Spatial Transcriptomics for the Analysis of Human Pituitary Development

Abstract The pituitary develops from oral ectoderm in contact with the adjacent hypothalamus. However, the precise mechanisms underlying pituitary development in concert with plural tissues are not fully understood, especially in human. A protocol to induce pituitary cells from human induced pluripotent stem cells (hiPSCs) has been established and applied to study pituitary development and disorders. In the method, oral ectoderm and hypothalamus are induced in one organoid, which enables recapitulation of the interactions between these tissues during embryonic development. It leads to self-organization of pituitary cells. Recently, spatial transcriptome technology has been developed and is suitable for the analysis of tissue interactions. Here, we utilized spatial transcriptomics to analyze pituitary organoids, especially focusing on the mechanisms regulating pituitary progenitor cell differentiation. Spatial transcriptomics revealed that the organoids consisted of several cell populations including hypothalamus, oral ectoderm, neural retina, and cortex neuron cells. Pituitary progenitor cells, characterized by the upregulation of LHX3, were included as part of the oral ectoderm population. Further analysis of the population identified human pituitary progenitor-specific genes including many causal genes for congenital hypopituitarism (CPH). Finally, using spatially resolved gene expression data, we examined the hypothalamic population that was in contact with pituitary progenitor cells and identified hypothalamic factors that might regulate progenitor cell differentiation in a paracrine manner. The genes upregulated in the pituitary progenitor and neighboring hypothalamus cell populations are potential causal gene candidates for CPH. In conclusion, spatial transcriptomics provides a novel platform to analyze tissue interaction networks during human pituitary development.

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Bone Marrow Niches for Skeletal Progenitor Cells and their Inhabitants in Health and Disease

Current Stem Cell Research & Therapy ◽

10.2174/1574888x14666190123161447 ◽

2019 ◽

Vol 14 (4) ◽

pp. 305-319 ◽

Cited By ~ 4

Author(s):

Marietta Herrmann ◽

Franz Jakob

Keyword(s):

Bone Marrow ◽

Progenitor Cells ◽

Progenitor Cell ◽

Adult Stem Cells ◽

Current Knowledge ◽

Cell Populations ◽

Surface Markers ◽

Bone Marrow Niches

The bone marrow hosts skeletal progenitor cells which have most widely been referred to as Mesenchymal Stem or Stromal Cells (MSCs), a heterogeneous population of adult stem cells possessing the potential for self-renewal and multilineage differentiation. A consensus agreement on minimal criteria has been suggested to define MSCs in vitro, including adhesion to plastic, expression of typical surface markers and the ability to differentiate towards the adipogenic, osteogenic and chondrogenic lineages but they are critically discussed since the differentiation capability of cells could not always be confirmed by stringent assays in vivo. However, these in vitro characteristics have led to the notion that progenitor cell populations, similar to MSCs in bone marrow, reside in various tissues. MSCs are in the focus of numerous (pre)clinical studies on tissue regeneration and repair.Recent advances in terms of genetic animal models enabled a couple of studies targeting skeletal progenitor cells in vivo. Accordingly, different skeletal progenitor cell populations could be identified by the expression of surface markers including nestin and leptin receptor. While there are still issues with the identity of, and the overlap between different cell populations, these studies suggested that specific microenvironments, referred to as niches, host and maintain skeletal progenitor cells in the bone marrow. Dynamic mutual interactions through biological and physical cues between niche constituting cells and niche inhabitants control dormancy, symmetric and asymmetric cell division and lineage commitment. Niche constituting cells, inhabitant cells and their extracellular matrix are subject to influences of aging and disease e.g. via cellular modulators. Protective niches can be hijacked and abused by metastasizing tumor cells, and may even be adapted via mutual education. Here, we summarize the current knowledge on bone marrow skeletal progenitor cell niches in physiology and pathophysiology. We discuss the plasticity and dynamics of bone marrow niches as well as future perspectives of targeting niches for therapeutic strategies.

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Co-Transplantation of Bone Marrow-MSCs and Myogenic Stem/Progenitor Cells from Adult Donors Improves Muscle Function of Patients with Duchenne Muscular Dystrophy

Cells ◽

10.3390/cells9051119 ◽

2020 ◽

Vol 9 (5) ◽

pp. 1119

Author(s):

Aleksandra Klimczak ◽

Agnieszka Zimna ◽

Agnieszka Malcher ◽

Urszula Kozlowska ◽

Katarzyna Futoma ◽

...

Keyword(s):

Skeletal Muscle ◽

Bone Marrow ◽

Duchenne Muscular Dystrophy ◽

Muscular Dystrophy ◽

Progenitor Cells ◽

Muscle Function ◽

Progenitor Cell ◽

Genetic Disorder ◽

Cell Populations ◽

Muscle Fibres

Duchenne muscular dystrophy (DMD) is a genetic disorder associated with a progressive deficiency of dystrophin that leads to skeletal muscle degeneration. In this study, we tested the hypothesis that a co-transplantation of two stem/progenitor cell populations, namely bone marrow-derived mesenchymal stem cells (BM-MSCs) and skeletal muscle-derived stem/progenitor cells (SM-SPCs), directly into the dystrophic muscle can improve the skeletal muscle function of DMD patients. Three patients diagnosed with DMD, confirmed by the dystrophin gene mutation, were enrolled into a study approved by the local Bioethics Committee (no. 79/2015). Stem/progenitor cells collected from bone marrow and skeletal muscles of related healthy donors, based on HLA matched antigens, were expanded in a closed MC3 cell culture system. A simultaneous co-transplantation of BM-MSCs and SM-SPCs was performed directly into the biceps brachii (two patients) and gastrocnemius (one patient). During a six-month follow-up, the patients were examined with electromyography (EMG) and monitored for blood kinase creatine level. Muscle biopsies were examined with histology and assessed for dystrophin at the mRNA and protein level. A panel of 27 cytokines was analysed with multiplex ELISA. We did not observe any adverse effects after the intramuscular administration of cells. The efficacy of BM-MSC and SM-SPC application was confirmed through an EMG assessment by an increase in motor unit parameters, especially in terms of duration, amplitude range, area, and size index. The beneficial effect of cellular therapy was confirmed by a decrease in creatine kinase levels and a normalised profile of pro-inflammatory cytokines. BM-MSCs may support the pro-regenerative potential of SM-SPCs thanks to their trophic, paracrine, and immunomodulatory activity. Both applied cell populations may fuse with degenerating skeletal muscle fibres in situ, facilitating skeletal muscle recovery. However, further studies are required to optimise the dose and timing of stem/progenitor cell delivery.

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Mass Cytometric Analysis of AML Stem and Early Progenitor Cells Reveals Karyotype and Genotype-Specific Cell Cycle Properties That Correlate with Known Responses to Chemotherapy

Blood ◽

10.1182/blood.v124.21.2359.2359 ◽

2014 ◽

Vol 124 (21) ◽

pp. 2359-2359

Author(s):

Gregory K. Behbehani ◽

Wendy J. Fantl ◽

Bruno C Medeiros ◽

Garry P. Nolan

Keyword(s):

Cell Cycle ◽

Stem Cells ◽

Bone Marrow ◽

Progenitor Cells ◽

Progenitor Cell ◽

S Phase ◽

The Other ◽

Cytotoxic Agents ◽

Phase Fraction ◽

Cell Populations

Abstract Introduction: Leukemic stem cells (LSCs) are recognized as important mediators of chemotherapy resistance and leukemia relapse. The postulated mechanism for this is the relative quiescence of these cell populations that renders them resistant to cytotoxic agents. This simple hypothesis, however, is supported almost entirely by indirect evidence, and fails to explain the large differences in relapse rates across different AML subtypes. To address this question, we have developed a mass cytometry (MCM) approach to assess the cell cycle of immunophenotypically complex primary samples from patients with AML. By processing samples immediately upon bone marrow harvest, we could determine if AML stem cells were quiescent in vivo and if the cell cycle properties of these cells varied between chemotherapy-responsive versus resistant AML subtypes. Methods: Bone marrow aspirates from 33 AML patients, 3 with APL, 2 with high-risk MDS, 5 with AML who achieved a CR with chemotherapy treatment, and 5 healthy donors (48 total samples) were incubated at 37°C for 15 minutes with 20uM Iodo-deoxyuridine (IdU) immediately after aspiration (<1 min), followed by fixation and storage. Samples were then analyzed with two overlapping 39-antibody MCM panels (50 markers total). Cellular barcoding was utilized to stain and analyze cells in tubes of 20 samples each, enabling direct comparison of samples to each other and to the healthy controls. Results: The high dimensionality of MCM enabled the simultaneous measurement of 25 surface markers and the identification of almost all immunophenotypic populations in human bone marrow. The use of barcoding, and the resultant ability to directly compare samples, enabled the detection of aberrant marker expression at very high resolution (2-3 fold changes). At least one surface marker aberrancy was detected in each AML sample. Unexpectedly, cell cycle analysis revealed that, compared to immunophenotypically similar normal cells, the average fraction of S-phase cells in AML samples was significantly lower. In both AML and healthy samples, the lowest S-phase fraction was found in fully differentiated populations and in hematopoietic stem cells (HSCs) while committed progenitor populations (myelo-monoblasts, promyelocytes, erythroblasts) exhibited the highest S-phase fraction. The HSC and early progenitor cell populations from patients with CBF AML (t(8;21) and inv(16)) demonstrated a significantly higher S-phase fraction than the same cell populations from the other AML samples (7.76% vs. 2.66%; p=0.0014). Furthermore, samples with FLT3-ITD mutations exhibited the lowest S-phase fraction in the HSC and early progenitor cell populations (0.63%), which was significantly lower than the S-phase fraction of the other AML samples (4.37%; p=9.3x10-4). Finally, a subset of patients (n=10) was being treated with hydroxyurea (HU) at the time of their bone marrow aspiration. The effect of HU treatment was manifest as a reduction in the IdU incorporation rate (with no change in S-phase fraction) in the cells of the treated patients. However, neither cell cycle arrest nor apoptosis were observed in these samples. This is in contrast with the commonly observed occurrence of both in leukemic cell lines treated in vitro with HU. Conclusions: By combining fresh sample processing with high-dimensional MCM analysis, we developed an innovative approach for the analysis of hematologic malignancies. Our results suggest that the relative sensitivity of CBF AML to cytotoxic chemotherapy may be the result of the increased fraction of S-phase cells within the HSC and early progenitor cell populations. Conversely, HSC and early progenitor cell populations from patients with FLT3-ITD mutations would be expected to be particularly resistant to cytarabine-based consolidation therapy due to the very low frequency of S-phase cells within these populations. This finding, combined with our observation that the stem and early progenitor cells from the FLT3-ITD samples have high expression of CD33, may provide a mechanistic explanation for the improved disease-free survival recently reported for FLT3-ITD AML patients treated with fractioned gemtuzumab ozogamicin in combination with standard therapy. Figure 1 Figure 1. Figure 2 Figure 2. Figure 3 Figure 3. Disclosures Behbehani: Fluidigm: Consultancy. Medeiros:Agios: Consulting - Ad board Other. Nolan:Fluidigm, Inc: Consultancy, Equity Ownership.

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The Adult Pituitary Shows Stem/Progenitor Cell Activation in Response to Injury and Is Capable of Regeneration

Endocrinology ◽

10.1210/en.2012-1152 ◽

2012 ◽

Vol 153 (7) ◽

pp. 3224-3235 ◽

Cited By ~ 56

Author(s):

Qiuli Fu ◽

Lies Gremeaux ◽

Raul M. Luque ◽

Daisy Liekens ◽

Jianghai Chen ◽

...

Keyword(s):

Pituitary Gland ◽

Progenitor Cells ◽

Transcriptional Activation ◽

Progenitor Cell ◽

Cell Activation ◽

Side Population ◽

Lineage Tracing ◽

Pituitary Cells ◽

Adult Age ◽

Gh Cells

The pituitary gland constitutes, together with the hypothalamus, the regulatory core of the endocrine system. Whether the gland is capable of cell regeneration after injury, in particular when suffered at adult age, is unknown. To investigate the adult pituitary's regenerative capacity and the response of its stem/progenitor cell compartment to damage, we constructed a transgenic mouse model to conditionally destroy pituitary cells. GHCre/iDTR mice express diphtheria toxin (DT) receptor after transcriptional activation by Cre recombinase, which is driven by the GH promoter. Treatment with DT for 3 d leads to gradual GH+ (somatotrope) cell obliteration with a final ablation grade of 80–90% 1 wk later. The stem/progenitor cell-clustering side population promptly expands after injury, concordant with the immediate increase in Sox2+ stem/progenitor cells. In addition, folliculo-stellate cells, previously designated as pituitary stem/progenitor cells and significantly overlapping with Sox2+ cells, also increase in abundance. In situ examination reveals expansion of the Sox2+ marginal-zone niche and appearance of remarkable Sox2+ cells that contain GH. When mice are left after the DT-provoked lesion, GH+ cells considerably regenerate during the following months. Double Sox2+/GH+ cells are observed throughout the regenerative period, suggesting recovery of somatotropes from stem/progenitor cells, as further supported by 5-ethynyl-2′-deoxyuridine (EdU) pulse-chase lineage tracing. In conclusion, our study demonstrates that the adult pituitary gland holds regenerative competence and that tissue repair follows prompt activation and plausible involvement of the stem/progenitor cells.

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Abstract 15440: Dynamics of Adipocyte and Endothelial Progenitor Cell Pools in Expanding Adipose Tissue

Circulation ◽

10.1161/circ.130.suppl_2.15440 ◽

2014 ◽

Vol 130 (suppl_2) ◽

Author(s):

Candice R Holden ◽

Marcin Wysoczynski ◽

Brian Sansbury ◽

Jason Hellmann ◽

Nagma Zafar ◽

...

Keyword(s):

Bone Marrow ◽

Adipose Tissue ◽

Stem Cell ◽

Progenitor Cells ◽

Progenitor Cell ◽

Stromal Vascular Fraction ◽

Cell Populations ◽

Adipose Organ ◽

The Impact

Objective: Obesity is a major risk factor for the development of several chronic diseases including type 2 diabetes and cardiovascular disease. Proper fat storage in white adipose tissue (WAT) is required to maintain insulin sensitivity and to preserve (cardio)vascular health. We hypothesize that endothelial and adipocyte progenitor cell populations (EPCs and APCs, respectively) must be appropriately balanced for physiological, as opposed to pathological, remodeling of WAT. Methods and Results: To determine the impact of nutrient excess on stem/progenitor cells in epididymal WAT, male C57BL/6J mice were placed on a high fat diet (HFD; 60% fat) for 12 weeks and changes in WAT stem cell populations were measured in the stromal vascular fraction by flow cytometry. Although the APC (CD24+/CD29+/Sca+/CD14-/CD45-) population, which has the capacity to differentiate into adipocytes both in vitro and in vivo , was not significantly changed with diet, Flk+/Sca+ EPCs were diminished, promoting a 4-fold decrease in the EPC/APC ratio (p <0.05, n = 6/group). To determine whether this deficit may be due to poor stem cell recruitment, mice were irradiated, and the bone marrow was repopulated with GFP+ donor marrow. The transplanted mice were then placed on a low fat diet (LFD; 10% fat) or HFD for 12 weeks, and WAT progenitor cells were again measured. Greater than 95% of the putative APCs in the WAT of HF-fed mice were GFP+ (p<0.0001, n=7-8/group), indicating a bone marrow-derived origin. Unexpectedly, less than 1% of the EPCs were GFP+ (p<0.001, n=7-8/group), which suggests that EPCs present in WAT are not derived from bone marrow in adult mice. Confocal analysis of WAT from HF-fed, bone marrow-transplanted mice showed little evidence of significant APC differentiation into triglyceride-laden adipocytes, suggesting that conditions associated with nutrient excess may impair the ability of the adipose organ to store fat properly. Conclusions: These results demonstrate that putative APCs, and not EPCs, in epididymal WAT are derived from bone marrow. Furthermore, our data suggest that conditions of nutrient excess promote an imbalance in EPCs and APCs, the stoichiometry of which may be critical for the development of new adipocytes and for proper storage of fat.

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Abstract 262: Deletion Of E2f1 Promotes Bone marrow Progenitor Cell Differentiation And Ischemic Cardiac Repair

Circulation Research ◽

10.1161/res.115.suppl_1.262 ◽

2014 ◽

Vol 115 (suppl_1) ◽

Author(s):

Shiyue Xu

Keyword(s):

Bone Marrow ◽

Cell Differentiation ◽

Oxidative Phosphorylation ◽

Progenitor Cells ◽

Progenitor Cell ◽

Cardiac Repair ◽

Pyruvate Dehydrogenase Kinase ◽

Expression Levels ◽

Significant Difference

Background: We have previously shown that knockout of E2F1 in mice enhances angiogenesis following induction of hind limb ischemia. Recent studies suggest that suppression of E2F1 enhances oxidative phosphorylation in a variety of cell types. Since an increase in oxidative phosphorylation in stem/progenitor cells is often associated with cell differentiation, we hypothesize that E2F1-deficiency may promote bone marrow (BM) progenitor cell differentiation thereby impact on ischemic cardiac repair. Methods and Results: We cultured bone marrow (BM) Lin- progenitor cells under hypoxic and normxic conditions for 24 h, then measured the expression of metabolism associated genes and evaluated cell proliferation and differentiation. We also performed adoptive BM transplantation to reconstitute BM of WT mice with E2F1-/- or WT BM, followed by surgical induction of myocardial infarction (MI), to compare the role of BM E2F1 in the cardiac repair in vivo. Notably, we found that the expression levels of pyruvate dehydrogenase kinase (PDK)-4 and PDK2, two critical inhibitors of mitochondrial oxidative phosphorylation, in the E2F1-/- BM Lin- progenitor cells is markedly lower than those in WT cells (P<0.01, n=4). After culture in the EPC differentiation medium for 7 days, the expression levels of EPC markers, CD31 and KDR , were significantly higher in E2F1-/- cells than in WT control cells (P<0.05, n=3). Although there was no significant difference in the proliferating rate between WT and E2F1-/- BM Lin- progenitor cells cultured in normoxia, when cultured under hypoxic condition the proliferating rate of E2F1-/- cells were markedly greater than that of WT cells (P<0.05, n=3). Consistently, the infarcted size in mice transplanted with E2F1-/-eGFPTg BM was significantly smaller than in mice transplanted with WTeGFPTg BM (P<0.01, n=5). Conclusions: Genetic deletion of E2F1 in the BM progenitor cells enhances oxidative metabolism that may result in enhanced differentiation towards endothelial lineage and increased proliferation in the ischemic/hypoxic tissue environment. Therefore, inhibition of E2F1 in BM progenitor cells may improve the recovery from cardiac ischemic injury.

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The expansion of adult stem/progenitor cells and their marker expression fluctuations are linked with pituitary plastic adaptation during gestation and lactancy

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00077.2016 ◽

2016 ◽

Vol 311 (2) ◽

pp. E367-E379 ◽

Cited By ~ 2

Author(s):

Alicia Maldré Vaca ◽

Carolina Beatriz Guido ◽

Liliana del Valle Sosa ◽

Juan Pablo Nicola ◽

Jorge Mukdsi ◽

...

Keyword(s):

Stem Cells ◽

Fluorescence Microscopy ◽

Progenitor Cells ◽

Protein Level ◽

Progenitor Cell ◽

Marginal Zone ◽

Cell Populations ◽

Double Immunostaining ◽

Cell Markers ◽

Physiological Conditions

Extensive evidence has revealed variations in the number of hormone-producing cells in the pituitary gland, which occur under physiological conditions such as gestation and lactancy. It has been proposed that new hormone-producing cells differentiate from stem cells. However, exactly how and when this takes place is not clear. In this work, we used immunoelectron microscopy to identify adult pituitary stem/progenitor cells (SC/P) localized in the marginal zone (MZ), and additionally, we detected GFRa2-, Sox2-, and Sox9-positive cells in the adenoparenchyma (AP) by fluorescence microscopy. Then, we evaluated fluctuations of SC/P mRNA and protein level markers in MZ and AP during gestation and lactancy. An upregulation in stemness markers was shown at term of gestation (AT) in MZ, whereas there were more progenitor cell markers in the middle of gestation and active lactancy. Concerning committed cell markers, we detected a rise in AP at beginning of lactancy (d1L). We performed a BrdU uptake analysis in MZ and AP cells. The highest level of BrdU uptake was observed in MZ AT cells, whereas in AP this was detected in d1L, followed by a decrease in both the MZ and AP. Finally, we detected double immunostaining for BrdU-GFRa2 in MZ AT cells and BrdU-Sox9 in the AP d1L cells. Taken together, we hypothesize that the expansion of the SC/P niche took place mainly in MZ from pituitary rats in AT and d1L. These results suggest that the SC niche actively participates in pituitary plasticity during these reproductive states, contributing to the origin of hormone cell populations.

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Persistent Expression of Notch2 Delays Gonadotrope Differentiation

Molecular Endocrinology ◽

10.1210/me.2005-0394 ◽

2006 ◽

Vol 20 (11) ◽

pp. 2898-2908 ◽

Cited By ~ 48

Author(s):

Lori T. Raetzman ◽

Bayly S. Wheeler ◽

Shelley A. Ross ◽

Paul Q. Thomas ◽

Sally A. Camper

Keyword(s):

Cell Differentiation ◽

Pituitary Gland ◽

Progenitor Cell ◽

Anterior Lobe ◽

Temporal Expression ◽

Cell Specification ◽

Pituitary Development ◽

Normal Pituitary Gland ◽

Gland Development ◽

Selection Of

Abstract Normal pituitary gland development requires coordination between maintenance of progenitor cell pools and selection of progenitors for differentiation. The spatial and temporal expression of Notch2 during pituitary development suggested that it could control progenitor cell differentiation in the pituitary. Consistent with this idea, Notch2 is not expressed in Prop1 mutants, and anterior pituitary progenitors in Prop1 mutants appear to be unable to transition from proliferation to differentiation properly, resulting in anterior lobe failed cell specification and evolving hypoplasia. To test the function of Notch2 directly, we used the αGSU subunit promoter to express activated NOTCH2 persistently in pre-gonadotropes and pre-thyrotropes of transgenic mice. At birth, there is a small reduction in the population of fully differentiated thyrotropes and almost no fully differentiated gonadotropes. The temporal and spatial expression of Hey1 suggests that it could be a mediator of this effect. Gonadotropes complete their differentiation program eventually, although expression of LH and FSH is mutually exclusive with NOTCH2 transgene expression. This demonstrates that activated Notch2 is sufficient to delay gonadotrope differentiation, and it supports the hypothesis that Notch2 regulates progenitor cell differentiation in the pituitary gland.

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A novel machine learning based approach for iPS progenitor cell identification

10.1101/744920 ◽

2019 ◽

Author(s):

Haishan Zhang ◽

Ximing Shao ◽

Yin Peng ◽

Yanning Teng ◽

Konda Mani Saravanan ◽

...

Keyword(s):

Machine Learning ◽

Image Analysis ◽

Progenitor Cells ◽

Progenitor Cell ◽

Early Stage ◽

Time Windows ◽

Underlying Mechanism ◽

Ips Cells ◽

Microscopic Image ◽

Induced Pluripotent

AbstractIdentification of induced pluripotent stem (iPS) progenitor cells, the iPS forming cells in early stage of reprogramming, could provide valuable information for studying the origin and underlying mechanism of iPS cells. However, it is very difficult to identify experimentally since there are no biomarkers known for early progenitor cells, and only about 6 days after reprogramming initiation, iPS cells can be experimentally determined via fluorescent probes. What is more, the ratio of progenitor cells during early reprograming period is below 5%, which is too low to capture experimentally in the early stage.In this paper, we propose a novel computational approach for the identification of iPS progenitor cells based on machine learning and microscopic image analysis. Firstly, we record the reprogramming process using a live cell imaging system after 48 hours of infection with retroviruses expressing Oct4, Sox2 and Klf4, later iPS progenitor cells and normal murine embryonic fibroblasts (MEFs) within 3 to 5 days after infection are labeled by retrospectively tracing the time-lapse microscopic image. We then calculate 11 types of cell morphological and motion features such as area, speed, etc., and select best time windows for modeling and perform feature selection. Finally, a prediction model using XGBoost is built based on the selected six types of features and best time windows. Our model allows several missing values/frames in the sample datasets, thus it is applicable to a wide range of scenarios.Cross-validation, holdout validation and independent test experiments showed that the minimum precision is above 52%, that is, the ratio of predicted progenitor cells within 3 to 5 days after viral infection is above 52%. The results also confirmed that the morphology and motion pattern of iPS progenitor cells is different from that of normal MEFs, which helps with the machine learning methods for iPS progenitor cell identification.Author SummaryIdentification of induced pluripotent stem (iPS) progenitor cells could provide valuable information for studying the origin and underlying mechanism of iPS cells. However, it is very difficult to identify experimentally since there are no biomarkers known for early progenitor cells, and only after about 6 days of induction, iPS cells can be experimentally determined via fluorescent probes. What is more, the percentage of the progenitor cells during the early induction period is below 5%, too low to capture experimentally in early stage. In this work, we proposed an approach for the identification of iPS progenitor cells, the iPS forming cells, based on machine learning and microscopic image analysis. The aim is to help biologists to enrich iPS progenitor cells during the early stage of induction, which allows experimentalists to select iPS progenitor cells with much higher probability, and furthermore to study the biomarkers which trigger the reprogramming process.

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Evaluation of Endothelial and Vascular-Derived Progenitor Cell Populations in the Proximal and Distal UCL of the Elbow: A Comparative Study

Orthopaedic Journal of Sports Medicine ◽

10.1177/2325967118777825 ◽

2018 ◽

Vol 6 (6) ◽

pp. 232596711877782 ◽

Cited By ~ 4

Author(s):

Salvatore J. Frangiamore ◽

Elizabeth R. Morris ◽

Alex C. Scibetta ◽

Jorge Chahla ◽

Gilbert Moatshe ◽

...

Keyword(s):

Progenitor Cells ◽

Progenitor Cell ◽

Smooth Muscle Actin ◽

Rank Test ◽

Cell Populations ◽

Fluorescent Staining ◽

Collateral Ligament ◽

Signed Rank ◽

Wilcoxon Signed Rank Test ◽

Signed Rank Test

Background: Vascular-derived progenitor and endothelial cell populations (CD31, CD34, CD146) are capable of multipotent differentiation at the site of injured ligamentous tissue to aid in the intrinsic healing response. Proximal ulnar collateral ligament (UCL) tears have been reported to have better healing capability when compared with distal UCL tears. Purpose: To compare the vascular composition of the proximal and distal insertions of the anterior bundle of the UCL of the elbow via known markers of endothelial and vascular-derived progenitor cells (CD31, CD34, CD146). Study Design: Descriptive laboratory study. Methods: UCLs were harvested from 10 nonpaired fresh-frozen human cadaveric elbows and transected into proximal and distal portions. Endothelial and vascular-derived progenitor cell densities were assessed with 4 staining groups: CD31 (immunohistochemistry) and CD31/α-smooth muscle actin (α-SMA), CD34/α-SMA, and CD146/α-SMA (immunofluorescence). CD31 immunohistochemistry identified endothelial progenitor cells in the UCL. Later staining of the same slides with α-SMA demonstrated the relationship of progenitor cells to the surrounding vasculature. Fluorescent staining was quantified by calculating the proportion of positively stained nuclei versus the total number of nuclei in the proximal and distal UCL. Results: CD31+ cells were present in the proximal and distal sections of all 10 UCLs. Fluorescent staining revealed no significant differences in the ratio of CD31 to total nuclei between the distal (median, 36% [range, 23%-53%]) and proximal UCL (39% [22%-56%]) ( P = .432, Wilcoxon signed-rank test). Similarly, no differences were seen between CD34 distal (39% [24%-64%]) and proximal regions (46% [28%-63%]) ( P = .846, Wilcoxon signed-rank test) or CD146 distal (40% [12%-65%]) and proximal regions (40% [22%-51%]) ( P ≥ .999, Wilcoxon signed-rank test). Conclusion: Analysis of UCL tissues demonstrated equal distributions of vascular endothelial and vascular-derived progenitor cell markers throughout the proximal and distal UCL. Unlike that of the medial collateral ligament of the knee, the microvascular composition of the proximal and distal UCL insertions was not different, suggesting a well-vascularized ligament throughout its course. Clinical Relevance: These findings investigate one of the possible contributors to UCL healing after injury, which may provide insight into operative and nonoperative management of UCL injuries in the future. This study also indicates that reasons other than differences in progenitor cell density alone may explain the clinical healing differences seen between proximal and distal UCL tears. A better understanding of the microvascular environment and associated blood supply is warranted to understand the healing capability of the UCL.

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