circRNA TATA-box binding protein associated factor 15 acts as an oncogene to facilitate bladder cancer progression through targeting miR-502-5p/high mobility group box 3.

Circular RNAs (circRNAs) are key in regulating bladder cancer progression. This study explored the effects of circRNA TATA-box binding protein associated factor 15 (circTAF15) on bladder cancer progression. We enrolled 80 bladder cancer patients to examine the relationship between circTAF15 expression and clinical features. The function of circTAF15 on bladder cancer cell viability, proliferation, migration, invasion, and glycolysis was monitored by cell counting kit-8 assay, 5-Ethynyl-2'-deoxyuridine experiment, Transwell experiment, and glycolysis analysis. Dual luciferase reporter gene assay, RNA pull-down assay, and RNA immunoprecipitation assay were used to verify the binding between circTAF15 and miR-502-5p or between miR-502-5p and high mobility group box 3 (HMGB3). circTAF15 effect on in vivo growth of bladder cancer was investigated by xenograft tumor experiment. Quantitative real-time polymerase chain reaction, Western blot, and immunohistochemistry were implemented to investigate the expression levels of genes. circTAF15 was upregulated in bladder cancer patients, associated with unfavorable outcomes. circTAF15 knockdown attenuated bladder cancer cell viability, proliferation, migration, invasion, epithelial-mesenchymal transition, and glycolysis. circTAF15 suppressed miR-502-5p expression, and miR-502-5p inhibited HMGB3 expression. Low miR-502-5p expression was associated with unfavorable outcomes in bladder cancer patients. miR-502-5p silencing and HMGB3 overexpression counteracted the inhibition of circTAF15 knockdown on the malignant phenotype of bladder cancer cells. circTAF15 knockdown attenuated the in vivo growth of bladder cancer cells. circTAF15 enhanced the progression of bladder cancer through upregulating HMGB3 via suppressing miR-502-5p. circTAF15 may be a novel target to treat bladder cancer in the future.

Comparison of clinical outcomes with hip replacement versus PFNA in the treatment of intertrochanteric fractures in the elderly: A systematic review and meta-analysis (PRISMA).

BACKGROUND: The purpose of this meta-analysis was to evaluate the clinical efficacy and safety of HR and PFNA in the treatment of intertrochanteric fractures in the elderly. METHODS: We carried out this review according to the principle of preferred reporting items for systematic reviews and meta-analyses (PRISMA) guideline. The clinical randomized controlled trials (RCTs), prospective cohort studies, retrospective cohort studies (RCSs), and case-control studies involving HR and PFNA in the treatment of intertrochanteric fractures in the elderly from 2000 to 2020 were compared by searching Web of Science, Pubmed, the Cochrane Library, and Embase. The quality of the included cohort study (CS) lines was evaluated using the Newcastle-Ottawa Scale (NOS). The quality of the included RCT lines was evaluated using Jadad. Forest plots were drawn by RevMan5.4 software based on the results and the data were analyzed. RESULTS: After screening, a total of 9 articles were included, of which one was a clinical RCT and eight were RCSs with 1374 patients. The operative time of the PFNA group was shorter [WMD = 15.20; 95% CI (13.17, 17.23), P < .05] and the intraoperative blood loss was less [WMD = 178.81; 95% CI (97.24, 260.38), P < .05] than the HR group, while the first weight-bearing time of the HR group was shorter [WMD = -7.70; 95% CI (-10.54, -4.86), P  < .05] than the PFNA group. There was no significant difference in the length of hospital stay, HHS, postoperative orthopedic complications, and postoperative medical complications between the 2 groups. CONCLUSION: With the development of HR technology and minimally invasive technology, the trauma caused by surgery is decreasing. Under the premise of improving perioperative management, such as optimizing the preoperative preparation and postoperative management, shortening the operative time, reducing intraoperative blood loss, and actively managing co-existing diseases, HR has more advantages than PFNA in the treatment of senile intertrochanteric fractures.

Exosomal 2',3'-CNP from mesenchymal stem cells promotes hippocampus CA1 neurogenesis/neuritogenesis and contributes to rescue of cognition/learning deficiencies of damaged brain.

Mesenchymal stem cells (MSCs) have been used in clinical studies to treat neurological diseases and damage. However, implanted MSCs do not achieve their regenerative effects by differentiating into and replacing neural cells. Instead, MSC secretome components mediate the regenerative effects of MSCs. MSC-derived extracellular vesicles (EVs)/exosomes carry cargo responsible for rescuing brain damage. We previously showed that EP4 antagonist-induced MSC EVs/exosomes have enhanced regenerative potential to rescue hippocampal damage, compared with EVs/exosomes from untreated MSCs. Here we show that EP4 antagonist-induced MSC EVs/exosomes promote neurosphere formation in vitro and increase neurogenesis and neuritogenesis in damaged hippocampi; basal MSC EVs/exosomes do not contribute to these regenerative effects. 2',3'-Cyclic nucleotide 3'-phosphodiesterase (CNP) levels in EP4 antagonist-induced MSC EVs/exosomes are 20-fold higher than CNP levels in basal MSC EVs/exosomes. Decreasing elevated exosomal CNP levels in EP4 antagonist-induced MSC EVs/exosomes reduced the efficacy of these EVs/exosomes in promoting ß3-tubulin polymerization and in converting toxic 2',3'-cAMP into neuroprotective adenosine. CNP-depleted EP4 antagonist-induced MSC EVs/exosomes lost the ability to promote neurogenesis and neuritogenesis in damaged hippocampi. Systemic administration of EV/exosomes from EP4 -antagonist derived MSC EVs/exosomes repaired cognition, learning, and memory deficiencies in mice caused by hippocampal damage. In contrast, CNP-depleted EP4 antagonist-induced MSC EVs/exosomes failed to repair this damage. Exosomal CNP contributes to the ability of EP4 antagonist-elicited MSC EVs/exosomes to promote neurogenesis and neuritogenesis in damaged hippocampi and recovery of cognition, memory, and learning. This experimental approach should be generally applicable to identifying the role of EV/exosomal components in eliciting a variety of biological responses.

EP4 Antagonist-Elicited Extracellular Vesicles from Mesenchymal Stem Cells Rescue Cognition/Learning Deficiencies by Restoring Brain Cellular Functions.

Adult brains have limited regenerative capacity. Consequently, both brain damage and neurodegenerative diseases often cause functional impairment for patients. Mesenchymal stem cells (MSCs), one type of adult stem cells, can be isolated from various adult tissues. MSCs have been used in clinical trials to treat human diseases and the therapeutic potentials of the MSC-derived secretome and extracellular vesicles (EVs) have been under investigation. We found that blocking the prostaglandin E2 /prostaglandin E2 receptor 4 (PGE2 /EP4 ) signaling pathway in MSCs with EP4 antagonists increased EV release and promoted the sorting of specific proteins, including anti-inflammatory cytokines and factors that modify astrocyte function, blood-brain barrier integrity, and microglial migration into the damaged hippocampus, into the EVs. Systemic administration of EP4 antagonist-elicited MSC EVs repaired deficiencies of cognition, learning and memory, inhibited reactive astrogliosis, attenuated extensive inflammation, reduced microglial infiltration into the damaged hippocampus, and increased blood-brain barrier integrity when administered to mice following hippocampal damage. Stem Cells Translational Medicine 2019.

PGE2 /EP4 antagonism enhances tumor chemosensitivity by inducing extracellular vesicle-mediated clearance of cancer stem cells.

Cells expressing mesenchymal/basal phenotypes in tumors have been associated with stem cell properties. Cancer stem cells (CSCs) are often resistant to conventional chemotherapy. We explored overcoming mesenchymal CSC resistance to chemotherapeutic agents. Our goal was to reduce CSC numbers in vivo, in conjunction with chemotherapy, to reduce tumor burden. Analysis of clinical samples demonstrated that COX-2/PGE2 /EP4 signaling is elevated in basal-like and chemoresistant breast carcinoma and is correlated with survival and relapse of breast cancer. EP4 antagonism elicts a striking shift of breast cancer cells from a mesenchymal/CSC state to a more epithelial non-CSC state. The transition was mediated by EP4 antagonist-induced extracellular vesicles [(EVs)/exosomes] which removed CSC markers, mesenchymal markers, integrins, and drug efflux transporters from the CSCs. In addition, EP4 antagonism-induced CSC EVs/exosomes can convert tumor epithelial/non-CSCs to mesenchymal/CSCs able to give rise to tumors and to promote tumor cell dissemination. Because of its ability to induce a CSC-to-non-CSC transition, EP4 antagonist treatment in vivo reduced the numbers of CSCs within tumors and increased tumor chemosensitivity. EP4 antagonist treatment enhances tumor response to chemotherapy by reducing the numbers of chemotherapy-resistant CSCs available to repopulate the tumor. EP4 antagonism can collaborate with conventional chemotherapy to reduce tumor burden.

Transfer of Mammary Gland-forming Ability Between Mammary Basal Epithelial Cells and Mammary Luminal Cells via Extracellular Vesicles/Exosomes.

Cells can communicate via exosomes, ~100-nm extracellular vesicles (EVs) that contain proteins, lipids, and nucleic acids. Non-adherent/mesenchymal mammary epithelial cell (NAMEC)-derived extracellular vesicles can be isolated from NAMEC medium via differential ultracentrifugation. Based on their density, EVs can be purified via ultracentrifugation at 110,000 x g. The EV preparation from ultracentrifugation can be further separated using a continuous density gradient to prevent contamination with soluble proteins. The purified EVs can then be further evaluated using nanoparticle-tracking analysis, which measures the size and number of vesicles in the preparation. The extracellular vesicles with a size ranging from 50 to 150 nm are exosomes. The NAMEC-derived EVs/exosomes can be ingested by mammary epithelial cells, which can be measured by flow cytometry and confocal microscopy. Some mammary stem cell properties (e.g., mammary gland-forming ability) can be transferred from the stem-like NAMECs to mammary epithelial cells via the NAMEC-derived EVs/exosomes. Isolated primary EpCAMhi/CD49flo luminal mammary epithelial cells cannot form mammary glands after being transplanted into mouse fat pads, while EpCAMlo/CD49fhi basal mammary epithelial cells form mammary glands after transplantation. Uptake of NAMEC-derived EVs/exosomes by EpCAMhi/CD49flo luminal mammary epithelial cells allows them to generate mammary glands after being transplanted into fat pads. The EVs/exosomes derived from stem-like mammary epithelial cells transfer mammary gland-forming ability to EpCAMhi/CD49flo luminal mammary epithelial cells.

PGE2 /EP4 Signaling Controls the Transfer of the Mammary Stem Cell State by Lipid Rafts in Extracellular Vesicles.

Prostaglandin E2 (PGE2 )-initiated signaling contributes to stem cell homeostasis and regeneration. However, it is unclear how PGE2 signaling controls cell stemness. This study identifies a previously unknown mechanism by which PGE2 /prostaglandin E receptor 4 (EP4 ) signaling regulates multiple signaling pathways (e.g., PI3K/Akt signaling, TGFß signaling, Wnt signaling, EGFR signaling) which maintain the basal mammary stem cell phenotype. A shift of basal mammary epithelial stem cells (MaSCs) from a mesenchymal/stem cell state to a non-basal-MaSC state occurs in response to prostaglandin E receptor 4 (EP4 ) antagonism. EP4 antagonists elicit release of signaling components, by controlling their trafficking into extracellular vesicles/exosomes in a lipid raft/caveolae-dependent manner. Consequently, EP4 antagonism indirectly inactivates, through induced extracellular vesicle/exosome release, pathways required for mammary epithelial stem cell homeostasis, e.g. canonical/noncanonical Wnt, TGFß and PI3K/Akt pathways. EP4 antagonism causes signaling receptors and signaling components to shift from non-lipid raft fractions to lipid raft fractions, and to then be released in EP4 antagonist-induced extracellular vesicles/exosomes, resulting in the loss of the stem cell state by mammary epithelial stem cells. In contrast, luminal mammary epithelial cells can acquire basal stem cell properties following ingestion of EP4 antagonist-induced stem cell extracellular vesicles/exosomes, and can then form mammary glands. These findings demonstrate that PGE2 /EP4 signaling controls homeostasis of mammary epithelial stem cells through regulating extracellular vesicle/exosome release. Reprogramming of mammary epithelial cells can result from EP4 -mediated stem cell property transfer by extracellular vesicles/exosomes containing caveolae-associated proteins, between mammary basal and luminal epithelial cells. Stem Cells 2017;35:425-444.

Residue-specific annotation of disorder-to-order transition and cathepsin inhibition of a propeptide-like crammer from D. melanogaster.

Drosophila melanogaster crammer is a novel cathepsin inhibitor involved in long-term memory formation. A molten globule-to-ordered structure transition is required for cathepsin inhibition. This study reports the use of alanine scanning to probe the critical residues in the two hydrophobic cores and the salt bridges of crammer in the context of disorder-to-order transition and cathepsin inhibition. Alanine substitution of the aromatic residues W9, Y12, F16, Y20, Y32, and W53 within the hydrophobic cores, and charged residues E8, R28, R29, and E67 in the salt bridges considerably decrease the ability of crammer to inhibit Drosophila cathepsin B (CTSB). Far-UV circular dichroism (CD), intrinsic fluorescence, and nuclear magnetic resonance (NMR) spectroscopies show that removing most of the aromatic and charged side-chains substantially reduces thermostability, alters pH-dependent helix formation, and disrupts the molten globule-to-ordered structure transition. Molecular modeling indicates that W53 in the hydrophobic Core 2 is essential for the interaction between crammer and the prosegment binding loop (PBL) of CTSB; the salt bridge between R28 and E67 is critical for the appropriate alignment of the α-helix 4 toward the CTSB active cleft. The results of this study show detailed residue-specific dissection of folding transition and functional contributions of the hydrophobic cores and salt bridges in crammer, which have hitherto not been characterized for cathepsin inhibition by propeptide-like cysteine protease inhibitors. Because of the involvements of cathepsin inhibitors in neurodegenerative diseases, these structural insights can serve as a template for further development of therapeutic inhibitors against human cathepsins.