Perfis imunogenéticos e sua reação com as suscetibilidades à síndrome pós-aguda da Covid-19: estudo experimental com a proteína Spike em modelos murinos Th1 e Th2

Abstract

COVID-19, caused by SARS-CoV-2, has been associated with persistent extrapulmonary manifestations, including neuropsychiatric symptoms, characterizing Post-Acute COVID-19 Syndrome (PACS). The Spike protein, the main structural component of the virus, can cross the blood-brain barrier and induce neuroinflammation. Considering the distinct immunological profiles of BALB/c (Th2) and C57BL/6 (Th1) mouse strains, this study investigated the effects of Spike protein in these strains, focusing on persistent immune and behavioral alterations associated with PACS. We aimed to evaluate behavioral, neurochemical, and immunological alterations induced by exposure to Spike protein in BALB/c and C57BL/6 mice, examining the influence of Th1 and Th2 immune responses on the persistence of PACS symptoms. In the in vitro model, murine astrocytes and macrophages (RAW 264.7 cells) were exposed to Spike protein (10 μg) for analysis of cytotoxicity (MTT), morphology and nitric oxide production. In the in vivo model, male BALB/c and C57BL/6 mice received saline (control) or Spike (10 μg/animal, intraperitoneal, single exposure) and were evaluated in two phases: (i) acute-AF (up to 7 days after exposure); (ii) post-acute-FPA (40–45 days after exposure). Behavioral tests included assessment of locomotor activity (open field test), spatial learning (T-maze test), non-spatial declarative memory (object recognition test), anxiety-like behavior (zero maze test) and depressive-like behavior (forced swim test). Cytokines were analyzed in the hippocampus (Mouse XL Cytokine Array) and dopamine and its metabolite DOPAC in the striatum (HPLC). In in vitro tests, the protein showed a more pronounced cytotoxic effect on RAW 264.7 macrophages (81.70%) than on astrocytes (39.28%). Exposure to Spike induced marked behavioral changes in BALB/c mice. In the in vivo model, BALB/c mice exposed to Spike in the AF exhibited anxious-depressive behavior, with increased immobility time in the forced swim test (P<0.0001), decreased distance traveled (P<0.0001), and longer time in the open follow-up (Zero Maze, P<0.0001). There was a reduction in the number of rearings in the post-acute phase (P = 0.0299), an increase in locomotor activity and a significant increase in the time spent in the central zone of the open field (P = 0.0001), and impairment in working memory (T-maze test). Neurochemically, there was an increase in DOPAC levels, indicating greater dopaminergic turnover. Cytokine analysis revealed low expression of IL-1ra, IL-15 and BDNF in the hippocampus. In contrast, C57BL/6 demonstrated behavioral and neurochemical resilience, with a more robust inflammatory response (mean cytokine expression: 12.33 ± 2.33 vs. 3.15 ± 1.81 in BALB/c; P < 0.0001), with increased IL-6, IL-22, IL-23, MIP-3β, IL-1ra and BDNF, consistent with a Th1 profile. In FA, there was reduced time in the locomotor part (P=0.0362), greater number of grooming (P=0.0191), while in FPA there was cognitive deficit in object recognition. BALB/c animals showed greater susceptibility to SPAC, with behavioral changes, dopaminergic dysfunction and deficient immune response. In contrast, C57BL/6 showed greater behavioral resilience and more efficient immune response. The data reinforces the role of the immunogenetic profile in vulnerability to long COVID and the translational utility of the BALB/c model.