Streptozotocin (stz)

Diabetes was induced in rats by a single intraperitoneal injection of freshly prepared solution of 50 mg/kg streptozotocin (Sigma-Aldrich, Johannesburg, SA) in citrate buffer (0.1 M, pH 4.5) to overnight fasted rats. Diabetes was confirmed by stable hyperglycemia (>18 mmol/l) in the tail blood glucose after 5 days of streptozotocin (STZ) injection using a portable glucometer (Accu-Chek, Roche, Germany).

The rat fasted overnight and then was injected with streptozotocin (Sigma Aldrich, Co, USA) at a dose of 55 mg/kg BW intraperitoneal (i.p) that was dissolved in citrate buffer (0.1 M; pH 4.5). Three days after the streptozotocin injection, blood samples were taken through the lateral vein of the tail and tested for blood glucose levels by the glucometer (Accu-Check, Roche Diagnostics, Pvt. Ltd.). Rats with a glucose level of >250 mg/dL could be used as experimental animals.

All experiments and procedures were approved by the Animal Ethics Committee of Soochow University and were carried out in accordance with the Guide for the Care and Use of Laboratory Animals. Male Sprague–Dawley rats weighing 200–250 g were purchased from the Experimental Animal Center of Soochow University. The diabetic rat model was established by a single intraperitoneal injection of streptozotocin (65 mg/kg, Sigma) as previously described9 (link). The tail vein blood glucose levels were measured one week after streptozotocin injection using a glucometer (Accu-Chek, Roche Diagnostics). Only rats with blood glucose levels ≥16.7 mmol/l were considered diabetic in this study. Diabetic rats were intracoronary injected with lentivirus of pcDNA-H19 (DM + pcDNA-H19) or empty vector (DM + pcDNA vector), and were then kept for 12 weeks together with the normal rats (Control) and the diabetic rats without lentivirus injection (DM).

Male 8-week-old C57BL/6 mice were purchased from Japan SLC (Hamamatsu, Shizuoka, Japan) and maintained under specific pathogen-free conditions while consuming a standard diet and water ad libitum. Animals with unrelated health issues were excluded from the study. Animals were randomly allocated to normal and diabetic groups. The mice in the diabetic group were fasted for 6 h and then received a 0.2-mL intraperitoneal injection of streptozotocin (Sigma-Aldrich, 100 mg/kg body weight in citrate buffer at pH 4.5). One week after streptozotocin administration, the blood glucose levels were measured using an Accu-Chek glucose meter (Roche Diagnostics, Miami, FL, USA) and the mice with glycemia, glucose > 250 mg/dL, were considered diabetic [55 ]. The diabetic mice were monitored for polydipsia, polyuria and a high fasting blood glucose concentration. All procedures were approved by the Institutional Review Committee of Juntendo University and were conducted according to the National Institutes of Health Guide for the Care and Use of Laboratory Animals. The study was conducted according to the guidelines of the Declaration of Helsinki and approved by the Institutional Review Board of Juntendo University (protocol code 310070, approved on 21 February 2019).

Mice in C57Bl/6N genetic background were used for STZ-induced diabetes. Female 8–12 week old mice were given five consecutive daily intraperitoneal injections of 50mg/kg body weight STZ (Sigma) diluted in 10mM citrate buffer in order to induce diabetes. Control mice were injected with buffer alone. Blood glucose levels were examined seven days after the final STZ injection, and every week from that time on by obtaining blood from the lateral saphenous vein and measuring glucose concentration with a glucometer (Accu-Check instant, Boehringer Mannheim Corporation, Indianapolis, IN, USA). Mice with blood glucose levels greater than 300 mg/dl for two consecutive measurements were considered diabetic.

Mice were rendered diabetic by intraperitoneal injection of STZ (Boehringer-Mannheim, Mannheim, Germany) diluted in citrate buffer (10 mM, pH 4.5) at a dose of 100 mg/kg/day for 3 days, The control group (non-diabetic, ND, n = 10) was administered an equivalent volume of the vehicle citrate buffer. One week after STZ injection, glycemia was measured after 6 hours of food deprivation and animals with glucose levels greater than 250 mg/dL for at least 2 days were considered diabetic. Six weeks after STZ injections, diabetic mice were randomly divided and received either no treatment (vehicle soy oil, DV, n = 10) or oral quercetin (DQ, n = 10) at a dosage of 10 mg/kg per day for 4 weeks. This dosage was based on previous studies with hypertensive and diabetic animals [31 (link), 32 (link)].