P0942HYPOXEMIA AND UREMIA INDUCE OXIDATIVE IMBALANCE IN ENDOTHELIAL CELLS
Abstract Background and Aims Intradialytic hypoxemia is associated with oxidative stress, endothelial dysfunction, inflammation, higher erythropoietin requirements, and higher all-cause hospitalization and mortality in hemodialysis (HD) patients (Meyring-Wosten, et al., 2016). The aim of the present...
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| description | Abstract
Background and Aims
Intradialytic hypoxemia is associated with oxidative stress, endothelial dysfunction, inflammation, higher erythropoietin requirements, and higher all-cause hospitalization and mortality in hemodialysis (HD) patients (Meyring-Wosten, et al., 2016). The aim of the present study was to further expand our insights into the hypoxia-uremia axis by investigating the oxidative balance in endothelial cells (EC) under hypoxic and uremic conditions.
Method
Human umbilical EC were incubated with DMEM medium supplemented with 10% of fetal bovine serum (FBS; control) or with sera obtained from healthy subjects (S-CON) or HD patients (S-HD, 1:10), respectively, for 40 minutes, 4 hours, and 24 hours under normoxic (21% O2) or hypoxic (5% O2) conditions (Culture Chamber ProOx, Biospherix). EC were analyzed by flow cytometer (BD Accuri™ C6 Plus) to assess a) intracellular production of reactive oxygen species (ROS, DCFH-DA probe, Abcam); b) reduced glutathione (GSH) content (ThiolTracker Violet probe, Thermo Fisher Scientific).
Results
S-HD increased intracellular ROS production at all time points compared to S-CON. Moreover, ROS production was higher under hypoxic conditions. ROS production declined after 24 hours. S-HD induced slightly higher GSH content when compared to S-CON in normoxia (Table 1).
Conclusion
In our in vitro experiments, hypoxia and uremia jointly favor EC oxidative imbalance. This effect is particularly pronounced after 4 hours. Translational studies are warranted to explore the clinical relevance of our findings.
Table:
Baseline (of 100%): EC incubated in DMEM medium supplemented with 10% FBS + antibiotics
Normoxia (21% O2)
Hypoxia (5% O2)
ROS production [%]
28.9±2.4
52.3±7.2
GSH content [%]
8.4±1
8.1±0.8
Values after Incubation with S-CON or S-HD
40 minutes
4 hours
24 hours
ROS production
Normoxia
Hypoxia
Normoxia
Hypoxia
Normoxia
Hypoxia
S-CON [%]
102±5.2
121±6.32&
102.5±7.3
109.2±9.2
34.2±7.3
50.5±14.5
S-HD [%]
130.7±8.7*
146.2±8.3*&
150±13.9*
190.3±37.6*
71.8±11.4*
94.8±19.3*&
GSH content
Normoxia
Hypoxia
Normoxia
Hypoxia
Normoxia
Hypoxia
S-CON [%]
8.9±1.3&
6.5±2.3
7.6±1.2
7.4±1.6
7.9±1.5
8.1±1.2
S-HD [%]
12.7±1.8*&
6.3±0.9
9.3±1.6*
9.8±1.8*
10.4±1.3*&
7.5±0.8
*
p |
| doi_str_mv | 10.1093/ndt/gfaa142.P0942 |
| format | Article |
| fullrecord | <record><control><sourceid>oup_cross</sourceid><recordid>TN_cdi_crossref_primary_10_1093_ndt_gfaa142_P0942</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><oup_id>10.1093/ndt/gfaa142.P0942</oup_id><sourcerecordid>10.1093/ndt/gfaa142.P0942</sourcerecordid><originalsourceid>FETCH-LOGICAL-c1222-d5105164c246306efdd56489d2858dc515c6a1ab69b05b26fb523b815430fe7d3</originalsourceid><addsrcrecordid>eNqNkMFOhDAQhhujibj6AN54ANmdKW2BY4UqTbqwUTDriQAFo1F3A3rw7UV2H8DT_Jl8_yTzEXKNsESI_NWn_Vq99HWNjC43EDF6QhxkAjzqh_yUOBODHnCIzsnFOL4BQESDwCF6htPnTb5Vay1dmSVu-TBHnSVlrNx8qxNZ6Cfl6vWtNDKbdjpzVZbkRaqMlsaNlTGPl-Ssr9_H7uo4F6S8U0Wceia_17E0XouUUs9yBI6CtZQJH0TXW8sFCyNLQx7aliNvRY11I6IGeENF33DqNyFy5kPfBdZfEDzcbYfdOA5dX-2H1496-KkQqj8X1eSiOrqo5vemzs2hs_ve_wP_BU53WlI</addsrcrecordid><sourcetype>Aggregation Database</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype></control><display><type>article</type><title>P0942HYPOXEMIA AND UREMIA INDUCE OXIDATIVE IMBALANCE IN ENDOTHELIAL CELLS</title><source>Oxford University Press Journals All Titles (1996-Current)</source><source>EZB-FREE-00999 freely available EZB journals</source><source>Alma/SFX Local Collection</source><creator>Moreno-Amaral, Andrea N ; Carvalho Silva, Caroline ; Andrade, Gabriela Bohnen ; Monteiro, Julia ; Grobe, Nadja ; Silva Pecoits Filho, Roberto Flavio ; Stinghen, Andréa E M ; Kotanko, Peter</creator><creatorcontrib>Moreno-Amaral, Andrea N ; Carvalho Silva, Caroline ; Andrade, Gabriela Bohnen ; Monteiro, Julia ; Grobe, Nadja ; Silva Pecoits Filho, Roberto Flavio ; Stinghen, Andréa E M ; Kotanko, Peter</creatorcontrib><description><![CDATA[Abstract
Background and Aims
Intradialytic hypoxemia is associated with oxidative stress, endothelial dysfunction, inflammation, higher erythropoietin requirements, and higher all-cause hospitalization and mortality in hemodialysis (HD) patients (Meyring-Wosten, et al., 2016). The aim of the present study was to further expand our insights into the hypoxia-uremia axis by investigating the oxidative balance in endothelial cells (EC) under hypoxic and uremic conditions.
Method
Human umbilical EC were incubated with DMEM medium supplemented with 10% of fetal bovine serum (FBS; control) or with sera obtained from healthy subjects (S-CON) or HD patients (S-HD, 1:10), respectively, for 40 minutes, 4 hours, and 24 hours under normoxic (21% O2) or hypoxic (5% O2) conditions (Culture Chamber ProOx, Biospherix). EC were analyzed by flow cytometer (BD Accuri™ C6 Plus) to assess a) intracellular production of reactive oxygen species (ROS, DCFH-DA probe, Abcam); b) reduced glutathione (GSH) content (ThiolTracker Violet probe, Thermo Fisher Scientific).
Results
S-HD increased intracellular ROS production at all time points compared to S-CON. Moreover, ROS production was higher under hypoxic conditions. ROS production declined after 24 hours. S-HD induced slightly higher GSH content when compared to S-CON in normoxia (Table 1).
Conclusion
In our in vitro experiments, hypoxia and uremia jointly favor EC oxidative imbalance. This effect is particularly pronounced after 4 hours. Translational studies are warranted to explore the clinical relevance of our findings.
Table:
Baseline (of 100%): EC incubated in DMEM medium supplemented with 10% FBS + antibiotics
Normoxia (21% O2)
Hypoxia (5% O2)
ROS production [%]
28.9±2.4
52.3±7.2
GSH content [%]
8.4±1
8.1±0.8
Values after Incubation with S-CON or S-HD
40 minutes
4 hours
24 hours
ROS production
Normoxia
Hypoxia
Normoxia
Hypoxia
Normoxia
Hypoxia
S-CON [%]
102±5.2
121±6.32&
102.5±7.3
109.2±9.2
34.2±7.3
50.5±14.5
S-HD [%]
130.7±8.7*
146.2±8.3*&
150±13.9*
190.3±37.6*
71.8±11.4*
94.8±19.3*&
GSH content
Normoxia
Hypoxia
Normoxia
Hypoxia
Normoxia
Hypoxia
S-CON [%]
8.9±1.3&
6.5±2.3
7.6±1.2
7.4±1.6
7.9±1.5
8.1±1.2
S-HD [%]
12.7±1.8*&
6.3±0.9
9.3±1.6*
9.8±1.8*
10.4±1.3*&
7.5±0.8
*
p <0.05 compared to control for the same O2 concentration. & p<0.05 normoxia vs hypoxia.]]></description><identifier>ISSN: 0931-0509</identifier><identifier>EISSN: 1460-2385</identifier><identifier>DOI: 10.1093/ndt/gfaa142.P0942</identifier><language>eng</language><publisher>Oxford University Press</publisher><ispartof>Nephrology, dialysis, transplantation, 2020-06, Vol.35 (Supplement_3)</ispartof><rights>The Author(s) 2020. Published by Oxford University Press on behalf of ERA-EDTA. All rights reserved 2020</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,776,780,1578,27901,27902</link.rule.ids></links><search><creatorcontrib>Moreno-Amaral, Andrea N</creatorcontrib><creatorcontrib>Carvalho Silva, Caroline</creatorcontrib><creatorcontrib>Andrade, Gabriela Bohnen</creatorcontrib><creatorcontrib>Monteiro, Julia</creatorcontrib><creatorcontrib>Grobe, Nadja</creatorcontrib><creatorcontrib>Silva Pecoits Filho, Roberto Flavio</creatorcontrib><creatorcontrib>Stinghen, Andréa E M</creatorcontrib><creatorcontrib>Kotanko, Peter</creatorcontrib><title>P0942HYPOXEMIA AND UREMIA INDUCE OXIDATIVE IMBALANCE IN ENDOTHELIAL CELLS</title><title>Nephrology, dialysis, transplantation</title><description><![CDATA[Abstract
Background and Aims
Intradialytic hypoxemia is associated with oxidative stress, endothelial dysfunction, inflammation, higher erythropoietin requirements, and higher all-cause hospitalization and mortality in hemodialysis (HD) patients (Meyring-Wosten, et al., 2016). The aim of the present study was to further expand our insights into the hypoxia-uremia axis by investigating the oxidative balance in endothelial cells (EC) under hypoxic and uremic conditions.
Method
Human umbilical EC were incubated with DMEM medium supplemented with 10% of fetal bovine serum (FBS; control) or with sera obtained from healthy subjects (S-CON) or HD patients (S-HD, 1:10), respectively, for 40 minutes, 4 hours, and 24 hours under normoxic (21% O2) or hypoxic (5% O2) conditions (Culture Chamber ProOx, Biospherix). EC were analyzed by flow cytometer (BD Accuri™ C6 Plus) to assess a) intracellular production of reactive oxygen species (ROS, DCFH-DA probe, Abcam); b) reduced glutathione (GSH) content (ThiolTracker Violet probe, Thermo Fisher Scientific).
Results
S-HD increased intracellular ROS production at all time points compared to S-CON. Moreover, ROS production was higher under hypoxic conditions. ROS production declined after 24 hours. S-HD induced slightly higher GSH content when compared to S-CON in normoxia (Table 1).
Conclusion
In our in vitro experiments, hypoxia and uremia jointly favor EC oxidative imbalance. This effect is particularly pronounced after 4 hours. Translational studies are warranted to explore the clinical relevance of our findings.
Table:
Baseline (of 100%): EC incubated in DMEM medium supplemented with 10% FBS + antibiotics
Normoxia (21% O2)
Hypoxia (5% O2)
ROS production [%]
28.9±2.4
52.3±7.2
GSH content [%]
8.4±1
8.1±0.8
Values after Incubation with S-CON or S-HD
40 minutes
4 hours
24 hours
ROS production
Normoxia
Hypoxia
Normoxia
Hypoxia
Normoxia
Hypoxia
S-CON [%]
102±5.2
121±6.32&
102.5±7.3
109.2±9.2
34.2±7.3
50.5±14.5
S-HD [%]
130.7±8.7*
146.2±8.3*&
150±13.9*
190.3±37.6*
71.8±11.4*
94.8±19.3*&
GSH content
Normoxia
Hypoxia
Normoxia
Hypoxia
Normoxia
Hypoxia
S-CON [%]
8.9±1.3&
6.5±2.3
7.6±1.2
7.4±1.6
7.9±1.5
8.1±1.2
S-HD [%]
12.7±1.8*&
6.3±0.9
9.3±1.6*
9.8±1.8*
10.4±1.3*&
7.5±0.8
*
p <0.05 compared to control for the same O2 concentration. & p<0.05 normoxia vs hypoxia.]]></description><issn>0931-0509</issn><issn>1460-2385</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2020</creationdate><recordtype>article</recordtype><recordid>eNqNkMFOhDAQhhujibj6AN54ANmdKW2BY4UqTbqwUTDriQAFo1F3A3rw7UV2H8DT_Jl8_yTzEXKNsESI_NWn_Vq99HWNjC43EDF6QhxkAjzqh_yUOBODHnCIzsnFOL4BQESDwCF6htPnTb5Vay1dmSVu-TBHnSVlrNx8qxNZ6Cfl6vWtNDKbdjpzVZbkRaqMlsaNlTGPl-Ssr9_H7uo4F6S8U0Wceia_17E0XouUUs9yBI6CtZQJH0TXW8sFCyNLQx7aliNvRY11I6IGeENF33DqNyFy5kPfBdZfEDzcbYfdOA5dX-2H1496-KkQqj8X1eSiOrqo5vemzs2hs_ve_wP_BU53WlI</recordid><startdate>20200601</startdate><enddate>20200601</enddate><creator>Moreno-Amaral, Andrea N</creator><creator>Carvalho Silva, Caroline</creator><creator>Andrade, Gabriela Bohnen</creator><creator>Monteiro, Julia</creator><creator>Grobe, Nadja</creator><creator>Silva Pecoits Filho, Roberto Flavio</creator><creator>Stinghen, Andréa E M</creator><creator>Kotanko, Peter</creator><general>Oxford University Press</general><scope>AAYXX</scope><scope>CITATION</scope></search><sort><creationdate>20200601</creationdate><title>P0942HYPOXEMIA AND UREMIA INDUCE OXIDATIVE IMBALANCE IN ENDOTHELIAL CELLS</title><author>Moreno-Amaral, Andrea N ; Carvalho Silva, Caroline ; Andrade, Gabriela Bohnen ; Monteiro, Julia ; Grobe, Nadja ; Silva Pecoits Filho, Roberto Flavio ; Stinghen, Andréa E M ; Kotanko, Peter</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c1222-d5105164c246306efdd56489d2858dc515c6a1ab69b05b26fb523b815430fe7d3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2020</creationdate><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Moreno-Amaral, Andrea N</creatorcontrib><creatorcontrib>Carvalho Silva, Caroline</creatorcontrib><creatorcontrib>Andrade, Gabriela Bohnen</creatorcontrib><creatorcontrib>Monteiro, Julia</creatorcontrib><creatorcontrib>Grobe, Nadja</creatorcontrib><creatorcontrib>Silva Pecoits Filho, Roberto Flavio</creatorcontrib><creatorcontrib>Stinghen, Andréa E M</creatorcontrib><creatorcontrib>Kotanko, Peter</creatorcontrib><collection>CrossRef</collection><jtitle>Nephrology, dialysis, transplantation</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Moreno-Amaral, Andrea N</au><au>Carvalho Silva, Caroline</au><au>Andrade, Gabriela Bohnen</au><au>Monteiro, Julia</au><au>Grobe, Nadja</au><au>Silva Pecoits Filho, Roberto Flavio</au><au>Stinghen, Andréa E M</au><au>Kotanko, Peter</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>P0942HYPOXEMIA AND UREMIA INDUCE OXIDATIVE IMBALANCE IN ENDOTHELIAL CELLS</atitle><jtitle>Nephrology, dialysis, transplantation</jtitle><date>2020-06-01</date><risdate>2020</risdate><volume>35</volume><issue>Supplement_3</issue><issn>0931-0509</issn><eissn>1460-2385</eissn><abstract><![CDATA[Abstract
Background and Aims
Intradialytic hypoxemia is associated with oxidative stress, endothelial dysfunction, inflammation, higher erythropoietin requirements, and higher all-cause hospitalization and mortality in hemodialysis (HD) patients (Meyring-Wosten, et al., 2016). The aim of the present study was to further expand our insights into the hypoxia-uremia axis by investigating the oxidative balance in endothelial cells (EC) under hypoxic and uremic conditions.
Method
Human umbilical EC were incubated with DMEM medium supplemented with 10% of fetal bovine serum (FBS; control) or with sera obtained from healthy subjects (S-CON) or HD patients (S-HD, 1:10), respectively, for 40 minutes, 4 hours, and 24 hours under normoxic (21% O2) or hypoxic (5% O2) conditions (Culture Chamber ProOx, Biospherix). EC were analyzed by flow cytometer (BD Accuri™ C6 Plus) to assess a) intracellular production of reactive oxygen species (ROS, DCFH-DA probe, Abcam); b) reduced glutathione (GSH) content (ThiolTracker Violet probe, Thermo Fisher Scientific).
Results
S-HD increased intracellular ROS production at all time points compared to S-CON. Moreover, ROS production was higher under hypoxic conditions. ROS production declined after 24 hours. S-HD induced slightly higher GSH content when compared to S-CON in normoxia (Table 1).
Conclusion
In our in vitro experiments, hypoxia and uremia jointly favor EC oxidative imbalance. This effect is particularly pronounced after 4 hours. Translational studies are warranted to explore the clinical relevance of our findings.
Table:
Baseline (of 100%): EC incubated in DMEM medium supplemented with 10% FBS + antibiotics
Normoxia (21% O2)
Hypoxia (5% O2)
ROS production [%]
28.9±2.4
52.3±7.2
GSH content [%]
8.4±1
8.1±0.8
Values after Incubation with S-CON or S-HD
40 minutes
4 hours
24 hours
ROS production
Normoxia
Hypoxia
Normoxia
Hypoxia
Normoxia
Hypoxia
S-CON [%]
102±5.2
121±6.32&
102.5±7.3
109.2±9.2
34.2±7.3
50.5±14.5
S-HD [%]
130.7±8.7*
146.2±8.3*&
150±13.9*
190.3±37.6*
71.8±11.4*
94.8±19.3*&
GSH content
Normoxia
Hypoxia
Normoxia
Hypoxia
Normoxia
Hypoxia
S-CON [%]
8.9±1.3&
6.5±2.3
7.6±1.2
7.4±1.6
7.9±1.5
8.1±1.2
S-HD [%]
12.7±1.8*&
6.3±0.9
9.3±1.6*
9.8±1.8*
10.4±1.3*&
7.5±0.8
*
p <0.05 compared to control for the same O2 concentration. & p<0.05 normoxia vs hypoxia.]]></abstract><pub>Oxford University Press</pub><doi>10.1093/ndt/gfaa142.P0942</doi><oa>free_for_read</oa></addata></record> |
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| title | P0942HYPOXEMIA AND UREMIA INDUCE OXIDATIVE IMBALANCE IN ENDOTHELIAL CELLS |
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