High-Quality Biobanks: Pivotal Assets for Reproducibility of OMICS-Data in Biomedical Translational Research.

Human biospecimen samples (HBS) and associated data stored in biobanks (also called "biotrusts," "biorepositories," or "biodistributors") are very critical resources for translational research. As HBS quality is decisive to the reproducibility of research results, biobanks are also key assets for new developments in precision medicine. Biobanks are more than infrastructures providing HBS and associated data. Biobanks have pioneered in identifying and standardizing sources of preanalytical variations in HBS, thus paving the way for the current biospecimen science. To achieve this milestone, biobankers have successively assumed the role of "detective," and then "architect," to identify new detrimental impact of preanalytical variables on the tissue integrity. While standardized methods in omics are required to be practiced throughout research communities, the accepted best practices and standards on biospecimen handling are generally not known nor applied by researchers. Therefore, it is mandatory to raise the awareness within omics communities regarding not only the basic concepts of collecting, storing, and utilizing HBS today, but also to suggest insights on biobanking in the cancer omics context.

Aldehyde dehydrogenase activity of Wharton jelly mesenchymal stromal cells: isolation and characterization.

Mesenchymal stromal cells (MSCs) are promising tools in regenerative medicine and targeted therapies. Although different origins have been described, there is still huge need to find a valuable source harboring specific subpopulations of MSCs with precise therapeutic functions. Here, we isolated by fluorescence activated cell sorting technique, two populations of Wharton's jelly (WJ)-MSCs based on their aldehyde dehydrogenase (ALDH) activity. Two different ALDH activities (low vs. high) were thus observed. We then analyzed their gene expression profile for stemness, phenotype, response to hypoxia, angiogenesis, hematopoietic support, immunomodulation and multilineage differentiation abilities (osteogenesis, adipogenesis, and chondrogenesis). According to ALDH activity, many differences in the mRNA expression of these populations were noticed. In conclusion, we provide evidences that WJ harbors two distinct populations of MSCs with different ALDH activity. These populations seem to display specific functional competences that may be interesting for concise therapeutic applications.

Interactions of spatial strategies producing generalization gradient and blocking: A computational approach.

We present a computational model of spatial navigation comprising different learning mechanisms in mammals, i.e., associative, cognitive mapping and parallel systems. This model is able to reproduce a large number of experimental results in different variants of the Morris water maze task, including standard associative phenomena (spatial generalization gradient and blocking), as well as navigation based on cognitive mapping. Furthermore, we show that competitive and cooperative patterns between different navigation strategies in the model allow to explain previous apparently contradictory results supporting either associative or cognitive mechanisms for spatial learning. The key computational mechanism to reconcile experimental results showing different influences of distal and proximal cues on the behavior, different learning times, and different abilities of individuals to alternatively perform spatial and response strategies, relies in the dynamic coordination of navigation strategies, whose performance is evaluated online with a common currency through a modular approach. We provide a set of concrete experimental predictions to further test the computational model. Overall, this computational work sheds new light on inter-individual differences in navigation learning, and provides a formal and mechanistic approach to test various theories of spatial cognition in mammals.

Foreskin-derived mesenchymal stromal cells with aldehyde dehydrogenase activity: isolation and gene profiling.

BACKGROUND: Mesenchymal stromal cells (MSCs) become an attractive research topic because of their crucial roles in tissue repair and regenerative medicine. Foreskin is considered as a valuable tissue source containing immunotherapeutic MSCs (FSK-MSCs). RESULTS: In this work, we used aldehyde dehydrogenase activity (ALDH) assay (ALDEFLUOR™) to isolate and therefore characterize subsets of FSK-MSCs. According to their ALDH activity, we were able to distinguish and sort by fluorescence activated cell sorting (FACS) two subsets of FSK-MSCs (referred as ALDH+ and ALDH-). Consequently, these subsets were characterized by profiling the gene expression related to the main properties of MSCs (proliferation, response to hypoxia, angiogenesis, phenotype, stemness, multilineage, hematopoiesis and immunomodulation). We thus demonstrated by Real Time PCR several relevant differences in gene expression based on their ALDH activity. CONCLUSION: Taken together, this preliminary study suggests that distinct subsets of FSK-MSCs with differential gene expression profiles depending of ALDH activity could be identified. These populations could differ in terms of biological functionalities involving the selection by ALDH activity as useful tool for potent therapeutic applications. However, functional studies should be conducted to confirm their therapeutic relevance.

Aldehyde Dehydrogenase Activity in Adipose Tissue: Isolation and Gene Expression Profile of Distinct Sub-population of Mesenchymal Stromal Cells.

Thanks to their relative abundance and easier collection, adipose tissue (AT) is considered an alternative source for the isolation of mesenchymal stromal cells (MSCs). MSCs have great therapeutic values and are thus under investigations for several clinical indications such as regenerative medicine and immunomodulation. In this work, we aimed to identify, isolate and characterize AT-MSCs based on their aldehyde dehydrogenase (ALDH) activity known to be a classical feature of stem cells. FACS technology allowed to isolate two different populations of AT-MSCs according to their ALDH activity (referred as ALDH+ and ALDH-). Depending on their ALDH activity, the transcriptome analysis of both cell populations demonstrated a differential pattern of genes related to the main properties of MSCs (proliferation, response to hypoxia, angiogenesis, phenotype, stemness, multilineage, hematopoiesis, immunomodulation). Based on these profiling, both AT-MSC populations could differ in terms of biological responses and functionalities. Collectively, the use of ALDH for isolating and identifying sub-populations of MSCs with specific gene profile may represent an alternative method to provide solutions for targeted therapeutic applications.

Isolation and Characterization of Bone Marrow Mesenchymal Stromal Cell Subsets in Culture Based on Aldehyde Dehydrogenase Activity.

Mesenchymal stromal cells (MSCs) have particular properties that allow their use as therapeutic strategies for several cell-based applications. Historically, bone marrow (BM)-MSCs are isolated by culture adherence since specific cell surface markers are yet to be developed. This original work aimed to identify and characterize isolating expanded BM-MSCs based on their aldehyde dehydrogenase (ALDH) activity known to be a hallmark of stem cells and relevant for their isolation. We thus isolated by fluorescence-activated cell sorting technology two functionally different populations of BM-MSCs depending on their ALDH activity (ALDH+ and ALDH-). Transcriptome analysis and profiling clearly demonstrated that both populations of BM-MSCs present distinct pattern of genes related to the main properties of MSCs (proliferation, response to hypoxia, angiogenesis, phenotype, stemness, multilineage, hematopoiesis, immunomodulation) in an ALDH activity dependent manner. Both BM-MSC populations look to significantly differ in terms of biological responses and functionalities. More functional analyses are needed to understand and characterize the properties of these ALDH populations. Collectively, our results highlight ALDH activity as a potential feature for isolating and segregating functional and/or competent subset of BM-MSC populations, which may account for better and more efficient therapeutic issue.

Pharmacological enrollment of aldehyde dehydrogenase modulators to assist treating ischemia reperfusion-induced intestinal injury: is there a gap to be bridged?

This commentary highlights the research presented by Zhu et al. [1]. In this issue of the Clinical Science, the authors evaluated the protective effect of Alda-1 (a novel class of small molecule aldehyde dehydrogenase (ALDH2) activators) in the intestinal ischemia reperfusion (IR) injury. Remarkably, enhancing the ADLH2 activity by the use of Alda-1 can ameliorate several deleterious effects related to aldehydes, and may provide a better protection against an injury preestablished by IR. Together, an innovative metabolic strategy for treating patients with IR injury could be the use of ALDH modulators in a near future.

Infliximab and Dexamethasone Attenuate the Ductular Reaction in Mice.

Chronic hepatic injury is accompanied by a ductular response that is strongly correlated with disease severity and progression of fibrosis. To investigate whether anti-inflammatory drugs can modulate the ductular response, we treated mice suffering from a steatotic or cholestatic injury with anti-TNF-α antibodies (Infliximab) or glucocorticoids (Dexamethasone). We discovered that Dexamethasone and Infliximab can both modulate the adaptive remodeling of the biliary architecture that occurs upon liver injury and limit extracellular matrix deposition. Infliximab treatment, at least in these steatotic and cholestatic mouse models, is the safer approach since it does not increase liver injury, allows inflammation to take place but inhibits efficiently the ductular response and extracellular matrix deposition. Infliximab-based therapy could, thus, still be of importance in multiple chronic liver disorders that display a ductular response such as alcoholic liver disease or sclerosing cholangitis.

Macrophage Depletion Attenuates Extracellular Matrix Deposition and Ductular Reaction in a Mouse Model of Chronic Cholangiopathies.

Chronic cholangiopathies, such as primary and secondary sclerosing cholangitis, are progressive disease entities, associated with periportal accumulation of inflammatory cells, encompassing monocytes and macrophages, peribiliary extracellular matrix (ECM) deposition and ductular reaction (DR). This study aimed to elucidate the relevance of macrophages in the progression of chronic cholangiopathies through macrophage depletion in a 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) mouse model. One group of mice received a single i.p. injection of Clodronate encapsulated liposomes (CLOLipo) at day 7 of a 14 day DDC treatment, while control animals were co-treated with PBSLipo instead. Mice were sacrificed after 7 or respectively 14 days of treatment for immunohistochemical assessment of macrophage recruitment (F4/80), ECM deposition (Sirius Red, Laminin) and DR (CK19). Macrophage depletion during a 14 day DDC treatment resulted in a significant inhibition of ECM deposition. Porto-lobular migration patterns of laminin-rich ECM and ductular structures were significantly attenuated and a progression of DR was effectively inhibited by macrophage depletion. CLOLipo co-treatment resulted in a confined DR to portal regions without amorphous cell clusters. This study suggests that therapeutic options selectively directed towards macrophages might represent a feasible treatment for chronic cholestatic liver diseases.

Policing of gut microbiota by the adaptive immune system.

The intestinal microbiota is a large and diverse microbial community that inhabits the intestine, containing about 100 trillion bacteria of 500-1000 distinct species that, collectively, provide benefits to the host. The human gut microbiota composition is determined by a myriad of factors, among them genetic and environmental, including diet and medication. The microbiota contributes to nutrient absorption and maturation of the immune system. As reciprocity, the host immune system plays a central role in shaping the composition and localization of the intestinal microbiota. Secretory immunoglobulins A (sIgAs), component of the adaptive immune system, are important player in the protection of epithelium, and are known to have an important impact on the regulation of microbiota composition. A recent study published in Immunity by Fransen and colleagues aimed to mechanistically decipher the interrelationship between sIgA and microbiota diversity/composition. This commentary will discuss these important new findings, as well as how future therapies can ultimately benefit from such discovery.

The plastic cellular states of liver cells: Are EpCAM and Lgr5 fit for purpose?

Adult liver cells have been considered restricted regarding their fate and lineage potential. That is, hepatocytes have been thought able only to generate hepatocytes and duct cells, only duct cells. While this may be the case for the majority of scenarios in a state of quiescence or homeostasis, evidence suggests that liver cells are capable of interconverting between cellular states of distinct phenotypic traits. This interconversion or plasticity had been suggested by classical studies using cellular markers, but recently lineage tracing approaches have proven that cells are highly plastic and retain an extraordinary ability to respond differently to normal tissue homeostasis, to tissue repair, or when challenged to expand ex vivo or to differentiate upon transplantation. Stemness, as "self-renewal and multipotency," seems not to be limited to a particular cell type but rather to a cellular state in which cells exhibit a high degree of plasticity and can move back and forth in different phenotypic states. For instance, upon damage cells can dedifferentiate to acquire stem cell potential that allows them to self-renew, repopulate a damaged tissue, and then undergo differentiation. In this review, we will discuss the evidence on cellular plasticity in the liver, focusing our attention on two markers, epithelial cell adhesion molecule and leucine-rich repeat-containing G protein-coupled receptor 5, which identify cells with stem cell potential. (Hepatology 2016;64:652-662).

Osteopontin is a proximal effector of leptin-mediated non-alcoholic steatohepatitis (NASH) fibrosis.

INTRODUCTION: Liver fibrosis develops when hepatic stellate cells (HSC) are activated into collagen-producing myofibroblasts. In non-alcoholic steatohepatitis (NASH), the adipokine leptin is upregulated, and promotes liver fibrosis by directly activating HSC via the hedgehog pathway. We reported that hedgehog-regulated osteopontin (OPN) plays a key role in promoting liver fibrosis. Herein, we evaluated if OPN mediates leptin-profibrogenic effects in NASH. METHODS: Leptin-deficient (ob/ob) and wild-type (WT) mice were fed control or methionine-choline deficient (MCD) diet. Liver tissues were assessed by Sirius-red, OPN and αSMA IHC, and qRT-PCR for fibrogenic genes. In vitro, HSC with stable OPN (or control) knockdown were treated with recombinant (r)leptin and OPN-neutralizing or sham-aptamers. HSC response to OPN loss was assessed by wound healing assay. OPN-aptamers were also added to precision-cut liver slices (PCLS), and administered to MCD-fed WT (leptin-intact) mice to determine if OPN neutralization abrogated fibrogenesis. RESULTS: MCD-fed WT mice developed NASH-fibrosis, upregulated OPN, and accumulated αSMA+ cells. Conversely, MCD-fed ob/ob mice developed less fibrosis and accumulated fewer αSMA+ and OPN+ cells. In vitro, leptin-treated HSC upregulated OPN, αSMA, collagen 1α1 and TGFß mRNA by nearly 3-fold, but this effect was blunted by OPN loss. Inhibition of PI3K and transduction of dominant negative-Akt abrogated leptin-mediated OPN induction, while constitutive active-Akt upregulated OPN. Finally, OPN neutralization reduced leptin-mediated fibrogenesis in both PCLS and MCD-fed mice. CONCLUSION: OPN overexpression in NASH enhances leptin-mediated fibrogenesis via PI3K/Akt. OPN neutralization significantly reduces NASH fibrosis, reinforcing the potential utility of targeting OPN in the treatment of patients with advanced NASH.

Laminin-332 sustains chemoresistance and quiescence as part of the human hepatic cancer stem cell niche.

BACKGROUND & AIMS: Cancer stem cells (CSCs) are thought to be persistent in tumours due to their chemoresistance and to cause relapse and metastasis. Hepatic carcinomas displaying hepatic progenitor cell (HPC) features have been associated with a poor prognosis, though it remains unclear how CSCs relate to these different histological subtypes. METHODS: Candidate CSCs were isolated using the side population (SP) technique from primary tissue samples diagnosed as keratin(K)19-negative or -positive hepatocellular carcinoma (HCC) or as combined hepatocellular/cholangiocarcinoma and analysed for gene and protein expression. The effect of laminin-332 was analysed in vitro by using HCC cell lines and in vivo using a xenograft mouse model. RESULTS: The size of the SP correlated with the degree of HPC features found in human hepatic cancer, and also showed an elevated mRNA expression of biliary/HPC markers and the extracellular matrix marker LAMC2, the gene encoding the laminin γ2-chain. Immunopositivity for the γ2-chain of laminin-332 was seen in the extracellular matrix surrounding small HPC-like tumour cells with a low proliferation rate. In vitro, laminin-332 increased K19 expression, phosphorylated mTOR and decreased phospho-histone H3 expression, indicating reduced cell mitosis. The effect of laminin-332 was enhanced upon mTORC1 inhibition and diminished when inhibiting mTORC1+C2. Resistance to doxorubicin and sorafenib treatment, and the SP fraction increased in the coated condition. In vivo, laminin-332 reduced tumour growth and sustained K19 expression. CONCLUSIONS: In this study we identified a prominent role for laminin-332 as part of the specialised CSC niche in maintaining and supporting cell 'stemness', which leads to chemoresistance and quiescence.

Advances in hepatic stem/progenitor cell biology.

The liver is famous for its strong regenerative capacity, employing different modes of regeneration according to type and extent of injury. Mature liver cells are able to proliferate in order to replace the damaged tissue allowing the recovery of the parenchymal function. In more severe scenarios hepatocytes are believed to arise also from a facultative liver progenitor cell compartment. In human, severe acute liver failure and liver cirrhosis are also both important clinical targets in which regeneration is impaired, where the role of this stem cell compartment seems more convincing. In animal models, the current state of ambiguity regarding the identity and role of liver progenitor cells in liver physiology dampens the enthusiasm for the potential use of these cells in regenerative medicine. The aim of this review is to give the basics of liver progenitor cell biology and discuss recent results vis-à-vis their identity and contribution to liver regeneration.

Next generation of ALDH substrates and their potential to study maturational lineage biology in stem and progenitor cells.

High aldehyde dehydrogenase (ALDH) activity is a feature of stem cells from normal and cancerous tissues and a reliable universal marker used to isolate them. There are numerous ALDH isoforms with preferred substrate specificity variably expressed depending on tissue, cell type, and organelle and cell status. On the other hand, a given substrate may be metabolized by several enzyme isoforms. Currently ALDH activity is evidenced by using Aldefluor, a fluorescent substrate likely to be metabolized by numerous ALDH isoforms. Therefore, isolation techniques based on ALDH activity detection select a heterogeneous population of stem or progenitor cells. Despite active research in the field, the precise role(s) of different ALDH isoforms in stem cells remains enigmatic. Understanding the metabolic role of different ALDH isoform in the control of stem cell phenotype and cell fate during development, tissue homeostasis, or repair, as well as carcinogenesis, should open perspectives to significant discoveries in tissue biology. In this perspective, novel ALDH substrates are being developed. Here we describe how new substrates could be instrumental for better isolation of cell population with stemness potential and for defining hierarchy of cell populations in tissue. Finally, we speculate on other potential applications.

Role of liver progenitors in liver regeneration.

During massive liver injury and hepatocyte loss, the intrinsic regenerative capacity of the liver by replication of resident hepatocytes is overwhelmed. Treatment of this condition depends on the cause of liver injury, though in many cases liver transplantation (LT) remains the only curative option. LT for end stage chronic and acute liver diseases is hampered by shortage of donor organs and requires immunosuppression. Hepatocyte transplantation is limited by yet unresolved technical difficulties. Since currently no treatment is available to facilitate liver regeneration directly, therapies involving the use of resident liver stem or progenitor cells (LPCs) or non-liver stem cells are coming to fore. LPCs are quiescent in the healthy liver, but may be activated under conditions where the regenerative capacity of mature hepatocytes is severely impaired. Non-liver stem cells include embryonic stem cells (ES cells) and mesenchymal stem cells (MSCs). In the first section, we aim to provide an overview of the role of putative cytokines, growth factors, mitogens and hormones in regulating LPC response and briefly discuss the prognostic value of the LPC response in clinical practice. In the latter section, we will highlight the role of other (non-liver) stem cells in transplantation and discuss advantages and disadvantages of ES cells, induced pluripotent stem cells (iPS), as well as MSCs.