Solid malignancies have been speculated to depend on cancer stem cells (CSCs) for expansion and relapse after therapy. Here we report on quantitative analyses of lineage tracing data from primary colon cancer xenograft tissue to assess CSC functionality in a human solid malignancy. The temporally obtained clone size distribution data support a model in which stem cell function in established cancers is not intrinsically, but is entirely spatiotemporally orchestrated. Functional stem cells that drive tumour expansion predominantly reside at the tumour edge, close to cancer-associated fibroblasts. Hence, stem cell properties change in time depending on the cell location. Furthermore, although chemotherapy enriches for cells with a CSC phenotype, in this context functional stem cell properties are also fully defined by the microenvironment. To conclude, we identified osteopontin as a key cancer-associated fibroblast-produced factor that drives in situ clonogenicity in colon cancer.

Cancer evolution is predominantly studied by focusing on differences in the genetic characteristics of malignant cells within tumors. However, the spatiotemporal dynamics of clonal outgrowth that underlie evolutionary trajectories remain largely unresolved. Here, we sought to unravel the clonal dynamics of colorectal cancer (CRC) expansion in space and time by using a color-based clonal tracing method. This method involves lentiviral red-green-blue (RGB) marking of cell populations, which enabled us to track individual cells and their clonal outgrowth during tumor initiation and growth in a xenograft model. We found that clonal expansion largely depends on the location of a clone, as small clones reside in the center and large clones mostly drive tumor growth at the border. These dynamics are recapitulated in a computational model, which confirms that the clone position within a tumor rather than cell-intrinsic features, is crucial for clonal outgrowth. We also found that no significant clonal loss occurs during tumor growth and clonal dispersal is limited in most models. Our results imply that, in addition to molecular features of clones such as (epi-)genetic differences between cells, clone location and the geometry of tumor growth are crucial for clonal expansion. Our findings suggest that either microenvironmental signals on the tumor border or differences in physical properties within the tumor, are major contributors to explain heterogeneous clonal expansion. Thus, this study provides further insights into the dynamics of solid tumor growth and progression, as well as the origins of tumor cell heterogeneity in a relevant model system.

There are currently few recommendations on how to assess inter-arm blood pressure (BP) differences. The authors compared simultaneous with sequential measurement on mean BP, inter-arm BP differences, and within-visit reproducibility in 240 patients stratified according to age (<50 or ≥60 years) and BP (<140/90 mm Hg or ≥140/90 mm Hg). Three simultaneous and three sequential BP measurements were taken in each patient. Starting measurement type and starting arm for sequential measurements were randomized. Mean BP and inter-arm BP differences of the first pair and reproducibility of inter-arm BP differences of the first and second pair were compared between both methods. Mean systolic BP was 1.3AE7.5 mm Hg lower during sequential compared with simultaneous measurement (P<.01). However, the first sequential measurement was on average higher than the second, suggesting an order effect. Absolute systolic inter-arm BP differences were smaller on simultaneous (6.2AE6.7/3.3AE3.5 mm Hg) compared with sequential BP measurement (7.8AE7.3/4.6AE5.6 mm Hg, P<.01 for both). Within-visit reproducibility was identical (both r=0.60). Simultaneous measurement of BP at both arms reduces order effects and results in smaller inter-arm BP differences, thereby potentially reducing unnecessary referral and diagnostic procedures. J Clin Hypertens (Greenwich). 2013;15:839-844. ª2013 Wiley Periodicals, Inc.Inter-arm blood pressure (BP) differences have been established since the early 20s of the last century.1 The importance of assessing inter-arm BP differences is to prevent underestimation and undertreatment of hypertension because the arm with the highest BP should be taken as a reference. Therefore, guidelines on BP measurement recommend bilateral BP measurement at a patient's first visit.2,3 In addition, large inter-arm BP differences in systolic BP (SBP) may indicate the presence of atherosclerotic plaques and other vascular occlusive diseases and are associated with increased cardiovascular risk. 4,5 A recent meta-analysis of studies assessing the inter-arm BP differences showed that a SBP difference ≥15 mm Hg was associated with an increased risk of cardiovascular mortality.6 This is in line with another recent prospective study in hypertensive primary care patients, which showed that a BP difference of ≥10 mm Hg was an independent predictor of cardiovascular events and all-cause mortality after 10 years of follow-up.

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Summary The tissue dynamics that govern maintenance and regeneration of the pancreas remain largely unknown. In particular, the presence and nature of a cellular hierarchy remains a topic of debate. Previous lineage tracing strategies in the pancreas relied on specific marker genes for clonal labeling, which left other populations untested and failed to account for potential widespread phenotypical plasticity. Here we employed a tracing system that depends on replication-induced clonal marks. We found that, in homeostasis, steady acinar replacement events characterize tissue dynamics, to which all acinar cells have an equal ability to contribute. Similarly, regeneration following pancreatitis was best characterized by an acinar self-replication model because no evidence of a cellular hierarchy was detected. In particular, rapid regeneration in the pancreas was found to be driven by an accelerated rate of acinar fission-like events. These results provide a comprehensive and quantitative model of cell dynamics in the exocrine pancreas.

Effective treatments for pancreatic ductal adenocarcinoma (PDAC) are lacking, and targeted agents have demonstrated limited efficacy. It has been speculated that a rare population of cancer stem cells (CSCs) drives growth, therapy resistance, and rapid metastatic progression in PDAC. These CSCs demonstrate high clonogenicity in vitro and tumorigenic potential in vivo. However, their relevance in established PDAC tissue has not been determined. Here, we use marker-independent stochastic clonal labeling, combined with quantitative modeling of tumor expansion, to uncover PDAC tissue growth dynamics. We find that in contrast to the CSC model, all PDAC cells display clonogenic potential in situ. Furthermore, the proximity to activated cancer-associated fibroblasts determines tumor cell clonogenicity. This means that the microenvironment is dominant in defining the clonogenic activity of PDAC cells. Indeed, manipulating the stroma by Hedgehog pathway inhibition alters the tumor growth mode, revealing that tumor-stroma crosstalk shapes tumor growth dynamics and clonal architecture.

Pancreatic ductal adenocarcinoma (PDAC) is characterized by abundant stroma, the main cellular constituents of which are cancer-associated fibroblasts (CAFs). Stromatargeting agents have been proposed to improve the poor outcome of current treatments. However, clinical trials using these agents showed disappointing results.Abbreviations: CA19.9, carbohydrate antigen 19-9; CAF, cancer-associated fibroblast; CXCL12, platelet factor 12; ELISA, enzyme-linked immunosorbent assay; FABP4, fatty acid binding protein 4; GEO, Gene Expression Omnibus; hg38, homo sapiens (human) genome assembly GRCh38; HR, hazard ratio; IL6, interleukin-6; KD, knock-down; MEF, mouse embryonic fibroblast; MHC-II, antigen-presenting major histocompatibility complex class II; mm10, Mus musculus (house mouse) genome assembly GRCm38; OGN, human osteoglycin protein (gene; OGN); Ogn, mouse osteoglycin protein (gene; Ogn); PDAC, pancreatic ductal adenocarcinoma; PDGFR, platelet-derived growth factor receptor; PDX, patient-derived xenograft; RT-qPCR, quantitative reverse transcription-PCR.

Cell generation and renewal are essential processes to develop, maintain, and regenerate tissues. New cells can be generated from immature cell types, such as stem-like cells, or originate from more differentiated pre-existing cells that self-renew or transdifferentiate. The adult pancreas is a dormant organ with limited regeneration capacity, which complicates studying these processes. As a result, there is still discussion about the existence of stem cells in the adult pancreas. Interestingly, in contrast to the classical stem cell concept, stem cell properties seem to be plastic, and, in circumstances of injury, differentiated cells can revert back to a more immature cellular state. Importantly, deregulation of the balance between cellular proliferation and differentiation can lead to disease initiation, in particular to cancer formation. Pancreatic ductal adenocarcinoma (PDAC) is a lethal disease with a 5-year survival rate of only ~9%. Unfortunately, metastasis formation often occurs prior to diagnosis, and most tumors are resistant to current treatment strategies. It has been proposed that a specific subpopulation of cells, i.e., cancer stem cells (CSCs), are responsible for tumor expansion, metastasis formation, and therapy resistance. Understanding the underlying mechanisms of pancreatic stem cells during homeostasis and injury might lead to new insights to understand the role of CSCs in PDAC. Therefore, in this review, we present an overview of the current literature regarding the stem cell dynamics in the pancreas during health and disease. Furthermore, we highlight the influence of the tumor microenvironment on the growth behavior of PDAC.