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Objective To investigate the expression of SLC35A2 and PFDN2 in breast cancer and their relationship with clinical indicators and prognosis.Methods TCGA database and TIMER 2.0 database were used to analyze the differences of SLC35A2 and PFDN2 expression in breast cancer tissues and paracancerous tissues;K-M database was used to create the survival curves of patients in the high and low expression groups of the two.qRT-PCR and immunohistochemistry were used to detect the expression of SLC35A2 and PFDN2 in the cancerous and paracancerous tissues,and the expression differences,the relationship between their expression levels and the clinical observation indexes were statistically analyzed,and the independent prognostic factors of breast cancer were screened out;K-M survival analysis was used to compare the prognostic differences between the groups and create the survival curves.Results The expression levels of SLC35A2 and PFDN2 in breast cancer tissues were significantly higher than those in paracancerous tissues according to the results of biopsy,qRT-PCR and immuno-histochemistry,and the expression levels of SLC35A2 were significantly correlated with lymph node metastasis,while the expression of PFDN2 was significantly correlated with the diameter of the tumor and the metastasis of lymph nodes,and the expression of SLC35A2 and PFDN2 was an independent prognostic factor for breast cancer.patients had the worst prognosis.Conclusion The expression of SLC35A2 and PFDN2 in breast cancer tissues was closely related to clinical indicators and prognosis of breast cancer,and could be used as a potential target for breast cancer treatment.
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Objective:To evaluate the effects of electroacupuncture (EA) on mitochondrial fusion and fission during intestinal injury in mice with endotoxemia and the role of heme oxygenase-1 (HO-1).Methods:Fifty SPF-grade healthy male C57BL/6 mice, aged 8 weeks, weighing 18-22 g, were divided into 5 groups ( n=10 each) using a random number table method: control group (group C), endotoxemia group (group E), endotoxemia plus EA group (group E+ EA), endotoxemia plus EA plus hemin group (group E+ EA+ H) and endotoxemia plus EA plus Znpp-Ⅸ group (group E+ EA+ Znpp-Ⅸ). Lipopolysaccharide (LPS) was intraperitoneally injected to develop the model of endotoxemia.Before LPS injection, the HO-1 inducer hemin 100 mg/kg was intraperitoneally injected in group E+ EA+ H, and the HO-1 inhibitor zinc protoporphyrin Ⅸ 10 μmol/kg was intraperitoneally injected in group E+ EA+ Znpp-Ⅸ.At 4, 3, 2 and 1 days and 30 min prior to development of the model, Zusanli and Hegu acupoints were stimulated with electric stimulator (disperse-dense wave, frequency 2 Hz/15 Hz, at a voltage of 1 mA) for 30 min, retaining the needle until the end of the experiment on the day of development of the model.Mice were sacrificed at 6 h after development of the model, and the small intestinal tissue was obtained from the terminal ileum for examination of the pathological results (with a light microscope) and ultrastructure of mitochondria (with an electron microscope) and for determination of the levels of reactive oxygen species (ROS), ATP and diamine oxidase (DAO) and expression of dynamin-related protein 1 (Drp1), mitofusin 1 (Mfn1) and HO-1 (by Western blot). Results:Compared with group C, the level of ROS was significantly increased, ATP content and DAO activity were decreased, the expression of HO-1 and Drp1 was up-regulated, the expression of Mfn1 was down-regulated ( P<0.05), and pathological damage to small intestine tissues was found in group E. Compared with group E, the level of ROS was significantly decreased, ATP content and DAO activity were increased, the expression of HO-1 and Mfn1 was up-regulated, the expression of Drp1 was down-regulated ( P<0.05), and pathological damage to small intestine tissues was significantly attenuated in group E+ EA.Compared with group E+ EA, the level of ROS was significantly decreased, ATP content and DAO activity were increased, the expression of HO-1 and Mfn1 was up-regulated, the expression of Drp1 was down-regulated ( P<0.05), and pathological damage to small intestine tissues was significantly attenuated in group E+ EA+ H, and the level of ROS was significantly increased, ATP content and DAO activity were decreased, the expression of HO-1 and Mfn1 was down-regulated, the expression of Drp1 was up-regulated ( P<0.05), and pathological damage to small intestine tissues was accenuated in group E+ EA+ Znpp-Ⅸ. Conclusions:EA can promote mitochondrial fusion, inhibit mitochondrial fission, and alleviate intestinal damage in mice with endotoxemia, and the mechanism is related to the up-regulation of HO-1 expression.
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Diarrhea occurs in 2-50% of cases of COVID-19 ([~]8% is average across series). The diarrhea does not appear to account for the disease mortality and its contribution to the morbidity has not been defined, even though it is a component of Long Covid or post-infectious aspects of the disease. Even less is known about the pathophysiologic mechanism of the diarrhea. To begin to understand the pathophysiology of COVID-19 diarrhea, we exposed human enteroid monolayers obtained from five healthy subjects and made from duodenum, jejunum, and proximal colon to live SARS-CoV-2 and virus like particles (VLPs) made from exosomes expressing SARS-CoV-2 structural proteins (Spike, Nucleocapsid, Membrane and Envelope). Results: 1) Live virus was exposed apically for 90 min, then washed out and studied 2 and 5 days later. SARS-Cov-2 was taken up by enteroids and live virus was present in lysates and in the apical>>basolateral media of polarized enteroids 48 h after exposure. This is the first demonstration of basolateral appearance of live virus after apical exposure. High vRNA concentration was detected in cell lysates and in the apical and basolateral media up to 5 days after exposure. 2) Two days after viral exposure, cytokine measurements of media showed significantly increased levels of IL-6, IL-8 and MCP-1. 3) Two days after viral exposure, mRNA levels of ACE2, NHE3 and DRA were reduced but there was no change in mRNA of CFTR. NHE3 protein was also decreased. 4) Live viral studies were mimicked by some studies with VLP exposure for 48 h. VLPs with Spike-D614G bound to the enteroid apical surface and was taken up; this resulted in decreased mRNA levels of ACE2, NHE3, DRA and CFTR. 4) VLP effects were determined on active anion secretion measured with the Ussing chamber/voltage clamp technique. S-D614G acutely exposed to apical surface of human ileal enteroids did not alter the short-circuit current (Isc). However, VLPS-D614G exposure to enteroids that were pretreated for [~]24 h with IL-6 plus IL-8 induced a concentration dependent increase in Isc indicating stimulated anion secretion, that was delayed in onset by [~]8 min. The anion secretion was inhibited by apical exposure to a specific calcium activated Cl channel (CaCC) inhibitor (AO1) but not by a specific CFTR inhibitor (BP027); was inhibited by basolateral exposure to the K channel inhibit clortimazole; and was prevented by pretreatment with the calcium buffer BAPTA-AM. 5) The calcium dependence of the VLP-induced increase in Isc was studied in Caco-2/BBe cells stably expressing the genetically encoded Ca2+ sensor GCaMP6s. 24 h pretreatment with IL-6/IL-8 did not alter intracellular Ca2+. However, in IL-6/IL-8 pretreated cells, VLP S-D614G caused appearance of Ca2+waves and an overall increase in intracellular Ca2+ with a delay of [~]10 min after VLP addition. We conclude that the diarrhea of COVID-19 appears to an example of a calcium dependent inflammatory diarrhea that involves both acutely stimulated Ca2+ dependent anion secretion (stimulated Isc) that involves CaCC and likely inhibition of neutral NaCl absorption (decreased NHE3 protein and mRNA and decreased DRA mRNA).
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The spike D614G mutation increases SARS-CoV-2 infectivity, viral load, and transmission but the molecular mechanism underlying these effects remains unclear. We report here that spike is trafficked to lysosomes and that the D614G mutation enhances the lysosomal sorting of spike and the lysosomal accumulation of spike-positive punctae in SARS-CoV-2-infected cells. Spike trafficking to lysosomes is an endocytosis-independent, V-ATPase-dependent process, and spike-containing lysosomes drive lysosome clustering but display poor lysotracker labeling and reduced uptake of endocytosed materials. These results are consistent with a lysosomal pathway of coronavirus biogenesis and raise the possibility that a common mechanism may underly the D614G mutations effects on spike protein trafficking in infected cells and the accelerated entry of SARS-CoV-2 into uninfected cells.
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Expression-dependent, Spike-only vaccines have been developed, deployed, and shown to be effective in the fight against SARS-CoV-2. However, additional approaches to vaccine development may be needed to meet existing and future challenges posed by emerging Spike variant strains, as well as a likely need for different antigen-delivery systems that are safe and effective for regular, periodic re-administration. We report here the development of mRNA-loaded exosomes, demonstrate that they can mediate the functional expression of heterologous proteins in vitro and in vivo, and have fewer adverse effects than comparable doses of lipid nanoparticles. Furthermore, we applied this approach to the development of an exosome-based, multiplexed mRNA vaccine that drives expression of immunogenic SARS-CoV-2 Nucleocapsid and Spike proteins. This vaccine elicited long-lasting cellular and humoral responses to Nucleocapsid and to Spike, demonstrating that exosome-based mRNA formulations represent a previously unexplored platform in the fight against COVID-19 and other infectious diseases.
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Based on the theory of cancer stem cells (CSCs),people have been searching for the treatments of malignant cancers.Gastric cancer side population cells (SP) have the characteristics of CSCs.Searching for the molecular targeted therapy strategy of gastric cancer which embarks from the gastric cancer SP and is based on the theory of CSCs provides a new direction for the treatment,early diagnosis,therapeutic effect and prognosis of gastric cancer.