scholarly journals TissueGrinder, a novel technology for rapid generation of patient-derived single cell suspensions from solid tumors by mechanical tissue dissociation

2021 ◽  
Author(s):  
Stefan Scheuermann ◽  
Jonas Max Lehmann ◽  
Ramkumar Ramani Mohan ◽  
Christoph Reissfelder ◽  
Felix Rueckert ◽  
...  
Keyword(s):  
Success Rate ◽  
Morphological Characteristics ◽  
Pancreatic Tumors ◽  
Isolated Cells ◽  
Preparation Methods ◽  
Tissue Dissociation ◽  
Tumor Tissues ◽  
Resected Tumor ◽  
Rapid Generation ◽  
Tissue Characteristics

Introduction: Recent advances hold promise of making personalized medicine a step closer to implementation in clinical settings. However, traditional sample preparation methods are not robust and reproducible. In this study, the TissueGrinder, a novel mechanical semi-automated benchtop device, which can isolate cells from tissue in a very fast and enzyme-free way is tested for cell isolation from surgically resected tumor tissues. Methods: 33 surgically resected tumor tissues from various but mainly pancreatic, liver or colorectal origins were processed by both novel TissueGrinder and explant method. An optimized processing program for tumors from pancreatic, liver or colorectal cancer was developed. The viability and morphological characteristics of the isolated cells were evaluated microscopically. Expression of pancreatic cancer markers was evaluated in cells isolated from pancreatic tumors. Finally, the effect of mechanical stress on the cells was evaluated by assessing apoptosis markers via western blotting. Results: TissueGinder was more efficient in isolating cells from tumor tissue with a success rate of 75% when compared to explant method 45% in terms of cell outgrowth six weeks after processing. Cells isolated with TissueGinder had a higher abundance and were more heterogeneous in composition as compared to explant method. Mechanical processing of the cells with TissueGrinder does not lead to apoptosis but causes slight stress to the cells. Discussion: Our results show that TissueGrinder can process solid tumor tissues more rapidly and efficiently and with higher success rate compared to the conventionally used explant method. The results of the study suggest that the TissueGrinder might be a suitable method for obtaining cells, which is important for its application in individualized therapy. Due to the great variance in different tumor entities and the associated individual tissue characteristics, a further development of the dissociation protocol for other types of tumors and normal will be targeted.

DNA nick end labeling: a reliable marker for apoptosis?

1995 ◽  
Vol 53 ◽  
pp. 962-963
Author(s):  
Anne F. Bushnell ◽  
Sarah Webster ◽  
Lynn S. Perlmutter
Keyword(s):  
Cell Death ◽  
Cultured Cells ◽  
Apoptotic Cell ◽  
Morphological Characteristics ◽  
Detection Methods ◽  
Tissue Preparation ◽  
Preparation Methods ◽  
Dna Laddering ◽  
Disease States ◽  
Reliable Marker

Apoptosis, or programmed cell death, is an important mechanism in development and in diverse disease states. The morphological characteristics of apoptosis were first identified using the electron microscope. Since then, DNA laddering on agarose gels was found to correlate well with apoptotic cell death in cultured cells of dissimilar origins. Recently numerous DNA nick end labeling methods have been developed in an attempt to visualize, at the light microscopic level, the apoptotic cells responsible for DNA laddering.The present studies were designed to compare various tissue processing techniques and staining methods to assess the occurrence of apoptosis in post mortem tissue from Alzheimer's diseased (AD) and control human brains by DNA nick end labeling methods. Three tissue preparation methods and two commercial DNA nick end labeling kits were evaluated: the Apoptag kit from Oncor and the Biotin-21 dUTP 3' end labeling kit from Clontech. The detection methods of the two kits differed in that the Oncor kit used digoxigenin dUTP and anti-digoxigenin-peroxidase and the Clontech used biotinylated dUTP and avidinperoxidase. Both used 3-3' diaminobenzidine (DAB) for final color development.

scholarly journals Plasma interleukin-17 and alpha-fetoprotein combination effectively predicts imminent hepatocellular carcinoma occurrence in liver cirrhotic patients

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Kung-Hao Liang ◽  
Ming-Wei Lai ◽  
Yang-Hsiang Lin ◽  
Yu-De Chu ◽  
Chih-Lang Lin ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Characteristic Curve ◽  
Alpha Fetoprotein ◽  
Enzyme Linked Immunosorbent Assay ◽  
Interleukin 17 ◽  
Medical Need ◽  
Tumor Tissues ◽  
Resected Tumor ◽  
Cirrhotic Patients ◽  
A Prospective Study

Abstract Background Predicting imminent hepatocellular carcinoma (HCC) in liver cirrhotic patients is an unmet medical need. We aimed to investigate circulatory biomarkers and their optimum combinations in a prospective study. Methods We investigated plasma interleukin 17 (IL-17) concentrations, quantified using enzyme-linked immunosorbent assay (ELISA), for the prediction of HCC in a large cohort of 404 HCC-naïve liver cirrhotic patients regularly followed after recruitment. Additionally, IL-17 in surgically resected tumor tissues were evaluated using immunohistochemistry staining. Results IL-17 was detected in HCC tissues. The IL-17 concentrations in the peripheral blood do not have correlation with an extensive list of 31 common demographic, metabolic and liver function variables in the cohort of liver cirrhotic patients. Furthermore, patients stratified by IL-17 and alpha-fetoprotein (AFP) showed distinctive cumulative incidence of HCC. Imminent HCC, defined here as HCC occurrence within 1 year, can be predicted by IL-17 alone with an area under the receiver operating characteristic curve [AUC] of 0.762 (P = 0.002). An multivariate analysis showed that age, hepatitis C viral infection, AFP and IL-17 were four independent factors associated with imminent HCC (adjusted P = 0.03, 0.041, 0.024 and 0.008 respectively). An explicit risk score (R) combining the concentrations of two plasma biomarkers, AFP and IL-17, achieved a high AUC of 0.933 (95% confidence interval 0.893–0.972, P < 0.001) in predicting imminent HCC, with 100% sensitivity and 79.9% specificity at the optimum cutoff. The score is defined as: $${\text{R}} = (2.6914)*{\text{IL-17}} + (0.3909)*{\text{AFP}} - (0.80812875)*{\text{IL-17}}^{2} + (0.10288876884)*{\text{IL-17}}^{2} *{\text{AFP}}.$$ R = ( 2.6914 ) ∗ IL-17 + ( 0.3909 ) ∗ AFP - ( 0.80812875 ) ∗ IL-17 2 + ( 0.10288876884 ) ∗ IL-17 2 ∗ AFP . Conclusions The circulatory IL-17 concentration is a predictor of subsequent HCC occurrence in liver cirrhotic patients. The combination of AFP and IL-17 is highly effective in predicting imminent HCC within 1 year.

scholarly journals Metabolic and histochemical characteristics of fat and muscle tissues in homozygous or heterozygous pigs for the body composition QTL located on chromosome 7

2007 ◽  
Vol 30 (3) ◽  
pp. 232-241 ◽  
Author(s):  
Julie Demars ◽  
Juliette Riquet ◽  
Marie-Pierre Sanchez ◽  
Yvon Billon ◽  
Jean-François Hocquette ◽  
...  
Keyword(s):  
Body Composition ◽  
Carcass Composition ◽  
The Body ◽  
Chromosome 7 ◽  
Isolated Cells ◽  
Large White ◽  
Muscle Tissues ◽  
Metabolism Enzymes ◽  
Tissue Characteristics ◽  
Qtl Effects

Quantitative trait loci (QTL) influencing many traits including backfat thickness and carcass composition have been detected on porcine chromosome 7 (SSC7) in an F2 cross between Large White (LW) and Meishan (MS) pigs. However, the genes and controlled pathways underlying the QTL effects on body phenotype remain unknown. This study aimed at investigating the tissue characteristics at metabolic and cellular levels in pigs that were either homozygous or heterozygous for a body composition SSC7 QTL. A backcross pig (BC3) was first progeny tested to confirm its heterozygoty for the SSC7 QTL; results on all offspring ( n = 80) confirmed the QTL effects on body fatness. This boar was then mated with three sows known to be heterozygous for this QTL. In the subset of pigs per genotype, we found that heterozygous LWQTL7/MSQTL7 pigs had smaller adipocytes in backfat, together with a lower basal rate of glucose incorporation into lipids and lower activities of selected lipogenic enzymes in backfat isolated cells, compared with homozygous LWQTL7/LWQTL7 pigs. A higher number of adipocytes was also estimated in backfat of LWQTL7/MSQTL7 animals compared with LWQTL7/LWQTL7 pigs. The SSC7 QTL did not influence oxidative and glycolytic metabolisms of longissimus and trapezius muscles, as estimated by the activities of specific energy metabolism enzymes, or the myofiber type properties. Altogether, this study provides new evidence for an altered adipocyte cellularity in backfat of pigs carrying at least one MS allele for the SSC7 QTL. Some candidate genes known for their functions on adipocyte growth and differentiation are suggested.

Coexistence of rearranged during transfection (RET) variants and activating EGFR mutations with their molecular implications in lung adenocarcinomas.

2017 ◽  
Vol 35 (15_suppl) ◽  
pp. e20610-e20610
Author(s):  
Jeong-Oh Kim ◽  
Jung-Young Shin ◽  
Min Young Kim ◽  
Kyoung Hwa Son ◽  
Chan-Kwon Jung ◽  
...  
Keyword(s):  
Protein Expression ◽  
Lung Adenocarcinoma ◽  
Egfr Mutation ◽  
Fusion Gene ◽  
Signaling Molecules ◽  
Egfr Mutations ◽  
Downstream Signaling ◽  
Tumor Tissues ◽  
Resected Tumor ◽  
Lung Adenocarcinomas

e20610 Background: RET rearrangements have been identified in 1-2% of lung adenocarcinomas. The most common fusion is the KIF5B-RET, the function and roles of the RET fusion oncogene, and its downstream signaling molecules remain unclear. Methods: We constructed a tissue microarray (TMA) comprising 581 resected tumor tissues from lung adenocarcinoma patients and investigated them using FISH with RET break-apart and KIF5B-RET SY translocation probes. NanoString’s nCounter technology was used to assay RETtranscripts. We evaluated the protein expressions of RET and RET-related signaling molecules, including p-AKT and p-ERK, using TMA-based IHC staining. Results: Using FISH, we identified 51 cases (8.8%) of RET variants and 10 cases (1.7%) of KIF5B-RET fusion genes among the 581 cases. RET protein expression was lower in the group harboring KIF5B-RET fusion gene than that in the group harboring a wild type RET gene. We found the activating EGFR mutations in 11 (21.6%) cases of 51 RET variants. For the group with KIF5B-RET fusion gene, the expression of p-ERK was significantly lower in EGFR mutation subgroup with presence of RET protein compared to EGFR mutation subgroup with absence of RET protein. For the group with RET rearrangement, there were significant differences in the expression level of p-AKT (P = 0.028) and, p-ERK protein expression was remarkably increased, especially in cases with no RET protein expression. Conclusions: Taken together, the expression of p-ERK protein was meaningfully increased in the RET variants group regardless of RET protein expression. This result suggests that RET inhibitors combined with ERK inhibitors may be an effective treatment strategy for lung adenocarcinoma patients harboring the RET variants.

The clinical significance of KRAS monitoring in tumor tissues and blood of patients with pancreatic tumor.

2017 ◽  
Vol 35 (4_suppl) ◽  
pp. 334-334 ◽  
Author(s):  
Fumiaki Watanabe ◽  
Koichi Suzuki ◽  
Kosuke Ichida ◽  
Yuji Takayama ◽  
Taro Fukui ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Overall Survival ◽  
Treatment Outcome ◽  
Clinical Significance ◽  
Kras Mutation ◽  
Poor Prognosis ◽  
Pancreatic Tumor ◽  
Pancreatic Tumors ◽  
Tumor Tissues ◽  
Kras Status

334 Background: Monitoring of gene alterations in blood to track circulating tumor DNAs has been attempted for a clinical application. For example, KRAS monitoring in colorectal cancer provides a valuable biomarker for diagnosis and prediction of treatment outcome. While half of of colon cancer has RAS mutation, 90% of pancreatic cancer shows KRAS mutation, suggesting that most of pancreatic cancer is a good candidate for KRAS monitoring. In this study, we elucidated the clinical significance of KRASmonitoring in patients with pancreatic tumor during treatments. Methods: KRAS mutation in tumor tissues was determined by Scorpion ARMS or RASKET methods in 39 patients with pancreatic tumors. KRASmutation in blood (G12D, G12V, G12C, G12A, G12S, G12R, G13D, Q61L, Q61H) were investigated by using droplet digital polymerase chain reaction (ddPCR) in 24 patients treated with chemotherapy. Results: KRAS assessment in tumor tissues showed 33 patients with mutation and 6 patients without mutation. Thirty-three patients with mutation showed significantly poor prognosis of 49% in three years overall survival (OS) as compared with 100% in 6 patients with mutation (p=0.02). KRAS assessment in blood revealed that KRAS mutation was detected in 14 patients, but no detection was seen in 10 patients. Patients harboring KRAS mutation in blood exhibited significantly poor treatment outcome, including 12 patients with progressive disease, as compared with 10 patients without detection of mutation, including 6 patients with any treatment responses (stable disease in 4 patients and partial response in 2 patients, p=0.03). Fourteen patients with mutation in blood displayed poor prognosis of 20% in three years overall survival (OS), comparing to 69% in 10 patients without mutation in blood (p=0.06). Conclusions: KRAS status in tumor tissues was involved in prognosis; in addition, KRAS status in blood was implicated in treatment response of chemotherapy in patients with pancreatic tumor. KRAS monitoring in tumor tissues and blood provides useful information for the treatment strategy including chemotherapy in patients with pancreatic tumor.

scholarly journals A first record of Pestalotiopsis clavispora in Argan mass cutting propagation: Prevalence, prevention and consequences for plant production

2011 ◽  
Vol 90 (3) ◽  
pp. 117-120
Author(s):  
Mustapha Bakry ◽  
Guy Bussières ◽  
Mohammed S. Lamhamedi ◽  
Hank A. Margolis ◽  
Debra C. Stowe ◽  
...  
Keyword(s):  
Mortality Rate ◽  
Success Rate ◽  
Molecular Analysis ◽  
Morphological Characteristics ◽  
First Record ◽  
Plant Production ◽  
Mass Propagation ◽  
Cutting Propagation ◽  
Apical Buds ◽  
First Time

A trial involving the mass propagation of Argania spinosa cuttings was established following two protocols: in mini-bouturathèques without mist and in a greenhouse under mist. Symptoms of petiole necrosis, foliar yellowing and abundant black acervuli were observed under both protocols. These symptoms were responsible for a 90% mortality rate in the mini-bouturathèques while under the mist treatment premature fatal necrosis of the apical buds resulted in 100% mortality. The disease’s causal agent, Pestalotiopsis clavispora, was identified on the basis of its morphological characteristics and by molecular analysis. Alternating weekly treatments of systemic and contact fungicides resulted in a 41% success rate in controlling this pathogen, described for the first time on argan cuttings.

scholarly journals The High Affinity CXCR4 Inhibitor, BL-8040, Impairs the Infiltration, Migration, Viability, and Differentiation of Regulatory T Cells

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 2804-2804
Author(s):  
Michal Abraham ◽  
Inbal Mishalian ◽  
Abi Vainstein ◽  
Liron Shemesh-Darvish ◽  
Ella Sorani ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Death ◽  
Regulatory T Cells ◽  
Cd4 T Cells ◽  
Cancer Metastasis ◽  
Treg Cells ◽  
Pancreatic Tumors ◽  
Cxcr4 Antagonist ◽  
High Affinity ◽  
Tumor Tissues

Abstract Introduction: Regulatory T (Treg) cells, an immunosuppressive subset of CD4+ T cells characterized by the expression of the master transcription factor forkhead box protein P3 (FOXP3), are a component of the immune system with essential roles in maintaining self-tolerance. Treg cells which are indispensable for preventing autoimmunity, also suppress effective tumor immunity (Togashi et al. Nat Rev Clin Oncol 2019) Treg cells abundantly infiltrate into tumor tissues, present in the tumor microenvironment where they promote tumor development and progression by dampening antitumor immune responses. The abundantly infiltrate of Treg cells into tumor tissues is often associated with poor prognosis in cancer patients (Tanaka et al Eur. J. Immunol. 2019). The chemokine receptor CXCR4 and its ligand, stromal cell-derived factor-1 (SDF-1/CXCL12) are critically involved in immune cell trafficking. CXCR4 overexpression, which has been identified in multiple cancer types, also supports cancer metastasis, recurrence and therapeutic resistance. More importantly, CXCR4 was shown to enhance tumor immune evasion by recruiting Treg (Santagata et al. Oncotarget. 2017) Objective: To study the effect of the high affinity CXCR4 antagonist, BL-8040, on the biology of Treg cells. To study how BL-8040 affects the ability of these cells to penetrate into the tumors, their migratory ability, their survival and also the differentiation of naive T cells towards Treg. Methods:C57BL/6 mice bearing LivMet pancreatic tumors and control mice were used for in-vivo study. In-vitro study was done with CD4 +CD25 hiFOXP3 + (Treg) cells which were isolated from fresh whole blood. CD4 +CD25 - cells were served as T conventional cells (Tconv). Differentiation of Treg cells was done from Naïve CD4+ T cells which were isolated from cord blood and stimulated with anti-CD3/CD28, TGF-b, IL-2 with or without BL-8040 for 6 days. Results: When mice bearing pancreatic cancer were treated with BL-8040, we found a significant reduction in the number of infiltrating Treg into the tumor. Following treatment with BL-8040 there was no alteration in the number of Treg in the blood neither in control mice nor in mice bearing tumors. To further understand the mechanism by which BL-8040 effected Treg cells we isolated Treg and Tconv cells and found that Treg cells express lower level of CXCR4, as compared to Tconv (Figure1). Further to, when we compared their motility, we found that Treg migration less efficiently towards CXCL12. Despite this, BL8040 more efficiently suppressed CXCL12 induced migration of Treg compared to Tconv. 100 nM of BL8040 was found to inhibits the migration of 82 % from the Treg compared to only 56.6% of Tconv cells (Figure 2). CXCR4 involves classical pathways of cell survival. In order to study the role of CXCR4 in the viability of Treg, we incubated Treg and Tconv cells in the presence of BL-8040 for 24 hr. We found that Treg cells are more sensitive to BL-8040 treatment with 19.2% of cell death compared to only 3.5% of Tconv cell death (Figure 3). Treg are one of the lineages of T helper (Th) cells which differentiated from naïve CD4 T cells. We found that BL-8040 inhibits the differentiation of naive CD4 T cells toward Treg. 10uM of BL-8040 shows a 5-fold inhibition in Treg differentiation from naïve CD4 T cells (Figure 4). Conclusions: In this work we show that the CXCR4 antagonist, BL-8040, can act as an immunomodulator by affecting the biology of regulatory T cells. BL8040 reduce the amount of infiltrating Treg into the tumors, impaired the migratory capacity of Treg toward CXCL12 and induces their cell death. Furthermore, BL-8040 was found to inhibit the differentiation of naïve CD4 T cells toward Treg. Taking all these together, BL-8040 may therefore improve the anticancer immune response, without impairing the activity of Tconv and thus can potentially serve as an effective immunomodulatory agent for cancer. Figure 1 Figure 1. Disclosures Abraham: Biokine Therapeutics: Current Employment. Vainstein: BioLineRx LTD: Current Employment. Shemesh-Darvish: BioLineRx LTD: Current Employment. Sorani: BioLineRx LTD: Current Employment. Eizenberg: Biokine Therapeutics Ltd: Current Employment. Peled: Biokine Therapeutics Ltd: Current Employment; Gamida Cell: Research Funding.

scholarly journals Interrelation of HPV-infection of endometry carcinoma and its clinical-morphological features

2018 ◽  
Vol 5 (3) ◽  
pp. 75-82
Author(s):  
T. A. Zykova ◽  
T. I. Moiseenko ◽  
E. M. Frantsiyants ◽  
M. A. Vovkochina
Keyword(s):  
Morphological Characteristics ◽  
Tumor Grade ◽  
Hpv Infection ◽  
Disease Stage ◽  
Tissue Samples ◽  
Total Cohort ◽  
First Case ◽  
Tumor Tissues ◽  
The Status ◽  
Formalin Fixed Paraffin Embedded

Background. There are contradictory data on the detection of human papillomavirus (HPV) in the tumor tissue in endometrial cancer (EС). Objective: to assess HPV infection rates in EC tumor tissues and to establish the relationship between the status of HPV infection and tumor morphological characteristics. Materials and methods. 57 formalin-fixed paraffin-embedded (FFPE) tissue samples of EC patients aged 47–78 years were studied. HPV DNAs were found in 54.4 % of samples. Results and conclusion. We did not reveal an association between the HPV tumor status and age, metastasis or invasion depth. However, there was an interdependence between HPV infection and some morphological characteristics of the tumor: its histological type (adenocarcinomas with squamous cell differentiation were HPV-positive 1.8 times more frequent compared to adenocarcinomas without one; in the first case, tumor tissues were more often infected with HPV 16, and in the second case with HPV 18); tumor grade (in the total cohort and in serous-papillary adenocarcinomas, tumors with higher grades were more often HPV-infected: from 0 to 81.8 % and from 0 to 100 % respectively); disease stage (in the total cohort the percentage of HPV-positive tumors in stage II was 2.4 times and in stage III – 1.6 times higher than in stage I, and stage IA tumors were HPV-positive 2.3 times more often than IB tumors); type of tumor growth (tumors with infiltrative growth type were HPV-positive 1.7 times more often than with exophytic growth, and 2.2 times more often than with mixed growth). The achieved results do not allow us to conclude with confidence that HPV is the main tumor forming factor in EC.

scholarly journals Evaluation of a canine transmissible venereal tumour cell line with tumour immunity capacity but without tumorigenic property

2019 ◽  
Vol 63 (2) ◽  
pp. 225-233 ◽  
Author(s):  
Yareellys Ramos Zayas ◽  
Moisés Armides Franco Molina ◽  
Reyes Tamez Guerra ◽  
Cristina Rodríguez Padilla
Keyword(s):  
Cell Line ◽  
Financial Burden ◽  
Morphological Characteristics ◽  
Tumour Cells ◽  
Tumour Cell Line ◽  
Histopathological Analysis ◽  
Isolated Cells ◽  
Tumour Immunity ◽  
Canine Transmissible Venereal Tumour ◽  
Transmissible Venereal Tumour

AbstractIntroduction:Canine transmissible venereal tumour (CTVT) is a sexually transmitted tumour affecting dogs worldwide, imposing a financial burden on dog owners. A stable culture cell line in continuous passages for >18 months has only been achieved once. The present study investigated a stable CTVT cell line isolated from a bitch and its potential as a vaccine.Material and Methods:A biopsy from a 2-year-old mongrel bitch with CTVT was obtained for histopathological confirmation and isolation of tumour cells. The isolated cells were cultured to passage 55 and characterised by flow cytometry, with karyotyping by GTG-banding and by PCR detection of myc S-2 and LINE AS1. The isolated CTVT cell line was also used as a preventive vaccine in a canine model.Results:Histopathological analysis of the isolated tumour cells revealed typical CTVT characteristics. Constant proliferation and stable morphological characteristics were observed during culture. Phenotypic analysis determined the expression of HLA-DR+, CD5.1+, CD14+, CD45+, CD83+, CD163+, and Ly-6G-Ly-6C+. GTG-banding revealed a mean of 57 chromosomes in the karyotype with several complex chromosomal rearrangements. LINE-c-myc insertion in the isolated CTVT cell line at 550 bp was not detected. However, a 340-bp band was amplified. Isolated CTVT cell line inoculation at a concentration of 1×108did not induce tumour growth in bitches, nor did a challenge with primary CTVT cells.Conclusion:The present study successfully identified and isolated a stable CTVT cell line that may be useful in CTVT prevention.

scholarly journals Dissecting cell type-specific metabolism in pancreatic ductal adenocarcinoma

2020 ◽  
Author(s):  
Allison N. Lau ◽  
Zhaoqi Li ◽  
Laura V. Danai ◽  
Anna M. Westermark ◽  
Alicia M. Darnell ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Cancer Cells ◽  
Heterogeneous Systems ◽  
Cell Types ◽  
Ductal Adenocarcinoma ◽  
Pancreatic Tumors ◽  
Specific Cell ◽  
Isolated Cells ◽  
Pancreatic Cancer Cells ◽  
Turn Over

AbstractTumors are composed of many different cell types including cancer cells, fibroblasts, and immune cells. Dissecting functional metabolic differences between various cell types within a mixed population can be limited by the rapid turnover of metabolites relative to the time needed to isolate cells. To overcome this challenge, we traced isotope-labeled nutrients into macromolecules that turn over more slowly than metabolites. This approach was used to assess differences between cancer cell and fibroblast metabolism in pancreatic cancer organoid-fibroblast co-cultures and in pancreatic tumors. In these contexts, we find pancreatic cancer cells exhibit increased pyruvate carboxylation relative to fibroblasts, and that this flux depends on both pyruvate carboxylase and malic enzyme 1 activity. Consequently, expression of both enzymes in cancer cells is necessary for organoid and tumor growth, demonstrating that dissecting the metabolism of specific cell populations within heterogeneous systems can identify dependencies that may not be evident from studying isolated cells in culture or bulk tumor tissue.

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