The effect of dried okra (Abelmoschus esculentus L.) fruit extract supplementation on serum levels of key factors associated with autophagy and genes expression of ATG-5, LC3, and Beclin-1 in patients with diabetic nephropathy: A triple-blinded controlled clinical trial

Abstract

Abstract

Background and aims: Diabetes mellitus (DM) is a fast expanding public health issue which its epidemiology and consequences offer a significant worldwide health threat. Autophagy is a homeostatic process regulating the turnover of organelles and proteins inside cells that is essential for kidney physiology and homeostasis. Numerous studies have identified autophagy as a key factor in the development of diabetic nephropathy (DN). The okra’s advantageous influence on glycemic status, as well as some autophagy key biomarkers have been reported in a few animal and cell-line studies. Regarding the fact that there is no human study to examine the effect of okra on autophagy pathway, the objective of the present randomized controlled experiment (RCT) was to investigate the effects of dried okra extract (DOE) supplementation on the autophagy key factors in DN patients.

Materials and Methods: In the present triple-blinded RCT, eligible individuals were recruited from patients, referred to endocrine clinic of Imam Reza hospital in Tabriz and Tabriz Diabetes Association. After diagnosis, individuals at stage Ⅲa DN (based on protein excretion more than 0.3 g in 24-hour urine for more than three months or GFR = 45-60 ml/min/1.73 m2 with a history of retinopathy) were invited to the study, considering the inclusion and exclusion criteria. Participants were randomly assigned to either the DOE or placebo groups and asked to consume one 125mg DOE capsule or similar amounts of carboxymethyl cellulose (CMC) as placebo, before lunch for 10 weeks. Glycemic status, anthropometric variables and serum levels of mTOR, SIRT-1, and Beclin-1 were measured at baseline and at the end of the trial. The expressions of ATG-5, LC3, and Beclin-1 genes were measured in peripheral blood mononuclear cells, using Real-Time PCR technique. Statistical analyses were performed based on per protocol using SPSS 21 and STATA 16 softwares.

Results: At the end of the trial, within-group analysis revealed a significant decrease in fasting blood glucose (FBG) (P=0.04), glycated hemoglobin A1C (HbA1C) (P=0.02), and Homeostatic Model Assessment for Insulin Resistance (HOMA-IR) (P=0.01) in DOE group when compared to baseline values. The mean of serum insulin level in placebo group (mean:12.6 μIU/ml) after study period was significantly lower when compared to baseline values (mean: 16.5 μIU/ml) (P=0.01). Besides, our results demonstrated that energy intake (P=0.04) and carbohydrate intake (P=0.03) in the DOE group at the end of the study decreased significantly, compared to the beginning of the study; although no significant difference was observed between the two study groups. Between-group analysis did not show any significant effects of DOE supplements on anthropometric variables. Further, no significant difference was observed in genes expression of ATG-5 (P=0.16), LC3 (P=0.66), and Beclin-1 (P=0.44) in the study groups. Intergroup analysis also did not demonstrate any significant alteration in serum levels of mTOR, SIRT-1, and Beclin-1. There were no significinat difference serum levels of mTOR, SIRT-1, and Beclin-1 regarding between group analyses. Conclusions: Our findings indicated that DOE supplementation could not significantly alter serum levels of mTOR, SIRT1, and Beclin1. Besides, DOE supplementation had no effect on autophagy-related genes expression. More clinical trials should be conducted to better explore the relationship between DOE supplementation and autophagy biomarkers.

Key words: diabetic nephropathy, okra, autophagy, sirtuin-1, beclin-1, autophagy gene