Inhibition of ROS production in peripheral blood mononuclear cells from type 2 diabetic patients by autologous plasma depends on Akt/PKB signaling pathway.
OBJECTIVE
To compare the role of Akt/PKB signaling pathway in the modulation of reactive oxygen species (ROS) production by autologous plasma in peripheral blood mononuclear cells (PBMNC) from type 2 diabetic patients and healthy subjects.


METHODS
This study was approved by Santa Casa Ethical Committee and has included patients diagnosed with diabetes type 2 (DM2) and control group (non-diabetic) (ND). PBMNC were purified utilizing Ficoll-hypaque gradient. ROS was quantified by luminol-dependent chemiluminescence. The Akt/PKB phosphorylation was measured using a CASE kit. Statistical analyses were made with t Student test and chi-square (chi(2)). p<0.05 was considered significant.


RESULTS
12, 13-Phorbol dibutyrate (PDB) stimulated the production of higher levels of ROS in PBMNC from type 2 diabetic patients than that from healthy subjects. Autologous plasma, however, inhibited induced or not ROS production in PBMNC in both groups. The inhibition of PBMNC-ROS derived by autologous plasma from healthy subjects was higher than that from type 2 diabetic patients. Plasma phosphorylated (activated) Akt/PKB. The percentage of phosphorylation induced by autologous plasma in PBMNC from patients and healthy control were 14% and 93%, respectively. Inhibition of ROS production in PBMNC from DM2 were similar for PBMNC+plasma; PBMNC+Akti; and PBMNC+plasma+Akti. However, in ND control, plasma showed a higher ROS inhibition than Akti or plasma plus Akti.


CONCLUSIONS
Our results suggest that the low antioxidant capacity observed in autologous plasma from DM2 patients in conjunction with the decreased activation of PKB may cause an imbalance in the oxidizing/reducing responses, possible inducing an oxidative stress state, which could be associated with tissular damage.
Veloso
C A
CA
Núcleo de Pós-Graduação e Pesquisa, Hospital Santa Casa de Belo Horizonte, Belo Horizonte, Minas Gerais, Brazil.
Isoni
C A
CA
Borges
E A
EA
Mattos
R T
RT
Calsolari
M R
MR
Reis
J S
JS
Bosco
A A
AA
Chaves
M M
MM
Nogueira-Machado
J A
JA
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D003924:Diabetes Mellitus, Type 2 / Q000378:metabolism*; D005260:Female; D006801:Humans; D007962:Leukocytes / Q000187:drug effects / Q000378:metabolism*; D008875:Middle Aged; D019255:NADPH Oxidases / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism*; D015398:Signal Transduction
D017382:Reactive Oxygen Species; D019255:NADPH Oxidases; D051057:Proto-Oncogene Proteins c-akt; D011493:Protein Kinase C