Endothelial cell Orai1 is essential for endothelium-dependent contraction of mouse carotid arteries in normotensive and hypertensive mice.

Endothelium-dependent contraction (EDC) exists in blood vessels of normotensive animals, but is exaggerated in hypertension. An early signal in EDC is cytosolic Ca2+ rise in endothelial cells. In this study we investigated the functional role of Orai1, a major endothelial cell Ca2+ entry channel, in EDC. Hypertension model was established in WT mice by intake of L-NNA in the drinking water (0.5 g/L) for 4 weeks or osmotic pump delivery of Ang II (1.5 mg·kg-1·d-1) for 2 weeks. In TRPC5 KO mice, the concentration of L-NNA and Ang II were increased to 1 g/L or 2 mg·kg-1·d-1, respectively. Arterial segments were prepared from carotid arteries and aortas, and EDC was elicited by acetylcholine in the presence of Nω-nitro-L-arginine methyl ester. We showed that low concentration of acetylcholine (3-30 nM) initiated relaxation in phenylephrine-precontracted carotid arteries of both normotensive and hypertensive mice, while high concentration of acetylcholine (0.1-2 µM) induced contraction. Application of selective Orai1 inhibitors AnCoA4 (100 µM) or YM58483 (400 nM) had no effect on ACh-induced relaxation but markedly reduced acetylcholine-induced EDC. We found that EDC was increased in hypertensive mice compared with that of normotensive mice, which was associated with increased Orai1 expression in endothelial cells of hypertensive mice. Compared to TRPC5 and TRPV4, which were also involved in EDC, endothelial cell Orai1 had relatively greater contribution to EDC than either TRPC5 or TRPV4 alone. We identified COX-2, followed by PGF2α, PGD2 and PGE2 as the downstream signals of Orai1/TRPC5/TRPV4. In conclusion, Orai1 coordinates together with TRPC5 and TRPV4 in endothelial cells to regulate EDC responses. This study demonstrates a novel function of Orai1 in EDC in both normotensive and hypertensive mice, thus providing a general scheme about the control of EDC by Ca2+-permeable channels.

Thioridazine protects against disturbed flow-induced atherosclerosis by inhibiting RhoA/YAP-mediated endothelial inflammation.

Atherosclerotic diseases remain the leading cause of adult mortality and impose heavy burdens on health systems globally. Our previous study found that disturbed flow enhanced YAP activity to provoke endothelial activation and atherosclerosis, and targeting YAP alleviated endothelial inflammation and atherogenesis. Therefore, we established a luciferase reporter assay-based drug screening platform to seek out new YAP inhibitors for anti-atherosclerotic treatment. By screening the FDA-approved drug library, we identified that an anti-psychotic drug thioridazine markedly suppressed YAP activity in human endothelial cells. Thioridazine inhibited disturbed flow-induced endothelial inflammatory response in vivo and in vitro. We verified that the anti-inflammatory effects of thioridazine were mediated by inhibition of YAP. Thioridazine regulated YAP activity via restraining RhoA. Moreover, administration of thioridazine attenuated partial carotid ligation- and western diet-induced atherosclerosis in two mouse models. Overall, this work opens up the possibility of repurposing thioridazine for intervention of atherosclerotic diseases. This study also shed light on the underlying mechanisms that thioridazine inhibited endothelial activation and atherogenesis via repression of RhoA-YAP axis. As a new YAP inhibitor, thioridazine might need further investigation and development for the treatment of atherosclerotic diseases in clinical practice.

A Novel Probiotic-Based Oral Vaccine against SARS-CoV-2 Omicron Variant B.1.1.529.

COVID-19 pandemic, caused by the SARS-CoV-2 virus, is still affecting the entire world via the rapid emergence of new contagious variants. Vaccination remains the most effective prevention strategy for viral infection, yet not all countries have sufficient access to vaccines due to limitations in manufacturing and transportation. Thus, there is an urgent need to develop an easy-to-use, safe, and low-cost vaccination approach. Genetically modified microorganisms, especially probiotics, are now commonly recognized as attractive vehicles for delivering bioactive molecules via oral and mucosal routes. In this study, Lactobacillus casei has been selected as the oral vaccine candidate based on its' natural immunoadjuvant properties and the ability to resist acidic gastric environment, to express antigens of SARS-CoV-2 Omicron variant B.1.1.529 with B-cell and T-cell epitopes. This newly developed vaccine, OMGVac, was shown to elicit a robust IgG systemic immune response against the spike protein of Omicron variant B.1.1.529 in Golden Syrian hamsters. No adverse effects were found throughout this study, and the overall safety was evaluated in terms of physiological and histopathological examinations of different organs harvested. In addition, this study illustrated the use of the recombinant probiotic as a live delivery vector in the initiation of systemic immunity, which shed light on the future development of next-generation vaccines to combat emerging infectious diseases.

Precise Modulation of Intramolecular Aggregation-induced Electrochemiluminescence by Tetraphenylethylene-based Supramolecular Architectures.

The precisely modulated synthesis of programmable light-emitting materials remains a challenge. To address this challenge, we construct four tetraphenylethylene-based supramolecular architectures (SA, SB, SC, and SD), revealing that they exhibit higher electrochemiluminescence (ECL) intensities and efficiencies than the tetraphenylethylene monomer and can be classified as highly efficient and precisely modulated intramolecular aggregation-induced electrochemiluminescence (PI-AIECL) systems. The best-performing system (SD) shows a high ECL cathodic efficiency exceeding that of the benchmark tris(2,2'-bipyridyl)ruthenium(II) chloride in aqueous solution by nearly six-fold. The electrochemical characterization of these architectures in an organic solvent provides deeper mechanistic insights, revealing that SD features the lowest electrochemical band gap. Density functional theory calculations indicate that the band gap of the guest ligand in the SD structure is the smallest and most closely matched to that of the host scaffold. Finally, the SD system is used to realize ECL-based cysteine detection (detection limit=14.4 nM) in real samples. Thus, this study not only provides a precisely modulated supramolecular strategy allowing chromophores to be controllably regulated on a molecular scale, but also inspires the programmable synthesis of high-performance aggregation-induced electrochemiluminescence emitters.

Endogenous ion channels expressed in human embryonic kidney (HEK-293) cells.

Mammalian expression systems, particularly the human embryonic kidney (HEK-293) cells, combined with electrophysiological studies, have greatly benefited our understanding of the function, characteristic, and regulation of various ion channels. It was previously assumed that the existence of endogenous ion channels in native HEK-293 cells could be negligible. Still, more and more ion channels are gradually reported in native HEK-293 cells, which should draw our attention. In this regard, we summarize the different ion channels that are endogenously expressed in HEK-293 cells, including voltage-gated Na+ channels, Ca2+ channels, K+ channels, Cl- channels, nonselective cation channels, TRP channels, acid-sensitive ion channels, and Piezo channels, which may complicate the recording of the heterogeneously expressed ion channels to a certain degree. We noted that the expression patterns and channel profiles varied with different studies, which may be due to the distinct originality of the cells, cell culture conditions, passage numbers, and different recording protocols. Therefore, a better knowledge of endogenous ion channels may help minimize potential problems in characterizing heterologously expressed ion channels. Based on this, it is recommended that HEK-293 cells from unknown sources should be examined before transfection for the characterization of their functional profile, especially when the expression level of exogenous ion channels does not overwhelm the endogenous ion channels largely, or the current amplitude is not significantly higher than the native currents.

Arabidopsis ENDOMEMBRANE PROTEIN 12 contributes to the endoplasmic reticulum stress response by regulating K/HDEL receptor trafficking.

ENDOMEMBRANE PROTEIN 70 (EMP70) proteins constitute a 12-member superfamily in Arabidopsis thaliana, and are the most abundant protein species in plant Golgi proteomes. However, the physiological functions of EMPs in plants remain largely unknown. Here we have demonstrated that two AtEMP12 T-DNA insertion mutants are sensitive to ER (endoplasmic reticulum) stress as induced by tunicamycin and dithiothreitol treatments. Interestingly, the unfolded protein response (UPR) is constitutively activated in the knockout mutant emp12-1 under normal growth conditions, suggesting that the activation is a result of insufficient chaperones in the ER to aid protein folding. Indeed, we have further shown that BiP is secreted into the apoplast in emp12-1, while the K/HDEL receptor ERD2a, which regulates BiP trafficking, is exclusively localized in the ER in emp12-1, instead of its known ER-Golgi dual-localization. Given an enhanced retrograde transport of ERD2a, along with less dimerized receptor formed in the absence of EMP12, ERD2a may be prematurely returned to the ER without its bound ligands. Therefore, we propose that EMP12 may act as a novel regulator of the K/HDEL receptor to ensure an effective retrograde transport of K/HDEL ligands.

Development and validation of MRI-based radiomics signatures models for prediction of disease-free survival and overall survival in patients with esophageal squamous cell carcinoma.

OBJECTIVES: To develop and validate an optimal model based on the 1-mm-isotropic-3D contrast-enhanced StarVIBE MRI sequence combined with clinical risk factors for predicting survival in patients with esophageal squamous cell carcinoma (ESCC). METHODS: Patients with ESCC at our institution from 2015 to 2017 participated in this retrospective study based on prospectively acquired data, and were randomly assigned to training and validation groups at a ratio of 7:3. Random survival forest (RSF) and variable hunting methods were used to screen for radiomics features and LASSO-Cox regression analysis was used to build three models, including clinical only, radiomics only and combined clinical and radiomics models, which were evaluated by concordance index (CI) and calibration curve. Nomograms and decision curve analysis (DCA) were used to display intuitive prediction information. RESULTS: Seven radiomics features were selected from 434 patients, combined with clinical features that were statistically significant to construct the predictive models of disease-free survival (DFS) and overall survival (OS). The combined model showed the highest performance in both training and validation groups for predicting DFS ([CI], 0.714, 0.729) and OS ([CI], 0.730, 0.712). DCA showed that the net benefit of the combined model and of the clinical model is significantly greater than that of the radiomics model alone at different threshold probabilities. CONCLUSIONS: We demonstrated that a combined predictive model based on MR Rad-S and clinical risk factors had better predictive efficacy than the radiomics models alone for patients with ESCC. KEY POINTS: • Magnetic resonance-based radiomics features combined with clinical risk factors can predict survival in patients with ESCC. • The radiomics nomogram can be used clinically to predict patient recurrence, DFS, and OS. • Magnetic resonance imaging is highly reproducible in visualizing lesions and contouring the whole tumor.

TRPV1 channels regulate the automaticity of embryonic stem cell-derived cardiomyocytes through stimulating the Na+ /Ca2+ exchanger current.

Calcium controls the excitation-contraction coupling in cardiomyocytes. Embryonic stem cell-derived cardiomyocytes (ESC-CMs) are an important cardiomyocyte source for regenerative medicine and drug screening. Transient receptor potential vanilloid 1 (TRPV1) channels are nonselective cation channels that permeate sodium and calcium. This study aimed to investigate whether TRPV1 channels regulate the electrophysiological characteristics of ESC-CMs. If yes, what is the mechanism behind? By immunostaining and subcellular fractionation, followed by western blotting, TRPV1 was found to locate intracellularly. The staining pattern of TRPV1 was found to largely overlap with that of the sarco/endoplasmic reticulum Ca2+ -ATPase, the sarcoplasmic reticulum (SR) marker. By electrophysiology and calcium imaging, pharmacological blocker of TRPV1 and the molecular tool TRPV1ß (which could functionally knockdown TRPV1) were found to decrease the rate and diastolic depolarization slope of spontaneous action potentials, and the amplitude and frequency of global calcium transients. By calcium imaging, in the absence of external calcium, TRPV1-specific opener increased intracellular calcium; this increase was abolished by preincubation with caffeine, which could deplete SR calcium store. The results suggest that TRPV1 controls calcium release from the SR. By electrophysiology, TRPV1 blockade and functional knockdown of TRPV1 decreased the Na+ /Ca2+ exchanger (NCX) currents from both the forward and reverse modes, suggesting that sodium and calcium through TRPV1 stimulate the NCX activity. Our novel findings suggest that TRPV1 activity is important for regulating the spontaneous activity of ESC-CMs and reveal a novel interplay between TRPV1 and NCX in regulating the physiological functions of ESC-CMs.

Endolysosomal ion channel MCOLN2 (Mucolipin-2) promotes prostate cancer progression via IL-1ß/NF-κB pathway.

BACKGROUND: Prostate cancer (Pca) is the most common cancer type among males worldwide. Dysregulation of Ca2+ signaling plays important roles during Pca progression. However, there is lack of information about the role of endolysosomal Ca2+ -permeable channels in Pca progression. METHODS: The expression pattern of MCOLN2 was studied by immunohistochemistry and western blot. Cell viability assay, transwell assay and in vivo tumorigenesis were performed to evaluate the functional role of MCOLN2. Downstream targets of MCOLN2 were investigated by cytokine array, enzyme-linked immunosorbent assay, Ca2+ release experiments and luciferase reporter assays. RESULTS: We report that MCOLN2 expression is significantly elevated in Pca tissues, and associated with poor prognosis. Overexpression of MCOLN2 promoted Pca cells proliferation, migration and invasion. Importantly, knockdown of MCOLN2 inhibited Pca xenograft tumor growth and bone lesion development in vivo. In addition, MCOLN2 promoted the production and release of IL-1ß. Moreover, luciferase reporter assay and western blot revealed that MCOLN2 promoted Pca development by regulating the IL-1ß/NF-κB pathway. CONCLUSION: In summary, MCOLN2 is crucially involved in Pca progression. Mechanistically, MCOLN2 regulates Pca progression via IL-1ß/NF-κB pathway. Our study highlights an intriguing possibility of targeting MCOLN2 as potential therapeutic strategy in Pca treatment.

Nanostructured Lateral Boryl Substitution Conjugated Donor-Acceptor Oligomers for Visible-Light-Driven Hydrogen Production.

Poor charge separation is the main factor that limits the photocatalytic hydrogen generation efficiency of organic conjugated polymers. In this work, a series of linear donor-acceptor (D-A) type oligomers are synthesized by a palladium-catalyzed Sonogashira-Hagihara coupling of electron-deficient diborane unit and different dihalide substitution sulfur functionalized monomers. Such diborane-based A unit exerts great impact on the resulting oligomers, including distinct semiconductor characters with isolated lowest unoccupied molecular orbital (LUMO) orbits locating in diborane-containing fragment, and elevated LUMO level higher than water reduction potential. Relative to A-A type counterpart, the enhanced dipole polarization effect in D-A oligomers facilitates separation of photogenerated charge carriers, as evidenced by notably prolonged electron lifetime. Owing to π-π stacking of rigid backbone, the oligomers can aggregate into an interesting 2D semicrystalline nanosheet (≈2.74 nm), which is rarely reported in linear polymeric photocatalysts prepared by similar carbon-carbon coupling reaction. Despite low surface area (30.3 m2 g-1 ), such ultrathin nanosheet D-A oligomer offers outstanding visible light (λ > 420 nm) hydrogen evolution rate of 833 µmol g-1 h-1 , 14 times greater than its A-A analogue (61 µmol g-1 h-1 ). The study highlights the great potential of using boron element to construct D-A type oligomers for efficient photocatalytic hydrogen generation.

Serum exosomes mediate delivery of arginase 1 as a novel mechanism for endothelial dysfunction in diabetes.

Exosomes, abundant in blood, deliver various molecules to recipient cells. Endothelial cells are directly exposed to circulating substances. However, how endothelial cells respond to serum exosomes (SExos) and the implications in diabetes-associated vasculopathy have never been explored. In the present study, we showed that SExos from diabetic db/db mice (db/db SExos) were taken up by aortic endothelial cells, which severely impaired endothelial function in nondiabetic db/m+ mice. The exosomal proteins, rather than RNAs, mostly account for db/db SExos-induced endothelial dysfunction. Comparative proteomics analysis showed significant increase of arginase 1 in db/db SExos. Silence or overexpression of arginase 1 confirmed its essential role in db/db SExos-induced endothelial dysfunction. This study is a demonstration that SExos deliver arginase 1 protein to endothelial cells, representing a cellular mechanism during development of diabetic endothelial dysfunction. The results expand the scope of blood-borne substances that monitor vascular homeostasis.

The TRPC5 channel regulates angiogenesis and promotes recovery from ischemic injury in mice.

Ischemia-related diseases are a leading cause of death worldwide, and promoting therapeutic angiogenesis is key for effective recovery from hypoxia-ischemia. Given the limited success of angiogenic factors, such as vascular endothelial growth factor, in clinical trials, it is important to find more promising angiogenic targets. Here, using both cell- and tissue-based assays and a mouse model of injury-induced ischemia, we investigated the involvement of the transient receptor potential canonical 5 (TRPC5) ion channel in angiogenesis and the effects of a TRPC5 activator, the Food and Drug Administration-approved drug riluzole, on recovery from ischemic injury. We demonstrate that TRPC5 is involved in endothelial cell sprouting, angiogenesis, and blood perfusion in an oxygen-induced retinopathy model and a hind limb ischemia model. We found a potential regulatory link between nuclear factor of activated T cell isoform c3 and angiopoietin-1 that could provide the mechanistic basis for the angiogenic function of TRPC5. Importantly, treatment with riluzole, which can activate TRPC5 in endothelial cells, improved recovery from ischemia in mice. Our study reveals TRPC5 as a potential angiogenic target and suggests riluzole as a promising drug for managing ischemic diseases.

The activity of transient receptor potential channel C-6 modulates the differentiation of fat cells.

Previously, the V1-3 isoforms of the transient receptor potential channel (TRP) have been shown to promote or prevent adipocyte differentiation. In the current study, the C isoforms were screened for blocking adipogenesis. The hypothesis that the TRP classic or canonical (TRPC) deters adipocyte differentiation was investigated in 3T3-L1 cells employing the channel-specific activator and antagonist, silencing, and overexpression techniques. Fat accumulation in cells was visualized by Oil Red O staining. Intracellular calcium inflow was estimated by confocal microscopy. A high-fat (HF) feeding study was also performed on C57BL/6J mice to verify the findings in the cell model. Among the 6 C isoforms tested, only TRPC-6 inhibited the differentiation of fat cells. The phytochemical quercetin induced the channel protein expression. Calcium-imaging results also revealed that the flavonoid could trigger calcium inflow. Coadministration of quercetin (1 or 20 mg/kg body weight) in an HF diet prevented TRPC-6 from declining and attenuated phosphorylated (p)-PKB and PI3k, as well as the proliferation of visceral fat cells. The present study illustrated that TRPC-6 activation could perturb adipocyte differentiation. The food flavonoid quercetin was a TRPC-6 inducer and activator and it could prevent adipogenesis in mice.-Tan, Y. Q., Kwan, H. Y., Yao, X., Leung, L. K. The activity of transient receptor potential channel C-6 modulates the differentiation of fat cells.

TRPP2 and STIM1 form a microdomain to regulate store-operated Ca2+ entry and blood vessel tone.

BACKGROUND: Polycystin-2 (TRPP2) is a Ca2+ permeable nonselective cationic channel essential for maintaining physiological function in live cells. Stromal interaction molecule 1 (STIM1) is an important Ca2+ sensor in store-operated Ca2+ entry (SOCE). Both TRPP2 and STIM1 are expressed in endoplasmic reticular membrane and participate in Ca2+ signaling, suggesting a physical interaction and functional synergism. METHODS: We performed co-localization, co-immunoprecipitation, and fluorescence resonance energy transfer assay to identify the interactions of TRPP2 and STIM1 in transfected HEK293 cells and native vascular smooth muscle cells (VSMCs). The function of the TRPP2-STIM1 complex in thapsigargin (TG) or adenosine triphosphate (ATP)-induced SOCE was explored using specific small interfering RNA (siRNA). Further, we created TRPP2 conditional knockout (CKO) mouse to investigate the functional role of TRPP2 in agonist-induced vessel contraction. RESULTS: TRPP2 and STIM1 form a complex in transfected HEK293 cells and native VSMCs. Genetic manipulations with TRPP2 siRNA, dominant negative TRPP2 or STIM1 siRNA significantly suppressed ATP and TG-induced intracellular Ca2+ release and SOCE in HEK293 cells. Inositol triphosphate receptor inhibitor 2-aminoethyl diphenylborinate (2APB) abolished ATP-induced Ca2+ release and SOCE in HEK293 cells. In addition, TRPP2 and STIM1 knockdown significantly inhibited ATP- and TG-induced STIM1 puncta formation and SOCE in VSMCs. Importantly, knockdown of TRPP2 and STIM1 or conditional knockout TRPP2 markedly suppressed agonist-induced mouse aorta contraction. CONCLUSIONS: Our data indicate that TRPP2 and STIM1 are physically associated and form a functional complex to regulate agonist-induced intracellular Ca2+ mobilization, SOCE and blood vessel tone. Video abstract.

Resveratrol Stimulates the Na+-Ca2+ Exchanger on the Plasma Membrane to Reduce Cytosolic Ca2+ in Rat Aortic Smooth Muscle Cells.

Resveratrol is well known to exhibit vascular relaxant and antihypertensive effects. In this study, we determined the effects of resveratrol on the modulation of cytosolic [Ca] level and adenosine 5'-triphosphate-induced Ca release from the sarcoplasmic reticulum (SR) in rat aortic smooth muscle cells (ASMCs) and explored its underlying mechanisms. In this article, cytosolic [Ca] and SR [Ca] in ASMCs were determined by Fluo-4/acetoxymethyl and Mag-Fluo-4/acetoxymethyl respectively. Resveratrol (20, 50, and 100 µM) caused a rapid and substantial reduction in cytosolic [Ca] in ASMCs bathed in normal Hank's Balanced Salt Solution or Ca-free Hank's Balanced Salt Solution. Pretreatment with resveratrol reduced adenosine 5'-triphosphate-induced SR Ca release and SR Ca content. In the cells bathed in Na-free physiological saline, which favors the reverse mode of the Na-Ca exchanger (NCX), resveratrol induced an increase in cytosolic [Ca] and SR [Ca]. However, its effect on cytosolic [Ca] was inhibited by the selective NCX inhibitor, SEA0400. Our findings suggest that resveratrol reduces cytosolic [Ca] and SR [Ca] in ASMCs in normal physiological saline, which might be, at least in part, mediated by the NCX.

Comparison of Chest CT Manifestations of Coronavirus Disease 2019 (COVID-19) and Pneumonia Associated with Lymphoma.

Objective: To retrospectively compare the clinical features and chest computed tomography (CT) characteristics of coronavirus disease 2019 (COVID-19) and pneumonia in lymphoma patients. Materials and Methods: Ten lymphoma patients with pneumonia and 12 patients with COVID-19 infections were enrolled from January 15 to March 14, 2020. The clinical features were recorded. All pulmonary lesions on chest CT were assessed for location, shape, density and diffusion degree. Other typical CT features were also evaluated. Results: The most commonly observed patchy lesions were ground-glass opacities (GGOs) and mixed GGOs in both groups. Regarding the diffusion degree, 82% (92/112) of the lesions in the COVID-19 group were relatively limited, while 69% (52/75) of those in the lymphoma group were diffuse (p < 0.001). The proportions of interlobular septal thickening, vascular thickening, pleural involvement and fibrous stripes observed in the lymphoma cases were statistically compatible with those observed in the COVID-19 cases (p > 0.05). Air bronchograms were observed more frequently in COVID-19 patients (45%, 50/112) than in lymphoma patients with pneumonia (5%, 4/75) (p < 0.001). Halo sign (6%) and reversed halo sign (1%) were observed in several COVID-19 patients but not in lymphoma-associated pneumonia patients. Conclusion: Both lymphoma-associated pneumonia and COVID-19 generally manifested as patchy GGOs and mixed GGOs in more than one lobe. Compared to COVID-19, lymphoma-associated pneumonia tended to be relatively diffuse, with fewer air bronchograms, and no halo or reversed halo signs observed on chest CT.

Polycystin-2 Plays an Essential Role in Glucose Starvation-Induced Autophagy in Human Embryonic Stem Cell-Derived Cardiomyocytes.

Autophagy is a process essential for cell survival under stress condition. The patients with autosomal dominant polycystic kidney disease, which is caused by polycystin-1 or polycystin-2 (PKD2) mutation, display cardiovascular abnormalities and dysregulation in autophagy. However, it is unclear whether PKD2 plays a role in autophagy. In the present study, we explored the functional role of PKD2 in autophagy and apoptosis in human embryonic stem cell-derived cardiomyocytes. HES2 hESC line-derived cardiomyocytes (HES2-CMs) were transduced with adenoviral-based PKD2-shRNAs (Ad-PKD2-shRNAs), and then cultured with normal or glucose-free medium for 3 hours. Autophagy was upregulated in HES2-CMs under glucose starvation, as indicated by increased microtubule-associated protein 1 light chain 3-II level in immunoblots and increased autophagosome and autolysosome formation. Knockdown of PKD2 reduced the autophagic flux and increased apoptosis under glucose starvation. In Ca2+ measurement, Ad-PKD2-shRNAs reduced caffeine-induced cytosolic Ca2+ rise. Co-immunoprecipitation and in situ proximity ligation assay demonstrated an increased physical interaction of PKD2 with ryanodine receptor 2 (RyR2) under glucose starvation condition. Furthermore, Ad-PKD2-shRNAs substantially attenuated the starvation-induced activation of AMP-activated protein kinase (AMPK) and inactivation of mammalian target of rapamycin (mTOR). The present study for the first time demonstrates that PKD2 functions to promote autophagy under glucose starvation, thereby protects cardiomyocytes from apoptotic cell death. The mechanism may involve PKD2 interaction with RyR2 to alter Ca2+ release from sarcoplasmic reticulum, consequently modulating the activity of AMPK and mTOR, resulting in alteration of autophagy and apoptosis. Stem Cells 2018;36:501-513.

TRPV6 protects ER stress-induced apoptosis via ATF6α-TRPV6-JNK pathway in human embryonic stem cell-derived cardiomyocytes.

Human pluripotent stem cell-derived cardiomyocytes have potential applications in disease modeling and drug screening. Therefore, it is important to understand the mechanisms and signaling pathways underlying the survival and death of these cells. Endoplasmic reticulum (ER) stress is triggered by various cellular stresses that disturb protein folding in the ER. Cells cope with ER stress by activating the unfolded protein response (UPR), a homeostatic signaling network that orchestrates the recovery of ER function. In the present study, we hypothesized that ER stress may upregulate the expression of transient receptor potential channel TRPV6, which in turn serves to protect human embryonic stem cell-derived cardiomyocytes (hESC-CMs) from ER stress-induced apoptotic cell death. Indeed, we found that ER stress induced by thapsigargin and tunicamycin led to increased expression of TRPV6 via ATF6α signaling branch. siRNA-mediated knockdown of TRPV6 aggravated ER stress-induced apoptotic cell death, whereas overexpression of TRPV6 attenuated ER stress-induced apoptosis in hESC-CMs. Furthermore, the signaling pathway downstream of TRPV6 was MAPK-JNK. Taken together, these results provide strong evidence that, under ER stress, TRPV6 is upregulated to protect hESC-CMs from apoptotic cell death via ATF6α-TRPV6-JNK pathway.

Orai1 is critical for Notch-driven aggressiveness under hypoxic conditions in triple-negative breast cancers.

It is believed that hypoxia stimulates triple-negative breast cancers (TNBCs) metastasis, which is associated with a poor prognosis. However, the underlying mechanism remains unclear. Here, we demonstrated that hypoxia up-regulates both the levels of Orai1 and Notch1, and the increase in Orai1 is mediated by Notch1 signaling in TNBCs. Functionally, Orai1 caused a sustained elevation of intracellular Ca2+ via Store-operated Ca2+ entry (SOCE), then activated the calcineurin-nuclear factor of activated T-cell 4 (NFAT4, also named NFATc3) in hypoxic TNBCs. Furthermore, pharmacologic inhibition or gene-silencing studies showed that the aggressiveness mediated by Orai1 during hypoxia is dependent on the Notch1/Orai1/SOCE/NFAT4 signaling. Moreover, Orai1 signaling also mediated hypoxia-induced angiogenesis in TNBCs. Thus, our results revealed a novel role of Orai1 as an inducer of aggression and angiogenesis under hypoxic conditions, and this suggests a novel mechanism of hypoxia-induced invasion. It may be worthwhile to further explore the potential of using Orai1 signaling as new target for anti-tumor therapy in TNBCs.

Auto-oxidation promoted sp3 C-H arylation of glycine derivatives.

An auto-oxidation promoted sp3 C-H arylation reaction between N-aryl glycine derivatives and electron-rich arenes, leading to the formation of N-aryl α-aryl α-amino acid derivatives, is described. This atom-economical and environmentally benign reaction proceeds smoothly under mild reaction conditions and requires only Brønsted acid and oxygen (balloon). A plausible radical involved mechanism is proposed.