During the development of the mammalian neuromuscular junction, acetylcholine receptors (AChRs) become localized to the postsynaptic muscle membrane. As this process nears completion, the fetal form ...of the receptor, containing a γ subunit (composition α2βγδ) is gradually replaced by an ϵ subunit-containing adult form (α2βϵδ). To understand how this transition is controlled, we compared the expression and regulation of the AChR γ and ϵ subunits in developing, adult, and cultured muscles. Immunostaining with subunit-specific antibodies showed that replacement of γ subunit- by ϵ subunit-containing AChRs occurs largely during the first postnatal week in fast-twitch muscles, and occurs homogeneously throughout individual endplates. In the slow-twitch soleus, however, this transition is delayed, and in the multiply innervated slow fibers of extraocular muscle, γ subunit expression persists into adulthood. The transcriptional bases of the AChR subunit transition, and of these intermuscular variations, were demonstrated in mice bearing transgenes containing promoter elements from the AChR γ and ϵ subunit genes, each coupled to a nuclear-localized β-galactosidase (nlacZ) reporter. We show that transgene expression is stimulated by the nerve-derived inducer of AChR expression, ARIA, in myotubes cultured from γ–nlacZ as well as ϵ–nlacZ mice. However, the expression of γ–nlacZ, but not ϵ–nlacZ, is increased by treatment of myotubes with TTX, and the ARIA sensitivity of γ–nlacZ is dependent on the electrical state of the myotube. Thus, the promoters of the γ and ϵ subunit genes may integrate ARIA- and activity-dependent signals in different ways to generate their complementary patterns of expression.
AChR-inducing activity (ARIA)/heregulin, a ligand for erbB receptor tyrosine kinases (RTKs), is likely to be one nerve-supplied signal that induces expression of acetylcholine receptor (AChR) genes ...at the developing neuromuscular junction. Since some RTKs act through Ras and phosphatidylinositol 3-kinase (PI3K), we investigated the role of these pathways in ARIA signaling. Expression of activated Ras or Raf mimicked ARIA-induction of AChR ε subunit genes in muscle cells; whereas dominant negative Ras or Raf blocked the effect of ARIA. ARIA rapidly activated erk1 and erk2 and inhibition of both erks also abolished the effect of ARIA. ARIA stimulated association of PI3K with erbB3, expression of an activated PI3K led to ARIA-independent AChR ε subunit expression, and inhibition of PI3K abolished the action of ARIA. Thus, synaptic induction of AChR genes requires activation of both Ras/MAPK and PI3K signal transduction pathways.
Little is known about the sociodemographic and clinical profile of older adults with intellectual disabilities (ID) in Singapore. We studied the sociodemographic and clinical profile of older adults ...with ID and investigated factors associated with caregiver availability and identity in this population.
The study population involved all adults with ID aged ≥40 years receiving services from the Movement for the Intellectually Disabled of Singapore (MINDS), the largest such provider in Singapore. Information on sociodemographic and clinical profiles, functional status, and availability of caregivers were collected via interviewer-administered questionnaires from guardians of older adults with ID. Descriptive characteristics were computed and chi-square and logistic regression identifi ed predictors of caregiver availability and identity.
Participation was 95% (227/239). There were differences in client age, gender, and caregiver availability between recipients of residential and non-residential services (all P <0.05). Common comorbidities included hyperlipidaemia (17.6%), hypertension (15.9%), psychiatric diagnoses (16.3%) and epilepsy (10.6%). The majority were fully independent in basic activities of daily living, but only 21.1% were fully communicative. Only a small minority (9.4%) were exercising regularly. The majority (73.5%) of clients had a primary caregiver; almost equal proportions relied on either parents or siblings. Older client age was associated independently with the lack of a primary caregiver, independent of greater functional dependence and presence of medical comorbidities in the client.
Older adults with ID have multiple medical, functional, and social issues. More can be done to support the care of this unique group of adults with special needs.
To determine the role of telomere dysfunction and telomerase reactivation in generating pro-oncogenic genomic events and in carcinoma progression, an inducible telomerase reverse transcriptase (
...mTert
) allele was crossed onto a prostate cancer-prone mouse model null for
Pten
and
p53
tumor suppressors. Constitutive telomerase deficiency and associated telomere dysfunction constrained cancer progression. In contrast, telomerase reactivation in the setting of telomere dysfunction alleviated intratumoral DNA damage signaling and generated aggressive cancers with rearranged genomes and new tumor biological properties (bone metastases). Comparative oncogenomic analysis revealed numerous recurrent amplifications and deletions of relevance to human prostate cancer. Murine tumors show enrichment of the TGFβ/SMAD4 network and genetic validation studies confirmed the cooperative roles of
Pten, p53
and
Smad4
deficiencies in prostate cancer progression including skeletal metastases. Thus, telomerase reactivation in tumor cells experiencing telomere dysfunction enables full malignant progression and provides a mechanism for acquisition of cancer-relevant genomic events endowing new tumor biological capabilities.
Developing motor axons induce synaptic specializations in muscle fibers, including preferential transcription of acetylcholine receptor (AChR) subunit genes by subsynaptic nuclei. One candidate ...nerve-derived signaling molecule is AChR-inducing activity (ARIA)/heregulin, a ligand of the erbB family of receptor tyrosine kinases. Here, we asked whether ARIA and erbB kinases are expressed in patterns compatible with their proposed signaling roles. In developing muscle, ARIA was present not only at synaptic sites, but also in extrasynaptic regions of the muscle fiber. ARIA was synthesized, rather than merely taken up, by muscle cells, as indicated by the presence of ARIA mRNA in muscle and of ARIA protein in a clonal muscle cell line. ARIA-responsive myotubes expressed both erbB2 and erbB3, but little EGFR/erbB1 or erbB4. In adults, erbB2 and erbB3 were localized to the postsynaptic membrane. ErbB3 was restricted to the postsynaptic membrane perinatally, at a time when ARIA was still broadly distributed. Thus, our data are consistent with a model in which ARIA interacts with erbB kinases on the muscle cell surface to provide a local signal that induces synaptic expression of AChR genes. However, much of the ARIA is produced by muscle, not nerve, and the spatially restricted response may result from the localization of erbB kinases as well as of ARIA. Finally, we show that erbB3 is not concentrated at synaptic sites in mutant mice that lack rapsyn, a cytoskeletal protein required for AChR clustering, suggesting that pathways for synaptic AChR expression and clustering interact.
Clinical and genomic evidence suggests that the metastatic potential of a primary tumor may be dictated by pro-metastatic events that have additional oncogenic capability. To test this deterministic ...hypothesis, we adopted a comparative oncogenomics-guided function-based strategy involving (i) comparison of global transcriptomes of two genetically engineered mouse models with contrasting metastatic potential, (ii) genomic and transcriptomic profiles of human melanoma, (iii) functional genetic screen for enhancers of cell invasion and (iv) evidence of expression selection in human melanoma tissues. This integrated effort identified 6 genes that are potently pro-invasive and oncogenic. Further, we show that one such gene, ACP5, confers spontaneous metastasis
in vivo
, engages a key pathway governing metastasis and is prognostic in human primary melanomas.
Two independent studies by two separate research teams (from Hong Kong and Singapore) failed to detect any influenza RNA landing on, or inhaled by, a life-like, human manikin target, after exposure ...to naturally influenza-infected volunteers. For the Hong Kong experiments, 9 influenza-infected volunteers were recruited to breathe, talk/count and cough, from 0.1 m and 0.5 m distance, onto a mouth-breathing manikin. Aerosolised droplets exhaled from the volunteers and entering the manikin's mouth were collected with PTFE filters and an aerosol sampler, in separate experiments. Virus detection was performed using an in-house influenza RNA reverse-transcription polymerase chain reaction (RT-PCR) assay. No influenza RNA was detected from any of the PTFE filters or air samples. For the Singapore experiments, 6 influenza-infected volunteers were asked to breathe (nasal/mouth breathing), talk (counting in English/second language), cough (from 1 m/0.1 m away) and laugh, onto a thermal, breathing manikin. The manikin's face was swabbed at specific points (around both eyes, the nostrils and the mouth) before and after exposure to each of these respiratory activities, and was cleaned between each activity with medical grade alcohol swabs. Shadowgraph imaging was used to record the generation of these respiratory aerosols from the infected volunteers and their impact onto the target manikin. No influenza RNA was detected from any of these swabs with either team's in-house diagnostic influenza assays. All the influenza-infected volunteers had diagnostic swabs taken at recruitment that confirmed influenza (A/H1, A/H3 or B) infection with high viral loads, ranging from 105-108 copies/mL (Hong Kong volunteers/assay) and 104-107 copies/mL influenza viral RNA (Singapore volunteers/assay). These findings suggest that influenza RNA may not be readily transmitted from naturally-infected human source to susceptible recipients via these natural respiratory activities, within these exposure time-frames. Various reasons are discussed in an attempt to explain these findings.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Activating KRAS mutations and p16Ink4a inactivation are near universal events in human pancreatic ductal adenocarcinoma (PDAC). In mouse models, KrasG12D initiates formation of premalignant ...pancreatic ductal lesions, and loss of either Ink4a/Arf (p16Ink4a/p19Arf) or p53 enables their malignant progression. As recent mouse modeling studies have suggested a less prominent role for p16Ink4a in constraining malignant progression, we sought to assess the pathological and genomic impact of inactivation of p16Ink4a, p19Arf, and/or p53 in the KrasG12D model. Rapidly progressive PDAC was observed in the setting of homozygous deletion of either p53 or p16Ink4a, the latter with intact germ-line p53 and p19Arf sequences. Additionally, KrasG12D in the context of heterozygosity either for p53 plus p16Ink4a or for p16Ink4a/p19Arf produced PDAC with longer latency and greater propensity for distant metastases relative to mice with homozygous deletion of p53 or p16Ink4a/p19Arf. Tumors from the double-heterozygous cohorts showed frequent p16Ink4a inactivation and loss of either p53 or p19Arf. Different genotypes were associated with specific histopathologic characteristics, most notably a trend toward less differentiated features in the homozygous p16Ink4a/p19Arf mutant model. High-resolution genomic analysis revealed that the tumor suppressor genotype influenced the specific genomic patterns of these tumors and showed overlap in regional chromosomal alterations between murine and human PDAC. Collectively, our results establish that disruptions of p16Ink4a and the p19ARF-p53 circuit play critical and cooperative roles in PDAC progression, with specific tumor suppressor genotypes provocatively influencing the tumor biological phenotypes and genomic profiles of the resultant tumors. PUBLICATION ABSTRACT
Genetic and biochemical data have identified Smad4 as a key intracellular effector of the transforming growth factor β (TGFβ superfamily of secreted ligands. In mouse, Smad4 -null embryos do not ...gastrulate, a phenotype consistent with loss of other TGFβ-related signaling components. Chimeric analysis reveals a primary requirement for Smad4 in the extra-embryonic lineages; however, within the embryo proper, characterization of the specific roles of Smad4 during gastrulation and lineage specification remains limited. We have employed a Smad4 conditional allele to specifically inactivate the Smad4 gene in the early mouse epiblast. Loss of Smad4 in this tissue results in a profound failure to pattern derivatives of the anterior primitive streak, such as prechordal plate, node, notochord and definitive endoderm. In contrast to these focal defects, many well-characterized TGFβ- and Bmp-regulated processes involved in mesoderm formation and patterning are surprisingly unaffected. Mutant embryos form abundant extra-embryonic mesoderm, including allantois, a rudimentary heart and middle primitive streak derivatives such as somites and lateral plate mesoderm. Thus, loss of Smad4 in the epiblast results not in global developmental abnormalities but instead in restricted patterning defects. These results suggest that Smad4 potentiates a subset of TGFβ-related signals during early embryonic development, but is dispensable for others.
Genetic and biochemical data have identified Smad4 as a key intracellular effector of the transforming growth factor beta (TGFbeta superfamily of secreted ligands. In mouse, Smad4-null embryos do not ...gastrulate, a phenotype consistent with loss of other TGFbeta-related signaling components. Chimeric analysis reveals a primary requirement for Smad4 in the extra-embryonic lineages; however, within the embryo proper, characterization of the specific roles of Smad4 during gastrulation and lineage specification remains limited. We have employed a Smad4 conditional allele to specifically inactivate the Smad4 gene in the early mouse epiblast. Loss of Smad4 in this tissue results in a profound failure to pattern derivatives of the anterior primitive streak, such as prechordal plate, node, notochord and definitive endoderm. In contrast to these focal defects, many well-characterized TGFbeta- and Bmp-regulated processes involved in mesoderm formation and patterning are surprisingly unaffected. Mutant embryos form abundant extra-embryonic mesoderm, including allantois, a rudimentary heart and middle primitive streak derivatives such as somites and lateral plate mesoderm. Thus, loss of Smad4 in the epiblast results not in global developmental abnormalities but instead in restricted patterning defects. These results suggest that Smad4 potentiates a subset of TGFbeta-related signals during early embryonic development, but is dispensable for others.
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