Effects of the p16/cyclin D1/CDK4/Rb/E2F1 pathway on aberrant lung fibroblast proliferation in neonatal rats exposed to hyperoxia

Zhao,Shimeng; Chen,Zhiguang; Han,Shuang; Wu,Hongmin

doi:10.3892/etm.2021.10491

Effects of the p16/cyclin D1/CDK4/Rb/E2F1 pathway on aberrant lung fibroblast proliferation in neonatal rats exposed to hyperoxia

Authors:
- Shimeng Zhao
- Zhiguang Chen
- Shuang Han
- Hongmin Wu
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Affiliations: Department of Neonatology, The First Affiliated Hospital of China Medical University, Shenyang, Liaoning 110001, P.R. China
Published online on: July 26, 2021 https://doi.org/10.3892/etm.2021.10491
Article Number: 1057

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Abstract

p16^INK4a (p16) inhibits the vital G₁ to S phase transition during cell cycle progression through the p16/cyclin D1/CDK4/retinoblastoma(Rb)/E2F1 pathway. Hyperoxia can suppress the G₁/S checkpoint and induce more lung fibroblasts (LFs) to transition from the G₁ phase to the S phase and undergo cell proliferation. The present study investigated the rate of p16 gene promoter methylation and the protein expression levels of p16, cyclin D1, CDK4, Rb and E2F1 in LFs from the lungs of rats exposed to hyperoxia and normoxia on postnatal days 3, 7 and 14. In the hyperoxia‑exposed group, the methylation rate was 50 and 80% on days 7 and 14, respectively. Cyclin D1 and CDK4 overexpression was associated with p16 loss and Rb inactivation by phosphorylation. Rb phosphorylation induced E2F1 release in the G₁ phase, which promoted cell proliferation. No methylation was observed in the normoxia‑exposed group. These observations suggested that p16 loss may stimulate aberrant LF proliferation via the p16/cyclin D1/CDK4/Rb/E2F1 pathway.

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1	Chen CM: Therapy for neonatal hyperoxia-induced lung injury. Pediatr Neonatol. 55:329–330. 2014.PubMed/NCBI View Article : Google Scholar
2	Crapo JD, Barry BE, Foscue HA and Shelburne J: Structural and biochemical changes in rat lungs occurring during exposures to lethal and adaptive dose of oxygen. Am Rev Respir Dis. 122:123–143. 1980.PubMed/NCBI View Article : Google Scholar
3	Reddy GP: Cell cycle: Regulatory events in G1->S transition of mammalian cells. J Cell Biochem. 54:379–386. 1994.PubMed/NCBI View Article : Google Scholar
4	Al-Khalaf HH, Colak D, Al-Saif M, Al-Bakheet A, Hendrayani SF, Al-Yousef N, Kaya N, Khabar KS and Aboussekhra A: p16(INK4a) positively regulates cyclin D1 and E2F1 through negative control of AUF1. PLoS One. 6(e21111)2011.PubMed/NCBI View Article : Google Scholar
5	Vogelstein B and Kinzler KW: Cancer genes and the pathways they control. Nat Med. 10:789–799. 2004.PubMed/NCBI View Article : Google Scholar
6	Haller F, Löbke C, Ruschhaupt M, Cameron S, Schulten HJ, Schwager S, von Heydebreck A, Gunawan B, Langer C, Ramadori G, et al: Loss of 9p leads to p16(INK4A) down-regulation and enables RB/E2F1-dependent cell cycle promotion in gastrointestinal stromal tumours (GISTs). J Pathol. 215:253–262. 2008.PubMed/NCBI View Article : Google Scholar
7	Wu Z and Yu Q: E2F1-mediated apoptosis as a target of cancer therapy. Curr Mol Pharmacol. 2:149–160. 2009.PubMed/NCBI View Article : Google Scholar
8	Giacinti C and Giordano A: RB and cell cycle progression. Oncogene. 25:5220–5227. 2006.PubMed/NCBI View Article : Google Scholar
9	Tyagi E, Liu B, Li C, Liu T, Rutter J and Grossman D: Loss of p16 INK4A stimulates aberrant mitochondrial biogenesis through a CDK4/Rb-independent pathway. Oncotarget. 8:55848–55862. 2017.PubMed/NCBI View Article : Google Scholar
10	Zhao SM, Wu HM, Cao ML and Hna D: Primary culture of lung fibroblasts from hyperoxia-exposed rats and a proliferative characteristics study. Cytotechnology. 70:751–760. 2018.PubMed/NCBI View Article : Google Scholar
11	Zhao S, Cao M, Wu H, Hu Y and Xue X: 5-aza-2'-deoxycytidine inhibits the proliferation of lung fibroblasts in neonatal rats exposed to hyperoxia. Pediatr Neonatol. 58:122–127. 2017.PubMed/NCBI View Article : Google Scholar
12	Chen X, Zhang X and Pan J: Effect of montelukast on bronchopulmonary dysplasia (BPD) and related mechanisms. Med Sci Monit. 25:1886–1893. 2019.PubMed/NCBI View Article : Google Scholar
13	Qi XJ, Ning W, Xu F, Dang HX, Fang F and Li J: Fasudil, an inhibitor of Rho-associated coiled-coil kinase, attenuates hyperoxia-induced pulmonary fibrosis in neonatal rats. Int J Clin Exp Pathol. 8:12140–12150. 2015.PubMed/NCBI
14	Hu Y, Fu J and Xue X: Association of the proliferation of lung fibroblasts with the ERK1/2 signaling pathway in neonatal rats with hyperoxia-induced lung fibrosis. Exp Ther Med. 17:701–708. 2019.PubMed/NCBI View Article : Google Scholar
15	Herman JG, Graff JR, Myohanen S, Nelkin BD and Baylin SB: Methylation specific PCR: A novel PCR assay for methylation status of CpG islands. Proc Natl Acad Sci USA. 93:9821–9826. 1996.PubMed/NCBI View Article : Google Scholar
16	Huang Y, Wu G, Fan H, Ye J and Liu X: Electroacupuncture promotes chondrocyte proliferation via accelerated G1/S transition in the cell cycle. Int JMol Med. 31:1443–1448. 2013.PubMed/NCBI View Article : Google Scholar
17	Sharpless NE and DePinho RA: The INK4A/ARF locus and its two gene products. Curr Opin Genet Dev. 9:22–30. 1999.PubMed/NCBI View Article : Google Scholar
18	Williams RT, Barnhill LM, Kuo HH, Lin WD, Batova A, Yu AL and Diccianni MB: Chimeras of p14ARF and p16: Functional hybrids with the ability to arrest growth. PLoS One. 9(e88219)2014.PubMed/NCBI View Article : Google Scholar
19	Gil J and Peters G: Regulation of the INK4b-ARF-INK4a tumour suppressor locus: All for one or one for all. Nat Rev Mol Cell Biol. 7:667–677. 2006.PubMed/NCBI View Article : Google Scholar
20	Sharpless NE: INK4a/ARF: A multifunctional tumor suppressor locus. Mutat Res. 576:22–38. 2005.PubMed/NCBI View Article : Google Scholar
21	Yue X, Fu J, Xue X, Gao H, Liu D, Zong Z, Wang W and Yuan Z: Detection of p16 promoter methylation in premature rats with chronic lung disease induced by hyperoxia. Pediatr Int. 52:520–526. 2010.PubMed/NCBI View Article : Google Scholar
22	Rayess H, Wang MB and Srivatsan ES: Cellular senescence and tumor suppressor gene p16. Int J Cancer. 130:1715–1725. 2012.PubMed/NCBI View Article : Google Scholar
23	Thompson JF, Scolyer RA and Kefford RF: Cutaneous melanoma. Lancet. 365:687–701. 2005.PubMed/NCBI View Article : Google Scholar
24	Myong NH: Cyclin D1 overexpression, p16 loss, and pRb inactivation play a key role in pulmonary carcinogenesis and have a prognostic implication for the long-term survival in non-small cell lung carcinoma patients. Cancer Res Treat. 40:45–52. 2008.PubMed/NCBI View Article : Google Scholar
25	Dobashi Y, Goto A, Fukayama M, Abe A and Ooi A: Overexpression of cdk4/cyclin D1, a possible mediator of apoptosis and an indicator of prognosis in human primary lung carcinoma. Int J Cancer. 110:532–541. 2004.PubMed/NCBI View Article : Google Scholar
26	Mittnacht S: The retinoblastoma protein-from bench to bedside. Eur J Cell Biol. 84:97–107. 2005.PubMed/NCBI View Article : Google Scholar
27	Wang X, Huang K and Xu LN: Interaction among Rb/p16, Rb/E2F1 and HDAC1 proteins in gallbladder carcinoma. J Huazhong Univ Sci Technolog Med Sci. 29:729–731. 2009.PubMed/NCBI View Article : Google Scholar
28	Haluska FG, Tsao H, Wu H, Haluska FS, Lazar A and Goel V: Genetic alterations in signaling pathways in melanoma. Clin Cancer Res. 12:2301s–2307s. 2006.PubMed/NCBI View Article : Google Scholar
29	Palmieri G, Capone M, Ascierto ML, Gentilcore G, Stroncek DF, Casula M, Sini MC, Palla M, Mozzillo N and Ascierto PA: Main roads to melanoma. J Transl Med. 7(86)2009.PubMed/NCBI View Article : Google Scholar
30	Wikman H and Kettunen E: Regulation of the G1/S phase of the cell cycle and alterations in the RB pathway in human lung cancer. Expert Rev Anticancer Ther. 6:515–530. 2006.PubMed/NCBI View Article : Google Scholar
31	Thomas NS, Pizzey AR, Tiwari S, Williams CD and Yang J: p130, p107, and pRb are differentially regulated in proliferating cells and during cell cycle arrest by alpha-interferon. J Biol Chem. 273:23659–23667. 1998.PubMed/NCBI View Article : Google Scholar
32	Clijsters L, Hoencamp C, Calis JJA, Marzio A, Handgraaf SM, Cuitino MC, Rosenberg BR, Leone G and Pagano M: Cyclin F controls cell-cycle transcriptional outputs by directing the degradation of the three activator E2Fs. Mol Cell. 74:1264–1277.e7. 2019.PubMed/NCBI View Article : Google Scholar
33	Stanelle J, Stiewe T, Theseling CC, Peter M and Pützer BM: Gene expression changes in response to E2F1 activition. Nucleic Acides Res. 30:1859–1867. 2002.PubMed/NCBI View Article : Google Scholar
34	Han S, Park K, Bae BN, Kim KH, Kim HJ, Kim YD and Kim HY: E2F1expression is related with the poor survival of lymph node-positive breast cancer patients treated with fluorouracil, doxorubicin and cyclophosphamide. Breast Cancer Res Treat. 82:11–16. 2003.PubMed/NCBI View Article : Google Scholar
35	Eymin B, Gazzeri S, Brambilla C and Brambilla E: Distinct pattern of E2F1 expression in human lung tumours: E2F1 is upregulated in small cell lung carcinomal. Oncogene. 20:1678–1687. 2001.PubMed/NCBI View Article : Google Scholar