Microchimerism and multiple sclerosis: a study on the impact of the sex of offspring on clinical, radiological, and paraclinical features of maternal disease. A new point of view for the sex differences in Neurological disease

Background. Multiple sclerosis (MS) is a chronic autoimmune disorder characterised by inflammation and neurodegeneration and representing one of the most common causes of neurologic disability among young adults. Over the last 40 years, several authors have confirmed the existence of fetal cells in maternal blood and their pregnancy-related origin, demonstrating that pregnancy may establish a long-term, low-grade chimeric state in women. The biological and clinical significance of fetal microchimeric cells (fMCs) in maternal health is largely unknown, although a role in autoimmune diseases have been hypothesised. Aims. The overarching aim of my PhD dissertation was to investigate the role of the sex of offspring, considered an indirect marker of fMCs, in clinical, paraclinical, and radiological MS features. Methods. During my PhD course, I conducted two retrospective studies and one prospective study on female MS patients. proceeding from literature data on the association between pregnancy and microchimerism, subjects were classified according to their pregnancy status and sex of offspring as follow: (a) subjects with history of at least a male pregnancy and supposedly carrying XY microchimeric cells (XYp), (b) subjects with history of only female pregnancy and supposedly carrying XX microchimeric cells, but not XY microchimeric cells (XXp), and (c) nulliparous subjects supposedly without microchimeric cells (NLp). In the first project, I obtained information on pregnancy history for a population of 354 MS female patients, including 87 nulliparous subjects (NLp), 188 subjects with history of at least a male pregnancy (XYp), and 79 subjects with history of only female pregnancy (XXp). Medical records were used to collect clinical, radiological, and paraclinical data at onset, diagnosis, and last clinical follow-up for this large cohort of patients. In the second study I selected a subgroup of 54 patients from the previous cohort, including 26 NLp, 8 XXp, and 20 XYp. I processed their magnetic resonance imaging (MRI) scan using the Lesion Segmentation Tool toolbox and FreeSurfer software to obtain quantitative data on white matter, cortical, and subcortical areas. Additional clinical, radiological, and paraclinical data at onset, diagnosis, and last clinical follow-up were also collected using medical records and telephone interview. Finally, in the third project, I enrolled 43 patients in a prospective study. The study is still on-going and the population recruited so far includes 18 NLp, 19 XYp, and 6 XXp. All patients were classified according to their obstetric history and underwent a blood test analysis to determine the microchimeric group amplifying Y chromosome-specific sequences. Each patients underwent a baseline visit to collect clinical data, an MRI scan, and an optical coherence tomography (OCT) scan. Results. The first study showed that, at disease onset, NLp were younger than XYp and XXp and that the same group reported a longer disease duration when attending the first visit at MS Centre. In addition, data showed that NLp had less frequently a pyramidal onset when compared to XXp. Comparing XYp and XXp patients, I observed that XXp had higher ARR, a higher disability after 3 and 5 years of disease duration, and more severe ambulation scores at EDSS at 3 years of disease duration. In the second project, I observed that NLp had lower brain volumes in several cortical areas, as well as in some subcortical and white matter volumes. More specifically, comparing NLp and XXp, I found that the former group had larger 4th ventricle and smaller right pallidum and left enthorinal volumes. NLp also reported lower thickness in left paracentral cortex, left precuneus cortex, and right lateral occipital cortex when compared with XXp. A similar trend was observed comparing NLp and XYp: NLp group had lower thickness in left paracentral cortex, left pericalcarine cortex, and right paracentral cortex. Interestingly, at the comparison between XYp and XXp, I observed that the thickness was higher in XYp in the left cuneus cortex, left pericalcarine cortex, and left insula, while XXp had a higher thickness in the right lateral occipital cortex. In the last project, preliminary data showed that the risk of MS onset in post-partum was higher in XYp when compared to XXp. I also found that XXp patients had higher spine lesion load at diagnosis and registered higher ARR, while XYp had more frequently brainstem involvement at onset, presented more frequently with progressive MS phenotype at last clinical follow-up, and reached lower scores at PASAT. OCT revealed that, despite having a similar age and disease duration, XXp patients had lower RNFL and GCIPL thickness when compared with XYp, although the difference was not significant. I also found similar trends in XYp and XXp when these groups were compared to NLp. However, the RNFL and GCIPL was again higher in XYp when compared to NLp, while non-significant differences were detected between NLp and XXp. Overall, my results support the hypothesis that XY and XX fMCs could differently modulate the inflammatory and degenerative processes underlying MS. Discussion. The results reported in this thesis demonstrated that the sex of offspring could influence disease features in MS. Being most of the changes occurring during pregnancy not different in male and female pregnancies, the results obtained from two retrospective studies and the preliminary findings of the prospective study suggested that (1) fMCs could be one of the pregnancy-related factors modulating the disease onset and course, and (2) the sex chromosome of fMCs could play a role on the biological processes underlying MS. Conclusion. The hypothesis that a small percentage of cells with an XX or XY genotype could, through the expressions of sexual chromosome genes, regulate the maternal immune system and the repair mechanisms activated in the mother is fascinating and lends a fresh perspective to the sexual differences in neurological diseases. My findings suggested that XX and XY fMCs could, although marginally and likely interacting with other factors, be involved in MS inflammation and axonal degeneration, influence the immune system activation, and induce mechanism of repair. Discovering whether the presence of fetal non-self chimeric cells, and their chromosomes, may play a role in the modulation of the nervous system and its pathology is surely one of the more interesting challenges for the future.