Differential Effects of MitoVitE, α-Tocopherol and Trolox on Oxidative Stress, Mitochondrial Function and Inflammatory Signalling Pathways in Endothelial Cells Cultured under Conditions Mimicking Sepsis

Abstract

Sepsis is a life-threatening response to infection associated with inflammation, oxidative stress and mitochondrial dysfunction. We investigated differential effects of three forms of vitamin E, which accumulate in different cellular compartments, on oxidative stress, mitochondrial function, mRNA and protein expression profiles associated with the human Toll-like receptor (TLR) -2 and -4 pathways. Human endothelial cells were exposed to lipopolysaccharide (LPS)/peptidoglycan G (PepG) to mimic sepsis, MitoVitE, α-tocopherol, or Trolox. Oxidative stress, mitochondrial function, mitochondrial membrane potential and metabolic activity were measured. NFκB-P65, total and phosphorylated inhibitor of NFκB alpha (NFκBIA), and STAT-3 in nuclear extracts, interleukin (IL)-6 and IL-8 production in culture supernatants and cellular mRNA expression of 32 genes involved in Toll-like receptor-2 and -4 pathways were measured. Exposure to LPS/PepG caused increased total radical production (p = 0.022), decreased glutathione ratio (p = 0.016), reduced membrane potential and metabolic activity (both p < 0.0001), increased nuclear NFκB-P65 expression (p = 0.016) and increased IL-6/8 secretion (both p < 0.0001). MitoVitE, α- tocopherol and Trolox were similar in reducing oxidative stress, NFκB activation and interleukin secretion. MitoVitE had widespread downregulatory effects on gene expression. Despite differences in site of actions, all forms of vitamin E were protective under conditions mimicking sepsis. These results challenge the concept that protection inside mitochondria provides better protection.

Reply to Comment on “Differential Effects of MitoVitE, α-Tocopherol and Trolox on Oxidative Stress, Mitochondrial Function and Inflammatory Signalling Pathways in Endothelial Cells Cultured under Conditions Mimicking Sepsis. Antioxidants 2020, 9(3), 195”

“…Hegarty and Byrne are correct in that there are other biologically active forms of tocopherol; however, α-tocopherol is the most abundant in the body and therefore arguably the most important and a good choice for comparison, although we concur that investigating the relative effects of other tocopherols and tocotrienols would have been interesting. We agree that MitoVitE has been shown to be more protective than Trolox; nevertheless, it has been demonstrated that Trolox has potent antioxidant activity in several different models (references 7–9 in our paper [ 3 ]). We expected Trolox to be of less benefit than MitoVitE under conditions of sepsis in our model, but that was not what we found.…”

“…In our study [ 3 ], we used carboxy-DCFDA to measure the overall total cellular ROS production. Hegarty and Byrne suggest that we could have also used a mitochondrial-specific dye such as MitoSOX red.…”

SARS-CoV-2 infection pathogenesis is related to oxidative stress as a response to aggression

“…An in vitro sepsis model showed that vitamin E (α tocopherol), Trolox (soluble vitamin E), and motVit E (a polar derivative) reduced mitochondrial damage/dysfunction, NF-κB transcription molecule, and cytokine expression. MotVit E, which accumulates inside mitochondria, had a more efficient protective effect on genes linked to TLRs [60] . The treatment of animals with vitamin E in the model of acute sepsis also showed beneficial effects [61] .…”

Modulators of Oxidative Stress: Chemical and Pharmacological Aspects

“…Since organ dysfunction associated with sepsis is due, at least in part, to mitochondrial dysfunction and mitochondrial dysfunction is considered as main cause of oxidative stress in sepsis, antioxidants targeted to mitochondria have been considered as an efficient antioxidant protection in sepsis [ 12 ]. Minter et al [ 13 ] compared the positively charged antioxidant MitoVitE, which accumulates within mitochondria, with the membrane-bound α-tocopherol and water-soluble cytosolic Trolox with regard to their effects on oxidative stress, mitochondrial function and expression of key genes and proteins involved in the toll-like receptor (TLR)-2 and -4 inflammatory signalling pathways in the umbilical vein endothelial cell line HUVEC-C under conditions mimicking acute bacterial sepsis. Each of these three forms of vitamin E (5 μM each) was found to have similar reducing effects against induced oxidative stress and NFκB activation caused by exposure to lipopolysaccharide/peptidoglycan G. However, in difference to α-tocopherol and Trolox, MitoVitE had also widespread downregulatory effects on gene expression involved in TLR pathways, resulting in anti-inflammatory profile.…”