bluuebunny/crossref_metadata_2025_split huggingface dataset:
Suppression of DAPK1 benefits to ischemia brain injury via mechanism(s) of ER stress mediated apoptotic pathway
Authors: Kim Joo‐Heon , Won Jinyoung , Hong Yonggeun 
Published: November 2023

Stroke occurs as results of arterial occlusion in the brain, and leads to irreversible neuronal injury. Activation of death‐associated protein kinase 1 (DAPK1) induces endoplasmic reticulum (ER) stress and triggers apoptotic signal pathway. Here, we examined whether DAPK1 is involved in ER stress mediated apoptotic cell death in focal cerebral ischemia. To demonstrate the effect of DAPK1 on ER stress dependent cell death, the experimental animals were subjected to middle cerebral artery occlusion and injected with oxazolone‐based DAPK1 inhibitor. The experimental animals exhibited increase of total infarct volume and neurologic damage after focal cerebral ischemia. Molecular mechanism related with ER stress response and apoptosis was differently expressed in DAPK1 inhibitor‐treated group compared vehicle group. While the expression of BCL‐2 related to anti‐apoptosis was up regulated, the expression of BAX and caspase‐3 related to pro‐apoptosis was down regulated in DAPK1 inhibitor group compared to vehicle group. It was shown that neuronal apoptosis combined with altered expression of ER stress sensor marker including XBP1, ATF6 and GRP78/BiP in lesion. These results indicated that suppression of DAPK1 induces apoptotic cell death and protects against ischemic neuronal damage. Thus, DAPK1 would be a novel therapeutic approach for ischemic injury because suppression of DAPK1 induces neuronal cell rescue and promotes neural regeneration.Support or Funding InformationNRF□2013R1A2A2A01067169, KGM4611613 to Y.H., J‐H.K. was supported by development fund foundation (2015) from GSNU. Republic of Korea
Involvement of NF2 Signaling Pathway in β‐Adrenergic Receptor‐Stimulated Cardiac Myocyte Apoptosis
Authors: Dalal Suman , Connelly Barbara A. , Singh Mahipal , Singh Krishna 
Published: November 2023

β‐adrenergic receptor (β‐AR) stimulation induces cardiac myocyte apoptosis in vitro and in vivo. Tumor suppressor protein NF2 (neurofibromin 2, also known as merlin) is a member of the ezrin/radixin/moesin (ERM) family of proteins. Post‐translational modifications such as phosphorylation and sumoylation affect NF2 activity, subcellular localization and function. Role of NF2 in inhibition of cell growth and apoptosis is well established in a variety of cell types. However, the role of NF2 in β‐AR‐stimulated cardiac myocyte apoptosis remains to be investigated. Here we provide evidence that treatment of adult rat ventricular myocytes (ARVMs) with β‐AR agonist (isoproterenol, ISO, 10 μM) increases phosphorylation (serine 518) and sumoylation of NF2. Activation of adenylyl cyclase using forskolin mimicked the effects of ISO. ISO‐mediated increase in phosphorylation of NF2 was inhibited by protein kinase A inhibitor (H89) and β1‐AR receptor antagonist (CGP 20712A). Knockdown of NF2 using siRNA decreased ISO‐mediated increase in the number of apoptotic ARVMs (CTL, 8.3±0.6; ISO, 23.2±1.0*; negSiRNA+ISO, 21.3±1.2*; NF2‐SiRNA+ISO, 12.0±1.5$; *p<0.05 vs CTL; $p<0.05 vs ISO and negSiRNA+ISO, n=3–4). Adenoviral‐mediated expression of WT‐NF2 led to increased phosphorylation and sumoylation of NF2, and apoptosis. It also activated JNKs, a pro‐apoptotic kinase. Treatment with ISO and forskolin, or adenoviral‐mediated expression of NF2 increased phosphorylation (inactivation) of YAP, a downstream target of NF2. In H9C2 cardiomyocytes, knockdown of NF2 using siRNA decreased YAP phosphorylation. These data suggest that β‐AR stimulation affects post‐translational modification of NF2 via the involvement of PKA pathway, and NF2 plays a pro‐apoptotic role in β‐AR‐stimulated myocyte apoptosis.Support or Funding InformationSupported by Department of Veterans Affairs (BX002332), NIH R15HL129140 (NHLBI) and Institutional Research and Improvement account.
The Role of CIP4 in SH‐SY5Y Cell Proliferation and Differentiation
Authors: Suthivanich Phichaya , Saengsawang Witchuda 
Published: November 2023

Cdc42‐interacting protein 4 (CIP4) is a protein in the F‐BAR family, which plays important roles in various cellular events including endocytosis and membrane protrusion in several cell types. CIP4 has been shown to coordinate membrane dynamics and cytoskeletal reorganization in several cell types. CIP4 is expressed in several tissues including the developing brain. However, the role of CIP4 in the brain is still unclear. Here we showed that CIP4 was highly expressed in mouse brain during an early stage of development (embryonic day 10.5), which is a period that neural progenitor cells actively proliferate and differentiate. We then determined the roles of CIP4 during cell proliferation and differentiation using neuroblastoma (SH‐SY5Y) cells as a model. We showed that CIP4 level was significantly increased in differentiated cells, which was induced by retinoic acid compared with un‐differentiated cells (proliferating cells). Overexpression of CIP4 resulted in a decrease in cell proliferation. In addition, CIP4 knockdown completely abolished the ability of retinoic acid to induce differentiation of SH‐SY5Y cells. Together these results suggest that CIP4 may play an important role during an early stage of brain development, possibly by inhibiting cell proliferation and promoting cell differentiation.Support or Funding InformationThis work was supported by Faculty of Science, Mahidol University and a grant from Mahidol University to WS.
Xenotransplantation of human mesenchymal stem cells alters host gene expression following prolonged high fat feeding
Authors: Newell Christopher , Hittel Dustin S. , Nyamandi Vongai , Khan Aneal , Shearer Jane 
Published: November 2023

BackgroundInitially identified as plastic‐adherent cells, human mesenchymal stem cells (hMSCs) are non‐hematopoietic stem cells which are capable of differentiating into a multitude of cell lineages. Over the past decade the number of clinical trials involving hMSCs has steadily increased, resulting in hMSCs being the most commonly used cells in tissue engineering and regenerative medicine. Known to preferentially home to sites of injury and inflammation, administered hMSCs have exhibited promotion of host tissue repair through donor cell engraftment, release of cell signaling factors, and transfer of healthy organelles. As such, enhanced hMSC engraftment to host tissues and improved therapeutic potential may be induced by a state of low‐grade inflammation. Here we induce low‐grade inflammation through excessive nutrient consumption, achieved through prolonged high fat feeding.PurposeTo examine the impact of hMSC treatment on the liver tissue of mice following a prolonged high fat feeding regiment.MethodsAt 4 weeks of age, mice were placed on a high fat diet (60% kcal fat) for 20 weeks in order to elicit an obese phenotype. Following dietary manipulation, mice were randomly divided into two groups: hMSC treatment (HFM; n = 8) or saline control (HFS; n = 8). Mitochondrial metabolism, microarray gene expression profiling, and metabolomic profiling of host liver tissue were specifically highlighted for their important role in the obese phenotype.ResultsAs a measure of mitochondrial metabolic activity, maximal complex I oxygen consumption (supported by the addition of ADP) was higher in HFM animals (p<0.05). The marker of mitochondrial abundance, citrate synthase, was also elevated following hMSC administration (p<0.05). Microarray gene expression profiling identified 226 genes with significant differential expression between groups ‐ fold change (±1.2) and p‐value (<0.01). Specific gene networks and pathway analysis commonly identified cellular signalling and actin cytoskeleton regulation as upregulated processes in HFM mice (p<0.01).ConclusionsOur data indicate that xenogeneic hMSC treatment improves metabolic capacity of host liver and results in widespread host gene shifting. The upregulation of cell signaling and actin cytoskeleton regulation may provide mechanistic insight into how hMSCs impart their ability to migrate to areas of inflammation and transfer organelles to host tissues.Support or Funding InformationThis study was supported by NSERC (JS). This research was supported by PhD funding to CN from MitoCanada and Alberta Innovates – Health Solutions.
Sex Differences in Intestinal Epithelial Crypt Cell Proliferation
Authors: Zhou Weinan , Suriyavirun Natcha , Dailey Megan J. 
Published: November 2023

The intestinal epithelium plays an essential role in ingestive behaviors and immune homeostasis. The epithelium is continually renewed every few days by the proliferation and differentiation of stem cells located in the intestinal crypts. Studies have shown that the luminal nutrient content drives changes in the stem cell proliferative process, but it is unknown whether there are male‐female differences in these nutrient‐driven changes. To determine whether sex differences affect crypt cell proliferation, we utilized primary small intestinal epithelial crypts isolated from male and female C57BL/6J mice to investigate the differences in the gene expression of steroid hormone receptors, crypt cell proliferation, and crypt cell proliferation in response to steroid hormones. We found that estrogen receptor α was expressed in crypts from both male and female mice, while estrogen receptor β, androgen receptor, and progesterone receptor were not expressed in crypts from either sex. Crypt cell proliferation was higher in crypts isolated from female mice compared with crypts isolated form male mice, however, 17β‐Estradiol had no effect on crypt cell proliferation in either sex. These results suggest that sex differences may influence crypt cell proliferation, but these differences may not be driven by steroid hormones.Support or Funding InformationUSDA Hatch ILLU‐538‐926
Body on‐chip and 3D culture: An overview
Authors: Klingelhutz Aloysius John , Gourronc Francoise , Chaly Anna , Ankrum James 
Published: November 2023

The development of better methods to grow human tissue in conditions that more closely recapitulate normal physiology and anatomy (i.e. biomimetic technology) will greatly enhance drug and toxin testing as well as advance our understanding of biology. Two dimensional (2D) cell culture has been extremely valuable for research, but cells grown in 2D culture are not subject to the same mechanical and physiologic conditions as cells in tissue. While animal studies are an alternative, rodents and humans often exhibit large differences with regard to their responses to drugs and toxins. Furthermore, animal studies can be expensive. Growing cells in 3D conditions is challenging but advances are being made that allow researchers to do so. One relatively straightforward method relies on hanging drop technology to develop 3D spheroids. These spheroids often exhibit differential growth and differentiation as compared to 2D culture. Methods to generate and grow spheroids as well as our experience with the development of human adipose spheroids will be discussed. Tissue on‐chip technology in a microfluidics platform raises the possibility of incorporating 3D tissue into a system that allows one to carefully regulate what the tissues are exposed to and to measure what they secrete. Advances are being made in development of tissue on‐chip culture systems for lung, cardiac, and liver, as well as other tissues. These approaches could greatly advance the ability to test how different factors affect physiology. In addition, it may also be possible to make connections between different tissue types (i.e. body on‐chip), thus allowing an assessment of how one cell or tissue type affects another. Innovative methods for 3D culture and tissue/body on‐chip technology will be reviewed.Support or Funding InformationDevelopment of a human white fat biomimetic, University of Iowa Vice President for Research Internal Funding Initiation; Role of aryl hydrocarbon receptor in adipogenesis and diabetes, University of Iowa Fraternal Order of Eagles Diabetes Research Center Pilot Award
Macrophage migration inhibitory factor (MIF) but not its homologue D‐dopachrome tautomerase (D‐DT) promotes fibroblast motility in a CD44/CD74‐independent manner
Authors: Szczesniak Pawel , Meinhardt Andreas , Klug Jörg 
Published: November 2023

Macrophage migration inhibitory factor (MIF) and D‐dopachrome tautomerase (D‐DT) are paralogous pro‐inflammatory cytokines with chemokine‐like functions. Whereas cytosolic MIF plays a role in cell cycle, secreted MIF and D‐DT activate second messenger signalling through a CD44/CD74 receptor complex. To differentiate between receptor‐ and non‐receptor‐mediated events, COS‐7/M6 fibroblasts deficient in CD44/CD74 (WT) were employed to generate stable cell lines expressing CD44 and CD74. Chemokinesis (random single‐cell motility) was assessed in cells stimulated with native recombinant MIF/D‐DT, enzyme‐null recombinant MIF, inhibitors of MIF's tautomerase activity, and inhibitors of clathrin‐ and non‐clathrin‐dependent endocytosis. In the presence of CD44/CD74, both MIF and D‐DT stimulated chemokinesis but only MIF enhanced chemokinesis in the absence of receptors. The stimulatory effect of MIF on migration depended on its tautomerase activity and lipid raft/caveolae‐mediated, but not clathrin‐mediated endocytosis. To observe a direct effect of MIF and D‐DT on actin dynamics, actin polymerization was quantified in an in vitro actin assembly assay. MIF but not D‐DT decreased the rate of F‐actin assembly. By decreasing the rate of cell extract‐driven actin assembly, MIF phenocopies the function of F‐actin capping proteins. Moreover, stimulation with MIF increased the number of prominent F‐actin stress fibres in COS‐7/M6 WT. Taken together, MIF stimulates fibroblast chemokinesis independent of its receptors, likely by directly modulating the actin cytoskeleton. MIF‐mediated chemokinesis appears to depend on lipid raft/caveolae‐mediated endocytosis. D‐DT triggers chemokinesis only in the presence of MIF/D‐DT receptors – CD44/CD74 – implying D‐DT requires the classical receptor‐driven second‐messenger transduction pathway.Support or Funding InformationIntramural funding.
Lipid signaling regulates fibroblast migration and the actin cytoskeleton in response to stiffness and microtopography
Authors: Mkrtschjan Michael , Gaikwad Snehal , Dommaraju Sagar , Li Jieli , Russell Brenda 
Published: November 2023

Cell migration depends on mechanotransduction—the conversion of mechanical cues from the microenvironment to internal biochemical cellular signals, such as by protein phosphorylation. Lipid signals are more difficult to study than proteins, but also play a key role in mechanotransduction pathways. In this study, we apply bioengineering approaches to manipulate physical cues of topography and stiffness in order to reprogram cells, assessing the effects of lipid signaling (PIP2) on the actin cytoskeleton. Primary cardiac fibroblasts were grown on polyacrylamide or polydimethylsiloxane (PDMS) with stiffness of 10, 100, or 400 kPa and glass (>1 GPa). Micropost topography was fabricated from PDMS (400 kPa) with a 75 μm‐spaced tetragonal array. An increase in actin assembly rate was determined by the kfrap kinetic constant from fluorescence recovery after photobleaching, and was significantly higher for cells grown on the microtopography than on the flat surface, P<0.05, n=7. To test the role of lipid signaling, PIP2 availability was reduced by neomycin (500 μM) scavenging, or the PIP2 level was increased by PI3K inhibition with wortmannin (1 μM). The localization of actin and lamellipodin (a PIP2 / actin binding protein) depended on stiffness and topography and was further changed by drugs. The actin cytoskeleton had more prominent stress fibers extending to lamella edge with neomycin treatment; the lamellipodin distribution is being assessed. Fibroblast migration is known to decelerate as the surface stiffens, and was measured here through a gap created by removal of a barrier or by the traditional scratch assay. Migration velocity at all stiffnesses was significantly faster with reduction of PIP2 availability with neomycin treatment compared to untreated fibroblasts (P<.05, n= 6). With wortmannin treatment, the velocity is similar to the untreated cells. We conclude that primary cardiac fibroblasts respond to stiffness and microtopography via lipid signaling and PIP2 to regulate cell migration and the architecture of the actin cytoskeleton. Therefore, the use of drugs or manipulation of the microenvironment might be beneficial for control of fibrosis in disease.Support or Funding InformationHL62426.
Phyllanthus taxodiifolius Beille Extract Disrupted Microtubule Dynamics in Glioma Cells
Authors: Kwanthongdee Jaturon , Saengsawang Witchuda , Patoomratana Tuchinda 
Published: November 2023

Glioma is the most common, life threatening, malignant primary brain tumor. This tumor has the ability to spread and invade through surrounding areas in the brain. Cell migration and adhesion are the most important processes involved in cancer cell spreading and invasion. Phyllanthus taxodiifolius Beille extract has been studied for its anti‐cancer action in breast and lung cancer cells. However, its mechanisms of action are still unclear. In addition, the effect of Phyllanthus taxodiifolius Beille extract on brain cancer cell viability and invasiveness, especially in glioma cells, has never been determined. Here we investigated the effect of Phyllanthus taxodiifolius Beille extract on glioma cell migration and adhesion using wound‐healing cell migration and cell spreading assays, respectively. We also determined its effects on microtubule structure and dynamics using both in vitro microtubule polymerization assay, and live cell imaging techniques with Total Internal Fluorescence Microscope (TIRFM). The results showed that Phyllanthus taxodiifolius Beille extract reduced glioma cell viability, disrupted cell adhesion and delayed cell migration. In addition, the extract rapidly disrupted microtubule structure and dynamics in glioma cells. Real time imaging of microtubule end‐binding protein (EB3) in glioma cells demonstrated that the extract disrupted the binding and dynamics of EB3 at the tips of microtubules. This study provided supporting evidences for the anti‐cancer properties of Phyllanthus taxodiifolius Beille extract. It is also the first study to demonstrate that the Phyllanthus taxodiifolius Beille extract interfered microtubule structure and dynamics, and suggested the possibility to further study and develop Phyllanthus taxodiifolius Beille derived compounds into a novel anti‐cancer drug targeting microtubules.Support or Funding InformationThis work was supported by a grant from the Center of Excellence on Environmental Health and Toxicology (EHT) to WS. We also gratefully acknowledge support from the Faculty of Science and Faculty of Graduate Studies, Mahidol University (to WS and JK) and support from Sritrangtong scholarship program (to JK).
ATP and extracellular calcium regulates differentially the phagocytosis of S. aureus and C. glabrata by macrophages.
Authors: Pérez Gabriela , Hernandez‐Silva Cesar , Arreola Jorge , Pérez‐Cornejo Patricia 
Published: November 2023

Phagocytosis is a fundamental function of innate immune cells; many receptors have been shown to mediate phagocytosis of bacterial and fungal pathogen. However, if the mechanism of phagocytosis is common among bacterial and fungal pathogens is poorly defined. We studied the effects of extracellular ATP and extracellular Ca2+ on uptake of Staphylococcus aureus and Candida glabrata in phagocytosis by J774 macrophages. Our findings show that phagocytosis of S. aureus was blocked by ATP and the P2X7 receptor agonist BzATP. A438079, an antagonist of P2X7 receptors, partially reverted the effects of ATP on bacterial phagocytosis. This data suggests that other P2X or P2Y receptors might be important for phagocytosis de S. aureus. In contrast, phagocytosis of C. glabrata remained unaffected by ATP stimulation. To determine if P2X‐mediated Ca2+ entry into macrophages was blocking the engulfment of bacteria, we measured phagocytic activity in the absence or presence of 2 mM extracellular Ca2+ with or without ATP. Ca2+, in the absence of ATP, was required for engulfment of C. glabrata but not S. aureus. Adding ATP inhibited phagocytosis of S. aureus regardless of Ca2+, suggesting that Ca2+ entry was not important for inhibiting phagocytosis. On the other hand, phagocytosis of normal or hyper‐adherent C. glabrata mutants had an absolute requirement for extracellular Ca2+ due to yeast adhesion to macrophages mediated by Ca2+‐dependent adhesion proteins. Addition of extracellular Ba2+ did not replace Ca2+, this indicate the high selectivity for Ca2+ in the phagocytic process of C. glabrata. We conclude that Ca2+ entry no role in phagocytosis of S. aureus; while the effect of Ca2+ on C. glabrata phagocytosis was mediated for calcium‐dependent adhesins.Support or Funding InformationThis work was supported by CONACYT (219949 joint grant to PPC and JA) and FAI (C16‐FAI‐09‐37.37 to GP)
Postpartum Cervical Remodeling Entails Acute Alteration in Expression of Mechanical‐ and Immune‐Related Factors
Authors: Stanley Robert Lee , Burton Catherine , Shah Sonam , Yang Eileen , Sampathkumar Prashanth , Rao Sanjana , Del Castillo Val , Douglas Veronica , Frank Martin , Mowa Chishimba Nathan 
Published: November 2023

Because the cervix undergoes marked mechanical trauma during birth and remains briefly open postpartum, it requires a speedy and complete tissue repair to avoid immediate and long term obstetrical complications, such as infection and possibly preterm labor, respectively. Here, we attempt to unravel the key biological changes of postpartum cervical repair in mice cervix during the first 48h using a proteome‐wide technology, proteomics, as well as real time PCR and confocal immunofluorescence. The proteomics data revealed variable expression of several intermediate filaments, cytoskeletal modulators, and proteins with immune‐ and/or wound healing properties. Real time PCR verified the proteomics and confocal immunofluorescence data of 13 molecules. We conclude that postpartum cervical repair involves expression of multiple factors, notably immune‐ and mechanical‐related factors that could potentially be used as markers for determining the phase of postpartum cervical repair in obstetrical complications, such as cervical lacerations.Support or Funding InformationAppalachian State University Office of Student Research