1,078 research outputs found
Immunocytochemical staining of Drosophila larval body-wall muscles
No full textFirst author Preethi Ramachandran is a doctoral student in the Neuroscience Program in the Morningside Graduate School of Biomedical Sciences (GSBS) at UMass Medical School.Over the last two decades, the Drosophila larval neuromuscular junction (NMJ) has gained immense popularity as a model system for the study of synaptic development, function, and plasticity. With this model, it is easy to visualize synapses and manipulate the system genetically with a high degree of temporal and spatial control, which makes it ideal for resolving problems in synaptic physiology and development. This article describes a procedure for labeling various proteins with antibodies in dissected larval body-wall muscles and visualizing their localization and distribution in the brain, NMJ, and muscle.Neuroscienc
Dissection of Drosophila larval body-wall muscles
No full textFirst author Preethi Ramachandran is a doctoral student in the Neuroscience Program in the Morningside Graduate School of Biomedical Sciences (GSBS) at UMass Medical School.Over the last two decades, the Drosophila larval neuromuscular junction has gained immense popularity as a model system for the study of synaptic development, function, and plasticity. With this model, it is easy to visualize synapses and manipulate the system genetically with a high degree of temporal and spatial control, which makes it ideal for resolving problems in synaptic physiology and development. A number of different techniques have been used to dissect third-instar larval preparations to expose the body-wall muscles. Here, we describe a procedure that uses magnetic chambers and pins to allow for fine control in spreading the larval body wall.Neuroscienc
Representation of Women Characters in Preethi Shenoy’s ‘It’s All in the Planets’
No full text \u27It’s all in the Planets\u27 is a charismatic love story written by Preethi Shenoy. It was published in September 2016. The author of the novel, Preethi Shenoy is an Indian author, Speaker and famous blogger. In this paper, I am analysing this novel from the perspective of feminism, especially how the author presented her female characters. Feminism originated as an offshoot of the women\u27s suffrage movement. The major feminist theorist is Toril Moi, Simone de Beauvoir, Elaine Showalter etc. It is a movement against men’s ideologies and society’s certain representations. Their main concern was the representations of women in literature. Traditionally females are represented in literature as passive characters. Though the feminists broke all such representations, our literary works do not completely get rid of such representations. Many of the authors still present women as passive in their works. In this novel too there are such representations. Superficially it is a love story between two persons who are living in two different worlds. They have their tastes, opinions and attitudes. Even they have two life partners (living relationships). But ultimately fate joins them
A new species of the pseudoscorpion genus Megachernes (Pseudoscorpiones: Chernetidae) associated with a threatened Sri Lankan rainforest rodent, with a review of host associations of Megachernes
No full textHarvey, Mark S., Ratnaweera, Pamoda B., Udagama, Preethi V., Wijesinghe, Mayuri R. (2012): A new species of the pseudoscorpion genus Megachernes (Pseudoscorpiones: Chernetidae) associated with a threatened Sri Lankan rainforest rodent, with a review of host associations of Megachernes. Journal of Natural History (J. Nat. Hist.) 46 (41-42): 2519-2535, DOI: 10.1080/00222933.2012.707251, URL: http://dx.doi.org/10.1080/00222933.2012.70725
사람의 침윤성 암세포에서 후성학적 BTG2/TIS21/PC3 발현 조절과 암 억제 기전 연구
No full textB cell translocation gene 2 (BTG2/TIS21/PC3) belongs to the family of antiproliferative (APRO) genes and reported as a tumor suppressor by our group and others. Expression of BTG2 is significantly reduced in cancers developed in various organs and tissues. EJ (bladder carcinoma cells), MKN-1 (gastric cancer cells) are highly invasive and metastatic cells with very less endogenous BTG2 expression due to epigenetic regulation. Significantly lower endogenous expression of BTG2 was observed in human muscle-invasive bladder cancers (MIBC) than matched normal tissues and non-muscle invasive bladder cancers (NMIBC). BTG2 expression was inversely correlated with increased expression of the DNA methyltransferases DNMT1 and DNMT3a in MIBC, but not NMIBC, suggesting a potential role for BTG2 expression in muscle invasion of bladder cancer. Over 90% of tumor tissues revealed strong methylation at CpG islands of the BTG2 gene, compared with no methylation in the normal tissues, implying epigenetic regulation of BTG2 expression in bladder carcinogenesis. BTG2 is constitutively expressed in mucous epithelium and parietal cells of gastric glands in stomach, and the expression was increased in mucous epithelium with H. pylori infection as opposed to loss in human gastric adenocarcinoma. Indeed, adenoviral transduction of BTG2 significantly inhibited Tipα activity in MKN-1 and MGT-40, human and mouse gastric cancer cells, respectively, thereby downregulated TNFα expression and Erk1/2 phosphorylation via reducing nucleolin, Tipα receptor, expression. Chromatin immunoprecipitation proved that BTG2 inhibited Sp1 expression and it’s binding to the promoter of nucleolin gene. In addition, BTG2 expression significantly reduced membrane localized nucleolin expression in cancer cells and the loss of BTG2/TIS21 expression rather induced cytoplasmic nucleolin availability in gastric cancer tissues, evidenced by immunoblot and immunohistochemistry. The higher expression of BTG2 and the lower nucleolin expression accompanied with the better overall survival of the poorly differentiated gastric cancer patients.B cell translocation gene 2(BTG2/TIS21/PC3)는 antiproliferative gene(APRO) 계열에 속하며 본 연구팀을 포함한 많은 연구자들에 의해 BTG2 의 암억제 기능이 연구되어 왔다. BTG2 는 여러 조직과 기관에서 발생한 암에서 그 발현이 감소되어 있고, 특히 침윤(invasion)과 전이(metastasis)를 잘하는 암세포인 EJ (bladder carcinoma cells)와 MKN-1 (gastric cancer cells)에서도 후성유전학적(epigenetic regulation)으로 BTG2 발현이 매우 감소되어 있다. 최근 사람에서, 이런 침윤성과 전이성이 높은 암에서 BTG2 가 침윤을 억제하고 침윤을 조장하는 단백질과 mRNA 발현과는 음의 상관도를 보인다는 보고가 많기에, 본 저자는 BTG2 가 어떤 기전으로 침윤성 암을 조절하는지를 중점적으로 연구하였다. 사람의 근육침범 방광암(muscleinvasive bladder cancers, MIBC)에서 비-근육침범 방광암(non-muscle invasive bladder cancers, NMIBC)보다 BTG2 의 발현이 현저히 감소되어 있다는 것을 발견하였다. 이런 MIBC 에서의 BTG2 의 감소는 반대로 DNA 메틸기전달효소 DNMT1 과 DNMT3a 의 증가와 연관이 있었고, 이런 음의 상관관계는 NMIBC 에서는 발견되지 않음으로써 BTG2 이 방광암의 근육 침범에 분명한 역할을 하고 있다는 가능성을 보였다. 방광암 조직들의 90%에서 BTG2 유전자의 CpG 섬에 강한 메틸화를 보였고, 반대로 정상 방광조직에서는 메틸화가 발견되지 않음으로써, 방광암발생과정에서 BTG2 이 후성학적 조절을 받는다는 사실도 증명했다. 한편, BTG2 는 위 점막의 상피세포와 위샘의 벽세포(Parietal cell)에서 기본적으로 발현되고 있으며, 이 유전자의 발현은 H. pylori 감염시 위 점막 상피세포에서 증가되고, 반대로 위선암(gastric adenocarcinoma)에서는 감소됨을 발견했다. 사람 암세포 MKN-1 과 쥐 암세포 MGT-40 에서 BTG2 를 adenoviral transduction 을 통해 발현시키면 Tumor necrosis factor- α inducing protein (Tip α ) 활성이 억제되었고, 이는 Tip α receptor 인 nucleolin 을 감소시킴으로 TNFα의 발현과 Erk1/2 의 인산화를 억제시켰다. BTG2 는 Sp1 의 발현과 또한 염색질 면역침전(Chromatin immunoprecipitation) 실험을 통해 nucleolin 의 promoter 에 결합하는 Sp1 도 감소시킴을 확인했다. 또한 암세포에서 BTG2 를 발현시키면 세포막에 위치한 nucleolin 의 발현이 감소되었고, 반대로 BTG2 발현을 감소시키면 세포질 내에 nucleolin 의 발현이 증가함을 면역조직화학법과 면역블롯검사를 통해 위암조직에서 밝혔다. 이런 BTG2 의 강한 발현과 nuclolin 의 발현 저하는 저분화 위암 환자에서의 높은 생존율(overall survival)과 연관이 있었다. 더욱이 이런 BTG2 의 발현증가는 침윤성 암세포에서도 암 진행과 성장을 억제했다. 따라서, 본 저자는 본 연구를 통해 BTG2 유전자가 침윤성 암 환자를 위한 강력한 치료로 사용될 수 있으리라 제안한다.ABSTRACT i
TABLE OF CONTENTS iii
LIST OF FIGURES vi
ABBREVIATIONS viii
I. INTRODUCTION 1
II. EXPERIMENTAL METHODS 7
A. Tissues and cell cultures 7
B. RNA isolation and reverse transcription 8
C. Real-time and RT-PCR analyses 8
D. Methylation-specific PCR (MSP) and unmethylation-specific PCR (USP) analyses 9
E. Western blotting 9
F. DNMT activity assay 10
G. Chromatin immunoprecipitation (ChIP) analysis 10
H. Cloning of CpG islands and sequencing analyses 10
I. Immunoprecipitation/immunoblot analysis 11
J. Regulation of gene expression 11
K. Tumorigenesis study 12
L. Invasion assay 13
M. Immunohistochemistry analysis 14
N. Preparation of recombinant Tipα protein 15
O. Subcellular fractionation 15
P. Statistical analysis 15
III. RESULTS 16
A. Downregulation of BTG2 expression in human MIBC by DNA methylation of BTG2 gene 16
B. Inverse correlations between the expressions of BTG2 vs DNMT1 and DNMT3a in MIBC 18
C. Upregulation of BTG2 expression in EJ bladder cancer cells upon Decitabine treatment 22
D. Chromatin remodeling at the promoter and intron of BTG2 gene after decitabine treatment 27
E. Sp1, the transcription factor, for BTG2 gene upon decitabine treatment 28
F. Induction of BTG2 expression by knockdown of DNMT1 in EJ cells 32
G. Downregulation of tumorigenesis and cell cycle arrest by BTG2 overexpression 35
H. Inhibition of tumor invasion by BTG2 via downregulation of DNMT1
expression 38
I. Direct effect of BTG2/TIS21 expression on the inhibition of cancer invasiveness 41
J. Immunohistochemical findings 44
K. Induction of BTG2 expression by decitabine treatment in various cancer cell lines in addition to APRO gene expression analyses 46
L. Expression of BTG2/TIS21 is increased in mucous epithelium infected with H. pylori, but lost in human gastric adenocarcinoma 48
M. Absence of endogenous BTG2/TIS21 expression in the gastric cancer cells 51
N. Expression of BTG2/TIS21 is epigenetically regulated in gastric adenocarcinoma 52
O. Inhibition of cancer cell proliferation by BTG2/TIS21 gene 54
P. Inhibition of Tipα activity by BTG2/TIS21 in human gastric cancer cells 55
Q. Inhibition of Tipα activity by downregulation p-ERK1/2 in human gastric cancer cells 57
R. Downregulation of nucleolin expression by BTG2/TIS21 59
S. Expression of nucleolin, Tipα receptor, was reduced by BTG2 expression via inhibiting Sp1binding to the nucleolin promoter 61
T. Inverse regulation of Tipα -induced TNFα expression by NCL and BTG2 64
U. Reciprocal expression of BTG2/TIS21 and nucleolin expressions in normal and cancer regions 68
V. Inverse regulation of overall survival of gastric cancer patients by BTG2/TIS21 and NCL genes 71
IV. DISCUSSION 74
V. SUMMARY 80
VI. CONCLUSIONS 82
VII. REFERENCES 83
국문요약 95Docto
Stabilisation limits of turbulent premixed flames by nanosecond repetitively pulsed discharges
No full textThe mechanism of flame stabilisation using nanosecond repetitively pulsed (NRP) plasma discharges in a turbulent, premixed methane–air flame at high velocities was investigated, focusing on the lean extinction limits. In contrast to the majority of the existing studies that considered NRP discharges as an assistance to conventional stabilisers, here, no other flame-holding method is used. High-speed (10 kHz) OH* chemiluminescence showed that the plasma discharges produce individual OH*-pockets that merge together at a high enough frequency to form a continuous flame sheet. Increasing the discharge repetition frequency from 5 kHz to 10 kHz improves flame stability, but no change in flame structure and stability was observed when the frequency was increased beyond 10 kHz. Change in the plasma energy level in the range studied had little effect on the flame structure. The lean extinction limit was quantified at various flow velocities, equivalence ratios, discharge frequencies and energy levels. It was observed that the trend of the extinction equivalence ratio with bulk velocity was similar to that of a conventional bluff body stabilised flame and that plasma-only stabilised flame was equally effective at certain operating conditions. An effort to correlate the stabilisation limits by a conventional Damköhler number Da* was made but was not satisfactory due to the presence of a characteristic flameholder lengthscale present in the Da* expression. A modified Da* was proposed to take the spark frequency effects into account, but this was not successful either. In contrast, the spread of the extinction data was smaller when a critical Karlovitz number was used, hence offering a way to extrapolate the present data to other conditions. The experiments demonstrate that NRP discharges can be used as an alternative stabilisation method for high-speed turbulent premixed flames
sj-docx-1-tag-10.1177_17562848221141501 – Supplemental material for Impact of recurrent hospitalization for Clostridioides difficile on longitudinal outcomes in patients with inflammatory bowel diseases: a nationally representative cohort
No full textSupplemental material, sj-docx-1-tag-10.1177_17562848221141501 for Impact of recurrent hospitalization for Clostridioides difficile on longitudinal outcomes in patients with inflammatory bowel diseases: a nationally representative cohort by Preethi G. Venkat, Nghia H. Nguyen, Jiyu Luo, Alexander S. Qian, Sahil Khanna and Siddharth Singh in Therapeutic Advances in Gastroenterology</p
sj-docx-3-tag-10.1177_17562848221141501 – Supplemental material for Impact of recurrent hospitalization for Clostridioides difficile on longitudinal outcomes in patients with inflammatory bowel diseases: a nationally representative cohort
No full textSupplemental material, sj-docx-3-tag-10.1177_17562848221141501 for Impact of recurrent hospitalization for Clostridioides difficile on longitudinal outcomes in patients with inflammatory bowel diseases: a nationally representative cohort by Preethi G. Venkat, Nghia H. Nguyen, Jiyu Luo, Alexander S. Qian, Sahil Khanna and Siddharth Singh in Therapeutic Advances in Gastroenterology</p
sj-docx-2-tag-10.1177_17562848221141501 – Supplemental material for Impact of recurrent hospitalization for Clostridioides difficile on longitudinal outcomes in patients with inflammatory bowel diseases: a nationally representative cohort
No full textSupplemental material, sj-docx-2-tag-10.1177_17562848221141501 for Impact of recurrent hospitalization for Clostridioides difficile on longitudinal outcomes in patients with inflammatory bowel diseases: a nationally representative cohort by Preethi G. Venkat, Nghia H. Nguyen, Jiyu Luo, Alexander S. Qian, Sahil Khanna and Siddharth Singh in Therapeutic Advances in Gastroenterology</p
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