Limits...
Shear stress attenuates apoptosis due to TNFα, oxidative stress, and serum depletion via death-associated protein kinase (DAPK) expression.

Rennier K, Ji JY - BMC Res Notes (2015)

Bottom Line: This is correlated with a parallel decrease of DAPK expression and caspase activity compared to non-sheared cells.Interestingly, shear stress applied to cells prior to induction with apoptosis agents resulted in a higher suppression of apoptosis and DAPK and caspase activity, compared to applying shear stress post induction.Also, shear stress alone also induced higher apoptosis and DAPK expression, and the effect is sustained even after 18 hrs incubation in static condition, compared to non-sheared cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedical Engineering, Indiana University Purdue University Indianapolis, 723 West Michigan Street, SL-220 J, Indianapolis, IN, 46202, USA. krennier@purdue.edu.

ABSTRACT

Background: Misdirected apoptosis in endothelial cells participates in the development of pathological conditions such as atherosclerosis. Tight regulation of apoptosis is necessary to ensure normal cell function. The rate of cell turnover is increased at sites prone to lesion development. Laminar shear stress is protective against atherosclerosis, and helps suppress apoptosis induced by cytokines, oxidative stress, and serum depletion. Current Studies have shown that the pro-apoptotic DAPK expression and function to be regulated in part by shear stress, and that shearing cells already treated with cytokine tumor necrosis factor (TNF) α significantly reduced apoptosis. We investigate further the suppression of endothelial apoptosis by shear stress with other apoptotic triggers, and the involvement of DAPK and caspase 3/7.

Results: We have shown that exposure to shear stress (12 dynes/cm(2) for 6 hrs) suppressed endothelial apoptosis triggered by cytokine (TNFα), oxidative stress (H2O2), and serum depletion, either before or after a long term (18 hr) induction. This is correlated with a parallel decrease of DAPK expression and caspase activity compared to non-sheared cells. We found similar modulation of DAPK and apoptosis by shear stress with other pro-apoptotic signals. Changes in DAPK and caspase 3/7 are directly correlated to changes in apoptosis. Interestingly, shear stress applied to cells prior to induction with apoptosis agents resulted in a higher suppression of apoptosis and DAPK and caspase activity, compared to applying shear stress post induction. This is correlated with a higher expression and activation of DAPK in cells sheared at the end of 24-hr experiment. Also, shear stress alone also induced higher apoptosis and DAPK expression, and the effect is sustained even after 18 hrs incubation in static condition, compared to non-sheared cells.

Conclusions: Overall, we show that laminar shear stress inhibits various apoptosis pathways by modulating DAPK activity, as well as caspase activation, in a time-dependent manner. Shear stress could target DAPK as a converging point to exert its effects of suppressing endothelial apoptosis. The temporal shear stress stimulation of DAPK and its role in different apoptosis pathways may help identify key mechanisms of the endothelial mechanotransduction pathway.

Show MeSH

Related in: MedlinePlus

Quantified cell apoptosis results using TUNEL staining. A: Analysis of TUNEL in pre-sheared cells versus cells treated with stimulus (TNFα, H2O2, or serum depletion). B: Analysis of TUNEL results in cells sheared post treatment with stimulus (TNFα, H2O2, or serum depletion). For all apoptosis data: * P < 0.05 compared to Control BAEC, + P < 0.05 compared to Static + TNFα, # P < 0.05 compared to Static + H2O2, ∆ P < 0.05 compared to Static – Serum.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC4374420&req=5

Fig4: Quantified cell apoptosis results using TUNEL staining. A: Analysis of TUNEL in pre-sheared cells versus cells treated with stimulus (TNFα, H2O2, or serum depletion). B: Analysis of TUNEL results in cells sheared post treatment with stimulus (TNFα, H2O2, or serum depletion). For all apoptosis data: * P < 0.05 compared to Control BAEC, + P < 0.05 compared to Static + TNFα, # P < 0.05 compared to Static + H2O2, ∆ P < 0.05 compared to Static – Serum.

Mentions: We added TNFα (25 ng/ml), H2O2 (0.5 mM), and serum depletion (0.5% FBS) to induce apoptosis, in combination with shear stress conditioning (12 dynes/cm2). Our apoptosis study combined long term inductions (18 hrs) with either pre-shearing or subsequent post-shearing for 6 hrs, to examine the suppression of apoptosis due to shear stress. Apoptosis was analyzed by TUNEL staining, quantified using flow cytometry, and the fold increase in apoptosis for each sample is calculated (Figure 4). Each apoptotic stimulus induced an approximately two-fold increase in endothelial apoptosis compared to control BAECs under static condition. Fold induction of apoptosis ranged from 2.24 to 2.96-fold increase. Conversely, shear stress exposure either before or after cytokine or biochemical induction significantly decreased overall apoptosis compared to static cells under the same conditions (Figure 4).Figure 4


Shear stress attenuates apoptosis due to TNFα, oxidative stress, and serum depletion via death-associated protein kinase (DAPK) expression.

Rennier K, Ji JY - BMC Res Notes (2015)

Quantified cell apoptosis results using TUNEL staining. A: Analysis of TUNEL in pre-sheared cells versus cells treated with stimulus (TNFα, H2O2, or serum depletion). B: Analysis of TUNEL results in cells sheared post treatment with stimulus (TNFα, H2O2, or serum depletion). For all apoptosis data: * P < 0.05 compared to Control BAEC, + P < 0.05 compared to Static + TNFα, # P < 0.05 compared to Static + H2O2, ∆ P < 0.05 compared to Static – Serum.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4374420&req=5

Fig4: Quantified cell apoptosis results using TUNEL staining. A: Analysis of TUNEL in pre-sheared cells versus cells treated with stimulus (TNFα, H2O2, or serum depletion). B: Analysis of TUNEL results in cells sheared post treatment with stimulus (TNFα, H2O2, or serum depletion). For all apoptosis data: * P < 0.05 compared to Control BAEC, + P < 0.05 compared to Static + TNFα, # P < 0.05 compared to Static + H2O2, ∆ P < 0.05 compared to Static – Serum.
Mentions: We added TNFα (25 ng/ml), H2O2 (0.5 mM), and serum depletion (0.5% FBS) to induce apoptosis, in combination with shear stress conditioning (12 dynes/cm2). Our apoptosis study combined long term inductions (18 hrs) with either pre-shearing or subsequent post-shearing for 6 hrs, to examine the suppression of apoptosis due to shear stress. Apoptosis was analyzed by TUNEL staining, quantified using flow cytometry, and the fold increase in apoptosis for each sample is calculated (Figure 4). Each apoptotic stimulus induced an approximately two-fold increase in endothelial apoptosis compared to control BAECs under static condition. Fold induction of apoptosis ranged from 2.24 to 2.96-fold increase. Conversely, shear stress exposure either before or after cytokine or biochemical induction significantly decreased overall apoptosis compared to static cells under the same conditions (Figure 4).Figure 4

Bottom Line: This is correlated with a parallel decrease of DAPK expression and caspase activity compared to non-sheared cells.Interestingly, shear stress applied to cells prior to induction with apoptosis agents resulted in a higher suppression of apoptosis and DAPK and caspase activity, compared to applying shear stress post induction.Also, shear stress alone also induced higher apoptosis and DAPK expression, and the effect is sustained even after 18 hrs incubation in static condition, compared to non-sheared cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedical Engineering, Indiana University Purdue University Indianapolis, 723 West Michigan Street, SL-220 J, Indianapolis, IN, 46202, USA. krennier@purdue.edu.

ABSTRACT

Background: Misdirected apoptosis in endothelial cells participates in the development of pathological conditions such as atherosclerosis. Tight regulation of apoptosis is necessary to ensure normal cell function. The rate of cell turnover is increased at sites prone to lesion development. Laminar shear stress is protective against atherosclerosis, and helps suppress apoptosis induced by cytokines, oxidative stress, and serum depletion. Current Studies have shown that the pro-apoptotic DAPK expression and function to be regulated in part by shear stress, and that shearing cells already treated with cytokine tumor necrosis factor (TNF) α significantly reduced apoptosis. We investigate further the suppression of endothelial apoptosis by shear stress with other apoptotic triggers, and the involvement of DAPK and caspase 3/7.

Results: We have shown that exposure to shear stress (12 dynes/cm(2) for 6 hrs) suppressed endothelial apoptosis triggered by cytokine (TNFα), oxidative stress (H2O2), and serum depletion, either before or after a long term (18 hr) induction. This is correlated with a parallel decrease of DAPK expression and caspase activity compared to non-sheared cells. We found similar modulation of DAPK and apoptosis by shear stress with other pro-apoptotic signals. Changes in DAPK and caspase 3/7 are directly correlated to changes in apoptosis. Interestingly, shear stress applied to cells prior to induction with apoptosis agents resulted in a higher suppression of apoptosis and DAPK and caspase activity, compared to applying shear stress post induction. This is correlated with a higher expression and activation of DAPK in cells sheared at the end of 24-hr experiment. Also, shear stress alone also induced higher apoptosis and DAPK expression, and the effect is sustained even after 18 hrs incubation in static condition, compared to non-sheared cells.

Conclusions: Overall, we show that laminar shear stress inhibits various apoptosis pathways by modulating DAPK activity, as well as caspase activation, in a time-dependent manner. Shear stress could target DAPK as a converging point to exert its effects of suppressing endothelial apoptosis. The temporal shear stress stimulation of DAPK and its role in different apoptosis pathways may help identify key mechanisms of the endothelial mechanotransduction pathway.

Show MeSH
Related in: MedlinePlus