Typically, in case–control studies, healthy volunteers are included in the control group, but in this study we compared acute stroke with cerebral microangiopathy. This is due to the following consideration: acute cerebral ischemia itself is a factor causing rapid and significant changes in thiol homeostasis not only in the brain, but also in the blood plasma [13, 14]. Therefore, to more clearly reflect the effect of IS on plasma GSH homeostasis, it was more appropriate to use as a comparison group not healthy people characterized by a “normal” balance of thiols, but people at risk of stroke, which include patients with cerebral microangiopathy.
According to a number of studies, normal levels of tGSH and rGSH in plasma are ~ 4.3–9.1 and ~ 1.5–4.3 μM [24,25,26]. Our study has shown that in patients with acute stroke, these indicators are significantly lower than the normal level, and they are also lower than in patients with cerebral microangiopathy. This is consistent with the results of experimental studies, which showed that acute cerebral ischemia is accompanied by a significant decrease in the values of GSH . This is due to a decrease in its synthesis and / or intensive consumption by reason of a significant increase in the activity of GSH-dependent enzymes, for example, glutathione peroxidase . In addition, it was found that the activity of this enzyme in erythrocytes significantly decreased the risk of stroke . In addition, the level of glutathione peroxidase was negatively correlated with NIHSS on admission and after 7 days from stroke . Polymorphism of another GSH-metabolizing enzyme, glutathione S-transferase, is also associated with the risk of stroke .
It was shown on the rat brain ischemia/reperfusion model that intensification of GSH synthesis or its consumption attenuated oxidative stress and protected against the exacerbated brain damage [31, 32]. Part of GSH reacts with NO to form unstable nitrosoglutathione (GSNO). Pre-clinical studies showed positive neuroprotective effect of GSNO in animal stroke models . Other works have shown that glutathione-ethyl ester is able to effectively prevent oxidative damage of mitochondrial complex I caused by ischemia/reperfusion and thereby reduces the infarct size and improved neurological outcome [34, 35]. However, research on the effectiveness of GSH and the significance of its level for the diagnosis of various subtypes of stroke has not yet been conducted.
The results of our study showed that significant differences in the content of total aminothiols (cysteine, homocysteine, and GSH) exist between LA and CE groups. LA was characterized by elevated plasma levels of these metabolites, but a detailed analysis of these results is beyond the scope of this article. A lot of studies on the relationship between plasma total homocysteine level and stroke subtypes have been published; however, the results have been varied. Homocysteine has been recognized as an independent risk factor for stroke, and many studies have found a high incidence of elevated levels (hyperhomocysteinemia—HHcy) in all stroke subtypes [36,37,38,39]. An early study found no difference in this indicator among the subtypes of stroke . However, later, larger work and meta-analyses showed that the incidence of HHcy is higher in LA than in other subtypes, including CE [37, 38, 40,41,42], which is consistent with our results. In addition, CE is generally not characterized by a high incidence of HHcy , and an inverse association of CE risk with total homocysteine levels was revealed . The high homocysteine level in LA most likely reflects its proatherogenic role in the pathogenesis of stroke, and may also explain the increased levels of cysteine and GSH in plasma, since a significant part of homocysteine is converted to cysteine, and the latter, in turn, is required for the formation of GSH. In contrast, it was reported that the homocysteine level was higher in CE than in LA . There is a lack of data regarding the association of tGSH levels in different subtypes of stroke, which is due to the low content of this metabolite and the lack of sensitive routine methods for its determination in blood plasma. It has only recently been shown that low GSH levels are associated with neurological deficits in acute ischemic stroke .
Our results showed that low t- and rGSH levels may be a risk factor for increased neurological deficits in patients with LA, but not with CE stroke, even though CE was more commonly characterized by low tGSH level. In addition, a low level of tGSH is obviously associated with an increased risk of poor functional outcome in short term in LA, but in the case of CE this factor may play, on the contrary, a positive role.
In addition, a low level of rGSH can be considered a risk marker for decreased independence in patients with LA, but not CE in short term. It is unlikely that these results could be due to age, sex differences, or the different incidence of chronic cerebral ischemia between these groups of patients. It is highly likely that cysteine and/or homocysteine have a significant effect on the association of low GSH with stroke severity, but the limited sample size of this study did not allow such an analysis.
Correction of GSH levels, especially in LA, can have a positive therapeutic effect. Recently, in a small study, the positive effect of taking N-acetylcysteine, a compound that intensifies GSH synthesis, was shown on functional status and neurological deficits 3 months after acute ischemic stroke . In addition, it was shown that the infusion of N-acetylcysteine led to a rapid rise in GSH levels in the brain . However, it is not yet clear whether its protective effect was caused by the synthesis of GSH or the drug’s own antioxidant activity. The authors of these works also did not analyze its effectiveness among patients with LA and CE separately.
An increase in the level of total homocysteine and cysteine in LA patients may indicate a decrease in the bioavailability of cysteine as a rate-limiting substrate for GSH synthesis due to inhibition of the transport and/or synthesis of cysteine in cells. It is also interesting to note that in the LA group, in contrast to the CE group, we did not find significant associations of the total level of glutathione with cysteine and homocysteine, which may also indicate that this subtype of stroke is more commonly associated with systemic disruption of aminothiols metabolism.
The present work, according to our data, being the first pilot study to identify the diagnostic and prognostic potential of GSH in the subtypes of ischemic stroke, was carried out on a rather limited number of subjects, which did not allow us to investigate additional factors affecting the association of GSH with clinical indicators of stroke. Additional studies are required to help identify in more detail the role of plasma GSH as a risk marker for the unfavorable course of atherothrombotic stroke or explain its involvement in the pathology of stroke.