Vitamin D3 decreases TNF-α-induced inflammation in lung epithelial cells through a reduction in mitochondrial fission and mitophagy.

Previous work has shown an association between vitamin D3 deficiency and an increased risk for acquiring various inflammatory diseases. Vitamin D3 can reduce morbidity and mortality in these patients via different mechanisms. Lung inflammation is an important event in the initiation and development of respiratory disorders. However, the anti-inflammatory effects of vitamin D3 and the underlying mechanisms remained to be determined. The purpose of this study was to examine the effects and mechanisms of action of vitamin D3 (Vit. D) on the expression of intercellular adhesion molecule-1 (ICAM-1) in vitro and in vivo with or without tumor necrosis factor α (TNF-α) treatment. Pretreatment with Vit. D reduced the expression of ICAM-1 and leukocyte adhesion in TNF-α-treated A549 cells. TNF-α increased the accumulation of mitochondrial reactive oxygen species (mtROS), while Vit. D reduced this effect. Pretreatment with Vit. D attenuated TNF-α-induced mitochondrial fission, as shown by the increased expression of mitochondrial fission factor (Mff), phosphorylated dynamin-related protein 1 (p-DRP1), and mitophagy-related proteins (BCL2/adenovirus E1B 19 kDa protein-interacting protein 3, Bnip3) in A549 cells. Inhibition of DRP1 or Mff significantly decreased ICAM-1 expression. In addition, we found that Vit. D decreased TNF-α-induced ICAM-1 expression, mitochondrial fission, and mitophagy via the AKT and NF-κB pathways. Moreover, ICAM-1 expression, mitochondrial fission, and mitophagy were increased in the lung tissues of TNF-α-treated mice, while Vit. D supplementation reduced these effects. In this study, we elucidated the mechanisms by which Vit. D reduces the expression of adhesion molecules in models of airway inflammation. Vit. D might be served as a novel therapeutic agent for the targeting of epithelial activation in lung inflammation. Graphical Headlights: • The expression of DRP1 and Mff, mitochondrial fission-related proteins, was increased in TNF-α-treated A549 cells. • The expression of Bnip3 and LC3B, mitophagy-related proteins, was increased in TNF-α-treated A549 cells. • Vit. D pretreatment decreased TNF-α-induced inflammation through the reduction of mitochondrial fission and mitophagy in A549 cells.

Supplementation of nano-bubble curcumin extract improves gut microbiota composition and exercise performance in mice.

BACKGROUND: In a previous study, we evaluated the potential beneficial effect of nano-bubble curcumin extract (NCE) in reducing exercise-related injuries and improving performance. METHODS: In this study, we seek to investigate changes in the gut microbiota composition upon NCE supplementation in relation to health and exercise performance. Male ICR mice were divided into 3 groups (n = 8 per group) and orally administered NCE once daily for six weeks at 0 (vehicle), 3.075 (NCE-1X) and 15.375 g kg-1 day-1 (NCE-5X). The gut microbiota from the mice was analyzed using 16S rRNA gene sequencing. RESULTS: NCE-5X did not appear to obviously cluster with the vehicle group, although NCE-5X groups showed an increased Firmicutes/Bacteroidetes ratio compared with the vehicle group. In addition, anti-fatigue activity and exercise performance were evaluated by investigating the exhaustive swimming time, forelimb grip strength and serum levels of lactate, ammonia, glucose, blood urea nitrogen (BUN), creatine kinase (CK) and lactate dehydrogenase (LDH) after swimming. The NCE-1X and NCE-5X groups showed a significantly longer exhaustive swimming time and higher relative forelimb grip strength than the vehicle group. Tissue glycogen content, an important energy source for exercise, increased significantly with NCE supplementation. CONCLUSION: Taken together, our results indicate that NCE supplementation alters the gut microbiota composition and aids in overcoming physical fatigue. Curcumin may be acting on the gut microbiome to modulate the gut system towards improving exercise performance.

The Effects of Ergosta-7,9(11),22-trien-3ß-ol from Antrodia camphorata on the Biochemical Profile and Exercise Performance of Mice.

Antrodia camphorata (AC) is a rare and unique mushroom that is difficult to cultivate. Previous studies have demonstrated the bioactivity of the compound Ergosta-7,9(11),22-trien-3ß-ol (EK100) from AC in submerged culture. The purpose of this study is to evaluate the potential beneficial effects of EK100 on fatigue and ergogenic functions following physiological challenge. Male ICR (Institute of Cancer Research) mice were randomly divided into three groups (n = 8 per group) and orally administered EK100 for six weeks at 0 (Vehicle), 10 (EK100-1X), and 20 (EK100-2X) mg/kg/day. The six-week Ek100 supplementation significantly increased grip strength (P = 0.0051) in trend analysis. Anti-fatigue activity was evaluated using 15-min. acute exercise testing and measuring the levels of serum lactate, ammonia, glucose, blood urea nitrogen (BUN), and creatine kinase (CK) after a 15-min. swimming exercise. Our results indicate that AC supplementation leads to a dose-dependent decrease in serum lactate, ammonia, BUN, and CK activity after exercise and significantly increases serum glucose and glycogen content in liver tissues. Biochemical and histopathological data demonstrated that long term daily administration of EK100 for over six weeks (subacute toxicity) was safe. EK100's anti-fatigue properties appear to be through the preservation of energy storage, increasing blood glucose and liver glycogen content, and decreasing the serum levels of lactate, ammonia, BUN, and CK. EK100 could potentially be used to improve exercise physiological adaptation, promote health, and as a potential ergogenic aid in combination with different nutrient strategies.

Ludwigia octovalvis (Jacq.) raven extract supplementation enhances muscle glycogen content and endurance exercise performance in mice.

Ludwigia octovalvis extract (LOE) is a widely used traditional Chinese herbal medicine. To date, few studies have demonstrated the effect of LOE supplementation on exercise performance, physical fatigue and biochemical profile. The purpose of this study is to evaluate the potential beneficial effects of LOE extract on fatigue and ergogenic functions following physiological challenge. Male ICR mice from 3 groups (n=8 per group) were orally administered LOE for 4 weeks at 0 (vehicle), 61.5 (LOE-1X) or 307.5 (LOE-5X) mg/kg/day. LOE supplementation was able to dose-dependently increase endurance swimming time (P<0.0001) and decrease levels of serum lactate (P=0.0022), ammonia (P<0.0001), creatine kinase (P<0.0001), blood urea nitrogen (P<0.0001) and glucose utilization (P<0.0001) after acute exercise challenge. The glycogen in gastrocnemius muscle also increased with LOE treatment in a dose-dependent manner (P<0.0001). Biochemically, AST, ALT, LDH, CK, BUN, creatinine and UA levels were decreased with LOE treatment. Our study shows that 4-week supplementation with LOE increases muscle glycogen content storage to enhance exercise performance and anti-fatigue effects.

The effects of wild bitter gourd fruit extracts on ICAM-1 expression in pulmonary epithelial cells of C57BL/6J mice and microRNA-221/222 knockout mice: Involvement of the miR-221/-222/PI3K/AKT/NF-κB pathway.

BACKGROUND: The extracts from wild bitter gourd fruit (WBGE) were reported to possess numerous pharmacological activities. However, the anti-inflammatory effects of WBGE on human lung epithelial cells and the underlying mechanisms have not been determined. PURPOSE: To evaluate the molecular basis of the effects of WBGE on intercellular adhesion molecule-1 (ICAM-1) expression in alveolar epithelial (A549) cells, C57BL/6 wild-type (WT) mice and microRNA (miR)-221/-222 knockout (KO) mice with or without tumor necrosis factor (TNF-α; 3 ng/ml) treatment. STUDY DESIGN/METHODS: WT mice and miR-221/-222 KO mice were fed a control diet and divided into four groups (C: control mice; T: treated with TNF-α alone; WBGE/T: pretreated with WBGE and then stimulated with TNF-α; WBGE: treated with WBGE alone). The effects of WBGE on ICAM-1 expression and the related signals in A549 cells and mice with or without TNF-α treatment were examined by Western blot and immunofluorescent staining. RESULTS: WBGE significantly decreased the TNF-α-induced ICAM-1 expression in A549 cells through the inhibition of phosphoinositide 3-kinase (PI3K)/ protein kinase B (AKT)/ nuclear factor- kappa B (NF-κB)/ inhibitor of NF-κB (IκB) phosphorylation and decreased leukocyte adhesion. In addition, WBGE reduced endogenous ICAM-1 expression and upregulated miR-221/-222 expression. The overexpression of miR-222 decreased PI3K/AKT/NF-κB/IκB and ICAM-1 expression, which resulted in reducing monocyte adhesion. Moreover, WBGE reduced ICAM-1 expression in lung tissues of WT mice with or without TNF-α treatment and upregulated miR-221/222. WBGE did not affect the miR-221/-222 level and had little effect on ICAM-1 expression in miR-221/-222 KO mice. CONCLUSIONS: These results suggest that WBGE reduced ICAM-1 expression both under in vitro and in vivo conditions. The protective effects were mediated partly through the miR-221/-222/PI3K/AKT/NF-κB pathway.

Supplementation with Hualian No. 4 wild bitter gourd (Momordica charantia Linn. var. abbreviata ser.) extract increases anti-fatigue activities and enhances exercise performance in mice.

Hualian No. 4 wild bitter gourd (WBG) is a specific vegetable cultivated by the Hualien District Agricultural Research and Extension Station in Taiwan. WBG is commonly consumed as a vegetable and used as a popular folk medicine. However, few studies have demonstrated the effects of WBG supplementation on exercise performance, physical fatigue and the biochemical profile. The purpose of this study was to evaluate the potential beneficial effects of WBG extract on fatigue and ergogenic functions following physiological challenge. Three groups of male ICR mice (n=8 per group) were orally administered 0, 1 or 2.5 g/kg/day of WBG for 4 weeks. They were respectively designated the vehicle, WBG-1X and WBG-2.5X groups. WBG significantly decreased body weight (BW) and epididymal fat pad (EFP) weight. Concerning physical performance, WBG supplementation dose-dependently increased grip strength and endurance swimming time. Concerning anti-fatigue activity, WBG decreased levels of serum lactate, ammonia, creatine kinase and blood urea nitrogen, and economized glucose metabolism after acute exercise challenge. Glycogen in the liver and gastrocnemius muscle dose-dependently increased with WBG treatment. Concerning the biochemical profile, WBG treatment significantly decreased alanine aminotransferase (ALT), blood urea nitrogen (BUN) and urea acid (UA), and increased total protein (TP). Therefore, 4-week supplementation with WBG may decrease white adipose weight, enhance energy economy, increase glycogen storage to enhance exercise performance and reduce fatigue.

The protective effect of eupafolin against TNF-α-induced lung inflammation via the reduction of intercellular cell adhesion molecule-1 expression.

ETHNOPHARMACOLOGICAL RELEVANCE: Eupafolin, a major bioactive compound found in Phyla nodiflora, has the anti-inflammatory property. Upregulation of cell adhesion molecules in the lung airway epithelium is associated with the epithelium-leukocyte interaction and plays a critical role in the pathogenesis of lung airway inflammatory disorders. To investigate the effects of eupafolin on tumor necrosis factor-α (TNF-α)-induced intercellular cell adhesion molecule-1 (ICAM-1) expression in A549 human lung airway epithelial cells and the underlying mechanisms. MATERIALS AND METHODS: The effect of eupafolin on ICAM-1 expression in A549 cells were examined by Western blotting and immunofluorescent staining. The mice were injected intraperitoneally with or without eupafolin and then were left untreated or were injected intratracheally with TNF-α. To detect the effect of eupafolin on ICAM-1 expression, the lung tissues were also examined by Western blotting and immunohistochemical staining. RESULTS: Eupafolin pretreatment reduced the TNF-α-induced ICAM-1 expression and also the ERK1/2, JNK, p38, and AKT/PI3K phosphorylation. However, the increase in ICAM-1 expression with TNF-α treatment was unaffected by p38 and PI3K inhibitors. Eupafolin decreased the TNF-α-induced NF-κB p65 activation and its nuclear translocation. Furthermore, eupafolin reduced ICAM-1 expression in the lung tissues of TNF-α-treated mice. CONCLUSIONS: Eupafolin exerts its anti-inflammatory activity by suppressing the TNF-α-induced ICAM-1 expression and subsequent monocyte adhesion via AKT/ERK1/2/JNK phosphorylation and nuclear translocation of NF-κB p65. These results suggest that eupafolin may represent a novel therapeutic agent targeting epithelial activation in lung inflammation.

Magnolol reduced TNF-α-induced vascular cell adhesion molecule-1 expression in endothelial cells via JNK/p38 and NF-κB signaling pathways.

Expression of cell adhesion molecules by the endothelium and the attachment of leukocytes to these cells play major roles in inflammation and cardiovascular disorders. Magnolol, a major active component of Magnolia officinalis, has antioxidative and anti-inflammatory properties. In the present study, the effects of magnolol on the expression of vascular cell adhesion molecule-1 (VCAM-1) in human aortic endothelial cells (HAECs) and the related mechanisms were investigated. TNF-α induced VCAM-1 protein expression and mRNA stability were significantly decreased in HAECs pre-treated with magnolol. Magnolol significantly reduced the phosphorylation of ERK, JNK, and p38 in TNF-α-treated HAECs. The decrease in VCAM-1 expression in response to TNF-α treatment was affected by JNK and p38 inhibitors, not by an ERK inhibitor. Magnolol also attenuates NF-κB activation and the translocation of HuR (an RNA binding protein) in TNF-α-stimulated HAECs. The VCAM-1 expression was weaker in the aortas of TNF-α-treated apo-E deficient mice with magnolol treatment. These data demonstrate that magnolol inhibits TNF-α-induced JNK/p38 phosphorylation, HuR translocation, NF-κB activation, and thereby suppresses VCAM-1 expression resulting in reduced leukocyte adhesion. Taken together, these results suggest that magnolol has an anti-inflammatory property and may play an important role in the prevention of atherosclerosis and inflammatory responses.

Ganoderma lucidum Polysaccharides Reduce Lipopolysaccharide-Induced Interleukin-1 ß Expression in Cultured Smooth Muscle Cells and in Thoracic Aortas in Mice.

The expression of inflammatory cytokines on vascular walls is a critical event in vascular diseases and inflammation. The aim of the present study was to examine the effects of an extract of Ganoderma lucidum (Reishi) polysaccharides (EORPs), which is effective against immunological disorders, on interleukin- (IL-) 1 ß expression by human aortic smooth muscle cells (HASMCs) and the underlying mechanism. The lipopolysaccharide- (LPS-) induced IL-1 ß expression was significantly reduced when HASMCs were pretreated with EORP by Western blot and immunofluorescent staining. Pretreatment with 10 µ g/mL EORP decreased LPS-induced ERK, p38, JNK, and Akt phosphorylation. But the increase in IL-1 ß expression with LPS treatment was only inhibited by pretreatment with the ERK1/2 inhibitor, while the JNK and p38 inhibitors had no effect. In addition, EORP reduced the phosphorylation and nuclear translocation of nuclear factor- (NF-) κ B p65 in LPS-treated HASMCs. Furthermore, in vivo, IL-1 ß expression was strongly expressed in thoracic aortas in LPS-treated mice. Oral administration of EORP decreased IL-1 ß expression. The level of IL-1 ß expression in LPS-treated or in LPS/EORP-treated group was very low and was similar to that of the saline-treated group in toll-like receptor 4-deficient (TLR4(-/-)) mice. These findings suggest that EORP has the anti-inflammatory property and could prove useful in the prevention of vascular diseases and inflammatory responses.

Comparison of chemical compositions and osteoprotective effects of different sections of velvet antler.

ETHNOPHARMACOLOGICAL RELEVANCE: Velvet antlers (VA) have been claimed for centuries to have numerous medical benefits including strengthen bones. To investigate and compare the anti-osteoporotic activities from different sections of VA. MATERIALS AND METHODS: Fresh VA prepared from farmed sika deers (Cervus nippon) was divided into upper (VAU), middle (VAM), and basal (VAB) sections. The chemical constituents and anti-osteoporotic effect of different sections from VA were evaluated using ovariectomized rats. RESULTS: Levels of water-soluble extracts, diluted alcoholic extract, amino acids, testosterone, insulin-like growth factor (IGF)-1 and testosterone plus estradiol significantly differed among the different sections. Levels of these constituents were significantly higher in the upper section than in the basal section. Moreover, levels of testosterone and IGF-1 of the VAM were also significantly higher than those of the VAB. Calcium level increased downward from the tip with statistical significance. The strength of vertebrae increased in all VA-treated groups compared to the control, but only treatment with VAU and VAM increased the strength of the femur and the microarchitecure of the trabecular bone. Alkaline phosphatase levels of VAU- and VAM-treated groups significantly decreased, but osteocalcin did not significantly change. Moreover, VAU and VAM dose-dependently increased proliferation and mineralization of MC3T3-E1 cells. CONCLUSION: Our study provides strong evidence for the regional differences in the effectiveness of velvet antler in treating osteoporosis. However, further studies are needed to elucidate the bioactive chemical constituents associated with the anti-osteoporotic effects of velvet antler.

Effects of velvet antler with blood on bone in ovariectomized rats.

In traditional Chinese medicine (TCM), both velvet antlers (VA) and VA blood can tonify qi, essence, and marrow, nourish the blood, and invigorate bones and tendons. In TCM, the combination of VA and VA blood is believed to have superior pharmacological effects. Scientific evidence supporting the traditional therapeutic preference for redder antler is needed. The effectiveness of the combination therapy of VA middle sections (VAMs) and VA blood (VAM-B) was first examined in promoting proliferation of mouse osteoblastic cells (MC3T3-E1). The anti-osteoporotic activity of VAM-B (ratio of VAM:VA blood = 1:0.2) was evaluated with ovariectomized (OVX) rats at a dose of 0.2 g/kg. In VAM-B-treated OVX rats, the body weight decreased 10.7%, and the strength of vertebrae and the femur respectively increased 18.1% and 15.4%, compared to the control. VAM-B treatment also recovered the estrogen-related loss of the right tibial trabecular bone microarchitecture. Alkaline phosphatase (ALP) significantly decreased, but estradiol did not significantly change in serum of VAM-B-treated OVX rats. We also provide an effective strategy to enhance the anti-osteoporotic activity of VAM. In conclusion, our results provide scientific evidence supporting the traditional therapeutic preference of redder antler and indicate that VAM-B is a potential therapeutic agent for managing osteoporosis.