Research Progress on Early Molecular Diagnosis Methods for Type 1

Diabetes Mellitus (T1DM)

Jiaxu Tu

Shenzhen College of International Education, Shenzhen, China

Keywords: Biomarkers, Early Diagnosis, Type 1 Diabetes Mellitus.

Abstract: Type 1 diabetes mellitus (T1DM) is a chronic disease with a common symptom of hyperglycemia, which is

caused by the loss of pancreatic beta-cells, leading to insulin defi-ciency. Recently, there has been much

research regarding a wide range of detection methods for T1DM. The research methods involve fields such

as immunology, epigenetics and microbi-ology. With more and more biomarkers being detected, earlier

diagnosing markers appear more in researchers’ eyes. But none of them have nearly 100% specificity and

effectiveness, and they also do not fit completely with the general pattern. In this review, the writer refers to

methods such as general biomarkers, immunology, and genetics and discusses them. The writ-er gets clear

attributes and limitations for each method and objectively evaluates them for guid-ance in future research

direction. Much further research can be done to get a clearer under-standing of newly discovered biomarkers

and come up with more assertive detection methods like combination testing.

1 INTRODUCTION

Type 1 diabetes mellitus (T1DM) is a chronic disease

with a common symptom of hyperglycemia, which is

caused by the loss of pancreatic beta-cells, leading to

insulin deficiency. The peak morbidity of T1DM is at

the beginning of adolescence, but the incidence in

adulthood is increasing attributed to the expansion of

human lifespan (Vanderniet et al., 2022). The most

common type of T1DM is autoimmune T1DM,

defined by the body’s immune system mistakenly

attacking its own organs, which is the pancreas in this

case. Nowadays, T1DM already has a well-accepted

way of treating it – insulin injection. This can

effectively control the blood glucose level in the

patient’s body, but it still has limitations: the insulin

dosage cannot be adjusted accurately based on the

real-time glucose level, especially when there are

extreme blood sugar fluctuations happened

commonly in the late stage. So, accompanied by

multiple other factors, including severe complications

if T1DM has developed like microvascular damage,

early detection and management of the disease is

needed in order to prevent them. For instance,

identifying T1DM before the symptom starts can

hugely decrease the risk of having diabetic

ketoacidosis from 60% to less than 5% (Simmons &

Sims, 2023).

Mănescu et al. pointed out that the majority of

biomarkers currently studied and validated are late

indicators, which have limited benefit for controlling

the progression of the disease. Also, many reviews

indicate the prevalence of researching prediction

methods for T1DM is in stages 1 and 2: after

seroconversion and islet autoimmunity, but if the

disease can be diagnosed before stage 1, the focus can

be moved on to prevent it rather than delay or treat it,

also implying the significance of predicting the

incidence as early as possible (Mănescu et al., 2024).

Much research recently regarding the early detection

of T1DM will not only provide more accurate ways

of diagnosis but also have the potential to discover the

disease when stage 0 to enable the avoidance of the

disease. The methods of early detection of T1DM

include the monitoring of related biomarkers, like

GAD and IA-2 antibodies; immune cells, like CD8+

T cells and B cells; and gene expressions, like HLA

genes.

In this article, the author is going to review the

recent progression of the molecular diagnosis of

T1DM in the early stage, including the methods

mentioned above, to help clarify the future direction

of research and to provide future research with

reference.

112

Tu, J.

Research Progress on Early Molecular Diagnosis Methods for Type 1 Diabetes Mellitus (T1DM).

DOI: 10.5220/0014424000004933

Paper published under CC license (CC BY-NC-ND 4.0)

In Proceedings of the 1st International Conference on Biomedical Engineering and Food Science (BEFS 2025), pages 112-116

ISBN: 978-989-758-789-4

2 BIOMARKERS

1.1 Beta Cell-Related Biomarkers

1.1.1 GAD Antibodies

Glutamic acid decarboxylase (GAD) antibody

(GADA) is an easy tool to detect T1DM. GAD

decarboxylates glutamic acid to produce gamma

amino butyric acid(GABA), which is an inhibitory

neurotransmitter that suppresses glucagon secretion

on pancreatic alpha cells during hyperglycemic states.

Additionally, GABA acts as a growth factor on beta

cells and also promotes the conversion from alpha

cells to beta cells. One isoform of GAD,

GAD65/GAD2, is present mostly in the pancreatic

beta cells. While GADA binds with GAD65 in beta

cells, beta cells will be damaged, and insulin

production will be reduced (Keshavarzi et al., 2022).

According to the Finnish Type 1 Diabetes

Prediction and Prevention Study (DIPP), they found

biomarkers like GADA are present in people’s serum

before their diagnosis for an average of 1.5 years,

even in the first year of some of their lives. GADA

will remain increasing after T1DM pathogenesis

initiates, and finally declines at the late stages. Using

enzyme-linked immunosorbent assay (ELISA),

GADA detection has a sensitivity of 60.8% and

specificity of 100%, and T1DM diagnosis has 100%

of the positive predictive value. These properties give

GADA a crucial role in the diagnosis of the disease

(Keshavarzi et al., 2022). In Japan, GADA is

prescribed as the first biomarker when diagnosing

T1DM. However, only 90% of people with T1DM

will be detected with GADA, while other

autoantibodies are positive in results, so multiple

autoantibody testing is important for the accuracy of

diagnosing T1DM (Kawasaki, 2023).

1.1.2 IA-2 Antibodies

Insulinoma-associated protein 2 (IA-2), also known

as islet cell autoantigen 512 (ICA512), is a

transmembrane protein found at insulin secretory

granule membranes in pancreatic beta cells. Its role is

to regulate the content inside the insulin secretory

granule and to help with the growth of beta cells.

When cytotoxic T cells initiate the autoimmune attack

on islet cells in the pancreas, IA-2 antibodies and

other antibodies such as GADA will appear. In the

studies performed in Colorado from 1993 to 2006,

Finland from 1994 to 2009, and Germany from 1989

to 2006, they are the signs of 100% risk of T1DM

during the whole lifespan and can be observed readily

at stage 1(categorized by The Juvenile Diabetes

Research Foundation, American Diabetes

Association, and Endocrine Society) of T1DM when

autoimmunity starts with normoglycemia (Kawasaki,

2023).

1.1.3 C-peptide

Connective peptide (C-peptide) is an amino acid

sequence that connects the A chain and the B chain of

the proinsulin together. The preproinsulin

synthesized in the ribosomes of pancreatic beta cells

goes through several further modification processes

in the granular endoplasmic reticulum, Golgi bodies,

and clathrin-coated secretory granules with

convertases 1 and 2; with their help, a preproinsulin

will be finally converted to an insulin molecule and a

C-peptide residue, stored in the secretory granules of

beta cells then secreted to the blood. The amount of

C-peptide secreted is equal to the moles of insulin;

although there is clearance of it by the kidney and

liver, the amount is too small and can be neglected.

Thus, individuals with T1DM are normally observed

with C-peptide levels below the normal range

(0.2~0.4 nmol/L, dependent on body weight)

(Maddaloni et al., 2022).

Additionally, the proinsulin/C-peptide ratio

shows the potential of telling a lot of information: a

study of non-diabetic twins of parents with insulin-

dependent diabetes shows that they have a high value

of this ratio due to insulin processing abnormalities

caused by pancreatic beta cell dysfunction. But more

researches are needed to confirm its reliability

(Maddaloni et al., 2022).

1.2 Metabolic Biomarkers

1.2.1 Insulin and Blood Glucose

Insulin is a protein hormone that is secreted by the

pancreatic beta cells, and it plays an extremely crucial

role in glucoregulation. More specifically, it helps to

lower the body's glucose level by promoting its

uptake into muscle and adipose tissue in the form of

glycogen or to be oxidized. Also, insulin has

significant effects on lipid and protein metabolism.

So, a lower-than-normal insulin level might be

suspected as T1DM. If the disease progresses,

absolute insulin deficiency might result. This can be

measured by ELISA using blood, plasma, serum, or

saliva (Wolkowicz et al., 2020).

Research Progress on Early Molecular Diagnosis Methods for Type 1 Diabetes Mellitus (T1DM)

113

Glucose is the monosaccharide that is involved in

multiple metabolic pathways in the body. The

hyperglycemia, a condition with excess glucose

concentration in the bloodstream, might be an

indicator of T1DM as well. This can be measured by

Amperometry using capillary blood (Wolkowicz et

al., 2020).

Simultaneous detection of insulin and glucose can

now be accomplished by using self-monitoring chip-

based point-of-care devices and continuous glucose

monitoring (CGM) devices easily (Wolkowicz et al.,

2020).

But these two can only be detected when the

destruction of beta cells occurs, and the damage is

irreversible. Furthermore, these symptoms are not

limited to T1DM, which is not good for treating the

disease compared to more early detecting methods.

3 IMMUNOLOGICAL METHODS

3.1 Immune Cells

A class of T cells with CD8 surface markers with the

main function of directly killing infected and tumor

cells is called CD8+ T cells. They are also known as

cytotoxic T cells which release cytotoxins and

produce cytokines to do their function. For the

pathogenesis of T1DM, autoimmune T cells will

recognize the autoantigens on beta cells presented by

Antigen-Presenting Cells (APCs), and then convert

into effector T cells in a very quick process. CD8+ T

cells are the main contributor to the destruction of

islet beta cells. CD8+ T cells are present commonly

among all the human pancreas, but for people with

diabetes, their amount might increase. The specific

type of autoreactive CD8+ T cells that appear in most

T1DM patients is preproinsulin-reactive CD8+ T

cells, suggesting preproinsulin might be the primary

target in the early development of T1DM (Yang et al.,

2024).

Some researches recently suggested that

autoantibodies involved in T1DM are not pathogenic:

the primary reason for T1DM development is

autoimmune T cell responses (Herold et al., 2024).

3.2 B Cells

B cells are white blood cells that produce antibodies

and help with immunological memory, but as

mentioned above, B cells do not contribute to the

progression of T1DM much by secreting

autoantibodies to destroy islet beta cells. They

actually act as APCs that selectively recognize

insulin, which will promote the development of

disease in non-obese diabetic mice. The anergy of

high-affinity insulin-binding B cells is observed to be

lost both before and at the time of T1DM diagnosis.

It is worth noting that people diagnosed with diabetes

before the age of 7 have a common situation with a

high number of B cells infiltrating the pancreatic islet

cells and a fewer number of beta cells remaining

compared to the people diagnosed at an older age

(Herold et al., 2024).

Since T1DM is an autoimmune disease, T cells

and B cells work together, that initiate and worsen the

disease. As they are from the immune system of

human bodies, it makes T1DM harder to cure unless

using more invasive measures, but it could also bring

considerable risks to the patient.

4 GENE EXPRESSION AND

EPIGENETICS

4.1 Related Genes

4.1.1 HLA Genes

HLA (Human Leukocyte Antigen) is a group of

proteins found on the surface of the cell that associate

the presentation of exogenous and endogenous

peptides to T-cell receptors, indicating that they are

also a part of the major histocompatibility complex

(MHC). Specifically, HLA class I gene is responsible

for encoding the proteins that present endogenous

peptides, and HLA class II gene is responsible for

encoding the proteins that present exogenous

peptides. After antigen, HLA molecules, and T cell

receptors bind, they release a signal to initiate the T

cell response targeting the antigenetic peptide in the

beta cells. Thus, the effector function will begin and

contribute to T1DM progression. The risk factors of

T1DM are varied, and it is very complex to predict

the risk due to diverse data sets, which are hard to

conclude with a common pattern. Haplotype of HLA-

DR3-DQ2.5 is commonly linked with T1D but with

various strengths of association, as well as conserved

HLA haplotypes such as B8-DR3 and A1-B8-DR3.

There is also a very high-risk heterozygous genotype

with as large as 45% of people with the risk of T1DM

in some studies: DR3/DR4 (Noble, 2024). Overall,

predicting T1DM using the genetic method is quite

complicated as the pattern is not consistent due to

large population numbers with high heterogeneity

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and multiple alleles, even though some of them are

also varied across the population.

Also, to help with the prediction of T1DM, using

high-throughput gene expression profiling such as

microarrays and RNA sequencing can help with

identifying early biomarkers and tracking the disease

progression by measuring the protein-coding genes’

expression levels.

4.2 Epigenetics Role

4.2.1 DNA Methylation

DNA methylation refers to a biological process in

which one methyl group from the S-adenosyl-L-

methionine is transferred onto the C5 position of

cytosines, mostly on CpG sites in mammals, and is

catalyzed by DNA methyltransferases. It typically

functions in transcriptional regulation/repression and

maintaining genome integrity. Dysregulated DNA

regulation has been observed recently in T1DM. In

the genome-wide DNA methylation analysis of

purified CD14+ monocytes from monozygotic twin

pairs with different T1DM onset, researchers have

identified 132 T1DM-methylation variable positions

such as HLA-DQB1 and GAD2. They are strong

evidence as they can be found at an early stage before

the diagnosis and symptomatic T1DM. Further

findings contribute to proving the implications of

DNA methylation on the pathogenesis of T1DM

(Zhang et al., 2021).

4.2.2 miRNAs

micro RNA (miRNA) is a type of small non-coding

RNA (ncRNA) that regulates gene expression post-

transcriptionally by binding to the 3’-untranslated

region(UTR) of target mRNA transcripts, resulting in

mRNA cleavage or repression of productive

translation. In the case of T1DM, miRNAs affect the

autoimmune initiation, beta cell dysfunction, and

apoptosis. miR-98, miR-23b, and miR-590-5p have

been shown to be overexpressed in CD8+ T cells from

patients with T1DM. By trying to block these

miRNAs, the incidence rate of T1DM can be

effectively controlled (Zhang et al., 2021). This can

also provide insight for future treatment of T1DM,

but efficiency and feasibility are waiting to be tested.

5 DISCUSSION

This study highlights the potential of molecular

biomarkers like C-peptides, several antibodies,

insulin, glucose, etc. in the early detection of T1DM.

Higher presentation and overexpression of most

factors above will be suspected to the development of

T1DM, while the under presentation of factors like C-

peptide will also imply the risk of T1DM. Early

detection methods are varied, but ideally, they have

to be as early as possible since the destruction of islet

beta cells is nearly irreversible, attributed to their

complex structure and still limited understanding of

the full picture of this disease. Insulin and glucose

levels have significant restrictions to assist the early

detection and mitigation of the disease in prior as they

are symptomatic signs. Their attributes based on

mechanism and understanding of the disease make

them be found after mostly other biomarkers and

signs mentioned in this article. For antibodies and

genes, their heterogeneities among the population

make their accuracy to be low, which could lead to a

delayed diagnosis that causes exacerbation of the

disease and delayed management. So, some scientists

suggest that multiple tests at the same time might be

a solution to this problem, but more work should be

done to test their efficiency and figure out the best

combination of testing that can be used in future

diagnoses medically. A good thing is that more and

more genes and ncRNAs are discovered to have a

relation with T1DM, but this also indicates that a

wide range of effort should be put in to discover more

of their mechanism, application, and so on.

6 CONCLUSION

In the recent five years, there has been huge progress

in research regarding the measures of T1DM early

detection. Methods ranging from immunology to

epigenetics to microbiology are devoted to more

accurate and reliable prediction and detection of

T1DM, especially in the early stages, which enables

people to manage the disease prior to its exacerbation.

This article provides various reviews regarding the

mechanisms of ways of diagnosis. The writer

discussed their limitations and evaluated their future

directions of research by summarizing the emerging

biomarkers discovered and initially proved to have

the potential associated with detection. In this article,

not all the detection methods and biomarkers are

listed and discussed, and some of them are not rich in

Research Progress on Early Molecular Diagnosis Methods for Type 1 Diabetes Mellitus (T1DM)

115

detail. Combination detection seems to have strong

potential in future applications, so more research is

needed to develop this method, and then, the pattern

researchers will get can contribute to the finding of

the best and most general way of combination testing.

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