Cisplatin Treatment Cell Viability And Role

THP-1 cells were incubated with cisplatin (CspA) at 10, 20 or 100μg/ml. There are 2 types of cells, cisplatin-treated and mock-treated. Among the cisplatin-treated cells are the 3 categories: 10μg/ml Cspa, 20μg/ml Cspa, and 100μg/ml Cspa. The mock-treated cells have 80% cell viability, 10μg/ml Cspa have around 78% cell viability, 20μg/ml Cspa have around 68% cell viability, and 100μg/ml Cspa have around 10% cell viability.

THP-1 cells were mock-infected or infected with HCMV for 1h, prior to treatment with 100μg/ml of cisplatin (CspA). Cell viability was assessed 24h after incubation with cisplatin. The mock-treated cells have 78% cell viability, 10μg/ml Cspa have around 30% cell viability, 20μg/ml Cspa have around 78% cell viability, and 100μg/ml Cspa have around 72% cell viability.

There are 2 types of cells, cisplatin-treated and mock-treated. Among the cisplatin-treated cells are the 3 categories: 10μg/ml Cspa, 20μg/ml Cspa, and 100μg/ml Cspa. THP-1 cells were mock-infected or infected with HCMV for 1h, prior to treatment with 100μg/ml of cisplatin (CspA). All results are statistically significant.

CD34+ has been infected with the followings and with following percentage of cell death. Mock, 15% cell death, HCMV, around 8% cell death, ERK, around 17% cell death, ERK + HCMV, around 40% cell death, RAF, around 12% cell death, RAF + HCMV, around 17% cell death, Tpl2, around 15% cell death, Tpl2 + HCMV, around 32% cell death. (B) THP-1 has been infected with the followings and with following percentage of cell death. Mock, 18% cell death, HCMV, around 15% cell death, ERK, around 17% cell death, ERK + HCMV, around 58% cell death, RAF, around 18% cell death, RAF + HCMV, around 16% cell death, Tpl2, around 17% cell death, Tpl2 + HCMV, around 47% cell death. 4. The rationale of analysis in the Fig. 2C has been the fold change in the MCL-1RNA expression versus Mock with which the infections are measured.

The Fig. 2C tells us about the infections of HCMV that has been incubated with inhibitor or incubated with tpl2 inhibitor and then infected with HCMV. These infections are treated with DMSO by performing qRT-PCR for measuring MCL-1RNA expression. MCL-1 RNA expression is measured 2 hours post infection in THP1 cells. 6. (A) Cell death was measured by trypan blue exclusion. CD34+ or THP-1 cells have been infected with the followings and with following percentage of cell death. For the CD34+; Mock, 10% cell death, HCMV, around 8% cell death, ERK 1/2, around 17% cell death, ERK 1/2+ HCMV, around 50% cell death.

For THP-1, Mock, 8% cell death, HCMV, around 7% cell death, ERK 1/2, around 10% cell death, ERK 1/2+ HCMV, around 8% cell death. (B) A qRT-PCR for MCL-1 RNA expression was performed. CD34+ or THP-1 cells have been infected with the followings and with following percentage of fold change in MCL-1 RNA expression versus mock. . For the CD34+; Mock, 2% MCL-1 RNA expression was performed, HCMV, around 15% MCL-1 RNA expression was performed, ERK 1/2, around 2% MCL-1 RNA expression was performed, ERK 1/2+ HCMV, around 3% MCL-1 RNA expression was performed. For the THP-1 cells; Mock, 2% MCL-1 RNA expression was performed, HCMV, around 5% MCL-1 RNA expression was performed, ERK 1/2, around 1% MCL-1 RNA expression was performed, ERK 1/2+ HCMV, around 2% MCL-1 RNA expression was performed.

Fig 3B indicates a qRT-PCR for MCL-1 RNA expression performed on CD34+ and THP-1. On the other hand, the question 5 indicates performing of qRT-PCR for measuring MCL-1RNA expression. Thus, what has been measured are roughly the same with one major difference. In Figure 3B, qRT-PCR for MCL-1 RNA expression was performed on CD34+ and THP-1, while for the question 5, the qRT-PCR for MCL-1 RNA expression has been performed on THP-1 only. Also the infections measured are slightly different between Fig 3B and Question 5. However, what has been measured is identical i.e. qRT-PCR for MCL-1 RNA expression and thus Fig 3B’s impact will be on Question 5.

The first experiment conducted has been the THP-1 cells incubated with cisplatin (CspA) at 10, 20 or 100μg/ml. The second experiment was the THP-1 cells been mock-infected or infected with HCMV for 1h, prior to treatment with 100μg/ml of cisplatin (CspA). Cell viability was assessed 24h after incubation with cisplatin. These 2 tests were strategically significant tested with a t-test. The next measurement was the MCL-1RNA expression versus Mock. Then the experiment conducted was with infections of HCMV that has been incubated with inhibitor or incubated with tpl2 inhibitor and then infected with HCMV. These infections are treated with DMSO by performing qRT-PCR for measuring MCL-1RNA expression. MCL-1 RNA expression is measured 2 hours post infection in THP1 cells. However, the limitation of this experiment was CD34+ being excluded from the experiment. The next experiment was a qRT-PCR for MCL-1 RNA expression was performed. CD34+ or THP-1 cells have been infected. Thus, in this experiment both CD34+ and THP-1 cells have been included.

The most important limitation that was detected was in MCL-1 RNA expression measured 2 hours post infection in THP1 cells and where CD34+ was excluded. The subsequent experiment conducted with qRT-PCR for MCL-1 RNA expression included both CD34+ and THP-1 cells. Therefore, the limitation of the earlier experiment has been addressed in the subsequent experiment to make its mechanism more complete and meaningful.

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