Analysis of the pathogenesis of Rheumatoid Arthritis by transcriptomic methods

Rheumatoid Arthritis (RA) is a complex autoimmune inflammatory disease that affects between 0.5-1% of the world’s population, appearing more frequently in women between 40 – 60 years old.

Among the symptoms it causes is chronic synovial inflammation, which translates as chronic inflammation of the different joints of the body, with the consequent reduction in the quality of life that this entails for the affected  patients.

The laboratory study of RA

In recent years, many efforts have been put into the study of RA. However, the exact mechanism that produces it is yet to be known.

The laboratory study of RA can be approached from different angles: from a genomic point of view (at DNA level), transcriptomic (at the RNA level) and proteomic (at the protein level). However, genomic information is, by definition, static, and does not necessarily represent all the genes that are transcribed and translated into proteins in an individual and in a given environment.

For this reason, the transcriptomic and proteomic analysis has proven to be more accurate.

Transcriptomic analysis

To analyze and compare samples from healthy patients and patients with RA from a transcriptomic point of view, the first step consists of isolating the RNA from the samples. One of the methods to extract RNA is the one used by Trizol.

Briefly, for this method one has to start with the desired sample volume in a 15 mL tube, and alternate the addition of different reagents (trizol, chloroform, isopropanol, etc.) with the centrifugation of the samples. Generally the samples are centrifuged at 12,000G, 15 min and 4ºC; However, for washing, centrifugation at 7,500G, 5 min and 4ºC would be sufficient.

For these centrifugation steps The centrifuge Digicen 22 R has a wide range of angle fixed rotors, both for low revolutions, with capacity up to 32 tubes of 15 ml./15 ml. conical, and for microtubes, cryotubes and high speed tubes is the best option, its powerful refrigeration system allows it to keep the minimum chamber temperature below 4°C regardless of the type of rotor and the selected speed.

As a result of the centrifugations, two phases are formed in the tube, which are known as “supernatant” (upper phase) and “pellet” (sediment). Thus, after the last centrifugation, the supernatant is discarded and the pellet, which contains the extracted RNA, is stored for subsequent analysis by transcriptomic methods (microarray or RNAseq).

Finally, once the samples have been analyzed, the results are compared with each other, to determine the differences in the expression of certain genes of interest between samples from healthy patients and those obtained from patients diagnosed with RA.

The transcriptomic approach for the study of RA allows comparisons to be made between healthy patients and patients with RA in terms of the expression of certain genes of interest.

For this process, it is necessary to isolate the RNA from the test samples; process that requires different steps of chain centrifugation during which temperature stability must be ensured to avoid deterioration of the genetic material.