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Biological effect of ketamine in urothelial cell lines and global gene expression analysis in the bladders of ketamine‑injected mice.

Shen CH, Wang ST, Lee YR, Liu SY, Li YZ, Wu JD, Chen YJ, Liu YW - Mol Med Rep (2014)

Bottom Line: Ketamine arrested the cells in G1 phase and increased the sub‑G1 population, and also increased the barrier permeability of these cell lines.Global gene expression analysis of the animals' bladders following data screening identified ten upregulated genes and 36 downregulated genes induced by ketamine.A total of 52% of keratin family genes were downregulated, particularly keratin 6a, 13 and 14, which was confirmed by polymerase chain reaction analysis.

View Article: PubMed Central - PubMed

Affiliation: Department of Urology, Chiayi Christian Hospital, Chiayi 600, Taiwan, R.O.C.

ABSTRACT
Ketamine is used clinically for anesthesia but is also abused as a recreational drug. Previously, it has been established that ketamine‑induced bladder interstitial cystitis is a common syndrome in ketamine‑abusing individuals. As the mechanisms underlying ketamine‑induced cystitis have yet to be revealed, the present study investigated the effect of ketamine on human urothelial cell lines and utilized a ketamine‑injected mouse model to identify ketamine‑induced changes in gene expression in mice bladders. In the in vitro bladder cell line assay, ketamine induced cytotoxicity in a dose‑ and time‑dependent manner. Ketamine arrested the cells in G1 phase and increased the sub‑G1 population, and also increased the barrier permeability of these cell lines. In the ketamine‑injected mouse model, ketamine did not change the body weight and bladder histology of the animals at the dose of 30 mg/kg/day for 60 days. Global gene expression analysis of the animals' bladders following data screening identified ten upregulated genes and 36 downregulated genes induced by ketamine. A total of 52% of keratin family genes were downregulated, particularly keratin 6a, 13 and 14, which was confirmed by polymerase chain reaction analysis. Keratin 14 protein, one of the 36 ketamine‑induced downregulated genes, was also reduced in the ketamine‑treated mouse bladder, as determined by immunohistochemical analysis. This suggested that cytotoxicity and keratin gene downregulation may have a critical role in ketamine‑induced cystitis.

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Cytotoxicity of ketamine on SV-HUC1, RT4 and 5637 cells. (A) Cytotoxicity of 0–8 mM ketamine. The cells were treated with 0–8 mM ketamine for 24 h (black bar) and 48 h (gray bar), and the cell viability was analyzed by an MTT assay. (B–D) Cell cycle distribution changed following incubation with 0–4 mM ketamine for 24 and 48 h. The cells were collected for cell cycle analysis following ketamine treatment in (B) SV-HUC1, (C) RT4 and (D) 5637 cells. Quantification performed from three independent experiments. *P<0.05, **P<0.01, ***P<0.001, significant difference between the control and ketamine-treated cells.
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f1-mmr-11-02-0887: Cytotoxicity of ketamine on SV-HUC1, RT4 and 5637 cells. (A) Cytotoxicity of 0–8 mM ketamine. The cells were treated with 0–8 mM ketamine for 24 h (black bar) and 48 h (gray bar), and the cell viability was analyzed by an MTT assay. (B–D) Cell cycle distribution changed following incubation with 0–4 mM ketamine for 24 and 48 h. The cells were collected for cell cycle analysis following ketamine treatment in (B) SV-HUC1, (C) RT4 and (D) 5637 cells. Quantification performed from three independent experiments. *P<0.05, **P<0.01, ***P<0.001, significant difference between the control and ketamine-treated cells.

Mentions: Following ketamine treatment for 24 h, the IC50 value of ketamine was ~4, 2 and 3 mM in SV-HUC-1, RT4 and 5637 cells, respectively. At 48 h, the IC50 was ~3, 1.5 and 2 mM in the SV-HUC-1, RT4 and 5637 cells, respectively (Fig. 1A). These results suggested that ketamine is cytotoxic to urotheliums in a dose-dependent and time-dependent manner. Due to the identified cytotoxicity, ketamine-induced cell cycle changes were analyzed. In the SV-HUC-1 cells, ketamine dose-dependently increased the G1 phase cells at a dose higher than 1 mM and significantly increased the sub-G1 level at 4 mM (Fig. 1B). In the RT4 (Fig. 1C) and 5637 (Fig. 1D) cells, ketamine also arrested the cells in the G1 phase between 1 to 2 mM, and significantly increased the sub-G1 level at 4 mM. All of the above data suggested that ketamine induced G1 arrest and cytotoxicity in the human urothelial cells.


Biological effect of ketamine in urothelial cell lines and global gene expression analysis in the bladders of ketamine‑injected mice.

Shen CH, Wang ST, Lee YR, Liu SY, Li YZ, Wu JD, Chen YJ, Liu YW - Mol Med Rep (2014)

Cytotoxicity of ketamine on SV-HUC1, RT4 and 5637 cells. (A) Cytotoxicity of 0–8 mM ketamine. The cells were treated with 0–8 mM ketamine for 24 h (black bar) and 48 h (gray bar), and the cell viability was analyzed by an MTT assay. (B–D) Cell cycle distribution changed following incubation with 0–4 mM ketamine for 24 and 48 h. The cells were collected for cell cycle analysis following ketamine treatment in (B) SV-HUC1, (C) RT4 and (D) 5637 cells. Quantification performed from three independent experiments. *P<0.05, **P<0.01, ***P<0.001, significant difference between the control and ketamine-treated cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4262485&req=5

f1-mmr-11-02-0887: Cytotoxicity of ketamine on SV-HUC1, RT4 and 5637 cells. (A) Cytotoxicity of 0–8 mM ketamine. The cells were treated with 0–8 mM ketamine for 24 h (black bar) and 48 h (gray bar), and the cell viability was analyzed by an MTT assay. (B–D) Cell cycle distribution changed following incubation with 0–4 mM ketamine for 24 and 48 h. The cells were collected for cell cycle analysis following ketamine treatment in (B) SV-HUC1, (C) RT4 and (D) 5637 cells. Quantification performed from three independent experiments. *P<0.05, **P<0.01, ***P<0.001, significant difference between the control and ketamine-treated cells.
Mentions: Following ketamine treatment for 24 h, the IC50 value of ketamine was ~4, 2 and 3 mM in SV-HUC-1, RT4 and 5637 cells, respectively. At 48 h, the IC50 was ~3, 1.5 and 2 mM in the SV-HUC-1, RT4 and 5637 cells, respectively (Fig. 1A). These results suggested that ketamine is cytotoxic to urotheliums in a dose-dependent and time-dependent manner. Due to the identified cytotoxicity, ketamine-induced cell cycle changes were analyzed. In the SV-HUC-1 cells, ketamine dose-dependently increased the G1 phase cells at a dose higher than 1 mM and significantly increased the sub-G1 level at 4 mM (Fig. 1B). In the RT4 (Fig. 1C) and 5637 (Fig. 1D) cells, ketamine also arrested the cells in the G1 phase between 1 to 2 mM, and significantly increased the sub-G1 level at 4 mM. All of the above data suggested that ketamine induced G1 arrest and cytotoxicity in the human urothelial cells.

Bottom Line: Ketamine arrested the cells in G1 phase and increased the sub‑G1 population, and also increased the barrier permeability of these cell lines.Global gene expression analysis of the animals' bladders following data screening identified ten upregulated genes and 36 downregulated genes induced by ketamine.A total of 52% of keratin family genes were downregulated, particularly keratin 6a, 13 and 14, which was confirmed by polymerase chain reaction analysis.

View Article: PubMed Central - PubMed

Affiliation: Department of Urology, Chiayi Christian Hospital, Chiayi 600, Taiwan, R.O.C.

ABSTRACT
Ketamine is used clinically for anesthesia but is also abused as a recreational drug. Previously, it has been established that ketamine‑induced bladder interstitial cystitis is a common syndrome in ketamine‑abusing individuals. As the mechanisms underlying ketamine‑induced cystitis have yet to be revealed, the present study investigated the effect of ketamine on human urothelial cell lines and utilized a ketamine‑injected mouse model to identify ketamine‑induced changes in gene expression in mice bladders. In the in vitro bladder cell line assay, ketamine induced cytotoxicity in a dose‑ and time‑dependent manner. Ketamine arrested the cells in G1 phase and increased the sub‑G1 population, and also increased the barrier permeability of these cell lines. In the ketamine‑injected mouse model, ketamine did not change the body weight and bladder histology of the animals at the dose of 30 mg/kg/day for 60 days. Global gene expression analysis of the animals' bladders following data screening identified ten upregulated genes and 36 downregulated genes induced by ketamine. A total of 52% of keratin family genes were downregulated, particularly keratin 6a, 13 and 14, which was confirmed by polymerase chain reaction analysis. Keratin 14 protein, one of the 36 ketamine‑induced downregulated genes, was also reduced in the ketamine‑treated mouse bladder, as determined by immunohistochemical analysis. This suggested that cytotoxicity and keratin gene downregulation may have a critical role in ketamine‑induced cystitis.

Show MeSH
Related in: MedlinePlus