The Effects of Carbon Dots on Immune System Biomarkers, Using the Murine Macrophage Cell Line RAW 264.7 and Human Whole Blood Cell Cultures
Abstract
:1. Introduction
2. Materials and Methods
2.1. Synthesis and Characterization of Carbon Dots (CDs)
2.2. Preparation of CD Stock Solutions
2.3. RAW 264.7 Cells
2.3.1. RAW 264.7 Macrophage Assay
2.3.2. Cytotoxicity Assay
2.3.3. NO Determination
2.3.4. Mouse IL-6 Double Antibody Sandwich (DAS) Enzyme Linked Immunosorbent Assay (ELISA)
2.3.5. Mouse MIPs (MIP-1α, MIP-1β and MIP-2) DAS ELISAs
2.3.6. Mouse Proteome Profiling Assay
2.3.7. Quantification of Pixel Density for Cytokine and Chemokine Membranes
2.4. Whole Blood Cell (WBC) Culture
2.4.1. Blood Collection
2.4.2. Cell Culture
2.4.3. Cytotoxicity Assay
2.4.4. Cytokine Analysis using DAS ELISAs
2.4.5. Human MIP-1β DAS ELISA
2.4.6. Human Proteome Profiling
2.5. Statistical Analysis
3. Results
3.1. The Effects of CDs on RAW 264.7 Cells
3.1.1. The Effects of CDs on the Viability of RAW 264.7 Cells
3.1.2. The Effects of CDs on the Inflammatory Biomarker NO Using RAW 264.7 Cells
3.1.3. The Effects of CDs on the Inflammatory Biomarker IL-6 Using RAW 264.7 Cells
3.1.4. The Effects of CDs on MIP-1α Using RAW 264.7 Cells
3.1.5. The Effects of CDs on MIP-1β Using RAW 264.7 Cells
3.1.6. The Effects of CDs on MIP-2 Using RAW 264.7 Cells
3.2. The Effects of CDs on the Secretory Cytokine and Chemokine Profile of RAW 264.7 Cells
3.3. The Effects of CDs on WBCs
3.3.1. The Effects of CDs on Viability of WBCs
3.3.2. The Effects of CDs on the Inflammatory System Biomarker IL-6 Using WBCs
3.3.3. The Effects of CDs on the Inflammatory Chemokine, MIP-1β Using WBCs
3.3.4. The Effects of CDs on the Humoral Immune System Biomarker IL-10 Using WBCs
3.3.5. The Effects of CDs on the Cell Mediated Immune System Biomarker IFNγ Using WBCs
3.4. The Effects of CDs on the Secretory Cytokine and Chemokine Profile of WBCs
4. Discussion
5. Conclusions
Author Contributions
Acknowledgments
Conflicts of Interest
Appendix A
pH | Average | Standard Deviation |
---|---|---|
3.04 | −1.42 | 0.87 |
5.25 | −11.40 | 4.35 |
6.99 | −16.83 | 1.40 |
10.12 | −35.07 | 2.74 |
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Cytokines and Chemokines | Positive Control | Negative Control | 500 μg/mL CDs |
---|---|---|---|
Reference Spot | 100 ± 8.62 | 100 ± 10.49 | 100 ± 15.19 |
IP-10 | 114.74 ± 5.24 | 0 ± 0 | 8.20 ± 0.11 a,b |
G-CSF | 58.27 ± 2.55 | 0 ± 0 | 26.76 ± 0.51 a,b |
TNF-α | 102.01 ± 5.91 | 17.96 ± 0.13 | 101.00 ± 0.60 a |
GM-CSF | 47.32 ± 3.02 | 0 ± 0 | 3.72 ± 0.03 a,b |
IL-6 | 91.94 ± 3.60 | 0 ± 0 | 0 ± 0 b |
JE | 78.97 ± 4.48 | 0 ± 0 | 10.47 ± 0.01 a,b |
sICAM | 109.05 ± 6.00 | 34.76 ± 0.02 | 78.65 ± 0.98 a,b |
MIP-1α | 72.87 ± 4.22 | 57.00 ± 1.94 | 64.87 ± 1.87 a,b |
MIP-1β | 62.77 ± 3.35 | 37.68 ± 0.33 | 45.40 ± 0.81 a,b |
MIP-2 | 55.51 ± 2.75 | 0 ± 0 | 76.63 ± 0.06 a,b |
RANTES | 100.57 ± 4.42 | 17.93 ± 0.83 | 36.09 ± 0.78 a,b |
SDF-1 | 16.41 ± 1.61 | 21.75 ± 1.01 | 0 ± 0 a,b |
IL-27 | 54.97 ± 4.05 | 0 ± 0 | 0 ± 0 b |
IL-1ra | 55.84 ± 1.66 | 0 ± 0 | 14.96 ± 0.21 a,b |
IL-1β | 39.16 ± 3.31 | 0 ± 0 | 0 ± 0 b |
Cytokines and Chemokines | Positive Control | Negative Control | 500 μg/mL CDs |
---|---|---|---|
Reference Spot | 100 ± 7.28 | 100 ± 5.57 | 100 ± 11.35 |
IL-1ra | 16.92 ± 1.46 | 0 ± 0 | 12.57 ± 1.27 a,b |
MIF | 17.56 ± 1.51 | 10.57 ± 1.91 | 20.41 ± 4.00 a |
MCP-1 | 26.26 ± 2.54 | 0 ± 0 | 0 ± 0 b |
Serpin E1 | 32.40 ± 2.49 | 24.26 ± 2.26 | 31.81 ± 4.38 |
MIP-1α/β | 70.22 ± 4.11 | 0 ± 0 | 25.84 ± 2.17 a,b |
RANTES | 62.94 ± 2.55 | 34.28 ± 1.37 | 32.95 ± 2.75 b |
sICAM | 37.15 ± 0.59 | 35.98 ± 2.19 | 32.38 ± 4.52 |
IL-6 | 29.31 ± 4.81 | 0 ± 0 | 0 ± 0 b |
IL-8 | 50.27 ± 8.41 | 0 ± 0 | 13.56 ± 2.53 a,b |
IL-1β | 20.29 ± 0.99 | 0 ± 0 | 0 ± 0 b |
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Lategan, K.; Fowler, J.; Bayati, M.; Fidalgo de Cortalezzi, M.; Pool, E. The Effects of Carbon Dots on Immune System Biomarkers, Using the Murine Macrophage Cell Line RAW 264.7 and Human Whole Blood Cell Cultures. Nanomaterials 2018, 8, 388. https://doi.org/10.3390/nano8060388
Lategan K, Fowler J, Bayati M, Fidalgo de Cortalezzi M, Pool E. The Effects of Carbon Dots on Immune System Biomarkers, Using the Murine Macrophage Cell Line RAW 264.7 and Human Whole Blood Cell Cultures. Nanomaterials. 2018; 8(6):388. https://doi.org/10.3390/nano8060388
Chicago/Turabian StyleLategan, Kim, Jodi Fowler, Mohamed Bayati, Maria Fidalgo de Cortalezzi, and Edmund Pool. 2018. "The Effects of Carbon Dots on Immune System Biomarkers, Using the Murine Macrophage Cell Line RAW 264.7 and Human Whole Blood Cell Cultures" Nanomaterials 8, no. 6: 388. https://doi.org/10.3390/nano8060388