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. 2020 Dec 17:27:33-43.
doi: 10.1016/j.jot.2020.09.006. eCollection 2021 Mar.

Chondrocyte ferroptosis contribute to the progression of osteoarthritis

Xudong Yao  1 , Kai Sun  1 , Shengnan Yu  2 , Jiahui Luo  3 , Jiachao Guo  1 , Jiamin Lin  1 , Genchun Wang  1 , Zhou Guo  1 , Yaping Ye  1 , Fengjing Guo  1
Affiliations

Chondrocyte ferroptosis contribute to the progression of osteoarthritis

Xudong Yao et al. J Orthop Translat. .

Abstract

Background: Osteoarthritis (OA) is a complex process comprised of mechanical load, inflammation, and metabolic factors. It is still unknown that if chondrocytes undergo ferroptosis during OA and if ferroptosis contribute to the progression of OA.

Materials and methods: In our study, we use Interleukin-1 Beta (IL-1β) to simulate inflammation and ferric ammonium citrate (FAC) to simulate the iron overload in vitro. Also, we used the surgery-induced destabilized medial meniscus (DMM) mouse model to induce OA in vivo. We verify ferroptosis by its definition that defined by the Nomenclature Committee on Cell Death with both in vitro and in vivo model.

Results: We observed that both IL-1β and FAC induced reactive oxygen species (ROS), and lipid ROS accumulation and ferroptosis related protein expression changes in chondrocytes. Ferrostatin-1, a ferroptosis specific inhibitor, attenuated the cytotoxicity, ROS and lipid-ROS accumulation and ferroptosis related protein expression changes induced by IL-1β and FAC and facilitated the activation of Nrf2 antioxidant system. Moreover, erastin, the most classic inducer of ferroptosis, promoted matrix metalloproteinase 13 (MMP13) expression while inhibited type II collagen (collagen II) expression in chondrocytes. At last, we proved that intraarticular injection of ferrostatin-1 rescued the collagen II expression and attenuated the cartilage degradation and OA progression in mice OA model.

Conclusions: In summary, our study firstly proved that chondrocytes underwent ferroptosis under inflammation and iron overload condition. Induction of ferroptosis caused increased MMP13 expression and decreased collagen II expression in chondrocytes. Furthermore, inhibition of ferroptosis, by intraarticular injection of ferrostatin-1, in our case, seems to be a novel and promising option for the prevention of OA.

The translational potential of this article: The translation potential of this article is that we first indicated that chondrocyte ferroptosis contribute to the progression of osteoarthritis which provides a novel strategy in the prevention of OA.

Keywords: Ferroptosis; Ferrostatin-1; Inflammation; Iron overload; Osteoarthritis.

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Conflict of interest statement

The authors declare that they have no conflicts of interest.

Figures

Figure 1
Figure 1
Both IL-1β and FAC induced ferroptosis related protein expression changes in chondrocytes (A) The protein expression level of ACSL4, GPX4, P53, and SLC7A11, when treated with IL-1β, were detected by western blot (B) Band density ratios of ACSL4, GPX4, P53, and SLC7A11 to GAPDH in the western blots were quantified by densitometry (C) The protein expression level of ACSL4, GPX4, P53, and SLC7A11 when treated with FAC were detected by western blot (D) Band density ratios of ACSL4, GPX4, P53 and SLC7A11 to GAPDH in the western blots were quantified by densitometry (E) Total GPX4 protein level were evaluated by immunofluorescence staining in chondrocytes treated with IL-1β or FAC. ∗P ​< ​0.05 versus control. Error bars represent SD.
Figure 2
Figure 2
Ferrostatin-1 attenuated the cytotoxicity, ROS and lipid-ROS accumulation and ferroptosis related protein expression changes induced by IL-1β and FAC (A–B) Cell viability determined by CCK-8 assay (C) Intracellular ROS and lipid-ROS level detected by DCFH-DA and C11 BODIPY fluorescent probe (D) The protein expression level of ACSL4, GPX4, P53, and SLC7A11 when treated by IL-1β with 1 ​μM ferrostain-1 or equal volume of DMSO were detected by western blot (E) Band density ratios of ACSL4, GPX4, P53 and SLC7A11 to GAPDH in the western blots were quantified by densitometry (F) The protein expression level of ACSL4, GPX4, P53 and SLC7A11 when treated by FAC with 1 ​μM ferrostain-1 or equal volume of DMSO were detected by western blot (G) Band density ratios of ACSL4, GPX4, P53 and SLC7A11 to GAPDH in the western blots were quantified by densitometry. ∗P ​< ​0.05 versus control, #P ​< ​0.05 versus IL-1β or FAC treated group. Error bars represent SD.
Figure 3
Figure 3
(A) The total GPX4 protein level in the chondrocytes treated with IL-1β or in combination with ferrostain-1 was evaluated by immunofluorescence staining. The nuclei were stained with DAPI (B) The total GPX4 protein level in the chondrocytes treated with FAC or in combination with ferrostain-1were evaluated by immunofluorescence staining. The nuclei were stained with DAPI.
Figure 4
Figure 4
Erastin promoted matrix metalloproteinase 13 (MMP13) expression while inhibited Type II collagen (collagen II) expression in chondrocytes (A) The protein expression level of GPX4, collagen II and MMP13 when treated with erastin were detected by western blot (B) The band density ratio of GPX4, collagen II and MMP13 to β-actin in the western blots were quantified by densitometry (C) The protein expression level of collagen II and MMP13 when treated by IL-1β with 1 μM ferrostain-1 or equal volume of DMSO were detected by western blot (D-E) The band density ratio of collagen II and MMP13 to β-actin in the western blots were quantified by densitometry (F) The protein expression level of collagen II and MMP13 when treated by FAC with 1 μM ferrostain-1 or equal volume of DMSO were detected by western blot (G-H) The band density ratio of collagen II and MMP13 to β-actin in the western blots were quantified by densitometry.∗P < 0.05 versus control, #P < 0.05 versus IL-1β or FAC treated group. Ns P > 0.05 versus IL-1β or FAC treated group. Error bars represent SD.
Figure 5
Figure 5
Nrf2 antioxidant system and ferroptosis are mutually regulated under inflammation and iron overload condition (A) The protein expression level of Nrf2, HO-1, NQO-1 and Trx when treated by IL-1β with 1 μM ferrostain-1 or equal volume of DMSO were detected by western blot (B) The band density ratio of Nrf2, HO-1, NQO-1 and Trx to GAPDH in the western blots were quantified by densitometry (C) The protein expression level of Nrf2, HO-1, NQO-1 and Trx when treated by FAC with 1 μM ferrostain-1 or equal volume of DMSO were detected by western blot (D) The band density ratio of Nrf2, HO-1, NQO-1 and Trx to GAPDH in the western blots were quantified by densitometry. (E) The nuclear protein level of Nrf2 and HO-1 when treated by IL-1β with 1 μM ferrostain-1 or equal volume of DMSO were detected by western blot (F) The band density ratio of Nrf2 and HO-1 to Lamin B were quantified by densitometry. (G) The nuclear protein level of Nrf2 and HO-1 when treated by FAC with 1 μM ferrostain-1 or equal volume of DMSO were detected by western blot (H) The band density ratio of Nrf2 and HO-1 to Lamin B were quantified by densitometry. (I) The protein level of GPX4 when treated by IL-1β with si-Nrf2 or negative control were detected by western blot (J) The band density ratio of GPX4 to GAPDH were quantified by densitometry. (K) The protein level of GPX4 when treated by IL-1β with si-Nrf2 or negative control were detected by western blot (L) The band density ratio of GPX4 to GAPDH were quantified by densitometry.∗P < 0.05 versus control, #P < 0.05 versus IL-1β or FAC treated group. ns P > 0.05. Error bars represent SD.
Figure 6
Figure 6
Ferrostain-1 attenuated cartilage degradation and increased the Collagen II and GPX4 expression in the mouse OA model (A) Cartilage degradation was assessed by Safranin O/fast green staining (B) Immunohistochemistry staining of GPX4 and collagen II (C) Number of GPX4 positive cells per field under 200-time magnification (D) The number of collagen positive cells per field under 200-time magnification (E) The progression of OA was evaluated using the OARSI scores. ∗P ​< ​0.05 versus sham surgery group, #P ​< ​0.05 versus DMM group. ns P ​> ​0.05. Error bars represent SD.
Supplymentary figure
Supplymentary figure
The negative control of immunohistochemical staining.
See this image and copyright information in PMC

References

    1. James S.L., Abate D., Abate K.H., Abay S.M., Abbafati C., Abbasi N. Global, regional, and national incidence, prevalence, and years lived with disability for 354 diseases and injuries for 195 countries and territories, 1990-2017: a systematic analysis for the Global Burden of Disease Study 2017. LANCET (N AM ED) 2018;392(10159):1789–1858. 2018-11-10. - PMC - PubMed
    1. Musumeci G., Aiello F.C., Szychlinska M.A., Di Rosa M., Castrogiovanni P., Mobasheri A. Osteoarthritis in the XXIst century: risk factors and behaviours that influence disease onset and progression. Int J Mol Sci. 2015;16(3):6093–6112. 2015-03-16. - PMC - PubMed
    1. Mobasheri A., Batt M. An update on the pathophysiology of osteoarthritis. Ann Phys Rehabil Med. 2016;59(5–6):333–339. 2016-12-01. - PubMed
    1. Abramoff B., Caldera F.E. Osteoarthritis: pathology, diagnosis, and treatment options. Med Clin. 2020;104(2):293–311. 2020-03-01. - PubMed
    1. Gozzelino R., Arosio P. Iron homeostasis in health and disease. Int J Mol Sci. 2016;17(1) 2016-01-20. - PMC - PubMed

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