Sunil Jamuna Tripathi

Dysregulated dopamine (DA) signaling and redox homeostasis contributes to multiple neuropsychiatric and neurodegenerative disorders. Polymorphisms that influence the activity of catechol-O-methyltransferase (COMT), an enzyme critical for degrading DA in the dorsolateral prefrontal cortex, have been implicated in behavioral and neuropsychiatric alterations associated with schizophrenia (SCZ). Adverse neuropsychiatric effects have also been reported in Parkinson’s disease (PD) patients administered COMT inhibitors in combination with other DA-enhancing therapies. COMT exists as two isoforms: a soluble short isoform (S-COMT) and a membrane-bound long isoform (MB-COMT). These variants differ in their N-terminal domains, with MB-COMT being the predominant brain isoform. Here, unbiased proteomic and biochemical analyses show that genetic loss of MB-COMT disrupts DA signaling and perturbs pathways governing synaptic and mitochondrial function, iron and copper homeostasis, and redox balance. Limited proteolysis mass spectrometry (LiP-MS) further revealed that MB-COMT deficiency triggers widespread protein structural alterations, a molecular event commonly occurring in neurodegenerative conditions but not as well studied in neuropsychiatric diseases. Our results show that MB-COMT is a molecular hub that connects multiple cellular pathways whose differential dysregulation underlies the pathophysiology of complex neuropsychiatric diseases such as SCZ. Thus, MB-COMT is identified as a key regulator of brain DA biology, loss of which activates cellular stress response pathways, revealing potential targets for therapeutic intervention.

Cellular homeostasis relies on regulation of processes, including protein post-translational modifications (PTMs) and biomolecular condensation. Aging disrupts the equilibrium of these processes, increasing susceptibility to disease and mortality. Here we used chemoproteomic techniques to generate an atlas of cysteine PTMs in the mouse brain and showed that age-related increases in thiol oxidation promoted the formation of biomolecular condensates. By contrast, protein persulfidation, regulated by hydrogen sulfide production, inhibited biomolecular condensation, preserving protein function. Age-induced alterations in cysteine PTMs influenced the phase separation properties of synapsin 1 and G3BP2, leading to impaired neurotransmitter release and defective stress granule formation and resolution, features associated with aging and neurodegenerative diseases. Mice deficient in cystathionine γ-lyase, the enzyme responsible for hydrogen sulfide production, exhibited reduced lifespans and spontaneously developed protein aggregates with age. Our results highlight the therapeutic potential of protein persulfidation in reversal of dysregulated biomolecular condensation and suggest that sulfide donors could be used to mitigate age-related diseases.

Deterioration of the blood–brain barrier (BBB), including impaired neurovascular uncoupling, contributes to cognitive decline in aging. The BBB is formed principally by brain microvascular endothelial cells (ECs), and ECs throughout the body are enriched for the transcription factor Krüppel-like factor 4 (KLF4). Because KLF4 levels in ECs decrease with age, we tested whether that decline contributes to aging-related BBB deterioration, neurovascular dysfunction, and cognitive impairment. Using EC-specific Klf4 knockout mice (EC-K4KO), we show that loss of EC KLF4 accelerates multiple age-related brain pathologies. Indeed, middle-aged EC-K4KO mice display pathological features that are not normally observed until advanced age, including marked BBB leakage, impaired neurovascular coupling, loss of microvessels, increased oxidative damage, neuroinflammation, neurodegeneration, anxiety-like behavior, and cognitive deficits. Single-cell RNA sequencing of brain vasculature reveals dysregulation of immune response and barrier-related genes in ECs lacking KLF4, indicating that KLF4 maintains brain endothelial homeostasis by constraining proinflammatory and senescence programs at the chromatin level. Together, these results identify loss of EC KLF4 as a key driver of neurovascular decline and age-associated cognitive dysfunction.

Alzheimer’s disease (AD) and traumatic brain injury (TBI) are currently untreatable neurodegenerative disorders afflicting millions of people worldwide. These conditions are pathologically related, and TBI is one of the greatest risk factors for AD. Although blood–brain barrier (BBB) disruption drives progression of both AD and TBI, strategies to preserve BBB integrity have been hindered by lack of actionable targets. Here, we identify 15-hydroxyprostaglandin dehydrogenase (15-PGDH), an enzyme that catabolizes eicosanoids and other anti-inflammatory mediators, as a therapeutic candidate that protects the BBB. We demonstrate that 15-PGDH is enriched in BBB-associated myeloid cells and becomes markedly elevated in human and mouse models of AD and TBI, as well as aging, another major risk factor for AD. Pathological increase in 15-PGDH correlates with pronounced oxidative stress, neuroinflammation, and neurodegeneration, alongside profound BBB structural degeneration characterized by astrocytic endfeet swelling and functional impairment. Pharmacologic inhibition or genetic reduction of 15-PGDH in AD and TBI models strikingly mitigates oxidative damage, suppresses neuroinflammation, and restores BBB integrity. Most notably, inhibiting 15-PGDH not only halts neurodegeneration but also preserves cognitive function at levels indistinguishable from healthy controls. Remarkably, these neuroprotective effects in AD are achieved without affecting amyloid pathology, underscoring a noncanonical mechanism for treating AD. In a murine microglia cell line exposed to amyloid beta oligomer, major protection was demonstrated by multiple anti-inflammatory substrates that 15-PGDH degrades. Thus, our findings position 15-PGDH inhibition as a broad-spectrum strategy to protect the BBB and thereby preserve brain health and cognition in AD and TBI.

Rationale Anxiety is one of the most comorbid conditions with major depressive disorder (MDD). Depression-associated anxiety often stems from the dysfunctional hypothalamic-pituitary-adrenal (HPA) axis and its altered regulation by the amygdala. Furthermore, MDD is associated with altered glutamatergic processing leading to anxiety and impaired regulation of the HPA axis. Recent studies have demonstrated that N-acetyl cysteine (NAC), a pleiotropic drug, exerts antidepressant-like effect by modulation of hippocampal functions, periterminal release of glutamate, and/or redox systems. However, the effects of NAC on depression-associated anxiety, HPA axis hyperactivity, and amygdalar dysfunctions are relatively unknown. Objectives Accordingly, we evaluated the effect of NAC on neonatal clomipramine (CLI)-induced adulthood anxiety and accompanying changes in plasma corticosterone levels, amygdalar volumes, neuronal/glial densities, levels of monoamines, and their metabolites in the amygdalar complex. Results We found that chronic treatment with NAC reverses CLI-induced anhedonia and enhanced anxiety. Interestingly, attenuation of CLI-associated anxiety in NAC-treated rats were accompanied by a reversal of adrenal and spleen hypertrophy, and normalization of enhanced plasma corticosterone levels, indicating improved HPA axis functioning. Furthermore, NAC treatment was sufficient to reverse volumetric hypertrophy of basolateral amygdala (BLA), and altered noradrenaline (NA) metabolism in the amygdalar complex. The effects of NAC in the reversal of CLI-induced impairments were similar to that of fluoxetine (FLX). Conclusions We suggest that beneficial effects of NAC on antidepressive- and antianxiety-like behaviors are at least in part mediated via restoration of amygdalar and HPA axis functioning. Our results support the hypothesis that NAC might be evolved as a therapeutic strategy for reversal of amygdalar dysfunction in depression.