A Rheostat of Ceramide and Sphingosine-1-Phosphate as a Determinant of Oxidative Stress-Mediated Kidney Injury

Abstract

Reactive oxygen species (ROS) modulate sphingolipid metabolism, including enzymes that generate ceramide and sphingosine-1-phosphate (S1P), and a ROS-antioxidant rheostat determines the metabolism of ceramide-S1P. ROS induce ceramide production by activating ceramide-producing enzymes, leading to apoptosis, while they inhibit S1P production, which promotes survival by suppressing sphingosine kinases (SphKs). A ceramide-S1P rheostat regulates ROS-induced mitochondrial dysfunction, apoptotic/anti-apoptotic Bcl-2 family proteins and signaling pathways, leading to apoptosis, survival, cell proliferation, inflammation and fibrosis in the kidney. Ceramide inhibits the mitochondrial respiration chain and induces ceramide channel formation and the closure of voltage-dependent anion channels, leading to mitochondrial dysfunction, altered Bcl-2 family protein expression, ROS generation and disturbed calcium homeostasis. This activates ceramide-induced signaling pathways, leading to apoptosis. These events are mitigated by S1P/S1P receptors (S1PRs) that restore mitochondrial function and activate signaling pathways. SphK1 promotes survival and cell proliferation and inhibits inflammation, while SphK2 has the opposite effect. However, both SphK1 and SphK2 promote fibrosis. Thus, a ceramide-SphKs/S1P rheostat modulates oxidant-induced kidney injury by affecting mitochondrial function, ROS production, Bcl-2 family proteins, calcium homeostasis and their downstream signaling pathways. This review will summarize the current evidence for a role of interaction between ROS-antioxidants and ceramide-SphKs/S1P and of a ceramide-SphKs/S1P rheostat in the regulation of oxidative stress-mediated kidney diseases.

Keywords: Bcl-2 family proteins; apoptosis; ceramide; fibrosis; inflammation; kidney injury; mitochondria; proliferation; reactive oxygen species; sphingosine-1-phosphate.

Figure 1

Simplified metabolic pathways of sphingolipids. Cer is de novo synthesized at the surface of the ER by condensation of serine and palmitoyl-CoA mediated by SPT, forming 3-KdhSph. 3-KdhSph is then reduced to dhSph by 3-KR. DhSph is the substrate of CerS, forming dhCer which is converted into Cer by DES1. Cer is transferred to the Golgi by CERT or vesicular trafficking and converted into SM by SMS or C1P by CerK. Cer is converted into GlcCer by GCSs and further metabolized into complex GSLs. Cer is also phosphorylated into C1P by C1PP. GSLs and SM in the Golgi are transferred to the plasma membrane by vesicular trafficking, where SM is converted into Cer by sSMase or nSMase. C1P is transferred to the plasma membrane by C1PTP. In the plasma membrane, Cer can be produced from SM via sSMase and converted into Sph by nCDase or C1P via C1PP. Sph can be metabolized into S1P by SphKs. SM in the plasma membrane enters into the recycling pathway in the acid compartment of the endolysosome, where aSMase and GCase produce Cer, which is hydrolyzed into Sph by aCDase. Once released into the cytosol, Sph is reused for Cer synthesis by CerS or phosphorylated by SphKs to yield S1P. S1P is hydrolyzed back into Sph via S1PP or degraded by S1PL into EA1P and HD. Abbreviations; aCDase; acid ceramidase, aSMase; acid sphingomyelinase, Cer; ceramide, CerK; Cer kinase, CerS; Cer synthase, CERT; ceramide transport protein, C1P; Cer-1-phosphate, C1PP; C1P phosphatase, C1PTP; C1P transfer protein, DES; dihydroceramide desaturase, dhCer; dihydroceramide, dhSph; dihydrosphingosine, EA1P; ethanolamine-1-phosphate, ER; endoplasmic reticulum, GCase; glycosidase, GCSs; glycosylceramide synthases, GlcCer; glucosylceramide, GSLs; glycosphingolipids, HD; hexadecenal, 3-KdhSph; 3-ketodihydrosphingosine, 3-KR; 3-KdhSph reductase, nSMase; neutral sphingomyelinase, SM; sphingomyelin, SMS; SM synthase, sSMase; secretaory SMase, Sph; sphingosine, SphK; sphingosine kinase, S1P; sphingosine-1-phosphate, S1PL; S1P lyase, S1PP; S1P phosphatase, SPT; serine palmitoyltransferase. Dashed arrows indicate transportation to the plasma membrane.

Figure 2

Balance between ROS and antioxidants determines ceramide-S1P rheostat that regulates oxidant-induced kidney injury. A rheostat of ROS and antioxidants regulates enzymes that generate Cer and S1P directly or through the modulation of subcellular translocation of the enzymes and upstream signaling pathways, mitochondrial function including apoptotic/anti-apoptotic Bcl-2 family proteins, and proinflammatory cytokines. ROS activate Cer-producing enzymes and vice versa, while antioxidants inhibit these enzymes. ROS stimulate proinflammatory cytokines, which in turn induce Cer generation, mitochondrial dysfunction, upregulation of apoptotic and downregulation of anti-apoptotic Bcl-2 proteins, leading to apoptosis, while antioxidants have the opposite effect. Cer induces ROS production, mitochondrial dysfunction and increased ratio of apoptotic/anti-apoptotic Bcl-2 proteins and inhibition of antioxidants, leading to apoptosis. ROS inhibit SphK/S1P and vice versa, while antioxidants activate it. SphK/S1P has the opposite biological effects to Cer, including restoration of mitochondrial function and the ratio of apoptotic/anti-apoptotic Bcl-2 family proteins, thereby promoting survival, cell proliferation, inflammation and renal fibrosis. →: stimulation, dashed arrow: inhibition. Red arrow; apoptotic, blue arrow; anti-apoptotic.

Figure 3

Interaction between ROS/antioxidants and mitochondria in the regulation of apoptosis. Oxidant stress inhibits MRCC, which induces ROS production. ROS and antioxidants inhibit each other. ROS inhibit MRCC, resulting in Cer production, MPTP opening, MOMP, decreased VDAC expression, suggestive of VDAC closure, and a decreased Bcl-2/Bax ratio, leading to apoptosis, while antioxidants and Bcl-2 prevent these events. Cer inhibits MRCC, leading to ROS production. Both ROS and Cer activate Bax/Bak, which enhances the mitochondrial uptake of Ca²⁺ by enhancing the transfer of Ca²⁺ from the ER, and these events trigger MPTP opening, leading to apoptosis. ROS increase intracellular Ca²⁺, which further enhances ROS production, while Bcl-2 inhibits this event and apoptosis by increasing the capacity of mitochondria to store and buffer Ca²⁺. Inhibition of MPTP opening and MOMP prevents ROS production and restores antioxidant levels. In contrast, antioxidants restore MRCC and loss of MMP, increase the expression of VDAC and Bcl-2/Bax ratio, and prevent an increase in intracellular Ca²⁺ which induces loss of MMP, ameliorating apoptosis. Antioxidants prevent an ROS-induced increase in intracellular Ca²⁺ by increasing Bcl-2 expression, which prevents a rise in intracellular Ca²⁺ by increasing the capacity of mitochondria to buffer Ca²⁺, ameliorating apoptosis. Anti-apoptotic Bcl-2 functions as an antioxidant and inhibits ROS-induced Cer formation. Abbreviations; Cer; ceramide, MMP; mitochondrial membrane potential, MOMP; mitochondrial outer membrane permeability, MPTP; mitochondrial transition pore, MRCC; mitochondrial respiratory chain complex, ROS; reactive oxygen species, VDAC; voltage-dependent anion channel. Red arrow; apoptotic pathway, blue arrow; anti-apoptotic pathway, →; stimulation, dashed arrow; inhibition.

Figure 4

Mitochondria regulate ceramide-induced apoptosis. Cer inhibits MRCC and reduces the Bcl-2/Bax ratio, resulting in MPTP opening and MOMP, which are inhibited by Bcl-2. Cer together with Bax induces MPTP opening and MOMP, which sensitize mitochondria to Ca²⁺, leading to apoptosis. Cer induces release of Ca²⁺ from the ER, resulting in increased mitochondrial uptake of Ca²⁺, leading to apoptosis, while Bcl-2 prevents it. Bcl-2 also increases mitochondrial capacity to store and buffer Ca²⁺, thereby preventing apoptosis. Cer-induced increase in the ratio of Bax/Bcl-2 results in mitochondrial dysfunction and mitochondrial Ca²⁺ uptake, leading to apoptosis. Cer forms Cer channels. Bax/Bak enhances Cer channel formation, triggering MPTP opening and MOMP, leading to apoptosis, while Bcl-2/Bcl-xL disassembles Cer channels. Binding of Cer to VDAC induces MOMP and decreases Bcl-2/Bcl-xL expression. Cer dephosphorylates Bad (dBad) which is required for Cer-induced sensitization of MPTP opening. More Bad and less VDAC are associated with Bcl-xL at the MOM, which sensitizes MPTP to Ca²⁺, leading to apoptosis. Cer binds to tubulin and this formation induces VDAC closure, leading to mitochondrial dysfunction. Abbreviations; dBad; dephosphorylated Bad.—binding, red arrow; apoptotic pathway, blue arrow; anti-apoptotic pathway. →; stimulation, dashed arrow; inhibition. Black bar indicates binding of dBad to Bcl-xL.

Figure 5

Ceramide-induced apoptotic signaling pathways in oxidant-induced kidney injury. ROS-induced Cer activates proinflammatory cytokines (TNF-α, IL-1β), ERK, p38MAPK, SAPK/JNK through PKCζ, p53, Ca²⁺-dependent calpain, NF-kB through cPLA2/COX-2, and TIMP-1, which decreases MMP-1 that suppresses apoptosis, leading to apoptosis. All these factors function as apoptotic except for anti-apoptotic ERK, which is antagonistic to p38MAPK activation, and MMP-I that functions anti-apoptotic. Cer inhibits VEGF. VEGF and IGF-I suppress Cer-induced apoptosis by activating ERK and by inhibiting PKCζ which suppresses Cer-activated SAPK/JNK, respectively. TNF-α-induced Cer production activates PLA2/COX-2, leading to apoptosis. Cer-induced PLA2 activation cleaves AA, and COX-2 is converted into AA, which promotes apoptosis. The binding of Cer to cPLA2 increases AA release, and both cPLA2 and AA increase Cer production, resulting in apoptosis. Cer activates TNF-α-induced NF-κB, cPLA2, AA, LIMK-1 that regulates cytoskeletal organization and IL-1β via inhibition of PKC, leading to apoptosis. Cer inhibits PI3K/Akt/CREB, which functions as anti-apoptotic. Cer suppresses PKC-α that inhibits PP2A activity by decreasing PI3K, which subsequently activates PP2A and results in apoptosis. Cer-induced activation of PP2A inhibits SphK1 activity that functions as anti-apoptotic, leading to apoptosis. Cer increases SGK-1 without increasing its phosphorylation via p38MAPK/cAMP/PKA/PI3K and decreases Akt activity, while overexpression of SGK-1 prevents apoptosis by activating PI3K/Ak. Abbreviations; AA; arachidonic acid, COX-2; cyclooxygenase-2, CREB; cyclic adenosine monophosphate response element-binding protein, ERK; extracellular signal-regulated kinase, IL; interleukin, JNK; Jun N-terminal protein kinase, LIMK-1; LIM kinase-1, MAPK; mitogen-activated protein kinase, MMP-1; matrix metalloproteinase-1, NF-κB; nuclear factor-κB, PI3K; phosphatidylinositol 3-kinase, PKC; protein kinase C, cPLA2; cytosolic phospholipase A2, PP2A; protein phosphatase 2A, SAPK; stress-activated protein kinase, SGK-1; serum- and glucocorticoid-inducible protein kinase-1, SphK1; sphingosine kinase 1, STAT: signal transducer and activator of transcription, TIMP-1; tissue inhibitor of matrix metalloproteinase-1, TNF-α; tumor necrosis factor-α, VEGF; vascular endothelial growth factor. Dashed arrow indicates that Cer increases the expression but no phosphorylation of SGK-1. Red arrow; apoptotic, blue arrow; anti-apoptotic. →; stimulation, dashed arrow, inhibition.

Figure 6

Mitochondria regulate S1P-induced cell survival. SphK1 promotes survival, while SphK2 has the opposite effect. ROS inhibits SphK1/S1P and vice versa. SphK1 prevents ROS-induced apoptosis by preserving MRCC which inhibits MOMP and restores mitochondrial function, leading to a reduction in ROS production, upregulating anti-apoptotic Bcl-2/Bcl-xL and downregulating apoptotic Bax/Bim, while SphK2 has the opposite effect. SphK2 cooperates with C8-BID to stimulate Bax and MOMP, leading to apoptosis, which is inhibited by Bcl-xL. SphK2 activates and inhibits apoptotic Bak and anti-apoptotic Bcl-2 proteins (Bcl-2/Bcl-xL), respectively. SphK2 interacts with Bcl-xL, leading to the suppression of its anti-apoptotic effects. ROS-induced activation of SphK2 induces Ca²⁺ release from the ER, which is dependent on Bax/Bak and dispensable for SphK2 activation, leading to the activation of Bax/Bak and apoptosis. Red arrow; apoptotic pathway, blue arrow; anti-apoptotic pathway, →; stimulation, dashed arrow; inhibition.

Figure 7

S1P-induced signaling pathways regulate cell survival in oxidant-induced kidney injury. SphK1 prevents ROS-induced apoptosis by activating pro-survival HIF-1α/HSP27. HIF-1α activates SphK1. SphK1 functions as anti-apoptotic by activating ERK/PI3K/Akt/PKB and by inhibiting apoptotic p38MAPK, while SphK2 has the opposite effect. Conversely, ERK activates SphK1. SphK1 activates IGF/IGFBP-3 and vice versa and VEGF, ameliorating ROS-induced apoptosis by activating ERK/PI3K/Akt/PKB. SphK1 inhibits JNK and PP2A, thereby inhibiting apoptosis. Conversely, PP2A inhibits SphK1, leading to apoptosis. Abbreviations; HIF; hypoxia-inducible factor, HSP; heat-shock protein, IGF; insulin growth factor, IGFBP-3; IGF binding protein-3, PI3K; phosphatidylinositol 3-kinase, PKB; protein kinase B, TNF; tumor necrosis factor. Red arrow; apoptotic pathway, blue arrow; anti-apoptotic pathway, →; stimulation, dashed arrow; inhibition.

Figure 8

S1P-induced signaling pathways regulate cell proliferation, inflammation and fibrosis in oxidant-induced kidney injury. Cell proliferation: SphK1 promotes cell proliferation via ERK/PI3K/Akt/PKB and TGF-β1, while SphK2 reduces it by inhibiting ERK/PI3K/Akt/PKB. SphK1 activates PDGF by activating ERK, leading to cell proliferation. PDGF promotes cell proliferation by activating SphK1, while this event is inhibited by COX-2. Binding of IGF-II to M6P/IGF-IIR activates ERK through PKC-mediated SphK1 activity, leading to cell proliferation. Inflammation: SphK1/S1P reduces inflammation by inhibiting proinflammatory cytokines (TNF-α and IL-1β) or by activating HIF-1α and HSP27. In contrast, SphK2 enhances inflammation by increasing TGF-β1 expression, inflammatory cytokines (TNF-α and IL-1β). SphK2 together with Fyn promotes inflammation via STAT3/Akt pathway. Renal fibrosis: both SphK1 and SphK2 promote renal fibrosis via the activation of ERK/TGF-β1/CK2α/NF-κB and via Fyn-STAT3/Akt/TGF-β1, respectively. SphK1 also promotes fibrosis by inhibiting PP2A. Abbreviations; CK2α; casein kinase 2α, HIF; hypoxia-inducible factor, HSP; heat-shock protein, IGFII; insulin growth factor II, IGFIIR; IGFII receptor, NF-κB; nuclear factor-κB, PDGF; platelet-derived growth factor, PKB; protein kinase B, PP2A; protein phosphatase 2A, STAT3; signal transducer and activator of transcription 3, TGF; tumor growth factor. →; stimulation, dashed arrow; inhibition.