Cell Journal最新文献

{"title":"Melatonin Protects Mouse Type A Spermatogonial Stem Cells against Oxidative Stress via The Mitochondrial Thioredoxin System.","authors":"Somayeh Heidarizadi, Zahra Rashidi, Cyrus Jalili, Kamran Mansouri, Iraj Rashidi, Behzad Mahaki, Mohammadreza Gholami","doi":"10.22074/cellj.2023.2003766.1316","DOIUrl":"https://doi.org/10.22074/cellj.2023.2003766.1316","url":null,"abstract":"<p><strong>Objective: </strong>Mitochondrial oxidative stress is an important factor in infertility. The mitochondrial thioredoxin system plays an important role in this condition. N-acetyl-5-methoxy tryptamine (melatonin) plays a role in reducing oxidative stress and apoptosis in spermatogonial stem cells (SSCs). In this study, we explore the probable protective effects of melatonin on the mitochondrial thioredoxin system [thioredoxin 2 (Trx2)/Txnip] in SSCs under oxidative stress.</p><p><strong>Materials and methods: </strong>In this experimental study, SSCs were co-cultured two-dimensionally (2D) with Sertoli cells in DMEM culture medium that contained 10% fetal bovine serum (FBS), 1% antibiotics, and 10 ng/ml glial cell-derived neurotrophic factor (GDNF) for 30 days. The cultured cells were subsequently divided into four groups: control; melatonin (250 μM, 24 hours); melatonin (250 μM, 24 hours)+hydrogen peroxide (H₂O₂, 50 μM, 24 hours); and H₂O₂ (50 μM, 24 hours). Intracellular reactive oxygen species (ROS) production was determined by flow cytometry. Malondialdehyde (MDA) levels were measured by Fluorometry. The expressions of apoptotic and antioxidant genes and nuclear factor erythroid 2-related factor 2 (Nrf2), Trx2, and nicotinamide nucleotide transhydrogenase (NNT) proteins were determined by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot. Adenosine triphosphate (ATP) levels were measured by fluorometry.</p><p><strong>Results: </strong>Melatonin reduced H2O2-induced ROS levels and apoptosis in the SSCs. Melatonin also increased mRNA expression of <i>Nrf2, Trx2, NNT, Sirtuin 3 (Sirt3)</i>, and decreased mRNA expression of Txnip, and increased protein expressions of Nrf2, Trx2, NNT thereby increasing activity of the mitochondrial thioredoxin system. In addition, melatonin increased ATP levels.</p><p><strong>Conclusion: </strong>Melatonin increased <i>Trx2</i> expression through the <i>Nrf2</i> pathway. This study suggests that melatonin may protect SSCs from oxidative stress in diseases related to infertility.</p>","PeriodicalId":49224,"journal":{"name":"Cell Journal","volume":"25 11","pages":"741-752"},"PeriodicalIF":2.0,"publicationDate":"2023-11-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10711295/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138812004","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Adipose Tissue-Derived Mesenchymal Stem Cells Alter Metabolites of Brain Cholesterol Homeostasis in An Alzheimer's Model.","authors":"Mehrnaz Karimi Darabi, Zahra Nazeri, Arash Rafeeinia, Seyedeh Pardis Pezeshki, Alireza Kheirollah, Yaghoob Farbood, Maryam Adelipour, Shirin Azizidoost, Maryam Cheraghzadeh","doi":"10.22074/cellj.2023.1999622.1272","DOIUrl":"https://doi.org/10.22074/cellj.2023.1999622.1272","url":null,"abstract":"<p><strong>Objective: </strong>Disruption of cholesterol homeostasis in Alzheimer's disease (AD) plays a crucial role in disease pathogenesis, making it a potential therapeutic target. Mesenchymal stem cells (MSCs) show promise in treating cognitive impairment and provide a novel therapeutic approach. This study aims to investigate the effects of MSCs on specific metabolites associated with brain cholesterol homeostasis in an AD rat model.</p><p><strong>Materials and methods: </strong>In this experimental study, animals were divided into three groups: control, AD, and AD+MSCs. AD was induced using amyloid beta (Aβ) and confirmed through the Morris water maze (MWM) behavioural test and Congo red staining. MSCs were extracted, characterised via flow cytometry, subjected to osteoblast and adipose differentiation, and injected intraventricularly. The cholesterol metabolite levels were measured using gas chromatography-mass spectrometry (GC)-MS and compared among the groups.</p><p><strong>Results: </strong>Treatment with MSCs significantly improved memory function in the AD+MSCs group compared to the AD group and the number of beta-amyloid plaques decreased according to histological assessment. Disturbances in the brain cholesterol metabolites that included desmosterol, 7-dehydrocholesterol, 24S-hydroxycholesterol, 27-hydroxycholesterol and cholesterol were observed in the AD group compared to the control group. Treatment with MSCs resulted in significant alterations in the levels of these metabolites.</p><p><strong>Conclusion: </strong>The findings indicate that MSC therapy has the potential to improve AD by modulating brain cholesterol homeostasis and promoting the differentiation of stem cells into nerve cells. The results emphasize the importance of investigating the role of cholesterol metabolites in the context of MSC therapy to gain deeper insights into underlying mechanisms of the therapeutic efficacy of MSCs in AD.</p>","PeriodicalId":49224,"journal":{"name":"Cell Journal","volume":"25 11","pages":"764-771"},"PeriodicalIF":2.0,"publicationDate":"2023-11-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10711292/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138811997","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Spinal Cord Injury Affects Gene Expression of Transmembrane Proteins in Tissue and Release of Extracellular Vesicle in Blood: In Silico and <i>In Vivo</i> Analysis.","authors":"Yasmin Mirzaalikhan, Nasim Eslami, Amin Izadi, Faezeh Shekari, Sahar Kiani","doi":"10.22074/cellj.2023.2004115.1320","DOIUrl":"https://doi.org/10.22074/cellj.2023.2004115.1320","url":null,"abstract":"<p><strong>Objective: </strong>Spinal cord injury (SCI) can disrupt membrane transmission by affecting transmembrane channels or neurotransmitter release. This study aimed to explore gene expression changes of transmembrane proteins underlying SCI through bioinformatics approaches and confirming in SCI model in rats.</p><p><strong>Materials and methods: </strong>In this experimental study, the differentially expressed genes (DEGs) in acute and subacute SCI were obtained based on microarray data downloaded from the gene expression omnibus (GEO). Transmembrane proteins of DEGs were recognized by using the UniProt annotation and transmembrane helices prediction (TMHMM) methods. The model of SCI was established through a weight-dropping procedure in rats. To confirm the SCI model, hematoxylin and eosin (H and E) staining was performed. Total mRNA was extracted from spinal cord tissues, and the RNA expression profile of some of the significantly changed genes in the previous part that has been confirmed by real-time polymerase chain reaction (PCR). Blood was collected from rats before sacrificing. Extracellular vesicles (EVs) were isolated by high-speed centrifugation from plasma. For the assessment of protein expression, western blotting was used.</p><p><strong>Results: </strong>Based on bioinformatics analysis, we candidated a set of membrane proteins in SCI's acute and sub-acute phases, and confirmed significant upregulation in <i>Grm1, Nrg1, CD63, Enpp3,</i>and <i>Cxcr4</i> between the acute and control groups and downregulation in Enpp3 between acute and subacute groups at the RNA level. Considering CD63 as an EV marker, we examined the protein expression of <i>CD9</i> and <i>CD63</i> in the plasma-derived EVs, and CD9 has significant expression between acute and control groups. We also demonstrate no significant <i>CD63</i> and <i>Cxcr4</i> expressions between groups.</p><p><strong>Conclusion: </strong>Our results provide new insight into the relationship between candidate transmembrane protein expression and different stages of SCI using in-silico approaches. Also, results show the release of EVs in blood in each group after SCI helping enlarge strategies to enhance recovery following SCI.</p>","PeriodicalId":49224,"journal":{"name":"Cell Journal","volume":"25 11","pages":"772-782"},"PeriodicalIF":2.0,"publicationDate":"2023-11-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10711288/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138812013","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Association of <i>MGLL</i> Intronic C>T Single Nucleotide Polymorphism (rs782440) with Borderline Personality Disorder: A Case-Control Study.","authors":"Nazanin Hatami Bavarsad, Leila Jahangard, Masood Saidijam, Seyed Asaad Karimi, Ali Reza Soltanian, Elahe Shahriari, Saeid Afshar, Abdolrahman Sarihi","doi":"10.22074/cellj.2023.2004323.1321","DOIUrl":"https://doi.org/10.22074/cellj.2023.2004323.1321","url":null,"abstract":"<p><strong>Objective: </strong>From the perspective of etiology, borderline personality disorder (BPD) is a multifactorial and complex disorder, hence our understanding about the molecular basis and signaling of this disorder is extremely limited. The purpose of this study was evaluating the relationship between BPD and the Monoacylglycerol lipase <i>(MGLL)</i> polymorphism rs782440 in the population of Hamadan, Iran.</p><p><strong>Materials and methods: </strong>In this case-control study, 106 participants including 53 patients with BPD and 53 healthy control subjects were selected by psychiatrists in the Department of Psychiatry at Farshchian Sina Hospital in Hamadan. The BPD patients were selected based on the Diagnostic and Statistical Manual of Mental Disorders <i>(DSM-5)</i> form for diagnosing BPD patients. For genotyping, polymerase chain reaction (PCR) was used to amplify the desired region including the <i>(MGLL)</i> intronic C>T single nucleotide polymorphism (SNP) (rs782440) and afterward the amplicon was sequenced using the Sanger sequencing method. To determine the genotype of these patients, their sequences were aligned with the reference sequence of MGLL through the CLC genomic workbench software.</p><p><strong>Results: </strong>The results indicated that the frequency of TT in comparison to the CC genotype was significantly different (P=0.003) and the risk of BPD in change from the TT genotype to CC genotype was increased by 6.679%. Regarding the frequency of allele in this group, no significant difference was observed.</p><p><strong>Conclusion: </strong>This paper, has studied and reports for the first time, the association between <i>MGLL</i> SNP (rs782440) with BPD. The findings of the current research revealed that the TT genotype increases the risk of BPD compared to the CC genotype. Considering the lack of a suitable diagnostic biomarker for BPD, using this potential biomarker in the near future can be promising.</p>","PeriodicalId":49224,"journal":{"name":"Cell Journal","volume":"25 11","pages":"783-789"},"PeriodicalIF":2.0,"publicationDate":"2023-11-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10711293/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138812001","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Annexin A7 and Its Related Protein Suppressor of Death Domains Regulates Migration and Proliferation of Hca-P Cells.","authors":"Shaoqing Wang, Qingyang Bai, Xiuwen Yu, Feng Gao, Yurong Sun, Xianyan Wang","doi":"10.22074/cellj.2023.559724.1108","DOIUrl":"https://doi.org/10.22074/cellj.2023.559724.1108","url":null,"abstract":"<p><strong>Objective: </strong>This study was to investigate whether annexin A7 (AnnexinA7, <i>ANXA7</i>) and its co-related protein tumor cell death domain silencer [suppressor of death domains (SODD)] regulates the migratory phenotype of liver cancer cells.</p><p><strong>Materials and methods: </strong>In this experimental study, expression of <i>ANXA7</i> in Hca-P cells, <i>PANXA7</i> downregulated cells and <i>PANXA7</i> unrelated sequence cells was detected by real-time quantitative polymerase chain reaction (PCR) at mRNA level and western blotting at protein level. Transwell migration and invasion assays were performed to determine the migratory phenotype.</p><p><strong>Results: </strong>After inhibition of <i>ANXA7</i> expression, expression of SODD protein was also significantly decreased (P<0.05). Transwell cell transfer experiments showed that number of tumor cells that penetrated into the cell membrane was significantly reduced after <i>ANXA7</i> silencing (P<0.05). Transwell cell invasion assay showed that number of tumor cells penetrating into Matrigel was significantly reduced after <i>ANXA7</i> down-regulation (P<0.05). The CCK8 assay was measured at 0, 24 and 48 hours, and proliferation rate of <i>PANXA7</i> lower weir cells was slower than that of Hca-P cells and <i>PANXA7</i> non-related sequence cells (P<0.05).</p><p><strong>Conclusion: </strong>SODD expression was decreased with the down-regulation of <i>ANXA7</i>. Down-regulating ANXA7 in Hca-P cells decreased proliferation, migration and invasion of tumor cells.</p>","PeriodicalId":49224,"journal":{"name":"Cell Journal","volume":"25 11","pages":"801-808"},"PeriodicalIF":2.0,"publicationDate":"2023-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10711291/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138811999","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The Effect of Mesenchymal Stem Cells Derived-Conditioned Media in Combination with Oral Anti-Androgenic Drugs on Male Pattern Baldness: An Animal Study.","authors":"Majid Kamali-Dolat Abadi, Gholamhossein Yousefi, Farzaneh Dehghani, Ali Akbar Alizadeh, Abolfazl Jangholi, Mohammad Amin Moadab, Maryam Naseh, Shima Parsa, Golara Nasiri, Negar Azarpira, Mehdi Dianatpour","doi":"10.22074/cellj.2023.2008138.1377","DOIUrl":"https://doi.org/10.22074/cellj.2023.2008138.1377","url":null,"abstract":"<p><strong>Objective: </strong>Androgenetic alopecia (AGA) is a prevalent form of hair loss, mainly caused by follicular sensitivity to androgens. Despite developing different anti-androgen treatment options, the success rate of these treatments has been limited. Using animal models, this study evaluated the therapeutic effects of umbilical cord (UC) stem cell conditioned media (CM) combined with oral anti-androgens for hair regeneration.</p><p><strong>Materials and methods: </strong>In this experimental study, Poloxamer 407 (P407) was used as a drug carrier for subcutaneous testosterone injection. AGA models were treated with oral finasteride, oral flutamide, and CM injections. Samples were thoroughly evaluated and compared using histological, stereological, and molecular analyses.</p><p><strong>Results: </strong>Injecting CM-loaded hydrogel alone or combined with oral intake of anti-androgens improved hair regeneration. These treatments could promote hair growth by inducing hair follicles in the anagen stage and shortening the telogen and catagen phases. Furthermore, the combination treatment led to an upregulation of hair induction gene expression with a downregulation of inflammation genes.</p><p><strong>Conclusion: </strong>Through a reduction in inflammation, injection of CM-loaded hydrogel alone or combined with oral intake of anti-androgens induces the hair cell cycle with regeneration in damaged follicles. Hence, this could be a promising therapeutic method for AGA patients.</p>","PeriodicalId":49224,"journal":{"name":"Cell Journal","volume":"25 11","pages":"790-800"},"PeriodicalIF":2.0,"publicationDate":"2023-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10711289/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138812018","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Royan Institute First Attempts: Autotransplantation of Vitrified Human Ovarian Tissue in Cancer Patients.","authors":"Naeimeh Sadat Abtahi, Bita Ebrahimi, Firouzeh Ghaffari, Rohollah Fathi, Mojtaba Rezazadeh Valojerdi, Abolfazl Mehdizadehkashi, Sepideh Khodaverdi, Azar Yahyaei, Maziar Faridi","doi":"10.22074/cellj.2023.2000360.1289","DOIUrl":"https://doi.org/10.22074/cellj.2023.2000360.1289","url":null,"abstract":"<p><p>Today, timely diagnosis and therapeutic progress open a road of hope for survival in cancerous patients. Increased knowledge about the various cytotoxic treatment's impacts on ovarian function and fertility has resulted in a surge in the number of patients seeking to preserve their fertility before starting the anti-cancer treatment process. In this regard, embryo cryopreservation can be recommended for fertility preservation when the woman is married and has adequate time for ovarian stimulation. If patients are prepubertal girls or not married women, oocytes or ovarian tissue can be frozen instead to be used in the future. In this regard, the first attempts for ovarian tissue transplantations were conducted in 2016 and in 2019 for two cancerous patients whose ovarian tissue was cryopreserved in the Royan Human Ovarian Tissue Bank (Tehran, Iran). Unfortunately, the transplantations did not result in a live birth.</p>","PeriodicalId":49224,"journal":{"name":"Cell Journal","volume":"25 11","pages":"809-812"},"PeriodicalIF":2.0,"publicationDate":"2023-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10711294/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138812010","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Integrative Bioinformatics Analysis of The Cell Division Cycle and Ribosomal Pathways in The Rat Varicocele: Implications for Drug Discovery.","authors":"Ali Nasr-Esfahani,&nbsp;Ali Valipour Motlagh,&nbsp;Minoo Adib,&nbsp;Kosar Pashaei,&nbsp;Mohammad Hossein Nasr-Esfahani","doi":"10.22074/cellj.2023.2004771.1329","DOIUrl":"10.22074/cellj.2023.2004771.1329","url":null,"abstract":"<p><strong>Objective: </strong>Varicocele is a common cause of male infertility, affecting a substantial proportion of infertile men. Recent studies have employed transcriptomic analysis to identify candidate genes that may be implicated in the pathogenesis of this condition. Accordingly, this study sought to leverage rat gene expression profiling, along with protein-protein interaction networks, to identify key regulatory genes, related pathways, and potentially effective drugs for the treatment of varicocele.</p><p><strong>Materials and methods: </strong>In this in-silico study, differentially expressed genes (DEGs) from the testicular tissue of 3 rats were screened using the edgeR package in R software and the results were compared to 3 rats in the control group. Data was obtained from GSE139447. Setting a -1<LogFC>1 and P<0.05 as cutoff points for statistical significance, up and down-regulated genes were identified. Based on Cytoscape plugins, protein-protein interaction (PPI) networks were drawn, and hub genes were highlighted. ShinyGO was used for pathway enrichment. Finally, effective drugs were identified from the drug database.</p><p><strong>Results: </strong>Among the 1277 DEGs in this study, 677 genes were up-regulated while 600 genes were down-regulated in rats with varicocele compared to the control group. Using protein-protein interaction networks, we identified the top five up-regulated genes and the top five down-regulated genes. Enrichment analysis showed that the up-regulated genes were associated with the cell division cycle pathway, while the down-regulated genes were linked to the ribosome pathway. Notably, our findings suggested that dexamethasone may be a promising therapeutic option for individuals with varicocele.</p><p><strong>Conclusion: </strong>The current investigation indicates that in varicocele the cell division cycle pathway is up-regulated while the ribosome pathway is down-regulated compared to controls. Based on these findings, dexamethasone could be considered a future candidate drug for the treatment of individuals with varicocele.</p>","PeriodicalId":49224,"journal":{"name":"Cell Journal","volume":"25 10","pages":"727-737"},"PeriodicalIF":2.0,"publicationDate":"2023-10-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/e6/da/Cell-J-25-727.PMC10591260.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"49684101","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"CRISPR/Cas9-Mediated Generation of <i>COL7A1</i>-Deficient Keratinocyte Model of Recessive Dystrophic Epidermolysis Bullosa.","authors":"Farzad Alipour,&nbsp;Mana Ahmadraji,&nbsp;Elham Yektadoost,&nbsp;Parvaneh Mohammadi,&nbsp;Hossein Baharvand,&nbsp;Mohsen Basiri","doi":"10.22074/cellj.2023.1989321.1225","DOIUrl":"10.22074/cellj.2023.1989321.1225","url":null,"abstract":"<p><strong>Objective: </strong>Recessive dystrophic epidermolysis bullosa (RDEB) is a genetic skin fragility and ultimately lethal blistering disease caused by mutations in the <i>COL7A1</i> gene which is responsible for coding type VII collagen. Investigating the pathological mechanisms and novel candidate therapies for RDEB could be fostered by new cellular models. The aim of this study was to employ CRISPR/Cas9 technology in the development of immortalized COL7A1-deficient keratinocyte cell lines intended for application as a cellular model for RDEB in <i>ex vivo</i> studies.</p><p><strong>Materials and methods: </strong>In this experimental study, we used transient transfection to express <i>COL7A1</i> -targeting guide RNA (gRNA) and Cas9 in HEK001 immortalized keratinocyte cell line followed by enrichment with fluorescent-activated cell sorting (FACS) via GFP expressing cells (GFP+ HEK001). Homogenous single-cell clones were then isolated, genotyped, and evaluated for type VII collagen expression. We performed a scratch assay to confirm the functional effect of <i>COL7A1</i> knockout.</p><p><strong>Results: </strong>We achieved 46.1% (P<0.001) efficiency of in/del induction in the enriched transfected cell population. Except for 4% of single nucleotide insertions, the remaining in/dels were deletions of different sizes. Out of nine single expanded clones, two homozygous and two heterozygous <i>COL7A1</i>-deficient cell lines were obtained with defined mutation sequences. No off-target effect was detected in the knockout cell lines. Immunostaining and western blot analysis showed lack of type VII collagen (COL7A1) protein expression in these cell lines. We also showed that <i>COL7A1</i>-deficient cells had higher motility compared to their wild-type counterparts.</p><p><strong>Conclusion: </strong>We reported the first isogenic immortalized <i>COL7A1</i>-deficient keratinocyte lines that provide a useful cell culture model to investigate aspects of RDEB biology and potential therapeutic options.</p>","PeriodicalId":49224,"journal":{"name":"Cell Journal","volume":"25 10","pages":"665-673"},"PeriodicalIF":2.0,"publicationDate":"2023-10-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/48/45/Cell-J-25-665.PMC10591263.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"49684100","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Characterization of CAR T Cells Manufactured Using Genetically Engineered Artificial Antigen Presenting Cells.","authors":"Ali Sayadmanesh,&nbsp;Mohamad Azadbakht,&nbsp;Kheirollah Yari,&nbsp;Ali Abedelahi,&nbsp;Hajar Shafaei,&nbsp;Dariush Shanehbandi,&nbsp;Behzad Baradaran,&nbsp;Mohsen Basiri","doi":"10.22074/cellj.2023.2001712.1304","DOIUrl":"10.22074/cellj.2023.2001712.1304","url":null,"abstract":"<p><strong>Objective: </strong>Chimeric antigen receptor (CAR) T cell therapy has recently emerged as a promising approach for the treatment of different types of cancer. Improving CAR T cell manufacturing in terms of costs and product quality is an important concern for expanding the accessibility of this therapy. One proposed strategy for improving T cell expansion is to use genetically engineered artificial antigen presenting cells (aAPC) expressing a membrane-bound anti-CD3 for T cell activation. The aim of this study was to characterize CAR T cells generated using this aAPC-mediated approach in terms of expansion efficiency, immunophenotype, and cytotoxicity.</p><p><strong>Materials and methods: </strong>In this experimental study, we generated an aAPC line by engineering K562 cells to express a membrane-bound anti-CD3 (mOKT3). T cell activation was performed by co-culturing PBMCs with either mitomycin C-treated aAPCs or surface-immobilized anti-CD3 and anti-CD28 antibodies. Untransduced and CD19-CARtransduced T cells were characterized in terms of expansion, activation markers, interferon gamma (IFN-γ) secretion, CD4/CD8 ratio, memory phenotype, and exhaustion markers. Cytotoxicity of CD19-CAR T cells generated by aAPCs and antibodies were also investigated using a bioluminescence-based co-culture assay.</p><p><strong>Results: </strong>Our findings showed that the engineered aAPC line has the potential to expand CAR T cells similar to that using the antibody-based method. Although activation with aAPCs leads to a higher ratio of CD8+ and effector memory T cells in the final product, we did not observe a significant difference in IFN-γ secretion, cytotoxic activity or exhaustion between CAR T cells generated with aAPC or antibodies.</p><p><strong>Conclusion: </strong>Our results show that despite the differences in the immunophenotypes of aAPC and antibody-based CAR T cells, both methods can be used to manufacture potent CAR T cells. These findings are instrumental for the improvement of the CAR T cell manufacturing process and future applications of aAPC-mediated expansion of CAR T cells.</p>","PeriodicalId":49224,"journal":{"name":"Cell Journal","volume":"25 10","pages":"674-687"},"PeriodicalIF":2.0,"publicationDate":"2023-10-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/75/fa/Cell-J-25-674.PMC10591261.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"49684099","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}