Role of label-retaining cells in estrogen-induced endometrial regeneration

Candidate stem/progenitor cells have been identified in mouse endometrium as label-retaining cells (LRCs). The role of endometrial stem/progenitor cells in initiating estrogen-stimulated endometrial growth in prepubertal and cycling mice was investigated following a single 17beta-estradiol (E2) injection in bromodeoxyuridine (BrdU)-labeled and -chased (LRC), ovariectomised mice. Proliferating (BrdU(+)/Ki-67(+)) and mitotic (BrdU(+)/PH3(+)) epithelial LRCs were first detected in prepubertal mice 8 hours following E2 treatment, initiating the proliferative response. In contrast, all epithelial LRCs and 16% of epithelial cells in cycling mice proliferated within 2 hours. In cycling mice, 12% of stromal LRCs initiated a proliferative response 8 hours after E2. Proliferating epithelial LRCs and most stromal LRCs (85%) lacked estrogen receptor-alpha (ESR1). These findings suggest that endometrial epithelial LRCs function as stem/progenitor cells by receiving proliferative signals from neighboring ESR1(+) niche cells to initiate the growth of the epithelium during development, while mature epithelial cells may undergo self-replication in cycling endometrium.postprin

The inducible caspase-9 suicide gene system as a 'safety switch' to limit on-target, off-tumor toxicities of chimeric antigen receptor T cells

Published online: 28 October 2014.Immune modulation has become a central element in many cancer treatments, and T cells genetically engineered to express chimeric antigen receptors (CAR) may provide a new approach to cancer immunotherapy. Autologous CAR T cells that have been re-directed toward tumor-associated antigens (TAA) have shown promising results in phase 1 clinical trials, with some patients undergoing complete tumor regression. However, this T-cell therapy must carefully balance effective T-cell activation, to ensure antitumor activity, with the potential for uncontrolled activation that may produce immunopathology. An inducible Caspase 9 (iCasp9) "safety switch" offers a solution that allows for the removal of inappropriately activated CAR T cells. The induction of iCasp9 depends on the administration of the small molecule dimerizer drug AP1903 and dimerization results in rapid induction of apoptosis in transduced cells, preferentially killing activated cells expressing high levels of transgene. The iCasp9 gene has been incorporated into vectors for use in preclinical studies and demonstrates effective and reliable suicide gene activity in phase 1 clinical trials. A third-generation CAR incorporating iCasp9 re-directs T cells toward the GD2 TAA. GD2 is over-expressed in melanoma and other malignancies of neural crest origin and the safety and activity of these GD2-iCAR T cells will be investigated in CARPETS and other actively recruiting phase 1 trials.Tessa Gargett and Michael P. Brow

Higher order contributions to the effective action of N=2 super Yang-Mills

We apply heat kernel techniques in N=1 superspace to compute the one-loop effective action to order $F^5$ for chiral superfields coupled to a non-Abelian super Yang-Mills background. The results, when combined with those of hep-th/0210146, yield the one-loop effective action to order $F^5$ for any N=2 super Yang-Mills theory coupled to matter hypermultiplets.Comment: 23 pages, references adde

The influence of crosswind tidal currents on Langmuir circulation in a shallow ocean

Author Posting. © American Geophysical Union, 2011. This article is posted here by permission of American Geophysical Union for personal use, not for redistribution. The definitive version was published in Journal of Geophysical Research 116 (2011): C08005, doi:10.1029/2011JC006971.Langmuir circulation (LC) is a turbulent process driven by wind and surface waves that plays a key role in transferring momentum, heat, and mass in the oceanic surface layer. On the coastal shelves the largest-scale LC span the whole water column and thus couple the surface and bottom boundary layers and enhance turbulent mixing. Observations and large eddy simulations (LES) of a shallow coastal ocean demonstrate that these relatively large scale Langmuir cells are strongly influenced by crosswind tidal currents. Two mechanisms by which crosswind tidal shear may distort and disrupt Langmuir cells are proposed. The first mechanism involves cell shearing due to differential advection across the whole cell. For the second mechanism, middepth vertical LC currents advect sheared mean crosswind current, leading to the attraction of upwelling and downwelling regions, so that LC cells are unsustainable when both regions overlap. Scaling arguments indicate that LC cells are more susceptible to crosswind shear distortion for smaller LC surface velocity convergence and greater cell aspect ratio (vertical to horizontal LC scale), which is consistent with the results obtained from the observations and LES. These results imply that scaling of LC characteristics in a coastal ocean differs from that in the open ocean, which has important practical implications for parameterizing enhanced mixing due to LC.This research was supported by the Office of Naval Research through grant N00014‐06‐1‐0178 (A.P., J.T.). Author T.K. received support from Faculty Startup Funds of the University of Delaware College of Earth, Ocean, and Environment

Increase in DNA vaccine efficacy by virosome delivery and co-expression of a cytolytic protein

The potential of DNA vaccines has not been realised due to suboptimal delivery, poor antigen expression and the lack of localised inflammation, essential for antigen presentation and an effective immune response to the immunogen. Initially, we examined the delivery of a DNA vaccine encoding a model antigen, luciferase (LUC), to the respiratory tract of mice by encapsulation in a virosome. Virosomes that incorporated influenza virus haemagglutinin effectively delivered DNA to cells in the mouse respiratory tract and resulted in antigen expression and systemic and mucosal immune responses to the immunogen after an intranasal (IN) prime/intradermal (ID) boost regimen, whereas a multidose ID regimen only generated systemic immunity. We also examined systemic immune responses to LUC after ID vaccination with a DNA vaccine, which also encoded one of the several cytolytic or toxic proteins. Although the herpes simplex virus thymidine kinase, in the presence of the prodrug, ganciclovir, resulted in cell death, this failed to increase the humoral or cell-mediated immune responses. In contrast, the co-expression of LUC with the rotavirus non-structural protein 4 (NSP4) protein or a mutant form of mouse perforin, proteins which are directly cytolytic, resulted in increased LUC-specific humoral and cell-mediated immunity. On the other hand, co-expression of LUC with diphtheria toxin subunit A or overexpression of perforin or NSP4 resulted in a lower level of immunity. In summary, the efficacy of DNA vaccines can be improved by targeted IN delivery of DNA or by the induction of cell death in vaccine-targeted cells after ID delivery.Tessa Gargett, Branka Grubor-Bauk, Darren Miller, Tamsin Garrod, Stanley Yu, Steve Wesselingh, Andreas Suhrbier, and Eric J Gowan

Nonlocal transport due to Langmuir circulation in a coastal ocean

Author Posting. © American Geophysical Union, 2012. This article is posted here by permission of American Geophysical Union for personal use, not for redistribution. The definitive version was published in Journal of Geophysical Research 117 (2012): C12007, doi:10.1029/2012JC008340.We present observations and simulations of large-scale velocity structures associated with turbulent boundary layer dynamics of a coastal ocean. Special purpose acoustic Doppler current profiler measurements revealed that such structures were frequently present, in spite of complex coastal environmental conditions. Large eddy simulation results are only consistent with these observations if the Langmuir circulation (LC) effect due to wave-current interaction is included in the model. Thus, model results indicate that the observed large-scale velocity structures are due to LC. Based on these simulations, we examine the shift of energetics and transport from a local regime for purely shear-driven turbulence to a nonlocal regime for turbulence with LC due to coherent large-scale motions that span the whole water column. Without LC, turbulent kinetic energy (TKE) dissipation rates approximately balance TKE shear production, consistent with solid wall boundary layer turbulence. This stands in contrast to the LC case for which the vertical TKE transport plays a dominant role in the TKE balance. Conditional averages argue that large-scale LC coherent velocity structures extract only a small fraction of energy from the wavefield but receive most of their energy input from the Eulerian shear. The analysis of scalar fields and Lagrangian particles demonstrates that the vertical transport is significantly enhanced with LC but that small-scale mixing may be reduced. In the presence of LC, vertical scalar fluxes may be up gradient, violating a common assumption in oceanic boundary layer turbulence parameterizations.This work was supported by the U.S. National Science Foundation (Grant OCE-1130678). CBLAST-Low analysis was supported by the Office of Naval Research under grants N00014-03-1- 0681 and N00014-06-1-0178 to the Woods Hole Oceanographic Institution. Author T.K. received support from Faculty Startup Funds of the School of Marine Science and Policy, University of Delaware.2013-06-1

Cytolytic DNA vaccine encoding lytic perforin augments the maturation of- and antigen presentation by- dendritic cells in a time-dependent manner

The use of cost-effective vaccines capable of inducing robust CD8+ T cell immunity will contribute significantly towards the elimination of persistent viral infections and cancers worldwide. We have previously reported that a cytolytic DNA vaccine encoding an immunogen and a truncated mouse perforin (PRF) protein significantly augments anti-viral T cell (including CD8+ T cell) immunity. Thus, the current study investigated whether this vaccine enhances activation of dendritic cells (DCs) resulting in greater priming of CD8+ T cell immunity. In vitro data showed that transfection of HEK293T cells with the cytolytic DNA resulted in the release of lactate dehydrogenase, indicative of necrotic/lytic cell death. In vitro exposure of this lytic cell debris to purified DCs from naïve C57BL/6 mice resulted in maturation of DCs as determined by up-regulation of CD80/CD86. Using activation/proliferation of adoptively transferred OT-I CD8+ T cells to measure antigen presentation by DCs in vivo, it was determined that cytolytic DNA immunisation resulted in a time-dependent increase in the proliferation of OT-I CD8+ T cells compared to canonical DNA immunisation. Overall, the data suggest that the cytolytic DNA vaccine increases the activity of DCs which has important implications for the design of DNA vaccines to improve their translational prospects.Danushka K. Wijesundara, Wenbo Yu, Ben J. C. Quah, Preethi Eldi, John D. Hayball, Kerrilyn R. Diener, Ilia Voskoboinik, Eric J. Gowans, and Branka Grubor-Bau

Positron emission tomographic imaging of tumor cell death using zirconium-89-labeled APOMAB(R) following cisplatin chemotherapy in lung and ovarian cancer xenograft models

Published online 06 July 2021Purpose Early detection of tumor treatment responses represents an unmet clinical need with no approved noninvasive methods. DAB4, or its chimeric derivative, chDAB4 (APOMAB®) is an antibody that targets the Lupus associated antigen (La/SSB). La/SSB is over-expressed in malignancy and selectively targeted by chDAB4 in cancer cells dying from DNA-damaging treatment. Therefore, chDAB4 is a unique diagnostic tool that detects dead cancer cells and thus could distinguish between treatment responsive and nonresponsive patients. Procedures In clinically relevant tumor models, mice bearing subcutaneous xenografts of human ovarian or lung cancer cell lines or intraperitoneal ovarian cancer xenografts were untreated or given chemotherapy followed 24h later by chDAB4 radiolabeled with [⁸⁹Zr]ZrIV. Tumor responses were monitored using bioluminescence imaging and caliper measurements. [⁸⁹Zr]Zr-chDAB4 uptake in tumor and normal tissues was measured using an Albira SI Positron-Emission Tomography (PET) imager and its biodistribution was measured using a Hidex gamma-counter. Results Tumor uptake of [⁸⁹Zr]Zr-chDAB4 was detected in untreated mice, and uptake significantly increased in both human lung and ovarian tumors after chemotherapy, but not in normal tissues. Conclusion Given that tumors, rather than normal tissues, were targeted after chemotherapy, these results support the clinical development of chDAB4 as a radiodiagnostic imaging agent and as a potential predictive marker of treatment response.Vasilios Liapis, William Tieu, Nicole L. Wittwer, Tessa Gargett, Andreas Evdokiou, Prab Takhar, Stacey E. Rudd, Paul S. Donnelly, Michael P. Brown, Alexander H. Staudache

Bone Marrow-Derived Cells from Male Donors Do Not Contribute to the Endometrial Side Population of the Recipient

Accumulated evidence demonstrates the existence of bone marrow-derived cells origin in the endometria of women undergoing bone marrow transplantation (BMT). In these reports, cells of a bone marrow (BM) origin are able to differentiate into endometrial cells, although their contribution to endometrial regeneration is not yet clear. We have previously demonstrated the functional relevance of side population (SP) cells as the endogenous source of somatic stem cells (SSC) in the human endometrium. The present work aims to understand the presence and contribution of bone marrow-derived cells to the endometrium and the endometrial SP population of women who received BMT from male donors. Five female recipients with spontaneous or induced menstruations were selected and their endometrium was examined for the contribution of XY donor-derived cells using fluorescent in situ hybridization (FISH), telomapping and SP method investigation. We confirm the presence of XY donor-derived cells in the recipient endometrium ranging from 1.7% to 2.62%. We also identify 0.45–0.85% of the donor-derived cells in the epithelial compartment displaying CD9 marker, and 1.0–1.83% of the Vimentin-positive XY donor-derived cells in the stromal compartment. Although the percentage of endometrial SP cells decreased, possibly being due to chemotherapy applied to these patients, they were not formed by XY donor-derived cells, donor BM cells were not associated with the stem cell (SC) niches assessed by telomapping technique, and engraftment percentages were very low with no correlation between time from transplant and engraftment efficiency, suggesting random terminal differentiation. In conclusion, XY donor-derived cells of a BM origin may be considered a limited exogenous source of transdifferentiated endometrial cells rather than a cyclic source of BM donor-derived stem cells

On the Background Field Method Beyond One Loop: A manifestly covariant derivative expansion in super Yang-Mills theories

There are currently many string inspired conjectures about the structure of the low-energy effective action for super Yang-Mills theories which require explicit multi-loop calculations. In this paper, we develop a manifestly covariant derivative expansion of superspace heat kernels and present a scheme to evaluate multi-loop contributions to the effective action in the framework of the background field method. The crucial ingredient of the construction is a detailed analysis of the properties of the parallel displacement propagators associated with Yang-Mills supermultiples in N-extended superspace.Comment: 32 pages, latex, 7 EPS figures. v2: references, comments added, typos corrected, incorrect `skeleton' conjecture in sect. 3 replaced by a more careful treatment. v3: typos corrected, final version published in JHE