Effects of Steaming Time and Frequency for Manufactured Red Liriope platyphylla on the Insulin Secretion Ability and Insulin Receptor Signaling Pathway

Abstract

In oriental medicine, Liriope platyphylla (LP) has long been regarded as a curative herb useful for the treatment of diabetes, asthma, and neurodegenerative disorders. The principal objective of this study was to assess the effects of steaming time and frequency for manufactured Red LP (RLP) on insulin secretion ability and insulin receptor signaling pathway. To achieve our goal, several types of LPs manufactured under different conditions were applied to INS cells and streptozotocin (STZ)-induced diabetic ICR mice, after which alterations in insulin concentrations were detected in the culture supernatants and sera. The optimal concentration for the investigation of insulin secretion ability was found to be 50 ug/mL of LP. At this concentration, maximum insulin secretion was observed in the INS cells treated with LP extract steamed for 3 h (3-SLP) with two repeated steps (3 h steaming and 24 h air-dried) carried out 9 times (9-SALP); no significant changes in viability were detected in any of the treated cells. Additionally, the expression and phosphorylation levels of most components in the insulin receptor signaling pathway were increased significantly in the majority of cells treated with steaming-processed LP as compared to the cells treated with LP prepared without steaming. With regard to glucose transporter (GLUT) expression, alterations of steaming time induced similar responses on the expression levels of GLUT-2 and GLUT-3. However, differences in steaming frequency were also shown to induce dose-dependent responses in the expression level of GLUT-2 only; no significant differences in GLUT-3 expression were detected under these conditions. Furthermore, these responses observed in vitro were similarly detected in STZ-induced diabetic mice. 24-SLP and 9-SALP treatment applied for 14 days induced the down-regulation of glucose concentration and upregulation of insulin concentration. Therefore, these results indicated that the steaming processed LP may contribute to the relief of diabetes symptoms and should be regarded as an excellent candidate for a diabetes treatment.

Keywords: Liriope platyphylla; diabetes; glucose; insulin; steaming process.

Figure 1

Optimal concentration of steamed Liriope platyphylla on the toxicity and insulin secretion of INS cells. Cells were cultured with one of 8 different concentrations in dH₂O for 24 hrs. Activity of cell viability was measured using an MTT assay (A). The culture supernatants were collected from each of the cells. Insulin concentration in the supernatant was measured with an anti-insulin ELISA kit (B). The values of data represented the means±SD of three experiments. ^*P<0.05 is the significance level relative to the vehicle-treated group.

Figure 2

Effects of the different steaming time and frequency of Liriope platyphylla on toxicity and insulin secretion in INS cells. Cells were cultured with one of the 5 different extracts manufactured under the different concentrations and two ginseng types in dH₂O for 24 hrs. Activity of cell viability was measured via MTT assays (A). The culture supernatants were collected from each of the cells. Insulin concentration in the supernatant was measured using an anti-insulin ELISA kit (B). The values of data represented the means±SD of three experiments. ^*P<0.05 is the significance level relative to the vehicle-treated group.

Figure 3

Effects of the different steaming times of Liriope platyphylla on the down-stream signaling pathway of the insulin receptor signaling pathway via Western blotting. Total cell lysates were prepared from INS cells treated with vehicle, KWG, KRG and five different LPs prepared under different steaming conditions, as described in the Materials and Methods section. Fifty micrograms of protein per sample were immunoblotted with antibodies for each protein. Three samples were assayed in triplicate via Western blotting. The values are expressed as means±SD. ^*P<0.05 is the significance level compared to the vehicle-treated group.

Figure 4

Effects of the different steaming frequencies of Liriope platyphylla on down-stream signaling pathway of insulin receptor signaling pathway via Western blotting. Total cell lysates were prepared from INS cells treated with vehicle, KWG, KRG and six different LPs prepared under different steaming conditions, as described in the Materials and Methods sections. Fifty micrograms of protein per sample were immunoblotted with antibodies for each protein. Three samples were assayed in triplicate via Western blotting. The values are expressed as the means±SD. ^*P<0.05 is the significance level compared to the vehicle-treated group.

Figure 5

Effects of steamed Liriope platyphylla on glucose metabolism of streptozotocin-induced diabetic model animals. The blood was collected from the abdominal veins of streptozotocin-induced diabetic model mice after steaming Liriope platyphylla for 14 days. The glucose concentration was measured at three different times using the sensitive strip of the Blood Glucose Monitoring System (A). The insulin concentration was detected in the serum of streptozotocin-induced diabetic model animals on the final day (B). The values of data were expressed as the means±SD of three experiments. ^*P<0.05 is the significance level compared to the vehicle-treated group.