scholarly journals Colorectal Adenocarcinoma Cell Culture in a Microfluidically Controlled Environment with a Static Molecular Gradient of Polyphenol

Molecules ◽  
2021 ◽  
Vol 26 (11) ◽  
pp. 3215
Author(s):  
Roman G. Szafran ◽  
Kazimierz Gąsiorowski ◽  
Benita Wiatrak
Keyword(s):  
Cell Culture ◽  
Cell Viability ◽  
Degradation Products ◽  
Microfluidic System ◽  
Small Height ◽  
Low Concentrations ◽  
Cell Mortality ◽  
Affect Cell Viability ◽  
The Future ◽  
Static Conditions

To study the simultaneous effect of the molecular gradient of polyphenols (curcumin, trans-resveratrol, and wogonin) and biological factors released from tumor cells on apoptosis of adjacent cells, a novel microfluidic system was designed and manufactured. The small height/volume of microfluidic culture chambers and static conditions allowed for establishing the local microenvironment and maintaining undisturbed concentration profiles of naturally secreted from cells biochemical factors. In all trials, we observe that these conditions significantly affect cell viability by stimulating cell apoptosis at lower concentrations of polyphenols than in traditional multiwell cultures. The observed difference varied between 20.4–87.8% for curcumin, 11.0–37.5% for resveratrol, and 21.7–62.2% for wogonin. At low concentrations of polyphenols, the proapoptotic substances released from adjacent cells, like protein degradation products, significantly influence cell viability. The mean increase in cell mortality was 38.3% for microfluidic cultures. Our research has also confirmed that the gradient microsystem is useful in routine laboratory tests in the same way as a multiwell plate and may be treated as its replacement in the future. We elaborated the new repetitive procedures for cell culture and tests in static gradient conditions, which may become a gold standard of new drug investigations in the future.

scholarly journals Cell Viability and Immune Response to Low Concentrations of Nickel and Cadmium: An In Vitro Model

2020 ◽  
Vol 17 (24) ◽  
pp. 9218
Author(s):  
Ahra Kim ◽  
SangJin Park ◽  
Joo Hyun Sung
Keyword(s):  
Heavy Metals ◽  
Immune Response ◽  
Cell Viability ◽  
Clinical Manifestations ◽  
A549 Cells ◽  
Low Concentrations ◽  
Affect Cell Viability ◽  
Cytokine Levels ◽  
Vitro Model

Environmental exposure to low concentrations of heavy metals is common in the general population, but the toxicity, immune response mechanisms, and the effects of single and mixed metal exposures have not been clearly identified. In this study, A549 cells and Raw264.7 cells were exposed to low concentrations of the heavy metals nickel (Ni) and cadmium (Cd) for 24, 48, and 72 h, and then cell viability and cytokine levels were analyzed. We found that exposure to low concentrations of Ni (50 nM) or Cd (10 nM) alone did not affect cell viability. However, mixing them together decreased cell viability. In addition, the levels of IL-10, IL-12, and TNF-α decreased with single (only Cd) and mixed (Ni and Cd) exposures. These results show that exposure to low concentrations of heavy metals could affect the normal immune response, even without obvious clinical manifestations. Therefore, chronic exposure to heavy metals might have adverse effects on overall health.

Islet-on-a-chip: Biomimetic micropillar-based microfluidic system for three-dimensional pancreatic islet cell culture

2021 ◽  
Vol 183 ◽  
pp. 113215
Author(s):  
Patrycja Sokolowska ◽  
Kamil Zukowski ◽  
Justyna Janikiewicz ◽  
Elzbieta Jastrzebska ◽  
Agnieszka Dobrzyn ◽  
...  
Keyword(s):  
Cell Culture ◽  
Pancreatic Islet ◽  
Islet Cell ◽  
Three Dimensional ◽  
Microfluidic System ◽  
Pancreatic Islet Cell ◽  
Islet Cell Culture

scholarly journals A glutaminyl cyclase-catalyzed α-synuclein modification identified in human synucleinopathies

2021 ◽  
Author(s):  
Maike Hartlage-Rübsamen ◽  
Alexandra Bluhm ◽  
Sandra Moceri ◽  
Lisa Machner ◽  
Janett Köppen ◽  
...  
Keyword(s):  
Cell Viability ◽  
Neurodegenerative Disorder ◽  
Lewy Bodies ◽  
Size Exclusion ◽  
Lewy Neurites ◽  
Enzymatic Assays ◽  
Aβ Peptides ◽  
Glutaminyl Cyclase ◽  
Affect Cell Viability ◽  
Exclusion Chromatography

AbstractParkinson’s disease (PD) is a progressive neurodegenerative disorder that is neuropathologically characterized by degeneration of dopaminergic neurons of the substantia nigra (SN) and formation of Lewy bodies and Lewy neurites composed of aggregated α-synuclein. Proteolysis of α-synuclein by matrix metalloproteinases was shown to facilitate its aggregation and to affect cell viability. One of the proteolysed fragments, Gln79-α-synuclein, possesses a glutamine residue at its N-terminus. We argue that glutaminyl cyclase (QC) may catalyze the pyroglutamate (pGlu)79-α-synuclein formation and, thereby, contribute to enhanced aggregation and compromised degradation of α-synuclein in human synucleinopathies. Here, the kinetic characteristics of Gln79-α-synuclein conversion into the pGlu-form by QC are shown using enzymatic assays and mass spectrometry. Thioflavin T assays and electron microscopy demonstrated a decreased potential of pGlu79-α-synuclein to form fibrils. However, size exclusion chromatography and cell viability assays revealed an increased propensity of pGlu79-α-synuclein to form oligomeric aggregates with high neurotoxicity. In brains of wild-type mice, QC and α-synuclein were co-expressed by dopaminergic SN neurons. Using a specific antibody against the pGlu-modified neo-epitope of α-synuclein, pGlu79-α-synuclein aggregates were detected in association with QC in brains of two transgenic mouse lines with human α-synuclein overexpression. In human brain samples of PD and dementia with Lewy body subjects, pGlu79-α-synuclein was shown to be present in SN neurons, in a number of Lewy bodies and in dystrophic neurites. Importantly, there was a spatial co-occurrence of pGlu79-α-synuclein with the enzyme QC in the human SN complex and a defined association of QC with neuropathological structures. We conclude that QC catalyzes the formation of oligomer-prone pGlu79-α-synuclein in human synucleinopathies, which may—in analogy to pGlu-Aβ peptides in Alzheimer’s disease—act as a seed for pathogenic protein aggregation.

scholarly journals Zinc, Zinc Transporters, and Cadmium Cytotoxicity in a Cell Culture Model of Human Urothelium

Toxics ◽  
2021 ◽  
Vol 9 (5) ◽  
pp. 94
Author(s):  
Soisungwan Satarug ◽  
Scott H. Garrett ◽  
Seema Somji ◽  
Mary Ann Sens ◽  
Donald A. Sens
Keyword(s):  
Dna Methylation ◽  
Cell Culture ◽  
Cell Viability ◽  
Zinc Transporters ◽  
Cell Culture Model ◽  
Induced Expression ◽  
Glutathione Biosynthesis ◽  
Culture Model ◽  
A Cell ◽  
Cd Exposure

We explored the potential role of zinc (Zn) and zinc transporters in protection against cytotoxicity of cadmium (Cd) in a cell culture model of human urothelium, named UROtsa. We used real-time qRT-PCR to quantify transcript levels of 19 Zn transporters of the Zrt-/Irt-like protein (ZIP) and ZnT gene families that were expressed in UROtsa cells and were altered by Cd exposure. Cd as low as 0.1 µM induced expression of ZnT1, known to mediate efflux of Zn and Cd. Loss of cell viability by 57% was seen 24 h after exposure to 2.5 µM Cd. Exposure to 2.5 µM Cd together with 10–50 µM Zn prevented loss of cell viability by 66%. Pretreatment of the UROtsa cells with an inhibitor of glutathione biosynthesis (buthionine sulfoximine) diminished ZnT1 induction by Cd with a resultant increase in sensitivity to Cd cytotoxicity. Conversely, pretreatment of UROtsa cells with an inhibitor of DNA methylation, 5-aza-2’-deoxycytidine (aza-dC) did not change the extent of ZnT1 induction by Cd. The induced expression of ZnT1 that remained impervious in cells treated with aza-dC coincided with resistance to Cd cytotoxicity. Therefore, expression of ZnT1 efflux transporter and Cd toxicity in UROtsa cells could be modulated, in part, by DNA methylation and glutathione biosynthesis. Induced expression of ZnT1 may be a viable mechanistic approach to mitigating cytotoxicity of Cd.

scholarly journals Intestinal and Hepatic Uptake of Dietary Peroxidized Lipids and Their Decomposition Products, and Their Subsequent Effects on Apolipoprotein A1 and Paraoxonase1

Antioxidants ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 1258
Author(s):  
Xueting Jiang ◽  
Pragney Deme ◽  
Rajat Gupta ◽  
Dmitry Litvinov ◽  
Kathryn Burge ◽  
...  
Keyword(s):  
Cell Culture ◽  
Culture Media ◽  
Degradation Products ◽  
Apolipoprotein A1 ◽  
Paraoxonase 1 ◽  
Pon1 Activity ◽  
Atherosclerotic Cardiovascular Disease ◽  
Metabolic Fate ◽  
Cell Culture Media ◽  
Decomposition Products

Both pro- and antiatherosclerotic effects have been ascribed to dietary peroxidized lipids. Confusion on the role of peroxidized lipids in atherosclerotic cardiovascular disease is punctuated by a lack of understanding regarding the metabolic fate and potential physiological effects of dietary peroxidized lipids and their decomposition products. This study sought to determine the metabolic fate and physiological ramifications of 13-hydroperoxyoctadecadienoic acid (13-HPODE) and 13-HODE (13-hydroxyoctadecadienoic acid) supplementation in intestinal and hepatic cell lines, as well as any effects resulting from 13-HPODE or 13-HODE degradation products. In the presence of Caco-2 cells, 13-HPODE was rapidly reduced to 13-HODE. Upon entering the cell, 13-HODE appears to undergo decomposition, followed by esterification. Moreover, 13-HPODE undergoes autodecomposition to produce aldehydes such as 9-oxononanoic acid (9-ONA). Results indicate that 9-ONA was oxidized to azelaic acid (AzA) rapidly in cell culture media, but AzA was poorly absorbed by intestinal cells and remained detectable in cell culture media for up to 18 h. An increased apolipoprotein A1 (ApoA1) secretion was observed in Caco-2 cells in the presence of 13-HPODE, 9-ONA, and AzA, whereas such induction was not observed in HepG2 cells. However, 13-HPODE treatments suppressed paraoxonase 1 (PON1) activity, suggesting the induction of ApoA1 secretion by 13-HPODE may not represent functional high-density lipoprotein (HDL) capable of reducing oxidative stress. Alternatively, AzA induced both ApoA1 secretion and PON1 activity while suppressing ApoB secretion in differentiated Caco-2 cells but not in HepG2. These results suggest oxidation of 9-ONA to AzA might be an important phenomenon, resulting in the accumulation of potentially beneficial dietary peroxidized lipid-derived aldehydes.

scholarly journals A High Throughput Approach Based on Dynamic High Pressure for the Encapsulation of Active Compounds in Exosomes for Precision Medicine

2021 ◽  
Vol 22 (18) ◽  
pp. 9896
Author(s):  
Eugenia Romano ◽  
Paolo Antonio Netti ◽  
Enza Torino
Keyword(s):  
Cell Culture ◽  
High Pressure ◽  
Culture Media ◽  
Preliminary Test ◽  
Bilayer Membrane ◽  
Microfluidic System ◽  
Culture Model ◽  
Before And After ◽  
Dynamic High Pressure

In recent decades, endogenous nanocarrier-exosomes have received considerable scientific interest as drug delivery systems. The unique proteo-lipid architecture allows the crossing of various natural barriers and protects exosomes cargo from degradation in the bloodstream. However, the presence of this bilayer membrane as well as their endogenous content make loading of exogenous molecules challenging. In the present work, we will investigate how to promote the manipulation of vesicles curvature by a high-pressure microfluidic system as a ground-breaking method for exosomes encapsulation. Exosomes isolated from Uppsala 87 Malignant Glioma (U87-MG) cell culture media were characterized before and after the treatment with high-pressure homogenization. Once their structural and biological stability were validated, we applied this novel method for the encapsulation in the lipidic exosomal bilayer of the chemotherapeutic Irinotecan HCl Trihydrate-CPT 11. Finally, we performed in vitro preliminary test to validate the nanobiointeraction of exosomes, uptake mechanisms, and cytotoxic effect in cell culture model.

scholarly journals Low-Power Red and Infrared Laser Effects on Cells Deficient in DNA Repair

2019 ◽  
Vol 10 (3) ◽  
pp. 157-162
Author(s):  
Lucas Kiyoshi da Fonseca Iwahara ◽  
Flavia de Paoli ◽  
Adenilson de Souza da Fonseca
Keyword(s):  
Escherichia Coli ◽  
Dna Repair ◽  
Low Power ◽  
Cell Viability ◽  
Bacterial Cells ◽  
Wild Type ◽  
Low Level ◽  
Affect Cell Viability ◽  
Study Results ◽  
Emission Mode

Introduction: Low-level lasers are successfully used to prevent and treat diseases in soft oral and bone tissues, particularly diseases in oral cavity caused by chemotherapy and radiotherapy in oncology. However, controversy exists as to whether these lasers induce molecular side effects, mainly on DNA. The aim of this work was to assess the effects of low-power lasers on mutant Escherichia coli cells in DNA repair. Methods: Escherichia coli wild type cultures as well as those lacking recombination DNA repair (recA- ) and la SOS responses (lexA- ) irradiated with lasers at different energy densities, powers, and emission modes for cell viability and morphology assessment were used in this study. Results: Laser irradiation: (i) did not affect cell viability of non-mutant and lexA- cells but decreased viability in recA- cultures; (ii) altered morphology of wild type and lexA, depending on the energy density, power, emission mode, and wavelength. Conclusion: Results show that low-level lasers have lethal effects on both recombination DNA repair and SOS response bacterial cells but do not induce morphological modifications in these cells.

scholarly journals Specific identification of fibrin polymers, fibrinogen degradation products, and crosslinked fibrin degradation products in plasma and serum with a new sensitive technique

Blood ◽  
1985 ◽  
Vol 65 (3) ◽  
pp. 589-597
Author(s):  
DG Connaghan ◽  
CW Francis ◽  
DA Lane ◽  
VJ Marder
Keyword(s):  
Degradation Products ◽  
Sodium Dodecyl ◽  
Fibrinolytic Therapy ◽  
Electrophoretic Patterns ◽  
Fibrin Degradation Products ◽  
Low Concentrations ◽  
Combined Action ◽  
Derivatives Of ◽  
Normal Controls ◽  
Immunological Identification

A new method is described for identifying low concentrations of circulating derivatives of fibrinogen and fibrin, even when present in heterogeneous mixtures. This technique is applicable to plasma and serum and uses electrophoresis in 2% agarose in the presence of sodium dodecyl sulfate (SDS) followed by immunological identification of separated derivatives, using radiolabeled antifibrinogen antiserum and autoradiography. Unique electrophoretic patterns distinguish plasmic derivatives of crosslinked fibrin from those of fibrinogen and also identify crosslinked fibrin polymers produced by the combined action of thrombin and factor XIII on fibrinogen. The assay is sensitive to a concentration of 0.1 micrograms/mL of fibrinogen in serum or plasma. Fibrin polymers, plasmic degradation products of fibrinogen, and plasmic degradation products of crosslinked fibrin were detected in the plasma or serum of a patient with disseminated intravascular coagulation. Plasmic derivatives of both fibrinogen and crosslinked fibrin appeared in serum in the course of fibrinolytic therapy for pulmonary embolism, whereas during acute myocardial infarction a marked increase in the proportion of fibrin polymers in plasma was found in comparison with normal controls. Thus, the procedure can distinguish between the simultaneous processes of fibrin polymer formation, fibrinogenolysis, and fibrinolysis, and is sufficiently sensitive to detect relevant quantities of derivatives in pathologic conditions.

Glucose Degradation Products and Peritoneal Membrane Function

2001 ◽  
Vol 21 (2) ◽  
pp. 201-207 ◽  
Author(s):  
Janusz Witowski ◽  
Thorsten O. Bender ◽  
Gerhard M. Gahl ◽  
Ulrich Frei ◽  
Achim Jörres
Keyword(s):  
Peritoneal Dialysis ◽  
Cell Viability ◽  
Degradation Products ◽  
Membrane Function ◽  
Peritoneal Mesothelial Cells ◽  
Glucose Degradation Products ◽  
And Function ◽  
The Impact ◽  
Dialysis Fluids

Background The bioincompatibility of peritoneal dialysis fluids (PDF) in current use has been partially attributed to the presence of glucose degradation products (GDPs), which are generated during heat sterilization of PDF. Several of the GDPs have been identified and we have recently demonstrated that these GDPs per se may impair the viability and function of human peritoneal mesothelial cells (HPMC) in vitro. It is also possible that GDP-related toxicity is further exacerbated by the milieu of PDF. We review the current literature on GDP and present the results of experiments comparing the impact of heat- and filter-sterilized PDF on the viability and function of HPMC. Methods Peritoneal dialysis fluids with low (1.5%) and high (4.25%) glucose concentrations were laboratory prepared according to the standard formula and sterilized either by heat (H-PDF; 121°C, 0.2 MPa, 20 minutes) or filtration (F-PDF; 0.2 μ). The buildup of GDP was confirmed by UV absorbance at 284 nm. Confluent HPMC monolayers were exposed to these solutions mixed 1:1 with standard M199 culture medium. After 24 hours, cell viability was assessed with the MTT assay, and interleukin-1β–stimulated monocyte chemotactic protein-1 (MCP-1) release with specific immunoassay. Results Exposure of HPMC to H-PDF resulted in a significant decrease in cell viability, with solutions containing 4.25% glucose being more toxic than 1.5% glucose-based PDF (27.4% ± 3.4% and 53.4% ± 11.0% of control values, respectively). In contrast, viability of HPMC exposed to F-PDF was not different from that of control cells. Moreover, treatment with H-PDF impaired the release of MCP-1 from HPMC to a significantly greater degree compared to F-PDF (17.4% and 24.9% difference for low and high glucose PDF, respectively). Conclusions Exposure of HPMC to H-PDF significantly impairs cell viability and the capacity for generating MCP-1 compared to F-PDF. This effect is likely to be mediated by GDPs present in H-PDF but not in F-PDF.

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