الفهرس 
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Abstract
Multiple sclerosis is the most common demyelinating autoimmune disease affecting central nervous system and causing a range of physical, mental, and psychological problems.
Multiple sclerosis pathological events include inflammation, demyelination, axonal loss and gliosis. Its pathophysioloigy is mainly cell mediated with inflammatory cellular infiltrates that mainly contain Th1, Th17, cytotoxic T lymphocytes (CTL), macrophages including activated microgilia, and other inflammatory cells.
The disease has different types of disease courses including clinically isolated syndrome, radiologically isolated syndrome, relapsing remitting, primary progressive, secondary progressive, progressive relapsing and even pediatric MS which should not be confused with acute disseminated encephalomyelitis in children.
T helper 17 cells are pioneers concerning trafficking the blood brain barrier BBB to invade the CNS, They can use their cytokines as IL17 and IL22 and chemokine receptors such as CCR6, that binds to CCL20 on choroid plexus epithelium, and CCR4, that binds to CCL17 and CCL22 in CSF of MS patients.
Follicular T helper cells have a crucial role in germinal center reactions which includes B cell and macrophage activation, antibodies class switching and affinity maturation, and production of long lived plasma cells and B memory cells.
CD278 or inducible T cell co stimultor is crucial for T fh cells proliferation, differentiation, and cytokine production like IL21, while CXCR5 chemokine receptor recruits the to the B cell follicles to produce a germinal center reaction.; thus, by increasing CXCL13 levels in CSF, more Tfh cells accumulation will occur and increasing EDSS scores will be achieved.
The major roles of B cells in MS disease progression include neuronal and oligodendrocytes specific self antigen presentation specially in ectopic CNS existing germinal centers, proinflammatory cytokines production, contribution to the maintenance of ectopic germinal centers, and production of class switched affinity matured autoantibodies against CNS antigens and against anti inflammatory cytokines by long lived plasma cells, which are responsible for epitope spreading, complement activation, and recruitment of cell mediated immune players. Including CTLs and macrophases, which take the upper hand in farther disease progression.
According to the 2017 revision of the McDonalds criteria, the presence of at least 2 CSF specific OCBs can replace dissemination in time for accelerating diagnosis of MS.
IgG index is the ratio between CSF/serum IgG to CSF/serum albumin, and as the IgG is a large molecule, it cannot easily cross the BBB; accordingly, elevated OCBs without identical match in serum reefers to intrathecal production of autoantibodies. In fact, IgG
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index larger than 0,66 is considered as an evidence of intrathecal production of OCBs and vice versa.
Expanded disability status scale is a well known method through which accumulated MS disabilities can be quantified. It depends on neurological examinations regarding seven functional systems in addition to ambulation. Only visual and sphincter functions need conversion before calculation. Generally, impaired ambulation starts to occur above 4.5 and score 10 refers to death.
As MS is an inflammatory disease that is characterized by neuroinflammation, BBB breakdown, edema, axonal and neuronal loss, and gradual CNS tissue erosion, MRI can easily clarify these changes through comparing fatty healthy tissue to demyelinated or totally lost tissues.In general, Gadolium enhanced T1 weighted imaging (Gd T1WI) hyperintense lesions mark BBB breakdown, T1WI hypointense lesions or black holes indicates permanent nerve damage, T2WI hyperintense lesions detect demyelination and edema, T2 FLAIR is used to clarify periventricular and cortical hyperintense demyelination lesions without being confused by bright CSF signals, and Gd enhanced T1WI spinal cord lesions, especially when the entire cord is scaned,show better correlations with EDSS score increase.
MS lesions are commonly affecting special white matter regions including periventricular, cortical and juxtacortical, the corpus callosum, infratentorial reasons such as pons and cerebellum, and the spinal cord.
The McDonald criteria are first described in April 2001. It has 3 revisions in 2005,2010,and 2017. It depends mainly on the presence of at least 2 MRI separated lesions at MS characteristic sites either disseminated in space and/or time.
Immunophenotyping is the analysis of heterogeneous cell populations in order to identify and quantify the expression of investigated cell populations. It depends on protein expression analysis of cell identification cell surface markers and intracellular lineage specific cytokines using several antibodies together each of which is conjugated to a different fluorescent dye.
Lumber puncture or spinal tap is a needle mediated sampling of cerebrospinal fluid from the spinal cord for lab testing. Blood clotting problems should be taken into consideration before CSF sampling. As it is an invasive procedure, it can cause annoying complications including headache, backache, lower limb pain, herniation, and possibility of infections.
This study aimed at investigating the correlation between the CSF expression levels of T helper 17 cells, follicular T helper cells, and plasma cells, and multiple sclerosis disease progression. It included 26 MS patients who diagnosed according to the 2017 revision of McDonald s criteria as a patients group and 8 individuals who had pseudotumor cerebri / idiopathic intracranial hypertension as a control group.
With respect to the types of disease courses, 57.7 % of patients (15 patients) showed RRMS course, while 42.3 % (11 patients) of them had SPMS disease course.
Referring to MS disease progression, in terms of EDSS scores among patients group, the mean score was 4.38±1.71. Roughly half of them scored moderate progression (3 till 5
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scores), 30,8 % of them had sever progression(more than 5 scores), while 19.2 recorded mild symptoms (scores less than3).
Regarding IgG index values among patients, the taken positivity threshold was 0.7. The mean value was 1.04±0.54, nearly half of them 57.7% scored less than 1, 7.7% of them had high values (more than 2), and about one third of them 34.6% scored moderate positive values.
In our study, there was no statistically significant correlation between EDSS and IgG index values; thus, IgG index cannot be taken as a measuring tool for estimating disease progression.
This study illustrated a statistically significant difference in respect to plasma cell expression profiles in CSF in favor of patients group with p value of 0.001.
In the present study, there was a statistically significant difference in favor of patients group concerning the expression of CD4 positive Th cells in CSF samples with p equals to 0.007.
Concerning the difference between patients and control groups with regard to follicular T helper cells expression in CSF samples, a statistically significant difference was recorded in favor of the patients group with p value of 0.026.By analyzing Tfh expression profiles in depth in CSF samples, both CD278 and CXCR5 surface markers expression profiles revealed statistically significant differences in favor of patients group with p values of 0.028 and 0.0001 respectively.
In respect to Th 17 expression patterns differences in CSF samples between patients and control groups, an obvious statistically significant difference was observed in favor of the patients group with p value of 0.0001. Talking in depth about this issue, all the CSF expression patterns of the Th17 cell surface markers, including CD194, CD196, and CD4/IL23R gated cells, experienced statistically significant differences in favor of the patients group with p values of 0.0001 for both CCR4 and CCR6, and 0.002 for CD4/IL23R gated cells.
In our study, among the patients group, the plasma cell expression profiles showed a statistically significant negative correlation with EDSS scores increase with p value of 0.033. This reveals the importance of cell mediated rather than humoral immunity in MS disesse progression.
This study illustrated a statistically significant positive correlation between CD4 positive T helper cells expression levels in general and MS disease progression represented by EDSS scores raise with p value of.046.
Ironically, the CSF expression profiles of CD4/IL23R gated cells revealed a non statistically significant correlation with EDSS scores increase with p value of 0.589. This may be due to the small sized patients group.
Our study illustrated a statistically significant inverse correlation between the CSF Th17 expression levels among the patients group and MS disease progression represented by the EDSS scores elevation 0.029. By analyzing the expression levels of Th17 cell surface markers, CD194 experienced a statistically significant negative correlation with
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EDSS scores increase with p value of 0..039, while CD196 expression levels did not correlate significantly with EDSS scores elevation and MS disease exacerbation.
Regarding the correlation between follicular T helper cells accumulation in CSF in the patients group and MS disease progression, not only did CSF expression profiles show a statistically significant positive correlation with EDSS values raise, with p value of.018, but also their cell surface marker CD278 expression profiles revealed a statistically significant correlation with MS disease progression with p value of 0. 022; however, CXCR5 CSF expression levels did not achieve a statistically significant correlation with increasing EDSS scores and disease progression.
As expected, neither the CSF expression profiles of plasma cells, Th17, and follicular T helper cells nor the CSF expression profiles of their cell surface markers could record any statistically significant correlation with the increase of IgG index values. These results emphasize the statistically non significant correlation between IgG index values and MS disease progression represented by EDSS scores increase.
6.2 Conclusions
from the current study the following were concluded:
The expanded disability status scale (EDSS) as a measuring tool of multiple sclerosis (MS) disease progression had no statistically significant correlation with IgG indexes values increase.
Plasma cell trafficking to CNS had a significant difference in favor of patients group; thus, it can be considered as a common feature distinguishing MS patients. Nonetheless, their accumulation in CNS of MS patients was inversely correlated with EDSS score increase and disease progression paving the way for cell mediated immunity to take the upper hand in the management of MS disease progression.
CNS existing CD4 expressing T helper cell expression profiles showed a positive correlation with EDSS scores raise and MS disease progression referring to their ability to reinforce both humoral / fluid mediated and cell mediated immunity.
As Th17 cells are pioneers in trafficking blood brain barrier BBB, their accumulation in patients CSF scored a statistically significant difference in favor of patients group; nevertheless, as the disease progress, their CSF Th17 expression experienced a significant negative correlation with increasing EDSS scores, which indicate disease progression; accordingly, CSF th17 profiles are potent indicators of MS disease onset not progression.
Owing to the small size of the patients group, interleukin 23 receptor IL23R expression revealed a non significant correlation with MS disease progression which was represented by EDSS scores increase.
Follicular T helper cells illustrated a statistically significant difference in favor of patients group; additionally, their expression among patients showed a significant positive correlation with EDSS score raise and MS disease exacerbation; accordingly, Tfh cells can be used as excellent indicators of both disease existence and disease progression.
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The simultaneous decease in CSF Th17 and increase in Tfh expression profiles accompanying EDSS score increase and MS disease progression, may reflect a skewing behavior of Th17 cells toward Tfh expression profiles
Despite the importance of oligoclonal bands OCBs in CSF and IgG index in accelerating MS diagnosis, IgG index values showed no significant correlation with EDSS scores, which reflects disease deterioration.
As MS disease is mainly cell mediated, IgG index values scored neither any statistically significant correlation with all cell types under investigation nor their surface markers expression profiles in CSF of MS patients; hence, IgG index is useless in monitoring disease progression.
6.3 Recommendations
1- Using a large sample size to increase the authenticity of the statistically significant correlations and achieve more obvious diversions between MS patents and healthy controls.
2- Expand the study patients base to include all different disease courses of MS from clinically isolated syndromes to progressive relapsing and pediatric MS.
3- Performing a large meta analysis studies in order to emphasize the actual correlation, if present, between EDSS and IgG indexes among MS patients.
4- Plasma cell studying is not a proper choice for estimating the exacerbation of MS disease due to the fact that MS is mainly a cell mediate autoimmune inflammatory disease.
5- Estimating more types of B cell populations in addition to plasma cells in CSF of MS patients including germinal center B cells, as CD83 expressing, DC-SIGN antigen presenting B cells, plasmoblasts and also myelin specific memory cells. This will expand our understanding of the variable roles of B cell populations in MS disease progression.
6- As autoimmune response in MS is mainly cell mediated, studying the correlation between cell mediated responsible cells such as CTL, natural killers NK, activated macrophages etc., with increasing disease progression, represented by EDSS scores, will be more objective.
7- Shedding more lights on Tfh subpopulations(TH1 like, Th2 like, and Th17 like) and monitoring their behaviors during disease exacerbation in CSF of MS patients will clarify the mysterious scene of skewing towards Tfh profiles; hence, targeting CSF Tfh cells by receptors specific medications can mitigate disease progression.
8- Estimating expression and functional aspects of regulatory T cells populations, including natural and induced Treg, in peripheral blood and CSF of MS patients. In addition, correlating their abundance and function, especially in CSF, among MS patients will increase our understanding of MS immunological onset and progression as an autoimmune disorder.