Modulation of Ki67 and myogenic regulatory factor expression by tocotrienol-rich fraction ameliorates myogenic program of senescent human myoblasts

IntroductionReplicative senescence results in dysregulation of cell proliferation and differentiation, which plays a role in the regenerative defects observed during age-related muscle atrophy. Vitamin E is a well-known antioxidant, which potentially ameliorates a wide range of age-related manifestations. The aim of this study was to determine the effects of tocotrienol-rich fraction (TRF) in modulating the expression of proliferation- and differentiation-associated proteins in senescent human myoblasts during the differentiation phase.Material and methodsHuman skeletal muscle myoblasts were cultured until senescence. Young and senescent cells were treated with TRF for 24 h before and after differentiation induction, followed by evaluation of cellular morphology and efficiency of differentiation. Expression of cell proliferation marker Ki67 protein and myogenic regulatory factors MyoD and myogenin were determined.ResultsOur findings showed that treatment with TRF significantly improved the morphology of senescent myoblasts. Promotion of differentiation was observed in young and senescent myoblasts with TRF treatment as shown by the increased fusion index and larger size of myotubes. Increased Ki67 and myogenin expression with TRF treatment was also observed in senescent myoblasts, suggesting amelioration of the myogenic program by TRF during replicative senescence.ConclusionsTRF modulates the expression of regulatory factors related to proliferation and differentiation in senescent human myoblasts and could be beneficial for ameliorating the regenerative defects during aging.     

Neurosteroids promote phosphorylation and membrane insertion of extrasynaptic GABAA receptors

Neurosteroids are synthesized within the brain and act as endogenous anxiolytic, anticonvulsant, hypnotic, and sedative agents, actions that are principally mediated via their ability to potentiate phasic and tonic inhibitory neurotransmission mediated by γ-aminobutyric acid type A receptors (GABA_ARs). Although neurosteroids are accepted allosteric modulators of GABA_ARs, here we reveal they exert sustained effects on GABAergic inhibition by selectively enhancing the trafficking of GABA_ARs that mediate tonic inhibition. We demonstrate that neurosteroids potentiate the protein kinase C-dependent phosphorylation of S443 within α4 subunits, a component of GABA_AR subtypes that mediate tonic inhibition in many brain regions. This process enhances insertion of α4 subunit-containing GABA_AR subtypes into the membrane, resulting in a selective and sustained elevation in the efficacy of tonic inhibition. Therefore, the ability of neurosteroids to modulate the phosphorylation and membrane insertion of α4 subunit-containing GABA_ARs may underlie the profound effects these endogenous signaling molecules have on neuronal excitability and behavior.Neurosteroids are synthesized de novo in the brain from cholesterol, or steroid hormone precursors. Raising neurosteroid levels in the CNS causes anxiolysis, sedation/hypnosis, anticonvulsant action, and anesthesia and reduces depressive-like behaviors (1–3). Accordingly, dysregulation of neurosteroid signaling is associated with premenstrual dysphoric disorder, panic disorder, depression, schizophrenia, and bipolar disorder. Neurosteroids exert the majority of their actions via potentiating the activity of γ-aminobutyric acid receptors (GABA_ARs), which mediate the majority of fast synaptic inhibition in the adult brain. Accordingly, at low nanomolar concentrations they potentiate GABA-dependent currents, whereas at micromolar concentrations they directly activate GABA_ARs (4–8).GABA_ARs are Cl⁻-preferring pentameric ligand-gated ion channels that assemble from eight families of subunits: α(1–6), β(1–3), γ(1–3), δ, ε, ө, π, and ρ(1–3) (9, 10). Receptor subtypes composed of α1–3βγ subunits largely mediate synaptic or phasic inhibition, whereas those constructed from α4–6β1–3, with or without γ/δ subunits, are principal determinants of tonic inhibition (11–13). Neurosteroids have been shown to bind GABA_ARs at an allosteric site distinct from that of GABA, benzodiazepines, or barbiturates (9, 14). Hosie et al. identified residues located within the transmembrane domain of GABA_AR α and β subunits that are critical for the direct activation (α1–6; Threonine 236, β1–3; Tyrosine 284) and allosteric potentiation (α1–6 Asparagine 407, and α1–6 Glutamine 246) of neurosteroids (15–17). Accordingly, mutation of glutamine 241 (Q241) within the α1–6 subunits prevents allosteric potentiation of GABA_AR composed of αβγ and αβδ subunits by neurosteroids (15, 16).In addition to modulating channel gating, neurosteroids exert potent effects on the expression levels of GABA_ARs (1, 18–20). Moreover, in the hippocampus, prolonged exposure to physiological concentrations of neurosteroids has been shown to enhance the tonic conductance mediated by extrasynaptic GABA_ARs containing the α4/δ subunits, while having little effect on the phasic conductance mediated by synaptic GABA_ARs (6, 21). However, the molecular mechanisms by which neurosteroids regulate GABA_AR expression levels remain unknown.Here, we reveal that neurosteroids act to increase the PKC-dependent phosphorylation of serine 443 (S443) within the intracellular domain of the α4 subunit. This process leads to increased insertion of α4 subunit-containing GABA_ARs into the plasma membrane and a selective enhancement of tonic inhibition. Thus, our experiments reveal a previously unidentified molecular mechanism by which neurosteroids exert sustained effects on GABAergic inhibition by selectively increasing α4-containing GABA_ARs in the membrane and therefore potentiate tonic inhibition.     

Occult colon cancer with initial cutaneous metastatic manifestation: PET/CT detection

Skin metastasis as initial clinical presentation of occult internal organ malignancy is of rare occurrence, predominantly encountered in breast and lung cancer. The search for the occult causative primary malignancy is usually difficult with conventional anatomy-based cross-sectional imaging. The authors present a case of cecal cancer with thoracic and abdominal cutaneous metastasis demonstrated by PET/CT.     

¹⁸F-FDG PET/CT imaging of the pancreas: spectrum of diseases

Since the introduction of integrated positron emission tomography-computed tomography (PET/CT), it has a great impact on the field of oncology. Comparing to other conventional scanners, only PET/CT is capable of providing important information on accurate detecting, staging/restaging, and post-therapeutic monitoring of many cancers. Many studies have demonstrated that PET/CT changes the management in approximately 30% of all cancer patients. Because 2-((18)F)-fluoro-2-deoxy-D-glucose (FDG) is a nonspecific tracer, understanding the PET/CT limitations and pitfalls for various pancreatic conditions can lead to more accurate interpretation of PET/CT images, which ultimately would impact patient care. As a result, it is important for radiologists and other clinicians to familiarize themselves with a wide spectrum of pancreatic PET/CT findings simulating cancer from benign entities.