Characterizing Long COVID in Children and Adolescents.

Importance: Most research to understand postacute sequelae of SARS-CoV-2 infection (PASC), or long COVID, has focused on adults, with less known about this complex condition in children. Research is needed to characterize pediatric PASC to enable studies of underlying mechanisms that will guide future treatment. Objective: To identify the most common prolonged symptoms experienced by children (aged 6 to 17 years) after SARS-CoV-2 infection, how these symptoms differ by age (school-age [6-11 years] vs adolescents [12-17 years]), how they cluster into distinct phenotypes, and what symptoms in combination could be used as an empirically derived index to assist researchers to study the likely presence of PASC. Design, Setting, and Participants: Multicenter longitudinal observational cohort study with participants recruited from more than 60 US health care and community settings between March 2022 and December 2023, including school-age children and adolescents with and without SARS-CoV-2 infection history. Exposure: SARS-CoV-2 infection. Main Outcomes and Measures: PASC and 89 prolonged symptoms across 9 symptom domains. Results: A total of 898 school-age children (751 with previous SARS-CoV-2 infection [referred to as infected] and 147 without [referred to as uninfected]; mean age, 8.6 years; 49% female; 11% were Black or African American, 34% were Hispanic, Latino, or Spanish, and 60% were White) and 4469 adolescents (3109 infected and 1360 uninfected; mean age, 14.8 years; 48% female; 13% were Black or African American, 21% were Hispanic, Latino, or Spanish, and 73% were White) were included. Median time between first infection and symptom survey was 506 days for school-age children and 556 days for adolescents. In models adjusted for sex and race and ethnicity, 14 symptoms in both school-age children and adolescents were more common in those with SARS-CoV-2 infection history compared with those without infection history, with 4 additional symptoms in school-age children only and 3 in adolescents only. These symptoms affected almost every organ system. Combinations of symptoms most associated with infection history were identified to form a PASC research index for each age group; these indices correlated with poorer overall health and quality of life. The index emphasizes neurocognitive, pain, and gastrointestinal symptoms in school-age children but change or loss in smell or taste, pain, and fatigue/malaise-related symptoms in adolescents. Clustering analyses identified 4 PASC symptom phenotypes in school-age children and 3 in adolescents. Conclusions and Relevance: This study developed research indices for characterizing PASC in children and adolescents. Symptom patterns were similar but distinguishable between the 2 groups, highlighting the importance of characterizing PASC separately for these age ranges.

Astrocytes Do Not Forfeit Their Neuroprotective Roles After Surviving Intense Oxidative Stress.

In order to fulfill their evolutionary role as support cells, astrocytes have to tolerate intense oxidative stress under conditions of brain injury and disease. It is well known that astrocytes exposed to mild oxidative stress are preconditioned against subsequent stress exposure in dual hit models. However, it is unclear whether severe oxidative stress leads to stress tolerance, stress exacerbation, or no change in stress resistance in astrocytes. Furthermore, it is not known whether reactive astrocytes surviving intense oxidative stress can still support nearby neurons. The data in this Brief Report suggest that primary cortical astrocytes surviving high concentrations of the oxidative toxicant paraquat are completely resistant against subsequent oxidative challenges of the same intensity. Inhibitors of multiple endogenous defenses (e.g., glutathione, heme oxygenase 1, ERK1/2, Akt) failed to abolish or even reduce their stress resistance. Stress-reactive cortical astrocytes surviving intense oxidative stress still managed to protect primary cortical neurons against subsequent oxidative injuries in neuron/astrocyte co-cultures, even at concentrations of paraquat that otherwise led to more than 80% neuron loss. Although our previous work demonstrated a lack of stress tolerance in primary neurons exposed to dual paraquat hits, here we show that intensely stressed primary neurons can resist a second hit of hydrogen peroxide. These collective findings suggest that stress-reactive astroglia are not necessarily neurotoxic, and that severe oxidative stress does not invariably lead to stress exacerbation in either glia or neurons. Therefore, interference with the natural functions of stress-reactive astrocytes might have the unintended consequence of accelerating neurodegeneration.

Evidence for cross-hemispheric preconditioning in experimental Parkinson's disease.

Dopamine loss and motor deficits in Parkinson's disease typically commence unilaterally and remain asymmetric for many years, raising the possibility that endogenous defenses slow the cross-hemispheric transmission of pathology. It is well-established that the biological response to subtoxic stress prepares cells to survive subsequent toxic challenges, a phenomenon known as preconditioning, tolerance, or stress adaptation. Here we demonstrate that unilateral striatal infusions of the oxidative toxicant 6-hydroxydopamine (6-OHDA) precondition the contralateral nigrostriatal pathway against the toxicity of a second 6-OHDA infusion in the opposite hemisphere. 6-OHDA-induced loss of dopaminergic terminals in the contralateral striatum was ablated by cross-hemispheric preconditioning, as shown by two independent markers of the dopaminergic phenotype, each measured by two blinded observers. Similarly, loss of dopaminergic somata in the contralateral substantia nigra was also abolished, according to two blinded measurements. Motor asymmetries in floor landings, forelimb contacts with a wall, and spontaneous turning behavior were consistent with these histological observations. Unilateral 6-OHDA infusions increased phosphorylation of the kinase ERK2 and expression of the antioxidant enzyme CuZn superoxide dismutase in both striata, consistent with our previous mechanistic work showing that these two proteins mediate preconditioning in dopaminergic cells. These findings support the existence of cross-hemispheric preconditioning in Parkinson's disease and suggest that dopaminergic neurons mount impressive natural defenses, despite their reputation as being vulnerable to oxidative injury. If these results generalize to humans, Parkinson's pathology may progress slowly and asymmetrically because exposure to a disease-precipitating insult induces bilateral upregulation of endogenous defenses and elicits cross-hemispheric preconditioning.

Transmission of α-synucleinopathy from olfactory structures deep into the temporal lobe.

BACKGROUND: α-synucleinopathy emerges quite early in olfactory structures such as the olfactory bulb and anterior olfactory nucleus (OB/AON) in Parkinson's disease. This may contribute to smell impairments years before the commencement of motor symptoms. We tested whether α-synucleinopathy can spread from the OB/AON to regions of the limbic telencephalon that harbor connections with olfactory structures. FINDINGS: α-synuclein fibrils were infused into the OB/AON. Inclusions containing pathologically phosphorylated α-synuclein (pSer129) were observed three months later in the piriform and entorhinal cortices, amygdala, and hippocampal formation. The retrograde tract-tracer FluoroGold confirmed the existence of first-order afferents at these sites. Some sites harbored FluoroGold(+) neurons but no inclusions, suggestive of selective vulnerabilities. Multiple areas close to the injection site but not connected with the OB/AON remained free of inclusions, suggesting a lack of widespread uptake of fibrils from interstitial diffusion. Two independent pSer129 antibodies revealed the same labeling patterns and preadsorption control experiments confirmed a loss of pSer129 staining. Dense total α-synuclein (but not pSer129) staining was apparent in the OB/AON 1.5 h following fibril infusions, suggesting that pSer129(+) staining did not reflect exogenously infused material. Waterbath sonication of fibrils for 1 h improved α-synucleinopathy transmission relative to 1 min-long probe sonication. Electron microscopy revealed that longer sonication durations reduced fibril size. The Thioflavin stain labeled cells at the infusion site and some, but not all inclusions contained ubiquitin. Three-dimensional confocal analyses revealed that many inclusions ensconced NeuN(+) neuronal nuclei. Young and aged mice exhibited similar topographical spread of α-synucleinopathy. CONCLUSIONS: 1) α-synucleinopathy in this model is transmitted through some, but not all neuroanatomical connections, 2) pathology is largely confined to first-order afferent sites at three months and this is most parsimoniously explained by retrograde transport, and 3) transmission in aged animals is largely similar to that in young control animals at three months post-infusion.

Astrocytes Surviving Severe Stress Can Still Protect Neighboring Neurons from Proteotoxic Injury.

Astrocytes are one of the major cell types to combat cellular stress and protect neighboring neurons from injury. In order to fulfill this important role, astrocytes must sense and respond to toxic stimuli, perhaps including stimuli that are severely stressful and kill some of the astrocytes. The present study demonstrates that primary astrocytes that managed to survive severe proteotoxic stress were protected against subsequent challenges. These findings suggest that the phenomenon of preconditioning or tolerance can be extended from mild to severe stress for this cell type. Astrocytic stress adaptation lasted at least 96 h, the longest interval tested. Heat shock protein 70 (Hsp70) was raised in stressed astrocytes, but inhibition of neither Hsp70 nor Hsp32 activity abolished their resistance against a second proteotoxic challenge. Only inhibition of glutathione synthesis abolished astrocytic stress adaptation, consistent with our previous report. Primary neurons were plated upon previously stressed astrocytes, and the cocultures were then exposed to another proteotoxic challenge. Severely stressed astrocytes were still able to protect neighboring neurons against this injury, and the protection was unexpectedly independent of glutathione synthesis. Stressed astrocytes were even able to protect neurons after simultaneous application of proteasome and Hsp70 inhibitors, which otherwise elicited synergistic, severe loss of neurons when applied together. Astrocyte-induced neuroprotection against proteotoxicity was not elicited with astrocyte-conditioned media, suggesting that physical cell-to-cell contacts may be essential. These findings suggest that astrocytes may adapt to severe stress so that they can continue to protect neighboring cell types from profound injury.