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Objective: This study investigates the effects of ZNF862 on the proliferation and apoptosis of human gingival fibroblasts and their related mechanisms. Background: As a major transcription factor family, zinc finger proteins (ZFPs) regulate cell differentiation, growth, and apoptosis through their conserved zinc finger motifs, which allow high flexibility and specificity in gene regulation. In our previous study, ZNF862 mutation was associated with hereditary gingival fibromatosis. Nevertheless, little is known about the biological function of ZNF862. Therefore, this study was aimed to reveal intracellular localization of ZNF862, the influence of ZNF862 on the growth and apoptosis of human gingival fibroblasts (HGFs) and its potential related mechanisms. Methods: Immunohistochemistry, immunofluorescence staining, and western blotting were performed to determine the intracellular localization of ZNF862 in HGFs. HGFs were divided into three groups: ZNF862 overexpression group, ZNF862 interference group, and the empty vector control group. Then, the effects of ZNF862 on cell proliferation, migration, cell cycle, and apoptosis were evaluated. qRT‐PCR and western blotting were performed to further explore the mechanism related to the proliferation and apoptosis of HGFs. Results: ZNF862 was found to be localized in the cytoplasm of HGFs. In vitro experiments revealed that ZNF862 overexpression inhibited HGFs proliferation and migration, induced cell cycle arrest at the G0/G1‐phase and apoptosis. Whereas, ZNF862 knockdown promoted HGFs proliferation and migration, accelerated the transition from the G0/G1 phase into the S and G2/M phase and inhibited cell apoptosis. Mechanistically, the effects of ZNF862 on HGFs proliferation and apoptosis were noted to be dependent on inhibiting the cyclin‐dependent kinase inhibitor 1A (p21)‐retinoblastoma 1 (RB1) signaling pathway and enhancing the B‐cell lymphoma‐extra‐large (Bcl‐xL)‐Caspase 3 signaling pathway. Conclusion: Our results for the first time reveal that ZNF862 is localized in the cytoplasm of HGFs. ZNF862 can inhibit the proliferation of HGFs by inhibiting the p21‐RB1 signaling pathway, and it also promotes the apoptosis of HGFs by enhancing the Bcl‐xL‐Caspase 3 signaling pathway. [ABSTRACT FROM AUTHOR]

Background: JAK inhibitors are well known for the treatment of rheumatoid arthritis (RA), but whether they can be used to treat pulmonary fibrosis, a common extra‐articular disease of RA, remains to be clarified. Methods: A jak2 inhibitor, CEP33779 (CEP), was administered to a rat model of RA‐associated interstitial lung disease to observe the degree of improvement in both joint swelling and pulmonary fibrosis. HFL1 cells were stimulated with TGF‐β1 to observe the expression of p‐JAK2. Then, different concentrations of related gene inhibitors (JAK2, TGFβ‐R1/2, and p‐STAT3) or silencers (STAT3, JAK2) were administered to HFL1 cells, and the expression levels of related proteins were detected to explore the underlying mechanisms of action. Results: CEP not only reduced the degree of joint swelling and inflammation in rats but also improved lung function, inhibited the pro‐inflammatory factors IL‐1β and IL‐6, reduced lung inflammation and collagen deposition, and alleviated lung fibrosis. CEP decreased the expression levels of TGFβ‐R2, p‐SMAD, p‐STAT3, and ECM proteins in rat lung tissues. TGF‐β1 induced HFL1 cells to highly express p‐JAK2, with the most pronounced expression at 48 h. The levels of p‐STAT3, p‐SMAD3, and ECM‐related proteins were significantly reduced after inhibition of either JAK2 or STAT3. Conclusion: JAK2 inhibitors may be an important and novel immunotherapeutic drug that can improve RA symptoms while also delaying or blocking the development of associated pulmonary fibrotic disease. The mechanism may be related to the downregulation of p‐STAT3 protein via inhibition of the JAK2/STAT signaling pathway, which affects the phosphorylation of SMAD3. [ABSTRACT FROM AUTHOR]

Objective To detect the expression changes of interleukin-10 (IL-10) and transforming growth factor-β1 (TGF-β1) during the development of deep vein thrombosis in mice, and to explore the application value of them in thrombus age estimation. Methods The mice in the experimental group were subjected to ligation of inferior vena cava. The mice were sacrificed by excessive anesthesia at 1 d, 3 d, 5 d, 7 d, 10 d, 14 d and 21 d after ligation, respectively. The inferior vena cava segment with thrombosis was extracted below the ligation point. The mice in the control group were not ligated, and the inferior vena cava segment at the same position as the experimental group was extracted. The expression changes of IL-10 and TGF-β1 were detected by immunohistochemistry (IHC), Western blotting and real-time qPCR. Results IHC results revealed that IL-10 was mainly expressed in monocytes in thrombosis and TGF-β1 was mainly expressed in monocytes and fibroblast-like cells in thrombosis. Western blotting and real-time qPCR showed that the relative expression levels of IL-10 and TGF-β1 in each experimental group were higher than those in the control group. The mRNA and protein levels of IL-10 reached the peak at 7 d and 10 d after ligation, respectively. The mRNA expression level at 7 d after ligation was 4.72±0.15 times that of the control group, and the protein expression level at 10 d after ligation was 7.15±0.28 times that of the control group. The mRNA and protein levels of TGF-β1 reached the peak at 10 d and 14 d after ligation, respectively. The mRNA expression level at 10 d after ligation was 2.58±0.14 times that of the control group, and the protein expression level at 14 d after ligation was 4.34±0.19 times that of the control group. Conclusion The expressions of IL-10 and TGF-β1 during the evolution of deep vein thrombosis present time-dependent sequential changes, and the expression levels of IL-10 and TGF-β1 can provide a reference basis for thrombus age estimation. [ABSTRACT FROM AUTHOR]

Objective: To screen and analyze the function of specific CircRNAs involved in pathological bone formation in patients with ankylosing spondylitis (AS). Methods: From September 2019 to October 2020, hip capsule tissues obtained from 3 patients with AS developed hip joint fusion and 3 patients with femoral neck fracture (FNF) were obtained. The circular RNA expressions of hip capsule were analyzed by Arraystar CircRNA chip. qRT‐PCR analysis wan performed to identify the expression patterns of differentially expression CircRNAs. Results: Our findings showed that there were 25 up‐regulated and 39 down‐regulated differential CircRNAs. Among these CircRNAs, we screened 10 highest up‐regulatedCircRNAs and 13 lowest down‐regulated CircRNAs (Fold Change≥2,P<0.05). In further verification analysis, hsa_circ_0067103, hsa_circ_0004496, and hsa_circ_0002649, ACTG1 were significantly upregulated, while hsa_circ_0020273, hsa_circ_0005699, and hsa_circ_0048764 were markly downregulated in AS tissue than FNF controls. Conclusion: The expression of CircRNAs involved of pathological bone formation in AS were significantly different from those of control group. These differentially expressed Circular RNAs may be closely related to the occurrence and development of pathological bone formation in AS. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) are types of endogenous non‐coding small RNAs found in eukaryotes that are 18–25 nucleotides long. miRNAs are considered to be key regulatory factors of the expression of target mRNA. The roles of miRNAs involved in the regulation of anthocyanin accumulation in pigmented potatoes have not been systematically reported. In this study, the differentially expressed miRNAs and their target genes involved in the accumulation of anthocyanin during different developmental stages in purple potato (Solanum tuberosum L.) were identified using small RNA (sRNA) and degradome sequencing. A total of 275 differentially expressed miRNAs were identified in the sRNA libraries. A total of 69,387,200 raw reads were obtained from three degradome libraries. The anthocyanin responsive miRNA–mRNA modules were analyzed, and 37 miRNAs and 23 target genes were obtained. Different miRNAs regulate the key enzymes of anthocyanin synthesis in purple potato. The structural genes included phenylalanine ammonia lyase, chalcone isomerase, flavanone 3‐hydroxylase, and anthocyanidin 3‐O‐glucosyltransferase. The regulatory genes included WD40, MYB, and SPL9. stu‐miR172e‐5p_L‐1R‐1, stu‐miR828a, stu‐miR29b‐4‐5p, stu‐miR8019‐5p_L‐4R‐3, stu‐miR396b‐5p, stu‐miR5303f_L‐7R + 2, stu‐miR7997a_L‐3, stu‐miR7997b_L‐3, stu‐miR7997c_L + 3R‐5_2ss2TA3AG, stu‐miR156f‐5p_L + 1, stu‐miR156a, stu‐miR156a_R‐1, stu‐miR156e, stu‐miR858, stu‐miR5021, stu‐miR828 and their target genes were validated by qRT‐PCR. They play important roles in the coloration and accumulation of purple potatoes. These results provide new insights into the biosynthesis of anthocyanins in pigmented potatoes. [ABSTRACT FROM AUTHOR]

Given the rising domestic demand and increasing global prices of corn and soybean, China is looking for alternatives for these imports to produce animal fodder. Kenaf (Hibiscus cannabinus L.) has great potential as a new forage source, due to abundant proteins, phenols and flavonoids in its leaves. However, few studies have evaluated the mechanism of protein synthesis in kenaf leaves. In the current work, compared with kenaf material "L332," the percentage of crude protein content in leaves of material "Q303" increased by 6.13%; combined with transcriptome and proteome data, the kenaf samples were systematically studied to obtain mRNA-protein correlation. Then, the genes/proteins related to protein synthesis in the kenaf leaves were obtained. Moreover, this work detected mRNA expression of 20 differentially expressed genes (DEGs). Meanwhile, 20 differentially expressed proteins (DEPs) related to protein synthesis were performed parallel reaction monitoring. Fructose-1,6-bisphosphatase (FBP), nitrite reductase (NirA), prolyl tRNA synthase (PARS) and glycine dehydrogenase (GLDC) presented increased mRNA and protein levels within kenaf leaves with high protein content. Based on the obtained findings, FBP , NirA , PARS , and GLDC genes may exert a vital function in the protein synthesis of kenaf leaves. The results provide a new idea for further studying the potential genes affecting the quality trait of protein content in kenaf leaves and provide gene resources and a theoretical foundation for further cultivating high protein kenaf varieties. [ABSTRACT FROM AUTHOR]

Glutathione S -transferases (GSTs) play important roles in metabolism and detoxification of plant cells. However, their functions during development are less well understood. Arabidopsis AtGSTU7 (AT2G29420) encodes a Tau class GST. Here we provide the AtGSTU7 was abundantly expressed in seeds and in roots at an early stage of germination. AtGSTU7 expression was repressed by exogenous ABA and promoted by osmotic stress. A null mutant of AtGSTU7 (atgstu7) accumulated higher contents of reduced GSH and decreased amounts of endogenous H 2 O 2 in seedlings. The atgstu7 plants showed decreased osmotic tolerance during seed germination, which was influenced by GSH and ABI3 gene expression. The results suggested that AtGSTU7 involvement in seed germination is mediated by GSH–ROS homeostasis and ABA signaling. • AtGSTU7 gene abundantly expressed in seeds and during seed germination. • AtGSTU7 gene responded to ABA and osmotic stresses. • atgstu7 accumulated reduced GSH and decreased endogenous H 2 O 2. • The atgstu7 plants decreased osmotic tolerance during seed germination influencing by GSH and ABI3 expression. • AtGSTU7 mediated seed germination by regulating GSH–ROS homeostasis and ABA signaling. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) are small non-coding RNA molecules that play an important role in the regulation of gene expression related to inflammatory responses. Human adipose stem cells are characterized by pluripotent differentiation potential and isolated from adipose tissues. These cells regulate inflammation mainly by interacting with immune cells and affecting the secretion of immune factors; details of this interaction are currently unknown. In the current study, we successfully established an acute inflammation model and a chronic inflammation model involving adipose stem cells. We used high-throughput miRNA microarray analysis to identify miRNAs that were significantly (p < 0.05) differentially expressed during both acute and chronic inflammation. Lipopolysaccharide (LPS) significantly (p < 0.05) reduced the expression of miR-223-3P and miR-2909, while promoting the production of pro-inflammatory cytokines, interleukin (IL) 6, IL-1β, and tumor necrosis factor (TNF)-α via the Toll-like receptor (TLR) 4/TLR2/nuclear factor (NF)-κB/signal transducer and activator of transcription (STAT) 3 signaling pathway in human adipose stem cells. Further, miR-223-3P expression was significantly (p < 0.05) reduced in human adipose stem cells during activation by IL-6 stimulation. The inducible down-regulation of miR-223-3P resulted in the activation of STAT3, which was directly targeted by miR-223-3P. STAT3 directly targeted TLR4 and TLR2, promoting the production of the pro-inflammatory cytokine, IL-6, and formed a positive feedback loop to regulate IL-6 levels. Similarly, TNF-α significantly (p < 0.05) increased the expression of miR-223-3p, with LPS and TLR4/TLR2/NF-κB/STAT3 forming a negative feedback loop to regulate TNF-α levels. In addition, miR-2909, which depends on NF-κB, targeted Krueppel-like factor (KLF) 4 to regulate the levels of pro-inflammatory cytokines, IL-6, IL-1β, and TNF-α. We conclude that miR-223-3p and miR-2909 form a complex regulatory network with pro-inflammatory factors and signaling pathways in adipose stem cells stimulated by LPS. These findings will inform the development of therapies against autoimmune and inflammatory diseases. [ABSTRACT FROM AUTHOR]

Ethnopharmacological relevance Sanmiao Wan (SMW) has been a basic prescription employed for the treatment for gout in the clinic since Yuan dynasty. Achyranthis bidentatae radix (ABR) is designed as a lower-guiding drug in SMW to augment the articular accumulation of active ingredients and improve the anti­inflammatory effect. Aim of the study Present study was undertaken to investigate the dose-response relationship of berberine in SMW between the articular concentration and anti-inflammatory effect in the knee joint under the lower-guiding of ABR. Materials and methods Rats were divided into control group, model group and SMW without or with low, medium and high doses of ABR groups. Rat model of acute gouty arthritis (AGA) was established by intra-articular injection of 0.2 mL monosodium urate crystal (20 mg/mL) inside knee joint cavity on day 2 during drug treatment slots. Knee joint swelling, synovial hyperplasia and inflammatory cell infiltration were investigated for anti-inflammatory study. The concentrations of berberine in rat plasma and tissues were determined by UPLC–MS/MS method. The effect of ABR on the expression levels of P-glycoprotein (P-gp) and MDR1 mRNA in the synovial tissues of knee joints in AGA rats was examined by Western blot and RT-qPCR assay, respectively. Results The distribution of berberine increased by 6.53%, 44.31% and 212.96% in the knee joint and 474.93%, 631.01% and 1063.3% in the ankle for SMW with low, medium and high doses of ABR groups, compared with SMW without ABR group. Similarly, the plasma level of berberine increased by 19.81%, 143.4% and 681.13%. On the contrary, the distribution of berberine evidently decreased 3.23, 10.61 and 46.21-fold in heart and 3.68, 6.74 and 24.78-fold in lung. SMW with different doses of ABR groups exhibited better efficiency than SMW without ABR group on ameliorating knee joint swelling, inhibiting synovial hyperplasia and alleviating inflammatory cell infiltration of AGA rats. The treatment with ABR could down-regulate the MDR1 mRNA and P-gp expressions of synovial tissues of knee joints in AGA rats. Conclusions The enhanced articular distribution of berberine in SMW was attributed to the lower-guiding effect of ABR, which could evidently increase the plasma concentration of berberine, improve the supply of blood of inflamed joint, reduce the distribution of berberine in heart and lung and significantly inhibit the MDR1 mRNA and P-gp expression of synovial tissues of knee joints in AGA rats. The dose-response relationship of berberine between the enhanced articular concentration and improved anti-inflammatory effect in the knee joint under the lower-guiding of ABR was observed for the first time. [ABSTRACT FROM AUTHOR]

Niu Huang Jie Du prescription (NHJD) is a traditional Chinese medicine (TCM) widely used in patients suffering from excessive inner fire toxin (Huo Du Nei Sheng) syndrome, such as sore throat, gingival swelling, and pain, mouth and tongue sores, etc. This formula contains realgar (As 4 S 4) which is one of the 28 toxic medicinal materials promulgated by the Chinese Ministry of Health. Many studies reported its toxicity on the liver and kidney, and the detoxification effect of NHJD. However, its detoxification mechanism is still unclear. To clarify the detoxification mechanism of NHJD to realgar, this study evaluated the detoxification effect of NHJD on realgar exposure in mice, and analyzed differences in mRNA expression profiles in liver tissues and associated functional predictions. ICR mice were administered with NHJD, realgar, and CMC-Na as blank control for 12 weeks, respectively. Liver injury was evaluated by histopathologic examination and liver mRNA gene were sequenced by Illumina. Differentially expressed gene, functionally enrichment and protein association network analysis were conducted. 43 genes were screened out, among which 15 genes in the realgar group were decreased, but the extent of the decline has been restored in the NHJD group. The remaining 28 genes have exactly the opposite trends. Functional module analysis revealed that those detoxification function-related genes were primarily for positive regulation of glutathione metabolism, P450 on the metabolism of exogenous compounds, oxidative stress and immune-related, etc. The results indicated that realgar mainly causes liver damage by changing the common enzymes of drug metabolism, especially the expression of genes related to CYPs, GSTs family, oxidative stress, and complement immunity, while the TCM prescription NHJD has a regulatory effect on the abnormal expression of corresponding genes. Our results will provide some clues for the detoxification mechanism of arsenic-containing TCM prescriptions. [Display omitted] • We focused on the detoxification mechanism of realgar-containing NHJD in mice. • RNA expression, KEGG, GO analysis and protein association network were conducted. • Enzymes of drug metabolism may be involved in the detoxification mechanism of NHJD. [ABSTRACT FROM AUTHOR]

Human leukocyte antigen-G (HLA-G) contributes to acceptance of allograts in solid organ/tissue transplantation. Most studies have determined that soluble HLA-G isoforms are systematically detected in serum/plasma of transplanted patients with significantly fewer episodes of acute and/or chronic rejection of allogeneic tissue/organ. Current models of the interactions of HLA-G and its specific receptors explain it as functioning in a monomeric form. However, in recent years, newd ata has revealed the ability of HLA-G to form disulide-linked dimeric complexes with high preferential binding and functional activities. Limited data are available on the role of soluble HLA-G dimers in clinical pathological conditions. We describe here the presence of soluble HLA-G dimers in kidney transplant patients. Our study showed that a high level of HLA-G dimers in plasma and increased expression of the membrane-bound form of HLA-G on monocytes are associated with prolongation of kidney allograt survival. We also determined that the presence of soluble HLA-G dimers links to the lower levels of proinlammatory cytokines, suggesting a potential role of HLA-G dimers in controlling the accompanying inlammatory state. [ABSTRACT FROM AUTHOR]

Abscisic acid (ABA) is an important stress phytohormone that plays an essential role in mediating the signaling networks associated with plant responses to various abiotic stresses. In the present study, we isolated a gene CrNCED1 encoding the rate-limiting enzyme of ABA synthesis, 9- cis-epoxycarotenoid dioxygenase (NCED), from Citrus reshni. Expression patterns of CrNCED1 varied among different tissues, in which higher levels were measured in the leaves than in the roots. The steady-state mRNA levels of CrNCED1 were progressively elevated by dehydration and ABA, only transiently induced by cold, but not affected by salt. To examine its function in stress tolerance, transgenic tobacco ( Nicotiana nudicaulis) plants constitutively overexpressing CrNCED1 were generated, which contained higher ABA levels than the wild type (WT) under both normal growth conditions and drought stress. The transgenic lines displayed enhanced tolerance to dehydration, drought, salt and oxidative stresses when compared with the WT. Lower levels of reactive oxygen species (HO and O) were detected in the transgenic plants than in the WT under dehydration and salt stress. On the contrary, transcript levels of several genes associated with ROS scavenging, osmoticum adjustment, and water maintenance, and activities of two antioxidant enzymes were higher in the transgenic plants relative to the WT under the dehydration stress. Taken together, CrNCED1 overexpression confers enhanced tolerance to multiple abiotic stresses, which may be, at least in part, ascribed to the positive activation of the stress-responsive genes. [ABSTRACT FROM AUTHOR]

Background: Transforming growth factor-β1 (TGF-β1) is a polypeptide member of the transforming growth factor β superfamily of cytokines and performs many cellular functions. Its overexpression may lead to renal fibrosis. Aim: This study planed to investigate the effects of TGF-β1 on the cell cycle and phenotype of mesangial cells. Methods: Rat mesangial cells were cultured together with different concentrations (0, 1, 2, 5, and 10 ng/mL) of TGF-β1 for specified times from 0 min to 72 h. 0 ng/mL TGF-β1 and 0 min served as controls. Cell cycles were assessed by flow cytometry and α-smooth muscle actin expression (α-SMA) protein expression by western blot analysis. All data were presented as Mean ± SD. Statistical analysis was performed by using one-way analysis of variance and correlation analysis. Results were considered significant at p < 0.05. Results: After 15 min of co-culture with different concentrations of TGF-β1, the percentage of mesangial cells in G0/G1 phase was significantly elevated compared to the control ( p < 0.05). 12 h co-culture induced cell hyperplasia, 24 h co-culture obvious up-regulation of α-SMA ( p < 0.01) and one or two cells' myofibroblast phenotype transition, and 36 h co-culture several cells' phenotype transition. Correlation analysis prompted that the TGF-β1-induced premature aging was time-dependent ( p < 0.01). Conclusion: TGF-β1 may induce mesangial cells' premature senescence and myofibroblast-like phenotype transformation time-dependently, which may contribute to the development of early stage of glomerulosclerosis. [ABSTRACT FROM AUTHOR]

Background: Transforming growth factor-β1 (TGF-β1) is a polypeptide member of the transforming growth factor β superfamily of cytokines and performs many cellular functions. Its overexpression may lead to renal fibrosis. Aim: This study planed to investigate the effects of TGF-β1 on the cell cycle and phenotype of mesangial cells. Methods: Rat mesangial cells were cultured together with different concentrations (0, 1, 2, 5, and 10 ng/mL) of TGF-β1 for specified times from 0 min to 72 h. 0 ng/mL TGF-β1 and 0 min served as controls. Cell cycles were assessed by flow cytometry and α-smooth muscle actin expression (α-SMA) protein expression by western blot analysis. All data were presented as Mean ± SD. Statistical analysis was performed by using one-way analysis of variance and correlation analysis. Results were considered significant at p < 0.05. Results: After 15 min of co-culture with different concentrations of TGF-β1, the percentage of mesangial cells in G0/G1 phase was significantly elevated compared to the control ( p < 0.05). 12 h co-culture induced cell hyperplasia, 24 h co-culture obvious up-regulation of α-SMA ( p < 0.01) and one or two cells' myofibroblast phenotype transition, and 36 h co-culture several cells' phenotype transition. Correlation analysis prompted that the TGF-β1-induced premature aging was time-dependent ( p < 0.01). Conclusion: TGF-β1 may induce mesangial cells' premature senescence and myofibroblast-like phenotype transformation time-dependently, which may contribute to the development of early stage of glomerulosclerosis. [ABSTRACT FROM AUTHOR]

The non-classical Human leukocyte antigen G (HLA-G) differs from classical HLA class I molecules by its low genetic diversity, a tissue-restricted expression, the existence of seven isoforms, and immuno-inhibitory functions. Most of the known functions of HLA-G concern the membrane-bound HLA-G1 and soluble HLA-G5 isoforms, which present the typical structure of classical HLA class I molecule: a heavy chain of three globular domains α-α-α non-covalently bound to β-2-microglobulin (B2M) and a peptide. Very little is known of the structural features and functions of other HLA-G isoforms or structural conformations other than B2M-associated HLA-G1 and HLA-G5. In the present work, we studied the capability of all isoforms to form homomultimers, and investigated whether they could bind to, and function through, the known HLA-G receptors LILRB1 and LILRB2. We report that all HLA-G isoforms may form homodimers, demonstrating for the first time the existence of HLA-G4 dimers. We also report that the HLA-G α-α structure, which constitutes the extracellular part of HLA-G2 and HLA-G6, binds the LILRB2 receptor but not LILRB1. This is the first report of a receptor for a truncated HLA-G isoform. Following up on this finding, we show that the α-α-Fc structure coated on agarose beads is tolerogenic and capable of prolonging the survival of skin allografts in B6-mice and in a LILRB2-transgenic mouse model. This study is the first proof of concept that truncated HLA-G isoforms could be used as therapeutic agents. [ABSTRACT FROM AUTHOR]

In this study, changes of reactive oxygen species (ROS) and the mRNA expression of catalase of the Pacific white shrimp, Litopenaeus vannamei, exposed to pH (5.4, 6.7, 8.0, and 9.3) stress was investigated at different stress time (24, 48, 72, 96, and 120 h). Level of malondialdehyde (MDA) in shrimp also were assessed. The results revealed that acidic (pH 5.4 and 6.7) or alkaline exposure (pH 9.3) induced production of ROS hemocytes and increase of MDA level in shrimp. Moreover, the catalase mRNA expression in hepatopancreas of L. vannamei was up-regulated in 24 h at pH 5.4, in 72 h at pH 6.7 and in 48 h at pH 9.3, whereas was down-regulated significantly after 72 h acidic (pH 5.4 and 6.7) or alkaline (pH 9.4) exposure. In the present study, there was the relationship between ROS and catalase mRNA expression under normal acidic and alkaline conditions. At pH 8, the increase of catalase transcripts due to up-regulation by ROS, whereas MDA level did not significantly change, suggesting activation of corresponding protective mechanisms of detoxifying ROS is essential for the proper functioning of cells and the survival of shrimps. [ABSTRACT FROM AUTHOR]

Abstract: T-helper type 1 (Th1) immune response is involved in the development of protective immunity against Mycobacterium tuberculosis. Thus, an increase in Th1 and cellular immune responses should lead to enhanced anti-mycobacterial activity. In this study, we aimed to improve Th1 immune responses to a DNA vaccine by adding potentially immunostimulatory nucleotide sequences into the transcribed region downstream of the antigen. The Mycobacterium leprae gene for hsp65, codon-optimized for expression in mammalian cells, was inserted into pVAX1 with and without 3′-sequences containing CpG and dsRNA motifs. When the plasmid contained both motifs, transfected murine macrophage-like RAW264.7 cells showed markedly increased levels of mRNA for immune molecules of Th1 (IFN-α, IL-12) and Th17 (IL-17, IL-23 and IL-6) responses and for T cell co-stimulatory molecules (CD80 and CD86) but not for a Th2 response (IL-4 and IL-10). Immunized mice showed substantially increased serum anti-Hsp65 IgG2a antibody levels and IFN-γ production by spleen cells, confirming enhancement of the Th1 response in vivo. Furthermore, when non-vaccinated mice were infected with H37Rv by low-dose aerosol challenge, and then 4 weeks later were treated with plasmids by intramuscular injection, the mice that had been treated with plasmids containing immunostimulatory motifs showed an enhanced reduction in mycobacterial loads in lung and spleen. We conclude that DNA vaccines may be made more highly immunogenic and more effective for treatment by including transcribed stimulatory sequences. [Copyright &y& Elsevier]

Background: As novel biomarkers for various cancers, microRNAs negatively regulate genes expression via promoting mRNA degradation and suppressing mRNA translation. miR-589 has been reported to be deregulated in several human cancer types. However, its biological role has not been functionally characterized in gastric cancer. Here, we aim to investigate the biological effect of miR-589 on gastric cancer and to reveal the possible mechanism. Methods: Real-time PCR was performed to evaluate the expression of miR-589 in 34 paired normal and stomach tumor specimens, as well as gastric cell lines. Functional assays, such as wound healing, transwell assays and in vivo assays, were used to detect the biological effect of miR-589 and LIFR. We determined the role of miR-589 in gastric cancer tumorigenesis in vivo using xenograft nude models. Dual-luciferase report assays and Chromatin immunoprecipitation (ChIP) assay were performed for target evaluation, and the relationships were confirmed by western blot assay. Result: MiR-589 expression was significantly higher in tumor tissues and gastric cancer cells than those in matched normal tissues and gastric epithelial cells, respectively. Clinically, overexpression of miR-589 is associated with tumor metastasis, invasion and poor prognosis of GC patients. Gain- and loss-of function experiments showed that miR-589 promoted cell migration, metastasis and invasion in vitro and lung metastasis in vivo. Mechanistically, we found that miR-589 directly targeted LIFR to activate PI3K/AKT/c-Jun signaling. Meanwhile, c-Jun bound to the promoter region of miR-589 and activated its transcription. Thus miR-589 regulated its expression in a feedback loop that promoted cell migration, metastasis and invasion. Conclusion: Our study identified miR-589, as an oncogene, markedly induced cell metastasis and invasion via an atypical miR-589-LIFR-PI3K/AKT-c-Jun feedback loop, which suggested miR-589 as a potential biomarker and/or therapeutic target for the gastric cancer management. [ABSTRACT FROM AUTHOR]

Background: Cervical cancer is a common leading cause of cancer related to women death worldwide. Cylindromatosis (CYLD) is known as an important tumor suppressor in various human cancers, and a deubiquitination enzyme (DUB) as well. Previously, we identified Skp2 as an E3 ligase of Aurora B ubiquitination, but the DUB of Aurora B still remains unknown. Methods: Aurora B ubiquitination site is identified through in vivo ubiquitination assay. Activity of Aurora B and CENPA was detected by immunoblotting (IB) and immunofluorescence (IF) assay. Protein-to-protein interaction was investigated by immunoprecipitation (IP). Cell chromosome dynamics was monitored by live-cell time-lapse Imaging. Cancer cell proliferation, colony formation, apoptosis, and cell invasion and migration assays were also performed. Protein level was checked by immunohistochemical (IHC) staining in clinical cervical cancer samples. Results: We identified Lysine 115 (K115) as the main Aurora B ubiquitination site for Skp2. We could also detect an interaction of Aurora B with the DUB CYLD. We found that CYLD promoted deubiquitination of Aurora B, and regulated Aurora B activity and function as well. Compared with control, we found it took more time for the cells to finish cell mitosis with CYLD over-expression. Furthermore, we found that CYLD deficiency promoted cervical cancer cell proliferation, colony formation, cell migration and invasion, and inhibited apoptosis instead, whereas it is just opposite with CYLD over-expression. In clinical cervical cancer samples, we showed a negative correlation of CYLD expression with Aurora B activation and histological cancer cell invasion. Furthermore, there was less CYLD abundance and higher Aurora B activity in advanced cancer samples compared with early stage. Conclusions: Our findings uncover CYLD as a novel potential DUB of Aurora B, which inhibits Aurora B activation and its subsequent function in cell mitosis, and also provide more evidence for its tumor suppressor function in cervical cancer. [ABSTRACT FROM AUTHOR]

E2F-mediated control of gene expression is believed to have an essential role in the control of cellular proliferation. Using a conditional gene-targeting approach, we show that the targeted disruption of the entire E2F activator subclass composed of E2f1, E2f2, and E2f3 in mouse embryonic fibroblasts leads to the activation of p53 and the induction of p53 target genes, including p21CIP1. Consequently, cyclin-dependent kinase activity and retinoblastoma (Rb) phosphorylation are dramatically inhibited, leading to Rb/E2F-mediated repression of E2F target gene expression and a severe block in cellular proliferation. Inactivation of p53 in E2f1-, E2f2-, and E2f3-deficient cells, either by spontaneous mutation or by conditional gene ablation, prevented the induction of p21CIP1 and many other p53 target genes. As a result, cyclin-dependent kinase activity, Rb phosphorylation, and E2F target gene expression were restored to nearly normal levels, rendering cells responsive to normal growth signals. These findings suggest that a critical function of the E2F1, E2F2, and E2F3 activators is in the control of a p53-dependent axis that indirectly regulates E2F-mediated transcriptional repression and cellular proliferation. [ABSTRACT FROM AUTHOR]

Realgar (As 2 S 2 or As 4 S 4) is a traditional Chinese medicine (TCM) containing arsenic. Existing studies have shown that it has genotoxicity under long-term use with large doses. Niuhuang Jiedu (NHJD) is a Chinese medicine prescription containing realgar and seven other TCMs. Whether the multiple TCMs combination in NHJD can reduce the genotoxicity induced by realgar in equivalent doses is still unknown. To research the effect of NHJD on realgar's genotoxicity and the possible mechanism involved based on the arsenic methylation metabolic pathway. Six groups (control, realgar (0.8 g/kg), NHJD (12.48 g/kg), as well as Glycyrrhiza uralensis Fisch (GU), Scutellaria baicalensis Georg (SB), Rheum palmatum L (RP) plus equivalent doses of realgar, respectively) were set up. ICR mice were intragastric administered for 12 weeks. First, genotoxicology tests were conducted to evaluate the effect of NHJD, GU, SB, and RP on reducing realgar's genotoxicity. The inorganic arsenic (iAs), dimethyl arsenic acid (DMA), and monomethyl arsenic acid (MMA) were determined by HPLC-AFS, and the iAs%, MMA%, DMA%, primary methylation index (PMI), etc. Were calculated. Meanwhile, the S-adenosyl methionine (SAM) and arsenate reductase (ARR) levels, the arsenic (+3)methyltransferase (As 3 MT), purine-nucleoside phosphorylase (PNP) , glutathione S-transfer omega1 (GSTO1) gene expression were detected, aimed to explore the possible alleviation mechanisms of NHJD. The combination of multiple TCMs in NHJD decreased the levels of MN‰, SPA%, and DNA damage caused by realgar, with similar effects observed when SB, RP, and GU were used separately with realgar. Notably, the iAs% significantly decreased, while DMA% and PMI notably increased in the NHJD and realgar + SB (or RP) groups compared to the realgar-only group (P < 0.05). Increases in SAM and ARR levels were observed across various groups, but only the ARR increase in the NHJD group was statistically significant. Moreover, significant increases in As 3 MT mRNA and GSTO1 mRNA were noted in the NHJD group, and PNP mRNA levels significantly rose in the realgar + SB group. This study revealed that NHJD could attenuate the genotoxic effects of realgar. The botanicals SB, RP, and GU within NHJD may be key contributors to this effect. Enhancements in arsenic methylation capabilities through increased levels of SAM and ARR and elevated gene expressions of As 3 MT, PNP, and GSTO1 suggest potential mechanisms behind these findings. [Display omitted] • NHJD reduces the increment of MN‰, SPA%, and DNA damage caused by realgar. • NHJD increases arsenic methylation capacity compared to equivalent doses of realgar. • SAM and ARR proteins may play roles in NHJD's alleviating realgar genotoxicity. • As 3 MT, PNP, and GSTO1 genes may be involved in the detoxification mechanisms of NHJD. • Nine chemical compounds were identified in the prepared NHJD decoction. [ABSTRACT FROM AUTHOR]

As a chronic infectious disease, periodontitis can cause gum recession, loss of alveolar bone, loosening of teeth, and even loss of teeth. Dental plaque biofilm is the initiating factor for the occurrence and development of periodontitis. Fusobacterium nucleatum (F. nucleatum) plays a vital role in the structure and ecology of dental plaque biofilms. It is a bridge between early and late colonization bacteria in dental plaque. Understanding the molecular mechanism of F. nucleatum during biofilm development is essential to control periodontitis. This study aimed to determine gene expression profiles of the F. nucleatum strain, ATCC 25586, in the planktonic and biofilm phase through RNA-sequencing approach. The results were confirmed by quantitative reverse transcriptase PCR (RT-qPCR). The results clearly illustrate the difference in gene expression of F. nucleatum under planktonic and biofilms. A total of 110 genes were differentially expressed by F. nucleatum in the biofilm state compared with the planktonic state. The 25 upregulated genes in the biofilm state were mainly related to carbohydrate and amino acid metabolism, while the 85 downregulated genes were primarily associated with cell growth, division, and oxidative stress; most of the upregulated genes of F. nucleatum involved in virulence and oral malodor. Furthermore, the transcriptome analysis and antibacterial activity test also identified Lysine might exhibit the antibacterial and antibiofilm activity of F. nucleatum for the first time. These new findings could provide caveats for future studies on the regulation and maintenance of plaque biofilm and the development of biomarkers for periodontitis. • The upregulated genes in biofilm were mainly related to carbohydrates, amino acid metabolism, and regulation. • The downregulated genes in biofilm state were mainly related to the transport and metabolism of inorganic ions, ribosomal proteins, cell division, and the cell cycle. • We found that Lysine can inhibit the planktonic growth and biofilm formation of F. nucleatum for the first time. [ABSTRACT FROM AUTHOR]

Sex differences play an important role in depression, the basis of which is an excessive stress response. We aimed at revealing the neurobiological sex differences in the same study in acute- and chronically-stressed rats. Female Sprague-Dawley (SD) rats were randomly divided into 6 groups: chronic unpredictable mild stress (CUMS), acute foot shock (FS) and controls, animals in all 3 groups were sacrificed in proestrus or diestrus. Male SD rats were randomly divided into 3 groups: CUMS, FS and controls. Comparisons were made of behavioral changes in CUMS and control rats, plasma levels of corticosterone (CORT), testosterone (T) and estradiol (E2), and of the hypothalamic mRNA-expression of stress-related molecules, i.e. estrogen receptor α and β, androgen receptor, aromatase, mineralocorticoid receptor, glucocorticoid receptor, corticotropin-releasing hormone, arginine vasopressin and oxytocin. CUMS resulted in disordered estrus cycles, more behavioral and hypothalamic stress-related molecules changes and a stronger CORT response in female rats compared with male rats. Female rats also showed decreased E2 and T levels after FS and CUMS, while male FS rats showed increased E2 and male CUMS rats showed decreased T levels. Stress affects the behavioral, endocrine and the molecular response of the stress systems in the hypothalamus of SD rats in a clear sexual dimorphic way, which has parallels in human data on stress and depression. [ABSTRACT FROM AUTHOR]

Highlights: [•] Skp2 orchestrates homing ability by maintaining the HSC binding to endosteal niche. [•] β-Catenin is downregulated in Skp2 difficient HSC. [•] Skp2 and β-catenin maintain HSC homing ability in a cell cycle independent manner. [•] Skp2 is a novel and critical regulator for HSC homing. [ABSTRACT FROM AUTHOR]