Nuclear pore proteins (Nups) form the only channels on the nuclear envelope that allow macromolecule transport between the cytoplasm and the nucleoplasm. Initial studies in the unicellular organism yeast have confirmed that deletions and/or mutations of multiple Nups result in deficiency in bulk transport between cellular compartments and subsequently lethality. Recent investigations regarding multicellular organisms, however, have revealed that mutations in Nups are often linked to cell-type specific defects and diseases. Therefore, there might be cell-type specific functions of Nups, for example in the context of differentiation. One potential cell-type specific function of Nups could be related to the regulation of genes that are activated in a cell-type dependent manner. Such hypothesis stems from the observation that microscopically, Nups appear to selectively interact with active genomic regions. To test this hypothesis, chromatin immunoprecipitation experiments for Nup98 have been performed in multiple cell types, including human cells at different stages of neural differentiation. It has been found that the patterns of Nup98-genome interaction are highly cell-type specific in the neural differentiation system. Further functional studies have demonstrated a role of Nup98 in regulating the expression of a cohort of genes active during neural development. As an alternative approach to investigate into the potential cell-type dependent behavior of Nups, dynamics of Nups were analyzed at different stages of muscle differentiation by live cell imaging. One Nup, Nup50, was found to exhibit decreased dynamics upon differentiation, serving as the first example of a Nup that has altered dynamics during development. Kifc1, a kinesin-superfamily protein that is predicted to be a nuclear motor, was found to confer the higher dynamics of Nup50 by physical interaction before differentiation. Upon differentiation, Kifc1 is degraded, which could explain the decrease in Nup50 mobility. Knocking down Nup50 decreased efficiency of muscle differentiation, whereas knocking down Kifc1 accelerated muscle differentiation. This suggests that Nup50 is differentially regulated during muscle development and is functionally important for differentiation. Together these studies revealed previously unidentified roles of Nup98 and Nup50 in differentiation, which might shed light on the cell-type specific diseases related to Nup mutations

The purpose of this study was to develop dosimetry verification procedures for volumetric-modulated arc therapy (VMAT)-based total marrow irradiation (TMI). The VMAT based TMI plans were generated for three patients: one child and two adults. The planning target volume (PTV) was defined as bony skeleton, from head to mid-femur, with a 3 mm margin. The plan strategy similar to published studies was adopted. The PTV was divided into head and neck, chest, and pelvic regions, with separate plans each of which is composed of 2-3 arcs/fields. Multiple isocenters were evenly distributed along the patient's axial direction. The focus of this study is to establish a dosimetry quality assurance procedure involving both two-dimensional (2D) and three-dimensional (3D) volumetric verifications, which is desirable for a large PTV treated with multiple isocenters. The 2D dose verification was performed with film for gamma evaluation and absolute point dose was measured with ion chamber, with attention to the junction between neighboring plans regarding hot/cold spots. The 3D volumetric dose verification used commercial dose reconstruction software to reconstruct dose from electronic portal imaging devices (EPID) images. The gamma evaluation criteria in both 2D and 3D verification were 5% absolute point dose difference and 3 mm of distance to agreement. With film dosimetry, the overall average gamma passing rate was 98.2% and absolute dose difference was 3.9% in junction areas among the test patients; with volumetric portal dosimetry, the corresponding numbers were 90.7% and 2.4%. A dosimetry verification procedure involving both 2D and 3D was developed for VMAT-based TMI. The initial results are encouraging and warrant further investigation in clinical trials.

The objective was to compare intensity-modulated radiation therapy (IMRT) with 3D conformal proton therapy (3DCPT) in the treatment of cervical cancer. In particular, each technique's ability to spare pelvic bone marrow (PBM) was of primary interest in this study. A total of six cervical cancer patients (3 postoperative and 3 intact) were planned and analyzed. All plans had uniform 1.0 cm CTV-PTV margin and satisfied the 95% PTV with 100% isodose (prescription dose = 45 Gy) coverage. Dose-volume histograms (DVH) were analyzed for comparison. The overall PTV and PBM volumes were 1035.9 ± 192.2 cc and 1151.4 ± 198.3 cc, respectively. In terms of PTV dose conformity index (DCI) and dose homogeneity index (DHI), 3DCPT was slightly superior to IMRT with 1.00 ± 0.001, 1.01 ± 0.02, and 1.10 ± 0.02, 1.13 ± 0.01, respectively. In addition, 3DCPT demonstrated superiority in reducing lower doses (i.e., V30 or less) to PBM, small bowel and bladder. Particularly in PBM, average V10 and V20 reductions of 10.8% and 7.4% (p = 0.001 and 0.04), respectively, were observed. However, in the higher dose range, IMRT provided better sparing (> V30). For example, in small bowel and PBM, average reductions in V45 of 4.9% and 10.0% (p = 0.048 and 0.008), respectively, were observed. Due to its physical characteristics such as low entrance dose, spread-out Bragg peak and finite particle range of protons, 3DCPT illustrated superior target coverage uniformity and sparing of the lower doses in PBM and other organs. Further studies are, however, needed to fully exploit the benefits of protons for general use in cervical cancer.

Transcription factor binding, chromatin modifications and large scale chromatin re-organization underlie progressive, irreversible cell lineage commitments and differentiation. We know little, however, about chromatin changes as cells enter transient, reversible states such as migration. Here we demonstrate that when human progenitor keratinocytes either differentiate or migrate they form complements of typical enhancers and super-enhancers that are unique for each state. Unique super-enhancers for each cellular state link to gene expression that confers functions associated with the respective cell state. These super-enhancers are also enriched for skin disease sequence variants. GRHL3, a transcription factor that promotes both differentiation and migration, binds preferentially to super-enhancers in differentiating keratinocytes, while during migration, it binds preferentially to promoters along with REST, repressing the expression of migration inhibitors. Key epidermal differentiation transcription factor genes, including GRHL3, are located within super-enhancers, and many of these transcription factors in turn bind to and regulate super-enhancers. Furthermore, GRHL3 represses the formation of a number of progenitor and non-keratinocyte super-enhancers in differentiating keratinocytes. Hence, chromatin relocates GRHL3 binding and enhancers to regulate both the irreversible commitment of progenitor keratinocytes to differentiation and their reversible transition to migration.