A Quantitative ELISA to Detect Anti-SARS-CoV-2 Spike IgG Antibodies in Infected Patients and Vaccinated Individuals.

There is an ongoing need for high-precision serological assays for the quantitation of anti-SARS-CoV-2 antibodies. Here, a trimeric SARS-CoV-2 spike (S) protein was used to develop an ELISA to quantify specific IgG antibodies present in serum, plasma, and dried blood spots (DBS) collected from infected patients or vaccine recipients. The quantitative S-ELISA was calibrated with international anti-SARS-CoV-2 immunoglobulin standards to provide test results in binding antibody units per mL (BAU/mL). The assay showed excellent linearity, precision, and accuracy. A sensitivity of 100% was shown for samples collected from 54 patients with confirmed SARS-CoV-2 infection more than 14 days after symptom onset or disease confirmation by RT-PCR and 58 vaccine recipients more than 14 days after vaccination. The assay specificity was 98.3%. Furthermore, antibody responses were measured in follow-up samples from vaccine recipients and infected patients. Most mRNA vaccine recipients had a similar response, with antibody generation starting 2-3 weeks after the first vaccination and maintaining positive for at least six months after a second vaccination. For most infected patients, the antibody titers increased during the second week after PCR confirmation. This S-ELISA can be used to quantify the seroprevalence of SARS-CoV-2 in the population exposed to the virus or vaccinated.

Aging triggers an upregulation of a multitude of cytokines in the male and especially the female rodent hippocampus but more discrete changes in other brain regions.

BACKGROUND: Despite widespread acceptance that neuroinflammation contributes to age-related cognitive decline, studies comparing protein expression of cytokines in the young versus old brains are surprisingly limited in terms of the number of cytokines and brain regions studied. Complicating matters, discrepancies abound-particularly for interleukin 6 (IL-6)-possibly due to differences in sex, species/strain, and/or the brain regions studied. METHODS: As such, we clarified how cytokine expression changes with age by using a Bioplex and Western blot to measure multiple cytokines across several brain regions of both sexes, using 2 mouse strains bred in-house as well as rats obtained from NIA. Parametric and nonparametric statistical tests were used as appropriate. RESULTS: In the ventral hippocampus of C57BL/6J mice, we found age-related increases in IL-1α, IL-1ß, IL-2, IL-3, IL-4, IL-6, IL-9, IL-10, IL-12p40, IL-12p70, IL-13, IL-17, eotaxin, G-CSF, interfeuron δ, KC, MIP-1a, MIP-1b, rantes, and TNFα that are generally more pronounced in females, but no age-related change in IL-5, MCP-1, or GM-CSF. We also find aging is uniquely associated with the emergence of a module (a.k.a. network) of 11 strongly intercorrelated cytokines, as well as an age-related shift from glycosylated to unglycosylated isoforms of IL-10 and IL-1ß in the ventral hippocampus. Interestingly, age-related increases in extra-hippocampal cytokine expression are more discreet, with the prefrontal cortex, striatum, and cerebellum of male and female C57BL/6J mice demonstrating robust age-related increase in IL-6 expression but not IL-1ß. Importantly, we found this widespread age-related increase in IL-6 also occurs in BALB/cJ mice and Brown Norway rats, demonstrating conservation across species and rearing environments. CONCLUSIONS: Thus, age-related increases in cytokines are more pronounced in the hippocampus compared to other brain regions and can be more pronounced in females versus males depending on the brain region, genetic background, and cytokine examined.

Loss of Function of Phosphodiesterase 11A4 Shows that Recent and Remote Long-Term Memories Can Be Uncoupled.

Systems consolidation is a process by which memories initially require the hippocampus for recent long-term memory (LTM) but then become increasingly independent of the hippocampus and more dependent on the cortex for remote LTM. Here, we study the role of phosphodiesterase 11A4 (PDE11A4) in systems consolidation. PDE11A4, which degrades cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), is preferentially expressed in neurons of CA1, the subiculum, and the adjacently connected amygdalohippocampal region. In male and female mice, deletion of PDE11A enhances remote LTM for social odor recognition and social transmission of food preference (STFP) despite eliminating or silencing recent LTM for those same social events. Measurement of a surrogate marker of neuronal activation (i.e., Arc mRNA) suggests the recent LTM deficits observed in Pde11 knockout mice correspond with decreased activation of ventral CA1 relative to wild-type littermates. In contrast, the enhanced remote LTM observed in Pde11a knockout mice corresponds with increased activation and altered functional connectivity of anterior cingulate cortex, frontal association cortex, parasubiculum, and the superficial layer of medial entorhinal cortex. The apparent increased neural activation observed in prefrontal cortex of Pde11a knockout mice during remote LTM retrieval may be related to an upregulation of the N-methyl-D-aspartate receptor subunits NR1 and NR2A. Viral restoration of PDE11A4 to vCA1 alone is sufficient to rescue both the LTM phenotypes and upregulation of NR1 exhibited by Pde11a knockout mice. Together, our findings suggest remote LTM can be decoupled from recent LTM, which may have relevance for cognitive deficits associated with aging, temporal lobe epilepsy, or transient global amnesia.

Identification of new PDE9A isoforms and how their expression and subcellular compartmentalization in the brain change across the life span.

3',5'-Cyclic nucleotide phosphodiesterases (PDEs) degrade 3',5' cyclic adenonosine monophosphate (cAMP) and 3',5' cyclic guanosine monophosphate (cGMP), with PDE9A having the highest affinity for cGMP. We show PDE9A6 and 3 novel PDE9 isoforms (PDE9X-100, PDE9X-120, and PDE9X-175) are reliably detected in the brain and lung of mice, whereas PDE9A2 and other isoforms are found elsewhere. PDE9A localizes to the membrane in all organs except the bladder, where it is cytosolic. Brain additionally shows PDE9 in the nuclear fraction. PDE9A mRNA expression/localization dramatically changes across neurodevelopment in a manner that is strikingly consistent between mice and humans (i.e., decreased expression in the hippocampus and cortex and inverted-U in the cerebellum). Study of the 4 PDE9 isoforms in the mouse brain from postnatal day 7 through 24 months similarly identifies dramatic effects of age on expression and subcellular compartmentalization that are isoform specific and brain region specific. Finally, PDE9A mRNA is elevated in the aged human hippocampus with dementia when there is a history of traumatic brain injury. Thus, brain PDE9 is localized to preferentially regulate nuclear- and membrane-proximal pools of cGMP, and its function likely changes across the life span.

Phosphodiesterase 11A (PDE11A), Enriched in Ventral Hippocampus Neurons, is Required for Consolidation of Social but not Nonsocial Memories in Mice.

The capacity to form long-lasting social memories is critical to our health and survival. cAMP signaling in the ventral hippocampal formation (VHIPP) appears to be required for social memory formation, but the phosphodiesterase (PDE) involved remains unknown. Previously, we showed that PDE11A, which degrades cAMP and cGMP, is preferentially expressed in CA1 and subiculum of the VHIPP. Here, we determine whether PDE11A is expressed in neurons where it could directly influence synaptic plasticity and whether expression is required for the consolidation and/or retrieval of social memories. In CA1, and possibly CA2, PDE11A4 is expressed throughout neuronal cell bodies, dendrites (stratum radiatum), and axons (fimbria), but not astrocytes. Unlike PDE2A, PDE9A, or PDE10A, PDE11A4 expression begins very low at postnatal day 7 (P7) and dramatically increases until P28, at which time it stabilizes to young adult levels. This expression pattern is consistent with the fact that PDE11A is required for social long-term memory (LTM) formation during adolescence and adulthood. Male and female PDE11 knockout (KO) mice show normal short-term memory (STM) for social odor recognition (SOR) and social transmission of food preference (STFP), but no LTM 24 h post training. Importantly, PDE11A KO mice show normal LTM for nonsocial odor recognition. Deletion of PDE11A may impair memory consolidation by impairing requisite protein translation in the VHIPP. Relative to WT littermates, PDE11A KO mice show reduced expression of RSK2 and lowered phosphorylation of S6 (pS6-235/236). Together, these data suggest PDE11A is selectively required for the proper consolidation of recognition and associative social memories.