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The EsmiRs follow-up study repeats the measurements and bio-sampling from the ERMA study and adds the following new measurements.

Fat Oxidation Capacity and Resting Energy Expenditure

Fat oxidation at rest and during exercise and resting energy expenditure are measured with indirect calorimetry. Indirect calorimetry relies on the assumption that fatty acid and carbohydrate oxidation lead to differences in cellular oxygen use and carbohydrate production. With breath-by-breath analysis of the breathing gases, substrate use and energy expenditure can be estimated noninvasively.

Peak Oxygen Consumption

A well-known proxy of cardiorespiratory fitness, peak oxygen consumption primarily results from cardiac output and arteriovenous oxygen difference. In the study, peak oxygen uptake is measured during a maximal graded incremental bicycle ergometer test.

Insulin Sensitivity

Insulin sensitivity is measured with a two hour oral glucose tolerance test during which blood samples are collected in every 30 minutes. Glucose and insulin concentrations are determined from the blood samples and the Matsuda index is calculated to quantify insulin sensitivity. Also, serum metabolites are determined from the samples with NMR spectroscopy to investigate changes in the serum metabolite profile following the glucose ingestion.

Arterial Stiffness

Each heart contraction causes a pulse wave that spreads from the heart to the periphery via arteries. Stiffening arteries increase the pulse wave velocity and change its form. These values can be measured noninvasively with a cuff technique similar to a blood pressure measurement. Results obtained with the noninvasive technique correlate strongly with the direct invasive technique. The study measures arterial stiffness at rest and following an exercise bout. 

Dietary Intake and Food Habits

Dietary intake of the participants will also be measured. The participants will record their food intake over three to four day periods before the physiological measurements. Food diaries will be analyzed for energy, macro- and micronutrient intakes. Dietary habits of the participants will also be screened by a questionnaire.

Biochemical Analyses from Blood and Tissue Samples

Blood samples collected over the course of the EsmiRs project are used to study the concentration of leukocytes, total cholesterol, different steroid hormones (e.g. estradiol and testosterone), and various cell-to-cell interaction-related molecular markers. Muscle samples taken as needle biopsies are used to investigate estrogen-associated muscle cell signaling cascades.

MicroRNAs

MicroRNAs, or miRNAs, are short RNA-molecules that participate in the epigenetic regulation of gene reading by binding with their target mRNA-molecules and transiently or permanently blocking corresponding protein translation. miRNAs can act as autocrine regulators inside the cells they have produced or they can be released into the bloodstream to act as endocrine messengers. Released miRNAs can serve as inter-tissue messengers whose concentration and quality can be measured in relation to certain diseases or conditions, including cancer and metabolic dysfunctions.

Next generation sequencing (NGS)

The EsmiRs project uses modern sequencing technology as a way to interpret complex miRNA messaging. Next generation sequencing produces accurate information and the exact sequence of nucleic acid based molecules in a fast and reliable manner. NGS is performed in collaboration with Central Finland Central Hospital’s laboratory of molecular pathology.

Cell Cultures/In Vitro Modelling

Primary myocytes extracted from biopsied tissues are used to study the effects of external estradiol supplements to the cells’ own hormone production in a controlled and adjustable environment.

Histology and Microscopy

Thin cross-sectional sections from the muscle biopsies are used for histology, which aims to find and visualize muscle stem cells’, also known as satellite cells, intracellular lipid accumulation as well as to measure cell size changes within the longitudinal study arm of the ERMA study. The targets are made visible with antibodies and fluorescent labels, which can then be imaged with confocal systems. Images are analyzed with computational software.

Protein Measurements

The biological sample pool contains a large amount of interesting proteins which can be separated and analyzed based on their size or quality. Electrophoresis and antibody-based Western Blots are routinely used to see the changes in the quantity and quality of proteins throughout different hormonal levels.

Quantitative PCR

Polymerase chain reaction (PCR) is used to study the amount of key RNA-molecules produced in the sample. The method can provide information from the reading activity of a gene or about what kind of RNA-transcripts are present in the tissue at a given point. Our research interest centres various miRNAs and, for example, apoptosis related gene transcripts .

ERMA study

The EsmiRs-study is a follow-up study largely based on previous ERMA study. The study has been conducted at the Gerontology Research Center of the Ä¢¹½Ö±²¥. EsmiRs study will utilize samples from ERMA study for biochemical analysis. 

University of Minnesota (USA)

EsmiRs study is done in collaboration with professor Dawn Lowe's research group, that is located at the University of Minnesota, Minneapolis, USA. Docent Eija Laakkonen, the PI of EsmiRs study, has visited Lowe's laboratory several times during the collaboration, and postdoctoral researcher Sira Karvinen spent a year in Lowe's laboratory for postdoctoral training (2016-2017). The collaboration enables EsmiRs project to utilize special skeletal muscle stimulation system ans samples from in vivo models available in Lowe's laboratory.

Central Finland Health Care District

The EsmiRs study utilizes for example the  Next Generation Sequencing (NGS) equipment available at the Central Finland Health Care District’s molecular pathology laboratory.