Cell nucleus

Long-term success of distraction osteogenesis depends on the ability of the surrounding soft tissues to tolerate distraction forces and to adapt to the resulting increase in skeletal length and volume. The hypothesis tested in this study was that mandibular elongation by distraction induces myocyte proliferation. Unilateral mandibular angle osteotomies were performed in 16 Yucatan minipigs. The hemimandibles were lengthened using semiburied distraction devices (Synthes Maxillofacial, Paoli, PA) with 0-day latency, 1, 2, or 4 mm/d distraction rates, and a neutral fixation period of twice the gap size in days. In 2 additional animals, the dissection and osteotomies were performed, and distraction devices were placed without activation (sham control). At the end of neutral fixation, tissues were taken from masseter muscle overlying the osteotomy and the equivalent region on the unoperated side (contralateral control). Proliferation of myocytes was estimated using immunohistochemical localization with antibodies against proliferating cell nuclear antigen (PCNA). Muscle overlying the distracted mandible showed 6-fold more PCNA-positive myocytes (16.8% +/- 11.3%) than the contralateral control side (2.8% +/- 1.1%, P < .0001). In the 2 sham-control animals, there was a low index of PCNA-positive myocytes on both the osteotomy (2%, 5%) and the contralateral sides (1%, 2%). The results of this study suggest that distraction of the porcine mandible by the protocol described induces myocyte proliferation in the masseter muscle. A proliferative response may contribute to improved long-term stability of mandibular expansion by distraction osteogenesis.

Mitogen- and stress-activated kinases (MSKs) are important substrates of the mitogen-activated protein kinase (MAPK)-activated protein kinase family. MSK1 and MSK2 are both nuclear serine/threonine protein kinases, with MSK1 being suggested to potentially play a role in breast cancer cell proliferation, cell cycle progression, cell migration, invasion and tumour growth. The aim of the current study was to assess MSK1 protein expression in breast cancer tumour specimens, evaluating its prognostic significance. A large cohort of 1902 early stage invasive breast cancer patients was used to explore the expression of MSK1. Protein expression was examined using standard immunohistochemistry on tissue microarrays. Low MSK1 protein expression was associated with younger age (P = 0.004), higher tumour grade (P < 0.001), higher Nottingham Prognostic Index scores (P = 0.007), negative ER (P < 0.001) and PR (P < 0.001) status, and with triple-negative (P < 0.001) and basal-like (P &...

Our recent study has shown that beetroot juice protects against N-nitrosodimethylamine (NDEA)-induced liver injury and increases the activity of phase II enzymes, suggesting the activation of the nuclear factor erythroid-2-related factor 2 (Nrf2)–antioxidant response element (ARE) pathway. The aim of the present study was to further explore the mechanism of the activity of beetroot by evaluating the cytoprotective effects of its major component. The influence of betanin (BET) on the activation of Nrf2 and the expression of GSTA, GSTP, GSTM, GSTT, NQO1 and HO-1 was assessed in two hepatic cell lines: non-tumour THLE-2 and hepatoma-derived HepG2 cell lines. The level of the tumour suppressor p53 in both cell lines and the methylation of GSTP in HepG2 cells were also evaluated. Treatment of both cell lines with 2, 10 and 20 μm of BET resulted in the translocation of Nrf2 from the cytosol to the nucleus. The mRNA and nuclear protein levels of Nrf2 and the binding of Nrf2 to ARE sequence...

A thorough understanding of the mechanisms of R-, C-and G-banding will come only from studies of the binding of Giemsa dyes to isolated and characterized preparations of heterochromatin and euchromatin. Since such studies require an exact knowledge of the optical characteristics of Giemsa, the spectral adsorption curves and extinction coefficients of Giemsa and its component dyes at various concentrations in the presence and absence of DNA were determined. — Although Giemsa is a complex mixture of thiazin dyes plus eosin; methylene blue, and azure A, B or C alone gave good banding. Thionin, with no methyl groups, gave poor or no banding. Eosin was not a necessary component for banding. — The most striking characteristic of the thiazin dyes is that they are strongly metachromatic, i.e., their adsorption spectra and extinction coefficients change as the concentration of the dye increases or as they bind to positively charged compounds (chromotropes). These changes, especially for methylene blue, are described in detail and allow a distinction between concentration dependent binding to DNA by intercalation and binding by side stacking.

Transient receptor potential channels (TRPC) are plasma membrane, non-selective cationic channels and have been proposed as candidates involved in the regulation of cellular Ca2+ influx. TRPC are involved in metabotropic glutamate receptor (mGluR)-mediated excitatory post-synaptic currents (EPSCs) in the dopaminergic neurons of the substantia nigra. We previously observed several subtypes of TRPC to be expressed at an mRNA level in

Previously we described a method to estimate the average number of virus genomes expressed in an infected cell. By analyzing the color spectrum of cells infected with a mixture of isogenic pseudorabies virus (PRV) recombinants expressing three fluorophores, we estimated that fewer than seven incoming genomes are expressed, replicated, and packaged into progeny per cell. In this report, we expand this work and describe experiments demonstrating the generality of the method, as well as providing more insight into herpesvirus replication. We used three isogenic PRV recombinants, each expressing a fluorescently tagged VP26 fusion protein (VP26 is a capsid protein) under the viral VP26 late promoter. We calculated a similar finite limit on the number of expressed viral genomes, indicating that this method is independent of the promoter used to transcribe the fluorophore genes, the time of expression of the fluorophore (early versus late), and the insertion site of the fluorophore gene in the PRV genome (UL versus US). Importantly, these VP26 fusion proteins are distributed equally in punctate virion assembly structures in each nucleus, which improves the signal-to-noise ratio when determining the color spectrum of each cell. To understand how the small number of genomes are distributed among the replication compartments, we used a two-color fluorescent in situ hybridization assay. Most viral replication compartments in the nucleus occupy unique nuclear territories, implying that they arose from single genomes. Our experiments suggest a correlation between the small number of expressed viral genomes and the limited number of replication compartments.

Analyses of genome orthologs in cancer on the background of tumor heterogeneity, coupled with the recent identification that the tumor propagating capacity resides within a very small fraction of cells (the tumor stem cells-TSCs), has not been achieved. Here, we describe a strategy to explore genetic drift in the mitochondrial genome accompanying varying stem cell dynamics in epithelial ovarian cancer. A major and novel outcome is the identification of a specific mutant mitochondrial DNA profile associated with the TSC lineage that is drastically different from the germ line profile. This profile, however, is often camouflaged in the primary tumor, and sometimes may not be detected even after metastases, questioning the validity of whole tumor profiling towards determining individual prognosis. Continuing mutagenesis in subsets with a mutant mitochondrial genome could result in transformation through a cooperative effect with nuclear genes - a representative example in our study is a tumor suppressor gene viz. cAMP responsive element binding binding protein. This specific profile could be a critical predisposing step undertaken by a normal stem cell to overcome a tightly regulated mutation rate and DNA repair in its evolution towards tumorigenesis. Our findings suggest that varying stem cell dynamics and mutagenesis define TSC progression that may clinically translate into increasing tumor aggression with serious implications for prognosis.

Parental (genomic) imprinting is the process by which the differential expression of maternal and paternal alleles at certain genetic loci in mammalian embryos occurs. Such loci are implicated in the control of fetal, placental and neonatal growth, and, more generally, in diverse aspects of fetal nutrient acquisition and maternal-fetal interactions. Not surprisingly, the aberrant expression of imprinted genes is implicated in a range of embryonic and fetal abnormalities. We outline how an evolutionary theory, based on classic parent-offspring conflict theory, relates to certain fetal growth abnormalities. In particular, we suggest that growth abnormalities resulting from the manipulation of preimplantation mammalian embryos in vitro (for example large calf syndrome) may reflect the occurrence of genetic conflict over the fetal growth programme in the early preimplantation period.

Store-operated calcium entry (SOCE) is the predominant Ca2+ entry mechanism in nonexcitable cells and controls a variety of physiological and pathological processes. Although significant progress has been made in identifying the components required for SOCE, the molecular mechanisms underlying it are elusive. The present study provides evidence for a direct involvement of kinase suppressor of Ras 2 (KSR2) in SOCE. Using lymphocytes and fibroblasts from ksr2−/− mice and shKSR2-depleted cells, we find that KSR2 is critical for the elevation of cytosolic Ca2+ concentration. Specifically, our results show that although it is dispensable for Ca2+-store depletion, KSR2 is required for optimal calcium entry. We observe that KSR2 deficiency affects stromal interaction molecule 1 (STIM1)/ORAI1 puncta formation, which is correlated with cytoskeleton disorganization. Of interest, we find that KSR2-associated calcineurin is crucial for SOCE. Blocking calcineurin activity impairs STIM1/ORAI1 pun...

There have been 2 putative prostatic cancer precursors, prostatic intraepithelial neoplasia (PIN) and atypical adenomatous hyperplasia (adenosis), but PIN remains as a well-known precancerous condition. To describe recent advances in knowledge of PIN and to better define the diagnostic criteria and differential diagnosis of PIN. Review of the pertinent literature and our experience. The presence of ductal/acinar epithelial changes including nuclear enlargement, prominent nucleoli, chromatin alterations, and luminal complexity is an easy way to identify the disorder. Four main patterns of high-grade PIN (HGPIN) have been described: tufting, micropapillary, cribriform, and flat. In addition, variants of HGPIN have also been described. Both HGPIN and prostatic carcinoma share an increased incidence and severity with advancing age and with high rates of occurrence in the peripheral zone of the prostate. Furthermore, HGPIN and prostate cancer share genetic and molecular markers as well, ...

Coiled bodies are nuclear organelles that contain components of at least three RNA-processing pathways: pre-mRNA splicing, histone mRNA 3′- maturation, and pre-rRNA processing. Their function remains unknown. However, it has been speculated that coiled bodies may be sites of splicing factor assembly and/or recycling, play a role in histone mRNA 3′-processing, or act as nuclear transport or sorting structures. To study the dynamics of coiled bodies in living cells, we have stably expressed a U2B"–green fluorescent protein fusion in tobacco BY-2 cells and in Arabidopsis plants. Time-lapse confocal microscopy has shown that coiled bodies are mobile organelles in plant cells. We have observed movements of coiled bodies in the nucleolus, in the nucleoplasm, and from the periphery of the nucleus into the nucleolus, which suggests a transport function for coiled bodies. Furthermore, we have observed coalescence of coiled bodies, which suggests a mechanism for the decrease in coiled bo...

Growing evidence suggests that oxidative stress is involved in the neuronal degeneration and can promote the aggregation of α-synuclein. However, the role of α-synuclein under physiological and pathological conditions remains poorly understood. In the present study, we examined the possible interaction between the α-synuclein and oxidative stress. In a dopaminergic cell line MES23.5, we have found that the 200μM H2O2

Subunit B of diphtheria toxin (DT), which consists of two domains: R (receptor-binding) and T (transmembrane), plays an important role in toxin-receptor binding on the cell-targets and in transportation of catalytic subunit A to the cell cytosol. Recombinant analogues of the subunit B are promising representatives in the unique class of transporting proteins, able to deliver different types of biologically active molecules to cell cytosol. In the development of these protein constructs understanding of the role of each of the DT fragments in determination of transporting pathways of endocytosed complex toxin-receptor is urgently required. We have studied in this work the T-domain effect on intracellular transport of recombinant fragments of DT. We have compared intracellular transport of the R-domain and the subunit B, the last one was consisted of both R-domain and T-domain. Recombinant fragments of DT used in this work were labeled with fluorescent proteins, which allowed applying...

The DJ-1 gene encodes a ubiquitous, highly conserved protein. Here, we show that DJ-1 mutations are associated with PARK7, a monogenic form of human parkinsonism. The function of the DJ-1 protein remains unknown, but evidence suggests its involvement in the oxidative stress response. Our findings indicate that loss of DJ-1 function leads to neurodegeneration. Elucidating the physiological role of DJ-1 protein may promote understanding of the mechanisms of brain neuronal maintenance and pathogenesis of Parkinson's disease.