Inflammatory cytokine levels in multiple system atrophy

Abstract Background: Multiple system atrophy (MSA) is a fatal neurodegenerative disease that progresses very rapidly and has a poor prognosis. Some studies indicate that the level of inflammatory cytokines may be related to MSA. However, no consistent conclusion has been drawn yet. The purpose of our research is to perform a meta-analysis to investigate whether the level of inflammatory cytokines is altered in MSA. Methods: Case-control studies on inflammatory cytokine levels in MSA will be searched in the following 3 databases: PubMed, Embase, and Web of Science from the database start time to March 17, 2020. Two independent authors will conduct research selection, data extraction, and quality evaluation. Data synthesis, subgroup analysis, sensitivity analysis, and the meta-analysis will be performed using Stata15.0 software. Results: This study will provide a comprehensive review of all studies on inflammatory cytokine levels in MSA. Conclusion: To the best of our knowledge, this study will be the first meta-analysis that provides the quantitative evidence of inflammatory cytokine levels in MSA. Registration Number: INPLASY202060034.


Introduction
Multiple system atrophy (MSA) is a fast progressive and fatal neurodegenerative disorder that is characterized clinically by autonomic failure, cerebellar ataxia, and parkinsonism in various combinations. [1,2] There are 2 main subtypes based on the clinical characteristics of MSA: (1) the MSA-P subtype with Parkinson's syndrome as the prominent manifestation and (2) the MSA-C subtype with the cerebellar ataxia symptoms as the prominent manifestations. [3,4] The prognosis of MSA is poor, with an average life span of 6 to 9 years from morbidity to death according to a report. [5][6][7] So far, there is only limited symptomatic treatment, and no effective drugs have been found to cure MSA. [8] Therefore, it is necessary to find biomarkers to help better understand the pathophysiology of MSA.
A great number of glial cytoplasmic inclusions in the oligodendroglia cytoplasm are a pathological hallmark of MSA. [9] Many studies have found that there are a greater number of activated microglia in MSA cases. [10,11] Potential toxic products include inflammatory cytokines and other inflammatory markers, produced and released by activated microglia. [12,13] Therefore, it is speculated that there is an inflammatory state in the brain of MSA, which may be associated with the neurophysiological cause of the disease. [14] To date, some studies have used easily accessible and less invasive peripheral blood to measure inflammatory cytokine levels in patients with MSA. [15][16][17][18]  because cerebrospinal fluid specimens are closely related to the brain and are not affected by drugs such as nonsteroidal anti-stimulation drugs. [19][20][21][22][23] Although some studies have evaluated inflammatory cytokines in MSA, the results are not always consistent. [15][16][17][18][19][20][21][22][23] Hall et al [23] found IL-8 is increased in patients with MSA compared to healthy controls. While Rydbirk et al [22] found no difference of IL-8 between patients with MSA and healthy controls. The results of individual studies can be quantitatively combined using metaanalysis techniques to increase the strength of the evidence. Thus, we will perform a meta-analysis to study the concentration of inflammatory cytokines in the peripheral blood and cerebrospinal fluid specimens of MSA patients.

Methods
In this systematic review and meta-analysis, the principles of the Preferred Reporting Items for Systematic reviews and Meta-Analyses checklist will be fully followed. [24] This protocol has been registered on the International Platform of Registered Systematic Review and Meta-Analysis Protocols (INPLASY) in June 2020, and its registration number is INPLASY202060034 (URL = https://inplasy.com/inplasy-2020-6-0034/).

Ethics approval
Because the data used in this paper are from published studies without the involvement of individual or animals' experiments, the ethical approval is not required.

2.2.
Eligibility criteria for study selection 2.2.1. Types of studies. We will choose case-control studies that compare the levels of cytokines between MSA patients and healthy controls. Inclusion criteria consisted of: (1) study design limited to case-control studies, (2) studies measuring peripheral blood or CSF inflammatory factor concentrations; (3) the inclusion of healthy subjects as controls.

Exclusion criteria included:
(1) Non-human studies, reviews, conference abstracts, editorials, or letters will be excluded unrelated to the research topic; (2) case reports and case series; (3) without healthy controls; (4) without necessary data; (5) the samples were collected before patients were diagnosed with MSA.

Types of participants. Patients (aged over 18 years)
diagnosed with MSA will be included in the study. Patients with other serious complications, a history of brain surgery, or other serious neurodegenerative diseases will be excluded from this study.

Types of interventions.
We will mainly study the differences in the level of inflammatory cytokines in cerebrospinal fluid or peripheral blood between MSA patients and healthy controls.

Type of comparators.
We will choose healthy controls, who has no disease.

Types of outcome measures.
Main results: Differences in the concentration of inflammatory cytokines in peripheral blood or cerebrospinal fluid between patients with MSA and healthy controls.

Search methods in the study
Two independent authors will search the following databases: PubMed, Embase, and Web of Science. The search strategy will include the following phrases:  The results of the above 3 database searches will be managed using endnote software (X7 version). Firstly, we will remove duplicate articles with the same author, title, and abstract (the same article is displayed in different databases). Secondly, we will screen the title and abstract of the article by 2 independent reviewers, select the studies that may meet the conditions, and exclude the studies that do not meet the selection criteria. Last but not least, we will filter the full text of the download. When the 2 reviewers disagree, we will discuss to solve it. If there are still objections, the third reviewer will analyze them. The reasons for the excluded articles will be recorded. The study selection process will be presented in the following Preferred Reporting Items for Systematic reviews and Meta-Analyses flow diagram (Fig. 1). Table 1 Search strategy for the PubMed database.

Number
Search terms 1  Multiple system atrophy  2  Multisystem atrophies  3  Atrophy, multisystem  4 Atrophies, multisystem 5 Multisystem atrophy 6 Multiple system atrophy syndrome 7 Multisystemic atrophies 8 Atrophy, multisystemic 9 Atrophies, multisystemic 10 Multisystemic atrophy 11 Multiple system atrophies 12 Atrophy We will extract the required data into Excel according to the established data extraction protocol. The contents collected will include: first author's name, year of publication, participants' country, age, number of patients and controls, female: male ratio, duration of disease, race, the mean and the standard deviation of cytokine concentration for patients with MSA and controls, United MSA Rating Scale. One reviewer will extract all the data, and another reviewer will independently verify the extraction method.

Risk of bias in included studies.
Two independent reviewers will independently assess the methodological quality of each included study. We will use the Newcastle-Ottawa scale checklist. [25] The range of this tool is from 0 to 9, which is from lowest quality to best quality. If there is a disagreement between the 2 reviewers, the issue will be resolved through discussion with the third reviewer.

Date analysis
We will use Stata 15.0 software to process and analyze the collected data. The Q test and I 2 will be used to test the heterogeneity of the study. When the heterogeneity is relatively high (I 2 > 50%), we will use a random-effects model to analyze the data; otherwise we will use a fixed-effects model.

Subgroup analysis
If the heterogeneity of the results is high and the data are sufficient, we will perform a subgroup analysis on the data in order to find the cause of the large heterogeneity. We will divide MSA patients into MSA-C and MSA-P subtypes according to clinical symptoms. At the same time, we will also analyze the race, nationality, duration of disease and other aspects of the subgroup analysis.

Sensitivity analysis
In order to ensure the robustness and reliability of the results, sensitivity analysis will be conducted by excluding highly biased studies.

Publication biases
To assess potential publication bias, if there is enough research, we will use funnel plot and Egger test.

Discussion
MSA is a very serious disease with a poor prognosis, so it is necessary to find effective biomarkers for diagnosing MSA.
Previous clinical studies have reported the relationship between inflammatory cytokines and patients with MSA. [15][16][17][18][19][20][21][22][23] However, there is no systematic comprehensive review and meta-analysis to discuss their relationship. This article will analyze the concentration of cytokines in the peripheral blood and cerebrospinal fluid in MSA patients. We hope to find biomarkers that provide a basis for auxiliary diagnosis of MSA.  24 14 AND 23 Table 3 Search strategy for the Web of Science database.  3  Cytokine  4  Chemokine  5  Interferon  6  Interleukin  7 Transforming growth factor 8 Tumor necrosis factor 9 C-reactive protein 10 2 OR 3 OR 4 OR 5 OR 6 OR 7 OR 8 OR 9 11 1 AND 10 MSA = multiple system atrophy.