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The Extract of Aster Koraiensis Prevents Retinal Pericyte Apoptosis in Diabetic Rats and Its Active Compound, Chlorogenic Acid Inhibits AGE Formation and AGE/RAGE Interaction

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ABSTRACT

Retinal capillary cell loss is a hallmark of early diabetic retinal changes. Advanced glycation end products (AGEs) are believed to contribute to retinal microvascular cell loss in diabetic retinopathy. In this study, the protective effects of Aster koraiensis extract (AKE) against damage to retinal vascular cells were investigated in streptozotocin (STZ)-induced diabetic rats. To examine this issue further, AGE accumulation, nuclear factor-kappaB (NF-κB) and inducible nitric oxide synthase (iNOS) were investigated using retinal trypsin digests from streptozotocin-induced diabetic rats. In the diabetic rats, TUNEL (Terminal deoxynucleotidyl transferase mediated dUTP Nick End Labeling)-positive retinal microvascular cells were markedly increased. Immunohistochemical studies revealed that AGEs were accumulated within the retinal microvascular cells, and this accumulation paralleled the activation of NF-κB and the expression of iNOS in the diabetic rats. However, AKE prevented retinal microvascular cell apoptosis through the inhibition of AGE accumulation and NF-κB activation. Moreover, to determine the active compounds of AKE, two major compounds, chlorogenic acid and 3,5-di-O-caffeoylquinic acid, were tested in an in vitro assay. Among these compounds, chlorogenic acid significantly reduced AGE formation as well as AGE/RAGE (receptor for AGEs) binding activity. These results suggest that AKE, particularly chlorogenic acid, is useful in inhibiting AGE accumulation in retinal vessels and exerts a preventive effect against the injuries of diabetic retinal vascular cells.

No MeSH data available.


The expression pattern of inducible NOS (iNOS). (A) Distribution of iNOS mRNA as detected by in situ hybridization. The diabetic retinal microvascular cells exhibited strong hybridization signal for iNOS (arrow). The AKE-treated capillary cells rarely exhibited signals compared to the diabetic cells; (B) Immunohistochemical localization of the iNOS protein. iNOS immunoreactivity (arrowhead) was observed in the cytoplasm of the diabetic retinal microvascular cells. The immunoreactivity in the AKE-treated retinal microvascular cells was decreased in intensity. NOR, normal rats; STZ, STZ-induced diabetic rats; AKE, AKE-treated diabetic rats. 400× magnification; Quantitative analysis of (C) iNOS mRNA and (D) iNOS protein signal intensities. The values in the bar graphs represent the mean ± the SEM, n = 8. * p < 0.01 vs. normal rats, #p < 0.01 vs. STZ-induced diabetic rats.
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nutrients-08-00585-f005: The expression pattern of inducible NOS (iNOS). (A) Distribution of iNOS mRNA as detected by in situ hybridization. The diabetic retinal microvascular cells exhibited strong hybridization signal for iNOS (arrow). The AKE-treated capillary cells rarely exhibited signals compared to the diabetic cells; (B) Immunohistochemical localization of the iNOS protein. iNOS immunoreactivity (arrowhead) was observed in the cytoplasm of the diabetic retinal microvascular cells. The immunoreactivity in the AKE-treated retinal microvascular cells was decreased in intensity. NOR, normal rats; STZ, STZ-induced diabetic rats; AKE, AKE-treated diabetic rats. 400× magnification; Quantitative analysis of (C) iNOS mRNA and (D) iNOS protein signal intensities. The values in the bar graphs represent the mean ± the SEM, n = 8. * p < 0.01 vs. normal rats, #p < 0.01 vs. STZ-induced diabetic rats.

Mentions: The localization of iNOS mRNA was detected by in situ hybridization. In the retinal trypsin digests of the diabetic rats, iNOS mRNA was expressed at markedly higher levels in the cytoplasm of the microvascular cells than in the digests of the normal rats (Figure 5A). Similarly, the iNOS protein was highly expressed in the cytoplasm of the retinal microvascular cells of the diabetic rats as assessed by immunohistochemistry (Figure 5B). However, the treatment with AKE markedly reduced the expression level of iNOS mRNA. Similarly, a remarkable reduction in the level of iNOS protein was observed in the AKE-treated diabetic retinal vessels (Figure 5C,D).


The Extract of Aster Koraiensis Prevents Retinal Pericyte Apoptosis in Diabetic Rats and Its Active Compound, Chlorogenic Acid Inhibits AGE Formation and AGE/RAGE Interaction
The expression pattern of inducible NOS (iNOS). (A) Distribution of iNOS mRNA as detected by in situ hybridization. The diabetic retinal microvascular cells exhibited strong hybridization signal for iNOS (arrow). The AKE-treated capillary cells rarely exhibited signals compared to the diabetic cells; (B) Immunohistochemical localization of the iNOS protein. iNOS immunoreactivity (arrowhead) was observed in the cytoplasm of the diabetic retinal microvascular cells. The immunoreactivity in the AKE-treated retinal microvascular cells was decreased in intensity. NOR, normal rats; STZ, STZ-induced diabetic rats; AKE, AKE-treated diabetic rats. 400× magnification; Quantitative analysis of (C) iNOS mRNA and (D) iNOS protein signal intensities. The values in the bar graphs represent the mean ± the SEM, n = 8. * p < 0.01 vs. normal rats, #p < 0.01 vs. STZ-induced diabetic rats.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5037569&req=5

nutrients-08-00585-f005: The expression pattern of inducible NOS (iNOS). (A) Distribution of iNOS mRNA as detected by in situ hybridization. The diabetic retinal microvascular cells exhibited strong hybridization signal for iNOS (arrow). The AKE-treated capillary cells rarely exhibited signals compared to the diabetic cells; (B) Immunohistochemical localization of the iNOS protein. iNOS immunoreactivity (arrowhead) was observed in the cytoplasm of the diabetic retinal microvascular cells. The immunoreactivity in the AKE-treated retinal microvascular cells was decreased in intensity. NOR, normal rats; STZ, STZ-induced diabetic rats; AKE, AKE-treated diabetic rats. 400× magnification; Quantitative analysis of (C) iNOS mRNA and (D) iNOS protein signal intensities. The values in the bar graphs represent the mean ± the SEM, n = 8. * p < 0.01 vs. normal rats, #p < 0.01 vs. STZ-induced diabetic rats.
Mentions: The localization of iNOS mRNA was detected by in situ hybridization. In the retinal trypsin digests of the diabetic rats, iNOS mRNA was expressed at markedly higher levels in the cytoplasm of the microvascular cells than in the digests of the normal rats (Figure 5A). Similarly, the iNOS protein was highly expressed in the cytoplasm of the retinal microvascular cells of the diabetic rats as assessed by immunohistochemistry (Figure 5B). However, the treatment with AKE markedly reduced the expression level of iNOS mRNA. Similarly, a remarkable reduction in the level of iNOS protein was observed in the AKE-treated diabetic retinal vessels (Figure 5C,D).

View Article: PubMed Central - PubMed

ABSTRACT

Retinal capillary cell loss is a hallmark of early diabetic retinal changes. Advanced glycation end products (AGEs) are believed to contribute to retinal microvascular cell loss in diabetic retinopathy. In this study, the protective effects of Aster koraiensis extract (AKE) against damage to retinal vascular cells were investigated in streptozotocin (STZ)-induced diabetic rats. To examine this issue further, AGE accumulation, nuclear factor-kappaB (NF-&kappa;B) and inducible nitric oxide synthase (iNOS) were investigated using retinal trypsin digests from streptozotocin-induced diabetic rats. In the diabetic rats, TUNEL (Terminal deoxynucleotidyl transferase mediated dUTP Nick End Labeling)-positive retinal microvascular cells were markedly increased. Immunohistochemical studies revealed that AGEs were accumulated within the retinal microvascular cells, and this accumulation paralleled the activation of NF-&kappa;B and the expression of iNOS in the diabetic rats. However, AKE prevented retinal microvascular cell apoptosis through the inhibition of AGE accumulation and NF-&kappa;B activation. Moreover, to determine the active compounds of AKE, two major compounds, chlorogenic acid and 3,5-di-O-caffeoylquinic acid, were tested in an in vitro assay. Among these compounds, chlorogenic acid significantly reduced AGE formation as well as AGE/RAGE (receptor for AGEs) binding activity. These results suggest that AKE, particularly chlorogenic acid, is useful in inhibiting AGE accumulation in retinal vessels and exerts a preventive effect against the injuries of diabetic retinal vascular cells.

No MeSH data available.