scholarly journals Application of ECIS to Assess FCCP-Induced Changes of MSC Micromotion and Wound Healing Migration

Sensors ◽  
2019 ◽  
Vol 19 (14) ◽  
pp. 3210 ◽  
Author(s):  
Sheng-Po Chiu ◽  
Yu-Wei Lee ◽  
Ling-Yi Wu ◽  
Tse-Hua Tung ◽  
Sofia Gomez ◽  
...  
Keyword(s):  
Time Series ◽  
Wound Healing ◽  
Cell Migration ◽  
Morphological Changes ◽  
Mitochondrial Dynamics ◽  
Human Mesenchymal Stem Cells ◽  
Atp Synthesis ◽  
Extracellular Flux ◽  
Sigmoid Curve ◽  
The Difference

Electric cell-substrate impedance sensing (ECIS) is an emerging technique for sensitively monitoring morphological changes of adherent cells in tissue culture. In this study, human mesenchymal stem cells (hMSCs) were exposed to different concentrations of carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone (FCCP) for 20 h and their subsequent concentration-dependent responses in micromotion and wound healing migration were measured by ECIS. FCCP disrupts ATP synthesis and results in a decrease in cell migration rates. To detect the change of cell micromotion in response to FCCP challenge, time-series resistances of cell-covered electrodes were monitored and the values of variance were calculated to verify the difference. While Seahorse XF-24 extracellular flux analyzer can detect the effect of FCCP at 3 μM concentration, the variance calculation of the time-series resistances measured at 4 kHz can detect the effect of FCCP at concentrations as low as 1 μM. For wound healing migration, the recovery resistance curves were fitted by sigmoid curve and the hill slope showed a concentration-dependent decline from 0.3 μM to 3 μM, indicating a decrease in cell migration rate. Moreover, dose dependent incline of the inflection points from 0.3 μM to 3 μM FCCP implied the increase of the half time for wound recovery migration. Together, our results demonstrate that partial uncoupling of mitochondrial oxidative phosphorylation reduces micromotion and wound healing migration of hMSCs. The ECIS method used in this study offers a simple and sensitive approach to investigate stem cell migration and its regulation by mitochondrial dynamics.

scholarly journals Chronic senescent human mesenchymal stem cells as possible contributor to the wound healing disorder after exposure to the alkylating agent sulfur mustard

2021 ◽  
Vol 95 (2) ◽  
pp. 727-747
Author(s):  
Simone Rothmiller ◽  
Niklas Jäger ◽  
Nicole Meier ◽  
Thimo Meyer ◽  
Adrian Neu ◽  
...  
Keyword(s):  
Stem Cells ◽  
Wound Healing ◽  
Mesenchymal Stem Cells ◽  
Alkylating Agent ◽  
Chronic Wounds ◽  
Sulfur Mustard ◽  
Morphological Changes ◽  
Human Mesenchymal Stem Cells ◽  
Age Related

AbstractWound healing is a complex process, and disturbance of even a single mechanism can result in chronic ulcers developing after exposure to the alkylating agent sulfur mustard (SM). A possible contributor may be SM-induced chronic senescent mesenchymal stem cells (MSCs), unable to fulfil their regenerative role, by persisting over long time periods and creating a proinflammatory microenvironment. Here we show that senescence induction in human bone marrow derived MSCs was time- and concentration-dependent, and chronic senescence could be verified 3 weeks after exposure to between 10 and 40 µM SM. Morphological changes, reduced clonogenic and migration potential, longer scratch closure times, differences in senescence, motility and DNA damage response associated genes as well as increased levels of proinflammatory cytokines were revealed. Selective removal of these cells by senolytic drugs, in which ABT-263 showed initial potential in vitro, opens the possibility for an innovative treatment strategy for chronic wounds, but also tumors and age-related diseases.

scholarly journals Detecting Effects of Low Levels of FCCP on Stem Cell Micromotion and Wound-Healing Migration by Time-Series Capacitance Measurement

Sensors ◽  
2021 ◽  
Vol 21 (9) ◽  
pp. 3017
Author(s):  
Si-Han Wang ◽  
Tse-Hua Tung ◽  
Sheng-Po Chiu ◽  
Hsin-Yi Chou ◽  
Yu-Han Hung ◽  
...  
Keyword(s):  
Time Series ◽  
Wound Healing ◽  
High Frequency ◽  
Human Mesenchymal Stem Cells ◽  
Capacitance Measurement ◽  
Cell Behavior ◽  
Impedance Sensing ◽  
Atp Production ◽  
Covered Electrodes ◽  
Dose Dependent Decrease

Electric cell–substrate impedance sensing (ECIS) has been used as a real-time impedance-based method to quantify cell behavior in tissue culture. The method is capable of measuring both the resistance and capacitance of a cell-covered microelectrode at various AC frequencies. In this study, we demonstrate the application of high-frequency capacitance measurement (f = 40 or 64 kHz) for the sensitive detection of both the micromotion and wound-healing migration of human mesenchymal stem cells (hMSCs). Impedance measurements of cell-covered electrodes upon the challenge of various concentrations of carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone (FCCP), from 0.1 to 30 μM, were conducted using ECIS. FCCP is an uncoupler of mitochondrial oxidative phosphorylation (OXPHOS), thereby reducing mitochondrial ATP production. By numerically analyzing the time-series capacitance data, a dose-dependent decrease in hMSC micromotion and wound-healing migration was observed, and the effect was significantly detected at levels as low as 0.1 μM. While most reported works with ECIS use the resistance/impedance time series, our results suggest the potential use of high-frequency capacitance time series for assessing migratory cell behavior such as micromotion and wound-healing migration.

Abstract 5379: Increased Myocardial Fatty Acid Uptake by Acyl-CoA Synthetase 1 Impairs Mitochondrial Function and Dynamics

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Heiko Bugger ◽  
Xiao Xuan Hu ◽  
Joseph Tuinei ◽  
Heather Theobald ◽  
William L Holland ◽  
...  
Keyword(s):  
Mitochondrial Dysfunction ◽  
Mitochondrial Membrane ◽  
Morphological Changes ◽  
Mitochondrial Dynamics ◽  
Mitochondrial Fission ◽  
Atp Synthesis ◽  
Fatty Acid Uptake ◽  
Membrane Lipid ◽  
Acid Uptake

Increased uptake and oxidation of fatty acids (FA) in diabetic hearts may contribute to mitochondrial dysfunction by promoting oxidative stress. To test this hypothesis, we investigated mice with cardiomyocyte-restricted overexpression of long-chain acyl-CoA synthetase 1 (MHC-ACS). In vivo PET studies revealed increased myocardial FA uptake (+120%; p<0.05) in MHC-ACS. Unexpectedly, FA oxidation and myocellular triglyceride content were unchanged, and PPARα-regulated FAO genes and PGC-1α-regulated OXPHOS genes were unaltered. However, cardiac content of the phospholipid precursors ceramide (+260%; p<0.05) and diacylglycerol (+40%; p<0.05) were increased. Mitochondrial cardiolipin content was decreased (−25%; p<0.05) and remodeled with substitution of 18:2 FA chains by unsaturated 22:6 FAs. Both ADP-stimulated mitochondrial O2 consumption (14.3±0.9 vs. 16.8±0.5 nmol/min/mgdw; p<0.05) and ATP synthesis (24.2±1.4 vs. 31.8±1.6 nmol/min/mgdw; p<0.05) were decreased in MHC-ACS in saponin-permeabilized cardiac fibers using palmitoyl-carnitine as a substrate. Mitochondrial superoxide production was increased by 75% (p<0.05). Electron microscopy in 3 to 24 week-old mice revealed increased mitochondrial number, which was highest at 3 weeks (3wk +210%, 12wk +120%, 24wk +130%; all p<0.05), and reduced mitochondrial size. Translocation of the mitochondrial fission protein dynamin-related protein 1 (Drp1) from cytosol to mitochondria was absent (p<0.05) in MHC-ACS, but not in controls. Mitochondrial membrane content of the fission protein fission 1 and the fusion proteins mitofusin 1 and 2 were unchanged. These morphological changes are consistent with increased mitochondrial fission and reduced Drp1 likely represents an adaptive response. Thus remodeling of the myocardial lipid pool and mitochondrial membrane lipid composition are associated with impaired mitochondrial dynamics and represents a novel mechanism for lipid-induced mitochondrial dysfunction in the heart.

scholarly journals Mitochondrial Dynamics in Placenta-Derived Mesenchymal Stem Cells Regulate the Invasion Activity of Trophoblast

2020 ◽  
Vol 21 (22) ◽  
pp. 8599 ◽  
Author(s):  
Jin Seok ◽  
Sujin Jun ◽  
Jung Ok Lee ◽  
Gi Jin Kim
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Mitochondrial Function ◽  
Mitochondrial Dynamics ◽  
Atp Synthesis ◽  
Trophoblast Invasion ◽  
Ubiquitin Protein Ligase ◽  
Extracellular Flux ◽  
Cellular Events ◽  
Cancerous Cells

Mitochondrial dynamics are involved in many cellular events, including the proliferation, differentiation, and invasion/migration of normal as well as cancerous cells. Human placenta-derived mesenchymal stem cells (PD-MSCs) were known to regulate the invasion activity of trophoblasts. However, the effects of PD-MSCs on mitochondrial function in trophoblasts are still insufficiently understood. Therefore, the objectives of this study are to analyze the factors related to mitochondrial function and investigate the correlation between trophoblast invasion and mitophagy via PD-MSC cocultivation. We assess invasion ability and mitochondrial function in invasive trophoblasts according to PD-MSC cocultivation by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and extracellular flux (XF) assay. Under PD-MSCs co-cultivation, invasion activity of a trophoblast is increased via activation of the Rho signaling pathway as well as Matrix metalloproteinases (MMPs). Additionally, the expression of mitochondrial function (e.g., reactive oxygen species (ROS), calcium, and adenosine triphosphate (ATP) synthesis) in trophoblasts are increased via PD-MSCs co-cultivation. Finally, PD-MSCs regulate mitochondrial autophagy factors in invasive trophoblasts via regulating the balance between PTEN-induced putative kinase 1 (PINK1) and parkin RBR E3 ubiquitin protein ligase (PARKIN) expression. Taken together, these results demonstrate that PD-MSCs enhance the invasion ability of trophoblasts via altering mitochondrial dynamics. These results support the fundamental mechanism of trophoblast invasion via mitochondrial function and provide a new stem cell therapy for infertility.

A Photoactivable Formaldehyde Donor with Fluorescence Monitoring Reveals Threshold to Arrest Cell Migration

2019 ◽  
Author(s):  
Lukas P Smaga ◽  
Nicholas W Pino ◽  
Gabriela E Ibarra ◽  
Vishnu Krishnamurthy ◽  
Jefferson Chan
Keyword(s):  
Wound Healing ◽  
Cell Migration ◽  
Fluorescence Enhancement ◽  
Biological Effects ◽  
Cellular Systems ◽  
Live Cells ◽  
First Report ◽  
Cell Lysate ◽  
Intracellular Release ◽  
Biological Analytes

Controlled light-mediated delivery of biological analytes enables the investigation of highly reactivity molecules within cellular systems. As many biological effects are concentration dependent, it is critical to determine the location, time, and quantity of analyte donation. In this work, we have developed the first photoactivatable donor for formaldehyde (FA). Our optimized photoactivatable donor, photoFAD-3, is equipped with a fluorescence readout that enables monitoring of FA release with a concomitant 139-fold fluorescence enhancement. Tuning of photostability and cellular retention enabled quantification of intracellular FA release through cell lysate calibration. Application of photoFAD-3 uncovered the concentration range necessary for arresting wound healing in live cells. This marks the first report where a photoactivatable donor for any analyte has been used to quantify intracellular release.

EVALUASI PROYEKSI DEMAND BARANG DAN PENUMPANG PELABUHAN DI PROVINSI MALUKU

2019 ◽  
Vol 19 (2) ◽  
pp. 101-110
Author(s):  
Adrian Firdaus ◽  
M. Dwi Yoga Sutanto ◽  
Rajin Sihombing ◽  
M. Weldy Hermawan
Keyword(s):  
Time Series ◽  
Projection Method ◽  
Development Plan ◽  
Master Plan ◽  
Port Development ◽  
Loading And Unloading ◽  
The Difference ◽  
Existing Data

Abstract Every port in Indonesia must have a Port Master Plan that contains an integrated port development plan. This study discusses one important aspect in the preparation of the Port Master Plan, namely the projected movement of goods and passengers, which can be used as a reference in determining the need for facilities at each stage of port development. The case study was conducted at a port located in a district in Maluku Province and aims to evaluate the analysis of projected demand for goods and passengers occurring at the port. The projection method used is time series and econometric projection. The projection results are then compared with the existing data in 2018. The results of this study show that the econometric projection gives adequate results in predicting loading and unloading activities as well as the number of passenger arrival and departure in 2018. This is indicated by the difference in the percentage of projection results towards the existing data, which is smaller than 10%. Whereas for loading and unloading activities, time series projections with logarithmic trends give better results than econometric projections. Keywords: port, port master plan, port development, unloading activities  Abstrak Setiap pelabuhan di Indonesia harus memiliki sebuah Rencana Induk Pelabuhan yang memuat rencana pengem-bangan pelabuhan secara terpadu. Studi ini membahas salah satu aspek penting dalam penyusunan Rencana Induk Pelabuhan, yaitu proyeksi pergerakan barang dan penumpang, yang dapat dipakai sebagai acuan dalam penentuan kebutuhan fasilitas di setiap tahap pengembangan pelabuhan. Studi kasus dilakukan pada sebuah pelabuhan yang terletak di sebuah kabupaten di Provinsi Maluku dan bertujuan untuk melakukan evaluasi ter-hadap analisis proyeksi demand barang dan penumpang yang terjadi di pelabuhan tersebut. Metode proyeksi yang dipakai adalah proyeksi deret waktu dan ekonometrik. Hasil proyeksi selanjutnya dibandingkan dengan data eksisting tahun 2018. Hasil studi ini menunjukkan bahwa proyeksi ekonometrik memberikan hasil yang cukup baik dalam memprediksi aktivitas bongkar barang serta jumlah penumpang naik dan turun di tahun 2018. Hal ini diindikasikan dengan selisih persentase hasil proyeksi terhadap data eksisting yang lebih kecil dari 10%. Sedangkan untuk aktivitas muat barang, proyeksi deret waktu dengan tren logaritmik memberikan hasil yang lebih baik daripada proyeksi ekonometrik. Kata-kata kunci: pelabuhan, rencana induk pelabuhan, pengembangan pelauhan, aktivitas bongkar barang

The Anti-tumor Activity and Mechanisms of rLj-RGD3 on Human Laryngeal Squamous Carcinoma Hep2 Cells

2020 ◽  
Vol 19 (17) ◽  
pp. 2108-2119
Author(s):  
Yang Jin ◽  
Li Lv ◽  
Shu-Xiang Ning ◽  
Ji-Hong Wang ◽  
Rong Xiao
Keyword(s):  
Wound Healing ◽  
Western Blot ◽  
Morphological Changes ◽  
In Vivo Studies ◽  
Migration And Invasion ◽  
Tunel Staining ◽  
Dna Ladder ◽  
Western Blot Assay

Background: Laryngeal Squamous Cell Carcinoma (LSCC) is a malignant epithelial tumor with poor prognosis and its incidence rate increased recently. rLj-RGD3, a recombinant protein cloned from the buccal gland of Lampetra japonica, contains three RGD motifs that could bind to integrins on the tumor cells. Methods: MTT assay was used to detect the inhibitory rate of viability. Giemsa’s staining assay was used to observe the morphological changes of cells. Hoechst 33258 and TUNEL staining assay, DNA ladder assay were used to examine the apoptotic. Western blot assay was applied to detect the change of the integrin signal pathway. Wound-healing assay, migration, and invasion assay were used to detect the mobility of Hep2 cells. H&E staining assay was used to show the arrangement of the Hep2 cells in the solid tumor tissues. Results: In the present study, rLj-RGD3 was shown to inhibit the viability of LSCC Hep2 cells in vitro by inducing apoptosis with an IC50 of 1.23µM. Western blot showed that the apoptosis of Hep2 cells induced by rLj- RGD3 was dependent on the integrin-FAK-Akt pathway. Wound healing, transwells, and western blot assays in vitro showed that rLj-RGD3 suppressed the migration and invasion of Hep2 cells by integrin-FAKpaxillin/ PLC pathway which could also affect the cytoskeleton arrangement in Hep2 cells. In in vivo studies, rLj-RGD3 inhibited the growth, tumor volume, and weight, as well as disturbed the tissue structure of the solid tumors in xenograft models of BALB/c nude mice without reducing their body weights. Conclusion: hese results suggested that rLj-RGD3 is an effective and safe suppressor on the growth and metastasis of LSCC Hep2 cells from both in vitro and in vivo experiments. rLj-RGD3 might be expected to become a novel anti-tumor drug to treat LSCC patients in the near future.

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