Search results for: expression analysis

Authors: Ricardo Alberto Chiong Zevallos, Eduardo Moraes Rego Reis

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Pancreatic adenocarcinoma (PDAC) patients have a less than 10% 5-year survival rate. PDAC has no defined diagnostic and prognostic biomarkers. Gemcitabine is the first-line drug in PDAC and several other cancers. Long non-coding RNAs (lncRNAs) contribute to the tumorigenesis and are potential biomarkers for PDAC. Although lncRNAs aren’t translated into proteins, they have important functions. LncRNAs can decoy or recruit proteins from the epigenetic machinery, act as microRNA sponges, participate in protein translocation through different cellular compartments, and even promote chemoresistance. The chromatin remodeling enzyme EZH2 is a histone methyltransferase that catalyzes the methylation of histone 3 at lysine 27, silencing local expression. EZH2 is ambivalent, it can also activate gene expression independently of its histone methyltransferase activity. EZH2 is overexpressed in several cancers and interacts with lncRNAs, being recruited to a specific locus. EZH2 can be recruited to activate an oncogene or silence a tumor suppressor. The lncRNAs misregulation in cancer can result in the differential recruitment of EZH2 and in a distinct epigenetic landscape, promoting chemoresistance. The relevance of the EZH2-lncRNAs interaction to chemoresistant PDAC was assessed by Real Time quantitative PCR (RT-qPCR) and RNA Immunoprecipitation (RIP) experiments with naïve and gemcitabine-resistant PDAC cells. The expression of several lncRNAs and EZH2 gene targets was evaluated contrasting naïve and resistant cells. Selection of candidate genes was made by bioinformatic analysis and literature curation. Indeed, the resistant cell line showed higher expression of chemoresistant-associated lncRNAs and protein coding genes. RIP detected lncRNAs interacting with EZH2 with varying intensity levels in the cell lines. During RIP, the nuclear fraction of the cells was incubated with an antibody for EZH2 and with magnetic beads. The RNA precipitated with the beads-antibody-EZH2 complex was isolated and reverse transcribed. The presence of candidate lncRNAs was detected by RT-qPCR, and the enrichment was calculated relative to INPUT (total lysate control sample collected before RIP). The enrichment levels varied across the several lncRNAs and cell lines. The EZH2-lncRNA interaction might be responsible for the regulation of chemoresistance-associated genes in multiple cancers. The relevance of the lncRNA-EZH2 interaction to PDAC was assessed by siRNA knockdown of a lncRNA, followed by the analysis of the EZH2 target expression by RT-qPCR. The chromatin immunoprecipitation (ChIP) of EZH2 and H3K27me3 followed by RT-qPCR with primers for EZH2 targets also assess the specificity of the EZH2 recruitment by the lncRNA. This is the first report of the interaction of EZH2 and lncRNAs HOTTIP and PVT1 in chemoresistant PDAC. HOTTIP and PVT1 were described as promoting chemoresistance in several cancers, but the role of EZH2 is not clarified. For the first time, the lncRNA LINC01133 was detected in a chemoresistant cancer. The interaction of EZH2 with LINC02577, LINC00920, LINC00941, and LINC01559 have never been reported in any context. The novel lncRNAs-EZH2 interactions regulate chemoresistant-associated genes in PDAC and might be relevant to other cancers. Therapies targeting EZH2 alone weren’t successful, and a combinatorial approach also targeting the lncRNAs interacting with it might be key to overcome chemoresistance in several cancers.

Keywords: epigenetics, chemoresistance, long non-coding RNAs, pancreatic cancer, histone modification

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Authors: Soumya S., Manju Nidagodu Jayakumar, Vellore Kannan Gopinath

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Dental pulp inflammation resulting from dental caries often leads to a pathologic condition known as irreversible pulpitis and the currently managed by root canal treatment. Extirpation of the entire pulp tissue is done during this procedure, and the canal space is filled with synthetic materials. Recent studies in the stem cell biology state that some portion of the irreversibly inflamed pulp tissue could be viable with progenitor cells, having the properties similar to that of Mesenchymal stem cells. Hence, we aim to isolate Dental Pulp Stem Cells (DPSCs) from patients diagnosed with severe irreversible pulpitis and characterize the cells for the MSC specific markers. The pulp tissue was collected from the dental clinic and subjected to collagenase/dispase digestion. The isolated cells were expanded in culture, and the phenotypic characterization was done using flow cytometry. MSC specific markers such as CD-90, CD-73, and CD-105 were analysed along with negative markers such as CD-14 and CD-45. The isolated cells expressed positive expression for CD markers with CD90 and CD105 ( > 95%) and CD73 (19%). The cells did not express the negative markers CD-14 and CD-45. The commercially available DPSCs from vital extracted teeth, preferably molar/wisdom teeth with large pulp cavity or incomplete root growth in young patients (aged 15-30 years) showed more than 90% expression for all the CD markers such as CD-90, 73 and 105, whereas negative for CD-14 and CD-45. The DPSCs isolated from inflamed pulp tissue showed a less expression for CD-73 compared to the commercially available DPSCs whereas, as the other two markers were found to show similar percentage of positive expression. This could be attributed to the fact that the pulp population is very heterogeneous and we used the pooled tissue from different patients. Hence the phenotypic characterization and comparison with the commercially available DPSCs proved that the inflamed pulp tissue is a good source of MSC like cells which can be utilized further for regenerative application.

Keywords: collagenase/dispase, dental pulp stem cells, flow cytometry, irreversible pulpitis

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Authors: Erfan Rahimi, Hadi Bahari Far, Mojgan Shikhpour

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Background: Urinary tract infection is one of the most common nosocomial infections, especially among women. E. coli is one of the main causes of urinary tract infections and one of the most common antibiotics to fight this bacterium is ampicillin. As conventional antibiotics led to bacterial antibiotic resistance, modification of the pure drugs can address this issue. The aim of this study was to prepare nanofluids containing carbon nanotubes conjugated with ampicillin to improve drug performance and reduce antibiotic resistance. Methods: Multi-walled carbon nanotubes (MWCNTs) were activated with thionyl chloride by reflux system and nanofluids containing antibiotics were prepared by ultrasonic method. The properties of the prepared nano-drug were investigated by general element analysis, infrared spectroscopy, Raman spectroscopy, scanning electron microscopy and transmission electron microscopy. After the treatment of the desired strain with nanofluid, microbial studies were performed to evaluate the antibacterial effects and molecular studies were carried out to measure the expression of the resistance gene AcrAB. Result: We have shown that the antimicrobial effect of ampicillin-functionalized MWCNTs at low concentrations performed better than that of the conventional drug in both resistant and ATCC strains. Also, a decrease in antibiotic resistance of bacteria treated with ampicillin-functionalized MWCNTs compared to the pure drug was observed. Also, ampicillin-functionalized MWCNTs downregulated the expression of AcrAB in treated bacteria. Conclusion: Because carbon nanotubes are capable of destroying the bacterial wall, which provides antibiotic resistance features in bacteria, their usage in the form of nanofluids can make lower dosages (about three times less) than that of the pure drug more effective. Additionally, the expression of the bacterial resistance gene AcrAB decreased, thereby reducing antibiotic resistance and improving drug performance against bacteria.

Keywords: urinary tract infection, antibiotic resistance, carbon nanotube, nanofluid

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Authors: Cheng-Cao Sun, Shu-Jun Li, Cuili Yang, Yongyong Xi, Liang Wang, Feng Zhang, De-Jia Li

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MicroRNAs (miRNAs) act as key regulators of multiple cancers. Hsa-miR-329 (miR-329) functions as a tumor suppressor in some malignancies. However, its role on lung cancer remains poorly understood. In this study, we investigated the role of miR-329 on the development of lung cancer. The results indicated that miR-329 was decreased in primary lung cancer tissues compared with matched adjacent normal lung tissues and very low levels were found in a non-small cell lung cancer (NSCLC) cell lines. Ectopic expression of miR-329 in lung cancer cell lines substantially repressed cell growth as evidenced by cell viability assay, colony formation assay and BrdU staining, through inhibiting cyclin D1, cyclin D2, and up-regulatiing p57(Kip2) and p21(WAF1/CIP1). In addition, miR-329 promoted NSCLC cell apoptosis, as indicated by up-regulation of key apoptosis gene cleaved caspase-3, and down-regulation of anti-apoptosis gene Bcl2. Moreover, miR-329 inhibited cellular migration and invasiveness through inhibiting matrix metalloproteinases (MMP)-7 and MMP-9. Further, oncogene MET was revealed to be a putative target of miR-329, which was inversely correlated with miR-329 expression. Furthermore, down-regulation of MET by siRNA performed similar effects to over-expression of miR-329. Collectively, our results demonstrated that miR-329 played a pivotal role in lung cancer through inhibiting cell proliferation, migration, invasion, and promoting apoptosis by targeting oncogenic MET.

Keywords: hsa-miRNA-329(miR-329), MET, non-small cell lung cancer (NSCLC), proliferation, apoptosis

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Authors: Leila Rashki Ghaleno, Mohamad Amin Hajari, Leila Montazeri, Abdolhossein Shahverdi, Mojtaba Rezazadeh Valojerdi

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Background: Spermatogonia stem cells (SSCs) exhibit pluripotency, enabling them to undergo differentiation into many cell lineages, including neurons, glia, endothelial cells, and hepatocytes when cultured in vitro. Although the specific mechanisms are not yet fully understood, it has been observed that biopolymer agents, such as hyaluronic acid (HA) and alginate (Alg), have the potential to induce transdifferentiation of SSCs. The current work aimed to examine the process of in vitro spermatogenesis and the conversion of mouse testicular cells into hepatocytes and erythrocyte-like cells utilizing the HA-Alg hydrogel. Method: After being extracted from the testes of a 5-day postpartum mouse (5 DPP), the testicular cells were separated into two enzymatic stages and then put into a composite hydrogel containing 0.5% HA and 1% alginate. On days 14 and 28 of culture, the colonies' growth, the cells' viability, and their histology were assessed. Result: Despite observing significant cell proliferation on day 14 and the development of circular-shaped organoids on day 28, it was noted that the organoids generated in the HA-Alg medium tended to maintain their circular morphology on day 28. Notably, the testicular cells underwent transdifferentiation into cell types resembling erythrocytes and hepatocytes. The hepatocyte-like cells exhibited the presence of glycogen and lipid deposits, indicating their hepatocyte-like characteristics. Interestingly, immunostaining analysis revealed the secretion of albumin and the presence of VEGFR on day 14. However, on day 28, albumin expression was not detected, while the expression of Sox9 (a marker for hepatocytes), Vegf, CD34, and C-kit (markers for erythrocytes) showed increased levels in the gene expression evaluation. Conclusion: The present findings indicated that HA-Alg could be a potent and effective agent for the transdifferentiation of testis cells into erythrocyte and hepatocyte-like cells, as recent studies have confirmed the transformation of SSCs into hepatocyte cells during in vitro culture.

Keywords: 3D culture, mouse testicular cell, hyaluronic acid, liver organoids

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Authors: Shangqing Song, Bin Xu, Yajun Cheng, Zhong Wang

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miRNAs derived from exosomes exist in a body fluid such as urine were regarded as potential biomarkers for various human cancers diagnosis and prognosis, as mature miRNAs can be steadily preserved by exosomes. However, its potential value in clear cell renal cell carcinoma (ccRCC) diagnosis and prognosis remains unclear. In the present study, differentially expressed miRNAs from urinal exosomes were identified by next-generation sequencing (NGS) technology. The 16 differentially expressed miRNAs were identified between ccRCC patients and healthy donors. To explore the specific diagnosis biomarker of ccRCC, we validated these urinary exosomes from 70 early-stage renal cancer patients, 30 healthy people and other urinary system cancers, including 30 early-stage prostate cancer patients and 30 early-stage bladder cancer patients by qRT-PCR. The results showed that urinary exosome miR-30c-5p could be stably amplified and meanwhile the expression of miR-30c-5p has no significant difference between other urinary system cancers and healthy control, however, expression level of miR-30c-5p in urinary exosomal of ccRCC patients was lower than healthy people and receiver operation characterization (ROC) curve showed that the area under the curve (AUC) values was 0.8192 (95% confidence interval was 0.7388-0.8996, P= 0.0000). In addition, up-regulating miR-30c-5p expression could inhibit renal cell carcinoma cells growth. Lastly, HSP5A was found as a direct target gene of miR-30c-5p. HSP5A depletion reversed the promoting effect of ccRCC growth casued by miR-30c-5p inhibitor, respectively. In conclusion, this study demonstrated that urinary exosomal miR-30c-5p is readily accessible as diagnosis biomarker of early-stage ccRCC, and miR-30c-5p might modulate the expression of HSPA5, which correlated with the progression of ccRCC.

Keywords: clear cell renal cell carcinoma, exosome, HSP5A, miR-30c-5p

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Authors: P. Narrima, C. Y. Looi, M. A. Mohd, H. M. Ali

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Persea declinata (Bl.) Kosterm is a member of the Lauraceae family, widely distributed in Southeast Asia. It is from the same genus with avocado (Persea americana Mill), which is widely consumed as food and for medicinal purposes. In the present study, we examined the anticancer properties of Persea declinata (Bl.) Kosterm bark methanolic crude extract (PDM). PDM exhibited a potent antiproliferative effect in MCF-7 human breast cancer cells, with an IC50 value of 16.68 µg/mL after 48h of treatment. We observed that PDM caused cell cycle arrest and subsequent apoptosis in MCF-7 cells, as exhibited by increased population at G0/G1 phase, higher lactate dehydrogenase (LDH) release, and DNA fragmentation. Mechanistic studies showed that PDM caused significant elevation in ROS production, leading to perturbation of mitochondrial membrane potential, cell permeability, and activation of caspases-3/7. On the other hand, real-time PCR and Western blot analysis showed that PDM treatment increased the expression of the proapoptotic molecule, Bax, but decreased the expression of prosurvival proteins, Bcl-2 and Bcl-xL, in a dose-dependent manner. These findings imply that PDM could inhibit proliferation in MCF-7 cells via cell cycle arrest and apoptosis induction, indicating its potential as a therapeutic agent worthy of further development.

Keywords: antiproliferative, apoptosis, MCF-7 human breast cancer, Persea declinata

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Authors: Nandhana Vivek, Bhaskar Gogoi, Ayyavu Mahesh

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Breast cancer metastasis plays a key role in cancer progression and fatality. The present study examines the potential causes of metastasis in breast cancer by investigating the novel interactions between genes and their pathways. The gene expression profile of GSE99394, GSE1246464, and GSE103865 was downloaded from the GEO data repository to analyze the differentially expressed genes (DEGs). Protein-protein interactions, target factor interactions, pathways and gene relationships, and functional enrichment networks were investigated. The proliferation pathway was shown to be highly expressed in breast cancer progression and metastasis in all three datasets. Gene Ontology analysis revealed 11 DEGs as gene targets to control breast cancer metastasis: LYN, DLGAP5, CXCR4, CDC6, NANOG, IFI30, TXP2, AGTR1, MKI67, and FTH1. Upon studying the function, genomic and proteomic data, and pathway involvement of the target genes, DLGAP5 proved to be a promising candidate due to it being highly differentially expressed in all datasets. The study takes a unique perspective on the avenues through which DLGAP5 promotes metastasis. The current investigation helps pave the way in understanding the role DLGAP5 plays in metastasis, which leads to an increased incidence of death among breast cancer patients.

Keywords: genomics, metastasis, microarray, cancer

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Authors: Valizadeh Gever Bita, Joel Caren, Louisa Huand, Yu-Cheng Zhu, Esmaeil Amiri

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Within a honey bee colony, queen is the sole reproducer of fertilized eggs, while queens are safeguarded by worker bees, trophallactic behavior and food sharing activities could expose them to agrochemicals. Here, we assessed the effects of three widely used pesticides—Acephate, Bifenthrin, and Chlorantraniliprole— on worker bees, to investigate indirect effects on the physiology and reproductive traits of queens as well as the eggs they produce. Using RT-qPCR we measured the expression of several detoxification and immune genes in adult worker bees, queens, and freshly laid eggs after pesticide exposure. These analyses aimed to elucidate the physiological changes in queens and potential transgenerational effects. While no significant changes in reproductive traits were observed following Chlorantraniliprole and Bifenthrin exposure, Acephate caused adverse effects on egg size, egg-laying activity, and queen weight. The expression of detoxification, immune and antioxidant-related genes in workers, queens and freshly laid eggs changed over time in response to these pesticides. The results of this investigation revealed that pesticides can cause negative impact on queen physiology and reproduction indirectly through their effects on exposed worker bees. These effects can potentially extend to the next generation of honey bees.

Keywords: apis mellifera, egg laying, detoxification enzymes, gene expression, honey bee queen

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Authors: Vanessa Funk, Steven Blum, Stephanie Cole, Jorge Maciel, Matthew Lux

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Paper-based synthetic gene circuits offer a new paradigm for programmable, fieldable biodetection. We demonstrate that by freeze-drying gene circuits with in vitro expression machinery, we can use complimentary RNA sequences to trigger colorimetric changes upon rehydration. We have successfully utilized both green fluorescent protein and luciferase-based reporters for easy visualization purposes in solution. Through several efforts, we are aiming to use this new platform technology to address a variety of needs in portable detection by demonstrating several more expression and reporter systems for detection functions on paper. In addition to RNA-based biodetection, we are exploring the use of various mechanisms that cells use to respond to environmental conditions to move towards all-hazards detection. Examples include explosives, heavy metals for water quality, and toxic chemicals.

Keywords: cell-free lysates, detection, gene circuits, in vitro

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Authors: Mukul Sharma, Madhusmita Das, Sundeep Chaitanya Vedithi

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Leprosy is a chronic human disease caused by Mycobacterium leprae, that cannot be cultured in vitro. Though treatable with multidrug therapy (MDT), recently, bacteria reported resistance to multiple antibiotics. Targeting DNA replication and repair pathways can serve as the foundation of developing new anti-leprosy drugs. Due to the absence of an axenic culture medium for the propagation of M. leprae, studying cellular processes, especially those belonging to DNA repair pathways, is challenging. Genomic understanding of M. Leprae harbors several protein-coding genes with no previously assigned function known as 'hypothetical proteins'. Here, we report identification and expression of known and hypothetical DNA repair genes from a human skin biopsy and mouse footpads that are involved in base excision repair, direct reversal repair, and SOS response. Initially, a bioinformatics approach was employed based on sequence similarity, identification of known protein domains to screen the hypothetical proteins in the genome of M. leprae, that are potentially related to DNA repair mechanisms. Before testing on clinical samples, pure stocks of bacterial reference DNA of M. leprae (NHDP63 strain) was used to construct standard graphs to validate and identify lower detection limit in the qPCR experiments. Primers were designed to amplify the respective transcripts, and PCR products of the predicted size were obtained. Later, excisional skin biopsies of newly diagnosed untreated, treated, and drug resistance leprosy cases from SIHR & LC hospital, Vellore, India were taken for the extraction of RNA. To determine the presence of the predicted transcripts, cDNA was generated from M. leprae mRNA isolated from clinically confirmed leprosy skin biopsy specimen across all the study groups. Melting curve analysis was performed to determine the integrity of the amplification and to rule out primer‑dimer formation. The Ct values obtained from qPCR were fitted to standard curve to determine transcript copy number. Same procedure was applied for M. leprae extracted after processing a footpad of nude mice of drug sensitive and drug resistant strains. 16S rRNA was used as positive control. Of all the 16 genes involved in BER, DR, and SOS, differential expression pattern of the genes was observed in terms of Ct values when compared to human samples; this was because of the different host and its immune response. However, no drastic variation in gene expression levels was observed in human samples except the nth gene. The higher expression of nth gene could be because of the mutations that may be associated with sequence diversity and drug resistance which suggests an important role in the repair mechanism and remains to be explored. In both human and mouse samples, SOS system – lexA and RecA, and BER genes AlkB and Ogt were expressing efficiently to deal with possible DNA damage. Together, the results of the present study suggest that DNA repair genes are constitutively expressed and may provide a reference for molecular diagnosis, therapeutic target selection, determination of treatment and prognostic judgment in M. leprae pathogenesis.

Keywords: DNA repair, human biopsy, hypothetical proteins, mouse footpads, Mycobacterium leprae, qPCR

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Authors: Hala Ali

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In the evolving landscape of contemporary art, the value of failure and experimentation plays a central role in reshaping artistic research. This paper explores an artistic residency where the focus shifted from traditional practices of ink on canvas to performance art using the human body as a medium of expression. This residency emphasized uncertainty, experimentation, and emotional expression as the core of the process. Through collaboration between a calligrapher and a visual artist, the performance engaged themes of seduction, silence, and the transition between reality and abstraction. In alignment with experimental art practices, the process itself became the artwork, embracing moments of failure and disruption as key components of creative exploration. This research integrates theories from neuroscience, psychology, and artistic failure, drawing on the insights of thinkers like John Cage, Samuel Beckett, and Cornelius Cardew to further contextualize the residency’s impact.

Keywords: embodied art, emotional communication, mindfulness in art, nonverbal communication, performance art

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Authors: Sigal Barkai

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'Ironic Historiography' is a hybrid notion combining criticism of historical narratives concerning the Israeli state with ironic artistic expression. The paper will deal with questions of identities of native Israeli visual artists who chose to live out of the country, or to move back and forth to and from it. It will examine the ways these wanderings are reflected in their work. The paper discusses the work of 4 contemporary artists who produce artworks in diverse techniques and media, such as video, performance and installation art. Yael Bartana, Erez Israeli and Tamir Zadok are artists who constantly deal with Israeli nationality and history in their artwork, using ironic components. In comparison, the paper will review the works of Mika Rottenberg, who is now a New York based artist. She is concerned with global social issues and neglected specific national identity altogether. All of them use visual irony as a means of reflecting and criticizing society. The analysis was done in awareness of the life stories of the artists, in an attempt to trace the ways they establish their identities through their art. It was pre-supposed that these identities will be shaped in the in-between space of being an Israeli citizen and a citizen of the world. The study asks how ironic expression appears in their work, what kind of irony do they use and in what ways does it serves them. The methodology combined visual analysis, interviews with the artists, and analyzation of secondary discourses in the media. As theoretical background various fields of knowledge were used, such as literature and language studies, Sociology, and Visual Culture studies. The findings point out that visual and artistic irony has many different goals in the use of historiographic fiction. It can bind an artist to his homeland and native society, or it can help her to detach. It helps healing breaches in the in-between space, or it can be used as a means to completely detach from any identification with a native origin.

Keywords: visual art, irony, identities, Israel

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Authors: Xinxiu Li, Chuqian Zheng, Yanyan Qian, Hong Fan

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Objectives: In hepatocellular carcinoma (HCC), oncogenes are continuously and robustly transcribed due to aberrant expression of essential components of the trans-acting super-enhancers (SE) complex. Preclinical and clinical trials are now being conducted on small-molecule inhibitors that target core-transcriptional components, including as transcriptional bromodomain protein 4 (BRD4) and cyclin-dependent kinase 7 (CDK7), in a number of malignant tumors. This study aims to explore whether co-overexpression of BRD4 and CDK7 is a potential marker of worse prognosis and a combined therapeutic target in HCC. Methods: The expression pattern of BRD4 and CDK7 and their correlation with prognosis in HCC were analyzed by RNA sequencing data and survival data of HCC patients from TCGA and GEO datasets. The protein levels of BRD4 and CDK7 were determined by immunohistochemistry (IHC), and survival data of patients were analyzed using the Kaplan-Meier method. The mRNA expression levels of genes in HCC cell lines were evaluated by quantitative PCR (q-PCR). CCK-8 and colony formation assays were conducted to assess cell proliferation of HCC upon treatment with BRD4 inhibitor JQ1 or/and CDK7 inhibitor THZ1. Results: It was shown that BRD4 and CDK7 were often overexpressed in HCCs and were associated with poor prognosis of HCC by analyzing the TCGA and GEO datasets. BRD4 or CDK7 overexpression was related to a lower survival rate. It's interesting to note that co-overexpression of CDK7 and BRD4 was a worse prognostic factor in HCC. Treatment with JQ1 or THZ1 alone had an inhibitory effect on cell proliferation; however, when JQ1 and THZ1 were combined, there was a more notable suppression of cell growth. At the same time, the combined use of JQ1 and THZ1 synergistically suppresses the expression of HCC driver genes. Conclusion: Our research revealed that BRD4 and CDK7 coupled can be a useful biomarker in HCC prognosis and the combination of JQ1 and THZ1 can be a promising therapeutic therapy against HCC.

Keywords: BRD4, CDK7, cell proliferation, combined inhibition

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Authors: Charalabos Antonatos, Mariza Panoutsopoulou, Georgios K. Georgakilas, Evangelos Evangelou, Yiannis Vasilopoulos

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The extended tissue damage and severe clinical outcomes of autoimmune diseases, accompanied by the high annual costs to the overall health care system, highlight the need for an efficient therapy. Increasing knowledge over the pathophysiology of specific chronic inflammatory diseases, namely Psoriasis (PsO), Inflammatory Bowel Diseases (IBD) consisting of Crohn’s disease (CD) and Ulcerative colitis (UC), and Rheumatoid Arthritis (RA), has provided insights into the underlying mechanisms that lead to the maintenance of the inflammation, such as Tumor Necrosis Factor alpha (TNF-α). Hence, the anti-TNFα biological agents pose as an ideal therapeutic approach. Despite the efficacy of anti-TNFα agents, several clinical trials have shown that 20-40% of patients do not respond to treatment. Nowadays, high-throughput technologies have been recruited in order to elucidate the complex interactions in multifactorial phenotypes, with the most ubiquitous ones referring to transcriptome quantification analyses. In this context, a random effects meta-analysis of available gene expression cDNA microarray datasets was performed between responders and non-responders to anti-TNFα therapy in patients with IBD, PsO, and RA. Publicly available datasets were systematically searched from inception to 10th of November 2020 and selected for further analysis if they assessed the response to anti-TNFα therapy with clinical score indexes from inflamed biopsies. Specifically, 4 IBD (79 responders/72 non-responders), 3 PsO (40 responders/11 non-responders) and 2 RA (16 responders/6 non-responders) datasetswere selected. After the separate pre-processing of each dataset, 4 separate meta-analyses were conducted; three disease-specific and a single combined meta-analysis on the disease-specific results. The MetaVolcano R package (v.1.8.0) was utilized for a random-effects meta-analysis through theRestricted Maximum Likelihood (RELM) method. The top 1% of the most consistently perturbed genes in the included datasets was highlighted through the TopConfects approach while maintaining a 5% False Discovery Rate (FDR). Genes were considered as Differentialy Expressed (DEGs) as those with P ≤ 0.05, |log2(FC)| ≥ log2(1.25) and perturbed in at least 75% of the included datasets. Over-representation analysis was performed using Gene Ontology and Reactome Pathways for both up- and down-regulated genes in all 4 performed meta-analyses. Protein-Protein interaction networks were also incorporated in the subsequentanalyses with STRING v11.5 and Cytoscape v3.9. Disease-specific meta-analyses detected multiple distinct pro-inflammatory and immune-related down-regulated genes for each disease, such asNFKBIA, IL36, and IRAK1, respectively. Pathway analyses revealed unique and shared pathways between each disease, such as Neutrophil Degranulation and Signaling by Interleukins. The combined meta-analysis unveiled 436 DEGs, 86 out of which were up- and 350 down-regulated, confirming the aforementioned shared pathways and genes, as well as uncovering genes that participate in anti-inflammatory pathways, namely IL-10 signaling. The identification of key biological pathways and regulatory elements is imperative for the accurate prediction of the patient’s response to biological drugs. Meta-analysis of such gene expression data could aid the challenging approach to unravel the complex interactions implicated in the response to anti-TNFα therapy in patients with PsO, IBD, and RA, as well as distinguish gene clusters and pathways that are altered through this heterogeneous phenotype.

Keywords: anti-TNFα, autoimmune, meta-analysis, microarrays

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Authors: Hala Raslan, Noha Eltaweel, Hanaa Rasmi, Solaf Kamel, May Magdy, Sherif Ismail, Khalda Amr

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Introduction: Rheumatoid arthritis (RA) is an inflammatory, autoimmune disorder with progressive joint damage. Osteoarthritis (OA) is a degenerative disease of the articular cartilage that shows multiple clinical manifestations or symptoms resembling those of RA. Genetic predisposition is believed to be a principal etiological factor for RA and OA. In this study, we aimed to measure the expression of the top deregulated miRNAs that might be the cause of pathogenesis in both diseases, according to our latest NGS analysis. Six of the deregulated miRNAs were selected as they had multiple target genes in the RA pathway, so they are more likely to affect the RA pathogenesis.Methods: Eighty cases were recruited in this study; 45 rheumatoid arthiritis (RA), 30 osteoarthiritis (OA) patients, as well as 20 healthy controls. The selection of the miRNAs from our latest NGS study was done using miRwalk according to the number of their target genes that are members in the KEGG RA pathway. Total RNA was isolated from plasma of all recruited cases. The cDNA was generated by the miRcury RT Kit then used as a template for real-time PCR with miRcury Primer Assays and the miRcury SYBR Green PCR Kit. Fold changes were calculated from CT values using the ΔΔCT method of relative quantification. Results were compared RA vs Controls and OA vs Controls. Target gene prediction and functional annotation of the deregulated miRNAs was done using Mienturnet. Results: Six miRNAs were selected. They were miR-15b-3p, -128-3p, -194-3p, -328-3p, -542-3p and -3180-5p. In RA samples, three of the measured miRNAs were upregulated (miR-194, -542, and -3180; mean Rq= 2.6, 3.8 and 8.05; P-value= 0.07, 0.05 and 0.01; respectively) while the remaining 3 were downregulated (miR-15b, -128 and -328; mean Rq= 0.21, 0.39 and 0.6; P-value= <0.0001, <0.0001 and 0.02; respectively) all with high statistical significance except miR-194. While in OA samples, two of the measured miRNAs were upregulated (miR-194 and -3180; mean Rq= 2.6 and 7.7; P-value= 0.1 and 0.03; respectively) while the remaining 4 were downregulated (miR-15b, -128, -328 and -542; mean Rq= 0.5, 0.03, 0.08 and 0.5; P-value= 0.0008, 0.003, 0.006 and 0.4; respectively) with statistical significance compared to controls except miR-194 and miR-542. The functional enrichment of the selected top deregulated miRNAs revealed the highly enriched KEGG pathways and GO terms. Conclusion: Five of the studied miRNAs were greatly deregulated in RA and OA, they might be highly involved in the disease pathogenesis and so might be future therapeutic targets. Further functional studies are crucial to assess their roles and actual target genes.

Keywords: MiRNAs, expression, rheumatoid arthritis, osteoarthritis

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Authors: Zunnu Raen Akhtar

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Cotton Leaf Curl Disease (CLCuD) is from Gemini virus and is transmitted through whiteflies in cotton. Transgenic cotton containing Antisense Coat Protein (ACP) has been found to show better results against CLCuD in cotton. In current research, Antisense Coat Protein was inserted in cotton plants to observe resistance developed in the cotton plants against CLCuD. T1 generation of plants were observed for its expression in plants. Tests were carried out to observe the expression of Antisense Coat Protein using Polymerase Chain Reaction (PCR) technique and by southern blotting. Whiteflies showing positive Cotton Leaf Curl Virus (CLCV) were reared and released in bioassay on ACP expressing cotton plants under laboratory as well as confined semi-field conditions. Results confirmed the expression of AC protein in PCR and southern blotting. Further laboratory results showed that cotton plants expressing AC protein showed rare incidence of CLCuD infection as compared to control. In the confined semi-field, similar results were observed in AC protein expressing cotton as compared to control. These results explicitly show that ACP can help to tackle the CLCuD issue in the future and further studies on biochemical processes involved in these plants and effects of ACP induction on non-target organisms should also be studied for eco-system.

Keywords: cotton, white flies, antisense coat protein, CLCV

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Authors: Yanyue Yuan

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In this paper, the author shares the findings of a pilot study that examines students’ creative expressions and their perceptions of creativity when engaged in project-based learning. The study is based on an elective course that the author co-designed and co-taught with a colleague to sixteen grade six and seven students over the spring semester in 2019. Using the Little Prince story as the main prompt, they facilitated students’ original creation of a storytelling concert that integrated script writing, music production, lyrics, songs, and visual design as a result of both individual and collaborative work. The author will share the specific challenges we met during the project, including learning cultures of the school, class management, teachers' and parents’ attitude, process-oriented versus product-oriented mindset, and facilities and logistical resources. The findings of this pilot study will inform the ongoing research initiative of exploring how we can foster creative learning in public schools in the Chinese context. While K-12 schools of China’s public education system are still dominated by exam-oriented and teacher-centered approaches, the author proposes that expression-based learning can be a starting point for promoting students’ creativity and can serve as experimental efforts to initiate incremental changes within the current education framework. The paper will also touch upon insights gained from collaborations between university and K-12 schools.

Keywords: creativity, expression-based learning, K-12, incremental changes

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Authors: Tiancheng Lan

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E10A is a kind of replication-defective adenovirus which carries the human endostatin gene to inhibit the growth of tumors. Kringle 5(K5) has almost the same function as angiostatin to also inhibit the growth of tumors since they are all the byproduct of the proteolytic cleavage of plasminogen. Tumor size increasing can be suppressed because both of the endostatin and K5 can restrain the angiogenesis process. Therefore, in order to improve the treatment effect on tumor, 2A peptide is used to construct a fusion gene carrying both E10A and K5. Using 2A peptide is an ideal strategy when a fusion gene is expressed because it can avoid many problems during the expression of more than one kind of protein. The overlap extension PCR is also used to connect 2A peptide with E10A and K5. The final construction of fusion gene E10A-2A-K5 can provide a possible new method of the anti-angiogenesis treatment with a better expression performance.

Keywords: E10A, Kringle 5, 2A peptide, overlap extension PCR

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Authors: Surbhi Surbhi, Andrea Erni, Gunter Meister, Harold Cremer, Christophe Beclin

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MicroRNAs (miRNAs) are short 20-23 nucleotide long non-coding RNAs; there are 2605 miRNA in humans and 1936 miRNA in mouse in total (miRBase). The nervous system expresses the most abundant miRNA and most diverse. MiRNAs play a role in many steps during neurogenesis, like cell proliferation, differentiation, neural patterning, axon pathfinding, etc. Moreover, in vitro studies suggested a role in the regulation of local translation at the synapse, thus controlling neuronal plasticity. However, due to the specific structure of miRNA molecules, an in-vivo confirmation of the general role of miRNAs in the control of neuronal plasticity is still pending. For example, their small size and their high level of sequence homology make difficult the analysis of their cellular and sub-cellular localization in-vivo by in-situ hybridization. Moreover, it was found that only 40% of the expressed miRNA molecules in a cell are included in RNA-Induced Silencing Complexes (RISC) and, therefore, involved in inhibitory interactions while the rest is silent. Definitively, the development of new tools is needed to have a better understanding of the cellular function of miRNAs, in particular their role in neuronal plasticity. Here we describe a new technique called in-vivo AGO-APP designed to investigate miRNA expression and function in-vivo. This technique is based on the expression of a small peptide derived from the human RISC-complex protein TNRC6B, called T6B, which binds all known Argonaute (Ago) proteins with high affinity allowing the efficient immunoprecipitation of AGO-bound miRNAs. We have generated two transgenic mouse lines conditionally expressing T6B either ubiquitously in the cell or targeted at the synapse. A comparison of the repertoire of miRNAs immuno-precipitated from mature neurons of both mouse lines will provide us with a list of miRNAs showing a specific activity at the synapse. The physiological role of these miRNAs will be subsequently addressed through gain and loss of function experiments.

Keywords: RNA-induced silencing complexes, TNRC6B, miRNA, argonaute, synapse, neuronal plasticity, neurogenesis

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Authors: Seung-Hwa Baek, In-Jung Nam, Sang-Han Lee

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The development of anti-melanogenic agents is important for the prevention of serious esthetic problem like a melasma, freckle, age spots, and chloasma. The aim of this study was to investigate the anti-melanogenic effect of sesamol, an active lignan isolated from sesame seed, by mushroom and cellular tyrosinase assay, melanin content and the analysis of melanogensis-related mRNA expressions in melana cells. Sesamol showed strong inhibitory activity against the mushroom tyrosinase in a dose-dependent manner. Intracellular tyrosinase inhibition activity was also confirmed by zymography. At a concentration of 50 μM, sesamol inhibited melanin production in melan-a cells with no cytoxicity while those of phenylthiourea (PTU) as a positive control were the same condition. Sesamol significantly inhibited the expression of melanogensis-related genes, such as tyrosinase, tyrosinase-related protein-1 (TRP-1), dopachrome tautomerase (Dct), microphthalmia-associated transcription factor (MITF) and melanocortin 1 receptor (MC1R). These findings indicate that sesamol could reduce melanin biosynthesis via the downregulation of tyrosinase activity and melanin production via subsequent gene expression of melanogenesis-related proteins. Together, these results suggest that the sesamol have strong potential in inhibiting melanin biosynthesis, in that the substance may be used as a new skin-whitening agent of cosmetic materials.

Keywords: sesamol, sesame seed, melanin biosynthesis, melanogenesis-related gene, skin-whitening agent

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Authors: Pelin Balcik Ercin

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Introduction: ZEB transcription factor family member ZEB2, has a role in epithelial to mesenchymal transition during development and metastasis. The altered circulating extracellular miRNAs expression is observed in diseases, and extracellular miRNAs have an important role in cancer cell microenvironment. In ChIP-Seq study, the expression of miR-2110 was found to be regulated by ZEB2. In this study, the effects of miR2110 on cell proliferation and migration of hepatocellular carcinoma (HCC) cells were examined. Material and Methods: SNU398 cells transfected with mimic miR2110 (20nM) (HMI0375, Sigma-Aldrich) and negative control miR (HMC0002, Sigma-Aldrich). MicroRNA isolation was accomplished with miRVANA isolation kit according to manufacturer instructions. cDNA synthesis was performed expression, respectively, and calibrated with Ct of controls. The real-time quantitative PCR (RT-qPCR) reaction was performed using the TaqMan Fast Advanced Master Mix (Thermo Sci.). Ct values of miR2110 were normalized to miR-186-5p and miR16-5p for the intracellular gene. Cell proliferation analysis was analyzed with the xCELLigence RTCA System. Wound healing assay was analyzed with the ImageJ program and relative fold change calculated. Results: The mimic-miR-2110 transfected SNU398 cells nearly nine-fold (log2) more miR-2110 expressed compared to negative control transfected cells. The mimic-miR-2110 transfected HCC cell proliferation significantly inhibited compared to the negative control cells. Furthermore, miR-2110-SNU398 cell migration capacity was relatively four-fold decreased compared to negative control-miR-SNU398 cells. Conclusion: Our results suggest the miR-2110 inhibited cell proliferation and also miR-2110 negatively affect cell migration compared to control groups in HCC cells. These data suggest the complexity of microRNA EMT transcription factors regulation. These initial results are pointed out the predictive biomarker capacity of miR-2110 in HCC.

Keywords: epithelial to mesenchymal transition, EMT, hepatocellular carcinoma cells, micro-RNA-2110, ZEB2

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Authors: Muhammad Imran, Sarfraz Shafiq, Xiangru Tang

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Alternative splicing (AS) is an important post-transcriptional regulatory mechanism to generate transcripts variability and proteome diversity in plants. Fragrant rice (Oryza sativa L.) has a high economic and nutritional value, and the application of micronutrients regulate 2-acetyl-1-pyrroline (2-AP) production, which is responsible for aroma in fragrant rice. However, no systematic investigation of AS events in response to micronutrients (Zn) has been performed in fragrant rice. Furthermore, the post-transcriptional regulation of genes involved in 2-AP biosynthesis is also not known. In this study, a comprehensive analysis of AS events under two gradients of Zn treatment in two different fragrant rice cultivars (Meixiangzhan-2 and Xiangyaxiangzhan) was performed. A total of 386 and 598 significant AS events were found in Meixiangzhan-2 treated with low and high doses of Zn, respectively. In Xiangyaxiangzhan, a total of 449 and 598 significant AS events were found in low and high doses of Zn, respectively. Go analysis indicated that these genes were highly enriched in physiological processes, metabolism, and cellular process in both cultivars. However, genotype and dose-dependent AS events were also detected in both cultivars. By comparing differential AS (DAS) events with differentially expressed genes (DEGs), we found a weak overlap among DAS and DEGs in both fragrant rice cultivars, indicating that only a few genes are post-transcriptionally regulated in response to Zn treatment. We further report that Zn differentially regulates the expression of 2-AP biosynthesis-related genes in both cultivars, and Zn treatment altered the editing frequency of SNPs in the genes involved in 2-AP biosynthesis. Finally, we showed that epigenetic modifications associated with active gene transcription are generally enriched over 2-AP biosynthesis-related genes. Taken together, our results provide evidence of the post-transcriptional gene regulation in fragrant rice in response to Zn treatment and highlight that the 2-AP biosynthesis pathway may also be post-transcriptionally regulated through epigenetic modifications. These findings will serve as a cornerstone for further investigation to understand the molecular mechanisms of 2-AP biosynthesis in fragrant rice.

Keywords: fragrant rice, 2-acetyl-1-pyrroline, gene expression, zinc, alternative splicing, SNPs

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Authors: Mrunmayee Das

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This study aims to explore the film 'Pyasa' directed by actor-director Guru Dutt, filmed during the 1950’s golden age of Hindi cinema. 'Pyasa' was filmed after a decade of India being a new nation and narrates the world-view of a poet dressed in western ideals, tasting modernity, uprooted from his familial and social moorings causing friction of being between survival and self- expression. The research is based on literature review to study the attitudes, particularly the post-colonial, informing the film. In terms of the structure, the relational study of the film and the historical background of that time came first. Further explorations deal with the use of image making, dialogue, and poetry in the form of songs facilitating the central theme of the human plight of poverty, not of material but of thought. The literature review establishes Dutt’s style of expressing melodic melodrama through a dance between light and shadow majorly in the form of song sequences signifying the greys of the society. It was found in this research that melodrama is created by the changing contrasts and use of close-ups. The song sequences convey the tensions of being a contemporary liberal educated youth and having to deal with the societal-ills of this world, which highlights the theme of compulsion towards self-destruction. It is concluded that Dutt’s 'Pyasa' is a autobiographical commentary on the state of a nation doing away with a borrowed identity and refashioning its own.

Keywords: cinema, Guru Dutt, post-colonial India, self-expression

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Authors: Grzegorz Kubinski

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In the proposed paper, we would like to present the phenomenon of disease and disability as an element of discourse redefining the contemporary canons of beauty and the category of normativity. In widely understood media, and above all in social media and fashion industry, the use of the disease as an aesthetic category has long been observed. There is an interesting case of promoting and maintaining a certain, ideal pattern of physical beauty, while at the same time very clear exploitation of various types of illnesses. The categories of disease and disabled body are shown as an element of the expression of the individuality and originality of one's own identity, while at the same time the disabled person is still experiencing social exclusion. Illness or body abnormality as an aesthetic category also functions as an ethical-political category. The analysis of the interrelations of these discourses will be presented on the example of selected projects present in social media, like Instagram or Facebook. We would like to present how old forms of 'curiosities' or 'abnormalities' turned into mainstream forms of a new aesthetic. For marginalized disabled people, there is a new form of expression and built their identity. But, there is an interesting point: are this contemporary forms of using disability and illness really new? Or maybe this is just another form of Wunderkammer or even cabinets of curiosities? We propose to analyze contemporary cultural and social context in order to clarify this issue. On the other hand, we would like to present some examples from personal interviews with disabled internet influencers and statements disabled persons concerning the role of the different body in society (e.g. #bodypositive, #perfeclyflawed).

Keywords: disability, new media, defect, fashion

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Authors: Hung-Chieh Chen, Kuo-Hui Wang, Tsu-Lin Yeh

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Proteomics represent the performance of genomic complement proteins and the protein level on functional genomics. This study adopted proteomic strategies for comparing serum proteins among three groups: elite sprinter (sprint runner group, SR), long-distance runners (long-distance runner group, LDR), and the untrained control group (control group, CON). Purposes: This study aims to identify elite sprinters and long-distance runners’ serum protein and to provide a comparison of their serum proteome’ composition. Methods: Serum protein fractionations that separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and analyzed by a quantitative nano-LC-MS/MS-based proteomic profiling. The one-way analysis of variance (ANOVA) and Scheffe post hoc comparison (α= 0.05) was used to determine whether there is any significant difference in each protein level among the three groups. Results: (1) After analyzing the 307 identified proteins, there were 26 unique proteins in the SR group, and 18 unique proteins in the LDR group. (2) For the LDR group, 7 coagulation function-associated proteins’ expression levels were investigated: vitronectin, serum paraoxonase/arylesterase 1, fibulin-1, complement C3, vitamin K-dependent protein, inter-alpha-trypsin inhibitor heavy chain H3 and von Willebrand factor, and the findings show the seven coagulation function-associated proteins were significantly lower than the group of SR. (3) Comparing to the group of SR, this study found that the LDR group’s expression levels of the 2 antioxidant proteins (afamin and glutathione peroxidase 3) were also significantly lower. (4) The LDR group’s expression levels of seven immune function-related proteins (Ig gamma-3 chain C region, Ig lambda-like polypeptide 5, clusterin, complement C1s subcomponent, complement factor B, complement C4-A, complement C1q subcomponent subunit A) were also significantly lower than the group of SR. Conclusion: This study identified the potential serum protein markers for elite sprinters and long-distance runners. The changes in the regulation of coagulation, antioxidant, or immune function-specific proteins may also provide further clinical applications for these two different track athletes.

Keywords: biomarkers, coagulation, immune response, oxidative stress

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Authors: Masoud Sakhinia, Sajjad Ahmad

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Introduction: Though knowledge of the compositional makeup and structure of the limbal niche has progressed exponentially during the past decade, much is yet to be understood. Identifying the precise profile and role of the stromal makeup which spans the ocular surface may inform researchers of the most optimum conditions needed to effectively expand LESCs in vitro, whilst preserving their differentiation status and phenotype. Limbal fibroblasts, as opposed to corneal fibroblasts are thought to form an important component of the microenvironment where LESCs reside. Methods: The corneal stroma was tissue engineered in vitro using both limbal and corneal fibroblasts embedded within a tissue engineered 3D collagen matrix. The effect of these two different fibroblasts on LESCs and hTCEpi corneal epithelial cell line were then subsequently determined using phase contrast microscopy, histolological analysis and PCR for specific stem cell markers. The study aimed to develop an in vitro model which could be used to determine whether limbal, as opposed to corneal fibroblasts, maintained the stem cell phenotype of LESCs and hTCEpi cell line. Results: Tissue culture analysis was inconclusive and required further quantitative analysis for remarks on cell proliferation within the varying stroma. Histological analysis of the tissue-engineered cornea showed a comparable structure to that of the human cornea, though with limited epithelial stratification. PCR results for epithelial cell markers of cells cultured on limbal fibroblasts showed reduced expression of CK3, a negative marker for LESC’s, whilst also exhibiting a relatively low expression level of P63, a marker for undifferentiated LESCs. Conclusion: We have shown the potential for the construction of a tissue engineered human cornea using a 3D collagen matrix and described some preliminary results in the analysis of the effects of varying stroma consisting of limbal and corneal fibroblasts, respectively, on the proliferation of stem cell phenotype of primary LESCs and hTCEpi corneal epithelial cells. Although no definitive marker exists to conclusively illustrate the presence of LESCs, the combination of positive and negative stem cell markers in our study were inconclusive. Though it is less traslational to the human corneal model, the use of conditioned medium from that of limbal and corneal fibroblasts may provide a more simple avenue. Moreover, combinations of extracellular matrices could be used as a surrogate in these culture models.

Keywords: cornea, Limbal Stem Cells, tissue engineering, PCR

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Authors: Pei-Hsiu Chiang, Ling Hong Huang, Hsin-I Chang

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In this research, we used UVA irradiation, which can penetrate into dermis and fibroblasts, the most abundant cells in dermis, to investigate the effect of UV light on dermis, such as inflammation, ECM degradation and elasticity loss. Moreover, this research is focused on the influence of hyaluronic acid (HA) on UVA treated dermal fibroblasts. We aim to establish whether HA can effectively relief ECM degradation, and restore the elasticity of UVA-damaged fibroblasts. Prolonged exposure to UVA radiation can damage fibroblasts and led variation in cell morphology and reduction in cell viability. Besides, UVA radiation can induce IL-1β expression on fibroblasts and then promote MMP-1 and MMP-3 expression, which can accelerate ECM degradation. On the other hand, prolonged exposure to UVA radiation reduced collagen and elastin synthesis on fibroblasts. Due to the acceleration of ECM degradation and the reduction of ECM synthesis, Atomic force microscope (AFM) was used to analyze the elasticity reduction on UVA-damaged fibroblasts. UVA irradiation causes photoaging on fibroblasts. UVA damaged fibroblasts with HA treatment can down-regulate the gene expression of MMP-1, MMP-3, and then slow down ECM degradation. On the other hand, HA may restore elastin and collagen synthesis in UV-damaged fibroblasts. Based on the slowdown of ECM degradation, UVA-damaged fibroblast elasticity can be effectively restored by HA treatment. In summary, HA can relief the photoaging conditions on fibroblasts, but may not be able to return fibroblasts to normal, healthy state. Although HA cannot fully recover UVA-damaged fibroblasts, HA is still potential for repairing photoaging skin.

Keywords: atomic force microscope, hyaluronic acid, UVA radiation, dermal fibroblasts

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Authors: Dani Sukkar, Ali Kanso, Philippe Laval-Gilly, Jairo Falla-Angel

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The honeybees' immune system is challenged by different risk factors that induce various responses. However, complex scenarios where bees are exposed to different pesticides simultaneously with immune activation are not well evaluated. The Toll pathway is one of the main signaling pathways studied in invertebrate immune responses, and it is a good indicator of the effect of such complex interactions in addition to key signaling elements of other pathways like Relish of the immune deficiency (IMD) pathway or Eater, the phagocytosis receptor or vitellogenin levels. Honeybee hemocytes extracted from 5th instar larvae were exposed to imidacloprid and/or amitraz with or without the presence of the zymosan a as an immune activator. The gene expression of multiple immune related genes were studied, including spaetzle, Toll, myD88, relish, eater and vitellogenin, by real-time polymerase chain reaction after RNA extraction. The results demonstrated that the Toll pathway is mainly affected by the pesticides; imidacloprid and amitraz, especially by their different combinations. Furthermore, immune activation by zymosan A, a fungal cell-wall component, acts to mitigate to some extent the effect of pesticides on the different levels of the Toll pathway. In addition, imidacloprid, amitraz, and zymosan A have complex and context-specific interactions depending on the levels of immune activation and the pathway evaluated affecting immune-gene expression differently.

Keywords: toll pathway, immune modulation, β-glucan, imidacloprid, amitraz, honeybees, immune genes

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Authors: K. Roushanger, A. Soleymanzadeh

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Weirs are structures often used in irrigation techniques, sewer networks and flood protection. However, the hydraulic behavior of this type of weir is complex and difficult to predict accurately. An accurate flow prediction over a weir mainly depends on the proper estimation of discharge coefficient. In this study, the Genetic Expression Programming (GEP) approach was used for predicting trapezoidal and rectangular sharp-crested side weirs discharge coefficient. Three different performance indexes are used as comparing criteria for the evaluation of the model’s performances. The obtained results approved capability of GEP in prediction of trapezoidal and rectangular side weirs discharge coefficient. The results also revealed the influence of downstream Froude number for trapezoidal weir and upstream Froude number for rectangular weir in prediction of the discharge coefficient for both of side weirs.

Keywords: discharge coefficient, genetic expression programming, trapezoidal weir

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