Lead inhibits microglial cell migration via suppression of store-operated calcium entry.

Lead (Pb) is a non-biodegradable environmental pollutant that can lead to neurotoxicity by inducing neuroinflammation. Microglial activation plays a key role in neuroinflammation, and microglial migration is one of its main features. However, whether Pb affects microglial migration has not yet been elucidated. Herein, the effect of Pb on microglial migration was investigated using BV-2 microglial cells and primary microglial cells. The results showed that cell activation markers (TNF-α and CD206) in BV-2 cells were increased after Pb treatment. The migration ability of microglia was inhibited by Pb. Both store-operated calcium entry (SOCE) and the Ca2+ release-activated Ca2+ (CRAC) current were downregulated by microglia treatment with Pb in a dose-dependent manner. However, there was no statistical difference in the protein levels of stromal interaction molecule (STIM) 1, STIM2, or Ca2+ release-activated Ca2+ channel protein (Orai) 1 in microglia. The external Ca2+ influx and cell migration ability were restored to a certain extent after overexpression of either STIM1 or its CRAC activation domain in microglia. These results indicated that Pb inhibits microglial migration by downregulation of SOCE and impairment of the function of STIM1.

Specific Inhibition of Orai1-mediated Calcium Signalling Resolves Inflammation and Clears Bacteria in an Acute Respiratory Distress Syndrome Model.

Rationale: Acute respiratory distress syndrome (ARDS) has an unacceptably high mortality rate (35%) and is without effective therapy. Orai1 is a Ca2+ channel involved in store-operated Ca2+ entry (SOCE), a process that exquisitely regulates inflammation. Orai1 is considered a druggable target, but no Orai1-specific inhibitors exist to date. Objectives: To evaluate whether ELD607, a first-in-class Orai1 antagonist, can treat ARDS caused by bacterial pneumonia in preclinical models. Methods: ELD607 pharmacology was evaluated in HEK293T cells and freshly isolated immune cells from patients with ARDS. A murine acute lung injury model caused by bacterial pneumonia was then used: mice were infected with Pseudomonas aeruginosa, Staphylococcus aureus, methicillin-resistant S. aureus, or multidrug-resistant P. aeruginosa and then treated with ELD607 intranasally. Measurements and Main Results: ELD607 specifically inhibited SOCE in HEK293T cells with a half-maximal inhibitory concentration of 9 nM. ELD607 was stable in ARDS airway secretions and inhibited SOCE in ARDS immune cells. In vivo, inhaled ELD607 significantly reduced neutrophilia and improved survival. Surprisingly, Orai1 inhibition by ELD607 caused a significant reduction in lung bacteria, including methicillin-resistant S. aureus. ELD607 worked as an immunomodulator that reduced cytokine levels, reduced neutrophilia, and promoted macrophage-mediated resolution of inflammation and clearance of bacteria. Indeed, when alveolar macrophages were depleted with inhaled clodronate, ELD607 was no longer able to resolve inflammation or clear bacteria. Conclusions: These data indicate that specific Orai1 inhibition by ELD607 may be a novel approach to reduce multiorgan inflammation and treat antibiotic-resistant bacteria.

Icaritin inhibits oxidative stress in murine astrocytes by binding to Orai1 to block store-operated calcium channel.

Previous study has shown that icaritin (ICT) has meaningful protective effect on cerebral ischemic stroke, and this study aimed to investigate its mechanism from the aspect of protecting astrocytes from oxidative stress. Murine primary astrocytes were pretreated by ICT and exposed to H2 O2 to induce oxidative stress. The results indicated that ICT inhibited H2 O2 -induced astrocytes apoptosis, decreased Bax and cleaved caspase-3, and increased Bcl-2. In addition, ICT inhibited H2 O2 -induced oxidative stress, increased mitochondrial membrane potential (ΔΨm ), and maintained mitochondrial morphology. ICT decreased the synthesis of malondialdehyde and increased the activity of glutathione peroxidase, catalase, and superoxide dismutase. Moreover, ICT suppressed the transient and resting intracellular Ca2+ overload. Further investigation revealed that ICT could target the combination with Orai1 to block store-operated calcium channel induced by H2 O2 . However, ICT did not enhance the protective effect of RO2959, a selective blocker of Orai1. These results indicate that ICT can play a neuroprotective role against oxidative stress injury by binding to Orai1 to block SOCC.

Inhibition of MCF-7 breast cancer cell proliferation by a synthetic peptide derived from the C-terminal sequence of Orai channel.

Intracellular Ca2+ signals play many important cellular functions such as migration, proliferation and differentiation. Store-operated Ca2+ entry (SOCE) is a major route of Ca2+ entry in nonexcitable cells. The activation of SOCE requires engagement between stromal interaction molecule 1 (STIM1) molecules on the endoplasmic reticulum and Ca2+ release-activated Ca2+ (CRAC) channel Orais (Orai1-3) on the plasma membrane. Accumulating evidence indicates that SOCE plays critical roles in cancer cell proliferation, invasion and metastasis. Here, we used the synthetic intracellular peptides derived from the C-termini of Orai channels to treat the breast cancer cells. We have found that Orai3-CT peptide exhibits stronger binding to STIM1 than Orai1-CT, and Orai3-CT peptide acts in a dominant negative fashion, blocking the STIM1-Orai1 interaction and reducing the Ca2+ entry and proliferation of breast cancer cells.