scholarly journals Heparin-Mimicking Polymer-Based In Vitro Platform Recapitulates In Vivo Muscle Atrophy Phenotypes

2021 ◽  
Vol 22 (5) ◽  
pp. 2488
Author(s):  
Hyunbum Kim ◽  
Ji Hoon Jeong ◽  
Mona Fendereski ◽  
Hyo-Shin Lee ◽  
Da Yeon Kang ◽  
...  
Keyword(s):  
Cell Culture ◽  
Wnt Signaling ◽  
Signaling Pathways ◽  
Muscle Atrophy ◽  
Skeletal Muscle Regeneration ◽  
In Vitro Cell Culture ◽  
Gene And Protein Expression ◽  
Cell Cell

The cell–cell/cell–matrix interactions between myoblasts and their extracellular microenvironment have been shown to play a crucial role in the regulation of in vitro myogenic differentiation and in vivo skeletal muscle regeneration. In this study, by harnessing the heparin-mimicking polymer, poly(sodium-4-styrenesulfonate) (PSS), which has a negatively charged surface, we engineered an in vitro cell culture platform for the purpose of recapitulating in vivo muscle atrophy-like phenotypes. Our initial findings showed that heparin-mimicking moieties inhibited the fusion of mononucleated myoblasts into multinucleated myotubes, as indicated by the decreased gene and protein expression levels of myogenic factors, myotube fusion-related markers, and focal adhesion kinase (FAK). We further elucidated the underlying molecular mechanism via transcriptome analyses, observing that the insulin/PI3K/mTOR and Wnt signaling pathways were significantly downregulated by heparin-mimicking moieties through the inhibition of FAK/Cav3. Taken together, the easy-to-adapt heparin-mimicking polymer-based in vitro cell culture platform could be an attractive platform for potential applications in drug screening, providing clear readouts of changes in insulin/PI3K/mTOR and Wnt signaling pathways.

scholarly journals Modulation of Calretinin Expression in Human Mesothelioma Cells Reveals the Implication of the FAK and Wnt Signaling Pathways in Conferring Chemoresistance towards Cisplatin

2019 ◽  
Vol 20 (21) ◽  
pp. 5391 ◽  
Author(s):  
Wörthmüller ◽  
Salicio ◽  
Oberson ◽  
Blum ◽  
Schwaller
Keyword(s):  
Wnt Signaling ◽  
Signaling Pathways ◽  
Signaling Pathway ◽  
Expression System ◽  
Single Agent ◽  
Wnt Signaling Pathway ◽  
Tumor Formation ◽  
Mesenchymal Transition

Malignant mesothelioma (MM) is an aggressive asbestos-linked neoplasm, characterized by dysregulation of signaling pathways. Due to intrinsic or acquired chemoresistance, MM treatment options remain limited. Calretinin is a Ca2+-binding protein expressed during MM tumorigenesis that activates the FAK signaling pathway, promoting invasion and epithelial-to-mesenchymal transition. Constitutive calretinin downregulation decreases MM cells’ growth and survival, and impairs tumor formation in vivo. In order to evaluate early molecular events occurring during calretinin downregulation, we generated a tightly controlled IPTG-inducible expression system to modulate calretinin levels in vitro. Calretinin downregulation significantly reduced viability and proliferation of MM cells, attenuated FAK signaling and reduced the invasive phenotype of surviving cells. Importantly, surviving cells showed a higher resistance to cisplatin due to increased Wnt signaling. This resistance was abrogated by the Wnt signaling pathway inhibitor 3289-8625. In various MM cell lines and regardless of calretinin expression levels, blocking of FAK signaling activated the Wnt signaling pathway and vice versa. Thus, blocking both pathways had the strongest impact on MM cell proliferation and survival. Chemoresistance mechanisms in MM cells have resulted in a failure of single-agent therapies. Targeting of multiple components of key signaling pathways, including Wnt signaling, might be the future method-of-choice to treat MM.

scholarly journals Determination of the Infectivity of Cryopreserved Theileria annu-lata Sporozoites in Tick Derived Stabilates Iran Ak-93 Strain, by In Vivo and In Vitro Methods

2019 ◽  
Author(s):  
Hossein MODIRROUSTA ◽  
Gholamreza HABIBI ◽  
Parviz SHAYAN ◽  
Asghar AFSHARI ◽  
Ali MIRJALILI ◽  
...  
Keyword(s):  
Cell Culture ◽  
Infected Cell ◽  
Cell Line ◽  
Challenge Test ◽  
Infected Cell Line ◽  
Rt Pcr ◽  
In Vitro Cell Culture ◽  
Microscopic Inspection

Background: The protozoan parasite Theileria annulata is the causative agent of tropical theileriosis in cattle. Vaccination is recommended by administration of attenuated schizont-infected cell lines. The expected protective immunity post-vaccination can be demonstrated by challenge test through inoculation of highly virulent infective sporozoites. The aim of this study was to produce Hyalomma anatolicum anatolicum tick infected with T. annulata (local strain) for preparation of tick-derived sporozoite stabilates for molecular characterization and infectivity test assay. Methods: A local T. annulata strain was used for experimental infection of calves. A field isolate of H. a. anatolicum was isolated, laboratory-reared and infected by blood-feeding on Theileria infected above-mentioned calves. The infectivity of calf, tick and prepared stabilate were confirmed by clinical signs of theileriosis, microscopic inspection, RT-PCR and in vitro cell culture. Results: The tick stabilate was prepared and cryopreserved in liquid nitrogen. The infectivity of the tick stabilate was verified by in vivo bioassay, in vitro cell culture infection, microscopic inspection in salivary glands and RT-PCR assay. The in vitro produced cell line in this study was characterized by T. annulata Cytochrome b gene analyzing. Conclusion: The infectivity of a new prepared tick-derived sporozoite stabilate was confirmed in susceptible calves; by microscopically, post mortem, tick microscopic and molecular assays. Moreover, naïve PBMCs were transformed and proliferated by T. annulata infected tick stabilate to immortal T. annulata schizont infected cell line. The potent infective sporozoite tick derived stabilate could be used for vaccine efficacy and challenge test as well as in vaccine development.

scholarly journals Garcinol Regulates EMT and Wnt Signaling Pathways In Vitro and In Vivo, Leading to Anticancer Activity against Breast Cancer Cells

2012 ◽  
Vol 11 (10) ◽  
pp. 2193-2201 ◽  
Author(s):  
Aamir Ahmad ◽  
Sanila H. Sarkar ◽  
Bassam Bitar ◽  
Shadan Ali ◽  
Amro Aboukameel ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Wnt Signaling ◽  
Anticancer Activity ◽  
Cancer Cells ◽  
Signaling Pathways ◽  
Breast Cancer Cells

scholarly journals The Extracellular Domain of Lrp5/6 Inhibits Noncanonical Wnt Signaling In Vivo

2009 ◽  
Vol 20 (3) ◽  
pp. 924-936 ◽  
Author(s):  
Vitezslav Bryja ◽  
Emma R. Andersson ◽  
Alexandra Schambony ◽  
Milan Esner ◽  
Lenka Bryjová ◽  
...  
Keyword(s):  
Wnt Signaling ◽  
Signaling Pathways ◽  
Extracellular Domain ◽  
Convergent Extension ◽  
Mouse Development ◽  
Downstream Target ◽  
Extracellular Domains ◽  
Antisense Morpholino

Lrp5/6 are crucial coreceptors for Wnt/β-catenin signaling, a pathway biochemically distinct from noncanonical Wnt signaling pathways. Here, we examined the possible participation of Lrp5/6 in noncanonical Wnt signaling. We found that Lrp6 physically interacts with Wnt5a, but that this does not lead to phosphorylation of Lrp6 or activation of the Wnt/β-catenin pathway. Overexpression of Lrp6 blocks activation of the Wnt5a downstream target Rac1, and this effect is dependent on intact Lrp6 extracellular domains. These results suggested that the extracellular domain of Lrp6 inhibits noncanonical Wnt signaling in vitro. In vivo, Lrp6−/− mice exhibited exencephaly and a heart phenotype. Surprisingly, these defects were rescued by deletion of Wnt5a, indicating that the phenotypes resulted from noncanonical Wnt gain-of-function. Similarly, Lrp5 and Lrp6 antisense morpholino-treated Xenopus embryos exhibited convergent extension and heart phenotypes that were rescued by knockdown of noncanonical XWnt5a and XWnt11. Thus, we provide evidence that the extracellular domains of Lrp5/6 behave as physiologically relevant inhibitors of noncanonical Wnt signaling during Xenopus and mouse development in vivo.

scholarly journals Accumulation of Tc-99m HL91 in Tumor Hypoxia: In Vitro Cell Culture and In Vivo Tumor Model

2008 ◽  
Vol 24 (9) ◽  
pp. 461-472 ◽  
Author(s):  
Bi-Fang Lee ◽  
Nan-Tsing Chiu ◽  
Chien-Chung Hsia ◽  
Lie-Hang Shen
Keyword(s):  
Cell Culture ◽  
Tumor Hypoxia ◽  
Tumor Model ◽  
In Vitro Cell Culture

Partner telomeres during anaphase in crane-fly spermatocytes are connected by an elastic tether that exerts a backward force and resists poleward motion

2002 ◽  
Vol 115 (7) ◽  
pp. 1541-1549 ◽  
Author(s):  
James R. LaFountain ◽  
Richard W. Cole ◽  
Conly L. Rieder
Keyword(s):  
Cell Culture ◽  
Strong Evidence ◽  
Laser Microbeam ◽  
In Vitro Cell Culture ◽  
Pulling Forces ◽  
As If

As chromosomes move polewards during anaphase in crane-fly spermatocytes,trailing arms commonly stretch backwards for a brief time, as if tethered to their partners. To test that notion, a laser microbeam was used to sever trailing arms and thereby release telomere-containing arm segments (called acentric fragments because they lack kinetochores) from segregating chromosomes. Analysis of the movement of acentric fragments after their release provided clear evidence that previously conjoined partners were indeed tethered at their telomeres and that tethers exerted backward forces that were sufficient to move the fragment across the equator and into the opposite half-spindle. To address concerns that tethers might be artifacts of in vitro cell culture, spermatocytes were fixed in situ, and stretched arms within fixed cells provided strong evidence for tethers in vivo. The substantial resistance that tethers impose on the poleward movement of chromosomes must normally be over-ridden by the poleward `pulling' forces exerted at kinetochores. In spermatocytes, poleward forces are supplied primarily by the`traction fibers' that are firmly attached to kinetochores through end-on attachments to the plus ends of kinetochore microtubules.

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