Human iN neuronal model of schizophrenia displays dysregulation of chromogranin B and related neuropeptide transmitter signatures.

Schizophrenia (SZ) is a serious mental illness and neuropsychiatric brain disorder with behavioral symptoms that include hallucinations, delusions, disorganized behavior, and cognitive impairment. Regulation of such behaviors requires utilization of neurotransmitters released to mediate cell-cell communication which are essential to brain functions in health and disease. We hypothesized that SZ may involve dysregulation of neurotransmitters secreted from neurons. To gain an understanding of human SZ, induced neurons (iNs) were derived from SZ patients and healthy control subjects to investigate peptide neurotransmitters, known as neuropeptides, which represent the major class of transmitters. The iNs were subjected to depolarization by high KCl in the culture medium and the secreted neuropeptides were identified and quantitated by nano-LC-MS/MS tandem mass spectrometry. Several neuropeptides were identified from schizophrenia patient-derived neurons, including chromogranin B (CHGB), neurotensin, and natriuretic peptide. Focusing on the main secreted CHGB neuropeptides, results revealed differences in SZ iNs compared to control iN neurons. Lower numbers of distinct CHGB peptides were found in the SZ secretion media compared to controls. Mapping of the peptides to the CHGB precursor revealed peptides unique to either SZ or control, and peptides common to both conditions. Also, the iNs secreted neuropeptides under both KCl and basal (no KCl) conditions. These findings are consistent with reports that chromogranin B levels are reduced in the cerebrospinal fluid and specific brain regions of SZ patients. These findings suggest that iNs derived from SZ patients can model the decreased CHGB neuropeptides observed in human SZ.

The mechanisms of chromogranin B-regulated Cl- homeostasis.

Chloride is the most abundant inorganic anions in almost all cells and in human circulation systems. Its homeostasis is therefore important for systems physiology and normal cellular activities. This topic has been extensively studied with chloride loaders and extruders expressed in both cell surfaces and intracellular membranes. With the newly discovered, large-conductance, highly selective Cl- channel formed by membrane-bound chromogranin B (CHGB), which differs from all other known anion channels of conventional transmembrane topology, and is distributed in plasma membranes, endomembrane systems, endosomal, and endolysosomal compartments in cells expressing it, we will discuss the potential physiological importance of the CHGB channels to Cl- homeostasis, cellular excitability and volume control, and cation uptake or release at the cellular and subcellular levels. These considerations and CHGB's association with human diseases make the CHGB channel a possible druggable target for future molecular therapeutics.

Chromogranin B in blood serum of patients with neuroendocrine tumors.

Neuroendocrine tumors (NETs) are a heterogeneous group of neoplasms from cells of the diffuse neuroendocrine system. Chromogranin B (CgB) is an acidic protein of the granin family, which can be used to detect the tumours of neuroendocrine nature. Analysis of levels and evaluation of the diagnostic efficiency of CgB in the blood serum of patients with NETs of various localizations. Patients with NETs (n=121) without specific treatment were examined. In the study were presented next localizations: 74 - pancreas, 20 - stomach, 12 - large intestine, 15 - other localizations (lungs, mammary gland, prostate gland, NETs with unidentified primary). 54 practically healthy donors were examined as control group. The determination of CgB in blood serum was performed with ELISA method on BEP 2000 analyzer using a standardized test system Human Chromogranin B (USCN, China). CgB levels in common NET group (median 18.9 ng/mL) were statistically significantly higher than in the control group (8.8 ng/mL). The highest median was obtained in group of intestinal NETs (21.2 ng/ml), which exceeded the median of the control group by more than 2.4 times. According to ROC analysis in the common NET group relative to the control group, the area under the curve AUC was 0.88 (95% CI 0.83-0.929). According to cut-off level of CgB - 15.8 ng/ml, the diagnostic sensitivity was 69.4%, with a specificity of 96.3%. The highest diagnostic sensitivity was in the group of the intestinal NETs (75.0%) and pancreas (71.2%). The study showed the significance of CgB as a potential biochemical marker of NETs with various localizations, alternative to CgA.

Value of Immunohistochemical Expression of Apelin, Succinate Dehydrogenase B, Chromogranin B, Human Epidermal Growth Factor Receptor-2, Contactin 4, and Succinyl-CoA Synthetase Subunit Beta in Differentiating Metastatic From Non-Metastatic Pheochromocytoma and Paraganglioma.

Objective: We aimed to retrospectively collect pathologically identified pheochromocytoma and paraganglioma (PPGL) tumor tissues from our center and investigate the expression of apelin and succinyl-CoA synthetase subunit beta (SUCLG2), human epidermal growth factor receptor-2 (HER2 or ERBB-2), contactin 4 (CNTN4), chromogranin B (CHGB), and succinate dehydrogenase B (SDHB) in metastatic and non-metastatic PPGLs, for exploring their roles in the diagnosis of metastatic PPGLs. Methods: A total of 369 patients with pathologically and surgically confirmed PPGLs at Xiangya Hospital, Central South University, between June 2010 and June 2020 were retrospectively included. Sixty patients-12 patients with metastatic PPGLs and 48 patients with non-metastatic PPGLs-were selected through propensity score matching (1:4) to reduce the effect of PPGL type, sex, and age. We observed and quantified the expression of apelin, SDHB, CHGB, ERBB-2, CNTN4, and SUCLG2 in paraffin-embedded samples using immunohistochemical staining. Results: No significant differences were observed between the metastatic group and non-metastatic group with respect to the expression of CNTN4 and SUCLG2. The expression of apelin, SDHB, CHGB, and ERBB-2 was significantly different between the two groups. The expression of apelin, SDHB, and CHGB was significantly lower in the metastatic group than that in the non-metastatic group (P < 0.001). ERBB-2 expression was significantly higher in the metastatic group than in the non-metastatic group (P = 0.042). Kaplan-Meier analysis revealed that patients with negative expression of apelin, SDHB, and CHGB showed significantly lower metastasis-free survival than those with positive expression. Multivariate Cox analysis revealed that SDHB and CHGB levels were independently associated with metastasis-free survival. Conclusion: The expression levels of apelin, CHGB, SDHB, and ERBB-2 may be predictive biomarkers for the diagnosis of metastatic PPGLs. Patients with negative expression of apelin, CHGB, and SDHB should be subjected to frequent postoperative follow-up procedures.

The infarction zone rather than the noninfarcted remodeling zone overexpresses angiotensin II receptor type 1 and is the main source of ventricular atrial natriuretic peptide.

Chromogranin B and inositol 1,4,5-trisphosphate-associated calcium signaling leading to increased natriuretic peptide production has been described in cardiac hypertrophy. Here, we performed left anterior descending coronary artery ligation in rats as a model for systolic heart failure and examined protein and gene expression clusters in the infarcted and noninfarcted myocardium and moreover under treatment with metoprolol. We found that atrial natriuretic peptide gene transcription was significantly more elevated in the infarcted compared with the noninfarcted myocardium. Chromogranin B, which facilitates calcium release from internal stores through the inositol 1,4,5-trisphosphate receptor, was upregulated in both areas. Interestingly, angiotensin II receptor type 1 gene transcription was significantly upregulated in the infarcted and unchanged in the noninfarcted myocardium. Nuclear factor ĸappa B as a calcium-dependent transcription factor showed increased activity in the infarction zone. The ß-adrenergic axis does not seem to be involved, as metoprolol treatment did not have a significant impact on any of these results. We conclude that region-specific upregulation of angiotensin II receptor type 1 is a major factor for increased atrial natriuretic peptide production in the infarcted anterior wall. This effect is most likely achieved through inositol 1,4,5-trisphosphate-mediated cytosolic calcium increase and subsequent nuclear factor ĸappa B activation, which is a known transcription factor for natriuretic peptides.

Changes of Protein Phosphorylation Are Associated with Synaptic Functions during the Early Stage of Alzheimer's Disease.

Alzheimer's disease is an irreversible neurodegenerative disorder for which we have limited knowledge of the mechanisms underlying its pathogenesis, especially the molecular events that trigger the deterioration of neuronal functions in the early stage. Protein phosphorylation and dephosphorylation are highly dynamic and reversible post-translational modifications that control protein signaling and hence neuronal functions, aberrations of which are implicated in various neurodegenerative diseases including Alzheimer's disease. We conducted a quantitative phosphoproteomic analysis in the brains of APP/PS1 mice, an Aß-deposition transgenic mouse model, at 3 months old, the stage at which amyloid pathology just initiates. Compared to the wild-type mouse brains, we found that changes in serine phosphorylation were predominant in the APP/PS1 mouse brains, and that the occurrence of proline-directed phosphorylation was most common among the overrepresented phosphopeptides. Further analysis of the 167 phosphoproteins that were significantly up- or downregulated in APP/PS1 mouse brains revealed the enrichment of these proteins in synapse-related pathways. In particular, Western blot analysis validated the increased phosphorylation of chromogranin B, a protein enriched in large dense-core vesicles, in APP/PS1 mouse brains. These findings collectively suggest that changes in the phosphoprotein network may be associated with the deregulation of synaptic functions during the pathogenesis of Alzheimer's disease.

Chromogranin B regulates early-stage insulin granule trafficking from the Golgi in pancreatic islet ß-cells.

Chromogranin B (CgB, also known as CHGB) is abundantly expressed in dense core secretory granules of multiple endocrine tissues and has been suggested to regulate granule biogenesis in some cell types, including the pancreatic islet ß-cell, though the mechanisms are poorly understood. Here, we demonstrate a critical role for CgB in regulating secretory granule trafficking in the ß-cell. Loss of CgB impairs glucose-stimulated insulin secretion, impedes proinsulin processing to yield increased proinsulin content, and alters the density of insulin-containing granules. Using an in situ fluorescent pulse-chase strategy to track nascent proinsulin, we show that loss of CgB impairs Golgi budding of proinsulin-containing secretory granules, resulting in a substantial delay in trafficking of nascent granules to the plasma membrane with an overall decrease in total plasma membrane-associated granules. These studies demonstrate that CgB is necessary for efficient trafficking of secretory proteins into the budding granule, which impacts the availability of insulin-containing secretory granules for exocytic release.This article has an associated First Person interview with the first author of the paper.

Neuropeptidomics of the Rat Habenular Nuclei.

Conserved across vertebrates, the habenular nuclei are a pair of small symmetrical structures in the epithalamus. The nuclei functionally link the forebrain and midbrain by receiving input from and projecting to several brain regions. Each habenular nucleus comprises two major asymmetrical subnuclei, the medial and lateral habenula. These subnuclei are associated with different physiological processes and disorders, such as depression, nicotine addiction, and encoding aversive stimuli or omitting expected rewarding stimuli. Elucidating the functions of the habenular nuclei at the molecular level requires knowledge of their neuropeptide complement. In this work, three mass spectrometry (MS) techniques-liquid chromatography (LC) coupled to Orbitrap tandem MS (MS/MS), LC coupled to Fourier transform (FT)-ion cyclotron resonance (ICR) MS/MS, and matrix-assisted laser desorption/ionization (MALDI) FT-ICR MS-were used to uncover the neuropeptide profiles of the rodent medial and lateral habenula. With the assistance of tissue stabilization and bioinformatics, a total of 262 and 177 neuropeptides produced from 27 and 20 prohormones were detected and identified from the medial and lateral habenula regions, respectively. Among these neuropeptides, 136 were exclusively found in the medial habenula, and 51 were exclusively expressed in the lateral habenula. Additionally, novel sites of sulfation, a rare post-translational modification, on the secretogranin I prohormone are identified. The results demonstrate that these two small brain nuclei have a rich and differentiated peptide repertoire, with this information enabling a range of follow-up studies.

Identification of proteins that bind to the neuroprotective agent neoechinulin A.

Neoechinulin A is an indole alkaloid with several biological activities. We previously reported that this compound protects neuronal PC12 cells from cytotoxicity induced by the peroxynitrite generator 3-morpholinosydnonimine (SIN-1), but the target proteins and precise mechanism of action of neoechinulin A were unclear. Here, we employed a phage display screen to identify proteins that bind directly with neoechinulin A. Our findings identified two proteins, chromogranin B and glutaredoxin 3, as candidate target binding partners for the alkaloid. QCM analyses revealed that neoechinulin A displays high affinity for both chromogranin B and glutaredoxin 3. RNA interference-mediated depletion of chromogranin B decreased the sensitivity of PC12 cells against SIN-1. Our results suggested chromogranin B is a plausible target of neoechinulin A.

TFH-derived dopamine accelerates productive synapses in germinal centres.

Protective high-affinity antibody responses depend on competitive selection of B cells carrying somatically mutated B-cell receptors by follicular helper T (TFH) cells in germinal centres. The rapid T-B-cell interactions that occur during this process are reminiscent of neural synaptic transmission pathways. Here we show that a proportion of human TFH cells contain dense-core granules marked by chromogranin B, which are normally found in neuronal presynaptic terminals storing catecholamines such as dopamine. TFH cells produce high amounts of dopamine and release it upon cognate interaction with B cells. Dopamine causes rapid translocation of intracellular ICOSL (inducible T-cell co-stimulator ligand, also known as ICOSLG) to the B-cell surface, which enhances accumulation of CD40L and chromogranin B granules at the human TFH cell synapse and increases the synapse area. Mathematical modelling suggests that faster dopamine-induced T-B-cell interactions increase total germinal centre output and accelerate it by days. Delivery of neurotransmitters across the T-B-cell synapse may be advantageous in the face of infection.