The subG1 phase has been proposed to be a population of apoptotic cells (Darzynkiewicz et al 1992). Kratom Extract Amazon effects of MSE on cell cycle distribution of HEK 293 cells after 24 and 48 hours of treatment. Histograms are representative of three replicates of experiments with similar results best uk kratom vendor and analysed by Cellquest Pro software.
Surprisingly this time a similar outcome was observed for both SH-SY5Y and MCL-5 cells and the shifting of the whole populations was evident at much lower concentrations of MSE than in the previous PI staining in chapter 2. This phenomenon is obviously due to the treatment effects as the control and lowest concentration of the MSE tested as seen in fig. The hypothesis of plasma membrane opening is supported with this finding. This Kratom Extract Amazon phenomenon creates disadvantages for this assay as when the whole FACS profile shifts to the right side of the scale the determination of the stages of cell death is difficult to interpret as the cells are no longer located in specific quadrants. This observation is clearly in contrast with the previous cytological examinations which indicated that SH-SY5Y cells treated with high dose of MSE undergo apoptosis rather than necrosis.
The trypan blue assay and clonogenicity assay were employed as described in chapter 2 section 2. MSE and MIT are discussed as follows: Effects of naloxone on MSE and MIT treated cells: Fig. Naloxone also appears to successfully inhibit the MIT toxicity (Fig.
Based on these observations two possibilities are considered: 1) the effect is cell cycle arrest independent of p53 and p21 pathway or 2) the loss of these proteins could be due to the leakage due to the increased membrane permeability or through pore opening. The toxicity findings noted thus far are Kratom Extract Amazon consistent with my hypothesis in which the dose is the main factor in determining the level of the cytotoxicity seen. The cytotoxicity events initially seen as cell cycle arrest proceed to cell death with increasing doses of MSE and MIT.
A in the previous chapter (section 2. MSE in the presence of S9 reduced the colony formation to less than 10% of the vehicle treated control. A similar outcome was seen using S9 with L5178Y cells in this assay in the preliminary tests for selecting the range of concentrations performed prior to plating assessment. MSE was found to be too toxic with kratom online paypal RSG only 2% (Table 3. The results for MIT as shown in table 3. A and 3. B also revealed a negative outcome for genotoxicity under conditions with or without the presence of metabolic activation by S9.
There is another interesting finding to note apart from the toxicology implications of MSE and MIT as discussed above. M) stimulate cells to proliferate in most of the human cell lines examined. Thus this finding may support the pharmacology of the Mitragyna speciosa Korth leaves which produce stimulation effects when consumed at low doses. The stimulation effects claimed at low doses are based on anecdotal reports from users however the specific clinical pharmacology and controlled dosage for humans is still poorly understood. One of the main reasons for conducting toxicology kratom tea ingredients studies is to determine the risk or in other words to determine the potential for harm towards human health or the environment upon exposure to naturally occurring or synthetic agents.
Cell cycle is an essential process for all living organisms with the ultimate goal to create new cells Kratom Extract Amazon necessary for maintaining continued survival. Under normal circumstances the four phases of the cell cycle G1 S G2 and M phases are tightly regulated. The entry of the cell into each phase of cell cycle is carefully regulated by cell cycle checkpoints which act as the cell cycle control systems.
Fas)-mediated apoptosis: live and let die. Mitochondrial membrane permeabilization in cell death. Wildtype p53 is a cell cycle checkpoint determinant following irradiation.
Cambridge university press. La Quaglia M. Wild
type p53 protein undergoes cytoplasmic sequestration in undifferentiated neuroblastoma but no in differentiated tumors.
This finding suggests that the mode of the cell death of MIT treated cells is dependant on caspase 3 and 7 activation pathway. There were no significant differences in the subG1 population (apoptosis population) between treated groups (caspase 3 inhibitor caspase 8 inhibitor caspase 9 inhibitor and general caspase inhibitor treated with high dose of MSE) and the control and negative control groups. At this stage it seems that despite having high MIT content in the MSE the high dose MSE treatment in SH-SY5Y cells does not activate caspase enzymes.
MSE and control groups implying that this cell line expresses p53 protein and the lost of p53 protein seen at high doses was due to treatment effects. Parallel immuno blotting experiments were also carried out for MIT as shown in fig. There was no significant difference in the p53 levels noted over the dose range used however they appeared to be down regulated compared to the control group. The time course of MIT induced p53 change was also carried as shown in fig.