Project description:Metformin, a commonly used drug prescribed to treat type-2 diabetes, has been found to extend health span and delay cancer incidence and progression. Here, we report that starting chronic treatment with low dose of metformin (0.1% w/w in diet) at one year of age extends health and lifespan in male mice, while a higher dose (1% w/w) was toxic. Treatment with low dose metformin mimicked some of the benefits of calorie restriction, such as improved physical performance, increased insulin sensitivity, and reduced LDL and cholesterol levels. At a molecular level, metformin increased AMP-activated protein kinase activity without attenuation of the mitochondrial electron transport chain activities. Metformin treatment resulted in lower chronic inflammation and increased antioxidant protection, suggesting that the ability of metformin to improve health of laboratory animals may stem from these factors. Our results support that metformin supplementation could be beneficial in extending health and lifespan in humans. Groups of one-year old male B6C57/J mice were maintained on ad libitum AIN-93G SD diet, or a 40% caloric restriction of the AIN-93G SD diet, or an ad libitum AIN-93G diet supplemented with 0.1% w/w metformin for the rest of their lives. 5 mice from each of these 3 groups were selected and RNA was extracted from both muscle and liver tissue using 1.0mm glass beads in a Precellys 24 Tissue Homogenizer and Qiagen RNeasy Mini Kits for Fibrous Tissue according to manufacturer's specifications. Quality and quantity of the total RNA was checked with the Agilent 2100 bioanalyzer using RNA 6000 Nano chips. RNA samples were labeled using the Illumina TotalPrep RNA Amplification Kit. In short, 0.5g of total RNA was first converted into single-stranded cDNA with reverse transcriptase using an oligo-dT primer containing the T7 RNA polymerase promoter site and then copied to produce double-stranded cDNA molecules. The double stranded cDNA was cleaned and concentrated with the supplied columns and used in an overnight in-vitro transcription reaction where single-stranded RNA (cRNA) was generated and labeled by incorporation of biotin-16-UTP. Arrays were hybridized using a total of 0.75ug of biotin-labeled cRNA at 58 degrees C for 16 hours to Illumina's Sentrix MouseRef-8 v2 Expression BeadChips. Each BeadChip has ~24,000 well-annotated RefSeq transcripts with approximately 30-fold redundancy. The arrays were washed, blocked and the biotin labeled cRNA was detected by staining with streptavidin-Cy3. Arrays were scanned at a resolution of 0.8um using the Beadstation 500 X from Illumina and the data was extracted using the Illumina GenomeStudio software(v1.6.0). Any spots at or below the background were filtered out using an Illumina detection p value of 0.02 and above. The natural log of all remaining scores were used to find the avg and std of each array and the z-score normalization was calculated and presented below. Z-score = (raw value - avg)/std.

Project description:This model is supplementary material of publication "Physiologically based metformin pharmacokinetics model of mice and scale-up to humans for the estimation of concentrations in various tissues" by Darta Maija Zake, Linda Zaharenko, Janis Kurlovics, Vitalijs Komasilovs, Janis Klovins and Egils Stalidzans. The model is pre-set for simulation of a single peroral dose. This is a whole-body model representing the pharmacokinetics of metformin in the mouse body. The model is in the form of ordinary differential equations and describes metformin concentration in 20 compartments. The model consists of 20 compartments (“Compartments” in COPASI model) describing various tissues or tissue sub-compartments and body fluids of metformin action (venous and arterial plasma, intestine, kidney, heart, fat, muscle, brain, lungs, stomach, liver, portal vein, remainder urine and feces). Body weight and the weight of all compartments is expressed as a volume in mL and for the calculations it is assumed that 1mL = 1g. The volumes of most compartments are calculated as a fraction of the body weight/volume, and the fractions are determined from literature data, the volumes of the stomach lumen and intestine lumen are fixed and do not change depending on the body weight. Similarly, the volume of external urine and feces is set to 1mL, but those are “volumeless” compartments as they are only necessary for the calculation of metformin amount, not concentration. The model consists of 20 species (“Species” in COPASI model) that correspond to the metformin concentrations in the 20 compartments. The initial concentrations for all the species are 0 nmol/mL as metformin is not produced in the body and can only be detected after dose administration. The model consists of 33 reactions – they describe the transport processes of metformin in the body. The reactions include local parameters that are involved only in that particular reaction and global parameters – parameters that are used in multiple reactions or are calculated depending on another parameter e.g. scale-up coefficients. The model consists of 52 global quantities – parameters involved in multiple reactions or necessary for another parameter calculation: 1.Parameters describing metformin dose – either in peroral (Metformin Dose in Lumen in mg) or intravenous (Metformin Dose in Plasma in mg). 2.Parameter describing mice physiology – body weight (in mL), cardiac output, blood flow to different compartments described as Q”compartment_name” (for example Qliver describes blood flow to the liver compartment). Qgfr refers to the glomerular filtration rate. 3.Tissue:plasma partition coefficients (Ktp) that are necessary for the scale-up to humans. 4.Parameters involved in the calculation of metformin amount in mg, these parameters are named mg”Compartment_name” (for example mgLiver describes the metformin amount in mg in the liver tissues). The time points of dose release are defined as “events” in COPASI and can be changed as necessary. Time course simulations can be accessed through the section “Time Course” in this section the time duration and intervals can be changed. When time-course simulations are run three plots are created – Metformin amount in the 20 compartments, metformin concentrations in the compartments and reaction fluxes of all the reactions (see “Output Specifications” -> “Plots” to activate or deactivate plots).

Project description:This model is supplementary material of publication "Physiologically based metformin pharmacokinetics model of mice and scale-up to humans for the estimation of concentrations in various tissues" by Darta Maija Zake, Linda Zaharenko, JanisKurlovics, Vitalijs Komasilovs, Egils Stalidzans and Janis Klovins. This is a whole-body model representing the pharmacokinetics of metformin in the mouse body. The model is in the form of ordinary differential equations and describes metformin concentration in 20 compartments. The model consists of 20 compartments (“Compartments” in COPASI model) describing various tissues or tissue sub-compartments and body fluids of metformin action (venous and arterial plasma, intestine, kidney, heart, fat, muscle, brain, lungs, stomach, liver, portal vein, remainder urine and feces). Body weight and the weight of all compartments is expressed as a volume in mL and for the calculations it is assumed that 1mL = 1g. The volumes of most compartments are calculated as a fraction of the body weight/volume, and the fractions are determined from literature data, the volumes of the stomach lumen and intestine lumen are fixed and do not change depending on the body weight. Similarly, the volume of external urine and feces is set to 1mL, but those are “volumeless” compartments as they are only necessary for the calculation of metformin amount, not concentration. The model consists of 20 species (“Species” in COPASI model) that correspond to the metformin concentrations in the 20 compartments. The initial concentrations for all the species are 0 nmol/mL as metformin is not produced in the body and can only be detected after dose administration. The model consists of 33 reactions – they describe the transport processes of metformin in the body. The reactions include local parameters that are involved only in that particular reaction and global parameters – parameters that are used in multiple reactions or are calculated depending on another parameter e.g. scale-up coefficients. The model consists of 52 global quantities – parameters involved in multiple reactions or necessary for another parameter calculation: 1.Parameters describing metformin dose – either in peroral (Metformin Dose in Lumen in mg) or intravenous (Metformin Dose in Plasma in mg). 2.Parameter describing mice physiology – body weight (in mL), cardiac output, blood flow to different compartments described as Q”compartment_name” (for example Qliver describes blood flow to the liver compartment). Qgfr refers to the glomerular filtration rate. 3.Tissue:plasma partition coefficients (Ktp) that are necessary for the scale-up to humans. 4.Parameters involved in the calculation of metformin amount in mg, these parameters are named mg”Compartment_name” (for example mgLiver describes the metformin amount in mg in the liver tissues). The time points of dose release are defined as “events” in COPASI and can be changed as necessary. Time course simulations can be accessed through the section “Time Course” in this section the time duration and intervals can be changed. When time-course simulations are run three plots are created – Metformin amount in the 20 compartments, metformin concentrations in the compartments and reaction fluxes of all the reactions (see “Output Specifications” -> “Plots” to activate or deactivate plots). Also plotting the species result after 0.5 hours will reproduce the literature results.

			Action	DRS
	SRR29157915_1.fastq.gz	Fastqsanger.gz
	SRR29157915_2.fastq.gz	Fastqsanger.gz
	SRR29157916_1.fastq.gz	Fastqsanger.gz
	SRR29157916_2.fastq.gz	Fastqsanger.gz
	SRR29157917_1.fastq.gz	Fastqsanger.gz

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Metformin mice

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