scholarly journals An integrated systems biology approach to understanding the rules of keratinocyte colony formation

2007 ◽  
Vol 4 (17) ◽  
pp. 1077-1092 ◽  
Author(s):  
Tao Sun ◽  
Phil McMinn ◽  
Simon Coakley ◽  
Mike Holcombe ◽  
Rod Smallwood ◽  
...  
Keyword(s):  
Human Keratinocytes ◽  
Self Organization ◽  
Hacat Cells ◽  
Agent Based ◽  
Wound Model ◽  
Cell Substrate ◽  
Scratch Wound ◽  
Keratinocyte Cell Line Hacat ◽  
Keratinocyte Cell

Closely coupled in vitro and in virtuo models have been used to explore the self-organization of normal human keratinocytes (NHK). Although it can be observed experimentally, we lack the tools to explore many biological rules that govern NHK self-organization. An agent-based computational model was developed, based on rules derived from literature, which predicts the dynamic multicellular morphogenesis of NHK and of a keratinocyte cell line (HaCat cells) under varying extracellular Ca ++ concentrations. The model enables in virtuo exploration of the relative importance of biological rules and was used to test hypotheses in virtuo which were subsequently examined in vitro . Results indicated that cell–cell and cell–substrate adhesions were critically important to NHK self-organization. In contrast, cell cycle length and the number of divisions that transit-amplifying cells could undergo proved non-critical to the final organization. Two further hypotheses, to explain the growth behaviour of HaCat cells, were explored in virtuo —an inability to differentiate and a differing sensitivity to extracellular calcium. In vitro experimentation provided some support for both hypotheses. For NHKs, the prediction was made that the position of stem cells would influence the pattern of cell migration post-wounding. This was then confirmed experimentally using a scratch wound model.

scholarly journals Two new sphingomyelin analogues inhibit phosphatidylcholine biosynthesis by decreasing membrane-bound CTP: phosphocholine cytidylyltransferase levels in HaCaT cells

1995 ◽  
Vol 311 (3) ◽  
pp. 873-879 ◽  
Author(s):  
T Wieder ◽  
C Perlitz ◽  
M Wieprecht ◽  
R T C Huang ◽  
C C Geilen ◽  
...  
Keyword(s):  
High Ratio ◽  
Head Group ◽  
Hacat Cells ◽  
Phosphocholine Cytidylyltransferase ◽  
Ctp:Phosphocholine Cytidylyltransferase ◽  
Phosphatidylcholine Biosynthesis ◽  
Keratinocyte Cell Line Hacat ◽  
Keratinocyte Cell ◽  
Membrane Bound

The effects of two newly synthesized sphingomyelin analogues on phosphatidylcholine biosynthesis were investigated in the immortalized human keratinocyte cell line HaCaT. N-Acetyl-erythro-sphingosine-1-phosphocholine (AcSM) and N-octanoyl-erythro-sphingosine-1-phosphocholine (OcSM) inhibited the incorporation of choline into phosphatidylcholine with half-inhibitory concentrations (IC50) of 6 micrograms/ml and 10 micrograms/ml respectively. Further experiments revealed that AcSM and OcSM interfered with the translocation of the rate-limiting enzyme of phosphatidylcholine biosynthesis, CTP:phosphocholine cytidylyltransferase (EC 2.7.7.15), in HaCaT cells and inhibited cytidylyltransferase activity in vitro. Despite the fact that OcSM was a potent inhibitor of cytidylyltransferase in vitro, its effects on phosphatidylcholine biosynthesis and translocation of cytidylyltransferase in HaCaT cells were less pronounced as compared with AcSM. Finally, we showed that the comparatively strong effects of AcSM in cell culture experiments were due to the uptake of large amounts of this sphingomyelin analogue into the cells. The results presented demonstrate that the activity of cytidylyltransferase may be negatively regulated by a high ratio of choline head group-containing sphingolipids.

scholarly journals 5-fluorouracil Toxicity Mechanism Determination in Human Keratinocytes: in vitro Study on HaCaT Cell Line

2017 ◽  
Vol 118 (4) ◽  
pp. 128-138
Author(s):  
Jan Hartinger ◽  
Pavel Veselý ◽  
Martin Šíma ◽  
Irena Netíková ◽  
Eva Matoušková ◽  
...  
Keyword(s):  
Cell Line ◽  
In Vitro Study ◽  
Human Keratinocytes ◽  
Cell Types ◽  
Toxicity Mechanism ◽  
Keratinocyte Cell Line Hacat ◽  
Keratinocyte Cell ◽  
First Time ◽  
Fluorouracil Toxicity

5-fluorouracil (5-FU) and capecitabine therapy is often accompanied by palmar-plantar erythrodysesthesia (PPE) which is manifestation of 5-FU toxicity in keratinocytes. The main mechanisms of 5-FU action are thymidylate synthase (TS) inhibition which can be abrogated by thymidine and strengthened by calciumfolinate (CF) and incorporation of fluorouridinetriphosphate into RNA which can be abrogated by uridine. For proper PPE treatment 5-FU mechanism of action in keratinocytes needs to be elucidated. We used the 5-FU toxicity modulators uridine, thymidine and CF to discover the mechanism of 5-FU action in human keratinocyte cell line HaCaT. To measure the cellular viability, we used MTT test and RTCA test. CF did not augment 5-FU toxicity and 5-FU toxicity was weakened by uridine. Therefore, the primary mechanism of 5-FU toxicity in keratinocytes is 5-FU incorporation into RNA. The uridine protective effect cannot fully develop in the presence of CF. Thymidine addition to 5-FU and uridine treated cells not only prevents the toxicity-augmenting CF effect but it also prolongs the 5-FU treated cells survival in comparison to uridine only. Therefore, it can be assumed that in the presence of uridine the 5-FU toxicity mechanism is switched from RNA incorporation to TS inhibition. Although particular 5-FU toxicity mechanisms were previously described in various cell types, this is the first time when various combinations of pyrimidine nucleosides and CF were used for 5-FU toxicity mechanism elucidation in human keratinocytes. We suggest that for PPE treatment ointment containing uridine and thymidine should be further clinically tested.

Calcipotriol inhibits proliferation of human keratinocytes by downregulating STAT1 and STAT3 signaling

2016 ◽  
Vol 65 (2) ◽  
pp. 376-381 ◽  
Author(s):  
Wenli Liang ◽  
Zigang Lin ◽  
Li Zhang ◽  
Xuan Qin ◽  
Yuan Zhang ◽  
...  
Keyword(s):  
Messenger Rna ◽  
Human Keratinocytes ◽  
Cytokine Signaling ◽  
Hacat Cells ◽  
Molecular Signaling ◽  
Beneficial Effects ◽  
Protein Levels ◽  
Stat3 Signaling ◽  
Keratinocyte Cell Line Hacat

Psoriasis is an autoimmune disease, which is characterized by aberrantly high levels of inflammation, but the underlying pathogenic mechanisms are still not fully understood. Signal transducer and activator of transcription 1 (STAT1) and STAT3, and the downstream proteins suppressor of cytokine signaling 1 (SOCS1) and SOCS3, have been implicated in psoriasis disease progression. Calcipotriol, a synthetic derivative of vitamin D, has been used clinically to treat psoriasis, but the mechanism of action that underlies the beneficial effects of calcipotriol is still being explored. The objective of this study was to determine whether STAT1 and STAT3 signaling is involved in calcipotriol treatment. Using an in vitro immortal human keratinocyte cell line, HaCaT cells, as a psoriasis model, we examined the molecular signaling induced by calcipotriol treatment. We found that calcipotriol treatment or silencing of either STAT1 or STAT3 inhibited proliferation of HaCaT cells. Calcipotriol downregulated the expression of STAT1 and STAT3 at the messenger RNA (mRNA) and protein levels. The levels of phosphorylated STAT1 and STAT3 were also decreased, suggesting calcipotriol treatment inhibited STAT1 and STAT3 activation. Calcipotriol-mediated STAT inhibition was further substantiated by the downregulation of SOCS1 and SOCS3 at the mRNA and protein expression levels. Taken together, our results suggest a novel molecular mechanism for calcipotriol-mediated treatment effects in psoriasis.

scholarly journals Nutrients Bioaccessibility and Anti-inflammatory Features of Fermented Bee Pollen: A Comprehensive Investigation

2021 ◽  
Vol 12 ◽  
Author(s):  
Pasquale Filannino ◽  
Raffaella Di Cagno ◽  
Olimpia Vincentini ◽  
Daniela Pinto ◽  
Andrea Polo ◽  
...  
Keyword(s):  
Colon Adenocarcinoma ◽  
Human Keratinocytes ◽  
Protective Role ◽  
Protective Effects ◽  
Adenocarcinoma Cell Line ◽  
Inflammatory Stimuli ◽  
Keratinocyte Cell ◽  
Functional Features ◽  
Positive Effect

We compared raw bee-collected pollen (Raw-BCP), spontaneously fermented BCP (Unstarted-BCP), and BCP fermented with selected microbial starters (Started-BCP) to deepen whether fermentation may favorably affect the nutrients bioaccessibility and functional features of BCP. Under in vitro gastrointestinal batches, the highest serum-availability of phenolic compounds was found in Started-BCP, highlighting the positive effect exerted by selected microbial starters. The same effect was not found in spontaneously fermented BCP. In colon adenocarcinoma cell line-2 (Caco-2) cells stressed by a pro-inflammatory stimulus, the treatment with Started-BCP halted the increase of pro-inflammatory mediator’s level. Started-BCP counteracted efficiently the deleterious effects of inflammatory stimuli on the integrity of the Caco-2 cells monolayer and its barrier function. Started-BCP successfully counteracted the H2O2-induced intracellular accumulation of reactive oxygen species (ROS) in Caco-2 cells. A protective role against lipopolysaccharide (LPS)-induced inflammation was exerted by Started-BCP in human keratinocytes. The same protective effects on Caco-2 and keratinocyte cell lines were negligible after treatments with Raw-BCP or Unstarted-BCP.

scholarly journals HaCaT Cells as a Reliable In Vitro Differentiation Model to Dissect the Inflammatory/Repair Response of Human Keratinocytes

2017 ◽  
Vol 2017 ◽  
pp. 1-12 ◽  
Author(s):  
Irma Colombo ◽  
Enrico Sangiovanni ◽  
Roberta Maggio ◽  
Carlo Mattozzi ◽  
Stefania Zava ◽  
...  
Keyword(s):  
Cell Density ◽  
Skin Diseases ◽  
Inflammatory Responses ◽  
Human Keratinocytes ◽  
Suitable Model ◽  
Hacat Cells ◽  
Primary Human Keratinocytes ◽  
Proinflammatory Mediators ◽  
Cytokines And Chemokines

Cultured primary human keratinocytes are frequently employed for studies of immunological and inflammatory responses; however, interpretation of experimental data may be complicated by donor to donor variability, the relatively short culture lifetime, and variations between passages. To standardize the in vitro studies on keratinocytes, we investigated the use of HaCaT cells, a long-lived, spontaneously immortalized human keratinocyte line which is able to differentiate in vitro, as a suitable model to follow the release of inflammatory and repair mediators in response to TNFα or IL-1β. Different treatment conditions (presence or absence of serum) and differentiation stimuli (increase in cell density as a function of time in culture and elevation of extracellular calcium) were considered. ELISA and Multiplex measurement technologies were used to monitor the production of cytokines and chemokines. Taken together, the results highlight that Ca2+ concentration in the medium, cell density, and presence of serum influences at different levels the release of proinflammatory mediators by HaCaT cells. Moreover, HaCaT cells maintained in low Ca2+ medium and 80% confluent are similar to normal keratinocytes in terms of cytokine production suggesting that HaCaT cells may be a useful model to investigate anti-inflammatory interventions/therapies on skin diseases.

scholarly journals Bioimpedance Analysis of L929 and HaCaT Cells in Low Frequency Range

2018 ◽  
Vol 4 (1) ◽  
pp. 115-118 ◽  
Author(s):  
Viviane S. Teixeira ◽  
Jan-Patrick Kalckhoff ◽  
Wolfgang Krautschneider ◽  
Dietmar Schroeder
Keyword(s):  
Low Frequency ◽  
Steel Corrosion ◽  
Frequency Dispersion ◽  
Human Keratinocytes ◽  
Cell Types ◽  
Hacat Cells ◽  
Frequency Range ◽  
Corrosion Resistant ◽  
L929 Mouse

AbstractIn this work, Bioimpedance Spectroscopy (BIS) is used to study fluids and cell solutions. A n ew fourelectrode- terminal (4T) chamber using 3D printing and stainless steel corrosion resistant V4A was designed to measure the impedance of live cell solutions at the frequency range 0.1Hz- 1MHz. At f < 1kHz the double layer (DL) that builds at electrode’s surface raises the impedance substantially preventing the observation of the real impedance of the cells. The new 4T design circumvents the DL, is more robust and cheap, and allows for the repeatability of the results. Experiments were performed in vitro with two cell lines, L929 (mouse fibroblasts) and HaCaT (human keratinocytes). Results show that it is possible to distinguish between the two cell types by means of its BIS measurements in the new setup. Also, a low-frequency dispersion (α-dispersion) was observed in HaCaT cells solution, but not in L929. Furthermore, a potentiostat circuit model was developed in LTSpice to simulate the hardware setup and two different circuit models were used to fit cell’s data.

scholarly journals The Selection of NFκB Inhibitors to Block Inflammation and Induce Sensitisation to FasL-Induced Apoptosis in HNSCC Cell Lines Is Critical for Their Use as a Prospective Cancer Therapy

2019 ◽  
Vol 20 (6) ◽  
pp. 1306 ◽  
Author(s):  
Mario Scheurer ◽  
Roman Brands ◽  
Mohamed El-Mesery ◽  
Stefan Hartmann ◽  
Urs Müller-Richter ◽  
...  
Keyword(s):  
Cancer Therapy ◽  
Cell Lines ◽  
In Vitro Study ◽  
Keratinocyte Cell Line Hacat ◽  
Keratinocyte Cell ◽  
Nfκb Pathway ◽  
Induced Apoptosis ◽  
Selection Of ◽  
Hnscc Cell

Inflammation is a central aspect of tumour biology and can contribute significantly to both the origination and progression of tumours. The NFκB pathway is one of the most important signal transduction pathways in inflammation and is, therefore, an excellent target for cancer therapy. In this work, we examined the influence of four NFκB inhibitors—Cortisol, MLN4924, QNZ and TPCA1—on proliferation, inflammation and sensitisation to apoptosis mediated by the death ligand FasL in the HNSCC cell lines PCI1, PCI9, PCI13, PCI52 and SCC25 and in the human dermal keratinocyte cell line HaCaT. We found that the selection of the inhibitor is critical to ensure that cells do not respond by inducing counteracting activities in the context of cancer therapy, e.g., the extreme IL-8 induction mediated by MLN4924 or FasL resistance mediated by Cortisol. However, TPCA1 was qualified by this in vitro study as an excellent therapeutic mediator in HNSCC by four positive qualities: (1) proliferation was inhibited at low μM-range concentrations; (2) TNFα-induced IL-8 secretion was blocked; (3) HNSCC cells were sensitized to TNFα-induced cell death; and (4) FasL-mediated apoptosis was not disrupted.

scholarly journals Local Treatment of Hand-Foot Syndrome with Uridine/Thymidine:In VitroAppraisal on a Human Keratinocyte Cell Line HaCaT

2012 ◽  
Vol 2012 ◽  
pp. 1-6 ◽  
Author(s):  
J. Hartinger ◽  
P. Veselý ◽  
E. Matoušková ◽  
S. Argalacsová ◽  
L. Petruželka ◽  
...  
Keyword(s):  
Cell Line ◽  
Local Treatment ◽  
Local Therapy ◽  
Anticancer Therapy ◽  
Protective Activity ◽  
Human Keratinocyte ◽  
Human Keratinocyte Cell Line ◽  
Keratinocyte Cell Line Hacat ◽  
Keratinocyte Cell

5-fluorouracil (5-FU) is one of the most commonly used antineoplastic drugs in the anticancer therapy. The hand-foot (HF) syndrome (palmar-plantar erythrodysesthesia) is an adverse effect frequently related to long-term i.v. administration of 5-FU or its orally applicable prodrug capecitabine. Its severity can even lead to interruption of the otherwise effective anticancer therapy. Tentative practice in some clinics has shown that topical application of 10% uridine ointment is beneficial for calming down the HF syndrome. This study is focused on verifying the alleged protective activity of uridine in thein vitromodel of cultured human keratinocyte cell line HaCaT. We also tested the protective effects of thymidine alone or uridine-thymidine combination. The cellular viability time progression was measured in order to evaluate the effect of protective agents by three different types of cytopathogenicity tests—NTCA test (non-destructive test of cellular activity), modified MTT test and RTCA (real-time cell analyser, Roche). All three methods proved the ability of uridine and uridine-thymidine combination to protect keratinocytes against 5-FU damagein vitro. While thymidine alone did not show any remarkable effect, the thymidine-uridine combination demonstrated enhanced protective activity compared to uridine alone. Our findings provided the supporting rationale for using uridine or uridine-thymidine ointments in the HF syndrome local therapy.

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