Embryonic stem (ES) cells are pluripotent and of therapeutic potential in regenerative medicine. Understanding pluripotency at the molecular level should illuminate fundamental properties of stem ...cells and the process of cellular reprogramming. Through cell fusion the embryonic cell phenotype can be imposed on somatic cells, a process promoted by the homeodomain protein Nanog, which is central to the maintenance of ES cell pluripotency. Nanog is thought to function in concert with other factors such as Oct4 (ref. 8) and Sox2 (ref. 9) to establish ES cell identity. Here we explore the protein network in which Nanog operates in mouse ES cells. Using affinity purification of Nanog under native conditions followed by mass spectrometry, we have identified physically associated proteins. In an iterative fashion we also identified partners of several Nanog-associated proteins (including Oct4), validated the functional relevance of selected newly identified components and constructed a protein interaction network. The network is highly enriched for nuclear factors that are individually critical for maintenance of the ES cell state and co-regulated on differentiation. The network is linked to multiple co-repressor pathways and is composed of numerous proteins whose encoding genes are putative direct transcriptional targets of its members. This tight protein network seems to function as a cellular module dedicated to pluripotency.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Pluripotency of embryonic stem (ES) cells is maintained by transcription factors that form a highly interconnected protein interaction network surrounding the homeobox protein Nanog. Enforced ...expression of Nanog in mouse ES (mES) cells promotes self-renewal and alleviates their requirement for leukemia inhibitory factor (LIF). Understanding molecular mechanisms by which Nanog functions should illuminate fundamental properties of stem cells and the process of cellular reprogramming. Previously, we showed that Nanog forms multiple protein complexes in mES cells. Here, we demonstrate that Nanog dimerizes through its C-terminal domain rather than the homeodomain. Dimerization is required for interaction with other pluripotency network proteins. We also show that enforced expression of the Nanog dimer, but not the monomer, functionally replaces wild-type Nanog to sustain LIF-independent self-renewal of ES cells. Our results demonstrate that Nanog-Nanog homodimerization is a critical aspect of its function promoting stem cell pluripotency.
In the present report, we carried out clinical-scale editing in adult mobilized CD34+ hematopoietic stem and progenitor cells (HSPCs) using zinc-finger nuclease-mediated disruption of BCL11a to ...upregulate the expression of γ-globin (fetal hemoglobin). In these cells, disruption of the erythroid-specific enhancer of the BCL11A gene increased endogenous γ-globin expression to levels that reached or exceeded those observed following knockout of the BCL11A coding region without negatively affecting survival or in vivo long-term proliferation of edited HSPCs and other lineages. In addition, BCL11A enhancer modification in mobilized CD34+ cells from patients with β-thalassemia major resulted in a readily detectable γ-globin increase with a preferential increase in G-gamma, leading to an improved phenotype and, likely, a survival advantage for maturing erythroid cells after editing. Furthermore, we documented that both normal and β-thalassemia HSPCs not only can be efficiently expanded ex vivo after editing but can also be successfully edited post-expansion, resulting in enhanced early in vivo engraftment compared with unexpanded cells. Overall, this work highlights a novel and effective treatment strategy for correcting the β-thalassemia phenotype by genome editing.
Since the 1970s, texts on research methods in animal behavior advocate that researchers minimize potential observer bias in their studies. One way to minimize possible bias is to record or score ...behavioral data blind to treatment, group, or individual. Another way to reduce bias is for researchers to analyze subsets or entire sets of data independently of one another and to obtain high inter‐observer reliability of behavioral coding. We reviewed several hundred published articles from 1970, 1980, 1990, 2000, and 2010 in five leading animal behavior journals and found that these two methods for minimizing or eliminating bias were rarely reported (<10% of articles reviewed). In contrast, a journal focusing on human infant behavior research was far more rigorous in incorporating methods to avoid bias (>80% of articles reviewed). The lack of reporting attempts to minimize bias in animal behavior studies suggests that, at best, many researchers view blind analyses of data or inter‐rater reliability as unimportant components of research or, if carried out, unnecessary to report in a manuscript. At worst, the lack of reporting attempts to minimize bias suggests that some published behavioral research may be unreliable. We are aware of constraints imposed by fieldwork and data collecting issues that make blind data comparisons or inter‐rater reliability assessments sometimes difficult or unfeasible. However, given that research ethicists often emphasize the fundamental importance of trust and transparency in science, we urge authors, reviewers, and editors of manuscripts to ensure that at least one of these two methods of reducing and reporting observer bias occurs.
Background
With the increasing use of neoadjuvant treatment (NAT) for patients with early-stage breast cancer (ESBC), adequate clinical staging is essential to inform treatment. While the use of MRI ...with NAT has been proposed to help with accuracy of pre-treatment clinical staging, its impact in clinical practice remains controversial.
Methods
A prospective institutional database of patients with ESBC treated with NAT between May 2012 and December 2020 was analyzed in order to compare the management of patients who received an MRI prior to NAT to those who did not. The indications for MRI and correlation of MRI findings to conventional breast imaging were evaluated. The impact of MRI on management was compared between the MRI and non-MRI groups.
Results
A total of 530 patients met inclusion criteria. Of these, 186 (35.1%) had an MRI and 344 (64.9%) did not. The most frequent indication for MRI was the determination of disease extent (54.5%). Patients who had an MRI prior to neoadjuvant treatment were significantly more likely to be younger (47 years versus 57 years;
p
< 0.001) and have multifocal disease (32.3% versus 22.1%;
p
< 0.05). When compared to conventional imaging, MRI reported a greater extent of disease in the breast (37.6%), more nodal involvement (18.8%), and multifocal disease (15.1%). Additional diagnostic interventions were advised in 52.2% of patients who underwent MRI. Rates of mastectomies were greater in the MRI group (80.0% versus 58.9%;
p
< 0.05) in addition to more axillary dissections (28.0% versus 17.4%;
p
< 0.01). Rates of locoregional recurrences were low in both groups, with similar disease-free survival outcomes at 5 years.
Conclusion
MRI identified significantly more disease in contrast to conventional imaging and lead to more aggressive surgical management. Prospective studies evaluating the role of MRI before NAT and its impact on long-term outcomes are needed.
Purpose
Data exploring optimal sequencing of anthracyclines and taxanes as neoadjuvant chemotherapy (NACT) for breast cancer are limited and inconsistent. The objective of this study was to assess ...the real-world impact of sequence order on pathologic complete response (pCR) and clinical outcomes from NACT.
Methods
Patients with HER2-negative breast cancer treated with NACT from May 2012 to April 2020 were identified from a prospectively collected institutional database. The primary endpoint was to compare rates of pCR (ypT0/isN0) between patients who received anthracyclines followed by taxanes (AC-T) to those who received taxanes followed by anthracyclines (T-AC). Additional endpoints of interest included clinical complete response, downstaging, Neo-Bioscore, conversion to breast-conserving surgery eligibility, relapse-free survival, and overall survival between groups.
Results
Of the 283 patients who met eligibility criteria, 187 (66%) received AC-T and 96 (34%) received T-AC. Sequence order did not influence the primary endpoint of pCR rate (19% for AC-T vs. 21% for T-AC,
p
= 0.752). There were also no significant differences in secondary NACT efficacy outcomes between groups. In the overall cohort, pCR rate was higher in patients with triple-negative breast cancer (TNBC) (32% vs. 13% in hormone-positive cancer,
p
< 0.001) and grade 3 tumors (31% vs. 12% for grade 1–2 tumors,
p
< 0.001).
Conclusions
In this real-world analysis of HER2-negative breast cancer patients, there was no differential impact on pCR rate or clinical outcomes from NACT with sequence order of anthracyclines and taxanes. This supports the current variation in prescribing practice.
Embryonic stem (ES) cells offer insight into early developmental fate decisions, and their controlled differentiation may yield vast regenerative potential. The molecular determinants supporting ES ...cell self-renewal are incompletely understood. The homeodomain proteins Nanog and Oct4 are essential for mouse ES cell self-renewal. Using a high-throughput approach, we discovered DNaseI hypersensitive sites and potential regulatory elements along a 160-kb region of the genome that includes GDF3, Dppa3, and Nanog. We analyzed gene expression, chromatin occupancy, and higher-order chromatin structure throughout this gene locus and found that expression of the reprogramming factor Oct4 is required to maintain its integrity.
To develop stem/progenitor cell-based therapy for cystic fibrosis (CF) lung disease, it is first necessary to identify markers of human lung epithelial progenitor/stem cells and to better understand ...the potential for differentiation into distinct lineages. Here we investigated integrin α6β4 as an epithelial progenitor cell marker in the human distal lung. We identified a subpopulation of α6β4(+) cells that localized in distal small airways and alveolar walls and were devoid of pro-surfactant protein C expression. The α6β4(+) epithelial cells demonstrated key properties of stem cells ex vivo as compared to α6β4(-) epithelial cells, including higher colony forming efficiency, expression of stem cell-specific transcription factor Nanog, and the potential to differentiate into multiple distinct lineages including basal and Clara cells. Co-culture of α6β4(+) epithelial cells with endothelial cells enhanced proliferation. We identified a subset of adeno-associated virus (AAVs) serotypes, AAV2 and AAV8, capable of transducing α6β4(+) cells. In addition, reconstitution of bronchi epithelial cells from CF patients with only 5% normal α6β4(+) epithelial cells significantly rescued defects in Cl(-) transport. Therefore, targeting the α6β4(+) epithelial population via either gene delivery or progenitor cell-based reconstitution represents a potential new strategy to treat CF lung disease.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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