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C2D2 research 4a - Did Rheumatoid Arthritis Really Begin in 1800?

Project: Other projectOther internal award

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Description

Rheumatoid arthritis (RA) is an enigma for medicine and medical historians. It is claimed that RA is a new disease that first appeared in Europe in the 19thC. Why did it appear so late in Europe compared to the Americas? Does its late appearance give clues to its underlying cause?

The first series of patients in Europe displaying the classic debilitating joint inflammation associated with this autoimmune disease were recorded in 1800. In many individuals, the immune response is targeted toward citrullinated proteins - proteins modified by an enzyme produced by our own bodies, as well as by the oral bacteria Porphyromonas gingivalis. It has been suggested that increasing sugar consumption in the 19thC led to changes in oral bacteria, like P. gingivalis, triggering an immune response to citrullinated proteins in greater frequency. Interestingly, tobacco smoking is a significant non-genetic risk factor for RA, a custom which also increases in prevalence in the 18-19thC.

This research will apply macroscopic and biomolecular techniques to archaeological skeletons to investigate RA in historic times, through: (i) radiography, to diagnose RA; (ii) proteomics, to identify citrullinated proteins in RA skeletons; and (iii) biochemical analyses, to investigate dietary/lifestyle information (sugar consumption, tobacco use).

Layman's description

Rheumatoid arthritis (RA) is an enigma for medicine and medical historians. It is claimed that RA is a new disease that first appeared in Europe in the 19thC. Why did it appear so late in Europe compared to the Americas? Does its late appearance give clues to its underlying cause?

The first series of patients in Europe displaying the classic debilitating joint inflammation associated with this autoimmune disease were recorded in 1800. In many individuals, the immune response is targeted toward citrullinated proteins - proteins modified by an enzyme produced by our own bodies, as well as by the oral bacteria Porphyromonas gingivalis. It has been suggested that increasing sugar consumption in the 19thC led to changes in oral bacteria, like P. gingivalis, triggering an immune response to citrullinated proteins in greater frequency. Interestingly, tobacco smoking is a significant non-genetic risk factor for RA, a custom which also increases in prevalence in the 18-19thC.
This research will apply macroscopic and biomolecular techniques to archaeological skeletons to investigate RA in historic times, through: (i) radiography, to diagnose RA; (ii) proteomics, to identify citrullinated proteins in RA skeletons; and (iii) biochemical analyses, to investigate dietary/lifestyle information (sugar consumption, tobacco use).

Key findings

Proteomic analysis
Quantitative proteomics has been successfully applied to RA-affected synovial fluids and plasma in order to identify joint-specific disease biomarkers and explore RA pathogenesis (Mateos et al. 2012). Given the successful extraction of proteomic profiles from archaeological bone (e.g. Cappellini et al.2012; Buckley & Wadsworth 2014) and other tissues (e.g. Warinner et al. 2014a; Bona et al. 2014; Hendy et al. 2016), we selected samples of both Old and New World origin (UK and Mexico) and used quantitative proteomics (iTRAQ) to investigate whether protein indicators of RA may preserve in archaeological bone. We explored whether quantitative differences can be observed in affected and non-affected bones with regards to i) proteins associated with inflammation in the synovial fluid and ii) the presence of citrullinated peptides.

Applying quantitative shotgun proteomics to 8 samples of non-RA and putative RA-affected bones, we identified proteins derived from bone, cartilage and synovial fluid. The majority of detected proteins have roles in forming, or developing, the extracellular matrix, forming the structural constituents of the skeletal tissue. Many (e.g. BGLAP, SPP1, FMOD) associate with collagen fibrils or with hydroxyapatite. In addition, several proteins detected are expressed in cartilage, such as ACAN, CILP, CHAD, and COL1A9. We also detected a number of proteins which have been previously detected in synovial fluid, with several reported to be expressed in RA-affected synovial fluid, including BGLAP (Gevers et al. 1986), IBSP (Saxne et al. 1995), IGFBP1 (Matsumoto et al. 1996), GSN, AHSG (Biswas et al. 2013), VTN (Rosenblum & Carsons 1996), and THBS1 (Gotis-Graham et al. 1997). No key proteins involved in RA associated bone erosion (such as TNFα, IL-1, IL-6 and RANK-L) were detected. Whilst we did observe differences in the proteomic profiles of bone elements with putative RA indicators and those absent of RA erosive lesions, these differences may be the result of the different physiologies of the bone elements analysed, and not the result of RA.

We also attempted to identify elevated levels of citrullinated peptides in RA-affected individuals compared with non-affected individuals. We identified seven peptides with citrullination modifications, but there appears to be no differences between putative RA and RA negative bone elements. In RA affected individuals citrullination typically occurs in keratin, collagen, vimentin and actin (Olivares- Martínez et al. 2011). Although collagen was identified in our study, there were no confidently identified peptides with citrullination modifications from these proteins.

Py-GC/MS analysis
The possibility of a novel marker for tabacco smoking (and tea/coffee consumption) was investigated though the use of Pyrolysis Gas Chromatography Mass Spectrometry (Py-GC/MS). Samples of clay pipes were also tested fro the presence of nicotine. Unfortunately only one unusually well preserved sample gave a tentative identification of nicotine. This initial finding indicates the possible potential of the technique but further research using SIMS to target specific analytes is needed to investigate the use of this technique to identify smoking and tea/coffee consumption in the future.

Stable isotope analysis
A total of 43 adult individuals from five post-medieval sites in London, Yorkshire and Manchester were analysed for bulk δ13C and δ15N stable isotopes in bone collagen to assess the protein sources in the diet, following conventional methods. Results indicated that almost all individuals exhibited enrichment in 15N as seen in published values for post-medieval urban populations. Further work is needed to characterise the isotopic baseline for urban centres to explore these enriched data (BSc dissertation student supervised by Alexander is exploring this in 2017/18). Only a few individuals exhibited high δ13C values indicative of potential input of C4 foods, again indicative of published values form elsewhere in the U.K.

To assess the potential for identifying the consumption of carbohydrates and in particular, sugar cane (a C4 crop), we analysed the δ13C in the carbonate fraction of bone mineral of the 43 individuals, first applying ATR-FTIR to bone samples to assess bone mineral preservation. Initial analysis was inconclusive for assessing the consumption of C4 resources, however analysis is ongoing with a WRoCAH (AHRC) funded PhD student (Chidimuro, started Jan 2017 supervised by Alexander) taking this portion of the research forward.

In collaboration with Kate Britton (Archaeology, Aberdeen University) we investigated the consumption of hot (tea, coffee) and distilled (gin) drinks that popular during the period though analysis of δ18O in bone and tooth mineral phosphate (15 samples) and carbonate (33 samples). Initial results from this research were presented at the European Archaeologists Association conference 2016 and are being prepared for publications to be submitted to the Journal of Archaeological Science and The Journal of Archaeological Science Reports in later 2017/early 2018.

StatusFinished
Effective start/end date7/01/151/07/18

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