Key Event Relationship Overview
AOPs Referencing Relationship
|AOP Name||Adjacency||Weight of Evidence||Quantitative Understanding|
|Endocytic lysosomal uptake leading to liver fibrosis||adjacent||High|
Life Stage Applicability
|All life stages|
Key Event Relationship Description
Different substances can be uptaken by endocytosis and localized in lysosomes, while some of them can cause lysosomal disruption. Lysosomotropic agents are mostly weak, lipophilic bases that diffuse across lysosomal membrane, get protonated in the acidic milieu of lysosome and therefore get trapped inside (de Duve et al., 1974). They accumulate and cause the destabilization of lysosomal membranes by acting as surfactants, incorporating its hydrophobic tail in the membrane and with the hydrophilic head facing the interior of the lysosome (de Duve et al.,1974; Firestone et al.,1979). Their accumulation increase the intralysosomal pH, which has many consequences, including the prevention of the further uptake of lysosomotropic compounds, an increase in size and number of lysosomes and the overloading of lysosomes with non-digestible materials.
There are different mechanisms how lysosomotropic agents can disrupt lysosomal membrane. However, not all lysosomotropic agents disrupt lysosomes- for example ammonia salts, methylamine and related hydrophilic weak bases cause swelling of the lysosomes, but do not increase permeability of the membrane. Usually in order to do that, agent requires a certain degree of lipid solubility. The amine will accumulate in the lysosomes until its concentration is high enough to solubilize the lysosomal membrane (Dubowchik et al., 1995)
It has been demonstrated that as a result of protonated agents in lysosomes, there will be accumulation of non-permeable charged substances which will result in inflow of water by increased osmolarity (Bandyopadhyay et al., 2014). Inflow of water results in increase of size and can cause the rupture of lysosome.
Also, oxidative stress can cause destabilization of the lysosomal membrane and for this process, intra-lysosomal ferric ions are essential. They catalyse the formation of oxygen radicals from hydrogen peroxide (Zdoslek et al., 1993).
Evidence Supporting this KER
Trapping of lysosomotropic agents accumulates substances inside of the lysosomes, increases volume of these organelles, and big lysosomes are more prone to rupture (Ono et al., 2003). However, there are many mechanisms for lysosomotropic substances to provoke lysosomal disruption, but their prior uptake by lysosomes is essential.
Accumulation of different substances in lysosomes is causing severe dysfunction, increased permeability of lysosomal membrane and even their rupture. It was demonstrated that even accumulation of substances that are physiologically find in lysosomes can lead towards lysosomal dysfunction. Use of cathepsins' B and L inhibitors causes abnormal accumulation of pro-cathepsins, enlargement of lysosomes and their severe dysfunction (Jung et al., 2015)
Leu Leu OMe is one of the agents that accumulates in lysosomes, where it is converted to a membranolytic compound and increase permeability of lysosomal membrane (Uchimoto et al., 1999, Thiele and Lipsky, 1990)
Sphingosine is another lysosomotropic agent that accumulates within the lysosomes, where it permeabilizes the membrane via a detergent mechanism and provokes relocation of lysosomal enzymes to the cytosol (Kagedal et al., 2001). It induces lysosomal disruption in primary mouse hepatocytes as well as it permabilize isolated hepatic lysosomes in vitro (Werneburg et al., 2002) Also, exposure of J774 cells to ammonium chloride prior to sphingosine resulted in formation of NH3, which entered into lysosomes and became protonated and increased the pH of the organelle. This prevented the accumulation of sphingosine in the lysosome and provided protection against its lysosomolytic and apoptosis-inducing effects (Kagedal et al., 2001).
Considering nanomaterials (NMs) as a trigger for lysosomal damage, recent studies underpinned the importance of lysosomal NM uptake for NM-induced toxicity. Once the material is taken up by a cell and transported to the lysosome, the acidic milieu herein can either enhance solubility of a NM, or the material remains in its initial nano form. Both can induce toxicity, causing lysosomal swelling, followed by lysosomal disruption and the release of pro-apoptotic proteins (Cho et al., 2011; Cho et al., 2012; Wang et al., 2013).
H2O2 also changes the permeability of the lysosomal membrane. It reacts with redox-active iron in the lysosomes, and produces hydroxyl radicals in Fenton-type reactions (Kubota et al., 2010). These radicals can destabilize the membrane by lipid peroxidation and damage of lysosomal proteins. Presence of desferrioxamine, that binds iron, is protecting lysosomes from rupture (Brunk et al., 2001).
Hayashi et al. demonstrated the uptake of FITC-crotamine into endosomal compartment of cells, and increased accumulation in a time dependant manner. Only 15 minutes after the treatment it was possible to see morphological changes of the cell, disruption of the lipid bilayer of the lysosomal membrane and subsequent rupture of lysosomes. They presented that crotamine treatment triggered the release of cysteine cathepsins to the cell cytosol (Hayashi et al., 2008).
Uncertainties and Inconsistencies
Quantitative Understanding of the Linkage
Known modulating factors
Known Feedforward/Feedback loops influencing this KER
Domain of Applicability
Mouse (Werneburg et al., 2002; Kagedal et al., 2001)
Rat (Jung et al., 2015)
Hamster (Hayashi et al., 2008)
Human (Wang et al., 2013)
Bandyopadhyay D, Cyphersmith A, Zapata JA, Kim YJ, Payne CK. Lysosome transport as a function of lysosome diameter. PLOS ONE. (2014) 9:e86847.
Brunk UT, Neuzil J, Eaton JW. Lysosomal involvement in apoptosis, Redox Report, (2001) 6 (2): 91-97.
Cho W-S, Duffin R, Howie SEM, Scotton CJ, Wallace WAH, Macnee W, Bradley M, Megson IL, Donaldson K. Progressive severe lung injury by zinc oxide nanoparticles; the role of Zn2+ dissolution inside lysosomes. Part Fibre Toxicol (2011) 8:27.
Cho W-S, Duffin R, Thielbeer F, Bradley M, Megson IL, MacNee W, Poland CA, Tran CL, Donaldson K. Zeta potential and solubility to toxic ions as mechanisms of lung inflammation caused by metal/metal oxide nanoparticles. Toxicol Sci (2012) 126:469–477.
de Duve C, de Barsy T, Poole B, Trouet A, Tulkens P, Van Hoof F. Commentary. Lysosomotropic agents. Biochem. Pharmacol. (1974) 23:2495- 2531.
Dubowchik GM, Gawlak SL, Firestone RA. The in vitro effects of three lysosomotropic detergents against three human tumor cell lines Bioorg. Med. Chem. Lett. (1995) 5:893-898.
Firestone RA, Pisano JM, Bonney, RJ. Lysosomotropic agents. 1. Synthesis and cytotoxic action of lysosomotropic detergents. J. Med. Chem. (1979) 22: 1130- 1133.
Hayashi MAF, Nascimento FD, Kerkis A, Oliveira V, Oliveira EB, Pereira A, Rádis-Baptista G, Nader HB, Yamane T, Kerkis I, Tersariol ILS. Cytotoxic effects of crotamine are mediated through lysosomal membrane permeabilization. Toxicon. (2008) 52(3): 508–517.
Jung M, Lee J, Seo H-Y, Lim JS, Kim E-K. Cathepsin Inhibition-Induced Lysosomal Dysfunction Enhances Pancreatic Beta-Cell Apoptosis in High Glucose. PLoS ONE. (2015) 10(1):e011697.
Kagedal K, Zhao M, Svensson I, Brunk UT. Sphingosine-induced apoptosis is dependent on lysosomal proteases. The Biochemical journal. (2001) 359: 335-43.
Kubota C, Torii S, Hou N, Saito N, Yoshimoto Y, Imai H, Takeuchi T. Constitutive reactive oxygen species generation from autophagosome/lysosome in neuronal oxidative toxicity. J Biol Chem. (2010) 285(1):667-74.
Ono K, Kim SO, Han J. Susceptibility of lysosomes to rupture is a determinant for plasma membrane disruption in tumor necrosis factor alpha-induced cell death. Mol Cell Biol. (2003) 23: 665-76.
Thiele DL, Lipsky PE. Mechanism of L-leucyl-L-leucine methyl ester-mediated killing of cytotoxic lymphocytes: dependence on a lysosomal thiol protease, dipeptidyl peptidase I, that is enriched in these cells. Proc Natl Acad Sci U S A. (1990) 87(1): 83–87.
Uchimoto T, Nohara H, Kamehara R, Iwamura M, Watanabe N, Kobayashi Y. Mechanism of apoptosis induced by a lysosomotropic agent, L-Leucyl-L-Leucine methyl ester. Apoptosis. (1999) 4(5): 357–362.
Wang F, Bexiga MG, Anguissola S, Boya P, Simpson JC, Salvati A, Dawson KA: Time resolved study of cell death mechanisms induced by amine-modified polystyrene nanoparticles. Nanoscale (2013) 5:10868–76.
Werneburg NW, Guicciardi ME, Bronk SF, Gores GJ. Tumor necrosis factor-alpha-associated lysosomal permeabilization is cathepsin B dependent. Am J Physiol Gastrointest Liver Physiol. (2002) 283:G947–G956.
Zdolsek J, Zhang H, Roberg K, Brunk U, Sies H. H2O2-Mediated Damage to Lysosomal Membranes of J-774 Cells, Free Radical Research Communications, 1993, 18:2, 71-85