BACKGROUND: Habitat transitions have considerable consequences in organism homeostasis, as they require the adjustment of several concurrent physiological compartments to maintain stability and adapt to a changing environment. Within the range of molecules with a crucial role in the regulation of different physiological processes, neuropeptides are key agents. Here, we examined the coding status of several neuropeptides and their receptors with pleiotropic activity in Cetacea.
RESULTS: Analysis of 202 mammalian genomes, including 41 species of Cetacea, exposed an intricate mutational landscape compatible with gene sequence modification and loss. Specifically for Cetacea, in the 12 genes analysed we have determined patterns of loss ranging from species-specific disruptive mutations (e.g. neuropeptide FF-amide peptide precursor; NPFF) to complete erosion of the gene across the cetacean stem lineage (e.g. somatostatin receptor 4; SSTR4).
CONCLUSIONS: Impairment of some of these neuromodulators may have contributed to the unique energetic metabolism, circadian rhythmicity and diving response displayed by this group of iconic mammals.

Hematological and oncological diseases are still among the leading causes of childhood mortality. Expression of growth hormone-releasing hormone (GHRH) and its receptors (GHRH-R) has been previously demonstrated in various human tumors, but very limited findings are available about the presence and potential function of GHRH-Rs in oncological and hematological disorders of children. In this study, we aimed to investigate the expression of mRNA for GHRH and splice variant 1 (SV) of GHRH-R in 15 pediatric hematological/oncological specimens by RT-PCR. The presence and binding characteristics of GHRH-R protein were also studied by Western blot and ligand competition assays. Of the fifteen specimens studied, eleven pediatric samples (73%) showed the expression of mRNA for GHRH. These eleven samples also expressed mRNA for GHRH receptor SV1. GHRH-R protein was found to be expressed in two benign tumor samples and five malignant tumors examined by Western blot. The presence of specific, high affinity binding sites on GHRH-R was demonstrated in all of the seven human pediatric solid tumor samples investigated. Our results show that the expression of GHRH and SV1 of GHRH-R in hemato-oncological diseases in children can pave the way for further investigation of GHRH-Rs as potential molecular targets for diagnosis and therapy.

Ursolic acid (UA), a pentacyclic triterpene, exhibits diverse pharmacological effects, including potential treatment for allergic diseases. It downregulates thymic stromal lymphopoietin (TSLP) and disrupts mast cell signaling pathways. However, the exact molecular mechanism by which UA interferes with mast cell action remains unclear. Therefore, the current study aimed to uncover molecular entities underlying the effect of UA on mast cells and its potential antipruritic effect, specifically investigating its modulation of key molecules such as TRPV4, PAR2, and MRGPRX2, which are involved in TSLP regulation and sensation. Calcium imaging experiments revealed that UA pretreatment significantly suppressed MRGPRX2 activation (and its mouse orthologue MrgprB2), a G protein-coupled receptor predominantly expressed in mast cells. Molecular docking predictions suggested potential interactions between UA and MRGPRX2/MrgprB2. UA pretreatment also reduced mast cell degranulation through MRGPRX2 and MrgprB2-dependent mechanisms. In a dry skin mouse model, UA administration decreased tryptase and TSLP production in the skin, and diminished TSLP response in the sensory neurons. While PAR2 and TRPV4 activation enhances TSLP production, UA did not inhibit their activity. Notably, UA attenuated compound 48/80-induced scratching behaviors in mice and suppressed spontaneous scratching in a dry skin model. The present study confirms the effective inhibition of UA on MRGPRX2/MrgprB2, leading to reduced mast cell degranulation and suppressed scratching behaviors. These findings highlight the potential of UA as an antipruritic agent for managing various allergy- or itch-related conditions.

Artemisia capillaris Thunb. (A. capillaris) is a well-known traditional Chinese herbal medicine with a wide range of pharmacological effects, such as soothing the liver and gallbladder, heat clearance, and detoxifying. Hence, its extract is commonly added to various traditional Chinese medicine formulas. Traditional Chinese medicine injection (TCMI) is a mature pharmaceutical dosage form developed using TCM theory combined with modern science and technology. Notably, allergic reactions, especially pseudo‑allergic reactions (PARs), greatly limited the use of these injections. Therefore, screening pseudo‑allergic components in A. capillaris extract is clinically significant. In the present study, we proposed a two-dimensional screening and identification system based on mas-related G protein-coupled receptor X2-HALO-tag/cell membrane chromatography (MrgX2-HALO-tag/CMC) high performance liquid chromatography mass spectrometry (HPLC-MS); seven potential active components were screened from 75 % ethanol extract of A. capillaris: NCA, CA, CCA, 1,3-diCQA, ICA-B, ICA-A, and ICA-C. The receptor-ligand interactions between these seven compounds and MrgX2 protein were analyzed using frontal analysis and molecular docking technology. Furthermore, a mast cell degranulation-related assay was used to assess the pseudo‑allergic activity of these compounds. The screened compounds can serve as ligands of MrgX2, and this study provides a research basis for pseudo‑allergic reactions caused by TCMIs containing A. capillaris.

Mas-related G protein-coupled receptor X2 (MrgprX2) plays a crucial role in anaphylactoid reactions and allergic diseases. Some antagonists with reasonable potency and selectivity have been reported. Cell membrane chromatography (CMC) is effective for discovering ligands. Protein-tag-based CMC models (e.g., SNAP tags and HALO tags) have enhanced performance but also increased nonspecific adsorption of small molecules. The Avi tag, a short peptide sequence, binds biotin specifically via BirA catalysis. Our study showed that 2-iminobiotin (IB) can be a BirA substrate, enabling the development of a new cell membrane stationary phase (CMSP) based on the chemical properties (modifying carboxyl silica gel and specifically labeling the Avi tag) of IB. First, we constructed the MrgprX2-Avi-tag HEK293T cell line. Next, we synthesized IB-modified silica gel (SiO2-IB) stepwise. Finally, we immobilized Avi-tagged MrgprX2 cell membranes on SiO2-IB under BirA catalysis. We characterized the developed CMSP and used it to establish a MrgprX2-Avi-tag/CMC-HPLC/MS two-dimensional screening platform, successfully screening vitexicarpin fromViticis Fructus extract via a 2D/CMC platform. In vitro and in vivo experiments confirmed that vitexicarpin targets the MrgprX2 receptor, demonstrating antiallergic effects. Our IB-Avi tag-based CMC approach effectively decreased nonspecific adsorption of the screening materials. The Avi-tag-based 2D/CMC platform is suitable for screening potential drug candidates.

The kidney and brain play critical roles in the regulation of blood pressure. Neuropeptide FF (NPFF), originally isolated from the bovine brain, has been suggested to contribute to the pathogenesis of hypertension. However, the roles of NPFF and its receptors, NPFF-R1 and NPFF-R2, in the regulation of blood pressure, via the kidney, are not known. In this study, we found that the transcripts and proteins of NPFF and its receptors, NPFF-R1 and NPFF-R2, were expressed in mouse and human renal proximal tubules (RPTs). In mouse RPT cells (RPTCs), NPFF, but not RF-amide-related peptide-2 (RFRP-2), decreased the forskolin-stimulated cAMP production in a concentration- and time-dependent manner. Furthermore, dopamine D1-like receptors colocalized and co-immunoprecipitated with NPFF-R1 and NPFF-R2 in human RPTCs. The increase in cAMP production in human RPTCs caused by fenoldopam, a D1-like receptor agonist, was attenuated by NPFF, indicating an antagonistic interaction between NPFF and D1-like receptors. The renal subcapsular infusion of NPFF in C57BL/6 mice decreased renal sodium excretion and increased blood pressure. The NPFF-mediated increase in blood pressure was prevented by RF-9, an antagonist of NPFF receptors. Taken together, our findings suggest that autocrine NPFF and its receptors in the kidney regulate blood pressure, but the mechanisms remain to be determined.

Genomes and transcriptomes from diverse organisms are providing a wealth of data to explore the evolution and origin of neuropeptides and their receptors in metazoans. While most neuropeptide-receptor systems have been extensively studied in vertebrates, there is still a considerable lack of understanding regarding their functions in invertebrates, an extraordinarily diverse group that account for the majority of animal species on Earth. Cephalochordates, commonly known as amphioxus or lancelets, serve as the evolutionary proxy of the chordate ancestor. Their key evolutionary position, bridging the invertebrate to vertebrate transition, has been explored to uncover the origin, evolution, and function of vertebrate neuropeptide systems. Amphioxus genomes exhibit a high degree of sequence and structural conservation with vertebrates, and sequence and functional homologues of several vertebrate neuropeptide families are present in cephalochordates. This review aims to provide a comprehensively overview of the recent findings on neuropeptides and their receptors in cephalochordates, highlighting their significance as a model for understanding the complex evolution of neuropeptide signaling in vertebrates.

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Mast cells are immune cells minimally present in normal tendon tissue. The increased abundance of mast cells in tendinopathy biopsies and at the sites of tendon injury suggests an unexplored role of this cell population in overuse tendon injuries. Mast cells are particularly present in tendon biopsies from patients with more chronic symptom duration and a history of intensive mechanical loading. This study, therefore, examined the cross talk between mast cells and human tendon cells in either static or mechanically active conditions in order to explore the potential mechanistic roles of mast cells in overuse tendon injuries. A coculture of isolated human tenocytes and mast cells (HMC-1) combined with Flexcell Tension System for cyclic stretching of tenocytes was used. Additionally, human tenocytes were exposed to agonists and antagonists of substance P (SP) receptors. Mast cell degranulation was assessed by measuring β-hexosaminidase activity. Transwell and cell adhesion assays were used to evaluate mast cell migration and binding to tendon extracellular matrix components (collagen and fibronectin), respectively. Gene expressions were analyzed using real time qRT-PCR. Our results indicate that mechanical stimulation of human tenocytes leads to release of SP which, in turn, activates mast cells through the Mas-related G-protein-coupled receptor X2 (MRGPRX2). The degranulation and migration of mast cells in response to MRGPRX2 activation subsequently cause human tenocytes to increase their expression of inflammatory factors, matrix proteins and matrix metalloproteinase enzymes. These observations may be important in understanding the mechanisms by which tendons become tendinopathic in response to repetitive mechanical stimulation.

Mas-related G protein-coupled receptor X2 (MrgprX2) is acknowledged as a mast cell-specific receptor, playing a crucial role in orchestrating anaphylactoid responses through mast cell degranulation. It holds promise as a target for regulating allergic and inflammatory diseases mediated by mast cells. Polygonum cuspidatum (PC) has shown notable anti-anaphylactoid effects, while its pharmacologically active components remain unclear. In this study, we successfully utilized MrgprX2 high-expressing cell membrane chromatography (CMC), in conjunction with liquid chromatography-mass spectrometry (LC-MS), to identify active anti-anaphylactoid components in PC. Our study pinpointed polydatin, resveratrol, and emodin-8-O-β-d-glucoside as potential anti-anaphylactoid compounds in PC. Their anti-anaphylactoid activities were evaluated through β-aminohexosidase and histamine release assays, demonstrating a concentration-dependent inhibition for both β-aminohexosidase and histamine release. This approach, integrating MrgprX2 high-expression CMC with LC-MS, proves effective in screening potential anti-anaphylactoid ingredients in natural herbal medicines. The findings from this study illuminated the anti-anaphylactoid properties of specific components in PC and provided an efficient method for the drug development of natural products.