Background: Doxorubicin (DOX) is one of the most effective anti-neoplastic drugs although its clinical use is limited by the severe cardiotoxicity. Apoptosis and defective autophagy are believed to contribute to DOX-induced cardiotoxicity. Here we explored the effect of curcumin (Cur) on DOX-induced cardiac injury and the mechanism involved with a focus on oxidative stress, autophagy and pyroptosis. Methods: Kunming mice were challenged with DOX (3 mg·kg⁻¹, i.p. every other day) with cohorts of mice receiving Cur at 50, 100, 200 and 400 mg·kg⁻¹ via gavage daily. Serum levels of cardiac enzymes, such as aspartate amino transferase (AST), lactate dehydrogenase (LDH), creatine kinase (CK), and heart homogenate oxidative stress markers, such as superoxide dismutase (SOD) and malondialdehyde (MDA) were determined. Echocardiographic and cardiac contraction were examined. Apoptosis, pyroptosis, autophagy and Akt/mTOR-signalling proteins were detected using western blot or electron microscopy. Cardiac contractile properties were assessed including peak shortening, maximal velocity of shortening/relengthening (± dL/dt), time-to-PS, and time-to-90% relengthening (TR90). Superoxide levels were evaluated using DHE staining. GFP-LC3 was conducted to measure autophagosomes. Results: Our study showed that Cur protected against cardiotoxicity manifested by a significant decrease in serum myocardial enzymes and improvement of anti-oxidative capacity. Cur inhibited autophagy and offered overt benefit for cardiomyocyte survive against DOX-induced toxicity. Cur attenuated DOX-induced cardiomyocyte pyroptosis as evidenced by NLR family pyrin domain containing 3 (NLRP3), Caspase-1, and interleukin-18 levels. DOX impaired cardiac function (reduced fractional shortening, ejection fraction, increased plasma cTnI level and TR90, decreased PS and ± dL/dt), the effects of which were overtly reconciled by 100 mg·kg⁻¹ but not 50 mg·kg⁻¹ Cur. H9c2 cells exposure to DOX displayed increased intracellular reactive oxygen species (ROS) and autophagy, the effects of which were nullified by Cur. Autophagy activator rapamycin cancelled off Cur-induced protective effects. Conclusions: Our finding suggested that Cur rescued against DOX-induced cardiac injury probably through regulation of autophagy and pyroptosis in a mTOR-dependent manner.

Oxidative damage, inflammation, and apoptosis are the primary features of diabetic testicular damage. Curcumin protects against diabetic testicular injury, but the underlying mechanisms remain obscure. This study examined the effect of curcumin on type 2 diabetes mellitus- (T2DM-) induced testicular injury, oxidative stress, and apoptotic changes. T2DM rats were intraperitoneally injected with 40 mg/kg STZ after being fed a high-fat diet for 8 weeks. One week after STZ injection, 100 or 200 mg/kg curcumin was administered orally to the diabetic rats for 16 weeks. Histological changes in the testes were determined by HE staining. Serum testosterone was measured. Markers of superoxide levels, such as SOD activity and MDA content, and markers of cell death, including the expression of Bax, Bcl-2, and MAPK family members, were measured by molecular biology or immunohistochemical techniques. Degeneration and disruption of seminiferous tubule structure were observed in diabetic rats. Serum testosterone levels were markedly lower in diabetic rats than in control rats. Moreover, testicular apoptosis and Bax expression were much higher in diabetic rats than in control rats. Superoxide generation, the NADP⁺/NADPH ratio, and NADPH oxidase subunit expression, including expression of the gp91phox, p47phox, and p67phox subunits, increased, while antioxidant enzyme levels decreased in diabetic rats. Furthermore, the MAPK signaling pathway was activated in diabetic rats. Curcumin partially prevented diabetes-induced microstructural abnormalities and significantly increased serum testosterone levels compared to untreated T2DM rats. Additionally, curcumin reduced testicular apoptosis by regulating apoptotic proteins and markedly inhibited oxidative stress levels by downregulating MDA expression, decreasing NADPH activity, and restoring antioxidant enzymes. Remarkably, curcumin treatment also suppressed MAPK activation. Thus, curcumin may have therapeutic value in the treatment of diabetes-induced testicular injury due to its prevention of testicular apoptosis and attenuation of oxidative stress.

Approximately 90% of male diabetes mellitus patients have varying degrees of testicular dysfunction. The molecular mechanism underlying diabetes-induced testicular damage has not been thoroughly elucidated. In this research, we sought to determine the influence of metformin (Met) on diabetes-induced testicular injury and the mechanism involved with a focus on testicular dysfunction, apoptosis, autophagy, and prokineticin 2 (PK2) signalling. In our study, C57BL/6J mice were randomly divided into the normal control group, the diabetes group, and the Met-treated group. Streptozotocin (50 mg·kg^-1·d^-1) was injected intraperitoneally into the mice for 5 days in a row to induce type 1 diabetes, which was diagnosed by a blood glucose level≥16.7mmol/L after 7 days. The experimental animals were orally administered Met (250 mg·kg^-1·d^-1) for 16 weeks. Properties of testicular function, including sperm motility and the total concentration of epididymal sperm, were assessed. Changes in testicular structure, such as the blood-testis barrier, histological pathology, and organelles, were observed. The levels of apoptosis and expression of related proteins, such as Bax and Bcl-2, were measured. Moreover, autophagy-related proteins, including Beclin-1, p62, and LC3B, as well as the PK2/PKR pathway, which consists of PK2, PKR1, PKR2, AKT, and GSK3β, were analysed. Upon the induction of diabetes, reproductive capacity was significantly impaired and a disordered arrangement of testicular seminiferous tubules and destroyed organelles in spermatogenic cells was observed. Met administration preserved testicular function and structure. In addition, in mice with diabetes, the levels of PK2, PKR2, p-Akt, and p-GSK3β were significantly decreased at different times, while that of PKR1 was markedly increased, and these changes were normalized by Met. Furthermore, diabetic mice showed increased apoptosis and decreased autophagy in the testes, the effects of which were nullified by Met. These results suggest that Met rescues diabetes-induced testicular damage by attenuating apoptosis and inducing autophagy. This effect is likely mediated by the PK2/PKR/AKT/GSK3β signalling pathway.

Mitochondrial injury and defective autophagy are common in diabetic cardiomyopathy. Recent evidence supports benefits of glucagon‐like peptide‐1 (GLP‐1) agonists exendin‐4 (Exe) and liraglutide (LIRA) against diabetic cardiomyopathy. This study was designed to examine the effect of Exe and LIRA on glucose‐induced cardiomyocyte and mitochondrial injury, oxidative stress, apoptosis, and autophagy change. Cardiomyocytes isolated from adult mice and H9c2 myoblast cells were exposed to high glucose (HG, 33 mM) with or without Exe or LIRA. Cardiac contractile properties were assessed including peak shortening, maximal velocity of shortening/relengthening (±dL/dt), time to PS, and time‐to‐90% relengthening (TR₉₀). Superoxide levels, apoptotic proteins such as cleaved caspase‐3, Bax, and Bcl‐2, and autophagy proteins including Atg5, p62, Beclin‐1, LC3B, and mTOR/ULK1 were evaluated using Western blot. Mitochondrial membrane potential (MMP) changes were assessed using JC‐1, and autophagosomes were determined using GFP‐LC3. Cardiomyocyte exposure to HG exhibited prolonged TR₉₀ associated with significantly decreased PS and ±dL/dt, the effects of which were partly restored by GLP‐1 agonists, the effects of which were negated by the mTOR activator 3BDO. H9c2 cell exposure to HG showed increased intracellular ROS, apoptosis, MMP loss, dampened autophagy, and elevated p‐mTOR and p‐ULK1, the effects of which were nullified by the GLP‐1 agonists. These results suggested that GLP‐1 agonists rescued glucose toxicity likely through induction of mTOR‐dependent autophagy.

Diabetic cardiomyopathy (DCM) is a complication of diabetes that can cause damage to myocardial structure and function. Metformin (Met) is a widely used type 2 diabetes treatment drug that exerts cardioprotective effects through multiple pathways. Prokineticin 2 (PK2) is a small-molecule secreted protein that plays pivotal parts in cardiomyocyte survival and angiogenesis. However, the role of Met in regulating the PK2 signaling pathway in DCM remains unclear. This experiment explored the effects of Met on high glucose (HG)-induced injury through the PK2/PKR pathway in vivo and in vitro. Cardiomyocytes isolated from adult or AKT-knockout mice were treated with HG (33 mmol/L) and PK2 or AKT1/2 kinase inhibitor (AKT inhibitor). Heart contraction properties based on cell shortening were evaluated; these properties included the resting cell length, peak shortening (PS), maximum speed of shortening/relengthening (±dL/dt), time to 90% relengthening (TR₉₀), and time to peak shortening (TPS). Mice with streptozotocin-induced diabetes were treated with Met to evaluate cardiac function, myocardial structure, and the PK2/PKR and AKT/GSK3β pathways. Moreover, H9c2 cardiomyocytes were exposed to HG in the absence or presence of Met with or without the PK2 antagonist PKRA7 or the AKT inhibitor, and apoptotic proteins such as Bax and Bcl-2 and the PK2/PKR and AKT/GSK3β pathways were evaluated using western blot analysis. The prolongation of TR₉₀ and decreases in PS and ±dL/dt caused by HG were ameliorated by PK2 in cardiomyocytes, but the effects of PK2 were ameliorated or negated by the AKT inhibitor and in AKT-knockout mice. Diabetic mice showed metabolic abnormalities, aberrant myocardial enzyme levels, declines in myocardial systolic and diastolic function associated with myocardial fibrosis, and pronounced apoptosis, but these effects were greatly rescued by Met treatment. Moreover, PK2, PKR1, and PKR2 expression and p-AKT/AKT and p-GSK3β/GSK3β ratios were decreased in diabetic mice, and these decreases were attenuated by Met. Likewise, H9c2 cells exposed to HG showed reduced PK2/PKR expression and decreased p-AKT/AKT and p-GSK3β/GSK3β ratios, and these effects were nullified by Met. In addition, the effects of Met on cardiomyocytes exposed to HG were abolished after intervention with PKRA7 or the AKT inhibitor. These results suggest that Met can activate the PK2/PKR-mediated AKT/GSK3β pathway, thus improving cardiac function and alleviating apoptosis in DM mice.