F0F1 ATP synthase regulates extracellular calcium influx in human neutrophils by interacting with Cav2.3 and modulates neutrophil accumulation in the lipopolysaccharide-challenged lung.

BACKGROUND: Neutrophils form the first line of innate host defense against invading microorganisms. We previously showed that F0F1 ATP synthase (F-ATPase), which is widely known as mitochondrial respiratory chain complex V, is expressed in the plasma membrane of human neutrophils and is involved in regulating cell migration. Whether F-ATPase performs cellular functions through other pathways remains unknown. METHODS: Blue native polyacrylamide gel electrophoresis followed by nano-ESI-LC MS/MS identification and bioinformatic analysis were used to identify protein complexes containing F-ATPase. Then, the identified protein complexes containing F-ATPase were verified by immunoblotting, immunofluorescence colocalization, immunoprecipitation, real-time RT-PCR and agarose gel electrophoresis. Immunoblotting, flow cytometry and a LPS-induced mouse lung injury model were used to assess the effects of the F-ATPase-containing protein complex in vitro and in vivo. RESULTS: We found that the voltage-gated calcium channel (VGCC) α2δ-1 subunit is a binding partner of cell surface F-ATPase in human neutrophils. Further investigation found that the physical connection between the two proteins may exist between the F1 part (α and ß subunits) of F-ATPase and the α2 part of VGCC α2δ-1. Real-time RT-PCR and PCR analyses showed that Cav2.3 (R-type) is the primary type of VGCC expressed in human neutrophils. Research on the F-ATPase/Cav2.3 functional complex indicated that it can regulate extracellular Ca2+ influx, thereby modulating ERK1/2 phosphorylation and reactive oxygen species production, which are typical features of neutrophil activation. In addition, the inhibition of F-ATPase can reduce neutrophil accumulation in the lungs of mice that were intratracheally instilled with lipopolysaccharide, suggesting that the inhibition of F-ATPase may prevent neutrophilic inflammation-induced tissue damage. CONCLUSIONS: In this study, we identified a mechanism by which neutrophil activity is modulated, with simultaneous regulation of neutrophil-mediated pulmonary damage. These results show that surface F-ATPase of neutrophils is a potential innate immune therapeutic target.

Metformin scavenges formaldehyde and attenuates formaldehyde-induced bovine serum albumin crosslinking and cellular DNA damage.

Formaldehyde (FA) can be produced in the environment and by cell metabolism and has been classified as a carcinogen in animals and humans. Metformin is the most commonly used drug for the treatment of type 2 diabetes. Metformin also has potential benefit in cancer prevention and treatment. The aim of this study was to determine whether metformin can directly react with FA and attenuate its toxicity in vitro. Metformin was incubated at pH 7.4 and 37°C in the presence of FA, and the reaction mixture was analyzed by UV spectrophotometry, high-performance liquid chromatography (HPLC), and mass spectrometry. Fluorescence spectrophotometry, immunofluorescence, and western blot were used to measure FA-induced bovine serum albumin (BSA) crosslinking and DNA damage in HepG2 cells treated with or without metformin. According to the HPLC and mass spectrometry data, we speculate that the reaction of metformin with FA (1:1) initially results in the formation of a conjugated intermediate followed by the subsequent generation of a stable six-membered ring structure. Correspondingly, metformin attenuated FA-induced fluorescence in BSA as well as the aggregation of γH2AX in HepG2 cells. These results suggest that metformin can protect protein and DNA damage induced by FA at least partly through a direct reaction process.

Regular aerobic exercise-ameliorated troponin I carbonylation to mitigate aged rat soleus muscle functional recession.

NEW FINDINGS: What is the central question of this study? What is the biological role of carbonylation in muscle age-related functional decline and how might exercise affect the carbonylation process differently compared to habitual sedentary behaviour? What is the main finding and its importance? The carbonylation of troponin I (TNNI1), tropomyosin α-1 chain and α-actinin-1 demonstrated a relationship with muscle age-related functional decline. Exercise attenuated the decline by slowing the rate of carbonylation and promoting antioxidant reactions within the muscle. As exercise demonstrated the greatest effect on TNNI1, quantification of protein carbonyls in TNNI1 may be used as a potential biomarker of muscle age-related functional decline. ABSTRACT: This study investigated the biological role of carbonylation in muscle age-related functional decline and how regular aerobic exercise may affect the carbonylation process differently from habitual sedentary behaviour. Twenty-four healthy male Sprague-Dawley (SD) rats (mean age: 23 months) were randomly divided into an old-aged sedentary control group (O-SED) and an old-aged aerobic exercise group (O-EX). The O-EX group participated in regular aerobic exercise - treadmill running - with exercise intensity increased gradually from 50-55% to 65-70% of maximum oxygen consumption ( V̇O2max ) over 10 weeks. Rats' body weight, exercise behaviour index, morphology and oxidative stress were monitored. Avidin magnetic beads and electrospray ionization quadrupole time-of-flight mass spectrometry were used for gathering and separating carbonylated proteins while western blot tested for molecular targets. O-SED and O-EX rats both had 19 oxidative modification sites for protein carbonylation. In the O-SED group, 16 specific carbonylated proteins were identified, while 16 additional specific species were also found in the O-EX group, with all 28 species demonstrating oxidative modifications. The carbonylated proteins included troponin I (TNNI1; slow skeletal muscle), tropomyosin α1 and α-actinin 1. In particular, TNNI1 carbonylation modifications were found only in sedentary rats. Aerobic exercise increased TNNI1 and Ca2+ /calmodulin-dependent protein kinase IIα expression significantly. Observations suggested that quantification of TNNI1 carbonylation may be a potential biomarker of muscle age-related functional decline. Importantly, regular aerobic exercise appeared to have antioxidant effects in the muscle that reduced TNNI1 slow carbonylation and promoted Ca2+ /calmodulin-dependent protein kinase IIα (CAMK2) and TNNI1 expression for skeletal muscle contraction regulation, thus attenuating possible age-related skeletal muscle functional decline.

Hyperglycemia induces NF-κB activation and MCP-1 expression via downregulating GLP-1R expression in rat mesangial cells: inhibition by metformin.

Hyperglycemia impairs glucagon-like peptide-1 receptor (GLP-1R) signaling in multiple cell types and thereby potentially attenuates the therapeutic effects of GLP-1R agonists. We hypothesized that the downregulation of GLP-1R by hyperglycemia might reduce the renal-protective effects of GLP-1R agonists in diabetic nephropathy (DN). In this study, we examined the effects of high glucose on the expression of GLP-1R and its signaling pathways in the HBZY-1 rat mesangial cell line. We found that high glucose reduced GLP-1R messenger RNA (mRNA) levels in HBZY-1 cells and in the renal cortex in db/db mice comparing with control groups. In consistence, GLP-1R agonist exendin-4 induced CREB phosphorylation was attenuated by high glucose but not low glucose treatment, which is paralleled with abrogated anti-inflammatory functions in HBZY-1 cells linked with nuclear factor-κB (NF-κB) activation. In consistence, GLP-1R inhibition aggravated the high glucose-induced activation of NF-κB and MCP-1 protein levels in cultured HBZY-1 cells while overexpression of GLP-1R opposite effects. We further proved that metformin restored high glucose-inhibited GLP-1R mRNA expression and decreased high glucose evoked inflammation in HBZY-1 cells. On the basis of these findings, we conclude that high glucose lowers GLP-1R expression and leads to inflammatory responses in mesangial cells, which can be reversed by metformin. These data support the rationale of combinative therapy of metformin with GLP-1R agonists in DN.

A Multiplatform Inversion Estimation of Statewide and Regional Methane Emissions in California during 2014-2016.

California methane (CH4) emissions are quantified for three years from two tower networks and one aircraft campaign. We used backward trajectory simulations and a mesoscale Bayesian inverse model, initialized by three inventories, to achieve the emission quantification. Results show total statewide CH4 emissions of 2.05 ± 0.26 (at 95% confidence) Tg/yr, which is 1.14 to 1.47 times greater than the anthropogenic emission estimates by California Air Resource Board (CARB). Some of differences could be biogenic emissions, superemitter point sources, and other episodic emissions which may not be completely included in the CARB inventory. San Joaquin Valley (SJV) has the largest CH4 emissions (0.94 ± 0.18 Tg/yr), followed by the South Coast Air Basin, the Sacramento Valley, and the San Francisco Bay Area at 0.39 ± 0.18, 0.21 ± 0.04, and 0.16 ± 0.05 Tg/yr, respectively. The dairy and oil/gas production sources in the SJV contribute 0.44 ± 0.36 and 0.22 ± 0.23 Tg CH4/yr, respectively. This study has important policy implications for regulatory programs, as it provides a thorough multiyear evaluation of the emissions inventory using independent atmospheric measurements and investigates the utility of a complementary multiplatform approach in understanding the spatial and temporal patterns of CH4 emissions in the state and identifies opportunities for the expansion and applications of the monitoring network.

Proteomic Profile of Carbonylated Proteins Screen the Regulation of Calmodulin-Dependent Protein Kinases-AMPK-Beclin1 in Aerobic Exercise-Induced Autophagy in Middle-Aged Rat Hippocampus.

BACKGROUND: Carbonylation is an oxidative modification of the proteins and a marker of oxidative stress. The accumulation of toxic carbonylated proteins might be one of the onsets of pathogenesis in hippocampal aging or neurodegeneration. Enormous evidence indicates that regular aerobic exercise might alleviate the dysfunction of carbonylated proteins, but the adaptational mechanisms in response to exercise are unclear. OBJECTIVE: This study explored the carbonyl stress mechanism in the hippocampus using proteomics and the role of calmodulin-dependent protein kinases (CAMK)-AMP-activated protein kinase (AMPK)-Beclin1 signaling pathways in alleviating aging or improving function with regular aerobic exercise. METHODS: Twenty-four healthy 13-month-old male Sprague-Dawley rats (average 693.21 ± 68.85 g) were randomly divided into middle-aged sedentary control group (M-SED, n = 12) and middle-aged aerobic exercise runner group (M-EX, n = 12). The M-EX group participated in regular aerobic exercise - treadmill running - with exercise intensity increasing gradually from 50-55% to 65-70% of maximum oxygen consumption (V˙O2max) over 10 weeks. The targeted proteins of oxidative modification were profiled by avidin magnetic beads and electrospray ionization quadrupole time-of-flight mass spectrometry (ESI-Q-TOF-MS). Western blots were used to test for molecular targets. RESULTS: Regular aerobic exercise restores the intersessional habituation and rescues the hippocampus morphological structure in middle-aged rats. -ESI-Q-TOF-MS screened 56 carbonylated proteins only found in M-SED and 16 carbonylated proteins only found in M-EX, indicating aerobic exercise decreased carbonyl stress. Intriguingly, Ca2+/CAMK II alpha (CAMKIIα) was carbonylated only in the M-SED group at the oxidative modification site of 4-hydroxynonenal adducts, while regular aerobic exercise alleviated CAMKIIα carbonylation. Regular aerobic exercise significantly increased the expression and phosphorylated, active levels of CAMKIIα and AMPKα1. It also upregulated the expression of Beclin1 and microtubule-associated protein1-light chain 3 in the hippocampus. CONCLUSION: Quantification of CAMKIIα carbonylation may be a potential biomarker of the hippocampal senescence. Additionally, regular aerobic exercise-induced autophagy via the activation of CAMK-AMPK-Beclin1 signaling pathway may mitigate the hippocampal neurodegeneration or pathological changes by alleviating protein carbonylation (carbonyl stress).

Proteomic profile of carbonylated proteins in rat liver: discovering possible mechanisms for tetracycline-induced steatosis.

To investigate biochemical mechanisms for the tetracycline-induced steatosis in rats, targeted proteins of oxidative modification were profiled. The results showed that tetracycline induced lipid accumulation, oxidative stress, and cell viability decline in HepG2 cells only under the circumstances of palmitic acid overload. Tetracycline administration in rats led to significant decrement in blood lipids, while resulted in more than four times increment in intrahepatic triacylglycerol and typical microvesicular steatosis in the livers. The triacylglycerol levels were positively correlated with oxidative stress. Proteomic profiles of carbonylated proteins revealed 26 targeted proteins susceptible to oxidative modification and most of them located in mitochondria. Among them, the long-chain specific acyl-CoA dehydrogenase was one of the key enzymes regulating fatty acid ß-oxidation. Oxidative modification of the enzyme in the tetracycline group depressed its enzymatic activity. In conclusion, the increased influx of lipid into the livers is the first hit of tetracycline-induced microvesicular steatosis. Oxidative stress is an essential part of the second hit, which may arise from the lipid overload and attack a series of functional proteins, aggravating the development of steatosis. The 26 targeted proteins revealed here provide a potential direct link between oxidative stress and tetracycline-induced steatosis.

Calcineurin-NFAT Signaling and Neurotrophins Control Transformation of Myosin Heavy Chain Isoforms in Rat Soleus Muscle in Response to Aerobic Treadmill Training.

This study elucidated the role of CaN-NFAT signaling and neurotrophins on the transformation of myosin heavy chain isoforms in the rat soleus muscle fiber following aerobic exercise training. To do so, we examined the content and distribution of myosin heavy chain (MyHC) isoforms in the rat soleus muscle fiber, the activity of CaN and expression of NFATc1 in these fibers, and changes in the expression of nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and neutrophin-3 (NT-3) in the soleus and striatum following high-and medium-intensity aerobic treadmill training. Specific pathogen-free 2 month old male Sprague-Dawley (SD) rats were randomly divided into three groups: Control group (Con, n = 8), moderate-intensity aerobic exercise group (M-Ex, n = 8) and high-intensity aerobic exercise group (H-Ex, n = 8). We used ATPase staining to identify the muscle fiber type I and II, SDS-PAGE to separate and analyze the isoforms MyHCI, MyHCIIA, MyHCIIB and MyHCIIx, and performed western blots to determine the expression of NFATc1, NGF, BDNF and NT-3. CaN activity was measured using a colorimetric assay. In the soleus muscle, 8 weeks of moderate-intensity exercise can induce transformation of MyHC IIA and MyHC IIB to MyHC IIX and MyHC I (p < 0.01), while high-intensity treadmill exercise can induce transform MyHC IIx to MyHC IIB, MyHC IIA and MyHC I (p < 0.01). In comparison to the control group, CaN activity and NFATcl protein level were significantly increased in both the M-Ex and H-Ex groups (p < 0.05, p < 0.01), with a more pronounced upregulation in the M-Ex group (p < 0.05). Eight weeks of moderate- and high-intensity aerobic exercise induced the expression of NGF, BDNF and NT-3 in the soleus muscle and the striatum (p < 0.01), with the most significant increase in the H-Ex group (p < 0.01). In the rat soleus muscle, (1) CaN-NFATcl signaling contributes to the conversion of MyHC I isoform in response to moderate-intensity exercise; (2) Neurotrophins NGF, BDNF and NT-3 might play a role in the conversion of MyHC II isoform in response to high-intensity treadmill exercise. Key pointsEight weeks of moderate-intensity treadmill training induces the transformation MyHC IIA and MyHC IIB to MyHC IIX and MyHC I in the soleus muscles, while high-intensity exercise leads to transformation of MyHC IIX to MyHC IIA, MyHC IIB and MyHC I.MyHC I conversion in response to moderate-intensity aerobic exercise is mediated by calcineurin-NFATcl signaling.Eight weeks of moderate- and high-ntensity aerobic exercise induces the expression of NGF, BDNF and NT-3 in expression noted in rats subjected to high-intensity training. NGF and NT-3 expression in the striatum is lower than in the soleus muscle, while BDNF levels are similar. Neurotrophins may be involved in mediating MyHC II conversion in response to high-intensity aerobic exercise.

Proteomic profile of carbonylated proteins in rat liver: exercise attenuated oxidative stress may be involved in fatty liver improvement.

To screen target proteins of oxidative stress which mediate the effects of exercise on preventing nonalcoholic fatty liver disease (NAFLD), the methods for selecting carbonylated proteins were modified, and carbonylated proteins were profiled. The results showed that treadmill training reduced oxidative stress and the levels of intrahepatic triglyceride (IHTG). The changes in IHTG showed a significant positive correlation with oxidative stress as indicated by malondialdehyde level. Further results from proteomics illustrated that 17 functional proteins were susceptible to oxidative modification, and exercise protected three proteins from carbonylation. The latter three proteins may serve as both direct target proteins of oxidative stress and mediators contributing to the beneficial effects of exercise. In particular, a long-chain specific acyl-CoA dehydrogenase (ACADL) which was a key enzyme in lipid metabolism was not carbonylated and with higher activities in exercise group. These findings indicate that this modified technique is practical and powerful in selecting carbonylated proteins. Long-term treadmill training is effective in ameliorating oxidative stress and preventing the accumulation of IHTG. Among the 17 target proteins of oxidative modification, three proteins contribute to the beneficial effects of exercise. Preventing ACADL from carbonylation may be involved in the physiological mechanism of exercise-induced NAFLD improvement.

Isolation technique and proteomic analysis of the erythrocyte ghosts of red-eared turtle (Trachemys scripta).

To proceed proteomic analysis of erythrocyte of the red-eared turtle Trachemys scripta, a method for obtaining turtle erythrocyte ghosts (TEG) was first developed. After hypotonic lysis using a special buffer, forcing the erythrocyte through the syringe with an "N"-shaped needle, applying low speed homogenizing and differential centrifugation, highly purified TEG fractions were obtained. The isolated TEG proteins were treated with in-gel digestion separated by SDS-PAGE or in-solution digestion followed by HPLC predissociation, and then identified by nano-ESI-LC MS/MS techniques. A total of 169 TEG proteins was identified, validated, and categorized. Among these proteins, tubulins, and cell-surface-located F-type ATP synthase revealed important information into the super tolerance of Trachemys scripta in anoxia and low temperature exposure. Altogether, this study not only provided a method to isolate high quality TEG and a dataset of comprehensive characterization of TEG proteins, but also provides a tool for proteomic research of all nucleated red blood cells, and thus opened a new research field for exploring the mechanisms of super tolerance of turtles in harsh environment.

Subhealth: definition, criteria for diagnosis and potential prevalence in the central region of China.

BACKGROUND: A full evaluation of health conditions is necessary for the effective implementation of public health interventions. However, terms to address the intermediate state between health and disease are lacking, leading the public to overlook this state and thus increasing the risks of developing disease. METHODS: A cross-sectional health survey of 1,473 randomly recruited Chinese Han adults of both sexes living in the central region of China. The criteria for diagnosis of subhealth was defined as the presence of ≥ 1 of the following abnormalities: body mass index ≥ 25 kg/m2 or waist circumference ≥ 102 cm in men and 88 cm in women; systolic pressure 120-139 mmHg and/or diastolic pressure 80-89 mmHg; serum triglyceride level ≥ 150 mg/dL and/or total cholesterol level ≥ 200 mg/dL and/or high-density lipoprotein cholesterol level < 40 mg/dL in men and 50 mg/dL in women; serum glucose level 110-125 mg/dL; estimated glomerular filtration rate 60-89 ml/min/1.73 m2; levels of liver enzymes in liver function tests between 41-59 U/L, or with fatty liver disease but < 33% of affected hepatocytes; levels of oxidative stress biomarkers beyond the reference range of 95%; or problems with both sleep quality and psychological state. RESULTS: The prevalences of subhealth and disease in the central region of China were 36.6% and 43.1%, respectively. The prevalence of disease increased from 26.3% in participants aged 20-39 years, to 47.6% and 78.9% for participants aged 40-59 years and those aged 60 years or older, respectively. Compared with participants aged 20-39, the prevalences of health and subhealth in participants aged 60 years or older decreased by 86.7% and 60.3%, respectively. The prevalence of subhealth was increased in association with increases in lifestyle risk scores, while the prevalences of both health and disease were reduced. CONCLUSION: The prevalences of subhealth and disease are high in central China. Subhealth is associated with high lifestyle risk scores. Both the health care sector and the public should pay more attention to subhealth. Lifestyle modifications and/or psychological interventions are needed to ameliorate these conditions.

Role of neutrophils in different stages of atherosclerosis.

Neutrophils constitute the first line of defense in human immunity and can be attracted to inflamed and infected sites by various chemokines. As essential players in immune processes, neutrophils theoretically play integral roles in the course of chronic inflammation-induced atherosclerosis. However, because neutrophils are rarely found in atherosclerotic lesions, their involvement in the pathophysiological progression of atherosclerosis has been largely underestimated or ignored. Recent research has revealed convincing evidence showing the presence of neutrophils in atherosclerotic lesions and has revealed neutrophil contributions to different atherosclerosis stages in mice and humans. This review describes the underlying mechanisms of neutrophils in different stages of atherosclerosis and highlights potential neutrophil-targeted therapeutic strategies relevant to atherosclerosis. An in-depth understanding of neutrophils' roles in atherosclerosis pathology will promote exploration of new methods for the prevention and treatment of atherogenesis and atherothrombosis.

Naringenin protects pancreatic ß cells in diabetic rat through activation of estrogen receptor ß.

Naringenin is a citrus flavonoid that potently improves metabolic parameters in animal models of metabolic disorders, such as type 2 diabetes. Estrogen receptor (ER) activation promotes ß cell function and survival, thereby improving systemic glucose metabolism. In this study, we used a luciferase reporter assay, isolated rat islets and a diabetic rat model to investigate the effects of naringenin on ER signaling and the underlying mechanism of naringenin-mediated improvement of islet function in diabetes. Naringenin specifically activated ERß without affecting the activity of ERα, G protein-coupled estrogen receptor (GPER) or estrogen-related receptor (ERR) α/ß/γ. Additionally, treatment with naringenin enhanced glucose-stimulated insulin secretion in isolated rat islets. This effect was abrogated by PHTPP, an ERß antagonist. Transcriptomic analysis revealed that naringenin upregulated the expression of genes, such as Pdx1 and Mafa, which are closely linked to improved ß-cell function. In consistence, single administration of naringenin to normal rats elevated plasma insulin levels and improved glucose responses. These beneficial effects were blocked by PHTPP. In streptozocin-nicotinamide induced diabetic rats, treatment for 2 weeks with naringenin alone, but not in combination with PHTPP, significantly restored pancreatic ß cell mass and improved glucose metabolism. Collectively, these data support that naringenin specifically activate ERß to improve insulin secretion in the primary rat islets. Furthermore, naringenin administration also protected ß cell function and reversed glucose dysregulation in diabetic rats. These beneficial effects are at least partially dependent on the ERß pathway.

Niacin exacerbates ß cell lipotoxicity in diet-induced obesity mice through upregulation of GPR109A and PPARγ2: Inhibition by incretin drugs.

The widely used lipid-lowering drug niacin was reported to increase blood glucose in diabetes. How does niacin regulate ß Cell function in diabetic patients remains unclear. This study aimed to investigate the effect of niacin on ß cell lipotoxicity in vitro and in vivo. Niacin treatment sensitized the palmitate-induced cytotoxicity and apoptosis in INS-1 cells. In addition, palmitate significantly increased the niacin receptor GPR109A and PPARγ2 levels, which could be further boosted by niacin co-treatment, creating a vicious cycle. In contrast, knocking down of GPR109A could reverse both PPARγ2 expression and niacin toxicity in the INS-1 cells. Interestingly, we found that GLP-1 receptor agonist exendin-4 showed similar inhibitive effects on the GPR109A/PPARγ2 axis and was able to reverse niacin induced lipotoxicity in INS-1 cells. In diet-induced obesity (DIO) mouse model, niacin treatment resulted in elevated blood glucose, impaired glucose tolerance and insulin secretion, accompanied by the change of islets morphology and the decrease of ß cell mass. The combination of niacin and DPP-4 inhibitor sitagliptin can improve glucose tolerance, insulin secretion and islet morphology and ß cell mass, even better than sitagliptin alone. Our results show that niacin increased ß cell lipotoxicity partially through upregulation of GPR109A and PPARγ2, which can be alleviated by incretin drugs. We provide a new mechanism of niacin toxicity, and suggest that the combination of niacin and incretin may have better blood glucose and lipid control effect in clinical practice.

Plasma reactive carbonyl species levels and risk of non-alcoholic fatty liver disease.

BACKGROUND AND AIM: Oxidative stress plays a critical role in the pathogenesis of non-alcoholic fatty liver disease (NAFLD). However, there is still no large cohort study to explore the direct risk role of oxidative stress for NAFLD. This study is to test the hypothesis that elevated oxidative stress is a direct risk factor for the pathogenesis of NAFLD under controlling the potential effects of covariates. METHODS: The levels of serum cholesterol, serum triglyceride, fasting plasma glucose and plasma reactive carbonyl species (RCS) were measured from 1204 Chinese Han adults, and the questionnaire and physical examination were administered to those with known and suspected risk factors for NAFLD. RESULTS: Statistically significant high levels of blood pressure, fasting plasma glucose, serum cholesterol and triglyceride, body mass index, serum alanine aminotransferase and aspartate aminotransferase, and plasma RCS were observed in NAFLD subjects compared to healthy subjects (P < 0.01). Multivariate-adjusted odds ratio illustrated that, compared with the lowest quartile of plasma RCS levels, the highest quartile subjects had a 132% increase in the risk of developing NAFLD. Further results from multi-interaction analysis demonstrated that the underlying mechanism of the risk of NAFLD by unhealthy physical conditions and lifestyles might be, at least in part, through the oxidative stress. CONCLUSIONS: Our findings provide credible evidence from a large population that oxidative stress, as indicated by plasma RCS levels, may be a direct risk factor for developing NAFLD.

Regular Aerobic Exercise-Alleviated Dysregulation of CAMKIIα Carbonylation to Mitigate Parkinsonism via Homeostasis of Apoptosis With Autophagy.

This study investigated carbonylation of proteins with oxidative modification profiling in the striatum of aging and Parkinson disease (PD) rats, as well as the long-term effects of regular aerobic exercise on the carbonylation process and the damaging effects of PD vs habitual sedentary behavior. Regular aerobic exercise improved the PD rats' rotational behavior, increased tyrosine hydroxylase expression in both the striatum and substantia nigra pars compacta, and decreased α-synuclein expression significantly. Interestingly, apoptotic nuclei and autophagosomes were increased in the aerobic exercise PD rat striatum. Carbonylated protein Ca2+/calmodulin-dependent protein kinase alpha (CAMKIIα) was present in the middle-aged and aged groups but only in the sedentary, not the exercise, PD rat striatum. Notably, CAMKIIα was characterized by a 4-hydroxynonenal adduct. Regular aerobic exercise upregulated CAMKIIα expression level, activated the CAMK signaling pathway, and promoted the expression of autophagy markers Beclin1 and microtubule-associated proteins 1 A/1B light chain 3II. Aberrant carbonylation of CAMKII initiated age-related changes and might be useful as a potential biomarker of PD. Regular aerobic exercise alleviated protein carbonylation modification of CAMKIIα and regulated the CAMK signaling pathway, thereby affecting and regulating the homeostasis of apoptosis and autophagy in the striatum to alleviate the neurodegenerative process of PD lesions.

Carbonyl stress: malondialdehyde induces damage on rat hippocampal neurons by disturbance of Ca(2+) homeostasis.

The objective of this study was to investigate the influences of carbonyl stress induced by malondialdehyde (MDA), a typical intermediate of lipid peroxidation, on intracellular free Ca(2+) concentration ([Ca(2+)](i)) alterations in cultured hippocampal neurons of rat. The microphotographic study clearly demonstrated that the hippocampal neurons became gradually damaged following exposure to different concentrations of MDA. Further study indicated that the plasma membrane Ca(2+)-ATPase (PMCA) activity was inhibited by MDA in a concentration- and time-dependent manner. The supplementation of 100 microM MDA was found to cause a notable early phase increase of [Ca(2+)](i) in hippocampal neuron cultures followed by a more pronounced late-phase elevation of [Ca(2+)](i). Such effect of MDA was prevented by the addition of nimodipine, an inhibitor of L-type calcium channel or by an extracellular Ca(2+) chelator EGTA. The identification of the calcium signalling pathways were studied by applying U73122, an inhibitor of PL-C, and H-89, an inhibitor of protein kinase A (PKA), showing the involvement of PL-C/IP3 pathway but not the PKA/cAMP pathway. These results suggested that MDA-related carbonyl stress caused damages of rat hippocampal neurons by triggering Ca(2+) influx and influencing Ca(2+) homeostasis in cultured neurons, and also MDA may act as a signalling molecule regulating Ca(2+) release from intracellular stores.

The balance of apoptosis and autophagy via regulation of the AMPK signal pathway in aging rat striatum during regular aerobic exercise.

The objective was to analyze the effects of aerobic exercise on aging striatum stress resistance, and the adaptive mechanisms related to neurodegenerative diseases, and the occurrence, and development of neural degeneration. The 10-weeks of regular moderate-intensity aerobic exercise intervention were carried out in the aerobic exercise runner Sprague-Dawley rats. Apoptotic nuclei appeared in the striatum of aged rats, showing a tendency to relate to aging. The apoptotic index of the striatum in young, middle-aged, and old-aged rats of the aerobic exercise groups increased by 205.56%, 57%, and 68.24%. Autophagy markers Beclin l and LC 3-II expression, AMPKα1 and pAMPKα1 expression increased significantly in all age-exercise groups. The ratio of AMPKα1/pAMPKα1 increased after exercise, and the tendency of exercise to alter autophagy and cell apoptosis increased with aging. Then SirT2 mRNA was significantly upregulated in the aerobic exercise runner groups. In conclusion, we showed that the balance of autophagy and apoptosis were closely regulated by regular aerobic exercise, which affected the development of aging, and via regulation of the AMPK/SirT2 signaling pathway.

MicroRNA Expression Profiling Screen miR-3557/324-Targeted CaMK/mTOR in the Rat Striatum of Parkinson's Disease in Regular Aerobic Exercise.

This study aimed to screen the target miRNAs and to investigate the differential miR-3557/324-targeted signal mechanisms in the rats' model of Parkinson's disease (PD) with regular aerobic exercise. Rats were divided into sedentary control PD group (SED-PD, n = 18) and aerobic exercise PD group (EX-PD, n = 22). After 8 weeks of regular aerobic exercise, a 6-hydroxydopamine- (6-OHDA-) induced PD lesion model was constructed. Preregular aerobic exercises enhanced the injury resistance of rats with 6-OHDA-induced PD. The rotational behavior after injection of apomorphine hydrochloride was alleviated. Under the scanning electron microscopy, we found the neurons, axons, and villi of the striatum were clearly and tightly arranged, and neurons and axons significantly becoming larger. Tyrosine hydroxylase (TH) was increased significantly and α-synuclein protein expression was reduced in the EX-PD group compared to the SED-PD group. Screening from miRNA microarray chip, we further found upregulation of miR-3557 and downregulation of miR-324 were closely related to the calcium-modulating signaling pathway, remitting the progress of Parkinson's disease on aerobic exercise. Compared to the SED-PD group, Ca2+/calmodulin dependent protein kinase II (CaMK2α) was upregulated, but CaMKV and voltage-dependent anion-selective channel protein 1 (Vdac1) were significantly downregulated in the EX-PD group. Additionally, phosphatidylinositol-3-kinase (PI3K)/mammalian target of rapamycin (mTOR) expression were activated, and ubiquitin carboxy-terminal hydrolase L1 (UCH-L1) expression was upregulated in the EX-PD group. Conclusions: the adaptive mechanism of regular aerobic exercise delaying neurodegenerative diseases and lesions was that miR-3557/324 was activated to regulate one of its targets CaMKs signaling pathways. CaMKs, coordinated with mTOR pathway-related gene expression, improved UCH-L1 level to favor for delaying neurodegeneration or improving the pathogenesis of PD lesions.

A comparative study of artificial ceroid/lipofuscin from different tissue materials of rats.

The artificial ceroid/lipofuscin pigments originated from different organ tissues, including liver, brain, heart, and kidney of rats, and biomaterials were studied with improved fluorometric techniques. With all tissue materials exposed under ultraviolet (UV) light, a series of similar fluorescent colors were observed under microfluorometer. Analogous fluorescence spectra were also demonstrated with a three-dimensional (3-D) front-surface fluorometric technique despite of the tissue differences. Measured with 3-D fluorometry, relatively simple lipofuscin-like fluorophores were observed from the reactions of malondialdehyde (MDA) with critical biological macromolecules, such as bovine serum albumin (BSA) and DNA. Our results demonstrated that the biomaterials from different tissues have a similar fate under accelerated oxidative/carbonyl stresses but may be differentiated by a fluorescence intensity ratio.