Receptor Beta Research Articles

Single-cell transcriptomic analysis reveals dynamic activation of cellular signaling pathways regulating beige adipogenesis.

PDGFRA+ cells have been identified as adipocyte stem cells (ASCs) that differentiate into beige adipocytes in white adipose tissue (WAT) following thermogenic stimuli. To elucidate the molecular heterogeneity of ASCs, we conducted single-cell transcriptomic profiling of PDGFRA+ cells isolated from the inguinal WAT (iWAT) of mice treated with the beta3 adrenergic receptor agonist CL316243. Single-cell RNA-seq revealed nine major clusters, which were categorized into four groups: resting, proliferating, differentiating, and adipogenic factor-expressing cells (AFECs). Trajectory analysis revealed sequential activation of molecular pathways, including the Hedgehog and Notch signaling pathways, during beige adipogenesis. AFECs expressed Dpp4 and did not differentiate into adipocytes in culture or after transplantation. Furthermore, genetic lineage tracing studies indicated that DPP4+ cells did not differentiate into adipocytes in iWAT during CL316243-induced beige adipogenesis. However, high-fat diet feeding led to the recruitment of adipocytes from DPP4+ cells in iWAT. Overall, this study improved our understanding of the dynamic molecular basis of beige adipogenesis and the potential contribution of DPP4+ adipocyte lineages to the pathological expansion of WAT during diet-induced obesity.

Carvedilol suppresses coronary artery spasm in A-kinase anchoring protein 150 knockout mouse

Abstract Backgrounds A-kinase anchoring proteins (AKAPs) act as scaffold to regulate activation of protein kinases and subsequent downstream signaling. AKAP 150 interacts with protein kinase A and protein kinase C, and has a critical role in regulating L-type calcium channel activity. We previously reported enhanced phospholipase C activity as an important mechanism of coronary spastic angina (CSA), known as Prinzmetal’s angina. Coronary artery spasm was induced by intravenous ergometrine injection in mouse model of vascular smooth muscle cell (VSMC)-specific enhanced phospholipase C activity concomitantly with enhanced intracellular Ca2+ concentration ([Ca2+]i). Then, we hypothesized that coronary artery spasm would be inhibited by AKAP 150 knockout, however, coronary artery spasm was unexpectedly induced in AKAP 150 knockout (AKAP-KO) mice without affecting [Ca2+]i, suggesting that AKAP-KO mouse is an unique model of CSA independent of [Ca2+]i. Previous report showed that AKAP-KO mice have a high Ca2+/calmodulin-dependent protein kinase (CaMK) II activity, and carvedilol, a non-selective beta-adrenergic receptor antagonist, inhibits CaMK II activity. Aim We tested the hypothesis that carvedilol could prevent coronary artery spasm via inhibiting CaMK II activity, which is a key molecule to regulate VSMC contraction, in AKAP-KO mice. Methods and Results We pretreated 14-18-weeks-old AKAP-KO mice with intraperitoneal injection of carvedilol (2.5mg/kg), propranolol (20mg/kg), or vehicle (control). Coronary artery spasm, evaluated by ST-segment elevation on lead II of body surface electrocardiogram, was induced by intravenous ergometrine injection (30mg/kg) in all controls (5/5, 100%) and all propranolol-treated mice (5/5, 100%), whereas it tended to be inhibited in carvedilol-treated mice (3/6, 50%) (p=0.08 vs control). In isolated VSMCs obtained from the aorta of AKAO-KO mice, the changes in [Ca2+]i from baseline in response to acetylcholine (ACh, 100μM) did not differ among the groups of pretreatment with carvedilol (10μM), propranolol (10μM), or vehicle, indicating that coronary artery spasm in AKAP-KO mice was induced by Ca2+-independent mechanism. In response to ACh, the phosphorylation of CaMK II was tended to be decreased (p=0.07), and the phosphorylation of myosin light chain was decreased (p<0.05) in VSMCs treated with carvedilol compared with control. Conclusions Carvedilol may inhibit coronary artery spasm probably via inhibiting CaMK II activity in AKAP-KO mice. Our results may provide a new paradigm into the pathogenesis of CSA as a model of enhanced Ca2+ sensitivity and important clinical implications for the mechanism of calcium channel antagonist-refractory coronary artery spasm.

Evaluation of 18F FOL a PET tracer targeting folate receptor beta in a rat model of myocardial infarction

Abstract Background Expression of folate receptor-β (FR-β) on the activated macrophages is associated with inflammatory diseases. However, the role of FR-β in the immune response following myocardial infarction (MI) remains unknown. Purpose To investigate FR-β-targeted positron emission tomography (PET) tracer aluminium fluoride-18-labeled 1,4,7,-triazacylononane-1,4,7-triacetic acid conjugated folate ([18F]FOL) for imaging immune response in a rat model of MI. Methods Rats underwent PET scan from 20 to 40 minutes after injection of 50±6 MBq of [18F]FOL on 3 (n=6), 7 (n=21), 15 (n=9), and 90 (n=11) days after surgical ligation of the left coronary artery. As controls, rats were studied on 3 (n=6), 7 (n=14), 15 (n=5), and 90 (n=8) days after sham-operation. [18F]FDG PET was performed a day before [18F]FOL injection in order to localize myocardium and area of MI. After [18F]FOL PET, autoradiography, histology, and staining of CD68 were performed 3 (n=6), 7 (n=6), and 90 (n=11) days after MI in order to detect [18F]FOL uptake and activated macrophages in myocardial tissue sections. A subgroup of rats (n=15) underwent serial [18F]FOL PET 7 and 90 days after MI together with echocardiography in order to evaluate left ventricular function. Results The presence of MI was confirmed by histology in all rats after coronary ligation. PET images showed increased uptake of [18F]FOL in the area of MI. The uptake was significantly higher (p<0.001) in the area of MI than in the myocardium of sham-operated rats on day 3 (SUV 2.04±0.25 vs. 0.74±0.12), and remained increased on day 7 (SUV 1.39±0.33 vs. 0.68±0.12), day 15 (SUV 1.24±0.20 vs. 0.59±0.07), and day 90 (SUV 1.39±0.25 vs. 0.69±0.11). On day 3, pre-treatment with folate glucosamine (n=3) reduced the [18F]FOL signal by 50% in the area of MI indicating specific binding to FR-β. Autoradiography confirmed focally increased uptake of [18F]FOL in the area of MI. A positive correlation was found between uptake of [18F]FOL in PET images and areal percentage of CD68-positivity in the area of MI (r2=0.447, p<0.001). Uptake of [18F]FOL on day 7 after MI was associated with decline in left ventricular ejection fraction (R=-0.665, p<0.01) and increase in left ventricular systolic volume (R=0.561, p=0.02) between 7 and 90 days after MI. Conclusion Cardiac [18F]FOL PET detects increased macrophage-associated FR-β expression after MI in a rat model. FR-β expression peaks early and remains elevated up to 3 months in the infarcted area. Increased FR-β expression early after MI is associated with worsening of left ventricular systolic function late after MI.

Gamma irradiation green synthesis of (polyacrylamide/chitosan/silver nanoparticles) hydrogel nanocomposites and their using as antifungal against Candida albicans and anti-cancer modulator

Silver nanoparticles-loaded hydrogel nanocomposites are exploited for medicinal and pharmaceutical applications. Hydrogel nanocomposites were prepared from acrylamide (Am), chitosan (CS) and AgNO3 utilizing gamma rays. Diverse variables were applied in preparation of silver nanoparticles-laoded hydrogel nanocomposites of (PAm/CS)-AgNPs such as influence of radiation dose and influnece of CS concentration. Diverse techniques were utilized to characterize hydrogel nanocomposites; Fourier transform infrared spectroscopy (FTIR), thermal gravimetric analysis (TGA), X-ray diffraction (XRD), Transmission electron microscopy (TEM), energy dispersive X-ray (EDX) and scanning electron microscopy (SEM). Results confirmed formation of silver nanoparticles-loaded hydrogel nanocomposites of (PAm/CS)-AgNPs. Antifungal activity of (PAm/CS)-AgNPs hydrogel nanocomposites on viability of C. albicans was esitmated. Results displayed the efficient microbial inhibition activity of treatment against C. albicans compared to control. Furthermore, (PAm/CS)-AgNPs hydrogel nanocomposite against cervical cancer HeLa cell line was investigated. Cytotoxicity of (PAm/CS)-AgNPs hydrogel nanocomposites on prior cancer cell line empolyed to prohibition of cell growth assesssed by MTT test. HeLa cancer cell is treated by (PAm/CS)-AgNPs for 48 h exposed a potential apoptotic activity by noticeable up-regulation of p53 gene expression. Moreover, anticancer activity was investigated by down-regulation of platelet-based growth variable receptor beta (PDGFR-β), Bcl2, Cathepsine, and MMP-2 gene expression. antioxidant activity was investigated and results showed antioxidant activity of (PAm/CS) hydrogel and (PAm/CS)-AgNPs hydrogel nanocomposite are 87.8% and 62.9%, respectively.

High Levels of Uric Acid Upregulate Endothelin Receptors: The Role of MAPK Pathways in an in vitro Study

IntroductionUric acid (UA) is the end product of purine compounds' metabolism. There is overwhelming evidence linking hyperuricemia (high levels of UA) and cerebrovascular diseases, but the effect of high levels of UA on cerebral vessels is not fully understood. The aim of this research is to clarify how UA affect the endothelin (ET) receptor in rat cerebral arteries and the related mechanism.Material and methodsIn an in vitro setting, segments of rat cerebral arteries (n=12) were exposed to high levels of UA, either alone or in conjunction with MAPK pathway inhibitors. ET agonists were used to induce contractions that were then measured with a myograph. ET receptor expression was measured using RT-PCR (n=6), Western Blot (n=3), or immunohistochemistry (n=3) to quantify mRNA and protein levels.ResultsThe study revealed that high levels of UA notably increase ETA and ETB receptor-induced contractions and boost the expression of ET receptors in cerebral arteries when compared to that are fresh or cultured alone, suggesting that UA enhances ETA and ETB receptors. Additionally, the up-regulation of ETB receptors induced by UA was inhibited by the p38 inhibitor SB203580, the JNK inhibitor SP600125, and the ERK1/2 inhibitor U0126. SB203580 significantly blocked the increase in ETA receptor-mediated contractions induced by UA and the upregulation of ETA receptor. Neither SP600125 nor U0126 had such effect.ConclusionsHigh levels of UA stimulate the up-regulation of ET receptors in rat cerebral arteries in vitro through MAPK pathways. This study may offer novel perspectives on hyperuricemia-associated cerebrovascular diseases.

Topical application of beta-blockers accelerates epithelialization to mesh skin grafted full-thickness burn in sheep

Aim : Beta-adrenergic receptor blockers are conventionally used for the treatment of hypertension, tachycardia, and glaucoma. Research has shown that beta-blockers can accelerate wound epithelialization. In this study, we tested the efficacy of the beta-blocker timolol in an ovine model of grafted full-thickness burn wound healing, which closely mimics clinical scenarios. Methods : Six full-thickness burn wounds were created on the sheep’s posterior surface. Twenty-four hours later, eschars were excised and meshed skin was grafted (Day0). The wounds in the treatment group received topical application of timolol. Blood flow was measured using a blood perfusion imager. Cardiovascular hemodynamics and blood glucose levels were recorded daily. The epithelialization rate on Day 14 was determined by planimetric assay and analyzed by paired t -test. The days that the epithelialization rate exceeded 85%, 90%, and 95% were analyzed by survival analysis. To assess the potential influence of TGFβ, epithelial-mesenchymal transition (EMT), or myofibroblast activation (MFA) on wound healing, the RNA abundance of gene products related to these pathways was measured by reverse transcription and quantitative polymerase chain reaction (RT-qPCR). Results : The epithelialization rate on Day 14 was significantly higher in the treatment group. The days that the epithelialization rate exceeded 85%, 90%, and 95% were significantly shorter in the treatment group. There was no significant difference in wound blood flow or RNA abundance related to TGFβ, EMT, or MFA-related pathways among the groups at any time point. Conclusion : The results demonstrate that the beta-blocker timolol accelerates epithelialization of mesh skin grafted full-thickness burn wounds through a mechanism other than improving wound blood flow.

Neurovascular unit, neuroinflammation and neurodegeneration markers in brain disorders

Neurovascular unit (NVU) inflammation via activation of glial cells and neuronal damage plays a critical role in neurodegenerative diseases. Though the exact mechanism of disease pathogenesis is not understood, certain biomarkers provide valuable insight into the disease pathogenesis, severity, progression and therapeutic efficacy. These markers can be used to assess pathophysiological status of brain cells including neurons, astrocytes, microglia, oligodendrocytes, specialized microvascular endothelial cells, pericytes, NVU, and blood-brain barrier (BBB) disruption. Damage or derangements in tight junction (TJ), adherens junction (AdJ), and gap junction (GJ) components of the BBB lead to increased permeability and neuroinflammation in various brain disorders including neurodegenerative disorders. Thus, neuroinflammatory markers can be evaluated in blood, cerebrospinal fluid (CSF), or brain tissues to determine neurological disease severity, progression, and therapeutic responsiveness. Chronic inflammation is common in age-related neurodegenerative disorders including Alzheimer’s disease (AD), Parkinson’s disease (PD), and dementia. Neurotrauma/traumatic brain injury (TBI) also leads to acute and chronic neuroinflammatory responses. The expression of some markers may also be altered many years or even decades before the onset of neurodegenerative disorders. In this review, we discuss markers of neuroinflammation, and neurodegeneration associated with acute and chronic brain disorders, especially those associated with neurovascular pathologies. These biomarkers can be evaluated in CSF, or brain tissues. Neurofilament light (NfL), ubiquitin C-terminal hydrolase-L1 (UCHL1), glial fibrillary acidic protein (GFAP), Ionized calcium-binding adaptor molecule 1 (Iba-1), transmembrane protein 119 (TMEM119), aquaporin, endothelin-1, and platelet-derived growth factor receptor beta (PDGFRβ) are some important neuroinflammatory markers. Recent BBB-on-a-chip modeling offers promising potential for providing an in-depth understanding of brain disorders and neurotherapeutics. Integration of these markers in clinical practice could potentially enhance early diagnosis, monitor disease progression, and improve therapeutic outcomes.

In vitro and computational investigation of antioxidant and anticancer properties of Streptomyces coeruleofuscus SCJ extract on MDA-MB-468 triple-negative breast cancer cells

This study aimed to explore the antioxidant potential of the ethyl acetate extract of Streptomyces coeruleofuscus SCJ strain, along with its inhibitory effects on the triple-negative human breast carcinoma cell line (MDA-MB-468). The ethyl acetate extract’s total phenolic and flavonoid contents were quantified, and its antioxidant activity was investigated using DPPH (1,1-Diphenyl-2-picrylhydrazyl), ABTS (2,2’-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid), and FRAP (Ferric Reducing Antioxidant Power) assays. Furthermore, the cytotoxic effect of the organic extract from Streptomyces coeruleofuscus SCJ on MDA-MB-468 cancer cells was assessed via the crystal violet assay. In tandem, a thorough computational investigation was conducted to explore the pharmacokinetic properties of the identified components of the extract, utilizing the SwissADME and pKCSM web servers. Additionally, the molecular interactions between these components and Estrogen Receptor Beta, identified as a potential target, were probed through molecular docking studies. The results revealed that ethyl acetate extract of SCJ strain exhibited remarkable antioxidant activity, with 39.899 ± 1.56% and 35.798 ± 0.082% scavenging activities against DPPH and ABTS, respectively, at 1 mg/mL. The extract also displayed significant ferric reducing power, with a concentration of 1.087 ± 0.026 mg ascorbic acid equivalents per mg of dry extract. Furthermore, a strong positive correlation (p < 0.0001) between the antioxidant activity, the polyphenol and the flavonoid contents. Regarding anticancer activity, the SCJ strain extract demonstrated significant anticancer activity against TNBC MDA-MB-468 cancer cells, with an inhibition percentage of 62.76 ± 0.62%, 62.67 ± 0.93%, and 58.07 ± 4.82% at 25, 50, and 100 µg/mL of the extract, respectively. The HPLC-UV/vis analysis revealed nine phenolic compounds: gallic acid, sinapic acid, p-coumaric acid, cinnamic acid, trans-fereulic acid, syringic acid, chloroqenic acid, ellagic acid, epicatechin. Streptomyces coeruleofuscus SCJ showed promise for drug discovery, exhibiting antioxidant and anticancer effects.

Individual and combined effects of commercial glyphosate, atrazine and 2,4-D herbicides on the gerbil ventral prostate

Exposure to pesticides, individually or in a mixture, in drinking water is one of the main sources of human contamination, which causes adverse effects on the reproductive system. Our study aimed to investigate, the effects of a 90-day exposure to low concentrations of glyphosate (GLY), atrazine (ATZ), and 2,4-dichlorophenoxyacetic acid (2,4-D), in commercial formulations, on morphological, molecular, and hormonal parameters of the ventral prostate of gerbils (Meriones unguiculatus). The animals were exposed via drinking water to individual concentrations of GLY: 700 μg/L, ATZ: 3 μg/L, and 2,4-D: 70 μg/L, as well as to their mixture (MIX). Our findings showed an increase in prostatic complex relative weight in ATZ-exposed animals. Stereological and morphometric techniques indicated an increase in the percentage and thickness of muscular stroma, following an increase in the amount of collagen and reticular fibers in the MIX group. Histopathological analysis showed a decrease in the incidence of epithelial atrophy, subepithelial inflammation, and microacini in the MIX. On the other hand, ATZ-exposed animals showed an increase in hyperplasia and total prostatic intraepithelial neoplasia (PIN). The expression of caspase-3 decreased and estrogen receptor alpha (ERα) increased in the 2,4-D and MIX. Western blotting showed an increase in estrogen receptor beta (ERβ) expression in MIX-exposed animals. Testosterone levels decreased in animals from the GLY, ATZ and 2,4-D groups. Our findings provide evidence that individual or combined exposure to herbicides causes hormonal imbalance and morphological alterations, besides favoring the incidence of proliferative lesions in the prostate, predisposing the gland to more severe injuries.

The beta 2 adrenergic receptor cross-linked interactome identifies 14-3-3 proteins as regulating the availability of signaling-competent receptors.

The emerging picture of G protein-coupled receptor function suggests that the global signaling response is an integrated sum of a multitude of individual receptor responses, each regulated by their local protein environment. The beta 2 adrenergic receptor (B2AR) has long served as an example receptor in the development of this model. But the mechanism and the identity of the protein-protein interactions that govern the availability of receptors competent for signaling remains incompletely characterized. To address this question, we characterized the interactome of agonist-stimulated B2AR in HEK293 cells using FLAG co-immunoprecipitation coupled to SILAC labeling and mass spectrometry. Our B2AR cross-linked interactome identified 190 high-confidence proteins, including almost all known interacting proteins and six out of seven isoforms of the 14-3-3 family of scaffolding proteins. Inhibiting 14-3-3 proteins with the peptide difopein enhanced isoproterenol-stimulated adrenergic signaling via cAMP approximately three-fold, and increased both miniGs and arrestin recruitment to B2AR more than two fold each, without noticeably changing EC50 with respect to cAMP signaling or effector recruitment upon stimulation. Our results show that 14-3-3 proteins negatively regulate downstream signaling by inhibiting access of B2AR to effector proteins. We propose that 14-3-3 proteins maintain a dynamic pool of B2AR that has reduced signaling efficacy in response to acute agonist stimulation, limiting the amount of signaling-competent receptors at the plasma membrane. Significance Statement This study presents a new interactome of the agonist-stimulated beta 2 adrenergic receptor (B2AR), a paradigmatic GPCR that is both a model system for members of this class and an important signaling protein in respiratory, cardiovascular, and metabolic regulation. We identify 14-3-3 proteins as responsible for restricting B2AR access to signaling effectors and maintaining a receptor population that is insensitive to acute stimulation by agonists.

Sympathetic activity markers in patients with resistant hypertension with renal dysfunction

Aim. To study the informativeness of sympathetic activity markers and perirenal adipose tissue (PRAT) in assessing renal function decline in patients with resistant hypertension (RH).Material and methods. The study included 63 patients with RH of both sexes aged 40-80 years. The main group included 19 patients with chronic kidney disease (CKD). The control group consisted of 44 patients with RH without CKD. All patients were assessed for uric acid, blood lipid profile, beta-adrenergic reactivity of erythrocyte mem­branes, free metanephrine and normetanephrine in plasma and 24-hour urine. The renal function was assessed by the glomerular filtra­tion rate calculated using the CKD-EPI equation. Twenty-four hour blood pressure monitoring was performed on the brachial artery of one arm at intervals of 15 minutes during the day and 30 minutes at night. Kidney size and PRAT were calculated based on the magnetic reso­nance imaging.Results. In both groups of patients, a significant increase in blood normetanephrine and beta-adrenergic reactivity of erythrocyte mem­branes relative to the reference values was shown. Blood and 24-hour urine metanephrine were within the reference intervals. No differences were found between the sympathetic activity indices in the main and control groups. In the main group patients, the PRAT thickness was greater (p=0,013), and the kidney diameter was smaller (p=0,046) than in the control group patients.Conclusion. CKD in RH patients is not accompanied by additional significant changes in sympathetic regulation, assessed by the blood and 24-hour urine catecholamine levels, the degree of beta-adrenergic receptor desensitization and variability of blood pressure. PRAT increase according to magnetic resonance imaging in patients with RH is associated with a decrease in renal filtration function.

Interleukin-1 Beta rs16944 and rs1143634 and Interleukin-6 Receptor rs12083537 Single Nucleotide Polymorphisms as Potential Predictors of COVID-19 Severity.

Host genetic variation has been recognized as a key predictor of diverse clinical sequelae among severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected patients. Insights into the link between the Interleukin-6 receptor (IL-6R) and Interleukin-1 beta (IL-1β) genetic variation and severe coronavirus disease 2019 (COVID-19) are crucial for developing new predictors and therapeutic targets. We aimed to investigate the association of IL-6R rs12083537, IL-1β rs16944, and IL-1β rs1143634 SNPs with the severity of COVID-19. Our study was conducted on 300 COVID-19-negative individuals (control group) and 299 COVID-19-positive cases, classified into mild, moderate, and severe subgroups. Analyses of IL-1β (rs16944, rs1143634) and IL-6R (rs12083537) SNPs' genotypes were performed using qPCR genotyping assays. The IL-1β (rs16944) CC genotype and IL-6R (rs12083537) GG genotype were substantially related to COVID-19 severity, which was also associated with comorbidities and some laboratory parameters (p < 0.001). The IL-1β (rs1143634) TT genotype was found to be protective. Likewise, the IL-1β (rs16944) CC genotype was associated with increased mortality. IL-1β rs16944 and IL-6R rs12083537 SNPs are promising potential predictors of SARS-CoV-2 disease severity. Meanwhile, the rs1143634 SNP T allele was protective against severity and mortality risk.

Suppressing PDGFRβ Signaling Enhances Myocyte Fusion to Promote Skeletal Muscle Regeneration.

Muscle cell fusion is critical for forming and maintaining multinucleated myotubes during skeletal muscle development and regeneration. However, the molecular mechanisms directing cell-cell fusion are not fully understood. Here, we identify platelet-derived growth factor receptor beta (PDGFRβ) signaling as a key modulator of myocyte fusion in adult muscle cells. Our findings demonstrate that genetic deletion of Pdgfrβ enhances muscle regeneration and increases myofiber size, whereas PDGFRβ activation impairs muscle repair. Inhibition of PDGFRβ activity promotes myonuclear accretion in both mouse and human myotubes, whereas PDGFRβ activation stalls myotube development by preventing cell spreading to limit fusion potential. Transcriptomics analysis show that PDGFRβ signaling cooperates with TGFβ signaling to direct myocyte size and fusion. Mechanistically, PDGFRβ signaling requires STAT1 activation, and blocking STAT1 phosphorylation enhances myofiber repair and size during regeneration. Collectively, PDGFRβ signaling acts as a regenerative checkpoint and represents a potential clinical target to rapidly boost skeletal muscle repair.

Structure of endothelin ETB receptor–Gi complex in a conformation stabilized by unique NPxxL motif

Endothelin type B receptor (ETBR) plays a crucial role in regulating blood pressure and humoral homeostasis, making it an important therapeutic target for related diseases. ETBR activation by the endogenous peptide hormones endothelin (ET)−1–3 stimulates several signaling pathways, including Gs, Gi/o, Gq/11, G12/13, and β-arrestin. Although the conserved NPxxY motif in transmembrane helix 7 (TM7) is important during GPCR activation, ETBR possesses the lesser known NPxxL motif. In this study, we present the cryo-EM structure of the ETBR–Gi complex, complemented by MD simulations and functional studies. These investigations reveal an unusual movement of TM7 to the intracellular side during ETBR activation and the essential roles of the diverse NPxxL motif in stabilizing the active conformation of ETBR and organizing the assembly of the binding pocket for the α5 helix of Gi protein. These findings enhance our understanding of the interactions between GPCRs and G proteins, thereby advancing the development of therapeutic strategies.

Galectin-3 inhibition reduces fibrotic scarring and promotes functional recovery after spinal cord injury in mice

BackgroundIn the context of spinal cord injury (SCI), infiltrating macrophages assume prominence as the primary inflammatory cells within the lesion core, where the fibrotic scar is predominantly orchestrated by platelet-derived growth factor receptor beta (PDGFRβ+) fibroblasts. Galectin-3, a carbohydrate-binding protein of the lectin family, is notably expressed by infiltrating hematogenous macrophages and mediates cell-cell interactions. Although Galectin-3 has been shown to contribute to the endocytic internalization of PDGFRβ in vitro, its specific role in driving fibrotic scar formation after SCI has not been determined.MethodsWe employed a crush mid-thoracic (T10) SCI mouse model. Galectin-3 inhibition after SCI was achieved through intrathecal injection of the Galectin-3 inhibitor TD139 or in situ injection of lentivirus carrying Galectin-3-shRNA (Lv-shLgals3). A fibrosis-induced mice model was established by in situ injection of platelet-derived growth factor D (PDGFD) or recombinant Galectin-3 (rGalectin-3) into the uninjured spinal cord. Galectin-3 internalization experiments were conducted in PDGFRβ+ fibroblasts cocultured in conditioned medium in vitro.ResultsWe identified the spatial and temporal correlation between macrophage-derived Galectin-3 and PDGFRβ in fibroblasts from 3 to 56 days post-injury (dpi). Administration of TD139 via intrathecal injection or in situ injection of Lv-shLgals3 effectively mitigated fibrotic scar formation and extracellular matrix deposition within the injured spinal cord, leading to better neurological outcomes and function recovery after SCI. Furthermore, the fibrosis-inducing effects of exogenous PDGFD in the uninjured spinal cord could be blocked by TD139. In vitro experiments further demonstrated the ability of PDGFRβ+ fibroblasts to internalize Galectin-3, with Galectin-3 inhibition resulting in reduced PDGFRβ expression.ConclusionsOur finding underscores the pivotal role of macrophage-derived Galectin-3 in modulating the sustained internalized activation of PDGFRβ within fibroblasts, providing a novel mechanistic insight into fibrotic scarring post-SCI.

Exosomal let-7b-5p deriving from parietal epithelial cells attenuate renal fibrosis through suppression of TGFβR1 and ARID3a in obstructive kidney disease.

As renal progenitor cells, parietal epithelial cells (PECs) have demonstrated multilineage differentiation potential in response to kidney injury. However, the function of exosomes derived from PECs has not been extensively explored. Immunofluorescent staining of Claudin-1 was used to identify primary PECs isolated from mouse glomeruli. Transmission electron microscopy, nanoparticle tracking analysis, and western blotting were used to characterize the properties of PECs-derived exosomes (PEC-Exo). The therapeutic role of PEC-Exo in tubulointerstitial fibrosis was investigated in the unilateral ureteral obstruction (UUO) mouse model and TGF-β1-stimulated HK-2 cells. High-throughput miRNA sequencing was employed to profile PEC-Exo miRNAs. One of the most enriched miRNAs in PEC-Exo was knocked down by transfecting miRNA inhibitor, and then we investigated whether this candidate miRNA was involved in PEC-Exo-mediated tubular repair. The primary PECs expressed Claudin-1, PEC-Exo was homing to obstructed kidney, and TGF-β1 induced HK-2 cells. PEC-Exo significantly alleviated renal inflammation and ameliorated tubular fibrosis both invivo and invitro. Mechanistically, let-7b-5p, highly enriched in PEC-Exo, downregulated the protein levels of transforming growth factor beta receptor 1(TGFβR1) and AT-Rich Interaction Domain 3A(ARID3a) in tubular epithelial cells (TECs), leading to the inhibition of p21 and p27 to restoring cell cycle. Furthermore, administration of let-7b-5p agomir mitigated renal fibrosis invivo. Our findings demonstrated that PEC-derived exosomes significantly repressed the expression of TGFβR1 and ARID3a by delivering let-7b-5p, thereby alleviating renal fibrosis. This study provides novel insights into the role of PEC-Exo in the repair of kidney injury and new ideas for renal fibrosis intervention.

Estrogen Receptor Beta Agonist Influences Presynaptic NMDA Receptor Distribution in the Paraventricular Hypothalamic Nucleus Following Hypertension in a Mouse Model of Perimenopause.

Women become susceptible to hypertension as they transition to menopause (i.e., perimenopause); however, the underlying mechanisms are unclear. Animal studies using an accelerated ovarian failure (AOF) model of peri-menopause (peri-AOF) demonstrate that peri-AOF hypertension is associated with increased postsynaptic NMDA receptor plasticity in the paraventricular hypothalamic nucleus (PVN), a brain area critical for blood pressure regulation. However, recent evidence indicates that presynaptic NMDA receptors also play a role in neural plasticity. Here, using immuno-electron microscopy, we examine the influence of peri-AOF hypertension on the subcellular distribution of the essential NMDA GluN1 receptor subunit in PVN axon terminals in peri-AOF and in male mice. Hypertension was produced by 14-day slow-pressor angiotensin II (AngII) infusion. The involvement of estrogen signaling was investigated by co-administering an estrogen receptor beta (ERß) agonist. Although AngII induced hypertension in both peri-AOF and male mice, peri-AOF females showed higher cytoplasmic GluN1 levels. In peri-AOF females, activation of ERß blocked hypertension and increased plasmalemmal GluN1 in axon terminals. In contrast, stimulation of ERß did not inhibit hypertension or influence presynaptic GluN1 localization in males. These results indicate that sex-dependent recruitment of presynaptic NMDA receptors in the PVN is influenced by ERß signaling in mice during early ovarian failure.

Role of IL3RA in a Family with Lumbar Spinal Stenosis.

Lumbar spinal stenosis (LSS) is a degenerative spinal condition characterized by the narrowing of the spinal canal, resulting in low back pain (LBP) and limited leg mobility. Twin and family studies have suggested that genetics contributes to disease progression. However, the genetic causes of familial LSS remain unclear. We performed whole-exome and direct sequencing on seven female patients from a Han Chinese family with LBP, among whom four developed LSS. Based on our genetic findings, we performed gene knockdown studies in human chondrocytes to study possible pathological mechanisms underlying LSS. We found a novel nonsense mutation, c.417C > G (NM_002183, p.Y139X), in IL3RA, shared by all the LBP/LSS cases. Knockdown of IL3RA led to a reduction in the total collagen content of 81.6% in female chondrocytes and 21% in male chondrocytes. The expression of MMP-1, -3, and/or -10 significantly increased, with a more pronounced effect observed in females than in males. Furthermore, EsRb expression significantly decreased following IL3RA knockdown. Moreover, the knockdown of EsRb resulted in increased MMP-1 and -10 expression in chondrocytes from females. We speculate that IL3RA deficiency could lead to a reduction in collagen content and intervertebral disk (IVD) strength, particularly in females, thereby accelerating IVD degeneration and promoting LSS occurrence. Our results illustrate, for the first time, the association between IL3RA and estrogen receptor beta, highlighting their importance and impact on MMPs and collagen in degenerative spines in women.

Recurrent Visceral Leishmaniasis in a Case With Interleukin-12 Receptor Beta-1 Deficiency.

In this study, we present the case of a children who was followed up for recurrent visceral leishmaniasis and diagnosed with IL-12Rβ1 deficiency. A female patient who received Bacille Calmette-Guérin (BCG) vaccine 2 months after birth and developed visceral leishmaniasis at the age of 91 months was subsequently diagnosed with IL-12Rβ1 deficiency. The patient's diagnosis and treatment process were examined retrospectively. IL-12Rβ1 deficiency is an autosomal recessive disease characterized by susceptibility to recurrent and/or severe infections caused by weakly pathogenic mycobacteria and salmonella. Infections with other intramacrophagic organisms may also occur, although rarely. Based on this information, it is believed that the mutation in the IFN-γ/IL-12 axis in our patient predisposed her to recurrent Leishmania infections. This study adds to the limited literature on IL12RB1 deficiency as a cause of VL. Patients diagnosed with VL should be evaluated immunologically, as recurrent Leishmania infections may occur in those with IL-12Rβ1 defects.

Role of glucocorticoid receptor beta in the development of steroid resistance

Glucocorticoids (GCs) are the most commonly used anti-inflammatory drugs for the treatment of inflammatory diseases, despite the fact that the resistance to them is increasing due to various circumstances. GC insensitivity is an important clinical issue that is associated with life-threatening disease progression. Currently, there is no reliable biomarker that can be used to define steroid resistance. According to several studies, increased expression of the glucocorticoid receptor beta (GR-β) isoform is associated with steroid resistance. The purpose of this review is to analyse the correlation between GR-β expression and the occurrence of steroid therapy-resistant diseases and to determine its potential as a biomarker. This review summarizes studies that point out increased GR-β expression in steroid-resistant groups of patients compared to steroid-sensitive groups. Articles included in the review focus on asthma, nasal polyposis, ulcerative colitis, and other chronic inflammatory diseases in which resistance to steroid therapy is observed. GR-β levels have also been shown to be elevated in patients with allergic rhinitis, Crohn's disease and rheumatoid arthritis. In most studies, higher levels of GR-β were detected in peripheral blood mononuclear cells (PBMC) by reverse transcription polymerase chain reaction (RT-PCR). Increased expression of GR-β is mostly associated with steroid resistance. Thus, it can potentially be used as a biomarker of steroid resistance-related disorders.