Mucus produced by goblet cells in the gastrointestinal tract forms a biological barrier that protects the intestine from invasion by commensals and pathogens. However, the host-derived regulatory ...network that controls mucus secretion and thereby changes gut microbiota has not been well studied. Here, we identify that Forkhead box protein O1 (Foxo1) regulates mucus secretion by goblet cells and determines intestinal homeostasis. Loss of Foxo1 in intestinal epithelial cells (IECs) results in defects in goblet cell autophagy and mucus secretion, leading to an impaired gut microenvironment and dysbiosis. Subsequently, due to changes in microbiota and disruption in microbiome metabolites of short-chain fatty acids, Foxo1 deficiency results in altered organization of tight junction proteins and enhanced susceptibility to intestinal inflammation. Our study demonstrates that Foxo1 is crucial for IECs to establish commensalism and maintain intestinal barrier integrity by regulating goblet cell function.
The transient receptor potential vanilloid 3 channel (TRPV3) is abundantly expressed in epidermal keratinocytes and has important roles in sensory biology and skin health. Mg2+ deficiency causes skin ...disorders under certain pathological conditions such as type 2 diabetes mellitus. In this study, we investigated the effect of Mg2+ on TRPV3 in primary epidermal keratinocytes. Extracellular Mg2+ (Mg2+o) inhibited TRPV3-mediated membrane current and calcium influx. TRPV3 activation induced a calcium signaling pathway culminating in activation of the cAMP response element binding. TRPV3 inhibition by Mg2+o, the TRPV3 blocker ruthenium red, or TRPV3 siRNA suppressed this response. In TRPV3-expressing Chinese hamster ovary cells, both extracellular and intracellular Mg2+ inhibited TRPV3 single-channel conductance, but not open probability. Neutralization of an aspartic acid residue (D641) in the extracellular pore loop or two acidic residues (E679, E682) in the inner pore region significantly attenuated the inhibitory effect of extracellular or intracellular Mg2+ on TRPV3-mediated signaling, respectively. Our findings suggest that epidermal TRPV3 is tonically inhibited by both extracellular and intracellular Mg2+, which act on both sides of the channel pore loop. Mg2+ deficiency may promote the function of TRPV3 and contribute to the pathogenesis of skin diseases.
To the Editor: Chronic skin inflammation is considered the most prominent feature for clinical diagnosis of psoriasis, a long-lasting autoimmune disease characterized by patches of red, itchy, and ...scaly skin. Besides skin inflammation, up to 84% of patients with psoriasis suffer from chronic itch, which significantly impairs quality of life.1 Although recent exciting studies have identified a positive correlation between the intensity of psoriatic itch and the expression levels of nerve growth factor, neuropeptides, and many cytokines,2 the molecular and cellular mechanisms underlying psoriatic itch are not fully understood. Because mast cell-derived histamine is one of the best studied pruritogens, especially in allergic itch, and TRPV1 is the major downstream mediator of histaminergic itch, we asked whether mast cells are involved in hBD2-elicited itch by measuring hBD2-induced scratching in mast cell-deficient KitW-sh “sash” mice. To our surprise, hBD2 induced comparable scratching responses among wild-type, ccr2−/−, and ccr6−/− mice (Fig 2, A and B), suggesting that neither CCR2 nor CCR6 mediates hBD2-induced itch. Because Toll-like receptor 4 (TLR4) mediates murine beta-defensin 2-induced activation of dendritic cells and murine beta-defensin 2 and hBD2 share structural and functional similarities,9 we tested hBD2-induced scratching in Tlr4−/− mice. Consistent with behavioral testing, 10 μM hBD2 elicited a robust Ca2+i response in 4.8% of the skin-resident cells examined (Fig 2, E) and the Ca2+i response was completely absent from the skin-resident cells isolated from the Tlr4−/− and LysMcre; Tlr4f/f mice or Tlr4−/− DRG neurons (Fig 2, F-H). ...hBD2 also activated human skin-resident myeloid cells, which was nearly abolished by a selective TLR4 antagonist LPS-RS (see Fig E4 in this article's Online Repository at www.jacionline.org).
Retinoids are structurally related derivatives of vitamin A and are required for normal vision as well as cell proliferation and differentiation. Clinically, retinoids are effective in treating many ...skin disorders and cancers. Application of retinoids evokes substantial irritating side effects, including pain and inflammation; however, the precise mechanisms accounting for the sensory hypersensitivity are not understood. Here we show that both naturally occurring and synthetic retinoids activate recombinant or native transient receptor potential channel vanilloid subtype 1 (TRPV1), an irritant receptor for capsaicin, the pungent ingredient of chili peppers. In vivo, retinoids produced pain-related behaviors that were either eliminated or significantly reduced by genetic or pharmacological inhibition of TRPV1 function. These findings identify TRPV1 as an ionotropic receptor for retinoids and provide cellular and molecular insights into retinoid-evoked hypersensitivity. These findings also suggest that selective TRPV1 antagonists are potential therapeutic drugs for treating retinoid-induced sensory hypersensitivity.
How the enteric nervous system determines the pacing and propagation direction of neurogenic contractions along the colon remains largely unknown. We used a chemogenetic strategy to ablate enteric ...neurons expressing calretinin (CAL). Mice expressing human diphtheria toxin receptor (DTR) in CAL neurons were generated by crossing
mice with
-dependent
mice. Immunohistochemical analysis revealed treatment with diphtheria toxin incurred a 42% reduction in counts of Hu-expressing colonic myenteric neurons (
= 0.036), and 57% loss of CAL neurons (comprising ∼25% of all Hu neurons;
= 0.004) compared to control. As proportions of Hu-expressing neurons, CAL neurons that contained nitric oxide synthase (NOS) were relatively spared (control: 15 ± 2%, CAL-DTR: 13 ± 1%;
= 0.145), while calretinin neurons lacking NOS were significantly reduced (control: 26 ± 2%, CAL-DTR: 18 ± 5%;
= 0.010). Colonic length and pellet sizes were significantly reduced without overt inflammation or changes in ganglionic density. Interestingly, colonic motor complexes (CMCs) persisted with increased frequency (mid-colon interval 111 ± 19 vs. 189 ± 24 s, CAL-DTR vs. control, respectively,
< 0.001), decreased contraction size (mid-colon AUC 26 ± 24 vs. 59 ± 13 gram/seconds, CAL-DTR vs. control, respectively,
< 0.001), and lacked preferential anterograde migration (
< 0.001). The functional effects of modest calretinin neuron ablation, particularly increased neurogenic motor activity frequencies, differ from models that incur general enteric neuron loss, and suggest calretinin neurons may contribute to pacing, force, and polarity of CMCs in the large bowel.
The molecular mechanisms of chronic pain are poorly understood and effective mechanism-based treatments are lacking. Here, we report that mice lacking adenosine deaminase (ADA), an enzyme necessary ...for the breakdown of adenosine, displayed unexpected chronic mechanical and thermal hypersensitivity due to sustained elevated circulating adenosine. Extending from Ada−/− mice, we further discovered that prolonged elevated adenosine contributed to chronic pain behaviors in two additional independent animal models: sickle cell disease mice, a model of severe pain with limited treatment, and complete Freund’s adjuvant paw-injected mice, a well-accepted inflammatory model of chronic pain. Mechanistically, we revealed that activation of adenosine A2B receptors on myeloid cells caused nociceptor hyperexcitability and promoted chronic pain via soluble IL-6 receptor trans-signaling, and our findings determined that prolonged accumulated circulating adenosine contributes to chronic pain by promoting immune-neuronal interaction and revealed multiple therapeutic targets.
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•Adenosine (Ado) deaminase deficient mice displayed chronic pain•Excess Ado signaling via ADORA2B promotes chronic pain in three distinct models•ADORA2B activation induces immunoneuronal interaction and promotes chronic pain•IL-6 trans-signaling underlies ADORA2B-induced prolonged hypersensitive nociception
Hu et al. show that prolonged increase in plasma adenosine activates ADORA2B on myeloid cells, leading to an increase in circulating IL-6 and sIL-6R. The IL-6/sIL-6R complex trans-activates gp130 on DRG cells, leading to STAT3 phosphorylation and induction of neuronal TRPV1 expression that results in increased sensitivity and chronic pain.
H2S, a gaseous signalling molecule, relaxes blood vessels partly through activation of ATP-sensitive K+ channels. It is however unclear whether H2S or its donors could affect other ion transporting ...proteins. The present study examined the hypothesis that NaHS, a H2S donor inhibits voltage-sensitive Ca2+ channels and thus relaxes vascular smooth muscle cells (VSMC) in the cerebral arteries. NaHS dilated cerebral arteries from Sprague–Dawley rats with the same potency against pre-contraction by 5-HT and 60mmol/L KCl, which were unaffected by several K+ channel blockers, NG-nitro-l-arginine methyl ester or indomethacin, as assessed in wire myograph under an isometric condition. Likewise, NaHS also dilated cerebral arteries against myogenic constriction in pressurized myograph under an isobaric condition. NaHS concentration-dependently inhibited CaCl2-induced contraction in Ca2+-free, 60mM K+-containing Krebs solution. Patch clamp recordings showed that NaHS reduced the amplitude of l-type Ca2+ currents in single myocytes isolated enzymatically from the cerebral artery. Calcium fluorescent imaging using fluo-4 showed a reduced Ca2+i in 60mmol/L KCl-stimulated rat cerebral arteries in response to NaHS. H2S precursor l-cysteine-induced relaxation in cerebral arteries was inhibited by cystathionine γ–lyase (CSE) inhibitor dl-propargylglycine. CSE was expressed in cerebral arteries. In summary, NaHS dilates rat cerebral arteries by reducing l-type Ca2+ currents and suppressing Ca2+i of arterial myocyte, indicating that NaHS relaxes cerebral arteries primarily through inhibiting Ca2+ influx via Ca2+ channels
β Amyloid protein (Aβ) and acetylcholinesterase (AChE) have been shown to be closely implicated in the pathogenesis of Alzheimer's disease. In the current study, we investigated the effects of ...bis(7)‐tacrine, a novel dimeric AChE inhibitor, on Aβ‐induced neurotoxicity in primary cortical neurons. Bis(7)‐tacrine, but not other AChE inhibitors, elicited a marked reduction of both fibrillar and soluble oligomeric forms of Aβ‐induced apoptosis as evidenced by chromatin condensation and DNA specific fragmentation. Both nicotinic and muscarinic receptor antagonists failed to block the effects of bis(7)‐tacrine. Instead, nimodipine, a blocker of L‐type voltage‐dependent Ca2+ channels (VDCCs), attenuated Aβ neurotoxicity, whereas N‐, P/Q‐ or R‐type VDCCs blockers and ionotropic glutamate receptor antagonists did not. Fluorescence Ca2+ imaging assay revealed that, similar to nimodipine, bis(7)‐tacrine reversed Aβ‐triggered intracellular Ca2+ increase, which was mainly contributed by the extracellular Ca2+ instead of endoplasmic reticulum and mitochondria Ca2+. Concurrently, using whole cell patch‐clamping technique, it was found that bis(7)‐tacrine significantly reduced the augmentation of high voltage‐activated inward calcium currents induced by Aβ. These results suggest that bis(7)‐tacrine attenuates Aβ‐induced neuronal apoptosis by regulating L‐type VDCCs, offers a novel modality as to how the agent exerts neuroprotective effects.
Bis(12)-hupyridone (B12H), derived from the Chinese medicinal component huperzine A, was originally designed as a novel acetylcholinesterase (AChE) inhibitor. In this paper, we report that B12H (24-h ...pretreatment) effectively blocked glutamate-induced neuronal excitotoxicity in cerebellar granule neurons (CGNs). However, the huge discrepancy between the EC50 value and IC50 value of B12H, to protect against neuronal toxicity (0.09μM) and to block the NMDA receptor (21.8μM) respectively, suggests that the neuroprotection of B12H might be not primarily due to the blockade of the NMDA receptor. Pretreatment by specific antagonists of alpha7-nicotinic acetylcholine receptor (α7nAChR), but not muscarinic acetylcholine receptor (mAChR) or α4β2nAChR, decreased the neuroprotection of B12H. The neuroprotection of B12H could also be abolished by the pretreatment of specific PI3-K inhibitors. Furthermore, B12H restored the suppressed activation of the Akt pathway caused by glutamate as evidenced by the decreased expressions of pSer473-Akt and pSer9-GSK3β. All these results suggest that B12H substantially protected CGNs against glutamate-induced neuronal excitotoxicity via activating α7nAChR/PI3-K/Akt cascade.
Oxidative stress-induced apoptosis plays a critical role in the pathogenesis of various neurodegenerative disorders. In this study, the neuroprotective properties of bis(12)-hupyridone (B12H), a ...novel dimeric acetylcholinesterase (AChE) inhibitor modified from a naturally occurring monomeric analogue, huperzine A, on H2O2-induced neurotoxicity were investigated in cerebellar granule neurons (CGNs). Exposure of CGNs to H2O2 resulted in apoptosis which could be attenuated by the pre-treatment of B12H (0.3–5nM) in a concentration-dependent manner. Moreover, tacrine and neostigmine failed to prevent neurotoxicity, indicating that the neuroprotection of B12H might not be due to its inhibitory property of AChE enzymatic activity. Increased activation of extracellular signal-regulated kinase (ERK) and decreased activation of glycogen synthase kinase (GSK) 3β were observed after H2O2 exposure, and B12H reversed the altered activation of GSK3β, but not that of ERK. Furthermore, using vascular endothelial growth factor (VEGF), phospho-VEGF receptor-2 (VEGFR-2) antibody, a specific VEGFR-2 inhibitor (PTK787/ZK222584) and specific phosphoinositide 3-kinase inhibitors (LY294002 and wortmannin), it was found that VEGF prevented H2O2-induced neuronal loss from activating the VEGF/VEGFR-2 system and that the observed B12H neuroprotective effects might share the same signaling pathway. These findings strongly suggest that B12H prevents H2O2-induced neuronal apoptosis independent of inhibiting AChE, but through regulating VEGFR-2/Akt/GSK3β signaling pathway.
►Bis(12)-hupyridone prevents H2O2-induced apoptosis in cerebellar granule neurons. ►VEGF also prevents H2O2-induced apoptosis in cerebellar granule neurons. ►Bis(12)-hupyridone reverses the VEGFR-2/Akt/GSK3β signaling pathways reduced by H2O2.
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