Biological Procedures Online最新文献

{"title":"Visualisation of Bluetongue Virus in the Salivary Apparatus of Culicoides Biting Midges Highlights the Accessory Glands as a Primary Arboviral Infection Site.","authors":"Marc Guimerà Busquets, Faye V Brown, Simon T Carpenter, Karin E Darpel, Christopher J Sanders","doi":"10.1186/s12575-023-00221-2","DOIUrl":"10.1186/s12575-023-00221-2","url":null,"abstract":"<p><strong>Background: </strong>Arthropods transmit a wide range of pathogens of importance for the global health of humans, animals, and plants. One group of these arthropod vectors, Culicoides biting midges (Diptera: Ceratopogonidae), is the biological vector of several human and animal pathogens, including economically important livestock viruses like bluetongue virus (BTV). Like other arthropod-borne viruses (arboviruses), Culicoides-borne viruses must reach and replicate in the salivary apparatus, from where they can be transmitted to susceptible hosts through the saliva during subsequent blood feeding. Despite the importance of the salivary gland apparatus for pathogen transmission to susceptible animals from the bite of infected Culicoides, these structures have received relatively little attention, perhaps due to the small size and fragility of these vectors.</p><p><strong>Results: </strong>In this study, we developed techniques to visualize the infection of the salivary glands and other soft tissues with BTV, in some of the smallest known arbovirus vectors, Culicoides biting midges, using three-dimensional immunofluorescence confocal microscopy. We showed BTV infection of specific structures of the salivary gland apparatus of female Culicoides vectors following oral virus uptake, related visualisation of viral infection in the salivary apparatus to high viral RNA copies in the body, and demonstrated for the first time, that the accessory glands are a primary site for BTV replication within the salivary apparatus.</p><p><strong>Conclusions: </strong>Our work has revealed a novel site of virus-vector interactions, and a novel role of the accessory glands of Culicoides in arbovirus amplification and transmission. Our approach would also be applicable to a wide range of arbovirus vector groups including sand flies (Diptera: Psychodidae), as well as provide a powerful tool to investigate arbovirus infection and dissemination, particularly where there are practical challenges in the visualization of small size and delicate tissues of arthropods.</p>","PeriodicalId":8960,"journal":{"name":"Biological Procedures Online","volume":"25 1","pages":"27"},"PeriodicalIF":3.7,"publicationDate":"2023-11-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10626815/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"71477658","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A Novel Dual-Reporter System Reveals Distinct Characteristics of Exosome-Mediated Protein Secretion in Human Cells.","authors":"Christopher Olson, Pengyang Zhang, Joy Ku, Renceh Flojo, Darin Boyes, Biao Lu","doi":"10.1186/s12575-023-00219-w","DOIUrl":"10.1186/s12575-023-00219-w","url":null,"abstract":"<p><strong>Background: </strong>Exosomes, a special subtype of extracellular vesicles derived from human cells, serve as vital mediators of intercellular communication by transporting diverse bioactive cargos, including proteins and enzymes. However, the underlying mechanisms governing exosome secretion and regulation remain poorly understood. In this study, we employed a dual-reporter system consisting of bioluminescent Gaussia luciferase and fluorescent proteins to investigate the dynamics and regulation of exosome secretion in cultured human cells.</p><p><strong>Results: </strong>Our results demonstrated that the engineered dual-reporters effectively monitored both exosome-mediated and ER-Golgi-mediated secretory pathways in a specific and quantitative manner. Notably, we observed distinct characteristics of exosome-mediated protein secretion, including significantly lower capacity and different dynamics compared to the ER-Golgi pathway. This phenomenon was observed in human kidney 293T cells and liver HepG2 cells, emphasizing the conserved nature of exosome-mediated secretion across cell types. Furthermore, we investigated the impact of brefeldin A (BFA), an inhibitor of ER-to-Golgi membrane trafficking, on protein secretion. Interestingly, BFA inhibited protein secretion via the ER-Golgi pathway while stimulating exosome-mediated protein secretion under same experimental conditions.</p><p><strong>Conclusions: </strong>Collectively, our study highlights the utility of the dual-reporter system for real-time monitoring and quantitative analysis of protein secretion through conventional ER-Golgi and unconventional exosome pathways. Moreover, our findings unveil distinct features of exosome-mediated protein secretion, shedding light on its differential capacity, dynamics, and regulatory mechanisms compared to ER-Golgi-mediated proteins in human cells.</p>","PeriodicalId":8960,"journal":{"name":"Biological Procedures Online","volume":"25 1","pages":"25"},"PeriodicalIF":6.4,"publicationDate":"2023-09-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10510171/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"41092671","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Comparison of two protocols for the generation of iPSC-derived human astrocytes.","authors":"Patrycja Mulica, Carmen Venegas, Zied Landoulsi, Katja Badanjak, Sylvie Delcambre, Maria Tziortziou, Soraya Hezzaz, Jenny Ghelfi, Semra Smajic, Jens Schwamborn, Rejko Krüger, Paul Antony, Patrick May, Enrico Glaab, Anne Grünewald, Sandro L Pereira","doi":"10.1186/s12575-023-00218-x","DOIUrl":"10.1186/s12575-023-00218-x","url":null,"abstract":"<p><strong>Background: </strong>Astrocytes have recently gained attention as key contributors to the pathogenesis of neurodegenerative disorders including Parkinson's disease. To investigate human astrocytes in vitro, numerous differentiation protocols have been developed. However, the properties of the resulting glia are inconsistent, which complicates the selection of an appropriate method for a given research question. Thus, we compared two approaches for the generation of iPSC-derived astrocytes. We phenotyped glia that were obtained employing a widely used long, serum-free (\"LSF\") method against an in-house established short, serum-containing (\"SSC\") protocol which allows for the generation of astrocytes and midbrain neurons from the same precursor cells.</p><p><strong>Results: </strong>We employed high-content confocal imaging and RNA sequencing to characterize the cultures. The astrocytes generated with the LSF or SSC protocols differed considerably in their properties: while the former cells were more labor-intense in their generation (5 vs 2 months), they were also more mature. This notion was strengthened by data resulting from cell type deconvolution analysis that was applied to bulk transcriptomes from the cultures to assess their similarity with human postmortem astrocytes.</p><p><strong>Conclusions: </strong>Overall, our analyses highlight the need to consider the advantages and disadvantages of a given differentiation protocol, when designing functional or drug discovery studies involving iPSC-derived astrocytes.</p>","PeriodicalId":8960,"journal":{"name":"Biological Procedures Online","volume":"25 1","pages":"26"},"PeriodicalIF":6.4,"publicationDate":"2023-09-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10512486/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"41113426","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Immune Activation Effects at Different Irradiated Sites and Optimal Timing of Radioimmunotherapy in Patients with Extensive-Stage Small Cell Lung Cancer: a Real-World Analysis.","authors":"Min Wu, Shihao Wu, Yuetong Chen, Liangchao Sun, Jundong Zhou","doi":"10.1186/s12575-023-00217-y","DOIUrl":"10.1186/s12575-023-00217-y","url":null,"abstract":"<p><strong>Background: </strong>In view of the limited data on radiotherapy (RT) combined with immunotherapy in patients with extensive-stage small cell lung cancer (ES-SCLC), this study aimed to identify the immune activation effect on different sites and the survival outcomes of radioimmunotherapy at different treatment stages.</p><p><strong>Methods: </strong>Forty-five patients diagnosed with ES-SCLC were included in this retrospective analysis. We collected the overall survival (OS) of the patients,, recorded the blood cell counts before, during, and after RT, and derived blood index ratios such as the neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio (PLR), and systemic immune-inflammation index (SII). The datasets were analyzed using the Spearman rank correlation test, Kruskal-Wallis rank sum test and logistic regression.</p><p><strong>Results: </strong>Among the selected blood indices, the delta-NLR/PLR/Sll correlated with different irradiated organs, and the mean ranks of these three indices were the lowest in the brain-irradiated group during immunotherapy. Additionally, adjunct first-line immunotherapy with RT demonstrated a significant improvement compared to second- or third-line therapy and subsequent therapies.</p><p><strong>Conclusion: </strong>Our findings suggest that compared to other organs, the strongest immune activation effect occurs with brain RT, and ES-SCLC patients who received radioimmunotherapy (RIT) earlier achieved higher OS rates.</p>","PeriodicalId":8960,"journal":{"name":"Biological Procedures Online","volume":"25 1","pages":"24"},"PeriodicalIF":6.4,"publicationDate":"2023-09-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10503112/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10267589","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Correction: Circ‑ABCB10 knockdown inhibits the malignant progression of cervical cancer through microRNA‑128‑3p/ZEB1 axis.","authors":"Wei Feng, Dongya Zhang, Ruitao Zhang","doi":"10.1186/s12575-023-00216-z","DOIUrl":"10.1186/s12575-023-00216-z","url":null,"abstract":"","PeriodicalId":8960,"journal":{"name":"Biological Procedures Online","volume":"25 1","pages":"23"},"PeriodicalIF":6.4,"publicationDate":"2023-08-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10401802/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9946280","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A high-throughput screening system for SARS-CoV-2 entry inhibition, syncytia formation and cell toxicity.","authors":"Shine Varghese Jancy,&nbsp;Santhik Subhasingh Lupitha,&nbsp;Aneesh Chandrasekharan,&nbsp;Shankara Narayanan Varadarajan,&nbsp;Shijulal Nelson-Sathi,&nbsp;Roshny Prasad,&nbsp;Sara Jones,&nbsp;Sreekumar Easwaran,&nbsp;Pramod Darvin,&nbsp;Aswathy Sivasailam,&nbsp;Thankayyan Retnabai Santhoshkumar","doi":"10.1186/s12575-023-00214-1","DOIUrl":"https://doi.org/10.1186/s12575-023-00214-1","url":null,"abstract":"<p><strong>Background: </strong>The entry of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) into the host cell is mediated through the binding of the SARS-CoV-2 Spike protein via the receptor binding domain (RBD) to human angiotensin-converting enzyme 2 (hACE2). Identifying compounds that inhibit Spike-ACE2 binding would be a promising and safe antiviral approach against COVID-19.</p><p><strong>Methods: </strong>In this study, we used a BSL-2 compatible replication-competent vesicular stomatitis virus (VSV) expressing Spike protein of SARS-CoV-2 with eGFP reporter system (VSV-eGFP-SARS-CoV-2) in a recombinant permissive cell system for high-throughput screening of viral entry blockers. The SARS-CoV-2 permissive reporter system encompasses cells that stably express hACE2-tagged cerulean and H2B tagged with mCherry, as a marker of nuclear condensation, which also enables imaging of fused cells among infected EGFP positive cells and could provide real-time information on syncytia formation.</p><p><strong>Results: </strong>A limited high-throughput screening identified six natural products that markedly inhibited VSV-eGFP-SARS-CoV-2 with minimum toxicity. Further studies of Spike-S1 binding using the permissive cells showed Scillaren A and 17-Aminodemethoxygeldanamycin could inhibit S1 binding to ACE2 among the six leads. A real-time imaging revealed delayed inhibition of syncytia by Scillaren A, Proscillaridin, Acetoxycycloheximide and complete inhibition by Didemnin B indicating that the assay is a reliable platform for any image-based drug screening.</p><p><strong>Conclusion: </strong>A BSL-2 compatible assay system that is equivalent to the infectious SARS-CoV-2 is a promising tool for high-throughput screening of large compound libraries for viral entry inhibitors against SARS-CoV-2 along with toxicity and effects on syncytia. Studies using clinical isolates of SARS-CoV-2 are warranted to confirm the antiviral potency of the leads and the utility of the screening system.</p>","PeriodicalId":8960,"journal":{"name":"Biological Procedures Online","volume":"25 1","pages":"22"},"PeriodicalIF":6.4,"publicationDate":"2023-07-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10373420/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9889422","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Full spectrum flow cytometry-powered comprehensive analysis of PBMC as biomarkers for immunotherapy in NSCLC with EGFR-TKI resistance.","authors":"Juan Zhou, Xiangling Chu, Jing Zhao, Mengqing Xie, Jing Wu, Xin Yu, Yujia Fang, Yazhou Li, Xiyan Li, Chunxia Su","doi":"10.1186/s12575-023-00215-0","DOIUrl":"10.1186/s12575-023-00215-0","url":null,"abstract":"<p><strong>Background: </strong>Clinical studies suggest that immune checkpoint inhibitor (ICI) monotherapy has limited benefits in non-small cell lung cancer (NSCLC) patients after epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) failure. However, data about efficacy of ICI plus chemotherapy remain controversial, probably attributed to the heterogeneity among such population, and robust efficacy biomarkers are urgent to explore.</p><p><strong>Methods: </strong>A total of 60 eligible patients who received ICI plus chemotherapy after EGFR-TKI treatment failure were enrolled, 24 of whom peripheral blood mononuclear cell (PBMC) samples were collected at baseline and after 2 cycles of treatment. We have designed a 23-color-antibody panel to detect PBMC by full spectrum flow cytometry.</p><p><strong>Results: </strong>For EGFR-TKI resistant NSCLC patients: 1) ICI plus chemotherapy achieved an objective response rate (ORR) of 21.7% and a median progression-free survival (PFS) of 6.4 months. 2) clinical characteristics associated with worse efficacy included liver metastasis and platelet-to-lymphocyte ratio (PLR) > 200. 3) the proportion of immune cell subset associated with better efficacy was higher baseline effective CD4⁺T cells (E4). 4) the baseline expression of immune checkpoint proteins (ICPs) on cell subsets associated with better efficacy included: higher expression of CD25 on dendritic cells (DC) and central memory CD8⁺T cells (CM8), and higher expression of Lymphocyte activation gene 3 (LAG-3) on effective memory CD8⁺T cells (EM8). 5) the expression of ICPs after 2 cycles of treatment associated with better efficacy included: higher expression of CD25 on CD8⁺T/EM8 /natural killer (NK) cells. 6) the dynamic changes of ICPs expression associated with worse efficacy included: significantly decrease of T cell immunoglobulin and ITIM domain (TIGIT) expression on regular T cells (Tregs) and decrease of V-domain immunoglobulin suppressor of T cell activation (VISTA) expression on Th1. 7) a prediction model for the efficacy of ICI plus chemotherapy was successfully constructed with a sensitivity of 62.5%, specificity of 100%, and area under curve (AUC) = 0.817.</p><p><strong>Conclusions: </strong>Some EGFR-TKI-resistant NSCLC patients could indeed benefit from ICI plus chemotherapy, but most patients are primary resistant to immunotherapy. Comprehensive analysis of peripheral immune cells using full spectrum flow cytometry showed that compared to the proportion of cell subsets, the expression type and level of ICPs on immune cells, especially CD25, were significantly correlated with the efficacy of immunotherapy.</p>","PeriodicalId":8960,"journal":{"name":"Biological Procedures Online","volume":"25 1","pages":"21"},"PeriodicalIF":3.7,"publicationDate":"2023-07-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10364374/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10232948","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"LINC00853 contributes to tumor stemness of gastric cancer through FOXP3-mediated transcription of PDZK1IP1.","authors":"Xia Hu,&nbsp;Maoyuan Zhao,&nbsp;Shuangyuan Hu,&nbsp;Qingsong Liu,&nbsp;Wenhao Liao,&nbsp;Lina Wan,&nbsp;Feng Wei,&nbsp;Fangting Su,&nbsp;Yu Guo,&nbsp;Jinhao Zeng","doi":"10.1186/s12575-023-00213-2","DOIUrl":"https://doi.org/10.1186/s12575-023-00213-2","url":null,"abstract":"<p><strong>Background: </strong>The incidence and mortality of gastric cancer (GC) are high worldwide. Tumor stemness is a major contributor to tumorigenesis and development of GC, in which long non-coding RNAs (lncRNAs) are deeply involved. The purpose of this study was to investigate the influences and mechanisms of LINC00853 in the progression and stemness of GC.</p><p><strong>Methods: </strong>The level of LINC00853 was assessed based on The Cancer Genome Atlas (TCGA) database and GC cell lines by RT-PCR and in situ hybridization. An evaluation of biological functions of LINC00853 including cell proliferation, migration, and tumor stemness was conducted via gain-and loss-of-function experiments. Furthermore, RNA pull-down and RNA immunoprecipitation (RIP) assay were utilized to validate the connection between LINC00853 and the transcription factor Forkhead Box P3 (FOXP3). Nude mouse xenograft model was used to identify the impacts of LINC00853 on tumor development.</p><p><strong>Results: </strong>We identified the up-regulated levels of lncRNA-LINC00853 in GC, and its overexpression correlates with poor prognosis in GC patients. Further study indicated that LINC00853 promoted cell proliferation, migration and cancer stemness while suppressed cell apoptosis. Mechanistically, LINC00853 directly bind to FOXP3 and promoted FOXP3-mediated transcription of PDZK1 interacting protein 1(PDZK1IP1). Alterations of FOXP3 or PDZK1IP1 reversed the LINC00853-induced biological effects on cell proliferation, migration and stemness. Moreover, xenograft tumor assay was used to investigate the function of LINC00853 in vivo.</p><p><strong>Conclusions: </strong>Taken together, these findings revealed the tumor-promoting activity of LINC00853 in GC, expanding our understanding of lncRNAs regulation on GC pathogenesis.</p>","PeriodicalId":8960,"journal":{"name":"Biological Procedures Online","volume":"25 1","pages":"20"},"PeriodicalIF":6.4,"publicationDate":"2023-07-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10318836/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10117042","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Trastuzumab-resistant breast cancer cells-derived tumor xenograft models exhibit distinct sensitivity to lapatinib treatment in vivo.","authors":"Hao Liu,&nbsp;Sanbao Ruan,&nbsp;Margaret E Larsen,&nbsp;Congcong Tan,&nbsp;Bolin Liu,&nbsp;Hui Lyu","doi":"10.1186/s12575-023-00212-3","DOIUrl":"https://doi.org/10.1186/s12575-023-00212-3","url":null,"abstract":"<p><strong>Background: </strong>Resistance to HER2-targeted therapies, including the monoclonal antibody trastuzumab and tyrosine kinase inhibitor lapatinib, frequently occurs and currently represents a significant clinical challenge in the management of HER2-positive breast cancer. We previously showed that the trastuzumab-resistant SKBR3-pool2 and BT474-HR20 sublines were refractory to lapatinib in vitro as compared to the parental SKBR3 and BT474 cells, respectively. The in vivo efficacy of lapatinib against trastuzumab-resistant breast cancer remained unclear.</p><p><strong>Results: </strong>In tumor xenograft models, both SKBR3-pool2- and BT474-HR20-derived tumors retained their resistance phenotype to trastuzumab; however, those tumors responded differently to the treatment with lapatinib. While lapatinib markedly suppressed growth of SKBR3-pool2-derived tumors, it slightly attenuated BT474-HR20 tumor growth. Immunohistochemistry analyses revealed that lapatinib neither affected the expression of HER3, nor altered the levels of phosphorylated HER3 and FOXO3a in vivo. Interestingly, lapatinib treatment significantly increased the levels of phosphorylated Akt and upregulated the expression of insulin receptor substrate-1 (IRS1) in the tumors-derived from BT474-HR20, but not SKBR3-pool2 cells.</p><p><strong>Conclusions: </strong>Our data indicated that SKBR3-pool2-derived tumors were highly sensitive to lapatinib treatment, whereas BT474-HR20 tumors exhibited resistance to lapatinib. It seemed that the inefficacy of lapatinib against BT474-HR20 tumors in vivo was attributed to lapatinib-induced upregulation of IRS1 and activation of Akt. Thus, the tumor xenograft models-derived from SKBR3-pool2 and BT474-HR20 cells serve as an excellent in vivo system to test the efficacy of other HER2-targeted therapies and novel agents to overcome trastuzumab resistance against HER2-positive breast cancer.</p>","PeriodicalId":8960,"journal":{"name":"Biological Procedures Online","volume":"25 1","pages":"19"},"PeriodicalIF":6.4,"publicationDate":"2023-06-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10294508/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9720446","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Integrating Omics and CRISPR Technology for Identification and Verification of Genomic Safe Harbor Loci in the Chicken Genome.","authors":"Nima Dehdilani,&nbsp;Lena Goshayeshi,&nbsp;Sara Yousefi Taemeh,&nbsp;Ahmad Reza Bahrami,&nbsp;Sylvie Rival Gervier,&nbsp;Bertrand Pain,&nbsp;Hesam Dehghani","doi":"10.1186/s12575-023-00210-5","DOIUrl":"https://doi.org/10.1186/s12575-023-00210-5","url":null,"abstract":"<p><strong>Background: </strong>One of the most prominent questions in the field of transgenesis is 'Where in the genome to integrate a transgene?'. Escape from epigenetic silencing and promoter shutdown of the transgene needs reliable genomic safe harbor (GSH) loci. Advances in genome engineering technologies combined with multi-omics bioinformatics data have enabled rational evaluation of GSH loci in the host genome. Currently, no validated GSH loci have been evaluated in the chicken genome.</p><p><strong>Results: </strong>Here, we analyzed and experimentally examined two GSH loci in the genome of chicken cells. To this end, putative GSH loci including chicken HIPP-like (cHIPP; between DRG1 and EIF4ENIF1 genes) and chicken ROSA-like (cROSA; upstream of the THUMPD3 gene) were predicted using multi-omics bioinformatics data. Then, the durable expression of the transgene was validated by experimental characterization of continuously-cultured isogenous cell clones harboring DsRed2-ΔCMV-EGFP cassette in the predicted loci. The weakened form of the CMV promoter (ΔCMV) allowed the precise evaluation of GSH loci in a locus-dependent manner compared to the full-length CMV promoter.</p><p><strong>Conclusions: </strong>cHIPP and cROSA loci introduced in this study can be reliably exploited for consistent bio-manufacturing of recombinant proteins in the genetically-engineered chickens. Also, results showed that the genomic context dictates the expression of transgene controlled by ΔCMV in GSH loci.</p>","PeriodicalId":8960,"journal":{"name":"Biological Procedures Online","volume":"25 1","pages":"18"},"PeriodicalIF":6.4,"publicationDate":"2023-06-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10290409/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10072048","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}