Effective approaches to neuropsychiatric disorders require detailed understanding of the cellular composition and circuitry of the complex mammalian brain. Here, we present a paradigm for ...deconstructing the diversity of neurons defined by a specific neurotransmitter using a microfluidic dynamic array to simultaneously evaluate the expression of 96 genes in single neurons. With this approach, we successfully identified multiple molecularly distinct dopamine neuron subtypes and localized them in the adult mouse brain. To validate the anatomical and functional correlates of molecular diversity, we provide evidence that one Vip+ subtype, located in the periaqueductal region, has a discrete projection field within the extended amygdala. Another Aldh1a1+ subtype, located in the substantia nigra, is especially vulnerable in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) model of Parkinson’s disease. Overall, this rapid, cost-effective approach enables the identification and classification of multiple dopamine neuron subtypes, with distinct molecular, anatomical, and functional properties.
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•Single-cell gene profiling identified multiple midbrain dopamine neuron subtypes•A Vip-expressing subtype projects to specific nuclei within the extended amygdala•A subtype expressing Aldh1a1 is more vulnerable in a model of Parkinson’s disease
Poulin et al. used a single-cell gene-profiling approach to identify multiple molecularly distinct midbrain dopamine neuron subtypes. The findings have implications for targeted therapeutics of dopamine-related diseases understanding the differential vulnerability of dopamine neurons, generating stem-cell-derived dopamine neurons, and understanding the dopaminergic system by providing points of genetic access.
ABSTRACT
Background
Esophageal histology is critical for diagnosis and surveillance of disease activity in eosinophilic esophagitis (EoE). A validated noninvasive biomarker has not been identified. ...We aimed to determine the utility of blood and urine eosinophil‐associated proteins to diagnose EoE and predict esophageal eosinophilia.
Methods
Blood and urine were collected from children undergoing endoscopy with biopsy. Absolute eosinophil count (AEC), plasma eosinophil‐derived neurotoxin (EDN), eosinophil cationic protein (ECP), major basic protein‐1 (MBP‐1), galectin‐10 (CLC/GAL‐10), Eotaxin‐2 and Eotaxin‐3, and urine osteopontin (OPN) and matrix metalloproteinase‐9 (MMP‐9) were determined. Differences were assessed between EoE and control, and with treatment response. The capacity to predict EoE diagnosis and esophageal eosinophil counts was assessed.
Results
Of 183 specimens were collected from 56 EoE patients and 15 non‐EoE controls with symptoms of esophageal dysfunction; 33 EoE patients had paired pre‐ and post‐treatment specimens. Plasma (CLC/GAL‐10, ECP, EDN, Eotaxin‐3, MBP‐1) and urine (OPN) biomarkers were increased in EoE compared to control. A panel comprising CLC/GAL‐10, Eotaxin‐3, ECP, EDN, MBP‐1, and AEC was superior to AEC alone in distinguishing EoE from control. AEC, CLC/GAL‐10, ECP, and MBP‐1 were significantly decreased in patients with esophageal eosinophil counts <15/hpf in response to treatment. AEC, CLC/GAL‐10, ECP, EDN, OPN, and MBP‐1 each predicted esophageal eosinophil counts utilizing mixed models controlled for age, gender, treatment, and atopy; AEC combined with MBP‐1 best predicted the counts.
Conclusions
We identified novel panels of eosinophil‐associated proteins that along with AEC are superior to AEC alone in distinguishing EoE from controls and predicting esophageal eosinophil counts.
A panel of eosinophil‐associated plasma proteins along with AEC are superior to AEC alone in distinguishing EoE from non‐EoE. A panel of AEC and MBP‐1 predicted esophageal eosinophil counts superior to AEC alone longitudinally. Histologic responders to EoE treatment have a greater reduction in AEC, CLC/GAL‐10, ECP, and MBP‐1 than non‐responders. Abbreviations: EoE, eosinophilic esophagitis; AEC, absolute eosinophil count; CLC/GAL10, galectin‐10; ECP, eosinophil cationic protein; EDN, eosinophil‐derived neurotoxin; MBP‐1, major basic protein.
Recently, the interplay between autophagy and apoptosis has become an important factor in chemotherapy for cancer treatment. Inhibition of autophagy may be an effective strategy to improve the ...treatment of chemo-resistant cancer by consistent exposure to chemotherapeutic drugs. However, no reports have clearly elucidated the underlying mechanisms. Therefore, in this study, we assessed whether salinomycin, a promising anticancer drug, induces apoptosis and elucidated potential antitumor mechanisms in chemo-resistant prostate cancer cells. Cell viability assay, Western blot, annexin V/propidium iodide assay, acridine orange (AO) staining, caspase-3 activity assay, reactive oxygen species (ROS) production, and mitochondrial membrane potential were assayed. Our data showed that salinomycin alters the sensitivity of prostate cancer cells to autophagy. Pretreatment with 3-methyladenine (3-MA), an autophagy inhibitor, enhanced the salinomycin-induced apoptosis. Notably, salinomycin decreased phosphorylated of AKT and phosphorylated mammalian target of rapamycin (mTOR) in prostate cancer cells. Pretreatment with LY294002, an autophagy and PI3K inhibitor, enhanced the salinomycin-induced apoptosis by decreasing the AKT and mTOR activities and suppressing autophagy. However, pretreatment with PD98059 and SB203580, an extracellular signal-regulated kinases (ERK), and p38 inhibitors, suppressed the salinomycin-induced autophagy by reversing the upregulation of ERK and p38. In addition, pretreatment with
-acetyl-l-cysteine (NAC), an antioxidant, inhibited salinomycin-induced autophagy by suppressing ROS production. Our results suggested that salinomycin induces apoptosis, which was related to ROS-mediated autophagy through regulation of the PI3K/AKT/mTOR and ERK/p38 MAPK signaling pathways.
Highlights • This was the largest epidemiologic study to include objective determination of sleep in a multiethnic population of Black, Asian, Hispanic, and White participants. • Adverse sleep ...characteristics were observed in Black participants as compared with White participants. • Despite similar cardiovascular risk profiles, Asian participants reported more daytime sleepiness than did Whites.
Moutan Cortex, the root bark of Paeonia suffruticosa ANDREWS in Ranunculaceae, has widely demonstrated analgesic, anti-spasmodic, and anti-inflammatory effects in various cancer and immune cell ...lines. Oxidative stress is associated with development of several diseases, including liver disease. We prepared the water extract of Moutan Cortex (MCE) to investigate the cytoprotective activities and its mechanism. MCE protected hepatocytes from arachidonic acid (AA)+iron induced oxidative stress, as indicated by reactive oxygen species (ROS) production and cell viability analysis. MCE also suppressed mitochondrial dysfunction in AA+iron-treated human hepatocyte-derived hepatocellular carcinoma cell line, HepG2 cells. In addition, MCE treatment induces AMP-activated protein kinase (AMPK) and liver kinase B1 phosphorylation, which play a role in inhibition of oxidative stress induced cell death. Moreover, one of the MCE compounds, chlorogenic acid, exerted protective effects against oxidative stress and apoptosis. Taken together, MCE protected hepatocytes against AA+iron-induced oxidative stress through AMPK activation, and may be a candidate for the treatment of liver disease.
Progranulin (PGRN) is a secreted glycoprotein with multiple biological functions in early embryogenesis, anti‐inflammation, and neurodegeneration. A good model for the functional study of PGRN is the ...zebrafish with knockdown or knockout of grn, the gene encoding PGRN. Morpholino oligonucleotides (MOs) and zinc finger nucleases have been used to generate zebrafish grn models, yet they have shown inconsistent phenotypes due to either the neurotoxicity of the MOs or possible genetic compensation responses during gene editing. In this study, we generated stable grna (one of the major grn homologues of zebrafish) knockout zebrafish by using CRISPR/Cas9‐mediated genome editing. A grna sgRNA was designed to target the similar repeated sequence shared by exon 13, exon 15, and exon 19 in zebrafish. The F1 generation with the frameshift mutation of + 4 bp (the addition of 4 bp to exon15), which causes a premature termination, was obtained and subjected to morphological and behavioral evaluation. The grna knockout zebrafish showed neurodevelopmental defects, including spinal motor neurons with shorter axons, decreased sensory hair cells, thinning of the outer nuclear layer and thickening of the inner nuclear layer of the retina, decreased expression of rhodopsin in the cone cells, and motor behavior changes. Moreover, the phenotypes of grna knockout zebrafish could be rescued with the Tol2 system carrying the grna gene. The grna knockout zebrafish model generated in this study provides a useful tool to study PGRN function and has potential for high‐throughput drug screening for disease therapy.
Progranulin (PGRN) is a secreted, cysteine‐enriched glycoprotein with multiple important biological functions. So far, no reliable grn zebrafish models have been available to study PGRN function. In this study, we generated stable grna knockout zebrafish (grna being one of the major grn homologues of zebrafish) by using CRISPR/Cas9‐mediated genome editing. The grna knockout zebrafish showed neurodevelopmental defects, including spinal motor neurons with shorter axons, thinning of the outer nuclear layer and thickening of the inner nuclear layer of the retina, decreased expression of rhodopsin in the cone cells, and motor behavior changes. This grna knockout zebrafish model provides a reliable tool to study the function of PGRN. SpMN, spinal motor neuron.
Human bone marrow-derived mesenchymal stem cells (hBMSCs) present promising opportunities for therapeutic medicine. Carbon derivatives showed only marginal enhancement in stem cell differentiation ...toward bone formation. Here we report that red-light absorbing carbon nitride (C3N4) sheets lead to remarkable proliferation and osteogenic differentiation by runt-related transcription factor 2 (Runx2) activation, a key transcription factor associated with osteoblast differentiation. Accordingly, highly effective hBMSCs-driven mice bone regeneration under red light is achieved (91% recovery after 4 weeks compared to 36% recovery in the standard control group in phosphate-buffered saline without red light). This fast bone regeneration is attributed to the deep penetration strength of red light into cellular membranes via tissue and the resulting efficient cell stimulation by enhanced photocurrent upon two-photon excitation of C3N4 sheets near cells. Given that the photoinduced charge transfer can increase cytosolic Ca2+ accumulation, this increase would promote nucleotide synthesis and cellular proliferation/differentiation. The cell stimulation enhances hBMSC differentiation toward bone formation, demonstrating the therapeutic potential of near-infrared two-photon absorption of C3N4 sheets in bone regeneration and fracture healing.
Cow's milk protein (CMP) is the most common trigger of inflammation in children and adults with eosinophilic esophagitis (EoE). We sought to assess the clinical, endoscopic, and histologic efficacy ...of dietary elimination of all CMP-containing foods in EoE.
We performed a prospective observational study in children with EoE treated with the 1-food elimination diet (1FED), excluding all CMP. Children and their caretakers were educated by a registered dietitian regarding dietary elimination of all CMP-containing foods, with substitutions to meet nutritional needs for optimal growth and development, and daily meal planning. Upper endoscopy with biopsies was performed after 8 to 12 weeks of treatment. The primary end point was histologic remission, defined as fewer than 15 eosinophils per high-power field. Secondary end points were symptomatic, endoscopic, and quality-of-life (QOL) improvements.
Forty-one children (76% male; ages, 9 ± 4 years; 88% white) underwent 1FED education and post-treatment endoscopy with biopsies. Histologic remission occurred in 21 (51%) children, with a decrease in peak eosinophils per high-power field from a median of 50 (interquartile range, 35-70) to a median of 1 (interquartile range, 0-6; P < .0001). Endoscopic abnormalities improved in 24 (59%) patients, while symptoms improved in 25 (61%). Improved symptoms included chest pain, dysphagia, and pocketing/spitting out food. Parents perceived worse QOL, while children perceived improved QOL with the 1FED.
One-food elimination of CMP-containing foods from the diet induced histologic remission in more than 50% of children with EoE and led to significant improvement in symptoms and endoscopic abnormalities. The ease of implementation and adherence supports the 1FED as first-line dietary treatment.
Abstract
Background
Kawasaki disease (KD) is the leading cause of childhood acquired heart disease in developed nations and can result in coronary artery aneurysms and death. Clinical and ...epidemiologic features implicate an infectious cause but specific antigenic targets of the disease are unknown. Peripheral blood plasmablasts are normally highly clonally diverse but the antibodies they encode are approximately 70% antigen-specific 1–2 weeks after infection.
Methods
We isolated single peripheral blood plasmablasts from children with KD 1–3 weeks after onset and prepared 60 monoclonal antibodies (mAbs). We used the mAbs to identify their target antigens and assessed serologic response among KD patients and controls to specific antigen.
Results
Thirty-two mAbs from 9 of 11 patients recognize antigen within intracytoplasmic inclusion bodies in ciliated bronchial epithelial cells of fatal cases. Five of these mAbs, from 3 patients with coronary aneurysms, recognize a specific peptide, which blocks binding to inclusion bodies. Sera from 5/8 KD patients day ≥ 8 after illness onset, compared with 0/17 infant controls (P < .01), recognized the KD peptide antigen.
Conclusions
These results identify a protein epitope targeted by the antibody response to KD and provide a means to elucidate the pathogenesis of this important worldwide pediatric problem.
We prepared monoclonal antibodies from plasmablasts from children with Kawasaki disease (KD). The antibodies recognize inclusion bodies in fatal KD tissues containing a specific protein epitope. KD sera from week 2 of illness but not infant control sera recognize the epitope.
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