Distribution of lipid droplets in hippocampal neurons and microglia: impact of diabetes and exercise.

Neuroinflammation, aging, and neurodegenerative disorders are associated with excessive accumulation of neutral lipids in lipid droplets (LDs) in microglia. Type 2 diabetes mellitus (T2DM) may cause neuroinflammation and is a risk factor for neurodegenerative disorders. Here, we show that hippocampal pyramidal neurons contain smaller, more abundant LDs than their neighboring microglia. The density of LDs varied between pyramidal cells in adjacent subregions, with CA3 neurons containing more LDs than CA1 neurons. Within the CA3 region, a gradual increase in the LD content along the pyramidal layer from the hilus toward CA2 was observed. Interestingly, the high neuronal LD content correlated with less ramified microglial morphotypes. Using the db/db model of T2DM, we demonstrated that diabetes increased the number of LDs per microglial cell without affecting the neuronal LD density. High-intensity interval exercise induced smaller changes in the number of LDs in microglia but was not sufficient to counteract the diabetes-induced changes in LD accumulation. The changes observed in response to T2DM may contribute to the cerebral effects of T2DM and provide a mechanistic link between T2DM and neurodegenerative disorders.

Hepatoprotective potential of taxifolin in type 2 diabetic rats: modulation of oxidative stress and Bcl2/Bax/Caspase-3 signaling pathway.

BACKGROUND: Diabetes mellitus (DM) is a global metabolic problem. Several factors including hyperglycemia, oxidative stress, and inflammation play significant roles in the development of DM complications. Apoptosis is also an essential event in DM pathophysiology, -with B-cell lymphoma 2 (Bcl-2) and Bcl-2 associated X (Bax) determining apoptotic susceptibility. The present study aimed to elucidate the protective effects of two doses of taxifolin (TXF) on liver damage in diabetic rats and explore the possible mechanisms of action. METHODS AND RESULTS: DM was induced in eighteen rats through intraperitoneal injections of 50 mg/kg streptozotocin and 110 mg/kg nicotinamide. Diabetic rats received daily oral intubation of 25 and 50 mg/kg TXF for 3 months. In the untreated diabetic group, there was a significant increase in fasting and postprandial glucose levels, glycosylated hemoglobin A1C (HbA1c), tumor necrosis factor-alpha (TNF-α), and interleukin-6 (IL-6), while insulin and adiponectin levels decreased significantly. Both TXF doses mitigated hyperglycemia, regulated cytokine production, and increased insulin level. Gene expressions and protein levels of Bax, caspase 3, and cytochrome c were significantly increased, while Bcl-2 was significantly decreased in the livers of diabetic rats, effects that were significantly ameliorated after TXF treatment. The results of the TUNEL assay supported the apoptotic pathway. Additionally, TXF significantly decreased lipid peroxidation and enhanced antioxidant enzyme activity in diabetic rats. Liver enzymes and histopathological changes also showed improvement. CONCLUSIONS: TXF mitigated diabetes-associated hepatic damage by reducing hyperglycemia, oxidative stress, inflammation, and modulating anti-/pro-apoptotic genes and proteins. A dose of 50 mg/kg TXF was more effective than 25 mg/kg and is recommended for consumption.

Hyperglycemia-induced oxidative stress exacerbates mitochondrial apoptosis damage to cochlear stria vascularis pericytes via the ROS-mediated Bcl-2/CytC/AIF pathway.

OBJECTIVES: Diabetes is closely linked to hearing loss, yet the exact mechanisms remain unclear. Cochlear stria vascularis and pericytes (PCs) are crucial for hearing. This study investigates whether high glucose induces apoptosis in the cochlear stria vascularis and pericytes via elevated ROS levels due to oxidative stress, impacting hearing loss. METHODS: We established a type II diabetes model in C57BL/6J mice and used auditory brainstem response (ABR), Evans blue staining, HE staining, immunohistochemistry, and immunofluorescence to observe changes in hearing, blood-labyrinth barrier (BLB) permeability, stria vascularis morphology, and apoptosis protein expression. Primary cultured stria vascularis pericytes were subjected to high glucose, and apoptosis levels were assessed using flow cytometry, Annexin V-FITC, Hoechst 33342 staining, Western blot, Mitosox, and JC-1 probes. RESULTS: Diabetic mice showed decreased hearing thresholds, reduced stria vascularis density, increased oxidative stress, cell apoptosis, and decreased antioxidant levels. High glucose exposure increased apoptosis and ROS content in pericytes, while mitochondrial membrane potential decreased, with AIF and cytochrome C (CytC) released from mitochondria to the cytoplasm. Adding oxidative scavengers reduced AIF and CytC release, decreasing pericyte apoptosis. DISCUSSION: Hyperglycemia may induce mitochondrial apoptosis of cochlear stria vascularis pericytes through oxidative stress.

Hypoxia-inducible factor-1α/vascular endothelial growth factor signaling pathway-based ulcer-healing mechanism of Astragalus Aqueous extract in diabetic foot rats.

The main objective of this work was to investigate the mechanism of Astragalus aqueous extract ulcer healing in diabetic foot model rats through the hypoxia-inducible factor 1-alpha (HIF-1ɑ)/vascular endothelial growth factor (VEGF) signalling pathway. Fifty specific-pathogen-free male Sprague Dawley rats were divided into blank (A), model control (B), Astragalus extract (C) and mupirocin (D) treatment groups. Group A received a regular diet, whereas the other groups received a high-fat/high-sugar diet and intraperitoneal streptozotocin injections to induce diabetes. Diabetic foot ulcers were created via skin excision. Subsequently, ulcers were debrided daily. Groups B, C and D received wet saline gauze, wet gauze with Astragalus extract and gauze with mupirocin, respectively, on the affected area. Group A received no treatment. After 14 days, the rats were assessed for ulcer healing and general condition. Immunohistochemistry was used to detect HIF-1ɑ and VEGF levels in the dorsalis pedis artery, and ELISA was used to determine serum IL-6 and CRP levels. The results revealed that Groups C and D had significantly faster ulcer healing compared with Group B (p < 0.01), and ulcer healing was faster in Group C than in Group D (p < 0.01). Group C exhibited notably higher HIF-1ɑ and VEGF protein expression levels compared with Groups B and D (p < 0.01). IL-6 and CRP expression levels in Groups C and D were significantly lower than those in Group B (p < 0.01). In summary, Astragalus aqueous extract effectively treats diabetic foot ulcers by up-regulating HIF-1ɑ and VEGF expression, activating the HIF-1ɑ/VEGF pathway, improving local tissue ischaemia and hypoxia, promoting collateral circulation and enhancing dorsalis pedis artery formation, thereby accelerating ulcer repair in diabetic rats.

Ameliorating effect of Chinese jujube polyphenol on blood glucose oxidative stress in type 2 diabetic rats.

BACKGROUND: Type 2 diabetes mellitus (T2DM) is a common metabolic disease characterized by insulin resistance and insufficient relative insulin secretion, leading to elevated blood sugar and the development of diabetic complications. T2DM not only seriously affects people's health and quality of life, but also brings a heavy burden to society and economy. At present, the treatment of T2DM mainly relies on drug therapy, but these drugs often have problems such as side effects, resistance and high cost, and can not fully meet the needs and expectations of patients. Therefore, it is of great significance and value to find safe and effective natural medicines or functional foods to assist the treatment and prevention of T2DM. OBJECTIVE: Chinese jujube are a common fruit that contain abundant polyphenolic compounds, which exhibit multiple physiological activities, such as antioxidation, anti-inflammation, and blood glucose lowering. The objective of this study was to explore the impact of red date polyphenols on glycemic control and oxidative stress status in patients with type 2 diabetes mellitus (T2DM).

Antox targeting AGE/RAGE cascades to restore submandibular gland viability in rat model of type 1 diabetes.

Diabetes mellitus (DM) is a chronic disorder of glucose metabolism that threatens several organs, including the submandibular (SMG) salivary glands. Antox (ANX) is a strong multivitamin with significant antioxidant benefits. The goal of this study was to demonstrate the beneficial roles of ANX supplementation in combination with insulin in alleviating diabetic SMG changes. For four weeks, 30 rats were divided into equal five groups (n = 6): (1) control group; (2) diabetic group (DM), with DM induced by streptozotocin (STZ) injection (50 mg/kg i.p.); (3) DM + ANX group: ANX was administrated (10 mg/kg/day/once daily/orally); (4) DM + insulin group: insulin was administrated 1U once/day/s.c.; and (5) DM + insulin + ANX group: co-administrated insulin. The addition of ANX to insulin in diabetic rats alleviated hyposalivation and histopathological alterations associated with diabetic rats. Remarkably, combined ANX and insulin exerted significant antioxidant effects, suppressing inflammatory and apoptotic pathways associated with increased salivary advanced glycation end-product (AGE) production and receptor for advanced glycation end-product expression (RAGE) activation in diabetic SMG tissues. Combined ANX and insulin administration in diabetic rats was more effective in alleviating SMG changes (functions and structures) than administration of insulin alone, exerting suppressive effects on AGE production and frustrating RAGE downstream pathways.

The effect of Polygonum hyrcanicum Rech. f. hydroalcoholic extract on oxidative stress and nephropathy in alloxan-induced diabetic mice.

Diabetic nephropathy, characterized by inflammation and oxidative stress, poses a management challenge. This study investigates the effect of Polygonum hyrcanicum extract on diabetic nephropathy in alloxan-induced diabetic mice. In this experimental animal study, the P. hyrcanicum extract was prepared using continuous macerations. Thirty male Albino mice, divided into five groups, were induced with alloxan-induced diabetes. They received intraperitoneal injections of the plant extract (100 and 200 mg/kg) and metformin (300 mg/kg) for four weeks. Kidney and blood samples were collected to assess protein carbonyl, glutathione, lipid peroxidation, TNF-α and IL-6 levels. The amount of total flavonoid and phenolic content in the hydroalcoholic extract of P. hyrcanicum were 7.5 ± 0.3 mg of quercetin and 88.2 ± 1.3 mg gallic acid per gram of extract respectively. The antioxidant activity level of the hydroalcoholic extract was determined to be 1.78 ± 0.51 mM equivalent per gram of extract. Alloxan administration resulted in a significant reduction in glutathione levels and a significant increase in protein carbonyl, lipid peroxidation, TNF-α, and IL-6 levels. Hydroalcoholic extract of P. hyrcanicum effectively reduced oxidative stress markers and inflammatory cytokines (TNF-α, IL-6), indicating its potential in mitigating diabetic nephropathy. However, no significant difference in efficacy was observed between the 100 mg/kg and 200 mg/kg doses in terms of reducing these toxicities.

Potential of Sorghum Seeds in Alleviating Hyperglycemia, Oxidative Stress, and Glycation Damage.

Diabetes mellitus, characterized by dysregulated glucose metabolism, oxidative stress, and the formation of advanced glycation end products, poses a significant global health burden. In this study, we explored the potential of sorghum (Sorghum bicolor) seeds, known for their abundant phytochemical composition, as a natural remedy for diabetes and its associated damage. High-performance liquid chromatography/high-resolution mass spectrometry analysis revealed a remarkable phenolic richness in sorghum grains, including gallic acid, quercetin, and the predominant procyanidin B-1, with ecotype-specific variations in flavonoid distribution. Elemental analysis by ICP showed an abundance of macro-elements (Ca, K, Mg), trace elements (Fe, Mn, Si, Zn), and ultra-trace elements (B, Co, Cr, Cu, Mo, Se, V) essential for human health, supporting its therapeutic and nutritional potential. Additionally, the results demonstrated variable total phenolic contents (188-297 mg GAE/g dE) and total flavonoid contents (66-78 mg QE/g dE), with corresponding differences in antioxidant activities across the five ecotypes. Treatment with sorghum seed extract (SE1) significantly reduced oxidative stress markers, such as malondialdehyde (MDA)by 40% and hydrogen peroxide (H2O2) by 63%, in diabetic mice, compared to untreated diabetic controls. Moreover, sorghum extracts exhibited a remarkable increase in antioxidant enzyme activities, including a 50% increase in superoxide dismutase (SOD) activity and a 60% increase in glutathione peroxidase (GPx) activity, indicating their potential to bolster antioxidant defenses against diabetes-induced oxidative stress. These findings underscore the therapeutic potential of sorghum seeds in diabetes management and prevention, paving the way for the development of functional foods with enhanced health benefits.

New Glycotoxin Inhibitor from Sesuvium sesuvioides Mitigates Symptoms of Insulin Resistance and Diabetes by Suppressing AGE-RAGE Axis in Skeletal Muscle.

The current study intended to investigate the role of new natural compounds derived from the Sesuvium sesuvioides plant in mitigating symptoms of diabetes and insulin resistance in the diabetic mice model. Anti-advanced glycation activity, insulin, and adiponectin were quantified by enzyme-linked immunosorbent assay (ELISA). Glucose uptake was performed using enzymatic fluorescence assay, and glycogen synthesis was measured using PAS staining. Gene and protein expression was assessed using real time PCR (RT-PCR), and immunoblotting and fluorescent microscopy, respectively. The new flavonoid glycoside eupalitin 3-O-α-L-rhamnopyranosyl-(1→2)-ß-D-glucopyranoside 1 isolated from S. sesuvioides exhibited anti-AGE activity by reducing human glycated albumin in liver cells. In a diabetic mouse model treated with compound 1, we observed improved glucose tolerance, increased adiponectin levels, and decreased insulin resistance. We also observed alleviated AGEs induced reduction in glucose uptake and restored glycogen synthesis in the compound 1-treated diabetic mice muscles. Exploring the molecular mechanism of action in skeletal muscle tissue of diabetic mice, we found that 1 reduced AGE-induced reactive oxygen species and the inflammatory gene in the muscle of diabetic mice. Additionally, 1 exhibited these effects by reducing the gene and protein expression of receptor for advanced glycation end products (RAGE) and inhibiting protein kinase C (PKC) delta activation. This further led us to demonstrate that compound 1 reduced serine phosphorylation of IRS-1, thereby restoring insulin sensitivity. We conclude that a new flavonoid glycoside from S. sesuvioides could be a therapeutic target for the treatment of symptoms of insulin resistance and diabetes.

IL-10 Enhances the Inhibitory Effect of Adipose-Derived Stromal Cells on Insulin Resistance/Liver Gluconeogenesis by Treg Cell Induction.

The modulation of cellular phenotypes within adipose tissue provides a potential means for therapeutic intervention for diabetes. Endogenous interleukin-10 (IL-10) protects against diet-induced insulin resistance. We examined the effects and mechanisms of action of IL-10-treated adipose-derived stromal cells on diabetes-induced insulin resistance and liver gluconeogenesis. We harvested stromal vascular fractions (SVFs) from the adipose tissue of diabetic (Leprdb/db) mice and treated them with IL-10 in vitro. SVFs treated with 10 or 100 ng of IL-10 were injected into the inguinal adipose tissue of Leprdb/db mice. IL-10 treatment suppressed the mRNA expression of IL-6, IL-33, CCL2, TNF-α, and IL-1ß. Additionally, it suppressed the protein expression of IL-6, pmTOR, pJNK, and pNF-κB but enhanced Foxp3 mRNA expression in SVFs from diabetic mice. Meanwhile, IL-10 treatment repressed CCL2 and PDGFRα expression in adipose tissue macrophages (ATMs) and IL-6 expression in non-ATMs but increased the Foxp3 and IL-10 mRNA expression of ATMs from diabetic mice. Injection of IL-10-treated SVFs decreased the IL-6, IL-33, CCL2, IL-1ß, and CCL2 but enhanced the Foxp3 and IL-10 mRNA expression of adipose tissue from Leprdb/db mice. Furthermore, injection of IL-10-treated SVFs increased CD4+ regulatory T cells (Tregs) in SVFs and adipose IL-10 levels and suppressed plasma adiponectin levels and DPP4 activity in diabetic mice. Injection of IL-10-treated SVFs decreased hepatic G6PC and PCK1 mRNA expression and increased Akt activation, STAT3 phosphorylation in the liver, and glucose tolerance in diabetic mice. Our data suggest that IL-10 treatment decreases inflammation in adipose SVFs of diabetic mice. Injection of IL-10-treated SVFs into the adipose tissue decreased diabetes-induced gluconeogenesis gene expression, DPP4 activity, and insulin resistance by enhancing Treg cells in diabetic mice. These data suggest that IL-10-treated adipose stromal vascular cells could be a promising therapeutic strategy for diabetes mellitus.

Small RNA-Seq and real time rt-qPCR reveal islet miRNA released under stress conditions.

Replacement of beta cells through transplantation is a potential therapeutic approach for individuals with pancreas removal or poorly controllable type 1 diabetes. However, stress and death of beta cells pose significant challenges. Circulating miRNA has emerged as potential biomarkers reflecting early beta cell stress and death, allowing for timely intervention. The aim of this study was to identify miRNAs as potential biomarkers for beta cell health. Literature review combined with small RNA sequencing was employed to select islet-enriched miRNA. The release of those miRNA was assessed by RT-qPCR in vivo, using a streptozotocin induced diabetes mouse model and in vitro, through mouse and human islets exposed to varying degrees of hypoxic and cytokine stressors. Utilizing the streptozotocin induced model, we identified 18 miRNAs out of 39 candidate islet-enriched miRNA to be released upon islet stress in vivo. In vitro analysis of culture supernatants from cytokine and/or hypoxia stressed islets identified the release of 45 miRNAs from mouse and 8 miRNAs from human islets. Investigation into the biological pathways targeted by the cytokine- and/or hypoxia-induced miRNA suggested the involvement of MAPK and PI3K-Akt signaling pathways in both mouse and human islets. We have identified miRNAs associated with beta cell health and stress. The findings allowed us to propose a panel of 47 islet-related human miRNA that is potentially valuable for application in clinical contexts of beta cell transplantation and presymptomatic early-stage type 1 diabetes.

Transcriptomic Profile of Lin-Sca1+c-kit (LSK) cells in db/db mice with long-standing diabetes.

BACKGROUND: The Lin-Sca1+c-Kit+ (LSK) fraction of the bone marrow (BM) comprises multipotent hematopoietic stem cells (HSCs), which are vital to tissue homeostasis and vascular repair. While diabetes affects HSC homeostasis overall, the molecular signature of mRNA and miRNA transcriptomic under the conditions of long-standing type 2 diabetes (T2D;>6 months) remains unexplored. METHODS: In this study, we assessed the transcriptomic signature of HSCs in db/db mice, a well-known and widely used model for T2D. LSK cells of db/db mice enriched using a cell sorter were subjected to paired-end mRNA and single-end miRNA seq library and sequenced on Illumina NovaSeq 6000. The mRNA sequence reads were mapped using STAR (Spliced Transcripts Alignment to a Reference), and the miRNA sequence reads were mapped to the designated reference genome using the Qiagen GeneGlobe RNA-seq Analysis Portal with default parameters for miRNA. RESULTS: We uncovered 2076 out of 13,708 mRNAs and 35 out of 191 miRNAs that were expressed significantly in db/db animals; strikingly, previously unreported miRNAs (miR-3968 and miR-1971) were found to be downregulated in db/db mice. Furthermore, we observed a molecular shift in the transcriptome of HSCs of diabetes with an increase in pro-inflammatory cytokines (Il4, Tlr4, and Tnf11α) and a decrease in anti-inflammatory cytokine IL10. Pathway mapping demonstrated inflammation mediated by chemokine, cytokine, and angiogenesis as one of the top pathways with a significantly higher number of transcripts in db/db mice. These molecular changes were reflected in an overt defect in LSK mobility in the bone marrow. miRNA downstream target analysis unveils several mRNAs targeting leukocyte migration, microglia activation, phagosome formation, and macrophage activation signaling as their primary pathways, suggesting a shift to an inflammatory phenotype. CONCLUSION: Our findings highlight that chronic diabetes adversely alters HSCs' homeostasis at the transcriptional level, thus potentially contributing to the inflammatory phenotype of HSCs under long-term diabetes. We also believe that identifying HSCs-based biomarkers in miRNAs or mRNAs could serve as diagnostic markers and potential therapeutic targets for diabetes and associated vascular complications.

Loss of Cldn5 -and increase in Irf7-in the hippocampus and cerebral cortex of diabetic mice at the early symptomatic stage.

Analyzing changes in gene expression within specific brain regions of individuals with Type 2 Diabetes (T2DM) who do not exhibit significant cognitive deficits can yield valuable insights into the mechanisms underlying the progression towards a more severe phenotype. In this study, transcriptomic analysis of the cortex and hippocampus of mice with long-term T2DM revealed alterations in the expression of 28 genes in the cerebral cortex and 15 genes in the hippocampus. Among these genes, six displayed consistent changes in both the cortex and hippocampus: Interferon regulatory factor 7 (Irf7), Hypoxia-inducible factor 3 alpha (Hif-3α), period circadian clock 2 (Per2), xanthine dehydrogenase (Xdh), and Transforming growth factor ß-stimulated clone 22/TSC22 (Tsc22d3) were upregulated, while Claudin-5 (Cldn5) was downregulated. Confirmation of these changes was achieved through RT-qPCR. At the protein level, CLDN5 and IRF7 exhibited similar alterations, with CLDN5 being downregulated and IRF7 being upregulated. In addition, the hippocampus and cortex of the T2DM mice showed decreased levels of IκBα, implying the involvement of NF-κB pathways as well. Taken together, these results suggest that the weakening of the blood-brain barrier and an abnormal inflammatory response via the Interferon 1 and NF-κB pathways underlie cognitive impairment in individuals with long-standing T2DM.

Hyperglycemia-induced Sirt3 downregulation increases microglial aerobic glycolysis and inflammation in diabetic neuropathic pain pathogenesis.

BACKGROUND: Hyperglycemia-induced neuroinflammation significantly contributes to diabetic neuropathic pain (DNP), but the underlying mechanisms remain unclear. OBJECTIVE: To investigate the role of Sirt3, a mitochondrial deacetylase, in hyperglycemia-induced neuroinflammation and DNP and to explore potential therapeutic interventions. METHOD AND RESULTS: Here, we found that Sirt3 was downregulated in spinal dorsal horn (SDH) of diabetic mice by RNA-sequencing, which was further confirmed at the mRNA and protein level. Sirt3 deficiency exacerbated hyperglycemia-induced neuroinflammation and DNP by enhancing microglial aerobic glycolysis in vivo and in vitro. Overexpression of Sirt3 in microglia alleviated inflammation by reducing aerobic glycolysis. Mechanistically, high-glucose stimulation activated Akt, which phosphorylates and inactivates FoxO1. The inactivation of FoxO1 diminished the transcription of Sirt3. Besides that, we also found that hyperglycemia induced Sirt3 degradation via the mitophagy-lysosomal pathway. Blocking Akt activation by GSK69093 or metformin rescued the degradation of Sirt3 protein and transcription inhibition of Sirt3 mRNA, which substantially diminished hyperglycemia-induced inflammation. Metformin in vivo treatment alleviated neuroinflammation and diabetic neuropathic pain by rescuing hyperglycemia-induced Sirt3 downregulation. CONCLUSION: Hyperglycemia induces metabolic reprogramming and inflammatory activation in microglia through the regulation of Sirt3 transcription and degradation. This novel mechanism identifies Sirt3 as a potential drug target for treating DNP.

NR4A3 prevents diabetes induced atrial cardiomyopathy by maintaining mitochondrial energy metabolism and reducing oxidative stress.

BACKGROUND: Atrial cardiomyopathy (ACM) is responsible for atrial fibrillation (AF) and thromboembolic events. Diabetes mellitus (DM) is an important risk factor for ACM. However, the potential mechanism between ACM and DM remains elusive. METHODS: Atrial tissue samples were obtained from patients diagnosed with AF or sinus rhythm (SR) to assess alterations in NR4A3 expression, and then two distinct animal models were generated by subjecting Nr4a3-/- mice and WT mice to a high-fat diet (HFD) and Streptozotocin (STZ), while db/db mice were administered AAV9-Nr4a3 or AAV9-ctrl. Subsequently, in vivo and in vitro experiments were conducted to assess the impact of NR4A3 on diabetes-induced atrial remodeling through electrophysiological, biological, and histological analyses. RNA sequencing (RNA-seq) and metabolomics analysis were employed to unravel the downstream mechanisms. FINDINGS: The expression of NR4A3 was significantly decreased in atrial tissues of both AF patients and diabetic mice compared to their respective control groups. NR4A3 deficiency exacerbated atrial hypertrophy and atrial fibrosis, and increased susceptibility to pacing-induced AF. Conversely, overexpression of NR4A3 alleviated atrial structural remodeling and reduced AF induction rate. Mechanistically, we confirmed that NR4A3 improves mitochondrial energy metabolism and reduces oxidative stress injury by preserving the transcriptional expression of Sdha, thereby exerting a protective influence on atrial remodeling induced by diabetes. INTERPRETATION: Our data confirm that NR4A3 plays a protective role in atrial remodeling caused by diabetes, so it may be a new target for treating ACM. FUNDING: This study was supported by the major research program of National Natural Science Foundation of China (NSFC) No: 82370316 (to Q-S. W.), No. 81974041 (to Y-P. W.), and No. 82270447 (to Y-P. W.) and Fundation of Shanghai Hospital Development Center (No. SHDC2022CRD044 to Q-S. W.).

Pharmacokinetic/Pharmacodynamic modelling of Saxagliptin and its active metabolite, 5-hydroxy Saxagliptin in rats with Type 2 Diabetes Mellitus.

BACKGROUND AND PURPOSES: It is unclear whether the parent Saxagliptin (SAX) in vivo is the same as that in vitro, which is twice that of 5-hydroxy Saxagliptin (5-OH SAX). This study is to construct a Pharmacokinetic-Pharmacodynamic (PK-PD) link model to evaluate the genuine relationship between the concentration of parent SAX in vivo and the effect. METHODS: First, we established a reliable Ultra Performance Liquid Chromatography-Mass Spectrometry (UPLC-MS/MS) method and DPP-4 inhibition ratio determination method. Then, the T2DM rats were randomly divided into four groups, intravenous injection of 5-OH SAX (0.5 mg/kg) and saline group, intragastric administration of SAX (10 mg/kg) and Sodium carboxymethyl cellulose (CMC-Na) group. Plasma samples were collected at different time points for subsequent testing. Finally, we used the measured concentrations and inhibition ratios to construct a PK-PD link model for 5-OH SAX and parent SAX. RESULTS: A two-compartment with additive model showed the pharmacokinetic process of SAX and 5-OH SAX, the concentration-effect relationship was represented by a sigmoidal Emax model and sigmoidal Emax with E0 model for SAX and 5-OH SAX, respectively. Fitting parameters showed SAX was rapidly absorbed after administration (Tmax=0.11 h, t1/2, ka=0.07 h), widely distributed in the body (V ≈ 20 L/kg), plasma exposure reached 3282.06 ng*h/mL, and the elimination half-life was 6.13 h. The maximum plasma dipeptidyl peptidase IV (DPP-4) inhibition ratio of parent SAX was 71.47%. According to the final fitting parameter EC50, EC50, 5-OH SAX=0.46EC50, SAX(parent), it was believed that the inhibitory effect of 5-OH SAX was about half of the parent SAX, which is consistent with the literature. CONCLUSIONS: The PK-PD link model of the parent SAX established in this study can predict its pharmacokinetic process in T2DM rats and the strength of the inhibitory effect of DPP-4 based on non-clinical data.

Low bone turnover is associated with advanced glycation end-products, oxidative stress, and inflammation induced by type 2 diabetes mellitus.

Type 2 diabetes mellitus (T2DM) can lead to multiple complications. T2DM-related bone damage has been linked to abnormal bone turnover, but it cannot fully explain the mechanisms of T2DM bone disease. This study attempts to elucidate the underlying mechanisms of poor bone quality in T2DM. Hence, T2DM model was induced by a high-fat diet combined with a single streptozotocin injection in 7-week-old male SD rats. Osteoblasts derived from SD rats were cultured in high glucose to mimic hyperglycemia. Low bone turnover was observed in T2DM bone with elevated levels of advanced glycation end-products (AGEs) and receptor for AGEs (RAGE). Additionally, higher levels of oxidative stress and inflammatory factors were found in T2DM bone. AGEs content in bone was pairwise correlated with RAGE, hydrogen peroxide, and inflammatory factors. Serum levels of RAGE, oxidative stress, and inflammatory factors were higher in T2DM, while AGEs content tended to be lower. Besides, 35 differentially expressed metabolites were screened in T2DM serum. Osteoblasts exposed to high glucose displayed analogous abnormal changes in these biomarkers. Thus, low bone turnover in T2DM might be partially due to excess oxidative stress and inflammation induced by AGE-RAGE signaling. Furthermore, these biomarker levels in serum were mostly consistent with bone, demonstrating their possibility for predicting bone quality in T2DM.

Hesperetin-loaded chitosan nanoparticles ameliorate hyperglycemia by regulating key enzymes of carbohydrate metabolism in a diabetic rat model.

The study aimed to investigate the potential of hesperetin-loaded chitosan nanoparticles (HSPCNPs) in alleviating hyperglycemia by modulating key enzymes in diabetic rats. Chitosan nanoparticles loaded with hesperetin were prepared using the ionic gelation method and characterized with Electron microscope (SEM), zeta potential, particle size analysis, Fourier-transform infrared (FT-IR), Energy dispersive spectroscopy (EDS) and Encapsulation efficiency and Loading efficiency. To induce diabetes, rats were fed a high-fat beef tallow diet for 28 days, then given a single dose of streptozotocin (STZ) at 35 mg/kg b.w in 0.1 M citrate buffer (pH 4.0). Rats were treated with HSPCNPs at doses of 10, 20, and 40 mg/kg b.w. The analyzed parameters included body weight, food and water intake, plasma glucose and insulin, liver and skeletal muscle glycogen levels, and carbohydrate metabolism. SEM imaging revealed dimensions between 124.2 and 251.6 nm and a mean particle size of 145.0 nm. FT-IR analysis confirmed the presence of functional groups in the chitosan nanoparticles, and the zeta potential was 35.5 mV. HSPCNP 40 mg/kg b.w significantly (p < 0.05) reduced blood glucose levels and glycosylated hemoglobin, improving body weight, food intake, and reducing water intake. In diabetic rats, enzymes for carbohydrate metabolism like fructose 1,6-bisphosphatase, phosphoenolpyruvate carboxykinase, and glucose 6-phosphatase are evaluated in the liver, while glucose 6 phosphate dehydrogenase and hexokinase activity were significantly lower. Additionally, plasma insulin levels increased, indicating enhanced insulin sensitivity. The results show that HSPCNPs at 40 mg/kg b.w. ameliorate hyperglycemia to provide robust protection against diabetic complications and significantly improve metabolic health.

Nuclear factor erythroid 2-related factor 2 activation in streptozotocin-induced diabetic rats normalize renal hemodynamics and oxygen consumption.

Background: Diabetic kidney disease is a major contributor to end stage renal disease. A change in kidney oxygen homeostasis leading to decreased tissue oxygen tension is an important factor initiating alterations in kidney function in diabetes. However, the mechanism contributing to changed oxygen homeostasis is still unclear. Hyperglycemia-induced production of reactive oxygen species and an altered response to them have previously been demonstrated. In the present study, chronic treatment with DL-sulforaphane to induce nuclear factor erythroid 2-related factor 2 (Nrf2) expression, a master transcriptional regulator binding to antioxidant response elements inducing increased protection against reactive oxygen species, is studied. Methods: Sprague-Dawley rats were made diabetic using streptozotocin and either left untreated or received daily subcutaneous injections of DL-sulforaphane for 4 weeks. Age-matched non-diabetic rats served as controls. After 4 weeks of treatment, rats were anesthetized using thiobutabarbital, and kidney functions were studied in terms of glomerular filtration rate (GFR), renal blood flow (RBF), sodium transport, kidney oxygen consumption, and kidney oxygen tension. Mitochondria was isolated from kidney cortical tissue and investigated using high-resolution respirometry. Results: GFR was increased in diabetics but not RBF resulting in increased filtration fraction in diabetics. DL-sulforaphane treatment did not affect RBF and GFR in controls but decreased the same parameters in diabetics. Increased GFR resulted in increased sodium transport and oxygen consumption, hence decreased efficiency in diabetics compared to controls. Increased oxygen consumption in diabetics resulted in decreased cortical tissue oxygen tension. DL-sulforaphane treatment decreased oxygen consumption in diabetics, whereas transport efficiency was not significantly affected. DL-sulforaphane treatment increased cortical pO2 in diabetics. Conclusions: DL-sulforaphane treatment affects renal hemodynamics, improving cortical oxygen tension but not mitochondrial efficiency.

Canagliflozin Mitigates Diabetic Cardiomyopathy through Enhanced PINK1-Parkin Mitophagy.

Diabetic cardiomyopathy (DCM) is a major determinant of mortality in diabetic populations, and the potential strategies are insufficient. Canagliflozin has emerged as a potential cardioprotective agent in diabetes, yet its underlying molecular mechanisms remain unclear. We employed a high-glucose challenge (60 mM for 48 h) in vitro to rat cardiomyocytes (H9C2), with or without canagliflozin treatment (20 µM). In vivo, male C57BL/6J mice were subjected to streptozotocin and a high-fat diet to induce diabetes, followed by canagliflozin administration (10, 30 mg·kg-1·d-1) for 12 weeks. Proteomics and echocardiography were used to assess the heart. Histopathological alterations were assessed by the use of Oil Red O and Masson's trichrome staining. Additionally, mitochondrial morphology and mitophagy were analyzed through biochemical and imaging techniques. A proteomic analysis highlighted alterations in mitochondrial and autophagy-related proteins after the treatment with canagliflozin. Diabetic conditions impaired mitochondrial respiration and ATP production, alongside decreasing the related expression of the PINK1-Parkin pathway. High-glucose conditions also reduced PGC-1α-TFAM signaling, which is responsible for mitochondrial biogenesis. Canagliflozin significantly alleviated cardiac dysfunction and improved mitochondrial function both in vitro and in vivo. Specifically, canagliflozin suppressed mitochondrial oxidative stress, enhancing ATP levels and sustaining mitochondrial respiratory capacity. It activated PINK1-Parkin-dependent mitophagy and improved mitochondrial function via increased phosphorylation of adenosine monophosphate-activated protein kinase (AMPK). Notably, PINK1 knockdown negated the beneficial effects of canagliflozin on mitochondrial integrity, underscoring the critical role of PINK1 in mediating these protective effects. Canagliflozin fosters PINK1-Parkin mitophagy and mitochondrial function, highlighting its potential as an effective treatment for DCM.