MCOLN1/TRPML1 in the lysosome: a promising target for autophagy modulation in diverse diseases.

MCOLN1/TRPML1 is a nonselective cationic channel specifically localized to the late endosome and lysosome. With its property of mediating the release of several divalent cations such as Ca2+, Zn2+ and Fe2+ from the lysosome to the cytosol, MCOLN1 plays a pivotal role in regulating a variety of cellular events including endocytosis, exocytosis, lysosomal biogenesis, lysosome reformation, and especially in Macroautophagy/autophagy. Autophagy is a highly conserved catabolic process that maintains cytoplasmic integrity by removing superfluous proteins and damaged organelles. Acting as the terminal compartments, lysosomes are crucial for the completion of the autophagy process. This review delves into the emerging role of MCOLN1 in controlling the autophagic process by regulating lysosomal ionic homeostasis, thereby governing the fundamental functions of lysosomes. Furthermore, this review summarizes the physiological relevance as well as molecular mechanisms through which MCOLN1 orchestrates autophagy, consequently influencing mitochondria turnover, cell apoptosis and migration. In addition, we have illustrated the implications of MCOLN1-regulated autophagy in the pathological process of cancer and myocardial ischemia-reperfusion (I/R) injury. In summary, given the involvement of MCOLN1-mediated autophagy in the pathogenesis of cancer and myocardial I/R injury, targeting MCOLN1 May provide clues for developing new therapeutic strategies for the treatment of these diseases. Exploring the regulation of MCOLN1-mediated autophagy in diverse diseases contexts will surely broaden our understanding of this pathway and offer its potential as a promising drug target.Abbreviation: CCCP:carbonyl cyanide3-chlorophenylhydrazone; CQ:chloroquine; HCQ: hydroxychloroquine;I/R: ischemia-reperfusion; MAP1LC3/LC3:microtubule associated protein 1 light chain 3; MCOLN1/TRPML1:mucolipin TRP cation channel 1; MLIV: mucolipidosis type IV; MTORC1:MTOR complex 1; ROS: reactive oxygenspecies; SQSTM1/p62: sequestosome 1.

Women in the Medical Physics Workforce: Insights from Membership Trends of the American Association of Physicists in Medicine, 1993 to 2023.

PURPOSE: Women remain underrepresented in medical physics in the United States, and determinants of persisting disparities remain unclear. Here, we performed a detailed investigation of American Association of Physicists in Medicine (AAPM) membership trajectories to evaluate trends in Full membership with respect to gender, age, and highest degree. METHODS AND MATERIALS: Membership data, including gender, date of birth, highest degree, membership type, and years of active membership for 1993 to 2023 were obtained from AAPM. Group 1 included Full members who joined AAPM in 1993 or later. A subset of group 1 including only members who joined and left AAPM since 1993 (former members, group 1F) was used to calculate age at membership cessation and duration. Results were compared by gender and highest degree. A Kaplan-Meier analysis was also used to evaluate membership "survival" by age and highest degree. RESULTS: Complete data were available for 6647 current and former Full members (group 1), including 2211 former members (group 1F). On average, women became Full members at a significantly younger age than men (34.6 vs 37.5 years of age, P < .001) and ended their memberships (if applicable) at a significantly younger age than men (46.1 vs 50.1 years of age, P < .001). The Kaplan-Meier "survival" analysis showed that for a given age, women were at a significantly greater risk of membership cessation than men, and women with master's degrees had the lowest membership survival of any gender/degree subgroup. When analyzing by membership duration, there was no difference in survival by gender alone. Still, women with PhDs were found to have the greatest membership survival among gender/degree subgroups. CONCLUSIONS: Both gender and degree type influenced AAPM membership trajectories. Although we have offered a discussion of possible explanations, qualitative data collected from both continuing and departing AAPM members will be critical in the ongoing journey toward gender parity in the profession of medical physics.

Tumor necrosis factor α-induced protein 8-like-2 controls microglia phenotype via metabolic reprogramming in BV2 microglial cells and responses to neuropathic pain.

Microglial are major players in neuroinflammation that have recently emerged as potential therapeutic targets for neuropathic pain. Glucose metabolic programming has been linked to differential activation state and function in microglia. Tumor necrosis factor α-induced protein 8-like-2 (TNFAIP8L2) is an important component in regulating the anti-inflammatory response. However, the role of TNFAIP8L2 in microglia differential state during neuropathic pain and its interplay with glucose metabolic reprogramming in microglia has not yet been determined. Thus, we aimed to investigate the role of TNFAIP8L2 in the status of microglia in vitro and in vivo. BV2 microglial cells were treated with lipopolysaccharides plus interferon-gamma (LPS/IFNγ) or interleukin-4 (IL-4) to induce the two different phenotypes of microglia in vitro. In vivo experiments were conducted by chronic constriction injury of the sciatic nerve (CCI). We investigated whether TNFAIP8L2 regulates glucose metabolic programming in BV2 microglial cells. The data in vitro showed that TNFAIP8L2 lowers glycolysis and increases mitochondrial oxidative phosphorylation (OXPHOS) in inflammatory microglia. Blockade of glycolytic pathway abolished TNFAIP8L2-mediated differential activation of microglia. TNFAIP8L2 suppresses inflammatory microglial activation and promotes restorative microglial activation in BV2 microglial cells and in spinal cord microglia after neuropathic pain. Furthermore, TNFAIP8L2 controls differential activation of microglia and glucose metabolic reprogramming through the MAPK/mTOR/HIF-1α signaling axis. This study reveals that TNFAIP8L2 plays a critical role in neuropathic pain, providing important insights into glucose metabolic reprogramming and microglial phenotypic transition, which indicates that TNFAIP8L2 may be used as a potential drug target for the prevention of neuropathic pain.

Tumor necrosis factor-α-induced protein 8-like 2 alleviates morphine antinociceptive tolerance through reduction of ROS-mediated apoptosis and MAPK/NF-κB signaling pathways.

Chronic morphine tolerance is a repulsive barrier to the clinical treatment of pain. Whereas the underlying molecular mechanisms of morphine tolerance remain unknown. Here, we proposed that tumor necrosis factor-α-induced protein 8-like 2 (TIPE2) is an essential control point regarding the progression of chronic morphine antinociceptive tolerance. We found that TIPE2 levels in the lumbar spinal cord were significantly downregulated in the morphine tolerance mouse model. Specifically, decreased TIPE2 by morphine tolerance was primarily expressed in spinal neurons, while increased expression of spinal TIPE2 distinctly attenuated the chronic morphine antinociceptive tolerance and tolerance-associated hyperalgesia. We also observed that increased expression of spinal TIPE2 significantly reduced morphine tolerance-induced neuronal ROS production and apoptosis, along with the activation of MAPKs and NF-κB signaling pathways. Moreover, the increased TIPE2 expression inhibited neuronal activation and glial reactivity in the spinal dorsal horn after chronic morphine exposure. Additionally, TIPE2 overexpression in cultured SH-SY5Y cells significantly suppressed ROS production and apoptosis in response to morphine challenge. Therefore, we can conclude that the upregulation of spinal TIPE2 may attenuate the morphine antinociceptive tolerance via TIPE2-dependent downregulation of neuronal ROS, inhibition of neuronal apoptosis, suppression of MAPKs and NF-κB activation. TIPE2 may be a potential strategy for preventing morphine tolerance in the future studies and clinical settings.

Transmission and retention of antibiotic resistance genes (ARGs) in chicken and sheep manure composting.

Transmission of ARGs during composting with different feedstocks (i.e., sheep manure (SM), chicken manure (CM) and mixed manure (MM, SM:CM = 3:1 ratio) was studied by metagenomic sequencing. 53 subtypes of ARGs for 22 types of antibiotics were identified as commonly present in these compost mixes; among them, CM had higher abundance of ARGs, 1.69 times than that in SM, while the whole elimination rate of CM, MM and SM were 55.2%, 54.7% and 42.9%, respectively. More than 50 subtypes of ARGs (with 8.6%, 11.4% and 20.9% abundance in the initial stage in CM, MM and SM composting) were "diehard" ARGs, and their abundance grew significantly to 56.5%, 63.2% and 69.9% at the mature stage. These "diehard" ARGs were transferred from initial hosts of pathogenic and/or probiotic bacteria to final hosts of thermophilic bacteria, by horizontal gene transfer (HGT) via mobile gene elements (MGEs), and became rooted in composting products.

Bacterial community succession in aerobic-anaerobic-coupled and aerobic composting with mown hay affected C and N losses.

The primary objective of this work was to investigate how the dominant microbial species change and affect C and N losses under aerobic and aerobic-anaerobic-coupled composting of mown hay (MH, ryegrass) and corn stover (CS) mix. Results showed that C and N losses in aerobic compost of MH-CS were significantly decreased by 19.57-31.47% and 29.04-41.18%, respectively. 16S rRNA gene sequencing indicated that the bacterial microbiota showed significant differences in aerobic and aerobic-anaerobic-coupled composting. LEfSe analyses showed that aerobic composting promoted the growth of bacteria related to lignocellulosic degradation and nitrogen fixation, while aerobic-anaerobic-coupled composting promoted the growth of bacteria related to denitrification. Correlation analysis between bacterial community and environmental factors indicated that moisture content (MC) was the most important environmental factor influencing the differentiation of bacterial growth. KEGG analysis showed that aerobic composting enhanced the amino acid, carbohydrate, and other advantageous metabolic functions compared to that of aerobic-anaerobic-coupled composting. As a conclusion, the addition of 10-20% corn stover (w/w) to new-mown hay (ryegrass) appeared to inhibit anaerobic composting and prompt aerobic composting in MH-CS mix, which led to the effective utilization of mown hay as a resource for composting.

Dinuclear Group 4 Metal Complexes Bearing Anthracene-Bridged Bifunctional Amido-Ether Ligands: Remarkable Metal Effect and Cooperativity toward Ethylene/1-Octene Copolymerization.

Two types of bifunctional amido-ether ligands (syn-L and anti-L) with the rigid anthracene skeleton were designed to support dinuclear group 4 metal complexes. All organic ligands and organometallic complexes (syn-M2 and anti-M2; M = Hf, Zr, and Ti) were fully characterized by 1H and 13C NMR spectroscopies and elemental analyses. The anti-Hf2 complex showed two confirmations at room temperature with C2-symmetry or S2-symmetry that can inter-exchange, as indicated by VT NMR, while only a C2-symmetric isomer was observed for syn-Hf2 complex at room temperature. However, for Zr and Ti analogues, both syn and anti complexes exhibited only one conformation at room temperature. The molecular structures of complexes syn-Hf2, anti-Hf2, and syn-Ti2 in the solid state were further determined by single-crystal X-ray diffraction, revealing the distances between two metal centers in syn-M2 from 7.138 Å (syn-Ti2) to 7.321 Å (syn-Hf2) but a much farther separation in anti-M2 (8.807 Å in C2-symmetric anti-Hf2). The mononuclear complex (2-CH3O-C6H4-N-C14H9)Zr(NMe2)3 (mono-Zr1) was also prepared for control experiments. In the presence of alkyl aluminum (AlEt3) as the alkylating agent and trityl borate ([Ph3C][B(C6F5)4]) as the co-catalyst, all metal complexes were tested for copolymerization of ethylene with 1-octene at high temperature (130 °C). The preliminary polymerization results revealed that the activity was highly dependent upon the nature of metal centers, and syn-Zr2 showed the highest activity of 9600 kg(PE)·mol-1 (Zr)·h-1, which was about 17- and 2.2-fold higher than those of syn-Hf2 and syn-Ti2, respectively. Benefitting from both steric proximity and electronical interaction of two metal centers, syn-Zr2 exhibited significant cooperativity in comparison to anti-Zr2 and mono-Zr1, with regard to activity and molecular weight and 1-octene incorporation of resultant copolymers.

Continuous cropping of potato changed the metabolic pathway of root exudates to drive rhizosphere microflora.

For potato production, continuous cropping (CC) could lead to autotoxicity buildup and microflora imbalance in the field soil, which may result in failure of crops and reduction in yield. In this study, non-targeted metabolomics (via liquid chromatography with tandem mass spectrometry (LC-MS/MS)) combined with metagenomic profiling (via high-throughput amplicon sequencing) were used to evaluate correlations between metabolomics of potato root exudates and communities of bacteria and fungi around potato plants to illustrate the impacts of CC. Potato plants were grown in soil collected from fields with various CC years (0, 1, 4, and 7 years). Metabolomic analysis showed that the contents and types of potential autotoxins in potato root exudates increased significantly in CC4 and CC7 plants (i.e., grown in soils with 4 and 7 years of CC). The differentially expressed metabolites were mainly produced via alpha-linolenic acid metabolism in plant groups CC0 and CC1 (i.e., no CC or 1 year CC). The metabolomics of the groups CC4 and CC7 became dominated by styrene degradation, biosynthesis of siderophore group non-ribosomal peptides, phenylpropanoid biosynthesis, and biosynthesis of various plant secondary metabolites. Continuous cropping beyond 4 years significantly changed the bacterial and fungal communities in the soil around the potato crops, with significant reduction of beneficial bacteria and accumulation of harmful fungi. Correlations between DEMs and microflora biomarkers were established with strong significances. These results suggested that continuous cropping of potato crops changed their metabolism as reflected in the plant root exudates and drove rhizosphere microflora to directions less favorable to plant growth, and it needs to be well managed to assure potato yield.

Applying formative evaluation in the mentoring of student intern nurses in an emergency department.

Objective: To explore the effectiveness of formative evaluation in the mentoring of student nursing interns in an emergency department. Methods: A total of 144 intern nursing students in the emergency department of a tertiary care hospital in Fuzhou were selected as the study subjects from July 2020 to February 2021. Adopting quasi-experimental studies methods, the students were divided into the experiment group (n = 74) and the control group (n = 70), based on their practicing rotation times. Formative evaluation methods such as in-person interviews, clinical scenario simulations, and clinical operation skills exams were conducted in the experiment group, while traditional summative evaluation methods were adopted for the control group. At the end of the intern period, a unified examination paper on professional knowledge concerning the emergency department, a cardiopulmonary resuscitation skill assessment, and a self-rating scale of self-directed learning was employed to evaluate professional theory performance, clinical practice ability, self-directed learning ability, and academic satisfaction among the nursing students, respectively. Results: The professional theoretical performance, clinical practice ability assessment scores, academic satisfaction, and self-directed learning abilities of the nursing students were significantly higher in the experiment group compared with the control group (P < 0.05). Conclusion: The application of formative evaluation during the mentoring of student intern nurses in an emergency department improved their professional theoretical performance, clinical practice skills, academic satisfaction, and self-directed learning abilities.

Feedstock-dependent abundance of functional genes related to nitrogen transformation controlled nitrogen loss in composting.

The objective of this work was to explore how selection of feedstock affects nitrogen cycle genes during composting, which eventually determines the nitrogen loss. Four composting mixes (CM: chicken manure; SM: sheep manure; MM1/3: mixed manure with CM: SM = 1:3 w/w, MM3/1: CM: SM = 3:1 w/w) were investigated. Results showed that adding 25 % and 75 % SM to CM reduced 26.5 % and 57.9 % nitrogen loss, respectively. CM contained more ammonification genes and nrfA gene, while SM had more denitrification genes. Nitrogen fixation genes in CM were slightly higher than that in SM at the initial stage, but they sharply dropped off as the composting entered the high temperature stage. MM1/3 showed significantly reduced ammonification genes than CM, and increased nitrogen fixation and NH4+ assimilation genes. Therefore, adding SM to CM could change the abundance of genes and enzymes related to nitrogen cycle to reduce nitrogen loss.

TRPML1-induced autophagy inhibition triggers mitochondrial mediated apoptosis.

Previous studies have demonstrated that autophagy tightly regulates apoptosis. However, the underlying mechanism whereby autophagy regulates apoptosis remains unclear. Here, we discover a "autophagy inhibition-mitochondrial turnover disruption-ROS elevation-DNA damage-p53 transactivation-apoptosis" axis that explicates the process of autophagy modulating apoptosis. We found that autophagy inhibition induced by TRPML1, a cationic channel localized in the lysosome, results in accumulation of damaged mitochondria via blocking the mitophagic flux to lysosomes in human melanoma and glioblastoma cells. The disrupted mitochondria turnover leads to ROS elevation, which in turn causes severe damage to DNA in these cancer cells. Damage to DNA resulted from TRPML1-mediated autophagy inhibition subsequently activates p53, which ultimately triggers mitochondrial mediated apoptosis by modulating pro- and anti-apoptosis proteins in these cancer cells. As a result, by triggering apoptosis, TRPML1-induced autophagy inhibition greatly suppresses growth of human melanoma and glioma both in vitro and in vivo. In summary, our findings define the mechanism underling the regulation of autophagy inhibition in apoptosis and represent TRPML1 as a novel target for potentially treating melanoma and glioblastoma in the clinical setting.

Targeting MCOLN1/TRPML1 channels to protect against ischemia-reperfusion injury by restoring the inhibited autophagic flux in cardiomyocytes.

Accumulating evidence suggests that macroautophagy/autophagy dysfunction plays a critical role in myocardial ischemia-reperfusion (I/R) injury. However, the underlying mechanisms responsible for malfunctional autophagy in cardiomyocytes subjected to I/R are poorly understood. As a result, there are no effective therapeutic options that target autophagy to prevent myocardial I/R injury. We recently revealed that MCOLN1/TRPML1, a lysosomal cationic channel, directly contributes to the inhibition of autophagic flux in cardiomyocytes post I/R. We found that MCOLN1 is activated secondary to reactive oxygen species (ROS) elevation following I/R, which in turn induces the release of lysosomal zinc into the cytosol. This ultimately blocks autophagic flux in cardiomyocytes by disrupting the fusion between autophagosomes containing engulfed mitochondria and lysosomes. Furthermore, we discovered that the MCOLN1-mediated inhibition of autophagy induced by I/R impairs mitochondrial function, which results in further detrimental ROS release that directly contributes to cardiomyocyte death. More importantly, restoration of blocked autophagic flux in cardiomyocytes subjected to I/R achieved by blocking MCOLN1 channels significantly rescues cardiomyocyte death in vitro and greatly improves cardiac function of mice subjected to I/R in vivo. Therefore, targeting MCOLN1 represents a novel therapeutic strategy to protect against myocardial I/R injury.Abbreviations: I/R: ischemia-reperfusion; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MCOLN1/TRPML1: mucolipin TRP cation channel 1; ROS: reactive oxygen species; SQSTM1/p62: sequestosome 1.

Blunting TRPML1 channels protects myocardial ischemia/reperfusion injury by restoring impaired cardiomyocyte autophagy.

Accumulating evidence suggests that autophagy dysfunction plays a critical role in myocardial ischemia/reperfusion (I/R) injury. However, the underling mechanism of malfunctional autophagy in the cardiomyocytes subjected to I/R has not been well defined. As a result, there is no effective therapeutic option by targeting autophagy to prevent myocardial I/R injury. Here, we used both an in vitro and an in vivo I/R model to monitor autophagic flux in the cardiomyocytes, by exposing neonatal rat ventricular myocytes to hypoxia/reoxygenation and by subjecting mice to I/R, respectively. We observed that the autophagic flux in the cardiomyocytes subjected to I/R was blocked in both in vitro and in vivo models. Down-regulating a lysosomal cationic channel, TRPML1, markedly restored the blocked myocardial autophagic flux induced by I/R, demonstrating that TRPML1 directly contributes to the blocked autophagic flux in the cardiomyocytes subjected to I/R. Mechanistically, TRPML1 is activated secondary to ROS elevation following ischemia/reperfusion, which in turn induces the release of lysosomal zinc into the cytosol and ultimately blocks the autophagic flux in cardiomyocytes, presumably by disrupting the fusion between autophagosomes and lysosomes. As a result, the inhibited myocardial autophagic flux induced by TRPML1 disrupted mitochondria turnover and resulted in mass accumulation of damaged mitochondria and further ROS release, which directly led to cardiomyocyte death. More importantly, pharmacological and genetic inhibition of TRPML1 channels greatly reduced infarct size and rescued heart function in mice subjected to I/R in vivo by restoring impaired myocardial autophagy. In summary, our study demonstrates that secondary to ROS elevation, activation of TRPML1 results in autophagy inhibition in the cardiomyocytes subjected to I/R, which directly leads to cardiomyocyte death by disrupting mitochondria turnover. Therefore, targeting TRPML1 represents a novel therapeutic strategy to protect against myocardial I/R injury.

Autophagy inhibition mediated by MCOLN1/TRPML1 suppresses cancer metastasis via regulating a ROS-driven TP53/p53 pathway.

Compelling evidence has demonstrated that macroautophagy/autophagy plays an important role in regulating multiple steps of metastatic cascades; however, the precise role of autophagy in metastasis remains unclear. This study demonstrates that autophagy inhibition induced by MCOLN1/TRPML1 suppresses cancer metastasis by evoking the ROS-mediated TP53/p53 pathway. First, we found that MCOLN1-mediated autophagy inhibition not only profoundly inhibits both migration and invasion in malignant melanoma and glioma cell lines in vitro, but also suppresses melanoma metastasis in vivo. Second, our study reveals that autophagy inhibition induced by MCOLN1 leads to damaged mitochondria accumulation followed by large quantities of ROS release. Third, we demonstrate that the elevated ROS resulting from autophagy inhibition subsequently triggers TP53 activity, which in turn modulates expression of its downstream targets that are involved in a broad spectrum of the metastatic cascade to suppress metastasis including MMP members and TWIST. In summary, our findings have established a mechanism by which autophagy inhibition suppresses metastasis via the ROS-TP53 signaling pathway. More importantly, our study demonstrates that autophagy inhibition through stimulation of MCOLN1 could evidently be one of the therapeutic potentials for combating cancer metastasis.Abbreviations: 3-MA: 3-methyladenine; AA: amino acid; ATG5: autophagy related 5; ATG12: autophagy-related 12; Baf-A1: bafilomycin A1; CCCP: carbonyl cyanide m-chlorophenylhydrazone; CQ: chloroquine; DMEM: Dulbecco's Modified Eagle Medium; EMT: epithelial-mesenchymal transition; FBS: fetal bovine serum; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; HEK: human embryonic kidney; LAMP1: lysosomal-associated membrane protein 1; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MCOLN1/TRPML1: mucolipin TRP cation channel 1; MMP: matrix metallopeptidase; NC: negative control; NRK: normal rat kidney; PBS: phosphate-buffered saline; shRNA: short hairpin RNA; siRNA: short interfering RNA; SQSTM1/p62: sequestosome 1; ULK1: unc-51 like autophagy-activating kinase 1.

Stimulating TRPM7 suppresses cancer cell proliferation and metastasis by inhibiting autophagy.

The transient receptor potential melastatin-subfamily member 7 (TRPM7) is a ubiquitous cation channel possessing kinase activity. TRPM7 mediates a variety of physiological responses by conducting flow of cations such as Ca2+, Mg2+, and Zn2+. Here, we show that the activation of TRPM7 channel stimulated by chemical agonists of TRPM7, Clozapine or Naltriben, inhibited autophagy via mediating Zn2+ release to the cytosol, presumably from the intracellular Zn2+-accumulating vesicles where TRPM7 localizes. Zn2+ release following the activation of TRPM7 disrupted the fusion between autophagosomes and lysosomes by disturbing the interaction between Sxt17 and VAMP8 which determines fusion status of autophagosomes and lysosomes. Ultimately, the disrupted fusion resulting from stimulation of TRPM7 channels arrested autophagy. Functionally, we demonstrate that the autophagy inhibition mediated by TRPM7 triggered cell death and suppressed metastasis of cancer cells in vitro, more importantly, restricted tumor growth and metastasis in vivo, by evoking apoptosis, cell cycle arrest, and reactive oxygen species (ROS) elevation. These findings represent a strategy for stimulating TRPM7 to combat cancer.

MCOLN1/TRPML1 finely controls oncogenic autophagy in cancer by mediating zinc influx.

Macroautophagy/autophagy is elevated to ensure the high demand for nutrients for the growth of cancer cells. Here we demonstrated that MCOLN1/TRPML1 is a pharmaceutical target of oncogenic autophagy in cancers such as pancreatic cancer, breast cancer, gastric cancer, malignant melanoma, and glioma. First, we showed that activating MCOLN1, by increasing expression of the channel or using the MCOLN1 agonists, ML-SA5 or MK6-83, arrests autophagic flux by perturbing fusion between autophagosomes and lysosomes. Second, we demonstrated that MCOLN1 regulates autophagy by mediating the release of zinc from the lysosome to the cytosol. Third, we uncovered that zinc influx through MCOLN1 blocks the interaction between STX17 (syntaxin 17) in the autophagosome and VAMP8 in the lysosome and thereby disrupting the fusion process that is determined by the two SNARE proteins. Furthermore, we demonstrated that zinc influx originating from the extracellular fluid arrests autophagy by the same mechanism as lysosomal zinc, confirming the fundamental function of zinc as a participant in membrane trafficking. Last, we revealed that activating MCOLN1 with the agonists, ML-SA5 or MK6-83, triggers cell death of a number of cancer cells by evoking autophagic arrest and subsequent apoptotic response and cell cycle arrest, with little or no effect observed on normal cells. Consistent with the in vitro results, administration of ML-SA5 in Patu 8988 t xenograft mice profoundly suppresses tumor growth and improves survival. These results establish that a lysosomal cation channel, MCOLN1, finely controls oncogenic autophagy in cancer by mediating zinc influx into the cytosol.Abbreviation: Abbreviations: 3-MA: 3-methyladenine; AA: amino acid; ATG12: autophagy related 12; Baf-A1: bafilomycin A1; BAPTA-am: 1,2-bis(2-aminophenoxy)ethane-N, N,N',N'-tetraacetic acid tetrakis-acetoxymethyl ester; co-IP: coimmunoprecipitaion; CQ: chloroquine; DMEM: Dulbecco's Modified Eagle Medium; FBS: fetal bovine serum; GAPDH: glyceraldehyde- 3-phosphate dehydrogenase; HCQ: hydroxychloroquine; HEK: human embryonic kidney; LAMP1: lysosomal associated membrane protein 1; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MCOLN1/TRPML1: mucolipin TRP cation channel 1; MTORC1: mechanistic target of rapamycin kinase complex 1; NC: negative control; NRK: normal rat kidney epithelial cells; PBS: phosphate-buffered saline; PtdIns3K: phosphatidylinositol 3-kinase; RPS6KB/S6K: ribosomal protein S6 kinase B; shRNA: short hairpin RNA; siRNA: short interfering RNA; SNARE: soluble N-ethylmaleimide-sensitive factor (NSF) attachment protein receptor; SQSTM1/p62: sequestosome 1; STX17: syntaxin 17; TPEN: N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine; TTM: tetrathiomolybdate; ULK1: unc-51 like autophagy activating kinase 1; VAMP8: vesicle associated membrane protein 8; Zn2+: zinc.

The synergistic effects of opioid and neuropeptide B/W in rat acute inflammatory and neuropathic pain models.

The use of morphine is controversial due to the incidence of rewarding behavior, respiratory depression, and tolerance, leading to increased drug dose requirements, advancing to morphine addiction. To overcome these barriers, strategies have been taken to combine morphine with other analgesics. Neuropeptide B23 and neuropeptide W23 (NPB23 and NPW23) are commonly used to relieve inflammatory pain and neuropathic pain. As NPB23 and NPW23 system shares similar anatomical basis with opioid system at least in the spinal cord we hypothesized that NPB23 or NPW23 and morphine may synergistically relieve inflammatory pain and neuropathic pain. To test this hypothesis, we demonstrated that µ opioid receptor and NPBW1 receptor (receptor of NPB23 and NPW23) are colocalized in the superficial dorsal horn of the spinal cord. Secondly, co-administration of morphine witheitherNPB23 or NPW23 synergistically attenuated inflammatory and neuropathic pain. Furthermore, either NPB23 or NPW23 significantly reduced morphine-induced conditioned place preference (CPP) and constipation. We also found that phosphorylation of extracellular-regulated protein kinase (ERK1/2) following morphine was profoundly potentiated by the application of NPB23 or NPW23. Hence, combination of morphine with either NPB23 or NPW23 reduced dose of morphine required for pain relief in inflammatory and neuropathic pain, while effectively prevented some side-effects of morphine.

Hemodynamic management of a patient with a huge right atrium myxoma during thoracic vertebral surgery: A case report.

RATIONALE: Myxoma is the most common primary benign cardiac tumor, which could lead to some fatal complications because of its strategic position. PATIENT CONCERNS: The patient was admitted to our hospital due to sudden onset of palpitation, chest tightness, mild fever, night sweats, accompanied with bilateral lower extremities adynamia, and paralysis for 5 days, but no obvious syncope and edema. DIAGNOSES: Transthoracic echocardiography showed a giant mobile myxoma (72 × 58 mm) in the right atrium (RA). Magnetic resonance imaging revealed an erosive space-occupying lesion located between the first and third thoracic vertebrae. INTERVENTIONS: Thoracic vertebral lesions were resected immediately to rescue the incomplete paraplegia. After the patient was placed in the prone position, significant hemodynamics changes were observed due to the displacement of the huge RA myxoma. OUTCOMES: Stable hemodynamics was maintained during the operation through control of fluid infusion combined with vasoactive drugs. LESSONS: Change in body position may lead to obstruction of intracardiac blood flow in patients with giant myxoma. This clinical manifestation is rarely reported.

BACE1 gene silencing alleviates isoflurane anesthesia­induced postoperative cognitive dysfunction in immature rats by activating the PI3K/Akt signaling pathway.

Postoperative cognitive dysfunction (POCD) is a severe complication characterized by cognitive dysfunction following anesthesia and surgery. The aim of the present study was to investigate the effects of ß­site amyloid precursor protein cleavage enzyme 1 (BACE1) gene silencing on isoflurane anesthesia­induced POCD in immature rats via the phosphatidylinositol­3­kinase (PI3K)/protein kinase B (Akt) signaling pathway. Rat models were established and then transfected with BACE1 small interfering RNA and wortmannin (an inhibitor of PI3K). Blood gas analysis was performed, and a series of behavioral experiments were conducted to evaluate the cognitive function, learning ability and locomotor activity of rats. Reverse transcription quantitative polymerase chain reaction and western blot analysis were employed to determine the mRNA and protein expression of the associated genes. An ELISA was used to detect the inflammatory indicators and the content of amyloid precursor protein (APP) and amyloid­ß (Aß). Apoptosis of the hippocampal CA1 region was observed by terminal deoxynucleotidyl transferase dUTP nick­end labeling staining. Initially, it was revealed that the percentage of stagnation time in rats was increased by BACE1 gene silencing; the escape latency and swimming distance were markedly reduced from the 4th to the 6th day, the time the rats spent in first passing the target area was shortened, and the times of passing the target area were increased by BACE1 gene silencing, demonstrating that BACE1 gene silencing enhanced the spatial memory ability of rats. Additionally, it was determined that silencing BACE1 improved the pathological state induced by isoflurane anesthesia in immature rats, and attenuated the inflammatory response and the levels of APP and Aß in hippocampal tissues. Furthermore, it was suggested that silencing BACE1 may have promoted the activation of the PI3K/Akt signaling pathway, thereby inhibiting the apoptosis of the hippocampal CA1 region. Taken together, these results indicated that BACE1 gene silencing may improve isoflurane anesthesia­induced POCD in immature rats by activating the PI3K/Akt signaling pathway and inhibiting the Aß generated by APP.

Lidocaine sensitizes the cytotoxicity of 5-fluorouacil in melanoma cells via upregulation of microRNA-493.

Lidocaine is a well-documented local anesthetic that has been reported to sensitize the cytotoxicity of cisplatin in cancer cells. However, little information is available concerning whether lidocaine sensitizes the cytotoxicity of 5-fluorouracil (5-FU) in melanoma cells. The study was aimed to explore the effects and mechanisms of lidocaine on the sensitivity to 5-FU in the melanoma cell line SK-MEL-2. Cell viability and apoptosis were analyzed after administration of different concentrations of lidocaine, 5-FU, or the combinations. Expression of microRNA (miR)-493 was assessed following lidocaine administration. The target genes of miR-493 were verified by luciferase reporter assay, PCR, and Western blot. The effects of abnormal expression of miR-493 and/or SRY-Box 4 (SOX4) on cell viability, apoptosis, and key proteins in phosphatidylinositol-3-kinase (PI3K)/AKT and the Smad pathways were detected. The effects of (0-100 uM) lidocaine on cell viability and apoptosis was not obvious; however, lidocaine could significantly increase the cell viability and inhibit apoptosis in 5-FU-treated cells. In addition, lidocaine induced upregulation of miR-493 in a dose-dependent manner, and we confirmed that the effects of miR-493 on the sensitivity were by upregulating miR-493. Moreover, we verified that Sox4 was a target of miR-493, and Sox4 overexpression decreased the sensitivity to 5-FU. Besides, Sox4 overexpression increased the levels of p-PI3K, p-AKT, p-Smad2 and p-Smad3, and Sox4 suppression showed contrary results. Our results suggest that lidocaine sensitizes the cytotoxicity of 5-FU in melanoma cells via upregulation of miR-493, which might be involved in SOX4-mediated PI3K/AKT and Smad pathways.