Mutations inhibiting KDM4B drive ALT activation in ATRX-mutated glioblastomas.

Alternative Lengthening of Telomeres (ALT) is a telomere maintenance pathway utilised in 15% of cancers. ALT cancers are strongly associated with inactivating mutations in ATRX; yet loss of ATRX alone is insufficient to trigger ALT, suggesting that additional cooperating factors are involved. We identify H3.3G34R and IDH1/2 mutations as two such factors in ATRX-mutated glioblastomas. Both mutations are capable of inactivating histone demethylases, and we identify KDM4B as the key demethylase inactivated in ALT. Mouse embryonic stem cells inactivated for ATRX, TP53, TERT and KDM4B (KDM4B knockout or H3.3G34R) show characteristic features of ALT. Conversely, KDM4B over-expression in ALT cancer cells abrogates ALT-associated features. In this work, we demonstrate that inactivation of KDM4B, through H3.3G34R or IDH1/2 mutations, acts in tandem with ATRX mutations to promote ALT in glioblastomas.

Empowerment of nurses in antibiotic stewardship: a social ecological qualitative analysis.

BACKGROUND: Inappropriate antibiotics use and antimicrobial resistance (AMR) are increasingly becoming global health issues of great concern. Despite the established antibiotic stewardship programmes (ASPs) in many countries, limited efforts have been made to engage nurses and clearly define their roles in ASPs. AIM: An exploratory qualitative study was conducted to understand the facilitators and barriers that impact nurses' involvement and empowerment in antibiotic stewardship. METHODS: Focus group discussions (FGDs) were conducted with purposively sampled nurses from three major public hospitals in Singapore. FGDs were audio-recorded and transcribed verbatim. Data were analysed using Applied Thematic Analysis and interpreted using the Social Ecological Model. FINDINGS: At the intrapersonal level, nurses felt empowered in carrying out their roles in antibiotic administration. They saw themselves as gatekeepers to ensure that the prescribed antibiotics were administered appropriately. However, nurses felt they lacked the knowledge and expertise in antibiotic use and AMR prevention. At the interpersonal level, this deficit in knowledge and expertise in antibiotic use impacted how they were perceived by patients and caregivers as well as their interactions with the primary care team when voicing outpatient safety concerns and antibiotic administration suggestions. At the organizational level, nurses relied on drug administration guidelines to ensure appropriate antibiotic administration and as a safety net when physicians questioned their clinical practice. At the community level, nurses felt there was a lack of awareness and knowledge on antibiotic use among the general population. CONCLUSION: These findings provide important insights to harness the contributions of nurses, and to formally acknowledge and enlarge their roles in ASPs.

Racial Differences Affecting Night Time Blood Pressure Dipping Groups in Hypertensive Patients.

BACKGROUND: Normal blood pressure (BP) follows a circadian rhythm, with dipping of BP at night. However, little has been done to show how the dipping groups vary amongst the White and Asian population at different periods of the year. This study aims to examine the pattern of nocturnal dipping between the White and Asian population, as well as to compare it to the different timings of the year, between summer and winter. METHODS: Ambulatory Blood Pressure Monitor recordings were obtained from 220 patients, half were White patients obtained from Mercy University Hospital, Cork, Ireland and half were Asian patients from National Heart Centre, Singapore during the summer period from May to June and the winter period from October to December. RESULTS: Both the Irish and Singaporeans exhibit a decrease in total number of reverse dipper from summer to winter. However, the redistribution of reverse dipper was mainly to the dippers in Singapore, while in Ireland it was to both the extreme dipper and dipper. Irish seasonal changes also resulted in an increase in nocturnal diastolic pressure (95% CI, 0.72 to 6.03, 3.37 mm Hg; p<0.05) and a change in the duration of dipping at night (95% CI, 0.045 to 1.01, 0.53 Hours; p<0.05). CONCLUSION: Regardless of race or temperature, reverse dippers seem to decrease in winter. However, the racial differences dictate the redistribution of the fall in number of dippers. This has implications on how reverse dippers should be treated at different periods of the year.

Chemical welding of binary nanoparticles: room temperature sintering of CuSe and In2S3 nanoparticles for solution-processed CuInS(x)Se(1-x) solar cells.

Chemical welding of oppositely charged dissimilar metal chalcogenide nanomaterials is reported to produce a quaternary metal chalcogenide. CuSe and In2S3 nanoparticles were synthesized with opposite surface charges by stabilizing with polyacrylic acid and polydiallyldimethylammonium chloride. Upon mixing these nanoparticles at room temperature, the electrostatic attraction induced coalescence of these nanoparticles and led to the formation of CuInSxSe1-x nanoparticles.

Centromere protein b-null mice display decreasing reproductive performance through successive generations of breeding due to diminishing endometrial glands.

Centromere protein B is a highly conserved constitutive protein found at centromeres. Gene knockout studies in mice have unexpectedly identified Cenpb as a candidate gene involved in uterine function. The present study further explores the role of Cenpb in mice by intermating Cenpb-null mice over several generations. Breeding studies and analysis of uterine tissue indicate that Cenpb-null mice lose reproductive fitness over a number of generations due to a significant reduction in endometrial glands. These results suggest that Cenpb may play an important function in the short- and long-term maintenance of uterine integrity.

Construction of neocentromere-based human minichromosomes for gene delivery and centromere studies.

Human neocentromeres are fully functional centromeres that arise naturally in non-centromeric regions devoid of alpha-satellite DNA. We have successfully produced a series of minichromosomes by telomere-associated truncation of a marker chromosome mardel(10) containing a neocentromere. The resulting minichromosomes are either linear or circular in nature, and range in size from approximately 650 kb to 2 Mb. These minichromosomes exhibit full centromeric activity, bind to essential centromere proteins, and are mitotically stable over many generations. They provide a useful system for dissecting the functional domains of complex eukaryotic centromeres and as vectors for therapeutic gene delivery.

Construction of neocentromere-based human minichromosomes by telomere-associated chromosomal truncation.

Neocentromeres (NCs) are fully functional centromeres that arise ectopically in noncentromeric regions lacking alpha-satellite DNA. Using telomere-associated chromosome truncation, we have produced a series of minichromosomes (MiCs) from a mardel(10) marker chromosome containing a previously characterized human NC. These MiCs range in size from approximately 0.7 to 1.8 Mb and contain single-copy intact genomic DNA from the 10q25 region. Two of these NC-based Mi-Cs (NC-MiCs) appear circular whereas one is linear. All demonstrate stability in both structure and mitotic transmission in the absence of drug selection. Presence of a functional NC is shown by binding a host of key centromere-associated proteins. These NC-MiCs provide direct evidence for mitotic segregation function of the NC DNA and represent examples of stable mammalian MiCs lacking centromeric repeats.

IFN-gamma priming up-regulates IFN-stimulated gene factor 3 (ISGF3) components, augmenting responsiveness of IFN-resistant melanoma cells to type I IFNs.

IFN-stimulated gene factor 3 (ISGF3) mediates transcriptional activation of IFN-sensitive genes (ISGs). The component subunits of ISGF3, STAT1alphabeta, STAT2, and p48-ISGF3gamma, are tyrosine phosphorylated before their assembly into a complex. Subsequently, the ISGF3 complex is translocated to the nucleus. We have recently established that the responsiveness of human melanoma cell lines to type I IFNs correlates directly with their intracellular levels of ISGF3 components, particularly STAT1. In the present study, we show that pretreating IFN-resistant melanoma cell lines with IFN-gamma (IFN-gamma priming) before stimulation with type I IFN also results in increased levels of ISGF3 components and enhanced DNA-binding activation of ISGF3. In addition, IFN-gamma priming of IFN-resistant melanoma cell lines increased expression of type I IFN-induced ISG products, including ISG54, 2'-5'-oligoadenylate synthase, HLA class I, B7-1, and ICAM-1 Ags. Furthermore, IFN-gamma priming enhanced the antiviral effect of IFN-beta on the IFN-resistant melanoma cell line, MM96. These results support a role for IFN-gamma priming in up-regulating ISGF3, thereby augmenting the responsiveness of IFN-resistant melanoma cell lines to type I IFN and providing a molecular basis and justification for using sequential IFN therapy, as proposed by others, to enhance the use of IFNs in the treatment of melanoma.

Interferon-resistant human melanoma cells are deficient in ISGF3 components, STAT1, STAT2, and p48-ISGF3gamma.

The mechanism of IFN resistance was examined in three long-term cell lines, SK-MEL-28, SK-MEL-3, and MM96, exhibiting significant variation in responsiveness to the antiproliferative and antiviral effects of type I IFNs. The JAK-STAT components involved in IFN signal transduction were analyzed in detail. After exposure to IFN, activation of the IFN type I receptor-linked tyrosine kinases, JAK-1 and TYK-2, was detected at similar levels in both IFN-sensitive and IFN-resistant cell types, indicating that IFN resistance did not result from a deficiency in signaling at the level of receptor-associated kinase activation. However, analysis of ISGF3 transcription factor components, STAT1, STAT2, and p48-ISGF3gamma, revealed that their expression and activation correlated with cellular IFN responsiveness. The analysis was extended to also include IFN-sensitive primary melanocytes, three additional IFN-resistant melanoma cell lines, and seven cell cultures recently established from melanoma patient biopsies. It was consistently observed that the most marked difference in ISGF3 was a lack of STAT1 in the resistant versus the sensitive cells. Transfection of the IFN-resistant MM96 cell line to express increased levels of STAT1 protein partially restored IFN responsiveness in an antiviral assay. We conclude that a defect in the level of STAT1 and possibly all three ISGF3 components in IFN-resistant human melanoma cells may be a general phenomenon responsible for reduced cellular responsiveness of melanomas to IFNs.

The SH2 domains of Stat1 and Stat2 mediate multiple interactions in the transduction of IFN-alpha signals.

Analysis of the ability of IFN-alpha to rapidly stimulate genes has led to the identification of a new family of signal transducing factors (STFs). The STF activated by IFN-alpha consists of two members of the STAT (Signal Transducers and Activators of Transcription) family of signaling proteins, Stat1 and Stat2. Sequence comparison of these STATs has determined that they share several conserved domains, the most notable of which is an SH2 domain. Recently, studies have determined that these SH2 domains can mediate a specific association between a STAT and the cytoplasmic domain of the appropriate receptor. Once associated with the receptor, the STATs become activated by an associated tyrosine kinase, whereupon they dissociate from the receptor and dimerize to form active STFs. The SH2 domain has also been implicated in the formation of STAT dimers found in STFs, suggesting it may play multiple roles in signaling. We have carried out a detailed analysis on the role of the Stat1 and Stat2 SH2 domains in the mediation of IFN-alpha stimulated signals. These studies, which have determined that the SH2 domain of Stat1 and Stat2 can mediate homo- as well as heterodimerization, suggest that a single SH2 domain-phosphotyrosyl interaction is sufficient for dimerization. Moreover, they provide the first direct evidence that the target of the SH2 domain is the STAT tyrosine activation site. In addition, these studies implicate the SH2 domain in another step in the signaling cascade, namely mediation of the interaction between STATs and their activating kinases (i.e. the JAKs).