Obesity and hyperglycemia lead to impaired post-ischemic recovery after permanent ischemia in mice

Abstract

Objective: Obesity-induced diabetes has increased over the years and has become one of the risk factors for stroke. We investigated the influence of diet-induced obesity and hyperglycemia on permanent distal middle cerebral artery occlusion (pMCAO)-induced ischemic stroke in mice.

Methods: Male C57/Bl6 mice were treated with a high-fat/high-carbohydrate diet [HFCD/obese and hyperglycemia (O/H)] or a normal diet (control) for 3.5 months, subjected to pMCAO, and sacrificed after 7 days.

Results: Infarct volume analysis showed no differences between the O/H and control group, whereas neurological deficits were significantly higher in the O/H group compared to the control group. Sirtuin (Sirt1) was overexpressed and NADPH oxidase was reduced in the O/H group. O/H mice had significantly lower expression of Wnt and glycogen synthase kinase 3 α and β, a key component in the Wnt signaling pathway. Translocation of apoptosis inducing factor (AIF) to the nucleus was observed in both the O/H and control groups, but O/H mice showed a higher expression of AIF in the nucleus.

Conclusions: These data suggest that impaired Wnt signaling and active apoptosis result in reduced post-stroke recovery in obese and hyperglycemic mice.

Figure 1

HFCD-induced O/H affects recovery after 7 days of permanent ischemia. A) O/H group showed significantly higher neurological deficits as compared to control group. B) O/H animals showed significantly lower locomotor activity after 7 days of pMCAO as compared to control group. C) Grip strength was lower, but not significantly in O/H group as compared to control group. D) Infarct volume analysis in control and O/H groups showed no significant differences after 7 days of pMCAO. Data are expressed as mean ± SEM; * p<0.05, vs. control; Control n=12 and O/H n=13

Figure 2

HFCD treatment induced Sirt1 expression and after 7 days of pMCAO. A) Sirt1 expression was significantly reduced in control pMCAO mice. Sirt1 expression was significantly elevated in O/H pMCAO group as compared to control pMCAO group. C) NAD(P)H oxidase (NOX4) expression in sham O/H and control pMCAO was reduced significantly as compared to control group and O/H pMCAO group showed further significant decrease in the expression levels. (B and D) corresponding graph shows the densitometric analysis normalized to actin. Data are expressed as mean ± SEM; ^@ p<0.05, vs. sham-control; ^# p<0.05, vs. sham-O/H; * p<0.05, vs. pMCAO-control; Control n=6 and O/H n=6 (@@/##/** represents p<0.001)

Figure 3

Effect of HFCD-induced O/H on Wnt and GSK-3 α/β expression following pMCAO. A) Wnt expression was reduced non-significantly in sham O/H and significantly on control pMCAO as compared to sham control. On subjecting O/H mice to pMCAO, Wnt expression was significantly altered after 7 days as compared to control pMCAO group. C) Similarly, phosphorylation of GSK-3 α/β was significantly reduced in sham O/H and control pMCAO which were further reduced in O/H pMCAO. (B and D) corresponding graph shows the densitometric analysis normalized to actin and t-GSK. Data are expressed as mean ± SEM; ^@ p<0.05, vs. sham-control; ^# p<0.05, vs. sham-O/H; * p<0.05, vs. pMCAO-control; Control n=6 and O/H n=6 (@@/##/** represents p<0.001)

Figure 4

O/H induced by HFCD upregulates caspase independent mechanism via AIF translocation to nucleus after permanent ischemia. A) O/H group when subjected to pMCAO activated caspase independent mechanism by significantly upregulating AIF translocation to nucleus as compared to control pMCAO group. B) Corresponding graph shows the densitometric analysis normalized to histones. Data are expressed as mean ± SEM; * p<0.05, vs. control; Control n=6 and O/H n=6

Figure 5

HFCD alters non-amyloidogenic APP expression after 7 days of pMCAO. A) O/H group of mice showed no significant difference compared to control group subjected to pMCAO, however APP expression was significantly reduced as compared to respective sham groups. B) Corresponding graph shows the densitometric analysis normalized to actin. Data are expressed as mean ± SEM; ^# p<0.05, vs. sham-O/H; * p<0.05, vs. control; Control n=6 and O/H n=6