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Figures 4 and 5 show no additional change in response to radiation in combination with GNP compared to GNPs alone, similar to the DNA damage data in Figure 2. Comparable to mitochondrial membrane polarisation, levels of cardiolipin oxidation remain steady post irradiation.
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However, the lack of change in the levels of mitochondrial membrane polarisation and cardiolipin post irradiation further emphasises the significance of the cellular events prior to irradiation in GNP radiosensitisation.
At 4 hours post irradiation with GNPs membrane polarisation had returned to the same level as non-irradiated GNP treated samples across all cell lines.
Figure 4 Mitochondrial membrane polarisation after GNP and irradiation.
Mitochondrial membrane polarisation was measured by TMRE flow cytometry and made relative to untreated control after cells were treated with 1.9 nm GNPs and/or 2 Gy irradiation.
MDA-MB-231 and T98G cells both displayed an increase in mitochondrial membrane polarisation of 30%and25%5% respectively, 1 hour post irradiation when exposed to GNP in combination with 2 Gy irradiation, which was significant in MDA-MB-231 cells, however, this coincided with an increase in membrane polarisation upon irradiation alone.
Figure 1 Correlation between resting membrane polarisation and insulin resistance (ISI = Insulin sensitivity index; indicates ratio of glucose infusion rate to the serum insulin concentration during steady state).
Changes in mitochondrial membrane polarisation were measured by flow cytometry analysis following 24 hours exposure to GNPs with and without exposure to a single dose of 2 Gy (Figure 4).
In unirradiated cells, GNPs alone significantly reduced mitochondrial membrane polarisation relative to controls across all cell lines with decreases of 50%, 55%and25%5% in TMRE fluorescence in MDA-MB-231, DU-145 and T98G cells respectively.
This is in contrast to strong dosage effects evident for both sporozoite membrane polarisation (Fig. S5) and internal granularity (Fig. S6).
However, it must also be considered that membrane polarisation and sporozoite internal granularity are complex functions and while related to exocytosis may also be a function of other critical parameters affecting sporozoite infectivity including sporozoite membrane and vesicle integrity.
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