Strain persistence of invasive Candida albicans in chronic hyperplastic candidosis that underwent malignant change

Objectives: The aim of this study was to assess persistence and tissue invasion of Candida albicans strains isolated from a 65 year‐old patient with chronic hyperplastic candidosis (CHC), that subsequently developed into squamous cell carcinoma (SCC). Materials and Methods: C. albicans (n=7) were recovered from the oral cavity of the patient over seven years. Confirmation of CHC and SCC in this patient was achieved by histopathological examination of incisional biopsy tissue. DNA fingerprinting was performed on the seven isolates from the CHC patient together with a further eight isolates from patients with normal oral mucosa (n=2), chronic atrophic candidosis (n=1), SCC (n=1) and CHC (n=4). Genotyping involved the use of inter‐repeat PCR using the eukaryotic repeat primer 1251. Characterisation of the tissue invasive abilities of the isolates was achieved by infecting a commercially available reconstituted human oral epithelium (RHE; SkinEthic, Nice, France). After 24 h. C. albicans tissue invasion was assessed by histopathological examination. Results: DNA fingerprinting demonstrated strain persistence of C. albicans in the CHC patient over a seven year period despite provision of systemic antifungal therapy. The strain of C. albicans isolated from this patient was categorised as a high invader within the RHE compared to other isolates. Conclusions: Candidal strain persistence was evident in a patient with CHC over seven years. This persistence may be due to incomplete eradication from the oral cavity following antifungal therapy or subsequent recolonisation from other body sites or separate exogenous sources. The demonstration of enhanced in vitro tissue invasion by this particular strain may, in part, explain the progression to carcinoma.     

In vitro zygotic embryo culture of wild Musa acuminata ssp.malaccensis and factors affecting germination and seedling growth

In vitro zygotic embryo culture of wild banana significantly increased the germination compared to greenhouse grown seeds. Embryo orientation and BAP concentration significantly affected germination rate. These factors together with gelling agent, dark and light conditions and coconut water, also showed variable effects on the number of roots per plant, root length, shoot length, number of days to root emergence and number of days to shoot emergence.     

Regulation of global acetylation in mitosis through loss of histone acetyltransferases and deacetylases from chromatin

Histone acetylation, a reversible modification of the core histones, is widely accepted to be involved in remodeling chromatin organization for genetic reprogramming. Histone acetylation is a dynamic process that is regulated by two classes of enzymes, the histone acetyltransferases (HATs) and histone deacetylases (HDACs). Although promoter-specific acetylation and deacetylation has received most of the recent attention, it is superimposed upon a broader acting and dynamic acetylation that profoundly affects many nuclear processes. In this study, we monitored this broader histone acetylation as cells enter and exit mitosis. In contrast to the hypothesis that HATs and HDACs remain bound to mitotic chromosomes to provide an epigenetic imprint for postmitotic reactivation of the genome, we observed that HATs and HDACs are spatially reorganized and displaced from condensing chromosomes as cells progress through mitosis. During mitosis, HATs and HDACs are unable to acetylate or deacetylate chromatin in situ despite remaining fully catalytically active when isolated from mitotic cells and assayed in vitro. Our results demonstrate that HATs and HDACs do not stably bind to the genome to function as an epigenetic mechanism of selective postmitotic gene activation. Our results, however, do support a role for spatial organization of these enzymes within the cell nucleus and their relationship to euchromatin and heterochromatin postmitotically in the reactivation of the genome.     

Mast cell degranulation requires N-ethylmaleimide-sensitive factor-mediated SNARE disassembly

Mast cells possess specialized granules that, upon stimulation of surface FcR with IgE, fuse with the plasma membrane, thereby releasing inflammatory mediators. A family of membrane fusion proteins called SNAREs, which are present on both the granule and the plasma membrane, plays a role in the fusion of these granules with the plasma membrane of mast cells. In addition to the SNAREs themselves, it is likely that the SNARE accessory protein, N-ethylmaleimide-sensitive factor (NSF), affects the composition and structure of the SNARE complex. NSF is a cytoplasmic ATPase that disassembles the SNARE complexes. To investigate the role of NSF in mast cell degranulation, we developed an assay to measure secretion from transiently transfected RBL (rat basophilic leukemia)-2H3 mast cells (a tumor analog of mucosal mast cells). RBL-2H3 cells were cotransfected with a plasmid encoding a human growth hormone secretion reporter along with either wild-type NSF or an NSF mutant that lacks ATPase activity. Human growth hormone was targeted to and released from secretory granules in RBL-2H3 cells, and coexpression with mutant NSF dramatically inhibited regulated exocytosis from the transfected cells. Biochemical analysis of SNARE complexes in these cells revealed that overexpression of the NSF mutant decreased disassembly and resulted in an accumulation of SNARE complexes. These data reveal a role for NSF in mast cell exocytosis and highlight the importance of SNARE disassembly, or priming, in regulated exocytosis from mast cells.     

Heterochromatin is refractory to gamma-H2AX modification in yeast and mammals

Double-strand break (DSB) damage in yeast and mammalian cells induces the rapid ATM (ataxia telangiectasia mutated)/ATR (ataxia telangiectasia and Rad3 related)-dependent phosphorylation of histone H2AX (gamma-H2AX). In budding yeast, a single endonuclease-induced DSB triggers gamma-H2AX modification of 50 kb on either side of the DSB. The extent of gamma-H2AX spreading does not depend on the chromosomal sequences. DNA resection after DSB formation causes the slow, progressive loss of gamma-H2AX from single-stranded DNA and, after several hours, the Mec1 (ATR)-dependent spreading of gamma-H2AX to more distant regions. Heterochromatic sequences are only weakly modified upon insertion of a 3-kb silent HMR locus into a gamma-H2AX-covered region. The presence of heterochromatin does not stop the phosphorylation of chromatin more distant from the DSB. In mouse embryo fibroblasts, gamma-H2AX distribution shows that gamma-H2AX foci increase in size as chromatin becomes more accessible. In yeast, we see a high level of constitutive gamma-H2AX in telomere regions in the absence of any exogenous DNA damage, suggesting that yeast chromosome ends are transiently detected as DSBs.     

Further phenotypic delineation of subtelomeric (terminal) 4q deletion with emphasis on intracranial and reproductive anatomy

Hypoplastic left heart syndrome(HLHS) refers to the abnormal development of the left-sided cardiac structures, resulting in obstruction to blood flow from the left ventricular outflow tract. In addition, the syndrome includes underdevelopment of the left ventricle, aorta, and aortic arch, as well as mitral atresia or stenosis. HLHS has been reported to occur in approximately 0.016 to 0.036% of all live births. Newborn infants with the condition generally are born at full term and initially appear healthy. As the arterial duct closes, the systemic perfusion becomes decreased, resulting in hypoxemia, acidosis, and shock. Usually, no heart murmur, or a non-specific heart murmur, may be detected. The second heart sound is loud and single because of aortic atresia. Often the liver is enlarged secondary to congestive heart failure. The embryologic cause of the disease, as in the case of most congenital cardiac defects, is not fully known. The most useful diagnostic modality is the echocardiogram. The syndrome can be diagnosed by fetal echocardiography between 18 and 22 weeks of gestation. Differential diagnosis includes other left-sided obstructive lesions where the systemic circulation is dependent on ductal flow (critical aortic stenosis, coarctation of the aorta, interrupted aortic arch). Children with the syndrome require surgery as neonates, as they have duct-dependent systemic circulation. Currently, there are two major modalities, primary cardiac transplantation or a series of staged functionally univentricular palliations. The treatment chosen is dependent on the preference of the institution, its experience, and also preference. Although survival following initial surgical intervention has improved significantly over the last 20 years, significant mortality and morbidity are present for both surgical strategies. As a result pediatric cardiologists continue to be challenged by discussions with families regarding initial decision relative to treatment, and long-term prognosis as information on long-term survival and quality of life for those born with the syndrome is limited.     

Expression of the SNARE Protein SNAP-23 Is Essential for Cell Survival

Members of the SNARE-family of proteins are known to be key regulators of the membrane-membrane fusion events required for intracellular membrane traffic. The ubiquitously expressed SNARE protein SNAP-23 regulates a wide variety of exocytosis events and is essential for mouse development. Germline deletion of SNAP-23 results in early embryonic lethality in mice, and for this reason we now describe mice and cell lines in which SNAP-23 can be conditionally-deleted using Cre-lox technology. Deletion of SNAP-23 in CD19-Cre expressing mice prevents B lymphocyte development and deletion of SNAP-23 using a variety of T lymphocyte-specific Cre mice prevents T lymphocyte development. Acute depletion of SNAP-23 in mouse fibroblasts leads to rapid apoptotic cell death. These data highlight the importance of SNAP-23 for cell survival and describe a mouse in which specific cell types can be eliminated by expression of tissue-specific Cre-recombinase.     

Role of Nbs1 in the activation of the Atm kinase revealed in humanized mouse models

Nijmegen breakage syndrome (NBS) is a chromosomal fragility disorder that shares clinical and cellular features with ataxia telangiectasia. Here we demonstrate that Nbs1-null B cells are defective in the activation of ataxia-telangiectasia-mutated (Atm) in response to ionizing radiation, whereas ataxia-telangiectasia- and Rad3-related (Atr)-dependent signalling and Atm activation in response to ultraviolet light, inhibitors of DNA replication, or hypotonic stress are intact. Expression of the main human NBS allele rescues the lethality of Nbs1-/- mice, but leads to immunodeficiency, cancer predisposition, a defect in meiotic progression in females and cell-cycle checkpoint defects that are associated with a partial reduction in Atm activity. The Mre11 interaction domain of Nbs1 is essential for viability, whereas the Forkhead-associated (FHA) domain is required for T-cell and oocyte development and efficient DNA damage signalling. Reconstitution of Nbs1 knockout mice with various mutant isoforms demonstrates the biological impact of impaired Nbs1 function at the cellular and organismal level.     

Hip joint replacement surgery for idiopathic osteoarthritis aggregates in families

In order to determine whether there is a genetic component to hip or knee joint failure due to idiopathic osteoarthritis (OA), we invited patients (probands) undergoing hip or knee arthroplasty for management of idiopathic OA to provide detailed family histories regarding the prevalence of idiopathic OA requiring joint replacement in their siblings. We also invited their spouses to provide detailed family histories about their siblings to serve as a control group. In the probands, we confirmed the diagnosis of idiopathic OA using American College of Rheumatology criteria. The cohorts included the siblings of 635 probands undergoing total hip replacement, the siblings of 486 probands undergoing total knee replacement, and the siblings of 787 spouses. We compared the prevalence of arthroplasty for idiopathic OA among the siblings of the probands with that among the siblings of the spouses, and we used logistic regression to identify independent risk factors for hip and knee arthroplasty in the siblings. Familial aggregation for hip arthroplasty, but not for knee arthroplasty, was observed after controlling for age and sex, suggesting a genetic contribution to end-stage hip OA but not to end-stage knee OA. We conclude that attempts to identify genes that predispose to idiopathic OA resulting in joint failure are more likely to be successful in patients with hip OA than in those with knee OA.