JMIR Publications

ABSTRACT

Background:

BACKGROUND Type 2 diabetes (T2D) is a disease characterized by chronic hyperglycemia, which gives rise to metabolic and signaling abnormalities (1-3). According to Kayal and Graves, these metabolic and signaling abnormalities have been reported to cause dysregulated innate immunity (1). Chronic dysregulated immunity, therefore, includes the changes in immune cells such as neutrophils, lymphocytes, monocytes, eosinophils, and basophils (4-7). Upon activation, these immune cells play a different role, including secretion of inflammatory markers such as C-reactive protein (CRP), tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), P-Selectin, Cluster of differentiation 40 ligand (CD40L), and Fibrinogen (4, 5, 8-11). The chronic immune activation in T2D, therefore, result in a suppressed immune system (1, 12). According to Lam and LeRoith, a fundamental change in the population suffering from T2D is witnessed by the healthcare communities (13). This was confirmed by the International Diabetes Federation (IDF) statistics, reporting that in 2019, were 19 million diabetic people in Africa aged between 20 and 79 years (3). IDF also reported that 12 million Africans aged between 20-79 years living with undiagnosed diabetes in 2019 (3). South Africa is the highest with 4.6 million diabetic adults (20-79 yrs.) (3). In 2017, the Indian population had been reported to have the highest prevalence of diabetes in South Africa by 11-13%, followed by the coloured population by 8-10%, then blacks by 5-8%, and whites being lowest by 4% (14, 15). The Indian population has been shown to be high due to their strong diabetes genetic predisposition (14, 15). However, the onset of T2D arises from the progression of the pre-diabetes stage (16). This pre-diabetes stage has been reported to be an asymptomatic stage, creating a research complication in its documentation of the statistics and prevalence. There is less evidence on the changes in immune cells and selective inflammatory markers at the pre-diabetes stage (17-20). However, in our laboratory, research has been done on animals in addition to the available research reporting the metabolic and signaling abnormalities, including immune activation on the prediabetes stage (21-24). This then raised a debatable issue if the same abnormalities occur at the pre-diabetes stage in the human subject due to limitations in the animal subject even though the research mimicked the human diet. From the search done, we found no report or evidence of the systematic review that reports on the changes in immune cell concentration and the level of secretion of selective inflammatory markers that occur during immune activation at pre-diabetes stage. Therefore, it is of great opportunity to compile a systematic review that will yield an exhaustive synthesis obtained from the available studies that previously reported on immune cells and selective inflammatory markers of interest at pre-diabetes stage. Additionally, this systematic review will give reports on the impact of demographics on changes of immune cells and secretion of selective inflammatory markers at pre-diabetes stage.

Objective:

OBJECTIVES 1. To determine the changes in immune cells (neutrophils, lymphocytes, monocytes, eosinophils, basophils) concentration at the pre-diabetic stage. 2. To investigate if there are changes in concentration on selective inflammatory markers (CRP, TNF-α, IL-6, P-Selectin, CD40L, and Fibrinogen) at the pre-diabetes stage. 3. To determine the impact of demographics on immune changes and secretion of selective inflammatory markers at the pre-diabetic stage.

Methods:

METHODS This protocol was prepared by adhering to the preferred PRISMA 2015 guidelines for reporting protocols (PRISMA checklist attached in additional file). Systematic review registration The protocol has been registered with PROSPERO with registration number "CRD42020184828” dated 05-07-2020. Eligibility Criteria for the study A minimum of 100 population and the studies that report community-based clinical cross-sectional study will be eligible. The inclusion and exclusion criteria will be as follows. Inclusion: The information reported from non-diabetic adults within the ages of 25-45 of all ethnicities will be used. Exclusion: Information reported from people with a history of liver disease, kidney disease, heart disease, and depression will not be used. Information from pregnant women will also not be used. Additionally, no samples from professional sports athletes will be allowed in the study. The full-text article/reports that indicate that the subjects that were used were free from all the mentioned criteria will then be eligible. Pre-diabetes diagnosis criteria Diagnostic criteria for pre‐diabetes will be as follows (participants should meet one of the following diagnosis): fasting blood glucose (FBG): 5.6 ‐7.0 mmol/L; 2 h postprandial blood glucose (2 h ‐ OGTT): 7.8 ‐11.0mmol/L with Glycated hemoglobin (HbA1c): 5.7‐6.4%. Study design Information Sources Participants: Intervention: The clinical studies that involve observational studies if they will be cross-sectional, comparative cross-sectional, case-control, or cohort study designs that involve normal/non-diabetic and pre-diabetes reports. The reported information that involves specifically one or more immune cells (neutrophils, lymphocytes, monocytes, eosinophils, basophils) at the pre-diabetic stage will be eligible for this systematic review. Additionally, a study that reports information that involves at least one or more inflammatory markers of interest, which are CRP, TNF-α, IL-6, P-Selectin, CD40L, and Fibrinogen, will also be eligible for this systematic review. Comparators: In this systematic review, the eligible comparing control groups will be non-diabetic/normal control and T2D control. OUTCOMES This systematic review is expected to have the outcome as follows; 1. The changes in immune cells (neutrophils, lymphocytes, monocytes, eosinophils, basophils) concentration at the pre-diabetes stage (reported as odd ratio and 95 % confidence interval). 2. The changes in concentration of selective markers (CRP, TNF-α, IL-6, P-Selectin, CD40L, and Fibrinogen) at the pre-diabetes stage (reported as odd ratio and 95 % confidence interval). 3. Changes in immune cells and selective inflammatory markers due to demographic impact such as the effect of gender, age, and race (reported as the mean). Search Strategy The electronic search strategy will be used as an identification for studies involving cohorts that have been done that are related to the study of interest. This strategy will be accomplished by search on MEDLINE (from 1963 to 2020), COCHRANE library displaying results of trials from PubMed, CT.gov, EMBASE, and ICTRP (from 1963 to 2020), and African Journal Online (from 1998 to 2020). Additionally, to these search strategies, clinical MeSH headings, and text words will be applied to filter the available information. For all search done, the keywords to be used will be "pre-diabetes and immunity,” "pre-diabetes and immune cells,” "pre-diabetes and leucocytes," and "pre-diabetes and inflammation." Identification of eligible studies The title and abstracts of all the obtained results will be screened by reviewers (NCM, AMS & AK), and the studies that meet the eligibility criteria will then be selected. Each reviewer will be responsible for screening all the selected study reports before the decision making of the eligible reports. The PRISMA flow chart for selecting studies will then be provided on reports from the systematic review. Patient and Public Involvement No patient involved. Data management Study Records and data extraction The data of study records selected as eligible reports will then be extracted and recorded in an Excel file. The pre-defined list of variables to be considered in each and every report will be used as categories in an excel file. Considering the research of interest, the outcome of interest will mainly be the immune cell response and concentration of selective markers in both genders, at an age parameter of interest in all ethnicities. However, the value of the baseline characteristic of the data reported will also be considered. Therefore, the baseline characteristics of eligible research reports obtained will be author, year of publication, country, and study setting. The methodology of the study reported will also be considered with the categories (design, time period, sampling strategy, and whether participants are normal or pre-diabetic population) considered. Finally, the outcomes from different gender, ages, ethnicity, immune cell changes /inflammatory markers will then be extracted. Data simplification Studies that report on the immune cells (neutrophils, lymphocytes, monocytes, eosinophils, basophils) will be combined into a single group. Additionally, the studies that report on selective inflammatory markers (CRP, TNF-α, IL-6, P-Selectin, CD40L, and Fibrinogen) will also be combined into a single group. Risk of bias The potential risk of bias in individual studies will be obtained using the Downs and Black Checklist (25). The scores will be rated as follows; excellent (25–26), good (20–24), moderate (14–19), poor (11–13), and very poor (< 10). Three reviewers (NCM, AMS, and AK) will be responsible for the independent judgments, which will be based on the four domains of the Black and Downs checklist tool which is reporting bias (10 items), external validity (3 items), internal validity (6 items), and selection bias (7 items). In a situation where there will be a difference in opinions between NCM, AMS, and AK. PSN will then be responsible for adjudication.In situations where the data is not clear. The investigator reported that data will be contacted three times, if no response ,data will be then excluded from eligible report. Data synthesis For the meta-analysis of reported data, a forest plot will be used from Review Manager software version 5.4 (RevMan) (26, 27). Using this forest plot, eligible data from all reported studies will be analyzed depending on their sample size and the mean of the concentration of immune cells or inflammatory markers in pre-diabetic and control groups. Additionally, an odds ratio and confidence interval will be used to plot the forest plot where the solid lines will represent the 95% confidence interval. Each reported study will be represented as a horizontal line on the y-axis to list the primary author and year of study. The forest plot will also include the weight of the study results that will be automatically obtained using Review Manager software. Sensitivity analysis RevMan Forrest plot will also test for heterogeneity, where greater homogeneity will be indicated by a greater overlap between the confidence intervals (28). Using the forest plot, I2 will then be calculated where a value between 0 and 100 % will be obtained. A value obtained less than 25% will be considered to be an indication of a strong homogeneity, and a value obtained greater than 75 % will then be considered to be an indication of a strong heterogeneity. However, a value of 50 % will be considered as an average value. Assessment of Strength of Evidence NCM, AMS, and AK will then be responsible for the assessment of the strength of evidence. The studies included in the review will then be evaluated using the Grading of Recommendations Assessment, Development, and Evaluation approach (GRADE) (28-30). Furthermore, the summary of the finding (SoF) table will then be created using a GRADE pro tool.

Results:

The synthesis of previous study reports obtained from this systematic review and meta-analysis will clarify the complications on the immune system at pre-diabetes, such as the changes that have been reported on immune cells, which are neutrophils, lymphocytes, monocytes, eosinophils, basophils. This systematic review and meta-analysis will also give an outstanding synthesis of data from previous reports based on selective inflammatory markers of interest.

Conclusions:

The synthesis from this systematic review and meta-analysis will create a hallmark of an impact of demographics at the pre-diabetes stage. This will clarify changes that might be observed in a study of interest-based on eThekwini district (South Africa). Clinical Trial: Registration details: This protocol has been registered with the International Prospective Registry of Systematic Reviews (PROSPERO) registration number "CRD42020184828” dated 05-07-2020).