Excess dietary sugar contributes to metabolic disorders including type 2 diabetes mellitus (T2DM). The fruit fly, Drosophila melanogaster, has conserved insulin/IGF signalling, short generation time, and genetic tractability, making it a widely used model to study diet-induced metabolic dysfunction and to test interventions. High-sucrose feeding in flies reproduces key features of T2DM such as elevated circulating sugars, insulin resistance–like changes, increased adiposity, delayed development, reduced fertility, and shortened lifespan. Oxidative stress driven by chronic hyperglycemia is central to diabetic complications and can be assessed in flies through antioxidant enzyme activities and stress-responsive gene expression.
Metformin, a biguanide commonly used for T2DM, reduces hepatic glucose production and improves insulin sensitivity. At the cellular level it inhibits mitochondrial complex I, alters AMP/ATP ratios, and activates AMPK, with downstream effects on glucose uptake and stress response pathways. Metformin also has reported antioxidant effects, including modulation of superoxide dismutase (SOD) and catalase (CAT) activity. This study tested whether metformin modulates physiological, biochemical, behavioral, and molecular outcomes in a Drosophila model of diet-induced metabolic dysfunction caused by high-sucrose feeding.
Wild-type Oregon R D. melanogaster were maintained at 23 ± 1°C with a 12:12 h light/dark cycle on instant Drosophila medium supplemented with dry yeast. Adult flies were transferred to fresh food every four days under consistent environmental conditions. All experimental treatments were administered for 10 days and the complete experiment was independently repeated on three occasions using newly established cohorts.
Five experimental groups were defined:
Sucrose was prepared from analytical-grade sucrose and incorporated into the medium. Metformin hydrochloride was prepared as a 100 mM stock and added to food to achieve final concentrations of 5 mM or 15 mM. Food vials were freshly prepared and replaced every 48 hours during treatment.
To avoid confounding from mortality or destructive sampling, independent parallel cohorts were established for each endpoint. For adult endpoints, each condition used three biological replicate vials with 20 flies per vial (10 males and 10 virgin females). Measured endpoints after 10 days included pupation rate, adult survival, locomotor performance, trehalose levels, antioxidant enzyme activities (SOD and GST), and expression of stress-related genes (Mn-SOD, Cu/Zn-SOD, MTH, CAT, Hsp40, Hsp70) with RpS20 as the housekeeping reference.
Pupation rate was measured in synchronized developmental cohorts. Adults were allowed to oviposit during a defined collection window, and equal numbers of embryos were placed into replicate vials per group. Pupation rate was calculated as the percentage of embryos progressing to visible pupae. Survival rate for adults was assessed in dedicated cohorts with mortality recorded every 48 hours; survival at day 10 was expressed as percent live flies relative to the initial cohort size.
Locomotor ability was measured using a standardized negative geotaxis assay on Day 10 in mixed-sex groups of 20 flies. After acclimation, flies were gently tapped to the bottom of vertical glass tubes and the percentage that crossed an 8-cm mark within 10 seconds was recorded. Each biological replicate was tested three times and the mean value used for analysis.
Whole-fly homogenates were prepared from a dedicated adult cohort after 10 days of treatment. Trehalose, the primary circulating sugar in D. melanogaster, was quantified using a two-aliquot approach: free glucose was measured directly with a glucose oxidase-based colorimetric assay, and total glucose was measured after trehalase digestion to hydrolyze trehalose. Trehalose-derived glucose was calculated by subtraction against the free glucose value and converted using a glucose standard curve.
Antioxidant enzyme activities, including superoxide dismutase (SOD) and glutathione S-transferase (GST), were measured in homogenates to assess oxidative stress and antioxidant capacity.
Expression of selected stress- and antioxidant-related genes—Mn-SOD, Cu/Zn-SOD, MTH, CAT, Hsp40, and Hsp70—was quantified relative to the normal control group using RpS20 as the reference gene. Independent cohorts were used for molecular assays to ensure sufficient material and avoid confounding effects.
Feeding Drosophila high-sucrose diets (5% and 15%) for 10 days produced several diabetes-relevant outcomes. High-sucrose diets significantly reduced pupation and adult survival rates, impaired locomotor performance in the negative geotaxis assay, increased whole-fly trehalose levels, and decreased antioxidant enzyme activities (SOD and GST). At the molecular level, high-sucrose feeding was associated with downregulation of stress- and antioxidant-related genes, indicating suppressed stress-response capacity under sucrose-induced metabolic stress.
Within corresponding sucrose-treatment groups, addition of metformin partially attenuated multiple sucrose-induced alterations. Metformin-treated sucrose groups showed improvements in pupation and survival compared with sucrose-only groups, enhanced locomotor performance, partial normalization of trehalose levels, increased antioxidant enzyme activities (SOD and GST) relative to sucrose alone, and recovery of expression for several stress-related genes. These effects were reported as partial attenuation rather than full restoration to the normal diet baseline.
The findings demonstrate that sucrose-enriched diets induce metabolic disturbance and oxidative stress in D. melanogaster that parallel aspects of mammalian diet-induced insulin resistance and diabetic pathology. Measured endpoints spanned developmental success, adult survival, behavior, carbohydrate homeostasis, antioxidant capacity, and gene expression, providing an integrated view of diet-induced dysfunction.
Metformin produced consistent modulatory effects across physiological, biochemical, and molecular measures within corresponding sucrose groups, supporting its dual metabolic and redox-modulatory actions observed in other systems. The study reinforces the utility of Drosophila as a tractable, cost-effective model for screening antidiabetic interventions and exploring mechanisms linking diet, oxidative stress, and stress-responsive gene regulation.
High-sucrose diets induced developmental delays, reduced survival, locomotor deficits, trehalose accumulation, oxidative stress, and suppression of stress-related genes in Drosophila melanogaster. Metformin partially mitigated several of these sucrose-driven changes when added to the corresponding sucrose diets. These results support the use of D. melanogaster to model diet-induced metabolic dysfunction and to evaluate pharmacological modulation, while indicating that metformin can provide partial protective effects against sucrose-induced metabolic and oxidative disturbances.