Keywords: SARS-Cov-2 spike protein; mRNA vaccines; prion and prion-like diseases; p53; Wip1; autophagy; aging; senescence; COVID-19.
1. Introduction and Background
A significant percentage of patients suffering from SARS-CoV-2 develop neurological and cognitive impairments, sometimes lasting long after the infection has cleared. This condition has been named “long haul COVID disease,” or simply “long COVID,” also known as “PASC” (Post-Acute Sequelae of SARS CoV-2 infection). An international study quantified persistent long-COVID symptoms among 3,762 individuals following a SARS-CoV-2 infection. These authors wrote: “Memory and cognitive dysfunction, experienced by over 88% of respondents, were the most pervasive and persisting neurologic symptoms in this cohort, equally common across all ages, and with substantial impact on work and daily life. Memory and cognitive dysfunction, together with other commonly reported neuropsychiatric symptoms, may point to larger neurological issues involving both the central and peripheral nervous system” [1]. A post-mortem study of the brains of three patients who died from severe COVID-19 showed a large number of activated microglia that were associated with overexpression of inflammatory markers, including Interleukin-1β (IL-1β) and IL-6. The authors suggested that oxidative stress induced a glial-mediated neuroinflammatory response leading to neuronal injury [2].
A growing consensus attributes these symptoms to neurotoxic effects of the spike glycoprotein, particularly the S1 subunit [3]. The receptor-binding domain of SARS-CoV-2 spike S1 protein binds to heparin and to heparin-binding proteins [4]. Idrees and Kumar wrote in their conclusion: ”Our results indicate stable binding of the S1 protein to these aggregation-prone proteins which might initiate aggregation of brain protein and accelerate neurodegeneration” [4]. A study evaluating the amyloidogenic potential of the spike protein verified that the spike protein can cause amyloid-like fibrils to appear after the protein has been subjected to proteolysis. A specific segment that appeared following proteolysis, spike 194-213 (FKNIDGYFKI), was demonstrated both theoretically and experimentally to be amyloidogenic [5]. A study by Kruger et al. found that proteolysis resistant fibrin amyloid microclots accumulate in the blood in association with PASC, and this also suggests that the spike protein has amyloidogenic properties [6].
Direct experimental evidence of S1’s toxic effects in the brain comes from studies conducted by a team of Korean researchers, published in 2022 [7]. In the experiment, S1 subunits were introduced directly into the dorsal hippocampus of mice, and it was shown that the mice subsequently suffered from anxiety-like behavior and cognitive deficits. Further experiments both in vivo and in vitro found that the effects were mediated by microglia, which became activated following exposure. The microglia released excitatory cytokines, in particular IL-1β. IL-1β expression was upregulated more than seven-fold in the hippocampi of the exposed mice. Morphologically, the microglia of the exposed mice acquired the features of reactive microglia.
In this paper, we attempt to trace the likely biological pathways by which neuronal damage occurs in response to the spike protein, particularly S1. We will argue based on the emerging literature that toll-like receptor 4 signaling is central to the destructive reaction process. An important intermediary is the MAPK cascade. MAPK comprises four distinct pathways, a) the extracellular signal regulated kinase 1 and 2 (ERK1/2), b) the ERK-big MAP kinase 1(BMK1), c) the c-Jun NH2-terminal kinases (JNK) or stress activated protein kinases (SAPKs), and d) the p38 MAPKs. The ERK pathways are stimulated by growth factors, hormones and pro-inflammatory stimuli whereas the JNK and p38 MAPK are activated by cellular and environmental stress signals in addition to pro-inflammatory stimuli [8,9]. It is these latter two pathways that we will argue play a primary role in spike protein neurotoxicity.
Recent neurotoxicity studies indicate that the SARS-CoV-2 S1 subunit induces neuro-inflammation in microglial cells, a special type of macrophage in the central nervous system (CNS) [10,11]. The neuroinflammatory response is mediated by p38 MAPK and nuclear factor κ-light chain enhancer of activated B cells (NF-κB) activation, mainly through the pattern recognition receptor TLR4. In addition, the SARS CoV-2 S1 subunit elicits a pro-inflammatory response in murine and human macrophages by activating TLR4 receptor signaling. In this signaling process, both JNK and p38 are activated by phosphorylation [12]. It is important to note that infectious prions also activate the p38 MAPK pathway to induce their neurotoxicity effects [13]. The spike protein has prion-like characteristics that may contribute to its neurotoxicity. We will return to this topic in great detail later.