The down syndrome biomarker initiative (DSBI) pilot: proof of concept for deep phenotyping of Alzheimer’s disease biomarkers in down syndrome

To gain further knowledge on the preclinical phase of AD, we sought to characterize cognitive performance, volumetric MRI, amyloid PET, FDG PET, retinal amyloid, and plasma biomarkers in a cohort of non-demented adults with Down Syndrome (DS). The goal of the Down Syndrome Biomarker Initiative (DSBI) pilot is to test feasibility of this approach for future multicenter studies. We enrolled 12 non-demented participants with DS between the ages of 30-60 years old. Participants underwent extensive cognitive testing, volumetric MRI, amyloid PET 18F-florbetapir, 18F-fluorodeoxyglucose (18F-FDG) PET, and retinal amyloid imaging. In addition, plasma beta-amyloid species were measured and ApoE genotyping was performed. Consistent with previous autopsy studies, subjects demonstrated amyloid PET positivity reflecting fibrillar amyloid plaque deposition. Results from our multimodal analysis also suggest greater hippocampal atrophy with amyloid load. Additionally, we identified an inverse relationship between amyloid load and regional glucose metabolism. Cognitive and functional measures did not correlate with amyloid load in DS but did correlate with regional FDG PET measures. Retinal amyloid imaging demonstrated presence of plaques. Biomarkers of AD can be readily studied in adults with DS as in other preclinical AD populations. Importantly, all subjects in this feasibility study were able to complete all test procedures. The data indicate that a large, multicenter longitudinal study is feasible to better understand the trajectories of AD biomarkers in this enriched population. This trial is registered with ClinicalTrials.gov, number NCT02141971

A novel Alzheimer disease locus located near the gene encoding tau protein

This is the author accepted manuscript. The final version is available from the publisher via the DOI in this recordAPOE ε4, the most significant genetic risk factor for Alzheimer disease (AD), may mask effects of other loci. We re-analyzed genome-wide association study (GWAS) data from the International Genomics of Alzheimer's Project (IGAP) Consortium in APOE ε4+ (10 352 cases and 9207 controls) and APOE ε4- (7184 cases and 26 968 controls) subgroups as well as in the total sample testing for interaction between a single-nucleotide polymorphism (SNP) and APOE ε4 status. Suggestive associations (P<1 × 10-4) in stage 1 were evaluated in an independent sample (stage 2) containing 4203 subjects (APOE ε4+: 1250 cases and 536 controls; APOE ε4-: 718 cases and 1699 controls). Among APOE ε4- subjects, novel genome-wide significant (GWS) association was observed with 17 SNPs (all between KANSL1 and LRRC37A on chromosome 17 near MAPT) in a meta-analysis of the stage 1 and stage 2 data sets (best SNP, rs2732703, P=5·8 × 10-9). Conditional analysis revealed that rs2732703 accounted for association signals in the entire 100-kilobase region that includes MAPT. Except for previously identified AD loci showing stronger association in APOE ε4+ subjects (CR1 and CLU) or APOE ε4- subjects (MS4A6A/MS4A4A/MS4A6E), no other SNPs were significantly associated with AD in a specific APOE genotype subgroup. In addition, the finding in the stage 1 sample that AD risk is significantly influenced by the interaction of APOE with rs1595014 in TMEM106B (P=1·6 × 10-7) is noteworthy, because TMEM106B variants have previously been associated with risk of frontotemporal dementia. Expression quantitative trait locus analysis revealed that rs113986870, one of the GWS SNPs near rs2732703, is significantly associated with four KANSL1 probes that target transcription of the first translated exon and an untranslated exon in hippocampus (P≤1.3 × 10-8), frontal cortex (P≤1.3 × 10-9) and temporal cortex (P≤1.2 × 10-11). Rs113986870 is also strongly associated with a MAPT probe that targets transcription of alternatively spliced exon 3 in frontal cortex (P=9.2 × 10-6) and temporal cortex (P=2.6 × 10-6). Our APOE-stratified GWAS is the first to show GWS association for AD with SNPs in the chromosome 17q21.31 region. Replication of this finding in independent samples is needed to verify that SNPs in this region have significantly stronger effects on AD risk in persons lacking APOE ε4 compared with persons carrying this allele, and if this is found to hold, further examination of this region and studies aimed at deciphering the mechanism(s) are warranted

Dental Caries and Enamel Defects in Very Low Birth Weight Adolescents

OBJECTIVES: The purpose of this study was to examine developmental enamel defects and dental caries in very low birth weight adolescents with high risk (HR-VLBW) and low risk (LR-VLBW) compared to full-term (term) adolescents. METHODS: The sample consisted of 224 subjects (80 HR-VLBW, 59 LR-VLBW, 85 term adolescents) recruited from an ongoing longitudinal study. Sociodemographic and medical information was available from birth. Dental examination of the adolescent at the 14-year visit included: enamel defects (opacity and hypoplasia); decayed, missing, filled teeth of incisors and molars (DMFT-IM) and of overall permanent teeth (DMFT); Simplified Oral Hygiene Index for debris/calculus on teeth, and sealant presence. A caregiver questionnaire completed simultaneously assessed dental behavior, access, insurance status and prevention factors. Hierarchical analysis utilized the zero-inflated negative binomial model and zero-inflated Poisson model. RESULTS: The zero-inflated negative binomial model controlling for sociodemographic variables indicated that the LR-VLBW group had an estimated 75% increase (p < 0.05) in number of demarcated opacities in the incisors and first molar teeth compared to the term group. Hierarchical modeling indicated that demarcated opacities were a significant predictor of DMFT-IM after control for relevant covariates. The term adolescents had significantly increased DMFT-IM and DMFT scores compared to the LR-VLBW adolescents. CONCLUSION: LR-VLBW was a significant risk factor for increased enamel defects in the permanent incisors and first molars. Term children had increased caries compared to the LR-VLBW group. The effect of birth group and enamel defects on caries has to be investigated longitudinally from birth

Umbilical Cord-derived Mesenchymal Stem Cells modulate TNF and soluble TNF Receptor 2 (sTNFR2) in COVID-19 ARDS patients

OBJECTIVEWe aimed at explaining the mechanism of therapeutic effect of Umbilical Cord Mesenchymal Stem Cells (UC-MSC) in subjects with COVID-19 Acute Respiratory Distress Syndrome (ARDS). Patients with COVID-19 ARDS present with a hyperinflammatory response characterized by high levels of circulating pro-inflammatory mediators, including tumor necrosis factor α and β (TNFα and TNFβ). Inflammatory functions of these TNFs can be inhibited by soluble TNF Receptor 2 (sTNFR2). In patients with COVID-19 ARDS, UC-MSC appear to impart a robust anti-inflammatory effect, and treatment is associated with remarkable clinical improvements. We investigated the levels of TNFα, TNFβ and sTNFR2 in blood plasma samples collected from subjects with COVID-19 ARDS enrolled in our trial of UC-MSC treatment. PATIENTS AND METHODSWe analyzed plasma samples from subjects with COVID-19 ARDS (n=24) enrolled in a Phase 1/2a randomized controlled trial of UC-MSC treatment. Plasma samples were obtained at Day 0 (baseline, before UC-MSC or control infusion), and Day 6 post infusion. Plasma concentrations of sTNFR2, TNFα, and TNFβ were evaluated using a quantitative multiplex protein array. RESULTSOur data indicate that at Day 6 after infusion, UC-MSC recipients develop significantly increased levels of plasma sTNFR2 and significantly decreased levels of TNFα and TNFβ, compared to controls. CONCLUSIONSThese observations suggest that sTNFR2 plays a mechanistic role in mediating UC-MSC effect on TNFα and TNFβ plasma levels, determining a decrease in inflammation in COVID-19 ARDS

The down syndrome biomarker initiative (DSBI) pilot: proof of concept for deep phenotyping of Alzheimer's disease biomarkers in down syndrome.

To gain further knowledge on the preclinical phase of Alzheimer's disease (AD), we sought to characterize cognitive performance, neuroimaging and plasma-based AD biomarkers in a cohort of non-demented adults with down syndrome (DS). The goal of the down syndrome biomarker Initiative (DSBI) pilot is to test feasibility of this approach for future multicenter studies. We enrolled 12 non-demented participants with DS between the ages of 30-60 years old. Participants underwent extensive cognitive testing, volumetric MRI, amyloid positron emission tomography (PET; 18F-florbetapir), fluorodeoxyglucose (FDG) PET (18F-fluorodeoxyglucose) and retinal amyloid imaging. In addition, plasma beta-amyloid (Aβ) species were measured and Apolipoprotein E (ApoE) genotyping was performed. Results from our multimodal analysis suggest greater hippocampal atrophy with amyloid load. Additionally, we identified an inverse relationship between amyloid load and regional glucose metabolism. Cognitive and functional measures did not correlate with amyloid load in DS but did correlate with regional FDG PET measures. Biomarkers of AD can be readily studied in adults with DS as in other preclinical AD populations. Importantly, all subjects in this feasibility study were able to complete all test procedures. The data indicate that a large, multicenter longitudinal study is feasible to better understand the trajectories of AD biomarkers in this enriched population. This trial is registered with ClinicalTrials.gov, number NCT02141971

Carbonic anhydrase-8 regulates inflammatory pain by inhibiting the ITPR1-cytosolic free calcium pathway.

Calcium dysregulation is causally linked with various forms of neuropathology including seizure disorders, multiple sclerosis, Huntington’s disease, Alzheimer’s, spinal cerebellar ataxia (SCA) and chronic pain. Carbonic anhydrase-8 (Car8) is an allosteric inhibitor of inositol trisphosphate receptor-1 (ITPR1), which regulates intracellular calcium release fundamental to critical cellular functions including neuronal excitability, neurite outgrowth, neurotransmitter release, mitochondrial energy production and cell fate. In this report we test the hypothesis that Car8 regulation of ITPR1 and cytoplasmic free calcium release is critical to nociception and pain behaviors. We show Car8 null mutant mice (MT) exhibit mechanical allodynia and thermal hyperalgesia. Dorsal root ganglia (DRG) from MT also demonstrate increased steady-state ITPR1 phosphorylation (pITPR1) and cytoplasmic free calcium release. Overexpression of Car8 wildtype protein in MT nociceptors complements Car8 deficiency, down regulates pITPR1 and abolishes thermal and mechanical hypersensitivity. We also show that Car8 nociceptor overexpression alleviates chronic inflammatory pain. Finally, inflammation results in downregulation of DRG Car8 that is associated with increased pITPR1 expression relative to ITPR1, suggesting a possible mechanism of acute hypersensitivity. Our findings indicate Car8 regulates the ITPR1-cytosolic free calcium pathway that is critical to nociception, inflammatory pain and possibly other neuropathological states. Car8 and ITPR1 represent new therapeutic targets for chronic pain

Carbonic Anhydrase-8 Regulates Inflammatory Pain by Inhibiting the ITPR1-Cytosolic Free Calcium Pathway

<div><p>Calcium dysregulation is causally linked with various forms of neuropathology including seizure disorders, multiple sclerosis, Huntington’s disease, Alzheimer’s, spinal cerebellar ataxia (SCA) and chronic pain. Carbonic anhydrase-8 (Car8) is an allosteric inhibitor of inositol trisphosphate receptor-1 (ITPR1), which regulates intracellular calcium release fundamental to critical cellular functions including neuronal excitability, neurite outgrowth, neurotransmitter release, mitochondrial energy production and cell fate. In this report we test the hypothesis that Car8 regulation of ITPR1 and cytoplasmic free calcium release is critical to nociception and pain behaviors. We show Car8 null mutant mice (MT) exhibit mechanical allodynia and thermal hyperalgesia. Dorsal root ganglia (DRG) from MT also demonstrate increased steady-state ITPR1 phosphorylation (pITPR1) and cytoplasmic free calcium release. Overexpression of Car8 wildtype protein in MT nociceptors complements Car8 deficiency, down regulates pITPR1 and abolishes thermal and mechanical hypersensitivity. We also show that Car8 nociceptor overexpression alleviates chronic inflammatory pain. Finally, inflammation results in downregulation of DRG Car8 that is associated with increased pITPR1 expression relative to ITPR1, suggesting a possible mechanism of acute hypersensitivity. Our findings indicate Car8 regulates the ITPR1-cytosolic free calcium pathway that is critical to nociception, inflammatory pain and possibly other neuropathological states. Car8 and ITPR1 represent new therapeutic targets for chronic pain.</p></div

Gene transfer of V5-Car8^WT regulates nociception and produces analgesia in MT mice.

<p>Sciatic nerve injections of <i>AAV8-V5-Car8</i>^<i>WT</i> virus (1.5μl, 1.29E+14 genome copies /mL) and <i>AAV8-V5-Car8</i>^<i>MT</i> virus (1.5μl, 1.61E+14 genome copies /mL) were used in MT mice. (7A) The “up-down” method (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118273#sec002" target="_blank">Methods</a> for details) was used to measure mechanical responses by probing the plantar aspect of the hindpaw with von Frey filaments and determining the paw withdrawal threshold (grams). (7B) Thermal withdrawal response latencies were measured to radiant heat (70 units) applied to the plantar aspect of the hind paw (seconds). <i>AAV8-V5-Car8</i>^<i>WT</i> increased both basal mechanical thresholds (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118273#pone.0118273.g007" target="_blank">Fig. 7A</a>) and thermal latencies (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118273#pone.0118273.g007" target="_blank">Fig. 7B</a>), starting on day 7 after injection and lasting more than 28 days. Sciatic nerve injections of <i>AAV8-V5-Car8</i>^<i>MT</i> failed to affect nociception (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118273#pone.0118273.g007" target="_blank">Fig. 7A, B</a>) over the 28 d. (N = 8. * P<0.1; ** P<0.01; *** P< 0.001; Student <i>t</i>-test and two-way repeated measure ANOVA.)</p

DRG Car8 expression WT and MT animals.

<p>(<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118273#pone.0118273.g002" target="_blank">Fig. 2A-D</a>) Immunoreactivity for anti-Car8 (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118273#pone.0118273.g002" target="_blank">Fig. 2A, D</a>, green), anti-NF200 (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118273#pone.0118273.g002" target="_blank">Fig. 2B</a>, red), and anti-Car8 with anti-NF200 (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118273#pone.0118273.g002" target="_blank">Fig. 2C</a>) antibodies, respectively. The merged image (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118273#pone.0118273.g002" target="_blank">Fig. 2C</a>) is from A and B. Immunohistochemistry demonstrates Car8 is expressed in the WT DRG (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118273#pone.0118273.g002" target="_blank">Fig. 2A</a>) but little or none in the MT DRG (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118273#pone.0118273.g002" target="_blank">Fig. 2D</a>). Percentage of different size neurons of Car8-containing neurons (measuring neuronal somata with visible nuclei) in the WT DRG (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118273#pone.0118273.g002" target="_blank">Fig. 2E</a>). Small: <300 μm²; Medium: 300–700 μm²; Large: >700 μm². RT-PCR demonstrates <i>Car8</i> mRNA in DRG tissues (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118273#pone.0118273.g002" target="_blank">Fig. 2F</a>). No template (TEMP) was used as a negative control (CTRL). The cerebellum was used as positive control. N = 4. Scale bar: 100 μm.</p