Tau is a microtubule (MT)-stabilizing protein that is altered in Alzheimer's disease (AD) and other tauopathies. It is hypothesized that the hyperphosphorylated, conformationally altered, and multimeric forms of tau lead to a disruption of MT stability; however, direct evidence is lacking in vivo. In this study, an in vivo stable isotope-mass spectrometric technique was used to measure the turnover, or dynamicity, of MTs in brains of living animals. We demonstrated an age-dependent increase in MT dynamics in two different tau transgenic mouse models, 3xTg and rTg4510. MT hyperdynamicity was dependent on tau expression, since a reduction of transgene expression with doxycycline reversed the MT changes. Treatment of rTg4510 mice with the epothilone, BMS-241027, also restored MT dynamics to baseline levels. In addition, MT stabilization with BMS-241027 had beneficial effects on Morris water maze deficits, tau pathology, and neurodegeneration. Interestingly, pathological and functional benefits of BMS-241027 were observed at doses that only partially reversed MT hyperdynamicity. Together, these data suggest that tau-mediated loss of MT stability may contribute to disease progression and that very low doses of BMS-241027 may be useful in the treatment of AD and other tauopathies.

BackgroundMuscle mass can be measured directly in vivo by isotope dilution, using Creatine‐(methyl‐d3) monohydrate (D3‐Cr) by mouth followed by measurement of the steady‐state enrichment of D3‐creatinine (D3‐Crn) in urine. Isotope dilution methods require knowledge of the amount of tracer delivered to the pool of interest. In a subset of human subjects, a small amount of orally administered D3‐Cr ‘spills’ into urine after absorption and prior to transport into skeletal muscle cells. The objectives were to develop a method to correct for spillage to compare the estimate of muscle mass by D3‐Cr dilution to other assessments of fat‐free mass.MethodsSubjects (19 males, 23–81 years old; 20 females, 20–77 years old) ingested a single dose of 60 mg D3‐Cr and urine was collected prior to and daily for 4 days following the dose. Fasting morning urine samples was assessed for D3‐Cr, total Cr, D3‐Crn, and total Crn concentrations, as well as isotopic enrichments of D3‐Crn, by LC/MS. The 24‐h urine collections over 3 days after the dose of D3‐Cr were also performed to determine D3‐Cr spillage. Total body water, fat mass, and fat‐free mass were assessed by bioelectrical impedance spectroscopy (BIS).ResultsSpillage of D3‐Cr in the urine was greater in women than men. D3‐Crn enrichment and the ratio of Cr/Crn were used in an algorithm to calculate Cr pool size and muscle mass. Specifically, an algorithm was developed for the estimation of spillage based on the relationship between the fasting Cr/Crn ratio and the cumulative proportion of the D3‐Cr dose excreted over 3 days based on 24‐h urine collections. Muscle mass corrected using the algorithm based on fasting urine levels correlated (r = 0.9967, P < 0.0001) with that corrected by measuring D3‐Cr dose excreted. Muscle mass measured by D3‐Crn enrichment also correlated (r = 0.8579, P < 0.0001, algorithm corrected) with that measured by 24‐h Crn excretion. Muscle mass measured by D3‐Cr dilution method correlated with intracellular water by BIS, whether using spillage corrected by the algorithm (r = 0.9041, P < 0.0001) or measured by 3 day D3‐Cr losses (r = 0.91, P < 0.0001) and similarly correlated with fat‐free mass by BIA (r = 0.8857 and 0.8929, P < 0.0001, respectively).ConclusionsThe D3‐Cr dilution method is further validated here as a non‐invasive, easy‐to‐use test for measuring muscle mass. The technical issue of D3‐Cr spillage can be corrected for with a simple algorithm based on fasting spot urine samples. Muscle mass by Cr dilution potentially has broad applications in clinical and research settings.

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Bile acid sequestrants (BAS) such as colesevelam are effective therapies for lowering LDL-cholesterol and improving glycemic control in individuals with type 2 diabetes. It is however not clear if changes in glucose metabolism with BAS treatment are regulated via changes in whole body BA metabolism. We performed a post-hoc analysis of circulating bile acid levels to investigate whether changes in glucose and lipid parameters are associated with alterations in plasma bile acids (BA) concentrations with colesevelam treatment. Subjects with type 2 diabetes (n=49) treated with diet and exercise, sulfonylurea, metformin or a combination thereof, were treated with 3.75 g/day colesevelam for 12 weeks. Fasting bile acid concentrations were measured by LC/MS and glucose and lipid kinetics were measured using stable isotope techniques at baseline and post-treatment. Colesevelam treatment reduced fasting LDL-cholesterol and increased HDL-cholesterol and triglyceride concentrations. No changes were seen in fasting total cholesterol concentrations and de novo lipogenesis (DNL). Beta-cell function (HOMA-B), glycolytic disposal of oral glucose and fasting plasma glucose clearance improved significantly. Colesevelam treatment resulted in a smaller (-40% from baseline) and more hydrophilic fasting plasma BA pool. All BA’s measured except for cholic acid, glycocholic acid and taurocholic acid decreased with treatment. Changes in total BA concentrations were inversely correlated with changes in A1C and a positive correlation was seen with fasting insulin concentrations. Changes in BA concentrations did not correlated with changes in lipid parameters. These data suggest that altered bile acids levels may contribute to improved glycemic control with colesevelam treatment in subjects with type 2 diabetes.

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