scholarly journals Chinese Veterinary Medicine B307 Promotes Cardiac Performance and Skeletal Muscle Contraction via Enhancing Intracellular Calcium Levels and Neural Electrical Activity in Animal and Cell Models

2020 ◽  
Vol 2020 ◽  
pp. 1-10
Author(s):  
Chia-Ying Lien ◽  
Chen-Wen Lu ◽  
Chih-Hsiang Hsu ◽  
Tai-Yuan Chuang ◽  
Li-Yu Su ◽  
...  
Keyword(s):  
Veterinary Medicine ◽  
Electrical Activity ◽  
Intracellular Calcium ◽  
Cell Model ◽  
Color Doppler ◽  
Neuroblastoma Cell ◽  
Motor Functions ◽  
Electrical Activities

The study mainly investigated the effects of Chinese veterinary medicine B307 in cardiac and motor functions in animal models of pigeons and mice. Related cellular mechanisms were also studied in the neuroblastoma cell model of SH-SY5Y. Cardiac functions of pigeons and mice were examined by using moorFLPI Laser color Doppler imager and M-mode echocardiography, and motor functions were examined by using muscle electrical stimulation and force recording in the isolated breast muscle. Intracellular calcium levels and electrical activity of SH-SY5Y cells were examined by using Fura 2-AM fluorescence and MED64 system separately. Our results in vivo found that those pigeons under oral B307 treatment obviously enhanced subcutaneous microcirculation and contractile force and prolonged fatigue time in their breast muscles. Those mice under oral B307 treatment obviously elevated ejection fraction and cardiac output in their hearts. Our results in vitro showed that those SH-SY5Y cells under B307 treatment obviously increased intracellular calcium mobilization and electrical activities. These results revealed that improvement of cardiac and motor functions under B307 treatments may be caused by increasing electrical activities and intracellular calcium levels in neuromuscular cells and a similar mechanism may also occur in muscle cells. Thus, we suggested that B307 can be a functional Chinese veterinary medicine for flying pigeons.

scholarly journals Preclinical In Vitro Studies with 3D Spheroids to Evaluate Cu(DDC)2 Containing Liposomes for the Treatment of Neuroblastoma

Pharmaceutics ◽  
2021 ◽  
Vol 13 (6) ◽  
pp. 894
Author(s):  
Friederike Hartwig ◽  
Monika Köll-Weber ◽  
Regine Süss
Keyword(s):  
Cell Cultures ◽  
Colloidal Stability ◽  
Cell Model ◽  
Neuroblastoma Cell ◽  
Anticancer Therapy ◽  
In Vitro Studies ◽  
Molar Ratio ◽  
Anticancer Activities

Preclinical in vitro studies of drug candidates for anticancer therapy are generally conducted on well-established 2D cell models. Unfortunately, these models are unable to mimic the properties of in vivo tumors. However, in vitro 3D models (spheroids) have been proven to be superior in reflecting the tumor microenvironment. Diethyldithiocarbamate (DDC−) is the active metabolite of Disulfiram, an approved drug for alcoholism and repurposed for cancer treatment. DDC− binds copper in a molar ratio of 2:1 resulting in a water-insoluble Cu(DDC)2 complex exhibiting anticancer activities. Delivery of the Cu(DDC)2 complex using nanoparticulate carriers provides decisive advantages for a parental application. In this study, an injectable liposomal Cu(DDC)2 formulation was developed and the toxicity was compared with a 2D neuroblastoma and a 3D neuroblastoma cell model. Our results indicate that Cu(DDC)2 liposomes complied with the size requirements of nanoparticles for intravenous injection and demonstrated high drug to lipid ratios as well as colloidal stability upon storage. Furthermore, an efficient cytotoxic effect on neuroblastoma 2D cell cultures and a very promising and even more pronounced effect on 3D cell cultures in terms of neuroblastoma monoculture and neuroblastoma co-culture with primary cell lines was proven, highly encouraging the use of Cu(DDC)2 liposomes for anticancer therapy.

scholarly journals A New Cell Model for Investigating Prion Strain Selection and Adaptation

Viruses ◽  
2019 ◽  
Vol 11 (10) ◽  
pp. 888
Author(s):  
Alexandra Philiastides ◽  
Juan Manuel Ribes ◽  
Daniel Chun-Mun Yip ◽  
Christian Schmidt ◽  
Iryna Benilova ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Cell Model ◽  
Prion Diseases ◽  
Neuroblastoma Cell ◽  
Neuroblastoma Cell Line ◽  
Strain Selection ◽  
Prion Strain ◽  
Prion Strains

Prion diseases are fatal neurodegenerative diseases that affect humans and animals. Prion strains, conformational variants of misfolded prion proteins, are associated with distinct clinical and pathological phenotypes. Host-strain interactions result in the selective damage of distinct brain areas and they are responsible for strain selection and/or adaptation, but the underlying molecular mechanisms are unknown. Prion strains can be distinguished by their cell tropism in vivo and in vitro, which suggests that susceptibility to distinct prion strains is determined by cellular factors. The neuroblastoma cell line PK1 is refractory to the prion strain Me7, but highly susceptible to RML. We challenged a large number of clonal PK1 lines with Me7 and successfully selected highly Me7-susceptible subclones (PME) to investigate whether the prion strain repertoire of PK1 can be expanded. Notably, the Me7-infected PME clones were more protease-resistant when compared to RML-infected PME clones, which suggested that cell-adapted Me7 and RML are distinct prion strains. Strikingly, Me7-refractory cells, including PK1 and astrocytes in cortico-hippocampal cultures, are highly susceptible to prions, being derived from homogenates of Me7-infected PME cells, suggesting that the passage of Me7 in PME cells leads to an extended host range. Thus, PME clones represent a compelling cell model for strain selection and adaptation.

Endogenous hydrogen sulfide sulfhydrates IKKβ at cysteine 179 to control pulmonary artery endothelial cell inflammation

2019 ◽  
Vol 133 (20) ◽  
pp. 2045-2059 ◽  
Author(s):  
Da Zhang ◽  
Xiuli Wang ◽  
Siyao Chen ◽  
Selena Chen ◽  
Wen Yu ◽  
...  
Keyword(s):  
Hydrogen Sulfide ◽  
Endothelial Cell ◽  
Pulmonary Artery ◽  
Cell Model ◽  
Site Directed Mutagenesis ◽  
Critical Event ◽  
Hypertensive Rats ◽  
Pulmonary Artery Endothelial Cell

Abstract Background: Pulmonary artery endothelial cell (PAEC) inflammation is a critical event in the development of pulmonary arterial hypertension (PAH). However, the pathogenesis of PAEC inflammation remains unclear. Methods: Purified recombinant human inhibitor of κB kinase subunit β (IKKβ) protein, human PAECs and monocrotaline-induced pulmonary hypertensive rats were employed in the study. Site-directed mutagenesis, gene knockdown or overexpression were conducted to manipulate the expression or activity of a target protein. Results: We showed that hydrogen sulfide (H2S) inhibited IKKβ activation in the cell model of human PAEC inflammation induced by monocrotaline pyrrole-stimulation or knockdown of cystathionine γ-lyase (CSE), an H2S generating enzyme. Mechanistically, H2S was proved to inhibit IKKβ activity directly via sulfhydrating IKKβ at cysteinyl residue 179 (C179) in purified recombinant IKKβ protein in vitro, whereas thiol reductant dithiothreitol (DTT) reversed H2S-induced IKKβ inactivation. Furthermore, to demonstrate the significance of IKKβ sulfhydration by H2S in the development of PAEC inflammation, we mutated C179 to serine (C179S) in IKKβ. In purified IKKβ protein, C179S mutation of IKKβ abolished H2S-induced IKKβ sulfhydration and the subsequent IKKβ inactivation. In human PAECs, C179S mutation of IKKβ blocked H2S-inhibited IKKβ activation and PAEC inflammatory response. In pulmonary hypertensive rats, C179S mutation of IKKβ abolished the inhibitory effect of H2S on IKKβ activation and pulmonary vascular inflammation and remodeling. Conclusion: Collectively, our in vivo and in vitro findings demonstrated, for the first time, that endogenous H2S directly inactivated IKKβ via sulfhydrating IKKβ at Cys179 to inhibit nuclear factor-κB (NF-κB) pathway activation and thereby control PAEC inflammation in PAH.

scholarly journals Localization matters: nuclear-trapped Survivin sensitizes glioblastoma cells to temozolomide by elevating cellular senescence and impairing homologous recombination

2021 ◽  
Author(s):  
Thomas R. Reich ◽  
Christian Schwarzenbach ◽  
Juliana Brandstetter Vilar ◽  
Sven Unger ◽  
Fabian Mühlhäusler ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Nuclear Export ◽  
Cell Model ◽  
Chromosome Instability ◽  
Direct Interaction ◽  
High Grade Glioma ◽  
Strand Break ◽  
Γh2ax Foci

AbstractTo clarify whether differential compartmentalization of Survivin impacts temozolomide (TMZ)-triggered end points, we established a well-defined glioblastoma cell model in vitro (LN229 and A172) and in vivo, distinguishing between its nuclear and cytoplasmic localization. Expression of nuclear export sequence (NES)-mutated Survivin (SurvNESmut-GFP) led to impaired colony formation upon TMZ. This was not due to enhanced cell death but rather due to increased senescence. Nuclear-trapped Survivin reduced homologous recombination (HR)-mediated double-strand break (DSB) repair, as evaluated by γH2AX foci formation and qPCR-based HR assay leading to pronounced induction of chromosome aberrations. Opposite, clones, expressing free-shuttling cytoplasmic but not nuclear-trapped Survivin, could repair TMZ-induced DSBs and evaded senescence. Mass spectrometry-based interactomics revealed, however, no direct interaction of Survivin with any of the repair factors. The improved TMZ-triggered HR activity in Surv-GFP was associated with enhanced mRNA and stabilized RAD51 protein expression, opposite to diminished RAD51 expression in SurvNESmut cells. Notably, cytoplasmic Survivin could significantly compensate for the viability under RAD51 knockdown. Differential Survivin localization also resulted in distinctive TMZ-triggered transcriptional pathways, associated with senescence and chromosome instability as shown by global transcriptome analysis. Orthotopic LN229 xenografts, expressing SurvNESmut exhibited diminished growth and increased DNA damage upon TMZ, as manifested by PCNA and γH2AX foci expression, respectively, in brain tissue sections. Consequently, those mice lived longer. Although tumors of high-grade glioma patients expressed majorly nuclear Survivin, they exhibited rarely NES mutations which did not correlate with survival. Based on our in vitro and xenograft data, Survivin nuclear trapping would facilitate glioma response to TMZ.

scholarly journals β-elemene alleviates airway stenosis via the ILK/Akt pathway modulated by MIR143HG sponging miR-1275

2021 ◽  
Vol 26 (1) ◽  
Author(s):  
Guoying Zhang ◽  
Cheng Xue ◽  
Yiming Zeng
Keyword(s):  
Cell Cycle ◽  
Cell Viability ◽  
Cell Model ◽  
Protective Efficacy ◽  
Rabbit Model ◽  
Akt Pathway ◽  
Loss Of Function ◽  
Airway Stenosis

Abstract Background We have previously found that β-elemene could inhibit the viability of airway granulation fibroblasts and prevent airway hyperplastic stenosis. This study aimed to elucidate the underlying mechanism and protective efficacy of β-elemene in vitro and in vivo. Methods Microarray and bioinformatic analysis were used to identify altered pathways related to cell viability in a β-elemene-treated primary cell model and to construct a β-elemene-altered ceRNA network modulating the target pathway. Loss of function and gain of function approaches were performed to examine the role of the ceRNA axis in β-elemene's regulation of the target pathway and cell viability. Additionally, in a β-elemene-treated rabbit model of airway stenosis, endoscopic and histological examinations were used to evaluate its therapeutic efficacy and further verify its mechanism of action. Results The hyperactive ILK/Akt pathway and dysregulated LncRNA-MIR143HG, which acted as a miR-1275 ceRNA to modulate ILK expression, were suppressed in β-elemene-treated airway granulation fibroblasts; β-elemene suppressed the ILK/Akt pathway via the MIR143HG/miR-1275/ILK axis. Additionally, the cell cycle and apoptotic phenotypes of granulation fibroblasts were altered, consistent with ILK/Akt pathway activity. In vivo application of β-elemene attenuated airway granulation hyperplasia and alleviated scar stricture, and histological detections suggested that β-elemene's effects on the MIR143HG/miR-1275/ILK axis and ILK/Akt pathway were in line with in vitro findings. Conclusions MIR143HG and ILK may act as ceRNA to sponge miR-1275. The MIR143HG/miR-1275/ILK axis mediates β-elemene-induced cell cycle arrest and apoptosis of airway granulation fibroblasts by modulating the ILK/Akt pathway, thereby inhibiting airway granulation proliferation and ultimately alleviating airway stenosis.

scholarly journals Generation of tetracycline-controllable CYP3A4-expressing Caco-2 cells by the piggyBac transposon system

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Moe Ichikawa ◽  
Hiroki Akamine ◽  
Michika Murata ◽  
Sumito Ito ◽  
Kazuo Takayama ◽  
...  
Keyword(s):  
Small Intestine ◽  
Drug Absorption ◽  
Cell Model ◽  
Negative Effects ◽  
Piggybac Transposon ◽  
Drug Metabolizing Enzyme ◽  
Metabolizing Enzyme ◽  
Cyp3a4 Expression

AbstractCaco-2 cells are widely used as an in vitro intestinal epithelial cell model because they can form a monolayer and predict drug absorption with high accuracy. However, Caco-2 cells hardly express cytochrome P450 (CYP), a drug-metabolizing enzyme. It is known that CYP3A4 is the dominant drug-metabolizing enzyme in human small intestine. In this study, we generated CYP3A4-expressing Caco-2 (CYP3A4-Caco-2) cells and attempted to establish a model that can simultaneously evaluate drug absorption and metabolism. CYP3A4-Caco-2 cells were generated by piggyBac transposon vectors. A tetracycline-controllable CYP3A4 expression cassette (tet-on system) was stably transduced into Caco-2 cells, thus regulating the levels of CYP3A4 expression depending on the doxycycline concentration. The CYP3A4 expression levels in CYP3A4-Caco-2 cells cultured in the presence of doxycycline were similar to or higher than those of adult small intestine. The CYP3A4-Caco-2 cells had enough ability to metabolize midazolam, a substrate of CYP3A4. CYP3A4 overexpression had no negative effects on cell proliferation, barrier function, and P-glycoprotein activity in Caco-2 cells. Thus, we succeeded in establishing Caco-2 cells with CYP3A4 metabolizing activity comparable to in vivo human intestinal tissue. This cell line would be useful in pharmaceutical studies as a model that can simultaneously evaluate drug absorption and metabolism.

scholarly journals PapRIV, a BV-2 microglial cell activating quorum sensing peptide

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yorick Janssens ◽  
Nathan Debunne ◽  
Anton De Spiegeleer ◽  
Evelien Wynendaele ◽  
Marta Planas ◽  
...  
Keyword(s):  
Quorum Sensing ◽  
Cell Model ◽  
Dependent Manner ◽  
Communication Signals ◽  
Gram Positive Bacteria ◽  
A Cell ◽  
Gastro Intestinal Tract

AbstractQuorum sensing peptides (QSPs) are bacterial peptides produced by Gram-positive bacteria to communicate with their peers in a cell-density dependent manner. These peptides do not only act as interbacterial communication signals, but can also have effects on the host. Compelling evidence demonstrates the presence of a gut-brain axis and more specifically, the role of the gut microbiota in microglial functioning. The aim of this study is to investigate microglial activating properties of a selected QSP (PapRIV) which is produced by Bacillus cereus species. PapRIV showed in vitro activating properties of BV-2 microglia cells and was able to cross the in vitro Caco-2 cell model and reach the brain. In vivo peptide presence was also demonstrated in mouse plasma. The peptide caused induction of IL-6, TNFα and ROS expression and increased the fraction of ameboid BV-2 microglia cells in an NF-κB dependent manner. Different metabolites were identified in serum, of which the main metabolite still remained active. PapRIV is thus able to cross the gastro-intestinal tract and the blood–brain barrier and shows in vitro activating properties in BV-2 microglia cells, hereby indicating a potential role of this quorum sensing peptide in gut-brain interaction.

scholarly journals Validation of an Engineered Cell Model for In Vitro and In Vivo HIF-1α Evaluation by Different Imaging Modalities

2013 ◽  
Vol 16 (2) ◽  
pp. 210-223 ◽  
Author(s):  
A. Lo Dico ◽  
S. Valtorta ◽  
C. Martelli ◽  
S. Belloli ◽  
U. Gianelli ◽  
...  
Keyword(s):  
Cell Model ◽  
Imaging Modalities

scholarly journals An optimised GAS-pharyngeal cell biofilm model

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Heema K. N. Vyas ◽  
Jason D. McArthur ◽  
Martina L. Sanderson-Smith
Keyword(s):  
Crystal Violet ◽  
Biofilm Formation ◽  
Cell Model ◽  
Matrix Material ◽  
Three Dimensional ◽  
Growth Period ◽  
Optimal Growth ◽  
Abiotic Surfaces

AbstractGroup A Streptococcus (GAS) causes 700 million infections and accounts for half a million deaths per year. Biofilm formation has been implicated in both pharyngeal and dermal GAS infections. In vitro, plate-based assays have shown that several GAS M-types form biofilms, and multiple GAS virulence factors have been linked to biofilm formation. Although the contributions of these plate-based studies have been valuable, most have failed to mimic the host environment, with many studies utilising abiotic surfaces. GAS is a human specific pathogen, and colonisation and subsequent biofilm formation is likely facilitated by distinct interactions with host tissue surfaces. As such, a host cell-GAS model has been optimised to support and grow GAS biofilms of a variety of GAS M-types. Improvements and adjustments to the crystal violet biofilm biomass assay have also been tailored to reproducibly detect delicate GAS biofilms. We propose 72 h as an optimal growth period for yielding detectable biofilm biomass. GAS biofilms formed are robust and durable, and can be reproducibly assessed via staining/washing intensive assays such as crystal violet with the aid of methanol fixation prior to staining. Lastly, SEM imaging of GAS biofilms formed by this model revealed GAS cocci chains arranged into three-dimensional aggregated structures with EPS matrix material. Taken together, we outline an efficacious GAS biofilm pharyngeal cell model that can support long-term GAS biofilm formation, with biofilms formed closely resembling those seen in vivo.

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