Silence Research Articles

Construction of a lentiviral vector carrying the casitas B-lineage lymphoma (Cbl) -b short hairpin RNA (shRNA) and its effect on the biological behavior of A375 melanoma cells in vitro

Objective To construct a recombinant lentiviral vector carrying the casitas B-lineage lymphoma (Cbl) -b short hairpin RNA (shRNA) , and to evaluate its effect on the biological behavior of A375 melanoma cells in vitro. Methods Three specific shRNAs targeting Cbl-b gene and a negative control shRNA were designed and synthesized, and recombinant lentiviral vectors were constructed. A375 cells were divided into 5 groups to be transfected with 3 kinds of lentiviral vector expressing Cbl-b gene-specific shRNAs (CBLB-shRNA-1 group, CBLB-shRNA-2 group and CBLB-shRNA-3 group) , a lentiviral vector containing negative control shRNA (negative control group) , and an empty vector (blank control group) . Real-time fluorescence-based quantitative PCR and Western blot analysis were performed to determine the silencing efficiency at 72 hours after transfection. Cell counting kit-8 (CCK-8) assay was conducted to evaluate cellular proliferative activity at 24, 48, 72 and 96 hours after transfection, flow cytometry to detect cell apoptosis and cell cycle at 72 hours after transfection, and Transwell invasion assay to assess cellular invasive activity at 72 hours after transfection. Results Three recombinant lentiviral vectors containing Cbl-b shRNA were constructed successfully. As Western blot analysis revealed, the CBLB-shRNA-3 showed the highest silencing efficiency. CCK-8 assay indicated that the proliferative activity of A375 cells was significantly lower in the CBLB-shRNA-3 group than in the negative control group and blank control group at 72 and 96 hours after transfection (all P < 0.01) . Flow cytometry showed that the apoptosis rate of A375 cells was significantly higher in the CBLB-shRNA-3 group (22.73% ± 6.58%) than in the negative control group (6.08% ± 1.35%, P < 0.01) and blank control group (6.34% ± 1.07%, P < 0.01) . The CBLB-shRNA-3 group showed a significantly higher proportion of A375 cells at G1 phase, but a significantly lower proportion of A375 cells at S phase compared with the negative control group and blank control group (all P < 0.01) . Transwell assay showed that there were significant differences in the number of A375 cells crossing the artificial basement membrane (matrigel) at 72 hours after transfection among the negative control group, blank control group and CBLB-shRNA-3 group (76.60 ± 1.82, 73.20 ± 3.83, 19.60 ± 1.14, respectively; F = 794.50, P < 0.01) . Conclusions A recombinant CBLB-shRNA-3-expressing lentiviral vector which can efficiently silence Cbl-b gene has been successfully constructed. It can inhibit the proliferation, cell cycle progression and invasive activity of A375 cells, but promote the apoptosis of A375 cells. Key words: Melanoma; Proto-oncogene proteins c-cbl; RNA, small interfering; Cell proliferation; Apoptosis; Cell cycle; Gene, Cbl-b; A375 cells; Short hairpin RNA

Effect of microRNA- 193a on the proliferation and migration ability of human lung carcinoma A549 cells by promyosin

Objective To investigate the effect and the related mechanism of microRNA (miRNA, miR)- 193a on the proliferation and migration ability of human lung carcinoma A549 cells by promyosin (TPMF). Methods Western blotting was used to select the cell lines with the lowest expression of TPMF in lung cancer cell lines. The TPMF gene was silenced through TPMF Silencing Lentivirus (LV3- TPMF). green fluorescent protein (GFP) fluorescence detector and Western bolting were used to detect the efficiency of gene silence. Western blotting and reverse transcriptase-polymerase chain reaction (RT-PCR) were used to detect the correlation between miR- 193a and TPMF. Effect of TPMF on the invasion ability of A549 cells was examined by Transwell matrigel invasion assays. Effect of TPMF on the A549 cells proliferation ability was examined by cell clone assays. The correlation between miR- 193a and TPMF was analyzed by Western blotting and RT- PCR. The Effect of TPMF Silencing on Proliferation of Tumor in Nude Mice Survival Rate was detected. Results LV3- TPMF lentivirus could effectively inhibit the expression of TPMF [(88.29±8.62)% vs. (17.82±4.91)%, P=0.010]. Silencing the expression of TPMF could inhibit the invasion ability of human lung carcinoma A549 cells [(351.23±6.92)% vs. (83.26±3.57)%, P=0.023]. Silence of TPMF could inhibit the proliferation of human lung carcinoma A549 cells [(103.82±5.12)% vs. (22.34±1.91)%, P=0.030]. Silence of TPMF could extend the survival rates of nude mice. Conclusion MiR- 193a can inhibit the invasion and proliferation of lung carcinoma A549 cells by regulating the expression of TPMF. Key words: MicroRNA-193a; Lung carcinoma; Transwell; Promyosin; Survival rate

Beyond the Historical Record? Henry James in “The Master at St. Bartholomew’s Hospital

The article analyses the short-story “The Master at St Bartholomew’s Hospital 1914–1916” by Joyce Carol Oates (2007) in the broader context of the Jamesian biofiction, a series of novels and tales featuring Henry James as their protagonist. The addition of the prefix “bio-” to “fiction” points out the hybrid nature of these texts, which are a melange of biography, autobiography, criticism and fiction. Oates’s story not only epitomizes this hybridity, but it also proves to be an exploration of the potentiality of this subgenre to penetrate the mystery surrounding James’s persona and saturate the lacunae in his biography by resorting to what David Lodge defined as “the novelist’s licence”. The short-story is yet another evidence of Oates’s fascination with the unsaid in James’s life and prose, because it revolves around the silence into which he sank at the outbreak of the Great War, when he did not write anything in his pocket diaries for three months. In an attempt to go beyond the limits of the historical record, Oates gives insights into the mind of the author by depicting a Henry James in crisis – nagged by doubts about his artistic legacy – in an atmosphere of uncertainty enhanced by a complex intertextual play. The result is ‘a Henry James’ slightly divergent from the historical one: thus, the tale advocates the inaccessibility of the private life of a real individual. Nonetheless, the acknowledgement of this limit spurs the celebration of fictional imagination.

Borders and silence as forms of knowledge in Amitav Ghosh’s The Calcutta Chromosome

Forms of knowledge and our understanding of the world are central concerns in Amitav Ghosh's fiction. His novels often deconstruct epistemology derived from the Western philosophical tradition, with the aim of pointing at its relativity in spite of its claim to universality. This essay wants to demonstrate how The Calcutta Chromosome (1996) closely connects the production of an alternative form of knowledge (the counter-science) with a different way of perceiving and conceiving space. Focusing specifically on Calcutta's urban space, it will examine the correlation between space and epistemology in the novel through three spatial and gnoseological categories: borders, the labyrinth, and, finally, networks.

Terry Waite, Out of the Silence: Memories/Poems/Reflections

Terry Waite, <i>Out of the Silence: Memories/Poems/Reflections</i>

Interferon-α enhances etoposide-induced apoptosis and cell cycle arrest

Objective To study effect of interferon-α (IFN-α) on etoposide-induced cell cycle arrest and apoptosis in human osteosarcoma cells with its mechanisms. Methods Osteosacoma U2OS cells were treated with IFN-α and etoposide, alone or in combination, for 72 h. Growth inhibition was determined with trypan blue exclude test. Cell cycle arrest was evaluated with FACS. Apoptosis was detected through Hoechst33258 staining. The expression of p53 was determined with reverse transcriptase-polymerase chain reaction (RT-PCR) in mRNA level. small interfering RNA (siRNA) interference was used to silence p53. Western blotting was used to evaluate the expression of p53 and poly adenosine diphosphate-ribose polymerase (PARP). Results After treatment for 72 h, the growth inhibition was (2.29±1.03)%, (23.85±3.09)% and (50.51±3.77)% in IFN-α, Etoposide and combination group. IFN-α enhanced etoposide-induced growth inhibiton. There were statistic differences (P=0.008). There were apoptotic morphological changes in etoposide and combination groups. Cells in S phase were (19.68±3.62)% and (11.33±2.94)% in Etoposide and combination group, while G2/M cells were (65.02±2.55)% and (76.09±2.87)%. There were statistic differences (P=0.037). IFN-α did not further increase the expression of p53 in mRNA level. The inhibition rate was (55.72±3.88)%, (51.63±4.36)% and (36.40±3.95)% in blank, control-siRNA and p53-siRNA group after p53 silence. There were statistic differences (P=0.023). p53 silence largely decreased PARP cleavage in response to the combination. After pan cysteinyl aspartate-specific protease (Caspase) inhibitor Z-VAD-FMK pre-treatment, the growth inhibition rate was (3.52±1.98)%, (27.11±4.06)%, (56.44±4.78)%, (21.37±3.55)% and (23.81±4.22)% in Z-VAD-FMK, Etoposide, IFN-α+ Etoposide, etoposide+ Z-VAD-FMK and IFN-α+ Etoposide+ Z-VAD-FMK group. Z-VAD-FMK decreased growth inhibition caused by combination. There were statistic defferences (P=0.015). PARP cleavage caused by combination was also inhibited by Z-VAD-FMK. Conclusion IFN-α enhances etoposide-mediated growth inhibition, cell cycle arrest and apoptosis in human osteosarcoma U2OS cells. The growth inhibiton and apoptosis are p53-dependent and Caspase-dependent. Key words: Interferon-α; Etoposide; Osteosarcoma; Cell cycle arrest; Apoptosis

Automatic prosodic boundary labeling based on fusing the silence duration with the lexical features

Automatic prosodic boundary labeling based on fusing the silence duration with the lexical features

Klf10 silencing inhibited mechanical force induced osteogenic differentiation of human periodontal ligament cells

To investigate the effect of Klf10 silence on human periodontal ligament cells (HPDLCs) under mechanical force. HPDLCs were isolated and transfected with Klf10 siRNA, and then exposed to centrifugal force for 6 h at 631 r/min. Purmorphamine, an hedgehog signaling pathway agonist, was used for intervention. The activity of alkaline phosphatase (ALP) was detected by ELISA. RT-PCR and Western blot were performed to detect the mRNA and protein expression of Klf10, Runt related transcription factor 2 (Runx2), osteopontin (OPN) and osteocalcin (OCN). The protein expression of glioma associated oncogene homolog 1 (Gli1) and patched-1 (PTCH1) was detected by Western blot. SPSS 20.0 software package was used to analyze the data. Mechanical force increased mRNA and protein level of Klf10, Runx2, OPN and OCN, and elevated ALP activity significantly (P<0.05). Mechanical force also upregulated the protein expression of GLI1 and Ptch1 significantly (P<0.05). Compared with the control group, Klf10 siRNA transfection significantly decreased mRNA and protein level of Klf10 (P<0.05). Klf10 siRNA significantly inhibited the activity of ALP, and downregulated mRNA and protein expression of Runx2, OPN and OCN (P<0.05). Moreover, Klf10 siRNA significantly inhibited protein expression of GLI1 and Ptch1 (P<0.05), and purmorphamine obviously inhibited the effect of Klf10 siRNA (P<0.05). Klf10 silencing could inhibit bone differentiation of human periodontal ligament cells under mechanical force, which may be through regulation of hedgehog signaling pathway.

“Disgusting Details Which Are Best Forgotten”: Disclosures of Child Sexual Abuse in Twentieth-Century Britain

For those who by the end of the twentieth century came to be termed “survivors” of child sexual abuse, different genres and forms have been available to narrate and evaluate that abuse. This article explores the reception and practical results of such disclosures: the unpredictable effects of telling, and the strategies of containment, silencing, or disbelief that greeted disclosures. I make note of the ethical challenges of writing the history of child sexual abuse and conclude that twenty-first-century observers have been too ready to perceive much of the previous century as a period of profound silence in relation to child sexual abuse. At the same time, historical and sociological accounts have also been too ready to claim the final third of the twentieth century as a period of compulsive disclosure and fluency in constructing sexual selves. The history of child sexual abuse reveals significant barriers to disclosure in the 1970s and 1980s, despite new visibility of child sexual abuse in the media and through feminist sexual politics. Attention to such obstacles suggests the need to rethink narratives of “permissive” sexual change to acknowledge more fully the ongoing inequities and hierarchies in sexual candor and voice.

The Constitutional Convention, The Sequel

(Overture. A long loud crashing wonderful piece followed by silence. Darkness. The sound of water, as of a trickling stream or little fountain, and the occasional sound of a plucked string or other beautiful musical sound. A scrim fills the back of the stage. The scrim is a delicate pink at the moment. A stuffed deer stands upstage. Through the scrim are projected such rules of civility as these: THE POLITE DINNER GUEST ALWAYS SPEAKS WITH RESTRAINT; THE REFINED SMOKER ASKS PERMISSION BEFORE HE LIGHTS UP; THE GENTLEMAN DANCER IS CONSIDERATE OF HIS PARTNER; THE LADY WHO RIDES SIDESADDLE MAY WEAR GREEN INFORMALLY; THE WELL-REARED CHILD APPRECIATES THE VALUE OF PROPERTY.

"A World for Itself and for Its Silences": Silence and the Nation in The Days of Ziklag

"A World for Itself and for Its Silences": Silence and the Nation in The Days of Ziklag

Speech is Silver, Silence is Golden

Speech is Silver, Silence is Golden

Gothic Silence: S. Alice Callahan's &lt;em&gt;Wynema&lt;/em&gt;, the Battle of the Little Bighorn, and the Indigenous Unspeakable

Gothic Silence: S. Alice Callahan's &lt;em&gt;Wynema&lt;/em&gt;, the Battle of the Little Bighorn, and the Indigenous Unspeakable

Into the Silence: Hemingway, Woolf, and Beckett in the Wake of War

Into the Silence: Hemingway, Woolf, and Beckett in the Wake of War

The Effects of Trust in Supervisor, Work Intensity and Organizational Cynicism on Organizational Silence in Clinical Nurses

본 연구는 임상간호사의 조직침묵에 상사신뢰도, 업무강도와 조직냉소주의가 미치는 영향에 대한 서술적 조사연구이다. 자료수집은 2018년 2월부터 3월 까지 B광역시와 U광역시에 소재한 100병상 이상의 6개 병원에 근무하는 임상간호사 134명을 대상으로 실시하였다. 자료분석은 SPSS 25.0 program을 활용하여 임상간호사...

Silence and the Audibility of the Word: Contemplative Listening as a Fundamental Act of the New Evangelization. Part 3: Christ Reveals Man to Himself on Calvary

Silence and the Audibility of the Word: Contemplative Listening as a Fundamental Act of the New Evangelization. Part 3: Christ Reveals Man to Himself on Calvary

This Splintered Silence by Kayla Olson

This Splintered Silence by Kayla Olson

"Things to be Forgotten": African American Family Histories, Silences, and the American Historical Profession in the Post-emancipation Era

L’intervention de Kendra Field s'inscrivait dans le cadre de ses propres travaux de recherche consacres a la question de la memoire familiale africaine-americaine, a ses modalites de transmission et ses nombreuses implications a la fois au sein de la sphere privee mais aussi a l'interieur du monde universitaire. En effet, Kendra Field soutient que ses observations sur le sujet de l'histoire familiale – qui fut le point de depart de son ouvrage Growing Up with the Country : Family Race, and Na...

Silence and Fog. On Gesture, Time, and Historicity in the Films of Aleksandr Sokurov

In dieser Untersuchung analysiere ich Geste, Ikonografie und Landschaftsaufbau in Aleksandr Sokurovs Filmen, um ihren spezifischen Umgang mit der Zeit und ihre Verpflichtung gegenuber der Historizitat ihrer Bildsprache aufzuzeigen. Mein hermeneutischer Ausgangspunkt ist Aby Warburgs alternative Herangehensweise an die Historizitat von Bildern, vor allem das besondere Augenmerk auf die Montagemechanismen zusammen mit der anthropologischen Bestrebung, eine Geschichte der Gestenubertragung im Westen zu formulieren. Warburgs Projekt kann bestimmte Aspekte der Poetik Sokurovs beleuchten, insbesondere ihre Beziehung zu literarischen, bildlichen und filmischen Traditionen. Die Geste der Stille, das sogenannte signum harpocraticum, spielt eine zentrale Rolle in Sokurovs Filmografie, da sie die Bedingungen einer Medialitat der kinematografischen Geste an sich betont. Die Geste der Stille etabliert die Deixis einer historisch-politischen Offenheit und eines dialektischen Moments, durch welche die Figur des historischen Zeugen Gestalt annimmt. Als Gegenschuss zu diesem direkten Appell an den Zuschauer begunstigen Bilder von Nebel, Sturm und Gewitter nicht nur eine haptische Visualitat, sondern auch eine Darstellung der “Nebel” der Geschichte. In diesen Nebeln versucht Sokurov, eine Erinnerung herauszulocken und sich ihrer schlieslich zu “bemachtigen, wie sie im Augenblick einer Gefahr aufblitzt” (Walter Benjamin). Das Historische, das Politische und das Heilige liegen in Sokurovs Filmen zwischen der Geste der Stille und der Darstellung des Nebels.

Experimental study on inhibiting invasion of inflammatory breast cancer cell lines SUM149 and SUM190 by silencing RhoC gene with the help of RNA interference

Objective In this study, we silence RhoC gene in inflammatory breast cancer (IBC) SUM149, SUM190 cell lines and explore its effects on cell invasion of IBC by RNA interference (RNAi) technology so as to provide experimental principal for target gene therapy of IBC. Methods Human IBC cells were divided blank group, control siRNA group, RhoC-siRNA group by balance randomization. RhoC-siRNA was used to transfect human IBC SUM149, SUM190 cell lines. Microscopy was applied to analyze transfection efficiency. After transfecting for 48 hours, reverse transcriptase-polymerase chain reaction (RT-PCR) was made to detect mRNA expression levels of RhoC. Western blotting was also used to analyze the expression level of RhoC protein. After transfecting for 48 hours, the ability to invasion was analyzed by transwell hole. Results We used siRNA to transfect human IBC SUM149, SUM190 cell lines and found that transfection efficiency can exceed to 80%. After we transfected for 48 hours, the expression levels of RhoC protein in IBC SUM149 cell lines 0.375±0.164 in RhoC siRNA group were significantly lower than those 0.897±0.376 in control siRNA group (t=6.274, P=0.045). The expression levels of RhoC protein in IBC SUM190 cell lines 0.381±0.166 in RhoC siRNA group were significantly lower than those 0.902±0.386 in control siRNA group (t=6.891, P=0.028). However, there were no obvious differences between in blank group and control siRNA group. After we transfected for 48 hours, in IBC SUM149 cell lines, the ability to invasion 85.6±12.7 in RhoC siRNA group was markedly lower than that 340.5±43.4 in control siRNA group (t=12.275, P=0.034). In IBC SUM190 cell lines, the ability to invasion 92.8±15.6 in RhoC siRNA group was markedly lower than that 345.9±47.7 in control siRNA group (t=11.769, P=0.022). Conclusion RhoC is known as an important role in the carcinogenesis of IBC. With the help of RNAi technology, we can greatly inhibit the invasion of IBC SUM149, SUM190 cell lines by silencing RhoC gene. RNAi technology perhaps becomes an important gene therapy tool for IBC. Key words: Inflammatory breast cancer; RhoC; RNA interference; Gene silence