see citing works
Robach, P., Recalcati, S., Girelli, D., Campostrini, N., Kempf, T., Wollert, K. C., Corbella, M., Santambrogio, P., Perbellini, L., Brasse‐Lagnel, C., et al. (2013) · European Journal Of Haematology · doi:10.1111/ejh.12122 · Cited by 25 · see citing works
“Ferritin concentrations were determined in cell lysates by ELISA based on monoclonal antibodies specific for the human H and L subunits, as previously described (32).”
MethodSelf-citationBehavioral Characterization of Mouse Models of Neuroferritinopathy
Capoccia, S., Maccarinelli, F., Buffoli, B., Rodella, L. F., Cremona, O., Arosio, P., & Cirulli, F. (2015) · PLoS ONE · doi:10.1371/journal.pone.0118990 · Cited by 24 · see citing works
Self-citationFerritin as prognostic marker in multiple myeloma patients undergoing autologous transplantation
Strasser‐Weippl, K., & Ludwig, H. (2014) · Leukemia & lymphoma/Leukemia and lymphoma · doi:10.3109/10428194.2014.891025 · Cited by 23 · see citing works
Chapter Five – Neuroferritinopathy
Keogh, M. J., Morris, C. M., & Chinnery, P. F. (2013) · International review of neurobiology · Cited by 21 · see citing works
3‐Hydroxybutyrate dehydrogenase‐2 and ferritin‐H synergistically regulate intracellular iron
Liu, Z., Velpula, K. K., & Devireddy, L. R. (2014) · FEBS Journal · doi:10.1111/febs.12794 · Cited by 22 · see citing works
Ferritin as an Effective Prognostic Factor and Potential Cancer Biomarker
Szymulewska-Konopko, K., Reszeć-Giełażyn, J., & Małeczek, M. (2025) · Current Issues in Molecular Biology · doi:10.3390/cimb47010060 · Cited by 25 · see citing works
“It is involved in the proliferation [63,64], angiogenesis [64], and provision of iron [65,66].”
Pathophysiology of perioperative anaemia
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Neurodegeneration with Brain Iron Accumulation
Schneider, S. A. (2014) · Elsevier eBooks · doi:10.1016/b978-0-12-410529-4.00042-5 · Cited by 19 · see citing works
Yu, H., Zhang, S.-Z., Ma, Y., Fei, D.-Q., Li, B., Yang, L.-A., Wang, J., Li, Z., Muhammad, A., & Xu, J. (2017) · International Journal of Molecular Sciences · doi:10.3390/ijms18102126 · Cited by 17 · see citing works
“Treatment of HeLa cells with H2O2 could produce ROS, whereas overexpression of ferritin could reduce the cellular levels of ROS and oxidative stress [51].”
BackgroundHe, X., Zhang, Y., Wu, X., Xiao, S., & Yu, Z. (2010) · Molecular Biology Reports · doi:10.1007/s11033-010-0339-0 · Cited by 16 · see citing works
“In mammalian cells, over-expression of ferritin H subunit shows an enhanced ability to reduce anti-oxidative stress [12].”
BackgroundHarned, J., Ferrell, J., Lall, M. M., Fleisher, L. N., Nagar, S., Goralska, M., & McGahan, M. C. (2010) · Investigative Ophthalmology & Visual Science · doi:10.1167/iovs.09-3861 · Cited by 16 · see citing works
“Indeed, transfection of HeLa cells with siRNA for H- or L-chain ferritin far more effectively reduced ferritin chain synthesis than did antisense treatment.(9) In this earlier study, FLsiRNA had no direct effect on cellular iron homeostasis but enhanced cellular proliferation.”
BackgroundInfluentialFerritin Expression in Rat Hepatocytes and Kupffer Cells after Lead Nitrate Treatment
Yang, F., Yamada, T., Shimizu, T., Nanashima, N., Akita, M., Suto, K., & Tsuchida, S. (2009) · Toxicologic Pathology · doi:10.1177/0192623308328544 · Cited by 16 · see citing works
“1995), cell proliferation (Cozzi et al. 2004; Kikyo et al. 1994), and apoptosis (Cozzi et al.”
BackgroundIncreasing expression of H- or L-ferritin protects cortical astrocytes from hemin toxicity
Li, Z., Chen‐Roetling, J., & Regan, R. F. (2009) · Free Radical Research · doi:10.1080/10715760902942808 · Cited by 16 · see citing works
“[8] reported that iron availability was negatively regulated by increasing expression of H-ferritin by gene transfer, but that L-ferritin had no effect per se.”
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FTL: a novel predictor in gastric cancer.
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Iron Binding in the Ferroxidase Site of Human Mitochondrial Ferritin
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The Chemistry of Nature's Iron Biominerals in Ferritin Protein Nanocages
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- Authors
- Cozzi, A., Corsi, B., Levi, S., Santambrogio, P., Biasiotto, G., & Arosio, P.
- Affiliations
- IRCCS San Raffaele Roma (IT)
- Venue
- Blood
- Publisher
- American Society of Hematology
- Published
- 2004-03-15
- Volume
- 103
- Issue
- 6
- Pages
- 2377-2383
- ISSN
- 0006-4971 (ISSN-L)
- 1528-0020
- OpenAlex
- W1519013154
- Cited by
- 132
- References
- 43
- Citations by year
- 2026: 2 · 2025: 2 · 2024: 3 · 2023: 7 · 2022: 3 · 2021: 2 · 2020: 6 · 2019: 5 · 2018: 1 · 2017: 3 · 2016: 4 · 2015: 7 · 2014: 9 · 2013: 21 · 2012: 4
- FWCI
- 3.07
- Type
- journal article
- Language
- en
- Open access
- Licence
- other-oa
- Locations
- Blood (publishedVersion)
- PubMed (publishedVersion)
- INFM-OAR (INFN Catania) (submittedVersion, other-oa)
- Topics
- Health Sciences › Medicine › Hematology › Iron Metabolism and Disorders
- Health Sciences › Medicine › Pulmonary and Respiratory Medicine › Ferroptosis and cancer prognosis
- Life Sciences › Neuroscience › Neurology › Neurological diseases and metabolism
- Keywords
- H-ferritin, L-ferritin, HeLa cells, cell proliferation, iron homeostasis, iron availability, iron supplementation, chelation, siRNA knockdown
- MeSH terms
- Quaternary Ammonium Compounds, Buffers, Cell Division, Ferric Compounds, Ferritins, HeLa Cells, Humans, Iron, Transfection, Down-Regulation, Up-Regulation, Gene Expression, DNA, Complementary, Oxidative Stress, RNA, Small Interfering
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