Orally Bioavailable Curcumin Nanosystems (nCUR) Prevent Pancreatic Islets/Beta Cells from Cellular Stress Induced Inflammation and Apoptosis in In Vitro and In Vivo Models of Type 1 Diabetes

Type 1 diabetes (T1D) is characterized by a selective and progressive loss of insulin‐producing β‐cells and approaches to prevent this process will help conquer the devastating disease. Curcumin (CUR), a natural anti‐inflammatory is known for its ability to suppress diabetes associated inflammation and cell death, however, the doses tested very high due to poor oral bioavailability of CUR and therefore difficult to translate. We recently prepared biodegradable nanosystems encapsulating curcumin (nCUR) resulting in 9‐fold improvement in oral bioavailability. In the current study we tested nCUR for its ability to prevent pancreatic islets/beta cells from cellular stress induced inflammation and apoptosis in vitro and in vivo . Preparation of nCUR The nCUR were prepared as previously reported by our laboratory. CUR/nCUR pretreatment on STZ induced cell death in RIN‐m5f cells The RIN‐m5f cells were cultured according to manufacturer's protocols. The cells (5*10 4 /well in 24‐well plates) were pretreated with various concentrations of CUR or nCUR for 24 h and were challenged with 5 mM streptozotocin (STZ) (dissolved in citrate buffer, pH 4.5; consist of trisodium citrate 0.1 M and citric acid 0.1 M) and incubated for 24 h. For insulin secretion assay we used Krebs‐Ringer bicarbonate (KRB) medium consisted of 20 mmol/l D‐glucose, 10 mmol/l HEPES, and 0.5% BSA re‐incubated at 37°C for 1 h. The cells were washed with PBS+1% BSA twice after taking off from the laminar and 0.5 ml/ml propidium iodide was added and cell death was observed under EVOS microscope. In another set of experiment cells were fixed in 4% formaldehyde in PBS at room temperature for 30 min followed by washing with PBS. The effects of CUR/nCUR were evaluated for caspase 8 by immunocytochemistry on fixed cells. CUR/nCUR pretreatment on STZ induced beta cell death in vivo Three months old Sprague Dawley rats with average body weight of 250±15 g were used in the study. All animals were fed normal diet ad libitum throughout the study. After acclimatization for two weeks, the rats were pre‐treated with nCUR or CUR 10, 50 mg/kg 12 h prior STZ (60 mg/kg) challenge and the rats were sacrificed 72 h post STZ. The control groups received neither CUR nor STZ. The blood and pancreas were collected and processed for various markers and histology/immunohistochemistry. Summary of Results The particle are ~300 nm in size with ~80% encapsulation efficiency at 15% CUR loading w/w of polymer. The propidium iodide staining of the cells revealed that nCUR is more efficacious in preventing STZ induced cell death compared to CUR ( Figure 1) due to particle uptake into the cells and sustained action of the encapsulated CUR. We show both CUR and nCUR offered protection by inhibiting caspase 8 which is an essential pathway for STZ induced β‐cell apoptosis ( Figure 2). In diabetes rat model, pancreatic islets were identified by insulin immunohistochemical staining where a single dose of nCUR at 10 or 50 mg/kg seems to have preserved the cells better than the CUR group ( Figure 3). The blood glucose levels for nCUR group for both doses were significantly lower compared to CUR and STZ groups. Conclusion The nCUR offered better protection compared to CUR due to enhanced bioavailability of the nanoparticles. Further studies are underway to investigate the ability of nCUR to regenerate the beta cells on repeated dosing as well as combination therapies with insulin to halt the disease. Support or Funding Information The start‐up funds from Irma Lerma Rangel College of Pharmacy, Texas A&M Health Science Center is greatly appreciated.