Protective effects and mechanism of Panax Notoginseng saponins on oxidative stress-induced damage and apoptosis of rabbit bone marrow stromal cells.
OBJECTIVE
To investigate the effects and possible mechanism of Panax Notoginseng saponins (PNS) on oxidative stress-induced damage and apoptosis in bone marrow stromal cells (BMSCs).


METHODS
BMSCs were isolated and cultured from 2-month-old New Zealand rabbits by the density gradient centrifugation combined with adherent method. The third passage cells were used for subsequent experiments. Oxidative stress was induced in cultured BMSCs by H(2)O(2) (0.1 mmol/L). BMSCs were pretreated with 25-200 μg/mL PNS for 4 h before H(2)O(2) treatment. Proliferation of BMSCs was observed using MTT assay. Alkaline phosphatase (ALP) activity, as an index of early osteoblastic differentiation, was determined with an ALP assay kit. Flow cytometry was used to observe the apoptosis of BMSCs by staining with annexinV-FITC/propidium iodide. Oxidative stress level was examined by reactive oxygen species (ROS) assay. The protein expressions of Bax, Bcl-2 and Caspase-3 in BMSCs were analyzed by Western blotting.


RESULTS
PNS had different concentration-dependent effects on proliferation and osteoblast differentiation of BMSCs induced by H(2)O(2). A PNS concentration of 100 μg/mL was determined as the optimal effective concentration. PNS markedly attenuated H(2)O(2)-induced apoptosis rate from 41.91% to 14.67% (P<0.01). PNS significantly decreased ROS level induced by H(2)O(2) (P<0.01). Furthermore, pretreatment with PNS significantly reversed H(2)O(2)-induced inhibition of Bcl-2 expression and augmentation of Bax and Caspase-3 expression (P<0.01).


CONCLUSIONS
PNS had a protective effect on oxidative stress-induced damage and apoptosis in cultured rabbit BMSCs through scavenging ROS and regulating the Bcl-2/Bax pathway.
Department of Orthopaedics, Shaanxi Provincial People's Hospital, the Third Affiliated Hospital, Medical School of Xi'an Jiaotong University, Xi'an 710068, China.
Hui
H
Qiang
Chen
C
Zhang
Zhi-bin
ZB
Shi
Hua-qing
HQ
Yang
Kun-zheng
KZ
Wang
D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D001854:Bone Marrow Cells / Q000166:cytology* / Q000187:drug effects* / Q000201:enzymology; D053148:Caspase 3 / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D005434:Flow Cytometry; D005452:Fluoresceins / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000494:pharmacology; D042541:Intracellular Space / Q000187:drug effects / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects*; D053723:Panax notoginseng / Q000737:chemistry*; D020011:Protective Agents / Q000494:pharmacology*; D011817:Rabbits; D017382:Reactive Oxygen Species / Q000378:metabolism; D012503:Saponins / Q000494:pharmacology*; D017154:Stromal Cells / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D051028:bcl-2-Associated X Protein / Q000378:metabolism
D005452:Fluoresceins; D020011:Protective Agents; D017382:Reactive Oxygen Species; D012503:Saponins; D051028:bcl-2-Associated X Protein; C029569:diacetyldichlorofluorescein; D006861:Hydrogen Peroxide; D000469:Alkaline Phosphatase; D053148:Caspase 3