Research Progress of Drug Delivery Systems Targeting the Kidneys.

Chronic kidney disease (CKD) affects more than 10% of the global population, and its incidence is increasing, partially due to an increase in the prevalence of disease risk factors. Acute kidney injury (AKI) is an independent risk factor for CKD and end-stage renal disease (ESRD). The pathogenic mechanisms of CKD provide several potential targets for its treatment. However, due to off-target effects, conventional drugs for CKD typically require high doses to achieve adequate therapeutic effects, leading to long-term organ toxicity. Therefore, ideal treatments that completely cure the different types of kidney disease are rarely available. Several approaches for the drug targeting of the kidneys have been explored in drug delivery system research. Nanotechnology-based drug delivery systems have multiple merits, including good biocompatibility, suitable degradability, the ability to target lesion sites, and fewer non-specific systemic effects. In this review, the development, potential, and limitations of low-molecular-weight protein-lysozymes, polymer nanomaterials, and lipid-based nanocarriers as drug delivery platforms for treating AKI and CKD are summarized.

Endoplasmic Reticulum Stress in Systemic Lupus Erythematosus and Lupus Nephritis: Potential Therapeutic Target.

Systemic lupus erythematosus (SLE) is a complex autoimmune disease. Approximately one-third to two-thirds of the patients with SLE progress to lupus nephritis (LN). The pathogenesis of SLE and LN has not yet been fully elucidated, and effective treatment for both conditions is lacking. The endoplasmic reticulum (ER) is the largest intracellular organelle and is a site of protein synthesis, lipid metabolism, and calcium storage. Under stress, the function of ER is disrupted, and the accumulation of unfolded or misfolded proteins occurs in ER, resulting in an ER stress (ERS) response. ERS is involved in the dysfunction of B cells, macrophages, T cells, dendritic cells, neutrophils, and other immune cells, causing immune system disorders, such as SLE. In addition, ERS is also involved in renal resident cell injury and contributes to the progression of LN. The molecular chaperones, autophagy, and proteasome degradation pathways inhibit ERS and restore ER homeostasis to improve the dysfunction of immune cells and renal resident cell injury. This may be a therapeutic strategy for SLE and LN. In this review, we summarize advances in this field.

Research progress on endoplasmic reticulum homeostasis in kidney diseases.

The endoplasmic reticulum (ER) plays important roles in biosynthetic and metabolic processes, including protein and lipid synthesis, Ca2+ homeostasis regulation, and subcellular organelle crosstalk. Dysregulation of ER homeostasis can cause toxic protein accumulation, lipid accumulation, and Ca2+ homeostasis disturbance, leading to cell injury and even death. Accumulating evidence indicates that the dysregulation of ER homeostasis promotes the onset and progression of kidney diseases. However, maintaining ER homeostasis through unfolded protein response, ER-associated protein degradation, autophagy or ER-phagy, and crosstalk with other organelles may be potential therapeutic strategies for kidney disorders. In this review, we summarize the recent research progress on the relationship and molecular mechanisms of ER dysfunction in kidney pathologies. In addition, the endogenous protective strategies for ER homeostasis and their potential application for kidney diseases have been discussed.

Asiatic acid alleviates cisplatin-induced renal fibrosis in tumor-bearing mice by improving the TFEB-mediated autophagy-lysosome pathway.

Nephrotoxicity is a major side effect of cisplatin treatment of solid tumors in the clinical setting. Long-term low-dose cisplatin administration causes renal fibrosis and inflammation. However, few specific medicines with clinical application value have been developed to reduce or treat the nephrotoxic side effects of cisplatin without affecting its tumor-killing effect. The present study analyzed the potential reno-protective effect and mechanism of asiatic acid (AA) in long-term cisplatin-treated nude mice suffering from tumors. AA treatment significantly attenuated renal injury, inflammation, and fibrosis induced by long-term cisplatin injection in tumor-bearing mice. AA administration notably suppressed tubular necroptosis and improved the autophagy-lysosome pathway disruption caused by chronic cisplatin treatment in tumor-transplanted nude mice and HK-2 cells. AA promoted transcription factor EB (TFEB)-mediated lysosome biogenesis and reduced the accumulation of damaged lysosomes, resulting in enhanced autophagy flux. Mechanistically, AA increased TFEB expression by rebalancing Smad7/Smad3, whereas siRNA inhibition of Smad7 or TFEB abolished the effect of AA on autophagy flux in HK-2 cells. In addition, AA treatment did not weaken, but actually enhanced the anti-tumor effect of cisplatin, as evidenced by the promoted tumor apoptosis and inhibited proliferation in nude mice. In summary, AA alleviates cisplatin-induced renal fibrosis in tumor-bearing mice by improving the TFEB-mediated autophagy-lysosome pathway.

Autophagy in peritoneal fibrosis.

Peritoneal dialysis (PD) is a widely accepted renal replacement therapy for patients with end-stage renal disease (ESRD). Morphological and functional changes occur in the peritoneal membranes (PMs) of patients undergoing long-term PD. Peritoneal fibrosis (PF) is a common PD-related complication that ultimately leads to PM injury and peritoneal ultrafiltration failure. Autophagy is a cellular process of "self-eating" wherein damaged organelles, protein aggregates, and pathogenic microbes are degraded to maintain intracellular environment homeostasis and cell survival. Growing evidence shows that autophagy is involved in fibrosis progression, including renal fibrosis and hepatic fibrosis, in various organs. Multiple risk factors, including high-glucose peritoneal dialysis solution (HGPDS), stimulate the activation of autophagy, which participates in PF progression, in human peritoneal mesothelial cells (HPMCs). Nevertheless, the underlying roles and mechanisms of autophagy in PF progression remain unclear. In this review, we discuss the key roles and potential mechanisms of autophagy in PF to offer novel perspectives on future therapy strategies for PF and their limitations.

AMP-activated protein kinase α2 contributes to acute and chronic hyperuricemic nephropathy via renal urate deposition in a mouse model.

Hyperuricemia can induce acute and chronic kidney damage, but the pathological mechanism remains unclear. The potential role of AMP-activated protein kinase (AMPK) α2 in hyperuricemia-induced renal injury was investigated in this study. Acute and chronic hyperuricemic nephropathy was induced by administering intraperitoneal injections of uric acid and oxonic acid to AMPK α2 knockout and wild-type mice. Changes in renal function, histopathology, inflammatory cell infiltration, renal interstitial fibrosis, and urate deposition were analyzed. In both acute and chronic hyperuricemic nephropathy mouse models, knockout of AMPK α2 significantly reduced serum creatinine levels and renal pathological changes. The tubular expression of kidney injury molecule-1 was also reduced in hyperuricemic nephropathy mice deficient in AMPK α2. In addition, knockout of AMPK α2 significantly suppressed the infiltration of renal macrophages and progression of renal interstitial fibrosis in mice with chronic hyperuricemic nephropathy. Knockout of AMPK α2 reduced renal urate crystal deposition, probably through increasing the expression of the uric acid transporter, multidrug resistance protein 4. In summary, AMPK α2 is involved in acute and chronic hyperuricemia-induced kidney injury and may be associated with increased urate crystal deposition in the kidney.

Metformin improves renal injury of MRL/lpr lupus-prone mice via the AMPK/STAT3 pathway.

OBJECTIVE: Lupus nephritis (LN) is a major complication and cause of death among patients with SLE. This research used in vivo and in vitro experiments to explore the therapeutic potential of metformin in kidney injury from LN-induced inflammation. METHODS: In vivo study, 8-week-old MRL/MpJ-Faslpr/J (MRL/lpr) mice were randomly divided into two groups (n=12 each): daily administration of 0.3 mg/mL metformin in drinking water and control (water only). Body weight and urinary samples were measured biweekly. Mice were sacrificed after 8-week treatment to harvest serum, lymph nodes, spleen and kidneys. In vitro study, human kidney-2 (HK-2) cells were pretreated with 1 mM metformin for 1 hour and then stimulated with 20 µg/mL lipopolysaccharides (LPS) or 10 ng/mL tumour necrosis factor-α (TNF-α) for another 48 hours. Protein was collected for subsequent analysis. RESULTS: We found that metformin administration improved renal function in MRL/lpr lupus-prone mice, measured by decreased urea nitrogen and urinary proteins. Metformin reduced immunoglobulin G and complement C3 deposition in glomeruli. The treatment also downregulated systemic and renal inflammation, as seen in decreased renal infiltration of F4/80-positive macrophages and reduced splenic and renal MCP-1 (monocyte chemoattractant protein-1) and TNF-α, and renal IL-1ß (interleukin 1ß) expression. Metformin administration decreased renal expression of necroptosis markers p-RIPK1 (phosphorylated receptor-interacting protein kinase 1) and p-MLKL, along with tubular injury marker KIM-1 (kidney injury molecule-1) in lupus mice. In addition, metformin alleviated the necroptosis of HK-2 cells stimulated by LPS and TNF-α, evidencing by a decrease in the expression of necroptosis markers p-RIPK1, p-RIPK3 and p-MLKL, and the inflammasome-related markers NLRP3 (NLR family pyrin domain containing 3), ASC (apoptosis-associated speck-like protein containing a CARD), caspase-1. Mechanistically, metformin treatment upregulated p-AMPK (phosphorylated AMP-activated protein kinase) and downregulated p-STAT3 (phosphorylated signal transducer and activator of transcription 3) expression in the kidneys. Moreover, AMPKα2 knockdown abolished the protective effects of metformin in vitro. CONCLUSIONS: Metformin alleviated kidney injury in LN though suppressing renal necroptosis and inflammation via the AMPK/STAT3 pathway.

Smad3 deficiency improves islet-based therapy for diabetes and diabetic kidney injury by promoting ß cell proliferation via the E2F3-dependent mechanism.

Rationale: Poor ß cell proliferation is one of the detrimental factors hindering islet cell replacement therapy for patients with diabetes. Smad3 is an important transcriptional factor of TGF-ß signaling and has been shown to promote diabetes by inhibiting ß cell proliferation. Therefore, we hypothesize that Smad3-deficient islets may be a novel cell replacement therapy for diabetes. Methods: We examined this hypothesis in streptozocin-induced type-1 diabetic mice and type-2 diabetic db/db mice by transplanting Smad3 knockout (KO) and wild type (WT) islets under the renal capsule, respectively. The effects of Smad3KO versus WT islet replacement therapy on diabetes and diabetic kidney injury were examined. In addition, RNA-seq was applied to identify the downstream target gene underlying Smad3-regulated ß cell proliferation in Smad3KO-db/db versus Smad3WT-db/db mouse islets. Results: Compared to Smad3WT islet therapy, treatment with Smad3KO islets produced a much better therapeutic effect on both type-1 and type-2 diabetes by significantly lowering serum levels of blood glucose and HbA1c and protected against diabetic kidney injuries by preventing an increase in serum creatinine and the development of proteinuria, mesangial matrix expansion, and fibrosis. These were associated with a significant increase in grafted ß cell proliferation and blood insulin levels, resulting in improved glucose intolerance. Mechanistically, RNA-seq revealed that compared with Smad3WT-db/db mouse islets, deletion of Smad3 from db/db mouse islets markedly upregulated E2F3, a pivotal regulator of cell cycle G1/S entry. Further studies found that Smad3 could bind to the promoter of E2F3, and thus inhibit ß cell proliferation via an E2F3-dependent mechanism as silencing E2F3 abrogated the proliferative effect on Smad3KO ß cells. Conclusion: Smad3-deficient islet replacement therapy can significantly improve both type-1 and type-2 diabetes and protect against diabetic kidney injury, which is mediated by a novel mechanism of E2F3-dependent ß cell proliferation.

Characteristics and health risks of personal exposure to particle-bound PAHs for Hong Kong adult residents: From ambient pollution to indoor exposure.

Research on individual level polycyclic aromatic hydrocarbons (PAHs) exposure is scarce. Moreover, the independent contribution of ambient- and indoor-origin PAHs to personal exposure remains poorly studied. We performed simultaneous ambient, residential indoor, and personal exposure measurements in a panel of healthy adults to investigate particle-bound PAHs, focusing on their carcinogenic congeners (cPAHs). Average PAH concentrations were much higher in ambient and residential indoor than personal exposure, with distinct seasonal variations. We employed chrysene as a tracer to investigate residential indoor and personal PAHs exposure by origin. Personal cPAH exposure was largely attributable to ambient-origin exposures (95.8%), whereas a considerable proportion of residential indoor PAHs was likely attributable to indoor emissions (33.8%). Benzo[a]pyrene equivalent (BaPeq) concentrations of cPAH accounted for 95.2%-95.6% of total carcinogenic potential. Uncertainties in estimated PAHs (and BaPeq) exposure and cancer risks for adults were calculated using the Monte Carlo simulation. Cancer risks attributable to ambient, residential indoor, and personal cPAH inhalation exposures ranged from 4.0 × 10-6 to 1.0 × 10-5 . A time-activity weighted model was employed for personal PAH exposure estimations. Estimated cPAH exposures demonstrate high cancer risks for adults in Hong Kong, suggesting that exposure to indoor-generated PAHs should be of great concern to the general population.

A prediction model based on platelet parameters, lipid levels, and angiographic characteristics to predict in-stent restenosis in coronary artery disease patients implanted with drug-eluting stents.

BACKGROUND: The present study was aimed to establish a prediction model for in-stent restenosis (ISR) in subjects who had undergone percutaneous coronary intervention (PCI) with drug-eluting stents (DESs). MATERIALS AND METHODS: A retrospective cohort study was conducted. From September 2010 to September 2013, we included 968 subjects who had received coronary follow-up angiography after primary PCI. The logistic regression analysis, receiver operator characteristic (ROC) analysis, nomogram analysis, Hosmer-Lemeshow χ2 statistic, and calibration curve were applied to build and evaluate the prediction model. RESULTS: Fifty-six patients (5.79%) occurred ISR. The platelet distribution width (PDW), total cholesterol (TC), systolic blood pressure (SBP), low-density lipoprotein cholesterol (LDL-C), and lesion vessels had significant differences between ISR and non-ISR groups (all P < 0.05). And these variables were independently associated with ISR (all P < 0.05). Furthermore, they were identified as predictors (all AUC > 0.5 and P < 0.05) to establish a prediction model. The prediction model showed a good value of area under curve (AUC) (95%CI): 0.72 (0.64-0.80), and its optimized cut-off was 6.39 with 71% sensitivity and 65% specificity to predict ISR. CONCLUSION: The incidence of ISR is 5.79% in CAD patients with DES implantation in the Xinjiang population, China. The prediction model based on PDW, SBP, TC, LDL-C, and lesion vessels was an effective model to predict ISR in CAD patients with DESs implantation.

Lysosome Depletion-Triggered Autophagy Impairment in Progressive Kidney Injury.

BACKGROUND: Macroautophagy (autophagy) is a cellular recycling process involving the destruction of damaged organelles and proteins in intracellular lysosomes for efficient nutrient reuse. SUMMARY: Impairment of the autophagy-lysosome pathway is tightly associated with multiple kidney diseases, such as diabetic nephropathy, proteinuric kidney disease, acute kidney injury, crystalline nephropathy, and drug- and heavy metal-induced renal injury. The impairment in the process of autophagic clearance may induce injury in renal intrinsic cells by activating the inflammasome, inducing cell cycle arrest, and cell death. The lysosome depletion may be a key mechanism triggering this process. In this review, we discuss this pathway and summarize the protective mechanisms for restoration of lysosome function and autophagic flux via the endosomal sorting complex required for transport (ESCRT) machinery, lysophagy, and transcription factor EB-mediated lysosome biogenesis. KEY MESSAGE: Further exploring mechanisms of ESCRT, lysophagy, and lysosome biogenesis may provide novel therapy strategies for the management of kidney diseases.

Polymorphisms of rs2483205 and rs562556 in the PCSK9 gene are associated with coronary artery disease and cardiovascular risk factors.

PCSK9 plays a crucial role in lipid metabolism. This case-control study explored the associations of novel single nucleotide polymorphisms (SNPs) of the PCSK9 gene with coronary artery disease (CAD) (≥ 1 coronary artery stenosis ≥ 50%) and its risk factors in the Han population in Xinjiang, China. Four tag SNPs (rs11583680, rs2483205, rs2495477 and rs562556) of the PCSK9 gene were genotyped in 950 CAD patients and 1082 healthy controls. The distributions of genotypes in rs2483205 and rs562556 were significantly different between the groups (all p < 0.05). The TT genotype of rs2483205, GG genotype of rs562556, and their H4 (T-G) haplotype were associated with CAD [odds ratio (OR) 0.65, confidence interval (CI) 0.45-0.95, p = 0.024; 0.63, 0.45-0.90, p = 0.011; 0.50, 0.35-0.70, p < 0.001, respectively]. Additionally, the model (TT + CT vs. CC) of rs2483205 was associated with increased risk of obesity, and the G allele of rs562556 was associated with lower low-density lipoprotein cholesterol (LDL-C), blood glucose, body mass index (BMI), and mean platelet volume (MPV) (all p < 0.05). rs2483205, rs562556, and their H4 haplotype of the PCSK9 gene were associated with CAD. Additionally, rs2483205 is associated with obesity, and rs562556 is associated with LDL-C, blood glucose, BMI, and MPV.

Cyclosporine A blocks autophagic flux in tubular epithelial cells by impairing TFEB-mediated lysosomal function.

Cyclosporine A (CsA) is an immunosuppressor widely used for the prevention of acute rejection during solid organ transplantation. However, severe nephrotoxicity has substantially limited its long-term usage. Recently, an impaired autophagy pathway was suggested to be involved in the pathogenesis of chronic CsA nephrotoxicity. However, the underlying mechanisms of CsA-induced autophagy blockade in tubular cells remain unclear. In the present study, we observed that CsA suppressed the activation and expression of transcription factor EB (TFEB) by increasing the activation of mTOR, in turn promoting lysosomal dysfunction and autophagy flux blockade in tubular epithelial cells (TECs) in vivo and in vitro. Restoration of TFEB activation by Torin1-mediated mTOR inhibition significantly improved lysosomal function and rescued autophagy pathway activity, suppressing TEC injury. In summary, targeting TFEB-mediated autophagy flux represents a potential therapeutic strategy for CsA-induced nephrotoxicity.

Toxicological effects of personal exposure to fine particles in adult residents of Hong Kong.

Toxicological studies have demonstrated the associations between fine particle (PM2.5) components and various cytotoxic endpoints. However, few studies have investigated the toxicological effects of source-specific PM2.5 at the individual level. To investigate the potential impact of source-specific PM2.5 on cytotoxic effects, we performed repeated personal PM2.5 monitoring of 48 adult participants in Hong Kong during the winter and summer of 2014-2015. Quartz filters were analyzed for carbonaceous aerosols and water-soluble ions in PM2.5. Teflon filters were collected to determine personal PM2.5 mass and metal concentrations. The toxicological effects of personal PM2.5 exposure-including cytotoxicity, inflammatory response, and reactive oxygen species (ROS) production-were measured using A549 cells in vitro. Personal PM2.5 samples collected in winter were more effective than those collected in summer at inducing cytotoxicity and the expression of proinflammation cytokine IL-6. By contrast, summer personal PM2.5 samples induced high ROS production. We performed a series of statistical analyses, Spearman correlation and a source apportionment approach with a multiple linear regression (MLR) model, to explore the sources contributing most significantly to personal PM2.5 bioreactivity. Secondary inorganic species and transition metals were discovered to be weak-to-moderately associated with cytotoxicity (rs: 0.26-0.55; p < 0.01) and inflammatory response (rs: 0.26-0.44; p < 0.05), respectively. Carbonaceous aerosols (i.e., organic and elemental carbon; rs: 0.23-0.27; p < 0.05) and crustal material (Mg and Ca) was positively associated with ROS generation. The PMF-MLR models revealed that tailpipe exhaust and secondary sulfate contributed to ROS generation, whereas secondary nitrate was the major contributor to PM2.5 cytotoxicity and inflammation. These results improve and variate the arguments for practical policies designed to mitigate the risks posed by air pollution sources and to protect public health.

SMAD3 promotes autophagy dysregulation by triggering lysosome depletion in tubular epithelial cells in diabetic nephropathy.

Macroautophagy/autophagy dysregulation has been noted in diabetic nephropathy; however, the regulatory mechanisms controlling this process remain unclear. In this study, we showed that SMAD3 (SMAD family member 3), the key effector of TGFB (transforming growth factor beta)-SMAD signaling, induces lysosome depletion via the inhibition of TFEB-dependent lysosome biogenesis. The pharmacological inhibition or genetic deletion of SMAD3 restored lysosome biogenesis activity by alleviating the suppression of TFEB, thereby protecting lysosomes from depletion and improving autophagic flux in renal tubular epithelial cells in diabetic nephropathy. Mechanistically, we found that SMAD3 directly binds to the 3'-UTR of TFEB and inhibits its transcription. Silencing TFEB suppressed lysosome biogenesis and resulted in a loss of the protective effects of SMAD3 inactivation on lysosome depletion under diabetic conditions. In conclusion, SMAD3 promotes lysosome depletion via the inhibition of TFEB-dependent lysosome biogenesis; this may be an important mechanism underlying autophagy dysregulation in the progression of diabetic nephropathy.Abbreviations: AGEs: advanced glycation end products; ATP6V1H: ATPase H+ transporting V1 subunit H; CTSB: cathepsin B; ChIP: chromatin immunoprecipitation; Co-BSA: control bovine serum albumin; DN: diabetic nephropathy; ELISA: enzyme-linked immunosorbent assay; FN1: fibronectin 1; HAVCR1/TIM1/KIM-1: hepatitis A virus cellular receptor 1; LAMP1: lysosomal associated membrane protein 1; LMP: lysosome membrane permeabilization; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; NC: negative control; SIS3: specific inhibitor of SMAD3; SMAD3: SMAD family member 3; siRNA: small interfering RNA; SQSTM1/p62: sequestosome 1; TECs: tubular epithelial cells; TFEB: transcription factor EB; TGFB1: transforming growth factor beta 1; TGFBR1: transforming growth factor beta receptor 1; UTR: untranslated region; VPS11: VPS11 core subunit of CORVET and HOPS complexes.

Characteristics and toxicological effects of commuter exposure to black carbon and metal components of fine particles (PM2.5) in Hong Kong.

Epidemiological studies have demonstrated significant associations between traffic-related air pollution and adverse health outcomes. Personal exposure to fine particles (PM2.5) in transport microenvironments and their toxicological properties remain to be investigated. Commuter exposures were investigated in public transport systems (including the buses and Mass Transit Railway (MTR)) along two sampling routes in Hong Kong. Real-time sampling for PM2.5 and black carbon (BC), along with integrated PM2.5 sampling, were performed during the warm and cold season of 2016-2017, respectively. Commuter exposure to BC during 3-hour commuting time exhibited a wider range, from 3.4 to 4.6 µg/m3 on the bus and 5.5 to 8.7 µg/m3 in MTR cabin (p < .05). PM2.5 mass and major chemical constituents (including organic carbon (OC), elemental carbon (EC), and metals) were analyzed. Cytotoxicity, including cellular reactive oxygen species (ROS) production, was determined in addition to acellular ROS generation. PM2.5 treatment promoted the ROS generation in a concentration-dependent manner. Consistent diurnal variations were observed for commuter exposure to BC and PM2.5 components, along with cellular and acellular ROS generation, which marked with two peaks during the morning (08:00-11:00) and evening rush hours (17:30-20:30). Commuter exposures in the MTR system were characterized by higher levels of PM2.5 and elemental components (e.g., Ca, Cr, Fe, Zn, Ba) compared to riding the bus, along with higher cellular and acellular ROS production (p < .01). These metals were attributed to different sources: rail tracks, wheels, brakes, and crustal origin. Weak to moderate associations were shown for the analyzed transition metals with PM2.5-induced cell viability and cellular ROS. Multiple linear regression analysis revealed that Ni, Zn, Mn, Fe, Ti, and Co attributed to cytotoxicity and ROS generation. These findings underscore the importance of commuter exposures and their toxic effects, urging effective mitigating strategies to protect human health.

Overexpression of IκBα in cardiomyocytes alleviates hydrogen peroxide-induced apoptosis and autophagy by inhibiting NF-κB activation.

BACKGROUND: Inflammation and oxidative stress play predominant roles in the initiation and progression of ischaemia/reperfusion (I/R) injury, with nuclear factor kappa B (NF-κB) serving as a crucial mediator. Overexpression of the inhibitor of κB alpha (IκBα) gene is hypothesized to have protective effects against apoptosis and autophagy in cardiomyocytes subjected to hydrogen peroxide (H2O2) by inhibiting the NF-κB pathway. METHODS: The IκBαS32A, S36A gene was transfected via adeno-associated virus serotype 9 (AAV9) delivery into neonatal rat ventricular cardiomyocytes (NRVMs) prior to H2O2 treatment. NRVMs were divided into control, H2O2, GFP + H2O2, IκBα+H2O2, and pyrrolidine dithiocarbamate (PDTC) + H2O2 groups. Nuclear translocation of the NF-κB p65 subunit was evaluated by immunofluorescence and Western blotting. Cell viability was assessed by Cell Counting Kit-8 assay. Supernatant lactate dehydrogenase (LDH) and intracellular malondialdehyde (MDA) were measured to identify H2O2-stimulated cytotoxicity. Apoptosis was determined by Annexin V-PE/7-AAD staining, and the mitochondrial membrane potential (ΔΨm) was detected by JC-1 staining. Western blotting was used to detect apoptosis- and autophagy-related proteins. RESULTS: IκBα transfection significantly increased cell viability and ΔΨm but decreased the supernatant LDH and cellular MDA levels in cardiomyocytes exposed to H2O2. Meanwhile, IκBα overexpression decreased H2O2-induced apoptosis by upregulating the Bcl-2/Bax ratio and reduced autophagy by downregulating the expression of Beclin-1 and the LC3-II/LC3-I ratio. These effects partly accounted for the ability of IκBα to inhibit the NF-κB signalling pathway, as evidenced by decreases in p65 phosphorylation and nuclear translocation. Indeed, the effects of inactivation of NF-κB signalling with the specific inhibitor PDTC resembled the cardioprotective effects of IκBα during H2O2 stimulation. CONCLUSION: IκBα overexpression can ameliorate H2O2-induced apoptosis, autophagy, oxidative injury, and ΔΨm loss through inhibition of the NF-κB signalling pathway. These findings suggest that IκBα transfection can result in successful resistance to oxidative stress-induced damage by inhibiting NF-κB activation, which may provide a potential therapeutic target for the prevention of myocardial I/R injury.

Indoor, outdoor, and personal exposure to PM2.5 and their bioreactivity among healthy residents of Hong Kong.

Direct evidence about associations between fine particles (PM2.5) components and the corresponding PM2.5 bioreactivity at the individual level is limited. We conducted a panel study with repeated personal measurements involving 56 healthy residents in Hong Kong. Fractional exhaled nitric oxide (FeNO) levels were measured from these subjects. Out of 56 subjects, 27 (48.2%) participated in concurrent outdoor, indoor, and personal PM2.5 monitoring. Organic carbon (OC), elemental carbon (EC), particle bound-polycyclic aromatic hydrocarbons (PAHs), and phthalates were analyzed. Alteration in cell viability, lactic dehydrogenase (LDH), interleukin-6 (IL-6), and 8-isoprostane by 50 µg/mL PM2.5 extracts was determined in A549 cells in vitro. Moderate heterogeneities were shown in PM2.5 exposures and the corresponding PM2.5 bioreactivity across different sample types. Associations between the analyzed components and PM2.5 bioreactivity were assessed using the multiple regression models. Toxicological results revealed that indoor and personal exposure to OC as well as PAH compounds and their derivatives (e.g., Alkyl-PAHs, Oxy-PAHs) induced cell viability reduction and increase in levels of LDH, IL-6, and 8-isoprostane. Overall, OC in personal exposure played a dominant role in PM2.5-induced bioreactivity. Subsequently, we examined the associations of FeNO with IL-6 and 8-isoprostane levels using mixed-effects models. The results showed that per interquartile change in IL-6 and 8-isoprostane were associated with a 6.4% (p < 0.01) and 11.1% (p < 0.01) increase in FeNO levels, respectively. Our study explored the toxicological properties of chemical components in PM2.5 exposure, which suggested that residential indoors and personal OC and PAHs should be of great concern for human health. These findings indicated that further studies in inflammation and oxidative stress-related illnesses due to particle exposure would benefit from the assessment of in vitro PM2.5 bioreactivity.

Autophagy Inhibition Sensitizes Renal Tubular Epithelial Cell to G1 Arrest Induced by Transforming Growth Factor beta (TGF-ß).

BACKGROUND Cell cycle arrest and autophagy have been demonstrated to be involved in various transforming growth factor (TGF)-ß-mediated phenotype alterations of tubular epithelial cells (TECs) and tubulointerstitial fibrosis. But the relationship between cell cycle arrest and the autophagy induced by TGF-ß has not been explored well. MATERIAL AND METHODS The effects of autophagy inhibition on TGF-ß-induced cell cycle arrest in TECs were explored in vitro. Human kidney-2 (HK-2) cells were stimulated by TGF-ß with or without a combined treatment of autophagy inhibitor chloroquine (CQ) or bafilomycin A1 (Baf). RESULTS Autophagy inhibition by CQ or Baf promotes the suppression of growth in TGF-ß-treated HK-2 cells, as detected by the Cell Counting Kit-8 (CCK-8) method. In addition, CQ or Baf stimulation enhances G1 arrest in TGF-ß treated HK-2 cells, as investigated using propidium iodide (PI) staining and flow cytometry, which was further confirmed by a decrease in the expression of phosphorylated retinoblastoma protein (p-RB) and cyclin-dependent kinase 4 (CDK4). The upregulation of p21 induced by CQ or Baf may mediate an enhanced G1 arrest in TGF-ß treated HK-2 cells. Western blot analysis showed that TGF-ß-induced expression of extracellular matrix fibronectin was notably upregulated in the presence of autophagy inhibitors. CONCLUSIONS Inhibition of autophagy sensitizes the TECs to G1 arrest and proliferation suppression induced by TGF-ß that contributes to the induction of tubulointerstitial fibrosis.

Apolipoprotein A1 is associated with SYNTAX score in patients with a non-ST segment elevation myocardial infarction.

BACKGROUND: The study was designed to investigate lipid profile and SYNTAX score in patients with non-ST segment elevation myocardial infarction (NSTEMI). METHODS: 311 patients with NSTEMI were enrolled. The demographic, clinical data, blood samples and SYNTAX score were documented. The Pearson linear correlation was used to detect confounding factors linearly correlated with SYNTAX score. The significantly correlated confounding factors were put into the multiple linear regressions. RESULTS: The Pearson linear correlation showed that high-density lipoprotein- cholesterol (HDL-C) and apolipoprotein A1 (ApoA1) were significantly correlated with Syntax Score (r = - 0.119, P = 0.044 and r = - 0.182, P = 0.002, respectively). The multiple linear regressions for Syntax Score were built using HDL-C and ApoA1, respectively. After the adjustment of other significantly correlated confounding factors such as white blood cell count (WBC), myohemoglobin (MB), glutamic-oxalacetic transaminase (AST) and creatinine, the ApoA1 still showed significant association with Syntax Score (ß = - 0.151, P = 0.028). The area under curve was (AUC) 0.624 and the optimal cutoff value is 1.07 g/L when using ApoA1 to predict moderate and severe coronary artery lesions. The patients with ApoA1 ≥ 1.07 g/L and < 1.07 g/L have the Syntax Scores of 12.21 ± 11.58 and 16.33 ± 11.53, respectively (P = 0.001). CONCLUSIONS: The ApoA1 is the only lipid factor significantly associated with complexity of coronary artery lesion in patients with NSTEMI, the patients with ApoA1 < 1.07 g/L may have more complex coronary artery lesions.