Therapy directed against oncogenic FLT3 has been shown to induce response in patients with acute myeloid leukemia (AML), but these responses are almost always transient. To address the mechanism of ...FLT3 inhibitor resistance, we generated two resistant AML cell lines by sustained treatment with the FLT3 inhibitor sorafenib. Parental cell lines carry the FLT3-ITD (tandem duplication) mutation and are highly responsive to FLT3 inhibitors, whereas resistant cell lines display resistance to multiple FLT3 inhibitors. Sanger sequencing and protein mass-spectrometry did not identify any acquired mutations in FLT3 in the resistant cells. Moreover, sorafenib treatment effectively blocked FLT3 activation in resistant cells, whereas it was unable to block colony formation or cell survival, suggesting that the resistant cells are no longer FLT3 dependent. Gene expression analysis of sensitive and resistant cell lines, as well as of blasts from patients with sorafenib-resistant AML, suggested an enrichment of the PI3K/mTOR pathway in the resistant phenotype, which was further supported by next-generation sequencing and phospho-specific-antibody array analysis. Furthermore, a selective PI3K/mTOR inhibitor, gedatolisib, efficiently blocked proliferation, colony and tumor formation, and induced apoptosis in resistant cell lines. Gedatolisib significantly extended survival of mice in a sorafenib-resistant AML patient-derived xenograft model. Taken together, our data suggest that aberrant activation of the PI3K/mTOR pathway in FLT3-ITD-dependent AML results in resistance to drugs targeting FLT3.
Previous studies suggest physical activity improves cognition and lowers Alzheimer's disease (AD) risk. However, key AD pathogenic factors that are thought to be influenced by physical activity, ...particularly plasma amyloid-β (Aβ) and Aβ brain load, have yet to be thoroughly investigated. The objective of this study was to determine if plasma Aβ and amyloid brain deposition are associated with physical activity levels, and whether these associations differed between carriers and non-carriers of the apolipoprotein E (APOE) ε4 allele. Five-hundred and forty six cognitively intact participants (aged 60-95 years) from the Australian Imaging, Biomarkers and Lifestyle Study of Ageing (AIBL) were included in these analyses. Habitual physical activity levels were measured using the International Physical Activity Questionnaire (IPAQ). Serum insulin, glucose, cholesterol and plasma Aβ levels were measured in fasting blood samples. A subgroup (n=116) underwent (11)C-Pittsburgh compound B (PiB) positron emission tomography (PET) scanning to quantify brain amyloid load. Higher levels of physical activity were associated with higher high density lipoprotein (HDL) (P=0.037), and lower insulin (P<0.001), triglycerides (P=0.019) and Aβ1-42/1-40 ratio (P=0.001). After stratification of the cohort based on APOE ε4 allele carriage, it was evident that only non-carriers received the benefit of reduced plasma Aβ from physical activity. Conversely, lower levels of PiB SUVR (standardised uptake value ratio) were observed in higher exercising APOE ε4 carriers. Lower plasma Aβ1-42/1-40 and brain amyloid was observed in those reporting higher levels of physical activity, consistent with the hypothesis that physical activity may be involved in the modulation of pathogenic changes associated with AD.
To determine whether IL-6 increases lipolysis and fat oxidation in patients with type 2 diabetes and/or whether it exerts this effect independently of changes to the hormonal milieu, patients with ...type 2 diabetes (D) and healthy control subjects (CON) underwent recombinant human (rh)IL-6 infusion for 3 h. Rates of appearance (Ra) and disappearance (Rd) of U-(13C)palmitate and 6,6-(2H2)glucose were determined. rhIL-6 infusion increased (P < 0.05) palmitate Ra and Rd in a similar fashion in both groups. Neither plasma glucose concentration nor glucose Ra/Rd was affected by rhIL-6 infusion in either group, whereas rhIL-6 infusion resulted in a reduction (P < 0.05) in circulating insulin in D. Plasma growth hormone (GH) was increased (P < 0.05) by IL-6 in CON, and cortisol increased (P < 0.05) in response to IL-6 in both groups. To determine whether IL-6 was exerting its effect directly or through activation of these hormones, we performed cell culture experiments. Fully differentiated 3T3-L1 adipocytes were treated with PBS (control) IL-6, or IL-6 plus dexamethasone and GH. IL-6 treatment alone increased (P < 0.05) lipolysis, but this effect was reduced by the addition of dexamethasone and GH such that IL-6 plus dexamethasone and GH had blunted (P < 0.05) lipolysis compared with IL-6 alone. To assess whether IL-6 increases fat oxidation, L6 myotubes were treated with PBS (Control), IL-6, or AICAR, a compound known to increase lipid oxidation. Both IL-6 and AICAR markedly increased (P < 0.05) oxidation of (14)Cpalmitate compared with Control. Acute IL-6 treatment increased fatty acid turnover in D patients as well as healthy CON subjects. Moreover, IL-6 appears to be activating lipolysis independently of elevations in GH and/or cortisol and appears to be a potent catalyst for fat oxidation in muscle cells.
The Mediterranean diet (MeDi), due to its correlation with a low morbidity and mortality for many chronic diseases, has been widely recognised as a healthy eating model. We aimed to investigate, in a ...cross-sectional study, the association between adherence to a MeDi and risk for Alzheimer's disease (AD) and mild cognitive impairment (MCI) in a large, elderly, Australian cohort. Subjects in the Australian Imaging, Biomarkers and Lifestyle Study of Ageing cohort (723 healthy controls (HC), 98 MCI and 149 AD participants) completed the Cancer Council of Victoria Food Frequency Questionnaire. Adherence to the MeDi (0- to 9-point scale with higher scores indicating higher adherence) was the main predictor of AD and MCI status in multinominal logistic regression models that were adjusted for cohort age, sex, country of birth, education, apolipoprotein E genotype, total caloric intake, current smoking status, body mass index, history of diabetes, hypertension, angina, heart attack and stroke. There was a significant difference in adherence to the MeDi between HC and AD subjects (P < 0.001), and in adherence between HC and MCI subjects (P < 0.05). MeDi is associated with change in Mini-Mental State Examination score over an 18-month time period (P < 0.05) in HCs. We conclude that in this Australian cohort, AD and MCI participants had a lower adherence to the MeDi than HC participants.
5-Aminoimidazole-4-carboxamide-ribonucleoside (AICAR) and caffeine, which activate AMP-activated protein kinase (AMPK) and cause sarcoplasmic reticulum calcium release, respectively, have been shown ...to increase mitochondrial biogenesis in L6 myotubes. Nitric oxide (NO) donors also increase mitochondrial biogenesis. Since neuronal and endothelial NO synthase (NOS) are calcium dependent and are also phosphorylated by AMPK, we hypothesized that NOS inhibition would attenuate the activation of mitochondrial biogenesis in response to AICAR and caffeine. L6 myotubes either were not treated (control) or were exposed acutely or for 5 h/day over 5 days to 100 microM of N(G)-nitro-L-arginine methyl ester (L-NAME, NOS inhibitor), 100 microM S-nitroso-N-acetyl-penicillamine (SNAP) (NO donor) +/- 100 microM L-NAME, 2 mM AICAR +/- 100 microM L-NAME, or 5 mM caffeine +/- 100 microM L-NAME (n = 12/treatment). Acute AICAR administration increased (P < 0.05) phospho- (P-)AMPK, but also increased P-CaMK, with resultant chronic increases in peroxisome proliferator-activated receptor-gamma coactivator-1 alpha (PGC-1 alpha), cytochrome-c oxidase (COX)-1, and COX-4 protein expression compared with control cells. NOS inhibition, which had no effect on AICAR-stimulated P-AMPK, surprisingly increased P-CaMK and attenuated the AICAR-induced increases in COX-1 and COX-4 protein. Caffeine administration, which increased P-CaMK without affecting P-AMPK, increased COX-1, COX-4, PGC-1 alpha, and citrate synthase activity. NOS inhibition, surprisingly, greatly attenuated the effect of caffeine on P-CaMK and attenuated the increases in COX-1 and COX-4 protein. SNAP increased all markers of mitochondrial biogenesis, and it also increased P-AMPK and P-CaMK. In conclusion, AICAR and caffeine increase mitochondrial biogenesis in L6 myotubes, at least in part, via interactions with NOS.
There is no consensus for a blood-based test for the early diagnosis of Alzheimer's disease (AD). Expression profiling of small non-coding RNA's, microRNA (miRNA), has revealed diagnostic potential ...in human diseases. Circulating miRNA are found in small vesicles known as exosomes within biological fluids such as human serum. The aim of this work was to determine a set of differential exosomal miRNA biomarkers between healthy and AD patients, which may aid in diagnosis. Using next-generation deep sequencing, we profiled exosomal miRNA from serum (N=49) collected from the Australian Imaging, Biomarkers and Lifestyle Flagship Study (AIBL). Sequencing results were validated using quantitative reverse transcription PCR (qRT-PCR; N=60), with predictions performed using the Random Forest method. Additional risk factors collected during the 4.5-year AIBL Study including clinical, medical and cognitive assessments, and amyloid neuroimaging with positron emission tomography were assessed. An AD-specific 16-miRNA signature was selected and adding established risk factors including age, sex and apolipoprotein ɛ4 (APOE ɛ4) allele status to the panel of deregulated miRNA resulted in a sensitivity and specificity of 87% and 77%, respectively, for predicting AD. Furthermore, amyloid neuroimaging information for those healthy control subjects incorrectly classified with AD-suggested progression in these participants towards AD. These data suggest that an exosomal miRNA signature may have potential to be developed as a suitable peripheral screening tool for AD.
Urinary incontinence (UI) is highly prevalent in nursing and residential care homes (CHs) and profoundly impacts on residents' dignity and quality of life. CHs predominantly use absorbent pads to ...contain UI rather than actively treat the condition. Transcutaneous posterior tibial nerve stimulation (TPTNS) is a non-invasive, safe and low-cost intervention with demonstrated effectiveness for reducing UI in adults. However, the effectiveness of TPTNS to treat UI in older adults living in CHs is not known. The ELECTRIC trial aims to establish if a programme of TPTNS is a clinically effective treatment for UI in CH residents and investigate the associated costs and consequences.
This is a pragmatic, multicentre, placebo-controlled, randomised parallel-group trial comparing the effectiveness of TPTNS (target n = 250) with sham stimulation (target n = 250) in reducing volume of UI in CH residents. CH residents (men and women) with self- or staff-reported UI of more than once per week are eligible to take part, including those with cognitive impairment. Outcomes will be measured at 6, 12 and 18 weeks post randomisation using the following measures: 24-h Pad Weight Tests, post void residual urine (bladder scans), Patient Perception of Bladder Condition, Minnesota Toileting Skills Questionnaire and Dementia Quality of Life. Economic evaluation based on a bespoke Resource Use Questionnaire will assess the costs of providing a programme of TPTNS. A concurrent process evaluation will investigate fidelity to the intervention and influencing factors, and qualitative interviews will explore the experiences of TPTNS from the perspective of CH residents, family members, CH staff and managers.
TPTNS is a non-invasive intervention that has demonstrated effectiveness in reducing UI in adults. The ELECTRIC trial will involve CH staff delivering TPTNS to residents and establish whether TPTNS is more effective than sham stimulation for reducing the volume of UI in CH residents. Should TPTNS be shown to be an effective and acceptable treatment for UI in older adults in CHs, it will provide a safe, low-cost and dignified alternative to the current standard approach of containment and medication.
ClinicalTrials.gov, NCT03248362. Registered on 14 August 2017. ISRCTN, ISRCTN98415244. Registered on 25 April 2018. https://www.isrctn.com/.
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