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Inducible hydrogen sulfide synthesis in chondrocytes and mesenchymal progenitor cells: is H2S a novel cytoprotective mediator in the inflamed joint?

Fox B, Schantz JT, Haigh R, Wood ME, Moore PK, Viner N, Spencer JP, Winyard PG, Whiteman M - J. Cell. Mol. Med. (2012)

Bottom Line: Oxidative stress-induced cell death was significantly inhibited by GYY4137 treatment but increased by pharmacological inhibition of H(2)S synthesis or by CBS/CSE-siRNA treatment.These data suggest CSE is an inducible source of H(2)S in cultured HACs and MPCs.H(2)S may represent a novel endogenous mechanism of cytoprotection in the inflamed joint, suggesting a potential opportunity for therapeutic intervention.

View Article: PubMed Central - PubMed

Affiliation: Peninsula Medical School, University of Exeter, St. Luke's Campus, Exeter, Devon, UK.

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Effect of CSE and CBS protein knockdown on oxidative stress induced cell death. MPCs were treated with CSE-siRNA, CBS-siRNA or two non-coding controls (NCC-1 and NCC-2) as described in Materials and Methods. L-cysteine (1 mmol/l) was added as a substrate for CSE and CBS. GYY4137 (500 μmol/l) was added as a source of pharmacological (exogenous) H2S. Cell death was assessed by MTT assay after treatment with (A) SIN-1 (500 μmol/l), (B) H2O2 (200 μmol/l) and (C) 4-HNE (35 μmol/l). Data are expressed as mean ± S.D. of six determinations. ^^^P < 0.001, cf. L-cysteine only treatment, ++P < 0.01, +++P < 0.001, cf. siRNA + oxidant treatment.
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fig05: Effect of CSE and CBS protein knockdown on oxidative stress induced cell death. MPCs were treated with CSE-siRNA, CBS-siRNA or two non-coding controls (NCC-1 and NCC-2) as described in Materials and Methods. L-cysteine (1 mmol/l) was added as a substrate for CSE and CBS. GYY4137 (500 μmol/l) was added as a source of pharmacological (exogenous) H2S. Cell death was assessed by MTT assay after treatment with (A) SIN-1 (500 μmol/l), (B) H2O2 (200 μmol/l) and (C) 4-HNE (35 μmol/l). Data are expressed as mean ± S.D. of six determinations. ^^^P < 0.001, cf. L-cysteine only treatment, ++P < 0.01, +++P < 0.001, cf. siRNA + oxidant treatment.

Mentions: To confirm the molecular requirement for CSE- and CBS-derived H2S in mediating cytoprotection, we next examined the effects of siRNA-mediated CSE and CBS protein knockdown in chondrocytes. In CSE- and CBS-siRNA-treated cells the cytoprotective effect of L-cysteine against oxidative injury was significantly reduced but preserved in cells transfected with two non-coding siRNA (Fig. 5A and B). Small but statistically significant increases in cell death induced by SIN-1 (Fig. 5A), H2O2 (Fig. 5B) and 4-HNE (Fig. 5C) were observed with CSE and CBS-siRNA treatment, in agreement with the effects of PAG and AOAA in MPCs (Fig. 4C and D). Pharmacological H2S, supplied via GYY4137, further significantly inhibited oxidative stress-induced cell death when added to CSE-treated and CBS-siRNA-treated cells, that is, the lack of enzymatically generated H2S could be overcome by pharmacological H2S.


Inducible hydrogen sulfide synthesis in chondrocytes and mesenchymal progenitor cells: is H2S a novel cytoprotective mediator in the inflamed joint?

Fox B, Schantz JT, Haigh R, Wood ME, Moore PK, Viner N, Spencer JP, Winyard PG, Whiteman M - J. Cell. Mol. Med. (2012)

Effect of CSE and CBS protein knockdown on oxidative stress induced cell death. MPCs were treated with CSE-siRNA, CBS-siRNA or two non-coding controls (NCC-1 and NCC-2) as described in Materials and Methods. L-cysteine (1 mmol/l) was added as a substrate for CSE and CBS. GYY4137 (500 μmol/l) was added as a source of pharmacological (exogenous) H2S. Cell death was assessed by MTT assay after treatment with (A) SIN-1 (500 μmol/l), (B) H2O2 (200 μmol/l) and (C) 4-HNE (35 μmol/l). Data are expressed as mean ± S.D. of six determinations. ^^^P < 0.001, cf. L-cysteine only treatment, ++P < 0.01, +++P < 0.001, cf. siRNA + oxidant treatment.
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Related In: Results  -  Collection

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fig05: Effect of CSE and CBS protein knockdown on oxidative stress induced cell death. MPCs were treated with CSE-siRNA, CBS-siRNA or two non-coding controls (NCC-1 and NCC-2) as described in Materials and Methods. L-cysteine (1 mmol/l) was added as a substrate for CSE and CBS. GYY4137 (500 μmol/l) was added as a source of pharmacological (exogenous) H2S. Cell death was assessed by MTT assay after treatment with (A) SIN-1 (500 μmol/l), (B) H2O2 (200 μmol/l) and (C) 4-HNE (35 μmol/l). Data are expressed as mean ± S.D. of six determinations. ^^^P < 0.001, cf. L-cysteine only treatment, ++P < 0.01, +++P < 0.001, cf. siRNA + oxidant treatment.
Mentions: To confirm the molecular requirement for CSE- and CBS-derived H2S in mediating cytoprotection, we next examined the effects of siRNA-mediated CSE and CBS protein knockdown in chondrocytes. In CSE- and CBS-siRNA-treated cells the cytoprotective effect of L-cysteine against oxidative injury was significantly reduced but preserved in cells transfected with two non-coding siRNA (Fig. 5A and B). Small but statistically significant increases in cell death induced by SIN-1 (Fig. 5A), H2O2 (Fig. 5B) and 4-HNE (Fig. 5C) were observed with CSE and CBS-siRNA treatment, in agreement with the effects of PAG and AOAA in MPCs (Fig. 4C and D). Pharmacological H2S, supplied via GYY4137, further significantly inhibited oxidative stress-induced cell death when added to CSE-treated and CBS-siRNA-treated cells, that is, the lack of enzymatically generated H2S could be overcome by pharmacological H2S.

Bottom Line: Oxidative stress-induced cell death was significantly inhibited by GYY4137 treatment but increased by pharmacological inhibition of H(2)S synthesis or by CBS/CSE-siRNA treatment.These data suggest CSE is an inducible source of H(2)S in cultured HACs and MPCs.H(2)S may represent a novel endogenous mechanism of cytoprotection in the inflamed joint, suggesting a potential opportunity for therapeutic intervention.

View Article: PubMed Central - PubMed

Affiliation: Peninsula Medical School, University of Exeter, St. Luke's Campus, Exeter, Devon, UK.

Show MeSH
Related in: MedlinePlus