Circ_0081054 facilitates melanoma development via sponging miR-637 and regulating RAB9A.

BACKGROUND: Accumulating evidence announces that aberrantly expressed circRNAs were closely related to the development of human cancers. However, the role and mechanism of multiple circRNAs remain unclear. Our work aimed to disclose the functional role and mechanism of circ_0081054 in melanoma. METHODS: Quantitative real-time polymerase chain reaction assay was utilized to detect circ_0081054, microRNA-637 (miR-637) and RAB9A (member RAS oncogene family) mRNA expression. Cell proliferative ability was evaluated via Cell Counting Kit-8 and colony formation assay. Cell invasion was assessed by using wound healing assay. RESULTS: The significant upregulation of circ_0081054 was detected in melanoma tissues and cells. The proliferation, migration, glycolytic metabolism, and angiogenesis in melanoma cells were suppressed, while apoptosis was promoted following the silence of circ_0081054. In addition, circ_0081054 could target miR-637, and miR-637 inhibitor could reverse the effects of circ_0081054 deficiency. Furthermore, RAB9A was a target gene for miR-637 and RAB9A overexpression could reverse the effects of miR-637 overexpression. In addition, the deficiency of circ_0081054 hampered tumor growth in vivo. Moreover, circ_0081054 could regulate RAB9A expression by sponging miR-637. CONCLUSION: All results indicated that circ_0081054 promoted the malignant behaviors of melanoma cells partly by regulating the miR-637/RAB9A molecular axis.

Efficacy of acupoint application in patients with overactive bladder: A randomized, placebo-controlled pilot trial.

OBJECTIVE: Conventional treatments for alleviating the symptoms of Overactive bladder (OAB) have been reported to have limited efficacy and a high rate of side effects. Traditional Chinese medicine (TCM) has been used in Asia countries because of its low side effects and being easy to operate. To confirm the efficacy of acupoint application treatment for alleviating OAB symptoms, a randomized and placebo-controlled pilot trial was conducted in this study. METHODS: All participants were randomly allocated into a treatment group or control group, receiving either a "Dinggui" acupoint application or placebo treatment for 4 weeks. The outcome measures were OAB symptom scores (OABSS), OAB questionnaire (OAB-q) scores, and TCM syndrome scores. Urine nerve growth factor (NGF) level, NGF normalized to urine creatinine (NGF/Cr), and maximum flow rate (Qmax) were also measured to evaluate the OAB symptoms. RESULTS: In total, 69 participants were included with 34 in the treatment group and 35 in the placebo-treated group. Treatment with "Dinggui" acupoint application showed a statistically significant decrease in OABSS scores (8.10±1.54 to 3.67±1.77), OAB-q scores (61.43±13.93 to 38.13±15.42), and TCM syndrome scores (15.60±5.98 to 9.20±4.82). The NGF and NGF/Cr were also observed meaningful changes in a decrease from 379.68 to 136.17 pg/ml and from 0.30 to 0.16 pg/mg, respectively. The Qmax value showed a significant increase from 14.40 to 24.05 ml/s. CONCLUSIONS: Treatment with "Dinggui" acupoint application could be considered an effective and alternative therapy for OAB management. Further studies with larger sample sizes and longer treatment periods are needed to investigate.

Two coexisting pseudo-mirror heteromolecular telomeric G-quadruplexes in opposite loop progressions differentially recognized by a low equivalent of Thioflavin T.

The final 3'-terminal residue of the telomeric DNA G-overhang is inherently less precise. Here, we describe how alteration of the last 3'-terminal base affects the mutual recognition between two different G-rich oligomers of human telomeric DNA in the formation of heteromolecular G-quadruplexes (hetero-GQs). Associations between three- and single-repeat fragments of human telomeric DNA, target d(GGGTTAGGGTTAGGG) and probe d(TAGGGT), in Na+ solution yield two coexisting forms of (3 + 1) hybrid hetero-GQs: the kinetically favourable LLP-form (left loop progression) and the thermodynamically controlled RLP-form (right loop progression). However, only the adoption of a single LLP-form has been previously reported between the same probe d(TAGGGT) and a target variant d(GGGTTAGGGTTAGGGT) having one extra 3'-end thymine. Moreover, the flanking base alterations of short G-rich probe variants also significantly affect the loop progressions of hetero-GQs. Although seemingly two pseudo-mirror counter partners, the RLP-form exhibits a preference over the LLP-form to be recognized by a low equivalent of fluorescence dye thioflavin T (ThT). To a greater extent, ThT preferentially binds to RLP hetero-GQ than with the corresponding telomeric DNA duplex context or several other representative unimolecular GQs.

CircRNA circ_0067772 aggravates the malignant progression of cutaneous squamous cell carcinoma by regulating miR-1238-3p/FOXG1 axis.

BACKGROUND: Cutaneous squamous cell carcinoma (CSCC) is a severe malignancy derived from skin. Dysregulated circular RNAs (circRNAs) might play vital roles in tumor development. OBJECTIVE: Here, we aimed to explore the function of a novel circRNA circ_0067772 in CSCC. METHODS: Quantitative real-time PCR (qRT-PCR) or Western blot assay was performed to determine the expression of circ_0067772, microRNA (miR)-1238-3p and forkhead box protein G1 (FOXG1). Cell proliferation was assessed by Cell Counting Kit-8 (CCK-8) assay and colony formation assay. Transwell assay and wound healing assay were employed to examine cell metastasis. Flow cytometry was employed to monitor cell cycle and apoptosis. The target association between miR-1238-3p and circ_0067772 or FOXG1 was validated by dual-luciferase reporter assay. Moreover, role of circ_0067772 in vivo was investigated via xenograft model in nude mice. RESULTS: Circ_0067772 and FOXG1 were upregulated, while miR-1238-3p was downregulated in CSCC tissues and cells. Circ_0067772 knockdown conferred inhibitory effects on cell proliferation, migration and invasion of CSCC cells. MiR-1238-3p served as a target of circ_0067772, whose silencing could reverse circ_0067772 knockdown-induced inhibitory impact on the malignant cellular behaviors. Circ_0067772 positively regulated FOXG1 expression by antagonizing miR-1238-3p. Additionally, miR-1238-3p could repress CSCC cell proliferation, migration and invasion by targeting FOXG1. Also, circ_0067772 knockdown hindered CSCC tumor growth in vivo. CONCLUSION: Our study identified a novel oncogenic circRNA and the involvement of circ_0067772/miR-1238-3p/FOXG1 axis in CSCC development, providing a target for CSCC therapy.

NMR structural study on the self-trimerization of d(GTTAGG) into a dynamic trimolecular G-quadruplex assembly preferentially in Na+ solution with a moderate K+ tolerance.

Vast G-quadruplexes (GQs) are primarily folded by one, two, or four G-rich oligomers, rarely with an exception. Here, we present the first NMR solution structure of a trimolecular GQ (tri-GQ) that is solely assembled by the self-trimerization of d(GTTAGG), preferentially in Na+ solution tolerant to an equal amount of K+ cation. Eight guanines from three asymmetrically folded strands of d(GTTAGG) are organized into a two-tetrad core, which features a broken G-column and two width-irregular grooves. Fast strand exchanges on a timescale of second at 17°C spontaneously occur between folded tri-GQ and unfolded single-strand of d(GTTAGG) that both species coexist in dynamic equilibrium. Thus, this tri-GQ is not just simply a static assembly but rather a dynamic assembly. Moreover, another minor tetra-GQ that has putatively tetrameric (2+2) antiparallel topology becomes noticeable only at an extremely high strand concentration above 18 mM. The major tri-GQ and minor tetra-GQ are considered to be mutually related, and their reversible interconversion pathways are proposed accordingly. The sequence d(GTTAGG) could be regarded as either a reading frame shifted single repeat of human telomeric DNA or a 1.5 repeat of Bombyx mori telomeric DNA. Overall, our findings provide new insight into GQs and expect more functional applications.

Effects of a Caregiver Training Program on Oral Hygiene of Alzheimer's Patients in Institutional Care.

OBJECTIVES: To investigate the effects of a caregiver training program on the oral hygiene of caregivers and patients with Alzheimer's disease (AD) and to identify program components and parameters for accurate assessment of outcomes. DESIGN: Single-blinded prospective cohort study. SETTING AND PARTICIPANTS: Patients with AD and caregivers in nursing homes in the Greater Zhengzhou Area, China. METHODS: Initially 168 AD patient/caregiver pairs were recruited and randomly assigned to control, limited training, and comprehensive training groups. The mini-mental state examination, global deterioration scale, and Katz activities of daily living scale were conducted for patients with AD. Information on participants' oral hygiene habits and general oral health was collected. The modified Quigley-Hein Plaque Index (PI) and Gingival Index (GI) were used to assess oral hygiene and gingival health. Intervention included (1) an educational video showing the role of dental plaque and the modified Bass technique; and (2) caregivers practicing toothbrushing on themselves and patients with AD under professional guidance. Changes in oral hygiene and correlations between patient PI/GI and caregiver PI/GI were analyzed. RESULTS: After 6 weeks, complete data for 146 AD patient/caregiver pairs were collected. Before enrollment, most patients with AD had very poor oral hygiene. Compared with controls and limited training, only comprehensive training was able to achieve steady reduction in PI and GI scores in patients with AD, which still fell short of desirable levels (PI: 2.46 ± 0.52, GI: 1.24 ± 0.24, week 6). PI and GI scores in caregivers saw steady improvement only through comprehensive training (PI: 1.41 ± 0.38, GI: 0.88 ± 0.19, week 6). Number of training sessions had the greatest influence on both patient PI and GI scores. CONCLUSIONS AND IMPLICATIONS: Comprehensive caregiver training on toothbrushing skills is effective in improving the oral hygiene of caregivers and patients with AD in nursing homes. Additional evidence is needed to establish the optimal program structure.

LINC00858 facilitates the malignant development of Wilms' tumor by targeting miR-653-5p.

BACKGROUND: To uncover the clinical significance of LINC00858 in the development of Wilms' tumor and the potential molecular mechanism. METHODS: LINC00858 levels in Wilms' tumor species and cell lines were determined by quantitative real-time polymerase chain reaction (qRT-PCR). The clinical significance of LINC00858 in influencing pathological features and prognosis in patients with Wilms' tumor was analyzed. Proliferative and migratory changes in Wilms' tumor cells with LINC00858 knockdown were assessed. The downstream gene of LINC00858 was verified by luciferase assay, and its involvement in the development of Wilms' tumor was further explored. RESULTS: LINC00858 was highly expressed in Wilms' tumor tissues and cell lines. High level of LINC00858 was correlated to high rate of lymphatic metastasis and poor prognosis in patients with Wilms' tumor. Knockdown of LINC00858 suppressed proliferative and migratory potentials in HFWT and 17-94 cells. MiR-653-5p was targeted by LINC00858. It was lowly expressed in Wilms' tumor tissues and negatively regulated by LINC00858. Knockdown of miR-653-5p partially abolished the regulatory effects of LINC00858 on proliferative and migratory potentials in Wilms' tumor cells. CONCLUSIONS: LINC00858 is highly expressed in Wilms' tumor species, and correlated to lymphatic metastasis rate and overall survival in patients with Wilms' tumor. Knockdown of LINC00858 suppresses Wilms' tumor cells to proliferate and migrate via targeting miR-653-3p.

Juglanin administration protects skin against UVB­induced injury by reducing Nrf2­dependent ROS generation.

Extensive solar ultraviolet B (UVB) exposure of the skin results in inflammation and oxidative stress, which may contribute to skin cancer. Natural products have attracted attention for their role in the effective treatment of cutaneous neoplasia. Juglanin is purified from the crude extract of Polygonum aviculare, exhibiting anti­oxidant, anti­inflammatory and anti­cancer activities. Jugalanin was used in the current study to investigate whether it may ameliorate UVB irradiation­induced skin damage by reducing oxidative stress and suppressing the inflammatory response in vivo and in vitro. In the present study, hairless mice were exposed to UVB irradiation in the absence or presence of juglanin administration for 10 weeks. The findings indicated that juglanin inhibited UVB­induced hyperplasia and decreased infiltration in the skin of mice. UVB exposure­induced oxidative stress in mice and cells was inhibited by juglanin via enhancing anti­oxidant activity. Additionally, juglanin markedly reduced pro­inflammatory cytokine release, including cyclic oxidase 2, interleukin­1ß and tumor necrosis factor­α, triggered by chronic UVB irradiation. Juglanin­ameliorated skin damage was associated with its suppression of mitogen activated protein kinases (MAPKs), including p38, extracellular signal regulated 1/2, and c­Jun N­terminal kinases, as well as nuclear factor (NF)­κB signaling pathways, which was dependent on nuclear factor­E2­related factor 2 (Nrf2)­modulated reactive oxygen species generation. Taken together, these data indicate that juglanin protected against UVB­triggered oxidative stress and inflammatory responses by suppressing MAPK and NF­κB activation via enhancing Nrf2 activity.

Fisetin inhibits pristine-induced systemic lupus erythematosus in a murine model through CXCLs regulation.

Systemic lupus erythematosus (SLE) is associated with an increased risk of vascular complications. Lupus nephritis is a major manifestation of SLE in the clinic. Lupus nephritis is elevated by T helper type 17 (Th17) cells, the major pro­inflammatory T­cell subset, leading to autoimmunity modulation. Therapeutic treatments targeting leukocyte recruitment may be useful in attenuating vascular complications linked to SLE progression. 3,7,3',4'­Tetrahydroxyflavone (fisetin) is a flavonol and a member of the flavonoid polyphenols. It is present in various fruits and vegetables, including persimmons, apples, kiwis, grapes, onions, strawberries and cucumbers. In the present study, the effects of fisetin against SLE induced by pristane (PRI) were evaluated in mice. Fisetin was indicated to reduce PRI­induced anti­double stranded DNA, anti­ small nuclear ribonucleoprotein and the ratio of albumin to creatinine in urine. In addition, the chemokine (C­X­C motif) ligand (CXCL) signaling pathway was activated for PRI treatment, which was reversed by fisetin administration by reducing CXCL­1 and 2, chemokine (C­C motif) ligand 3, as well as CXC receptor 2 expression. In addition, the induction of inflammatory cytokines, including interleukin (IL)­6, tumor necrosis factor­α, IL­1ß, as well as the chemokine interferon­Î³, by PRI were downregulated by fisetin treatment in mice. Furthermore, Th17 cells and their associated cytokines were highly induced by PRI treatment, which was inhibited by fisetin administration. The present results indicated that fisetin may be an effective management for SLE by targeting the CXCL signaling pathway and regulating Th17 differentiation during lupus nephritis development.

Salidroside prevents skin carcinogenesis induced by DMBA/TPA in a mouse model through suppression of inflammation and promotion of apoptosis.

Salidroside (SR) is a main component of Rhodiola rosea L. and exhibits a variety of pharmacologic properties. The present study was carried out to explore the potential effect of SR against skin cancer induced by 7,12-dimethylbenz(a)anthracene (DMBA) and 12-O-tetradecanoylphorbol-13­acetate (TPA) in female Institute for Cancer Research (ICR) mice and to reveal the underlying molecular targets regulated by SR. The mice were randomly divided into 4 groups: control, DMBA/TPA, DMBA/TPA+SR (20 mg/kg) and DMBA/TPA+SR (40 mg/kg). SR was administered to mice five times a week after DMBA treatments. In our study, we found that SR dose-dependently ameliorated skin cancer incidence and the multiplicity in the animal models by reducing the release of inflammation-related cytokines, including tumor necrosis factor α (TNF-α), interleukin-1ß (IL-1ß), interleukin-18 (IL-18), interleukin-6 (IL-6), cyclooxygenase 2 (COX2) and transforming growth factor ß-1 (TGF-ß1). Suppression of the nuclear factor (NF)-κB signaling pathway by SR was effective to prevent skin carcinogenesis. Furthermore, TUNEL analysis indicated that compared to the DMBA/TPA group, enhanced apoptosis was observed in the DMBA/TPA+SR group. In addition, p53 expression levels were increased by SR in the DMBA/TPA-induced mice. Therefore, SR was effective for inducing apoptosis during skin cancer progression triggered by DMBA/TPA. Consistently, p21, p53 upregulated modulator of apoptosis (PUMA), Bax and caspase-3 were highly induced by SR to enhance the apoptotic response for preventing skin cancer. Moreover, in vitro, we found that SR dramatically reduced the inflammatory response, while enhancing the aoptotic response by blocking NF-κB and activating caspase-3 pathways, respectively. In addition, flow cytometric analysis further confirmed the induction of apoptosis by SR in DMBA-treated cells in vitro. Taken together, the in vivo and in vitro studies illustrated that SR might be a promising compound to reduce skin cancer risk.

IRF1 up-regulates isg15 gene expression in dsRNA stimulation or CSFV infection by targeting nucleotides -487 to -325 in the 5' flanking region.

Interferon (IFN)-stimulated gene 15 (ISG15) encodes a ubiquitin-like protein that is heavily involved in immune response elicitation. As an important member of interferon regulatory factor (IRF) family, IRF1 can activate the expression of multiple genes, including the human optineurin gene (Sudhakar et al., 2013). In this study, a sequence in the promoter region of the optineurin gene was compared to the 5' flanking region of the porcine isg15 gene. Porcine IRF1 also possesses antiviral activity against several swine viruses (Li et al., 2015), but the mechanism is not well understood. Herein, we report that porcine IRF1 and ISG15 were up-regulated in porcine kidney (PK-15) cells following stimulation with double-stranded RNA (dsRNA) or classical swine fever virus (CSFV) infection. We also found that siRNA-mediated knockdown of IRF1 expression resulted in lower ISG15 expression in response to polyinosinic:polycytidylic acid [poly(I:C)] or CSFV infection. The overexpression of IRF1 resulted in ISG15 up-regulation. IRF1 was shown to translocate to the nucleus in response to dsRNA stimulation. To further identify the functional domain of the isg15 gene that promotes IRF1 transcriptional activity, firefly luciferase and ISG15 reporter systems were constructed. The results of the firefly luciferase and ISG15 reporter assay suggested that IRF1 mediates the up-regulation of ISG15. Nucleotides -487 to -325, located in the 5' flanking region of the isg15 gene, constituted the promoter region of IRF1. ChIP assay indicated that IRF1 protein was able to interact with the DNA in the 5'fr of isg15 gene in cells. As an innate immune response protein with broad-spectrum antiviral activity, the up-regulation of ISG15 mediated by IRF1 in porcine cells is reported for the first time. These results warrant further investigation into the antiviral activity of porcine IRF1 against reported swine viruses.

Galangin inhibits the cell progression and induces cell apoptosis through activating PTEN and Caspase-3 pathways in retinoblastoma.

Retinoblastoma is reported as a rare cancer that occurs during childhood. Although several treatments are available for retinoblastoma, there is a need for alternative new treatment modalities for retinoblastoma with better safety and efficacy profile. Galangin (3,5,7-trihydroxyflavone), is a flavonoid compound, which is found in high concentration in lesser galangal. Galangin has been reported to have various bioactivities, including anti-inflammation, anti-oxidative stress and anti-cancer through various pathways. The objective of our study was to explore the effects of galangin on the suppression of retinoblastoma in vitro and in vivo. Using MTT analysis, transwell-chamber migration analysis, colony-forming analysis, wound healing analysis, immunofluorescent assay of KI-67, we found that galangin exhibited a suppressive effect on human retinoblastoma cell proliferation and migration. Moreover, PTEN, a tumor-suppressor, was increased by galangin in cancer cells and in tumor tissues isolated from retinoblastoma xenograft models. Additionally, galangin reduced protein kinase B (Akt) phosphorylation, which was associated with PTEN up-regulation. Galangin-reduced Akt activation and cell proliferation was abolished by PTEN knockdown, which might be associated with the over-expression of phosphatidylinositol-3,4,5-triphosphate (PIP3)/diphosphate product (PIP2). Furthermore, flow cytometry, Hoechst 33258 staining and western blot assays indicated that galangin could induce apoptosis through promoting Caspase-3 pathway, which was, at least partly, dependent on PTEN expression. Our data illustrated that galangin treatment suppressed the growth of retinoblastoma tumor in vivo by anti-proliferative and apoptogenic mechanisms. Thus, galangin might be a safe and promising non-chemotherapeutic drug, which could be useful as an adjuvant against retinoblastoma.

[Canine Circovirus Genome Cloning and Sequence Analysis].

Dog circovirus (DogCV) is a newly discovered mammalian circovirus. To investigate the genomic characteristics and genetic diversity of DogCV spreads in China, the first genome sequence of Chinese isolate, designated as JZ98/2014,was obtained by overlap PCR using the DNA extracted from dog serum as template for amplification. The nucleotide content and genome organization were subsequently analyzed. The results showed that the full-length genome of JZ98/2014 is 2063nt,and contains three major open reading frame: ORF V1 (encodes the 303 amino acid Rep protein),ORF C1(encodes the 270 amino acid Cap protein),and ORF C2(encodes 106 amino acids).JZ98/2014 shared 82.1%-89.5% homology with the complete genome sequences of DogCV isolates from America and Europe. The Rep gene and Cap gene of JZ98/2014 shared 82.1%-89.5%and 84.6%-89.1% homology, respectively, with the same genes from other DogCVs. Phylogenetic tree analysis indicated that there were several different genetic clades of DogCV spread in the world, and JZ98/2014 formed a clade by itself.