Impaired Autophagy Research Articles

Common Pathogenetic Pathways of Non-Alcoholic Fatty Liver Disease and Type 2 Diabetes Mellitus.

Type 2 diabetes mellitus (T2DM) and non-alcoholic fatty liver disease (NAFLD) are two cardinal manifestations of the metabolic syndrome, which is becoming a growing global pandemic and a health care burden. They constitute a pathogenetic duo, with complex interplay through interrelated, but still partly understood, pathophysiological pathways, which mainly involve lipid toxicity (expressed through increased hepatic de novo lipogenesis, hepatic and peripheral insulin resistance, upregulated lipolysis, lipoprotein abnormalities, hyperinsulinemia), impaired autophagy, mitochondrial dysfunction, endoplasmic reticulum stress, adipose tissue dysfunction with a consequent latent inflammatory state, inflammasome activation, genetic and epigenetic factors, altered gut microbiota and finally dietary factors. In this review, based on data from recent studies and focusing mainly on common molecular mechanisms, we will highlight the common pathophysiological grounds and the interplay between NAFLD and T2DM.

The SGLT2 inhibitor dapagliflozin improves kidney function in glycogen storage disease XI.

Glycogen storage disease XI, also known as Fanconi-Bickel syndrome (FBS), is a rare autosomal recessive disorder caused by mutations in the SLC2A2 gene that encodes the glucose-facilitated transporter type 2 (GLUT2). Patients develop a life-threatening renal proximal tubule dysfunction for which no treatment is available apart from electrolyte replacement. To investigate the renal pathogenesis of FBS, SLC2A2 expression was ablated in mouse kidney and HK-2 proximal tubule cells. GLUT2Pax8Cre+ mice developed time-dependent glycogen accumulation in proximal tubule cells and recapitulated the renal Fanconi phenotype seen in patients. In vitro suppression of GLUT2 impaired lysosomal autophagy as shown by transcriptomic and biochemical analysis. However, this effect was reversed by exposure to a low glucose concentration, suggesting that GLUT2 facilitates the homeostasis of key cellular pathways in proximal tubule cells by preventing glucose toxicity. To investigate whether targeting proximal tubule glucose influx can limit glycogen accumulation and correct symptoms in vivo, we treated mice with the selective SGLT2 inhibitor dapagliflozin. Dapagliflozin reduced glycogen accumulation and improved metabolic acidosis and phosphaturia in the animals by normalizing the expression of Napi2a and NHE3 transporters. In addition, in a patient with FBS, dapagliflozin was safe, improved serum potassium and phosphate concentrations, and reduced glycogen content in urinary shed cells. Overall, this study provides proof of concept for dapagliflozin as a potentially suitable therapy for FBS.

Treatment of calcific arterial disease via enhancement of autophagy using GSK343

Vascular calcification is a hallmark of atherosclerotic disease and serves as a strong predictor and risk factor for cardiovascular events. Growing evidence suggests that autophagy may play a protective role in early atherosclerosis. The precise effects of autophagy on VSMC-mediated calcification remain unknown. In this study, we utilized multi-omic profiling to investigate impaired autophagy at the transcriptional level as a key driver of VSMC calcification. Our findings revealed that impaired autophagy is an essential determinant of VSMC calcification. We observed that an osteogenic environment affects the open chromatin status of VSMCs, compromising the transcriptional activation of autophagy initiation genes. Invivo experiments involve pharmacological and genetic activation of autophagy using mouse models of spontaneous large (Mgp-/-) and small (Abcc6-/-) artery calcification. Taken together, these data advance our mechanistic understanding of vascular calcification and provide important insights for a broad range of cardiovascular diseases involving VSMC phenotype switch.

Novel TFG mutation causes autosomal-dominant spastic paraplegia and defects in autophagy

BackgroundMutations in the tropomyosin receptor kinase fused (TFG) gene are associated with various neurological disorders, including autosomal recessive hereditary spastic paraplegia (HSP), autosomal dominant hereditary motor and sensory neuropathy with...

Impaired autophagy in the lower airways and lung parenchyma in stable COPD.

There is increasing evidence of autophagy activation in COPD, but its role is complex and probably regulated through cell type-specific mechanisms. This study aims to investigate the autophagic process at multiple levels within the respiratory system, using different methods to clarify conflicting results reported so far. This cross-sectional study was performed on bronchial biopsies and peripheral lung samples obtained from COPD patients (30 and 12 per sample type, respectively) and healthy controls (25 and 22 per sample type, respectively), divided by smoking history. Subjects were matched for age and smoking history. We analysed some of the most important proteins involved in autophagosome formation, such as LC3 and p62, as well as some molecules essential for lysosome function, such as lysosome-associated membrane protein 1 (LAMP1). Immunohistochemistry was used to assess the autophagic process in both sample types. ELISA and transcriptomic analysis were performed on lung samples. We found increased autophagic stimulus in smoking subjects, regardless of respiratory function. This was revealed by immunohistochemistry through a significant increase in LC3 (p<0.01) and LAMP1 (p<0.01) in small airway bronchiolar epithelium, alveolar septa and alveolar macrophages. Similar results were obtained in bronchial biopsy epithelium by evaluating LC3B (p<0.05), also increased in homogenate lung tissue using ELISA (p<0.05). Patients with COPD, unlike the others, showed an increase in p62 by ELISA (p<0.05). No differences were found in transcriptomics analysis. Different techniques, applied at post-transcriptional level, confirm that cigarette smoke stimulates autophagy at multiple levels inside the respiratory system, and that autophagy failure may characterise COPD.

Impaired Autophagic Flux in Glucose-Deprived Cells: An Outcome of Lysosomal Acidification Failure Exacerbated by Mitophagy Dysfunction.

Autophagy dysfunction is associated with human diseases and conditions including neurodegenerative diseases, metabolic issues, and chronic infections. Additionally, the decline in autophagic activity contributes to tissue and organ dysfunction and aging-related diseases. Several factors, such as down-regulation of autophagy components and activators, oxidative damage, microinflammation, and impaired autophagy flux, are linked to autophagy decline. An autophagy flux impairment (AFI) has been implicated in neurological disorders and in certain other pathological conditions. Here, to enhance our understanding of AFI, we conducted a comprehensive literature review of findings derived from two well-studied cellular stress models: glucose deprivation and replicative senescence. Glucose deprivation is a condition in which cells heavily rely on oxidative phosphorylation for ATP generation. Autophagy is activated, but its flux is hindered at the autolysis step, primarily due to an impairment of lysosomal acidity. Cells undergoing replicative senescence also experience AFI, which is also known to be caused by lysosomal acidity failure. Both glucose deprivation and replicative senescence elevate levels of reactive oxygen species (ROS), affecting lysosomal acidification. Mitochondrial alterations play a crucial role in elevating ROS generation and reducing lysosomal acidity, highlighting their association with autophagy dysfunction and disease conditions. This paper delves into the underlying molecular and cellular pathways of AFI in glucose-deprived cells, providing insights into potential strategies for managing AFI that is driven by lysosomal acidity failure. Furthermore, the investigation on the roles of mitochondrial dysfunction sheds light on the potential effectiveness of modulating mitochondrial function to overcome AFI, offering new possibilities for therapeutic interventions.

Intermittent hypoxia therapy ameliorates beta-amyloid pathology via TFEB-mediated autophagy in murine Alzheimer's disease

BackgroundAlzheimer's disease (AD) is the most prevalent neurodegenerative disorder. Impaired autophagy in plaque-associated microglia (PAM) has been reported to accelerate amyloid plaque deposition and cognitive impairment in AD pathogenesis. Recent evidence suggests that the transcription factor EB (TFEB)-mediated activation of the autophagy–lysosomal pathway is a promising treatment approach for AD. Moreover, the complementary therapy of intermittent hypoxia therapy (IHT) has been shown to upregulate autophagy and impart beneficial effects in patients with AD. However, the effect of IHT on PAM remains unknown.Methods8-Month-old APP/PS1 mice were treated with IHT for 28 days. Spatial learning memory capacity and anxiety in mice were investigated. AD pathology was determined by the quantity of nerve fibers and synapses density, numbers of microglia and neurons, Aβ plaque deposition, pro-inflammatory factors, and the content of Aβ in the brain. TFEB-mediated autophagy was determined by western blot and qRT-PCR. Primary microglia were treated with oligomeric Aβ 1–42 (oAβ) combined with IHT for mechanism exploration. Differential genes were screened by RNA-seq. Autophagic degradation process of intracellular oAβ was traced by immunofluorescence.ResultsIn this study, we found that IHT ameliorated cognitive function by attenuating neuronal loss and axonal injury in an AD animal model (APP/PS1 mice) with beta-amyloid (Aβ) pathology. In addition, IHT-mediated neuronal protection was associated with reduced Aβ accumulation and plaque formation. Using an in vitro PAM model, we further confirmed that IHT upregulated autophagy-related proteins, thereby promoting the Aβ autophagic degradation by PAM. Mechanistically, IHT facilitated the nuclear localization of TFEB in PAM, with TFEB activity showing a positive correlation with Aβ degradation by PAM in vivo and in vitro. In addition, IHT-induced TFEB activation was associated with the inhibition of the AKT–MAPK–mTOR pathway.ConclusionsThese results suggest that IHT alleviates neuronal damage and neuroinflammation via the upregulation of TFEB-dependent Aβ clearance by PAM, leading to improved learning and memory in AD mice. Therefore, IHT may be a promising non-pharmacologic therapy in complementary medicine against AD.Graphical

The Protective Pathways Activated in Kidneys of αMUPA Transgenic Mice Following Ischemia\Reperfusion-Induced Acute Kidney Injury.

Despite the high prevalence of acute kidney injury (AKI), the therapeutic approaches for AKI are disappointing. This deficiency stems from the poor understanding of the pathogenesis of AKI. Recent studies demonstrate that αMUPA, alpha murine urokinase-type plasminogen activator (uPA) transgenic mice, display a cardioprotective pathway following myocardial ischemia. We hypothesize that these mice also possess protective renal pathways. Male and female αMUPA mice and their wild type were subjected to 30 min of bilateral ischemic AKI. Blood samples and kidneys were harvested 48 h following AKI for biomarkers of kidney function, renal injury, inflammatory response, and intracellular pathways sensing or responding to AKI. αMUPA mice, especially females, exhibited attenuated renal damage in response to AKI, as was evident from lower SCr and BUN, normal renal histology, and attenuated expression of NGAL and KIM-1. Notably, αMUPA females did not show a significant change in renal inflammatory and fibrotic markers following AKI as compared with wild-type (WT) mice and αMUPA males. Moreover, αMUPA female mice exhibited the lowest levels of renal apoptotic and autophagy markers during normal conditions and following AKI. αMUPA mice, especially the females, showed remarkable expression of PGC1α and eNOS following AKI. Furthermore, MUPA mice showed a significant elevation in renal leptin expression before and following AKI. Pretreatment of αMUPA with leptin-neutralizing antibodies prior to AKI abolished their resistance to AKI. Collectively, the kidneys of αMUPA mice, especially those of females, are less susceptible to ischemic I/R injury compared to WT mice, and this is due to nephroprotective actions mediated by the upregulation of leptin, eNOS, ACE2, and PGC1α along with impaired inflammatory, fibrotic, and autophagy processes.

LIPUS combined with TFSC alleviates premature ovarian failure by promoting autophagy and inhibiting apoptosis

Objective Premature ovarian failure (POF), also known as primary ovarian insufficiency, is a major cause of infertility in female worldwide. Excessive apoptosis and impaired autophagy in ovarian granulosa cells are the main pathological mechanisms of POF. The total flavonoids from semen cuscutae (TFSC) are often used in the treatment of gynecological endocrine disorders. In addition, low intensity pulsed ultrasound (LIPUS) is report as an effective method to improve ovarian function. This study aims to investigate the protective effect of POF by the combined use of TFSC and LIPUS. Methods POF rats model and granulosa cell model were successfully induced by tripterygium glycosides and cyclophosphamide, respectively. After that, model rats and cells received TFSC plus LIPUS administration. Then ovarian histomorphology, senescence, estrus cycle, and serum sex hormone levels were detected in rats. Ovarian tissue and granulosa cells autophagy and apoptosis levels were also assessed. Results Disturbed sex hormone levels, atrophied and senescent ovaries, and abnormal estrous cycle were found in POF rats. Meanwhile, cell autophagy was inhibited and cell apoptosis was activated in POF ovarian tissue and granulosa cells. However, TFSC combined with LIPUS improved these changes, and this combination treatment exhibited synergistic effects. The abnormal expression of the cell apoptosis-, autophagy-, and PI3K/AKT/mTOR signaling pathway-related proteins were also improved by combination treatment. Conclusion The study found that the combination of TFSC and LIPUS can alleviate POF by modulating cell autophagy and apoptosis. The findings may provide a viable scientific basis for POF treatment.

The STING-IRF3 Signaling Pathway, Mediated by Endoplasmic Reticulum Stress, Contributes to Impaired Myocardial Autophagic Flux After Ischemia/Reperfusion.

This study aimed to determine whether endoplasmic reticulum (ER) stress is involved in impaired autophagy after myocardial ischemia/reperfusion (M-I/R) and elucidate the underlying mechanisms. The expression levels of stimulator of interferon gene (STING) and interferon regulatory transcription factor 3 (IRF3) phosphorylation increased in M-I/R heart tissues and hypoxia-treated/reoxygenation-treated H9c2 cells. The ER stress inhibitor 4-phenylbutyric acid (4-PBA) significantly suppressed the stimulation of STING-IRF3 transcription and alleviated cardiac dysfunction caused by M-I/R injury. In addition, 4-PBA reversed ischemia-induced/reperfusion-induced autophagic flux dysfunction, as demonstrated by a decrease in p 62 and LC3 levels. Similarly, the protective effect of STING deficiency on myocardial cell damage was achieved by the recovery of autophagic flux. Conversely, the protective effect of 4-PBA against hypoxia/reoxygenation injury in cardiomyocytes was offset by STING overexpression, wherein the activated STING-IRF3 pathway promoted the expression of Rubicon (a negatively-regulated autophagic molecule) by binding to the Rubicon promoter. Rubicon ablation effectively counteracts the adverse effects of STING overexpression in cardiomyocytes. The data showed that STING-IRF3 signaling of ER stress receptors is particularly important in the progression of physiological M-I/R caused by the inhibition of autophagic flow in vivo and in vitro.

Ligand-dependent hedgehog signaling maintains an undifferentiated, malignant osteosarcoma phenotype

TP53 and RB1 loss-of-function mutations are common in osteosarcoma. During development, combined loss of TP53 and RB1 function leads to downregulation of autophagy and the aberrant formation of primary cilia, cellular organelles essential for the transmission of canonical Hedgehog (Hh) signaling. Excess cilia formation then leads to hypersensitivity to Hedgehog (Hh) ligand signaling. In mouse and human models, we now show that osteosarcomas with mutations in TP53 and RB1 exhibit enhanced ligand-dependent Hh pathway activation through Smoothened (SMO), a transmembrane signaling molecule required for activation of the canonical Hh pathway. This dependence is mediated by hypersensitivity to Hh ligand and is accompanied by impaired autophagy and increased primary cilia formation and expression of Hh ligand in vivo. Using a conditional genetic mouse model of Trp53 and Rb1 inactivation in osteoblast progenitors, we further show that deletion of Smo converts the highly malignant osteosarcoma phenotype to benign, well differentiated bone tumors. Conversely, conditional overexpression of SHH ligand, or a gain-of-function SMO mutant in committed osteoblast progenitors during development blocks terminal bone differentiation. Finally, we demonstrate that the SMO antagonist sonidegib (LDE225) induces growth arrest and terminal differentiation in vivo in osteosarcomas that express primary cilia and Hh ligand combined with mutations in TP53. These results provide a mechanistic framework for aberrant Hh signaling in osteosarcoma based on defining mutations in the tumor suppressor, TP53.

Chloroquine impairs maximal transdiaphragmatic pressure generation in old mice.

Aging results in increased neuromuscular transmission failure and denervation of the diaphragm muscle, as well as decreased force generation across a range of motor behaviors. Increased risk for respiratory complications in old age is a major health problem. Aging impairs autophagy, a tightly regulated multistep process responsible for clearing misfolded or aggregated proteins and damaged organelles. In motor neurons, aging-related autophagy impairment may contribute to deficits in neurotransmission, subsequent muscle atrophy, and loss of muscle force. Chloroquine is commonly used to inhibit autophagy. We hypothesized that chloroquine decreases transdiaphragmatic pressure (Pdi) in mice. Old mice (16-28 mo old; n = 26) were randomly allocated to receive intraperitoneal chloroquine (50 mg/kg) or vehicle 4 h before measuring Pdi during eupnea, hypoxia (10% O2)-hypercapnia (5% CO2) exposure, spontaneous deep breaths ("sighs"), and maximal activation elicited by bilateral phrenic nerve stimulation (Pdimax). Pdi amplitude and ventilatory parameters across experimental groups and behaviors were evaluated using a mixed linear model. There were no differences in Pdi amplitude across treatments during eupnea (∼8 cm H2O), hypoxia-hypercapnia (∼10 cm H2O), or sigh (∼36 cm H2O), consistent with prior studies documenting a lack of aging effects on ventilatory behaviors. In vehicle and chloroquine-treated mice, average Pdimax was 61 and 46 cm H2O, respectively. Chloroquine decreased Pdimax by 24% compared to vehicle (P < 0.05). There were no sex or age effects on Pdi in older mice. The observed decrease in Pdimax suggests aging-related susceptibility to impairments in autophagy, consistent with the effects of chloroquine on this important homeostatic process.NEW & NOTEWORTHY Recent findings suggest that autophagy plays a role in the development of aging-related neuromuscular dysfunction; however, the contribution of autophagy impairment to the maintenance of diaphragm force generation in old age is unknown. This study shows that in old mice, chloroquine administration decreases maximal transdiaphragmatic pressure generation. These chloroquine effects suggest a susceptibility to impairments in autophagy in old age.

PG545 Prevents Osteoarthritis Development by Regulating PI3K/AKT/mTOR Signaling and Activating Chondrocyte Autophagy

Introduction: Osteoarthritis (OA) is a degenerative disease common in the elderly and is characterized by joint pain, swelling, and restricted movement. In recent years, heparanase has been reported to play an important role in the development of osteoarthritic cartilage. PG545 is a heparan sulfate mimetic with heparanase inhibitory activity. In this study, the therapeutic effects and possible mechanisms of PG545 were investigated in a chondrocyte injury model induced by interleukin-1β (IL -1β). Methods: Following treatment with PG545 or the autophagy inhibitor 3-methyladenine (3-MA), chondrocyte viability was detected using Cell Counting Kit-8 and fluorescein diacetate/propidium iodide double staining. The apoptosis rate of chondrocytes was determined by flow cytometry. Expression of light chain 3 and P62 was monitored by immunofluorescence labeling. Western blot, lentivirus infection with red fluorescent protein and green fluorescent protein, and quantitative real-time polymerase chain reaction were used to determine the expression levels of chondrocyte markers, apoptosis-related factors, autophagy proteins, and key proteins of the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) pathway. The expression and activity of stress-specific enzymes such as malondialdehyde, superoxide dismutase, and catalase (CAT) were investigated. Chondrocytes with ATG5 knockdown were used to investigate the relationship between the therapeutic effect of PG545 and autophagy. The therapeutic effect of PG545 was verified in vivo. Results: PG545 had a significant protective effect on chondrocytes by reducing oxidative stress, apoptosis, and degradation of chondrocytes and increasing chondrocyte proliferation. PG545 was effective in inducing autophagy in IL-1β-treated cells, while 3-MA attenuated the effect. The PI3K/Akt/mTOR pathway may be involved in the promotion of autophagy and OA treatment by PG545. Conclusion: PG545 was able to restore impaired autophagy and autophagic flux via the PI3K/Akt/mTOR pathway, thereby delaying the progression of OA, suggesting that PG545 may be a novel therapeutic approach for OA.

The histone acetyl transferases CBP and p300 regulate stress response pathways in synovial fibroblasts at transcriptional and functional levels

The activation of stress response pathways in synovial fibroblasts (SF) is a hallmark of rheumatoid arthritis (RA). CBP and p300 are two highly homologous histone acetyl transferases and writers of activating histone 3 lysine 27 acetylation (H3K27ac) marks. Furthermore, they serve as co-factors for transcription factors and acetylate many non-histone proteins. Here we showed that p300 but not CBP protein expression was down regulated by TNF and 4-hydroxynonenal, two factors that mimic inflammation and oxidative stress in the synovial microenvironment. We used existing RNA-sequencing data sets as a basis for a further in-depth investigation of individual functions of CBP and p300 in regulating different stress response pathways in SF. Pathway enrichment analysis pointed to a profound role of CBP and/ or p300 in regulating stress response-related gene expression, with an enrichment of pathways associated with oxidative stress, hypoxia, autophagy and proteasome function. We silenced CBP or p300, and performed confirmatory experiments on transcriptome, protein and functional levels. We have identified some overlap of CBP and p300 target genes in the oxidative stress response pathway, however, with several genes being regulated in opposite directions. The majority of stress response genes was regulated by p300, with a specific function of p300 in regulating hypoxia response genes and genes encoding proteasome subunits. Silencing of p300 suppressed proteasome enzymatic activities. CBP and p300 regulated autophagy on transcriptome and functional levels. Whereas CBP was indispensable for autophagy synthesis, silencing of p300 affected late-stage autophagy. In line with impaired autophagy and proteasome function, poly-ubiquitinated proteins accumulated after silencing of p300.

Deletion of the Homocysteine Thiolactone Detoxifying Enzyme Bleomycin Hydrolase, in Mice, Causes Memory and Neurological Deficits and Worsens Alzheimer's Disease-Related Behavioral and Biochemical Traits in the 5xFAD Model of Alzheimer's Disease.

Bleomycin hydrolase (BLMH), a homocysteine (Hcy)-thiolactone detoxifying enzyme, is attenuated in Alzheimer's disease (AD) brains. Blmh loss causes astrogliosis in mice while the loss of histone demethylase Phf8, which controls mTOR signaling, causes neuropathy in mice and humans. To examine how Blmh gene deletion affects the Phf8/H4K20me1/mTOR/autophagy pathway, amyloid-β (Aβ) accumulation, and cognitive/neuromotor performance in mice. We generated a new mouse model of AD, the Blmh-/-5xFAD mouse. Behavioral assessments were conducted by cognitive/neuromotor testing. Blmh and Phf8 genes were silenced in mouse neuroblastoma N2a-APPswe cells by RNA interference. mTOR- and autophagy-related proteins, and AβPP were quantified by western blotting and the corresponding mRNAs by RT-qPCR. Aβ was quantified by western blotting (brains) and by confocal microscopy (cells). Behavioral testing showed cognitive/neuromotor deficits in Blmh-/- and Blmh-/-5xFAD mice. Phf8 was transcriptionally downregulated in Blmh-/- and Blmh-/-5xFAD brains. H4K20me1, mTOR, phospho-mTOR, and AβPP were upregulated while autophagy markers Becn1, Atg5, and Atg7 were downregulated in Blmh-/- and Blmh-/-5xFAD brains. Aβ was elevated in Blmh-/-5xFAD brains. These biochemical changes were recapitulated in Blmh-silenced N2a-APPswe cells, which also showed increased H4K20me1-mTOR promoter binding and impaired autophagy flux (Lc3-I, Lc3-II, p62). Phf8-silencing or treatments with Hcy-thiolactone or N-Hcy-protein, metabolites elevated in Blmh-/- mice, induced biochemical changes in N2a-APPswe cells like those induced by the Blmh-silencing. However, Phf8-silencing elevated Aβ without affecting AβPP. Our findings show that Blmh interacts with AβPP and the Phf8/H4K20me1/mTOR/autophagy pathway, and that disruption of those interactions causes Aβ accumulation and cognitive/neuromotor deficits.

The ameliorating effects of mesenchymal stem cells compared to α-tocopherol on apoptosis and autophagy in streptozotocin-induced diabetic rats: Implication of PI3K/Akt signaling pathway and entero-insular axis.

Bone marrow-derived mesenchymal stem cells (BM-MSCs) are considered a novel regenerative therapy that holds much potential. This study aimed to examine and compare the ameliorative effects of BM-MSCs compared to α-tocopherol (α-Toc) on apoptosis, autophagy, and β-cell function in a rat model of streptozotocin (STZ)-induced diabetes and further analyzed the implications and interrelations of the entero-insular axis, and type I phosphoinositide 3-kinase(PI3K)/Akt signaling.Forty adult male albino rats were categorized into four groups (n = 10, in each): control group, STZ-induced diabetic group (single i.p. injection of STZ 45 mg/kg), diabetic and treated with BM-MSCs injection, diabetic and treatment with α-Toc p.o. The serum glucose, insulin, nitric oxide (NO), and catalase (CAT) were measured. Histopathological examination of the pancreas, the expression levels of insulin, CD44, caspase-3, autophagy markers, P13K/Akt, and pancreas/duodenum homeobox protein 1, in pancreatic tissue, and glucose-dependent insulinotropic polypeptide (GIP) in the duodenum were detected by hematoxylin and eosin staining, immunofluorescence labeling, and by quantitative real-time polymerase chain reaction.The diabetic rats showed reduced insulin, hyperglycemia, nitrosative stress (NO, CAT), augmented apoptosis (caspase 3), impaired autophagy (p62/SQSTM1, LC3), downregulated PI3K/Akt pathway and increased GIP expression, and degeneration of pancreatic islets. Treatment with either BM-MSCs or α-Toc suppressed the nitrosative stress, reduced apoptosis, recovered autophagy, upregulated PI3K/Akt pathway, and subsequently increased insulin levels, decreased blood glucose, and downregulated GIP expression with partial restoration of pancreatic islets.Based on our findings, the cytoprotective effects of BM-MSCs and α-Toc in type 1-induced diabetes appeared to be related to repaired autophagy and recovered PI3K/Akt signaling. Moreover, we reported their novel effects on reversing intestinal GIP expression level. The effect of BM-MSCs was notably superior to that of α-Toc.

Punicalagin's Protective Effects on Parkinson's Progression in Socially Isolated and Socialized Rats: Insights into Multifaceted Pathway.

Parkinson's disease (PD) is a gradual deterioration of dopaminergic neurons, leading to motor impairments. Social isolation (SI), a recognized stressor, has recently gained attention as a potential influencing factor in the progress of neurodegenerative illnesses. We aimed to investigate the intricate relationship between SI and PD progression, both independently and in the presence of manganese chloride (MnCl2), while evaluating the punicalagin (PUN) therapeutic effects, a natural compound established for its cytoprotective, anti-inflammatory, and anti-apoptotic activities. In this five-week experiment, seven groups of male albino rats were organized: G1 (normal control), G2 (SI), G3 (MnCl2), G4 (SI + MnCl2), G5 (SI + PUN), G6 (MnCl2 + PUN), and G7 (SI + PUN + MnCl2). The results revealed significant changes in behavior, biochemistry, and histopathology in rats exposed to SI and/or MnCl2, with the most pronounced effects detected in the SI rats concurrently exposed to MnCl2. These effects were associated with augmented oxidative stress biomarkers and reduced antioxidant activity of the Nrf2/HO-1 pathway. Additionally, inflammatory pathways (HMGB1/RAGE/TLR4/NF-ᴋB/NLRP3/Caspase-1 and JAK-2/STAT-3) were upregulated, while dysregulation of signaling pathways (PI3K/AKT/GSK-3β/CREB), sustained endoplasmic reticulum stress by activation PERK/CHOP/Bcl-2, and impaired autophagy (AMPK/SIRT-1/Beclin-1 axis) were observed. Apoptosis induction and a decrease in monoamine levels were also noted. Remarkably, treatment with PUN effectively alleviated behaviour, histopathological changes, and biochemical alterations induced by SI and/or MnCl2. These findings emphasize the role of SI in PD progress and propose PUN as a potential therapeutic intervention to mitigate PD. PUN's mechanisms of action involve modulation of pathways such as HMGB1/RAGE/TLR4/NF-ᴋB/NLRP3/Caspase-1, JAK-2/STAT-3, PI3K/AKT/GSK-3β/CREB, AMPK/SIRT-1, Nrf2/HO-1, and PERK/CHOP/Bcl-2.

Oxalate disrupts monocyte and macrophage cellular function via Interleukin-10 and mitochondrial reactive oxygen species (ROS) signaling

Oxalate is a small compound found in certain plant-derived foods and is a major component of calcium oxalate (CaOx) kidney stones. Individuals that consume oxalate enriched meals have an increased risk of forming urinary crystals, which are precursors to CaOx kidney stones. We previously reported that a single dietary oxalate load induces nanocrystalluria and reduces monocyte cellular bioenergetics in healthy adults. The purpose of this study was to extend these investigations to identify specific oxalate-mediated mechanisms in monocytes and macrophages. We performed RNA-Sequencing analysis on monocytes isolated from healthy subjects exposed to a high oxalate (8 mmol) dietary load. RNA-sequencing revealed 1,198 genes were altered and Ingenuity Pathway Analysis demonstrated modifications in several pathways including Interleukin-10 (IL-10) anti-inflammatory cytokine signaling, mitochondrial metabolism and function, oxalic acid downstream signaling, and autophagy. Based on these findings, we hypothesized that oxalate induces mitochondrial and lysosomal dysfunction in monocytes and macrophages via IL-10 and reactive oxygen species (ROS) signaling which can be reversed with exogenous IL-10 or Mitoquinone (MitoQ; a mitochondrial targeted antioxidant). We exposed monocytes and macrophages to oxalate in an in-vitro setting which caused oxidative stress, a decline in IL-10 cytokine levels, mitochondrial and lysosomal dysfunction, and impaired autophagy in both cell types. Administration of exogenous IL-10 and MitoQ attenuated these responses. These findings suggest that oxalate impairs metabolism and immune response via IL-10 signaling and mitochondrial ROS generation in both monocytes and macrophages which can be potentially limited or reversed. Future studies will examine the benefits of these therapies on CaOx crystal formation and growth in vivo.

Improving lysosomal ferroptosis with NMN administration protects against heart failure.

Myocardial mitochondria are primary sites of myocardial energy metabolism. Mitochondrial disorders are associated with various cardiac diseases. We previously showed that mice with cardiomyocyte-specific knockout of the mitochondrial translation factor p32 developed heart failure from dilated cardiomyopathy. Mitochondrial translation defects cause not only mitochondrial dysfunction but also decreased nicotinamide adenine dinucleotide (NAD+) levels, leading to impaired lysosomal acidification and autophagy. In this study, we investigated whether nicotinamide mononucleotide (NMN) administration, which compensates for decreased NAD+ levels, improves heart failure because of mitochondrial dysfunction. NMN administration reduced damaged lysosomes and improved autophagy, thereby reducing heart failure and extending the lifespan in p32cKO mice. We found that lysosomal damage due to mitochondrial dysfunction induced ferroptosis, involving the accumulation of iron in lysosomes and lipid peroxide. The ameliorative effects of NMN supplementation were found to strongly affect lysosomal function rather than mitochondrial function, particularly lysosome-mediated ferroptosis. NMN supplementation can improve lysosomal, rather than mitochondrial, function and prevent chronic heart failure.

Insulin and aging - a disappointing relationship.

Experimental studies in animal models of aging such as nematodes, fruit flies or mice have observed that decreased levels of insulin or insulin signaling promotes longevity. In humans, hyperinsulinemia and concomitant insulin resistance are associated with an elevated risk of age-related diseases suggestive of a shortened healthspan. Age-related disorders include neurodegenerative diseases, hypertension, cardiovascular disease, and type 2 diabetes. High ambient insulin concentrations promote increased lipogenesis and fat storage, heightened protein synthesis and accumulation of non-functional polypeptides due to limited turnover capacity. Moreover, there is impaired autophagy activity, and less endothelial NO synthase activity. These changes are associated with mitochondrial dysfunction and oxidative stress. The cellular stress induced by anabolic activity of insulin initiates an adaptive response aiming at maintaining homeostasis, characterized by activation of the transcription factor Nrf2, of AMP activated kinase, and an unfolded protein response. This protective response is more potent in the long-lived human species than in short-lived models of aging research resulting in a stronger pro-aging impact of insulin in nematodes and fruit flies. In humans, resistance to insulin-induced cell stress decreases with age, because of an increase of insulin and insulin resistance levels but less Nrf2 activation. These detrimental changes might be contained by adopting a lifestyle that promotes low insulin/insulin resistance levels and enhances an adaptive response to cellular stress, as observed with dietary restriction or exercise.