FGF4 improves hepatocytes ferroptosis in autoimmune hepatitis mice via activation of CISD3.

Autoimmune hepatitis (AIH) is increasingly affecting human health but pharmacotherapies remain to be identified. Growing evidence reveals that ferroptosis, a newly recognized form of programmed cell death, is critical for AIH. However, the exact mechanisms of the ferroptotic cascade remain elusive. Data in this study showed that ferroptosis aggravation was associated with protectively-elevated fibroblast growth factor 4 (FGF4) expression in Concanavalin A (ConA)-induced AIH liver injury, with these effects being effectively reversed by Ferrostatin-1 (Fer-1). Moreover, hepatic Fgf4 depletion was more susceptible to lipid peroxidation and iron accumulation, as well as hepatic lesion and inflammation caused by ConA administration. Conversely, treatment with non-mitogenic recombinant FGF4 (rFGF4) mitigated liver damage and hepatocellular ferroptosis while being accompanied by the upregulation of CDGSH iron-sulfur domain-containing protein 3 (CISD3) in vivo and in vitro. Furthermore, CISD3 overexpression exhibited stronger resistance to ferroptosis while CISD3 knockdown reduced ferroptotic biomarkers cystine/glutamate transporter (xCT) and glutathione peroxidase 4（GPX4) in rFGF4-treated Erastin-induced AML12 cells. In addition, rFGF4 significantly enhanced the levels of heme oxygenase 1 (HO-1) and nuclear factor erythroid 2-related factor 2 (Nrf2) in ConA-induced AIH mice. Overall, this study showed that FGF4 can act as a phylactic role in AIH progression, with rFGF4 treatment inhibiting ferroptosis of hepatocytes by increasing CISD3 levels and activating Nrf2/HO-1 signaling.

R- Is Superior to S-Form of α-Lipoic Acid in Anti-Inflammatory and Antioxidant Effects in Laying Hens.

The development of single enantiomers with high efficiency and low toxic activity has become a hot spot for the development and application of drugs and active additives. The aim of the present study was to investigate the effectiveness of the application of α-lipoic acid with a different optical rotation to alleviate the inflammation response and oxidative stress induced by oxidized fish oil in laying hens. Sixty-four 124-week-old Peking Red laying hens were randomly allocated to four groups with eight replicates of two birds each. The normal group was fed basal diets supplemented with 1% fresh fish oil (FO), and the oxidative stress model group was constructed with diets supplemented with 1% oxidized fish oil (OFO). The two treatment groups were the S-form of the α-lipoic acid model with 1% oxidized fish oil (OFO + S-LA) and the R-form of the α-lipoic acid model with 1% oxidized fish oil (OFO + R-LA) added at 100 mg/kg, respectively. Herein, these results were evaluated by the breeding performance, immunoglobulin, immune response, estrogen secretion, antioxidant factors of the serum and oviduct, and pathological observation of the uterus part of the oviduct. From the results, diets supplemented with oxidized fish oil can be relatively successful in constructing a model of inflammation and oxidative stress. The OFO group significantly increased the levels of the serum inflammatory factor (TNF-α, IL-1ß, IL-6, and IFN-γ) and the oxidative factor MDA and decreased the activity of the antioxidant enzyme (T-AOC, T-SOD, GSH-Px, GSH, and CAT) in the oviduct. The addition of both S-LA and R-LA significantly reduced the levels of serum inflammatory factors (TNF-α, IL-1ß, IL-6, and IFN-γ), increased the activity of antioxidant indexes (T-AOC, T-SOD, GSH-Px, GSH, and CAT), and decreased the MDA contents in the serum and oviduct. Meanwhile, the supplementation of S-LA and R-LA also mitigated the negative effects of the OFO on the immunoglobulins (IgA and IgM) and serum hormone levels (P and E2). In addition, it was worth noting that the R-LA was significantly more effective than the S-LA in some inflammatory (IL-1ß) and antioxidant indices (T-SOD, GSH, and CAT). Above all, both S-LA and R-LA can alleviate the inflammation and oxidative damage caused by oxidative stress in aged laying hens, and R-LA is more effective than S-LA. Thus, these findings will provide basic data for the potential development of α-lipoic acid as a chiral dietary additive for laying hens.

Lipoamide Alleviates Oxidized Fish Oil-Induced Host Inflammatory Response and Oxidative Damage in the Oviduct of Laying Hens.

Fish oil (FO) is an important source of lipid in functional food and aquafeeds. However, the harmful effects of oxidized fish oil (OFO) on host metabolism and reproductive health are not yet clear. In addition, lipoamide (LAM) has been widely studied as an agent for alleviating various diseases associated with oxidative disruption. Therefore, in the current study, to investigate the effects of LAM in alleviating OFO-induced decline in reproductive performance and oxidative damage to the oviduct in laying hens. We constructed a 1% fresh FO model, a 1% OFO model, and a LAM model with 1% OFO (OFO + LAM) added at 100 mg/kg to explore the antioxidant effect of LAM. Herein, these results were evaluated by breeding performance, immune responses, estrogen, and antioxidant indices of serum samples, as well as the number of follicles and antioxidant parameters of oviducts. From the results, compared with the FO group, OFO significantly decreased the egg-laying rate, increased the contents of total protein (TP) and inflammatory factors [tumor necrosis factor α (TNF-α), interleukin (IL)-6, IL-8, and interferon γ (INF-γ)], and reduced the concentrations of anti-oxidation [total antioxidant (T-AOC), total superoxide dismutase (T-SOD), glutathione peroxidase (GSH-Px), glutathione (GSH), glutathione reductase (GR), catalase (CAT), and hydroxyl radical scavenging activity (HRSA)] in serum samples, as well as reduced the levels of anti-oxidation indexes in oviduct tissues (p < 0.05). Of note, the supplementation of LAM could significantly increase the laying performance, improve the levels of serum immunoglobulins (IgA, IgG, and IgM), serum estrogen [progesterone (P) and estradiol (E2)], and serum antioxidant parameters (T-AOC, T-SOD, GSH-Px, GSH, GR, CAT, and HRSA) and decrease the concentrations of serum inflammatory cytokines (TNF-α, IL-6, IL-8, and INF-γ) in laying hens following OFO administration (p < 0.05). In addition, LAM could dramatically increase the contents of antioxidant factors (p < 0.05) in oviducts and enhance the secretion capacity of the uterine part. Taken together, OFO caused host metabolic dysfunction, oxidative damage, uterine morphological abnormalities, and alterations of ovarian function. These results suggested that LAM administration could alleviate host metabolic dysfunctions and inflammatory damage, and then ameliorate oxidative damage in the oviduct induced by OFO, ultimately improving reproductive function.

Zearalenone toxicosis on reproduction as estrogen receptor selective modulator and alleviation of zearalenone biodegradative agent in pregnant sows.

BACKGROUND: Zearalenone (ZEA) is a resorcylic acid lactone derivative derived from various Fusarium species that are widely found in food and feeds. The molecular structure of ZEA resembles that of the mammalian hormone 17ß-oestradiol, thus zearalenone and its metabolites are known to compete with endogenous hormones for estrogen receptors binding sites and to activate transcription of oestrogen-responsive genes. However, the effect of long-term low-dose ZEA exposure on the reproductive response to Bacillus subtilis ANSB01G culture for first-parity gilts has not yet been investigated. This study was conducted to investigate the toxic effects of ZEA as an estrogen receptor selective modulator and the alleviating effects of Bacillus subtilis ANSB01G cultures as ZEA biodegraders in pregnant sows during their first parity. RESULTS: A total of 80 first-parity gilts (Yorkshire × Landrace) were randomly assigned to four dietary treatments during gestation: CO (positive control); MO (negative control, 246 µg ZEA/kg diet); COA (CO + B. subtilis ANSB01G culture with 2 × 109 CFU/kg diet); MOA (MO + B. subtilis ANSB01G culture with 2 × 109 CFU/kg diet). There were 20 replications per treatment with one gilt per replicate. Feeding low-dose ZEA naturally contaminated diets disordered most of reproductive hormones secretion and affected estrogen receptor-α and estrogen receptor-ß concentrations in serum and specific organs and led to moderate histopathological changes of gilts, but did not cause significant detrimental effects on reproductive performance. The addition of Bacillus subtilis ANSB01G culture to the diet can effectively relieve the competence of ZEA to estrogen receptor and the disturbance of reproductive hormones secretion, and then ameliorate toxicosis of ZEA in gilts. CONCLUSIONS: Collectively, our study investigated the effects of feeding low-dose ZEA on reproduction in pregnant sows during their first parity. Feeding low-dose ZEA could modulate estrogen receptor-α and -ß concentrations in specific organs, cause disturbance of reproductive hormones and vulva swelling, and damage organ histopathology and up-regulate apoptosis in sow models. Diet with Bacillus subtilis ANSB01G alleviated negative effects of the ZEA on gilts to some extent.

Effects of Different Ammonia Concentrations on Pulmonary Microbial Flora, Lung Tissue Mucosal Morphology, Inflammatory Cytokines, and Neurotransmitters of Broilers.

Atmospheric ammonia is one of the main environmental stressors affecting the performance of broilers. Previous studies demonstrated that high levels of ammonia altered pulmonary microbiota and induced inflammation. Research into the lung-brain axis has been increasing in recent years. However, the molecular mechanisms in pulmonary microbiota altered by ambient ammonia exposure on broilers and the relationship between microflora, inflammation, and neurotransmitters are still unknown. In this study, a total of 264 Arbor Acres commercial meal broilers (21 days old) were divided into 4 treatment groups (0, 15, 25, and 35 ppm group) with 6 replicates of 11 chickens for 21 days. At 7 and 21 D during the trial period, the lung tissue microflora was evaluated by 16S rDNA sequencing, and the content of cytokines (IL-1ß, IL-6, and IL-10) and norepinephrine (NE), 5-hydroxytryptamine (5-HT) in lung tissue were measured. Correlation analysis was established among lung tissue microflora diversity, inflammatory cytokines, and neurotransmitters. Results showed that the broilers were not influenced after exposure to 15 ppm ammonia, while underexposure of 25 and 35 ppm ammonia resulted in significant effects on pulmonary microflora, inflammatory cytokines, and neurotransmitters. After exposure to ammonia for 7 and 21 days, both increased the proportion of Proteobacteria phylum and the contents of IL-1ß and decreased the content of 5-HT. After exposure to ammonia for 7 days, the increase in Proteobacteria in lung tissue was accompanied by a decrease in 5-HT and an increase in IL-1ß. In conclusion, the microflora disturbance caused by the increase in Proteobacteria in lung tissue may be the main cause of the changes in inflammatory cytokines (IL-1ß) and neurotransmitters (5-HT), and the damage caused by ammonia to broiler lungs may be mediated by the lung-brain axis.

Effects of Ammonia on Gut Microbiota and Growth Performance of Broiler Chickens.

In order to investigate the influence of ammonia on broiler intestinal microflora and growth performance of broiler chickens, 288 21-day-old male Arbor Acres broilers with a similar weight were randomly divided into four groups with different NH3 levels: 0 ppm, 15 ppm, 25 ppm, and 35 ppm. The growth performance of each group was recorded and analyzed. Additionally, 16s rRNA sequencing was performed on the cecal contents of the 0 ppm group and the 35 ppm group broilers. The results showed the following: a decrease in growth performance in broilers was observed after 35 ppm ammonia exposure for 7 days and 25 ppm ammonia exposure for 14 days. At phylum level, the relative abundance of Proteobacteria phylum was increased after 35 ppm ammonia exposure. At genus level, ammonia increased the relative abundance of Escherichia-Shigella and decreased the relative abundance of Butyricicoccus, Parasutterella, Lachnospiraceae_UCG-010, Ruminococcaceae_UCG-013 and Ruminococcaceae_UCG-004. Negative correlation between Escherichia-Shigella and growth performance, and positive correlation between bacteria genera (including Butyricicoccus, Parasutterella, Lachnospiraceae_UCG-010, Ruminococcaceae_UCG-013 and Ruminococcaceae_UCG-004) and growth performance was observed. In conclusion, ammonia exposure caused changes in the structure of cecal microflora, and several species were either positively or negatively correlated with growth performance. These findings will help enhance our understanding of the possible mechanism by which ammonia affect the growth of broilers.

The alterations of tracheal microbiota and inflammation caused by different levels of ammonia exposure in broiler chickens.

Ammonia (NH3) is a known harmful gas and exists in haze, forming secondary organic aerosols. Exposure to ambient ammonia correlates with the respiratory tract infection, and microbiota in the upper respiratory tract is an emerging crucial player in the homeostatic regulation of respiratory tract infection, and microbiota perturbation is usually accompanied by the inflammatory reactions; however, the effects of different levels of ammonia exposure on tracheal microbiota and inflammation are unclear. A total of 288 22-day-old male Arbor Acres broilers were chosen and divided into 4 groups with 6 replicates of 12 chickens, and respectively exposed to ammonia at 0, 15, 25, and 35 ppm for 21-d trial period. Cytokines (interleukin (IL)-1ß, IL-6, and IL-10) in the trachea were measured at the 21 d of exposure to NH3. Tracheal microbiota at the 21 d was analyzed by the 16S rRNA gene analysis. The results showed that an increase in ammonia levels, even in 15 ppm, significantly decreased the alpha diversity and changed the bacterial community structure. Six genera (Faecalibacterium, Ruminococcus]_torques_group, unclassified_f__Lachnospiraceae, Ruminococcaceae_UCG-014, Streptococcus, Blautia) significantly increased, whereas Lactobacillus significantly decreased under different levels of ammonia exposure. We also observed positive associations of Faecalibacterium, Blautia, g__Ruminococcaceae_UCG-014, unclassified_f__Lachnospiraceae and Ruminococcus]_torques_group abundances with tracheal IL-1ß concentration. Moreover, an increase in ammonia levels, even in 15 ppm, caused respiratory tract inflammatory injury. The results indicated that 15 ppm ammonia exposure changed the composition of tracheal microbiota that caused the tracheal injury possibly through increasing the IL-1ß, which might make the broiler more sensitive to the changes of environment and pathogenic micro-organisms in the poultry house, and may be also a critical value that needs high alertness. Herein, the present experiment also suggested that the standard limit of ammonia concentration in adult poultry house is 15 ppm. This research provides an insight into the relationship between the upper respiratory tract microbiota and inflammation under ammonia exposure.

High mechanical strength gelatin composite hydrogels reinforced by cellulose nanofibrils with unique beads-on-a-string morphology.

This work prepared high mechanical strength gelatin composite hydrogels reinforced by cellulose nanofibrils with unique beads-on-a-string morphology. In detail, cellulose nanofibrils (H-Cel) with unique beads-on-a-string morphology were obtained by acid hydrolysis followed by intensive sonication. The D-H-Cel nanofibrils were prepared through oxidizing part of the non-esterified hydroxyl groups on the H-Cel into aldehyde groups. D-H-Cel were then mixed with gelatin and D-H-Cel/Gel composite hydrogels were produced. During the mixing, a giant network structure was constructed through the Schiff-base reaction between the aldehyde groups on the surface of D-H-Cel nanofibrils and the primary amino groups on gelatin macromolecular chains. Since the cellulose nanofibrils were covalently bonded to gelatin, the stress could be efficiently transferred between the reinforcing agent and matrix, resulting in a composite hydrogel with drastically increased mechanical strength. The compressive strength of D-40H-Cel/Gel hydrogel reached 3.398 MPa. SEM images showed a highly porous three-dimensional structure in the hydrogel with regulated pore size. The crosslinking indices were measured with ninhydrin assay. The composite hydrogels could maintain their shape well in buffer solution. The present work shows that natural polymer-based composite hydrogels with extremely high mechanical strength could be obtained by reinforcing with surface modified cellulose nanofibrils with unique beads-on-a-string morphology.

Increased expression of miR-338-3p impairs Treg-mediated immunosuppression in pemphigus vulgaris by targeting RUNX1.

Pemphigus vulgaris (PV) is a regulatory T cell (Treg)-associated autoimmune disease. Treg cells maintain immunosuppression by expressing the signature transcription factor FOXP3. MicroRNAs (miRNAs) have frequently emerged as regulators in Treg-mediated immunosuppression. We previously found that miR-338-3p was overexpressed in the peripheral blood mononuclear cells of patients with PV. Herein, we explored the role of miR-338-3p in Treg-mediated immunosuppression by quantitative real-time polymerase chain reaction, analysis of public microarray data, miRNA transfection, Western blotting, flow cytometry, and luciferase reporter assays. Increased expression of miR-338-3p was detected in CD4+ T cells of active PV patients compared with those in healthy controls. Moreover, the miR-338-3p level was positively related to disease severity. Bioinformatics prediction revealed that Runt-related transcription factor 1 (RUNX1), a gene activating FOXP3 expression, was a putative target of miR-338-3p. There was a reduction of FOXP3 and RUNX1 expression in the CD4+ T cells of patients with PV, along with significant correlations with the level of miR-338-3p. MiRNA transfection, mRNA and protein analysis, and luciferase reporter assays verified that miR-338-3p attenuated FOXP3 expression by targeting RUNX1. This study suggests that excessive expression of miR-338-3p attenuates the expression of FOXP3 by targeting RUNX1, contributing to Treg dysfunction in PV.

SELEX tool: a novel and convenient gel-based diffusion method for monitoring of aptamer-target binding.

BACKGROUND: Aptamers, single-stranded DNAs or RNAs, can be selected from a library containing random sequences using a method called Systematic Evolution of Ligands by EXponential Enrichment (SELEX). In SELEX, monitoring the enriching statuses of aptamer candidates during the process is a key step until today. Conformational change of an aptamer caused by target-binding in gel can be used to indicate its statuses of binding. RESULTS: In this study, an easy-to-implement gel-based diffusion method (GBDM) was developed to monitor the interaction between enriched aptamer candidates and their targets. In order to prove the concept, characterization of aptamers targeting their targets including protein (thrombin) and non-protein molecules (acetamiprid, ATP, atrazine, profenofos and roxithromycin), respectively, were performed using mini gels. Our method has advantages over the common methods including easy performed with labor- and time- saving in experimental operation. The concept has been proven by monitoring enrichment of dynamic aptamer candidate libraries targeting a small molecule 2,2-bis(4-chlorophenyl) acetic acid (DDA) during SELEX process. A mini gel cassette was designed and fabricated by our laboratory to make mini agarose gels for diffusion with different directions. CONCLUSIONS: These results indicate that GBDM, in particular, chasing diffusion is suitable for monitoring the interaction between enriched aptamer candidates and their targets. These pioneering efforts are helpful for novel aptamer selection by breaking through the technical bottleneck of aptamer development and helpful for development of novel aptasensors.

Yes-Associated Protein Promotes the Development of Condyloma Acuminatum through EGFR Pathway Activation.

OBJECTIVE: Investigate the role of Yes-associated protein (YAP1) in the development of condyloma acuminatum (CA). METHODS: We enrolled 30 male patients with CA and 20 healthy individuals as a control group, to compare the YAP1 expression in their tissue samples. Following this, we overexpressed and downregulated YAP1 expression in HaCaT cells to examine the migratory, proliferative, and apoptotic potential of HaCaT cells expressing different levels of YAP1. RESULTS: In the CA patient tissue samples, an increase in YAP1 expression can be observed. In vitro,the overexpression of YAP1 was shown to promote the growth and migration of HaCaT cells and to activate epidermal growth factor receptor (EGFR) pathway-associated proteins, while the downregulation of YAP1 inhibited cell growth and migration of these cells. CONCLUSIONS: YAP1 promotes the growth of keratinocytes in CA through the activation of the EGFR pathway, and it may mediate the development of human papilloma virus-associated diseases.

Modified nano microfibrillated cellulose/carboxymethyl chitosan composite hydrogel with giant network structure and quick gelation formability.

Carboxymethyl chitosan (CMCS) has good biocompatibility, biodegradability and water solubility. This work investigated a nanocellulose fibrils reinforced CMCS composite hydrogel with giant network structure and quick gelation formability. The microfibrillated cellulose (MFC) with diameter in nano scale and length in sub-micron scale was obtained through high-pressure homogenization. The hydroxyl groups on the MFC nanofibrils surface were partly oxidized into aldehyde groups. The dialdehyde MFC (DAMFC), remaining the fibril morphology and fine three-dimensional network structure, was mixed with CMCS. Due to the Schiff-base reaction between the aldehyde groups on DAMFC surface and amino groups on CMCS chains, a giant network was formed in DAMFC/CMCS. As a result, CMCS composite hydrogel could be formed in short time, although pure CMCS could not form hydrogel. In the composite, DAMFC acted as a reinforcing agent as well as a macroscopical crosslinking agent. The structure and properties of the composite hydrogels were investigated. The compression strength and the work of fracture of the hydrogel was increased dramatically as the DAMFC content increasing from 5 to 15 wt%, an increase of 330% and 338%, respectively. SEM images of the hydrogel showed a fine three-dimensional porous structure, which may provide a promising platform for tissue engineering scaffold.

iTRAQ-based quantitative proteomics analysis of the spleen reveals innate immunity and cell death pathways associated with heat stress in broilers (Gallus gallus).

Heat stress induces immune dysfunction and cell death, but the mechanisms by which this occurs are not fully understood. Therefore, the isobaric tags for relative and absolute quantification (iTRAQ) was used to identify differentially abundant proteins (DAPs) in the spleen between heat-stressed group and control group, and real time qPCR (RT-qPCR), and Parallel Reaction Monitoring (PRM) were performed to validate the differentially abundant proteins of interest. The results showed that nine down regulated DAPs related to innate immunity were enriched in the Toll-like receptor signaling pathway (IRF3 and CD40), NOD-like receptor signaling pathway (TNFAIP3, IL-18, CathL2, IRF3, IAP3 and CYBA), RIG-I-like receptor signaling pathway (TRIM25 and IRF3), and Cytosolic DNA-sensing pathway (IL-18, POLR3F and IRF3). Six down or up regulated DAPs related to cell death were enriched in apoptosis (CTSD, PARP3 and IAP3), ferroptosis (FTH) and necroptosis (FTH, CHMP1B, TNFAIP3, PARP3 and IAP3). In addition, compared with control group, heat stress significantly increased serums IL-1ß, IL-6, TNF-α, and IFN-α, as well as the splenocyte apoptosis rate, whereas significantly decreased serum IFN-ß. Taken together, these findings indicate that heat stress inhibits innate immunity and induces cell death through different pathways. SIGNIFICANCE: Our study identified potential signaling pathways and differentially abundant proteins related to the innate immunity and cell death of broilers under high temperature. These findings will facilitate a better understanding of the mechanisms of broiler response to heat stress and provide possible targets for alleviating heat stress in broiler production.

Quantitative proteomic analysis of deer antler stem cells as a model of mammalian organ regeneration.

The ability to activate and regulate stem cells during wound healing and tissue/organ regeneration is a promising field which could bring innovative approaches to regenerative medicine. The regenerative capacity of invertebrates has been well documented, however in mammals, stem cells that drive organ regeneration are rare. Deer antler is unique in providing a mammalian model of complete organ regeneration based on stem cells. The present study investigated the differentially regulated proteins (DRPs) between different antler stem cell populations (n = 3) using 2D-DIGE. Western blotting was used to validate the proteomics results. Comparative proteomics resulted in protein profiles which were similar for the biological replicates but different between the cells derived from two different stem cell niches involved in antler growth/regeneration and cells derived from facial periosteum. Ninety-two up- and down-regulated proteins were identified by MALDI-TOF MS. The work indicates that the epithelial-mesenchymal transition process may participate in the initiation of antler regeneration including the first stage of scar-less wound healing. Cell mobility is also highly regulated during antler regeneration. Energy and nucleotide metabolism may however be less active in antler regeneration as compared to that in antler generation phase. These results provide new insights into the underlying mechanisms of stem cell-based regeneration of mammalian organs.

Nonlocal effects on the structural transition of gold clusters from planar to three-dimensional geometries.

Conventional density functional theory calculations heavily bias planar structures in gold clusters, failing to predict the structural transition from planar to three-dimensional geometries in experimentally detected gold species. Inspired by recent progress in calculating the defect energies of coinage metals with nonlocal effect-enhanced hybrid functionals, we have studied nonlocal effects in gold clusters. Although the hybrid functional was accurate for bulk gold, it heavily biased the planar structure for gold clusters. By including dispersive interactions into semilocal density functional calculations, we obtained an accurate vacancy formation energy of 0.72 eV for bulk gold along with the correct structural transition for gold clusters. The transition was found to occur at Au12 - for gold anions and at Au8 + for gold cations, agreeing very well with the experimental results. For neutral gold clusters, we found the transition to occur at Au10, indicating the need for experimental verification. The results show the importance of nonlocal effects in the study of gold clusters, calling for further comprehensive theoretical and experimental studies to evaluate nonlocal effects in Au and other precious metals.

Usefulness of miRNA-338-3p in the diagnosis of pemphigus and its correlation with disease severity.

BACKGROUND: Pemphigus is a common life-threatening, autoimmune bullous disease effecting both cutaneous and mucous membranes. Previous diagnosis of pemphigus is based on clinical presentations, histopathology, immunofluorescence and enzyme-linked immunosorbent assay. Furthermore, no laboratory parameters could be used to indicate disease severity. MicroRNAs are endogenous small RNAs, which could be used as diagnostic biomarkers for some autoimmune diseases. Previously, miR-338-3p has been proven significantly up-regulated in pemphigus patients. METHODS: Pemphigus patients (including pemphigus vulgaris and pemphigus foliaceus) with active lesions and with remission, patients diagnosed as bullous pemphigoid and healthy volunteers were recruited, and miR-338-3p expression level was measured using reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR). Active pemphigus patients accepting treatment were followed up for at least 2 weeks to investigate the expression change of miR-338-3p during treatment period. Target genes of miR-338-3p were screened through computer-aided algorithm and verified by RT-qPCR, Western blot and Luciferase activity assay. RESULTS: MiR-338-3p was specifically increased in patients diagnosed as pemphigus with active lesions. The expression level of miR-338-3p gradually decreased after effective treatment. MiR-338-3p expression was independently correlated with disease severity defined by PDAI (Pemphigus Disease Area Index) or ABSIS (Autoimmune Bullous Skin Disorder Intensity Score) criteria. Up-regulation of miR-338-3p could significantly suppress RNF114 expression at mRNA and protein level in vitro. DISCUSSION: MiR-338-3p could be used as a diagnostic biomarker of pemphigus in addition to other traditional methods. Up-regulation of MiR-338-3p was associated with more severe condition in pemphigus. RNF114 is the target gene of miR-338-3p, which probably participates in the regulation of disease activity of pemphigus.

A proteomic approach reveals the differential protein expression in Drosophila melanogaster treated with red ginseng extract (Panax ginseng).

BACKGROUND: Red ginseng is a popularly used traditional medicine with antiaging effects in Asian countries. The present study aimed to explore the changes in protein expression underlying the mechanisms of life span extension and antiaging caused by red ginseng extract (RGE) in Drosophila melanogaster. METHODS: A proteomic approach of two-dimensional polyacrylamide gel electrophoresis (2-DE) was used to identify the differential abundance of possible target proteins of RGE in D. melanogaster. The reliability of the 2-DE results was confirmed via Western blotting to measure the expression levels of selected proteins. Proteins altered at the expression level after RGE treatment (1 mg/mL) were identified by matrix-assisted laser desorption/ionization-time of flight tandem mass spectrometry and by searching against the National Center for Biotechnology nonredundant and Uniprot protein databases. The differentially expressed proteins were analyzed using bioinformatics methods. RESULTS: The average survival life span of D. melanogaster was significantly extended by 12.60% with RGE treatment (1 mg/mL) compared to untreated flies. This followed increased superoxide dismutase level and decreased methane dicarboxylic aldehyde content. Based on the searching strategy, 23 differentially expressed proteins were identified (16 up-regulated and 7 down-regulated) in the RGE-treated D. melanogaster. Transduction pathways were identified using the Kyoto Encyclopedia of Genes and Genomes database, and included the hippo and oxidative phosphorylation pathways that play important roles in life span extension and antiaging process of D. melanogaster. CONCLUSION: Treatment with RGE in D. melanogaster demonstrated that mechanisms of life span extension and antiaging are regulated by multiple factors and complicated signal pathways.

Deer thymosin beta 10 functions as a novel factor for angiogenesis and chondrogenesis during antler growth and regeneration.

BACKGROUND: Deer antlers are the only known mammalian organ with vascularized cartilage that can completely regenerate. Antlers are of real significance as a model of mammalian stem cell-based regeneration with particular relevance to the fields of chondrogenesis, angiogenesis, and regenerative medicine. Recent research found that thymosin beta 10 (TMSB10) is highly expressed in the growth centers of growing antlers. The present study reports here the expression, functions, and molecular interactions of deer TMSB10. METHODS: The TMSB10 expression level in both tissue and cells in the antler growth center was measured. The effects of both exogenous (synthetic protein) and endogenous deer TMSB10 (lentivirus-based overexpression) on antlerogenic periosteal cells (APCs; nonactivated antler stem cells with no basal expression of TMSB10) and human umbilical vein endothelial cells (HUVECs; endothelial cells with no basal expression of TMSB10) were evaluated to determine whether TMSB10 functions on chondrogenesis and angiogenesis. Differences in deer and human TMSB10 in angiogenesis and molecular structure were determined using animal models and molecular dynamics simulation, respectively. The molecular mechanisms underlying deer TMSB10 in promoting angiogenesis were also explored. RESULTS: Deer TMSB10 was identified as a novel proangiogenic factor both in vitro and in vivo. Immunohistochemistry revealed that TMSB10 was widely expressed in the antler growth center in situ, with the highest expression in the reserve mesenchyme, precartilage, and transitional zones. Western blot analysis using deer cell lines further supports this result. Both exogenous and endogenous deer TMSB10 significantly decreased proliferation of APCs (P < 0.05), while increasing the proliferation of HUVECs (P < 0.05). Moreover, deer TMSB10 enhanced chondrogenesis in micromass cultures and nerve growth as assessed using a dorsal root ganglion model (P < 0.05). Deer TMSB10 was proangiogenic using models of chicken chorioallantoic membrane, tube formation, and aortic arch assay. At the molecular level, endogenous deer TMSB10 elevated the expression of vascular endothelial growth factor (VEGF), VEGF-B, VEGF-C, and VEGF-D, and VEGFR2 and VEGFR3 in HUVECs (P < 0.05). CONCLUSIONS: Deer TMSB10, in contrast to its human counterpart, was identified as a novel stimulating factor for angiogenesis, cartilage formation, and nerve growth, which is understandable given that deer antlers (as the arguably fastest mammalian growing tissue) may require this extra boost during a period of rapid growth and regeneration.

Increased expression of microRNA-338-3p contributes to production of Dsg3 antibody in pemphigus vulgaris patients.

Expression of microRNA-338-3p (miR-338-3p) was aberrantly elevated in pemphigus vulgaris (PV), although its role in PV is still unknown. The present study investigated the functional role and possible molecular mechanisms of miR-338-3p in PV. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was performed to detect miR-338-3p expression in peripheral blood mononuclear cells (PBMCs) from patients with PV. Correlation with disease severity and anti-desmoglein 3 antibody (anti-Dsg3) titers were analyzed in patients with PV. The effects of overexpression and knockdown of miR-338-3p expression in PBMCs and effects on Th1 and Th2 cytokines were also examined using ELISA. The luciferase reporter analysis, RT-qPCR and western blot analysis were applied to investigate potential and functional target genes. The data showed that miR-338-3p expression was significantly upregulated in PV and the upregulation of miR-338-3p associated with disease severity and a high anti-Dsg3 antibody titer. Expression of miR-338-3p/mimic in healthy PBMCs significantly downregulated Th1 cytokine (IFN-γ) and upregulated Th2 cytokines (IL-4 and IL-10), whereas knockdown of miR-338-3p expression in PBMCs from patients with PV induced the reverse effects. Overexpression of miR-338-3p suppressed cell viability. Luciferase reporter, RT-qPCR and western blot assays idnicated that TNFR1-associated death domain protein (TRADD) was the direct and functional target of miR-338-3p. Increased expression of miR-338-3p contributed to the production of Dsg3 antibody by inhibiting TRADD expression to induce an imbalance in Th1/Th2 cell functions. Taken together, this study suggests that miR-338-3p may be used as a potential therapeutic target for PV treatment.