T-type calcium channels, but not Cav3.2, in the peripheral sensory afferents are involved in acute itch in mice.

T-type calcium channels are prominently expressed in primary nociceptive fibers and well characterized in pain processes. Although itch and pain share many similarities including primary sensory fibers, the function of T-type calcium channels on acute itch has not been explored. We investigated whether T-type calcium channels expressed within primary sensory fibers of mouse skin, especially Cav3.2 subtype, involve in chloroquine-, endothelin-1- and histamine-evoked acute itch using pharmacological, neuronal imaging and behavioral analyses. We found that pre-locally blocking three subtypes of T-type calcium channels in the peripheral afferents of skins, yielded an inhibition in acute itch or pain behaviors, while selectively blocking the Cav3.2 channel in the skin peripheral afferents only inhibited acute pain but not acute itch. These results suggest that T-type Cav3.1 or Cav3.3, but not Cav3.2 channel, have an important role in acute itch processing, and their distinctive roles in modulating acute itch are worthy of further investigation.

Effect of Subanesthetic Dose of Esketamine on Perioperative Neurocognitive Disorders in Elderly Undergoing Gastrointestinal Surgery: A Randomized Controlled Trial.

Background: Perioperative neurocognitive disorders (PND), including delayed neurocognitive recovery (dNCR) and postoperative cognitive dysfunction (POCD), are common postoperative complications in elderly patients and adversely affect their prognosis. The study was designed to explore the effects of esketamine on postoperative cognitive function in elderly patients who underwent gastrointestinal surgery under general anesthesia and its potential mechanism. Methods: Eighty-four patients aged 65 and above undergoing gastrointestinal surgery were randomly divided into 2 groups: the esketamine group (group S) and the control group (group C). Group S received intravenous sub-anesthetic doses of esketamine (0.15 mg/kg) 5 minutes before the initiation of surgery, while group C received the same volume of saline. A battery of neuropsychological tests was used to assess cognitive function before surgery, 7 days, and 3 months after surgery. The primary outcome was the incidence of dNCR at 7 days postoperatively and POCD at 3 months postoperatively in both groups. The secondary outcome measures included changes in the levels of serum interleukin-6 (IL-6) and calcium-binding protein ß (S100ß) before and 1 day after surgery. Results: The incidence of dNCR in group S was lower than that of group C (18.15% vs 38.24% P=0.033). Contrarily, there was no difference in both groups regarding POCD 3 months postoperatively (6.06% vs 14.37% P=0.247). Plasma IL-6 and S100ß levels were significantly elevated in both groups on postoperative day 1 (p<0.05), but esketamine pretreatment reduced these levels to some extent compared with group C (p<0.05). Conclusion: Sub-anesthetic doses of esketamine might reduce the incidence of dNCR and improve early postoperative cognitive function in elderly patients undergoing gastrointestinal surgery, which might be related to the anti-neuroinflammation effects of esketamine.

Development and Validation of a Delirium Risk Prediction Model for Elderly Patients Undergoing Elective Orthopedic Surgery.

Purpose: This study aimed to develop and validate a post-operative delirium (POD) nomogram in a population of elderly patients undergoing elective orthopedic surgery. Patients and Methods: A predictive model was developed based on a training dataset of 474 elderly patients undergoing elective orthopedic surgery from March 2021 to May 2022. POD was identified using the Confusion Assessment Methods (CAM). The least absolute shrinkage and selection operator (LASSO) method was used to screen risk factors, and prediction models were created by combining the outcomes with logistic regression analysis. We employ bootstrap validation for internal validation to examine the model's repeatability. The results were validated using a prospective study on 153 patients operated on from January 2022 to May 2022 at another institution. Results: The predictors in the POD nomogram included age, the Mini-Mental State Examination(MMSE), sleep disorder, neurological disorders, preoperative serum creatinine (Pre-SCR), and ASA classification. The c-index of the model was 0.928 (95% confidence interval 0.898 ~ 0.957) and the bootstrap validation still achieved a high c-index of 0.912. The c-index of the external validation was 0.921. The calibration curve for the diagnostic probability showed good agreement between prediction by nomogram and actual observation. Conclusion: By combining preoperative and intraoperative clinical risk factors, we created a POD risk nomogram to predict the probability of POD in elderly patients who undergo elective orthopedic surgery. It could be a tool for guiding individualized interventions.

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.