2025: Low Fluoride Regulates Macrophage Polarization Through Mitochondrial Autophagy Mediated by PINK1/Parkin Axis

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NOTE: PINK1 = PTEN-induced putative kinase 1.

The PINK1/Parkin axis is a crucial cellular pathway involved in mitophagy, the selective degradation of damaged mitochondria via autophagy. It plays a key role in maintaining mitochondrial quality control and has been heavily studied in the context of Parkinson’s disease.

PINK1 is thyroid hormone-responsive (T3) (i.e. Zhang et al., 2021; Chang et a;., 2022).

SEE: viewtopic.php?f=3&t=5757

Xie F, Zhou J, Liu B, Zhao L, Lv C, Zhang Q, Yuan L, Sun D, Wei W - "Low Fluoride Regulates Macrophage Polarization Through Mitochondrial Autophagy Mediated by PINK1/Parkin Axis" Biomolecules 15(5):647 (2025). doi: 10.3390/biom15050647
https://www.mdpi.com/2218-273X/15/5/647

Abstract

Fluoride exposure has been shown to affect immune cell subsets and immune function, but its impact on macrophage polarization remains unclear. This study investigates the effects of low fluoride exposure on macrophage polarization and its underlying mechanisms through epidemiological surveys, animal experiments, and in vitro cell experiments. In the population-based epidemiological survey, we used mass cytometry to assess the impact of low fluoride exposure (0.570–2.027 mg/L) in the environment on human immune cell populations following the current water improvement and fluoride reduction measures. A rat fluorosis model was established by treating rats with sodium fluoride (NaF) in drinking water at concentrations of 0 mg/L, 5 mg/L, 10 mg/L, 25 mg/L, and 50 mg/L for 90 days, and morphological changes were assessed by hematoxylin–eosin (H&E) staining and transmission electron microscopy in the spleen of rats. Flow cytometry was used to analyze the proportion of macrophage subtypes in the spleen, while Western blot and immunofluorescence were performed to detect the expression of mitochondrial autophagy-related proteins. An M1 macrophage model was constructed in vitro by inducing THP-1 cells, and the effects of fluoride on macrophage-related cell markers and cytokines were assessed using flow cytometry and ELISA, respectively, following intervention with an autophagy inhibitor. Mitochondrial membrane potential and mitochondrial–lysosomal colocalization are analyzed through flow cytometry and confocal microscopy. The study aims to investigate the role of mitophagy in sodium fluoride-induced macrophage polarization. Epidemiological investigations revealed that low fluoride increases the proportion of blood monocytes, as well as the expression levels of CD68 (a macrophage surface marker), CD86 (an M1 macrophage marker), and the inflammatory cytokine IFN-γ in peripheral blood mononuclear cells (PBMCs). In the rats of NaF-treated groups, splenic tissues exhibited inflammatory infiltration, mitochondrial swelling, and increased autophagosome formation. Moreover, low fluoride activated the PINK1/Parkin-mediated mitophagy pathway, promoting an increase in the M2/M1 macrophage ratio. In vitro experiments further confirmed that autophagy inhibitors reversed the NaF-induced increase in the M2/M1 macrophage ratio. This study demonstrates that low fluoride induces inflammatory responses in the body and drives M1 macrophage polarization toward M2 macrophages via mitophagy. These findings highlight the potential immunological risks associated with low fluoride and provide mechanistic insights into the interplay among fluoride, mitophagy, and macrophage polarization.

Thyroid

Bi W, Jia J, Pang R, Nie C, Han J, Ding Z, Liu B, Sheng R, Xu J, Zhang J - "Thyroid hormone postconditioning protects hearts from ischemia/reperfusion through reinforcing mitophagy" Biomed Pharmacother 118:109220 (2019). doi: 10.1016/j.biopha.2019.109220.
https://pubmed.ncbi.nlm.nih.gov/31357081/
"Meanwhile, we found that THPostC [T3 postconditioning] stimulated PINK1/Parkin pathway, a critical regulator for mitophagy."

Chang H, Lin C, Li Z, Shen Y, Zhang G, Mao L, Ma C, Liu N, Lu H - "T3 alleviates neuroinflammation and reduces early brain injury after subarachnoid haemorrhage by promoting mitophagy via PINK 1-parkin pathway" Exp Neurol 357:114175 (2022)oi: 10.1016/j.expneurol.2022.114175
https://www.sciencedirect.com/science/a ... 862200200X
"We showed that in vivo T3 treatment promoted mitophagy, decreased microglial activation, alleviated neuroinflammation, and reduced neuronal apoptosis following SAH. Overall, this thyroid hormone (TH) exerts a protective effect on neurones after SAH via the PINK 1/PARKIN pathway."

Chi HC, Chen SL, Lin SL, Tsai CY, Chuang WY, Lin YH, Huang YH, Tsai MM, Yeh CT, Lin KH - "Thyroid hormone protects hepatocytes from HBx-induced carcinogenesis by enhancing mitochondrial turnover" Oncogene 36(37):5274-5284 (2017). doi: 10.1038/onc.2017.136.
https://www.nature.com/articles/onc2017136
"Using microarray data analysis, this protective effect of TH [Thyroid Hormone] was found to be mediated via activation of PTEN-induced kinase 1 (PINK1) in hepatocytes. PINK1, in turn, activated and recruited Parkin, an E3 ligase, to ubiquitinate MITO-associated HBx protein and trigger selective mitophagy. The pathological significance of the TH/PINK1 pathway in liver protection was confirmed by the concomitant decrease in expression of both TR and PINK1 in matched HCC tumor tissues and negatively correlated with aggressive progression of cancer and poor prognosis. Our data indicate that TH/PINK1/Parkin pathway has a critical role in protecting hepatocytes from HBx-induced carcinogenesis."

Singh BK, Sinha RA, Tripathi M, Mendoza A, Ohba K, Sy JAC, Xie SY, Zhou J, Ho JP, Chang CY, Wu Y, Giguère V, Bay BH, Vanacker JM, Ghosh S, Gauthier K, Hollenberg AN, McDonnell DP, Yen PM - "Thyroid hormone receptor and ERRα coordinately regulate mitochondrial fission, mitophagy, biogenesis, and function" Sci Signal 11(536):eaam5855 (2018). doi: 10.1126/scisignal.aam5855
https://pubmed.ncbi.nlm.nih.gov/29945885/
"TH induced the expression of genes regulating mitochondrial fission (Drp1) and mitophagy (Pink1 and Parkin)...in mouse livers."

Yu G, Tzouvelekis A, Wang R, Herazo-Maya JD, Ibarra GH, Srivastava A, de Castro JPW, DeIuliis G, Ahangari F, Woolard T, Aurelien N, Arrojo EDR, Gan Y, Graham M, Liu X, Homer RJ, Scanlan TS, Mannam P, Lee PJ, Herzog EL, Bianco AC, Kaminski N - "Thyroid hormone inhibits lung fibrosis in mice by improving epithelial mitochondrial function" Nat Med 24:39–49 (2018)
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5760280/

Zhang Y, Yu G, Kaminski N, Lee PJ - "PINK1 mediates the protective effects of thyroid hormone T3 in hyperoxia-induced lung injury" Am J Physiol Lung Cell Mol Physiol 320(6):L1118-L1125 (2001) doi: 10.1152/ajplung.00598.2020
https://pmc.ncbi.nlm.nih.gov/articles/PMC8285622/
https://journals.physiology.org/doi/pre ... 00598.2020

• Vamesu BM, Nicola T, Li R, Hazra S, Matalon S, Kaminski N, Ambalavanan N, Kandasamy J - "Thyroid hormone modulates hyperoxic neonatal lung injury and mitochondrial function" JCI Insight. 2023 Apr 24;8(8):e160697. doi: 10.1172/jci.insight.160697
  https://pmc.ncbi.nlm.nih.gov/articles/PMC10243814/
  "We found that inhaled T3 (given i.n.) attenuated hyperoxia-induced lung injury and mitochondrial dysfunction in newborn mice. T3 also reduced bioenergetic deficits in UC-MSCs obtained from both infants with no or mild BPD and those with moderate to severe BPD. T3 also increased the content of peroxisome proliferator-activated receptor γ coactivator 1α in lung homogenates of mice exposed to hyperoxia as well as mitochondrial potential in both NMLFs and UC-MSCs."

Gq/11

Lin Q, Li S, Jiang N, Shao X, Zhang M, Jin H, Zhang Z, Shen J, Zhou Y, Zhou W, Gu L, Lu R, Ni Z - "PINK1-parkin pathway of mitophagy protects against contrast-induced acute kidney injury via decreasing mitochondrial ROS and NLRP3 inflammasome activation" Redox Biol 26:101254 (2019). doi: 10.1016/j.redox.2019.101254
https://www.sciencedirect.com/science/a ... 1719302988

• NOTE: NLRP3 is regulated by Gq/11 - see:
  Kong R, Peng L, Bao H, Sun L, Feng Y, Li H, Wang D - "The role of Gαq in regulating NLRP3 inflammasome activation" Inflamm Res. 2024 Oct 26. doi: 10.1007/s00011-024-01961-x
  https://pubmed.ncbi.nlm.nih.gov/39455437/

Zhang Y, Sauler M, Shinn AS, Gong H, Haslip M, Shan P, Mannam P, Lee PJ - "Endothelial PINK1 mediates the protective effects of NLRP3 deficiency during lethal oxidant injury" J Immunol 192(11):5296-304 (2014). doi: 10.4049/jimmunol.1400653
https://pubmed.ncbi.nlm.nih.gov/24778451/

PFAS

Shang Y, Chen K, Ni H, Zhu X, Yuan X, Wang Y, Liu X, Cui Z, Niu Y, Shi Y, Wu H, Xia D, Wu Y - "Environmentally relevant concentrations of perfluorobutane sulfonate impair locomotion behaviors and healthspan by downregulating mitophagy in C. elegans" J Hazard Mater 480:135938 (2024) doi: 10.1016/j.jhazmat.2024.135938
https://pubmed.ncbi.nlm.nih.gov/39326150/
"The results furtherly revealed that PFBS exposure led to elevated levels of reactive oxygen species (ROS) and mitophagy impairment through downregulating pink-1/pdr-1 pathway."

Kalyn M, Lee H, Curry J, Tu W, Ekker M, Mennigen JA - "Effects of PFOS, F-53B and OBS on locomotor behaviour, the dopaminergic system and mitochondrial function in developing zebrafish (Danio rerio)" Environ Pollut 326:121479 (2023) doi: 10.1016/j.envpol.2023.121479
https://pubmed.ncbi.nlm.nih.gov/36958660/
"PFOS specifically reduces transcript abundance of dat and pink1."

Feng Y, Huang Y, Lu B, Xu J, Wang H, Wang F, Lin N - "The role of Drp1 - Pink1 - Parkin - mediated mitophagy in perfluorobutane sulfonate- induced hepatocyte damage" Ecotoxicol Environ Saf 285:117066 (2024) doi: 10.1016/j.ecoenv.2024.117066
https://www.sciencedirect.com/science/a ... 1324011424
"These findings highlight PFBS's capacity to inflict hepatocyte injury through mitochondrial disruption, positioning Drp1/Pink1/Parkin-mediated mitophagy as a crucial cellular defense mechanism against PFBS-induced toxicity."

Cadmium

Chen C, Chen Y, Zhai H, Xiao Y, Xu J, Gu Y, Han X, Wang C, Chen Q, Lu H - "Cadmium exposure induces skeletal muscle insulin resistance through the reactive oxygen species-mediated PINK1/Parkin pathway" Ecotoxicol Environ Saf 284:116954 (2024) doi: 10.1016/j.ecoenv.2024.116954
https://www.sciencedirect.com/science/a ... 1324010303

Sun J, Yu F, Wang T, Bian J, Liu Z, Zou H - "The role of DRP1- PINK1-Parkin-mediated mitophagy in early cadmium-induced liver damage" Toxicology. 466:153082 (2022) doi: 10.1016/j.tox.2021.153082
https://pubmed.ncbi.nlm.nih.gov/34952138/

Li D, Yang C, Sun L, Zhao Z, Liu J, Zhang C, Sun D, Zhang Q - "High fluoride aggravates cadmium-mediated nephrotoxicity of renal tubular epithelial cells through ROS-PINK1/Parkin pathway" Sci Total Environ 953:175927 (2024) doi: 10.1016/j.scitotenv.2024.175927
https://pubmed.ncbi.nlm.nih.gov/39236818/
"High F of 15 μg/mL aggravated Cd-mediated nephrotoxicity of renal tubular epithelial cells via the ROS-PINK1/Parkin pathway."

[admin]

Here are a few more studies on fluoride and PINK1/Parkin from the last few years. - Wendy

Dong Y, Sun X, He W, Xiang J, Qi X, Hong W, He Y, Guan Z - "Elevated Level of PINK1/Parkin-Mediated Mitophagy Pathway Involved to the Inhibited Activity of Mitochondrial Superoxide Dismutase in Rat Brains and Primary Hippocampal Neurons Exposed to High Level of Fluoride" Biol Trace Elem Res. 202(2):538-547 (2024) doi: 10.1007/s12011-023-03681-5
https://pubmed.ncbi.nlm.nih.gov/37193858/
"The results showed that the rats exposed to fluoride exhibited different degrees of dental fluorosis. In comparison to controls, the expressions of PINK1 and Parkin were significantly higher in the rat brains and primary neurons exposed to high fluoride."

Hu Y, Li Y, Li M, Zhao T, Zhang W, Wang Y, He Y, Zhao H, Li H, Wang T, Zhao Y, Wang J, Wang J - "Calcium supplementation attenuates fluoride-induced bone injury via PINK1/Parkin-mediated mitophagy and mitochondrial apoptosis in mice" J Hazard Mater 465:133411 (2024) doi: 10.1016/j.jhazmat.2023.133411
"Fluoride exposure caused the impairment of mitochondria and activation of PTEN-induced putative kinase1 (PINK1)/E3 ubiquitin ligase Park2 (Parkin)-mediated mitophagy and mitochondrial apoptosis in the bones, which were restored after blocking Parkin. "

Kumar S, Swamy RS, Bhushan R, Chhabra V, Shenoy S, Murti K, Singh SK, Kumar N - "Molecular and immunohistochemical alterations in fluoride-induced neurological impediment in adult rats" J Trace Elem Med Biol 86:127511 (2024). doi: 10.1016/j.jtemb.2024.127511
https://www.sciencedirect.com/science/a ... 2X24001317
"The confocal studies showed increased expression of inflammatory (α-Synuclein, PARKIN), apoptotic (LC3B, B.A.X., p53, KI67), and mitochondrial dysfunction (NeuN, DNM-1, M.F.N.) markers in fluoride-treated animals."

• 

Liang C, Gao Y, He Y, Han Y, Manthari RK, Tikka C, Chen C, Wang J, Zhang J - "Fluoride induced mitochondrial impairment and PINK1-mediated mitophagy in Leydig cells of mice: In vivo and in vitro studies" Environ Pollut 256:113438 (2020) doi: 10.1016/j.envpol.2019.113438
https://pubmed.ncbi.nlm.nih.gov/31672359/
"The results showed that fluoride decreased the mitochondrial membrane potential with a concomitant increase in the number of lysosomes. Meanwhile, fluoride exposure also increased the expressions of PINK1 and PHB2 in TM3 Leydig cells. These results revealed that fluoride could induce mitochondrial impairment and excessive PINK1/Parkin-mediated mitophagy in testicular cells, especially in Leydig cells, which could contribute to the elucidation of the mechanisms of F-induced male reproductive toxicity."

Song C, Zhang A, Zhang M, Song Y, Huangfu H, Jin S, Sun Y, Zhang C, Shi D, Wang J, Peng W, Luo Q - "Nrf2/PINK1-mediated mitophagy induction alleviates sodium fluoride-induced hepatic injury by improving mitochondrial function, oxidative stress, and inflammation" Ecotoxicol Environ Saf. 252:114646 (2023) doi: 10.1016/j.ecoenv.2023.114646
"NaF-induced mitophagy activation is regulated by PINK1/Parkin signaling...Nrf2 triggers PINK1 expression under NaF stimulation."

Tang Y, Zhang J, Hu Z, Xu W, Xu P, Ma Y, Xing H, Niu Q. - "PRKAA1 induces aberrant mitophagy in a PINK1/Parkin-dependent manner, contributing to fluoride-induced developmental neurotoxicity" Ecotoxicol Environ Saf 255:114772 (2023) doi: 10.1016/j.ecoenv.2023.114772.
https://www.sciencedirect.com/science/a ... 1323002762
"Our findings indicated that NaF causes aberrant mitophagy via PRKAA1 in a PINK1/Parkin-dependent manner, which triggers neuronal apoptosis."

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Fluoroquinolones & Macrophage Polarization & RAW 264.7 cells

Lang L, Zhang Y, Yang A, Dong J, Li W, Zhang G - "Macrophage polarization induced by quinolone antibiotics at environmental residue level" Int Immunopharmacol 106:108596 (2022). doi: 10.1016/j.intimp.2022.108596
https://papers.ssrn.com/sol3/papers.cfm ... id=3982289

Abstract

Widespread use of quinolone antibiotics leads to serious residues in the environment and toxicity effects. This paper studied the effects of three typical quinolone antibiotics ciprofloxacin, norfloxacin and pipemidic acid on the polarization of macrophages RAW264.7 cells. The experimental concentrations were 0.01, 0.1, 1, 10, 100 and 1000 μg/L according to the environmental residual level. By MTT assay, phagocytosis assay and migration assay, macrophages were found to exhibit hormesis effect of low dose promotion and high dose inhibition. The detection of macrophages surface markers showed that 0.1 μg/L antibiotics increased the secretion of pro-inflammatory cytokines and induced macrophages to be M1-type, and 1000 μg/L antibiotics significantly increased the secretion of anti-inflammatory cytokines and induced macrophages to be M2-type. In order to explore the relationship between low-dose excitatory effects and polarization, the mechanism of 0.1 μg/L antibiotics inducing macrophages to M1-type was further studied. Results showed that 0.1 μg/L quinolone antibiotics activated the key proteins in the PI3K/Akt, Notch1, JNK and JAK2/STAT3 signaling pathways to cause the secretion of pro-inflammatory cytokines and induce inflammation. In order to eliminate inflammation, macrophages number feedback increased, phagocytosis and migration function enhanced, showing hormesis effect. In vitro macrophages experiment confirmed that quinolone antibiotics with environmental residual concentration had immunotoxicity.

Compare to:

De la Fuente B, Vázquez M, Rocha RA, Devesa V, Vélez D - "Effects of sodium fluoride on immune response in murine macrophages" Toxicol In Vitro. 34:81-87 (2016) doi: 10.1016/j.tiv.2016.03.001
https://pubmed.ncbi.nlm.nih.gov/26965474/

Abstract

Excessive fluoride intake may be harmful for health, producing dental and skeletal fluorosis, and effects upon neurobehavioral development. Studies in animals have revealed effects upon the gastrointestinal, renal and reproductive systems. Some of the disorders may be a consequence of immune system alterations. In this study, an in vitro evaluation is made of fluoride immunotoxicity using the RAW 264.7 murine macrophage line over a broad range of concentrations (2.5-75mg/L). The results show that the highest fluoride concentrations used (50-75mg/L) reduce the macrophage population in part as a consequence of the generation of reactive oxygen and/or nitrogen species and consequent redox imbalance, which in turn is accompanied by lipid peroxidation. A decrease in the expression of the antiinflammatory cytokine Il10 is observed from the lowest concentrations (5mg/L). High concentrations (50mg/L) in turn produce a significant increase in the proinflammatory cytokines Il6 and Mip2 from 4h of exposure. In addition, cell phagocytic capacity is seen to decrease at concentrations of ≥20mg/L. These data indicate that fluoride, at high concentrations, may affect macrophages and thus immune system function - particularly with regard to the inflammation autoregulatory processes, in which macrophages play a key role.

PFAS

Kim HG, Kim SH, Kim TS, Park TW, Won R, Park HD, Choi SA, Jung YW - "Polyvinylidene Fluoride Alters Inflammatory Responses by Activation-induced Cell Death in Macrophages" Immune Netw 17(6):402-409 (2017) doi: 10.4110/in.2017.17.6.402
https://pubmed.ncbi.nlm.nih.gov/29302253/

Chang LP, Lai YS, Wu CJ, Chou TC - "Liquid perfluorochemical inhibits inducible nitric oxide synthase expression and nitric oxide formation in lipopolysaccharide-treated RAW 264.7 macrophages " J Pharmacol Sci 111(2):147-54 (2009)
https://pubmed.ncbi.nlm.nih.gov/19834286/

Kong B, Wang X, He B, Wei L, Zhu J, Jin Y, Fu Z - "8:2 fluorotelomer alcohol inhibited proliferation and disturbed the expression of pro-inflammatory cytokines and antigen-presenting genes in murine macrophages" Chemosphere 219:1052-1060 (2019) doi: 10.1016/j.chemosphere.2018.12.091
https://pubmed.ncbi.nlm.nih.gov/30558807/