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Synergistic Antiproliferative Effects of Chondroitin Sulfate and Fucoidan in Tumor-derived Spheroids: Insights From a 3D Cell Culture Approach

E Nováková

E Nováková

  • Comenius University Bratislava, Faculty of Natural Sciences, Department of Microbiology and VirologyBratislava, Slovak Republic
  • Comenius University Bratislava, Faculty of Pharmacy, Department of Galenic PharmacyBratislava, Slovak Republic
  • Biomedical Research Center of the Slovak Academy of Sciences, Institute of VirologyBratislava, Slovak Republic
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Nováková, E
, 
J Zima

J Zima

Comenius University Bratislava, Faculty of Pharmacy, Department of Galenic PharmacyBratislava, Slovak Republic
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Zima, J
, 
M Špaglová

M Špaglová

Comenius University Bratislava, Faculty of Pharmacy, Department of Galenic PharmacyBratislava, Slovak Republic
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Špaglová, M
, 
M Labudová

M Labudová

  • Comenius University Bratislava, Faculty of Natural Sciences, Department of Microbiology and VirologyBratislava, Slovak Republic
  • Biomedical Research Center of the Slovak Academy of Sciences, Institute of VirologyBratislava, Slovak Republic
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Labudová, M
 und 
M Šupolíková

M Šupolíková

  • Comenius University Bratislava, Faculty of Natural Sciences, Department of Microbiology and VirologyBratislava, Slovak Republic
  • Comenius University Bratislava, Faculty of Pharmacy, Department of Galenic PharmacyBratislava, Slovak Republic
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Šupolíková, M
  
20. Jan. 2025
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Artikel-Kategorie: Research paper
Online veröffentlicht: 20. Jan. 2025
Seitenbereich: 25 - 33
Eingereicht: 04. Dez. 2024
Akzeptiert: 12. Dez. 2024
DOI: https://doi.org/10.2478/afpuc-2024-0015
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© 2024 E Nováková et al., published by Sciendo
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Figure 1.

Comparison of non-tumor NIH3T3 cell line in 2D and 3D cell culture: A) non-tumor NIH3T3 cell line in 2D cell culture forming a cell monolayer (100× magnification); B) non-tumor NIH3T3 cell line in 3D cell culture forming a spheroid (100× magnification).
Comparison of non-tumor NIH3T3 cell line in 2D and 3D cell culture: A) non-tumor NIH3T3 cell line in 2D cell culture forming a cell monolayer (100× magnification); B) non-tumor NIH3T3 cell line in 3D cell culture forming a spheroid (100× magnification).

Figure 2.

Comparison of the Hepa1c1c7 tumor cell line in 2D and 3D cell culture: A) Hepa1c1c7 tumor cell line in 2D cell culture forming a cell monolayer (100× magnification); B) Hepa1c1c7 tumor cell line in 3D cell culture forming a spheroid (100× magnification).
Comparison of the Hepa1c1c7 tumor cell line in 2D and 3D cell culture: A) Hepa1c1c7 tumor cell line in 2D cell culture forming a cell monolayer (100× magnification); B) Hepa1c1c7 tumor cell line in 3D cell culture forming a spheroid (100× magnification).

Figure 3.

Spheroids from the non-tumor cell line NIH3T3 after treatment with A) chondroitin sulfate (CS), B) fucoidan (F), C) a combination of chondroitin sulfate and fucoidan (CS + F). Day 3 – first addition of substances; Day 7 – last addition of substances. Spheroids were photographed at 50× magnification. Up arrow (↑) – spheroids that increased their area compared to control spheroids; down arrow (↓) – spheroids that decreased their area compared to control spheroids; equal to (=) – spheroids that have approximately the same area compared to control spheroids.
Spheroids from the non-tumor cell line NIH3T3 after treatment with A) chondroitin sulfate (CS), B) fucoidan (F), C) a combination of chondroitin sulfate and fucoidan (CS + F). Day 3 – first addition of substances; Day 7 – last addition of substances. Spheroids were photographed at 50× magnification. Up arrow (↑) – spheroids that increased their area compared to control spheroids; down arrow (↓) – spheroids that decreased their area compared to control spheroids; equal to (=) – spheroids that have approximately the same area compared to control spheroids.

Figure 4.

Spheroids from the Hepa1c1c7 tumor cell line after treatment with A) chondroitin sulfate (CS), B) fucoidan (F), and C) a combination of chondroitin sulfate and fucoidan (CS + F). Day 3 – first addition of substances; Day 7 – last addition of substances. Spheroids were photographed at 50× magnification. Up arrow (↑) – spheroids that increased their area compared to control spheroids; down arrow (↓) – spheroids that decreased their area compared to control spheroids; equal to (=) – spheroids that have approximately the same area compared to control spheroids.
Spheroids from the Hepa1c1c7 tumor cell line after treatment with A) chondroitin sulfate (CS), B) fucoidan (F), and C) a combination of chondroitin sulfate and fucoidan (CS + F). Day 3 – first addition of substances; Day 7 – last addition of substances. Spheroids were photographed at 50× magnification. Up arrow (↑) – spheroids that increased their area compared to control spheroids; down arrow (↓) – spheroids that decreased their area compared to control spheroids; equal to (=) – spheroids that have approximately the same area compared to control spheroids.

Figure 5.

Flow cytometric analysis of non-tumor NIH3T3 cell lines treated with A) chondroitin sulfate (CS), B) fucoidan (F), and C) a combination of chondroitin sulfate and fucoidan (CS + F), and D) phosphate-buffered saline (PBS). Dead cells are shown on the top left, necrotic cells on the top right, live cells on the bottom left, and apoptotic cells are shown on the bottom right.
Flow cytometric analysis of non-tumor NIH3T3 cell lines treated with A) chondroitin sulfate (CS), B) fucoidan (F), and C) a combination of chondroitin sulfate and fucoidan (CS + F), and D) phosphate-buffered saline (PBS). Dead cells are shown on the top left, necrotic cells on the top right, live cells on the bottom left, and apoptotic cells are shown on the bottom right.

Figure 6.

Flow cytometric analysis of Hepa1c1c7 tumor cell line treated with A) chondroitin sulfate (CS), B) fucoidan (F), and C) a combination of chondroitin sulfate and fucoidan (CS + F), and D) phosphate-buffered saline (PBS). Dead cells are shown on the top left, necrotic cells on the top right, live cells on the bottom left, and apoptotic cells are shown on the bottom right.
Flow cytometric analysis of Hepa1c1c7 tumor cell line treated with A) chondroitin sulfate (CS), B) fucoidan (F), and C) a combination of chondroitin sulfate and fucoidan (CS + F), and D) phosphate-buffered saline (PBS). Dead cells are shown on the top left, necrotic cells on the top right, live cells on the bottom left, and apoptotic cells are shown on the bottom right.

The concentration of F and CS_

Dilution in DMEM CS (mmol/L) F (mmol/L) CS + F (mmol/L)
1:1 0.0620 0.0920 0.0620 0.0920
1:2 0.0310 0.0460 0.0310 0.0460
1:4 0.0155 0.0230 0.0155 0.0230

Quantitative representation of cells in spheroids prepared from the NIH3T3 cell line compared to Hepa1c1c7 cell line evaluated by flow cytometry (5000 cells)_

NIH3T3 cell line
Living cells (%) Apoptotic cells (%) Dead cells (%) Necrotic cells (%)
CS 66.43 ± 4.43 20.98 ± 1.03 9.79 ± 0.52 2.80 ± 0.18
F 13.53 ± 3.60 38.72 ± 4.09 1.50 ± 0.17 46.25 ± 2.97
CS + F 28.30 ± 1.65 19.10 ± 1.05 24.23 ± 5.28 28.37 ± 1.78
PBS 31.57 ± 3.81 32.67 ± 10.05 4.61 ± 0.50 31.15 ± 6.46
Hepa1c1c7 cell line
Living cells (%) Apoptotic cells (%) Dead cells (%) Necrotic cells (%)
CS 29.18 ± 3.69 29.57 ± 18.29 3.89 ± 0.55 37.36 ± 5.97
F 49.65 ± 4.77 27.66 ± 6.83 9.93 ± 0.25 12.75 ± 1.25
CS + F 41.52 ± 3.76 31.50 ± 6.55 19.84 ± 5.80 7.14 ± 0.83
PBS 50.00 ± 8.15 33.22 ± 2.52 7.09 ± 0.67 9.69 ± 0.45
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