Introduction

Diabetes has become a global epidemic disorder and the number of people with diabetes has been steadily increasing. Of these, type 2 diabetes mellitus (T2DM) accounts for the majority of diabetes cases. In 1985, only about 30Ā million people worldwide suffered from diabetes, however, by 2015, the number of the patients had increased to 392Ā million. The International Diabetes Federation estimates that approximately 463Ā million adults (ages 20ā€“79) worldwide have diabetes in 2019. Thatā€™s about 6% of the worldā€™s population. Chronic complications of diabetes are mainly related to lesions caused by microvascular injury, such as retinopathy, renal disease, cardiovascular disease and nerve damage (including peripheral nerve and autonomic neuropathy). Retinopathy affects approximately one-third of diabetic patients and is an important cause of visual impairment and blindness. Approximately 25% of T2DM patients suffer from diabetic nephropathy and end-stage renal disease. Nearly 50% of people with diabetes have diabetic neuropathy. The prevalence of erectile dysfunction in diabetic men ranges from 35 to 90%. Diabetic foot complications, including foot ulcers and amputations, are all the problems caused by microvascular injury1,2,3,4.

Vascular injury in diabetes is largely due to a combination of metabolic abnormalities, chronic hyperglycemia (high blood sugar levels), and related disturbances of various physiological pathways. The pathogenesis of diabetic vascular injury involves multiple mechanisms including: endothelial dysfunction, oxidative stress, inflammation, advanced glycation end products (AGE), increased platelet activation, dyslipidemia, atherosclerotic plaque formation, microvascular complications, impaired angiogenesis and wound healing. All of these mechanisms can cause damage to tiny blood vessels, leading to a series of vascular injury diseases5. Diabetic retinal, renal and neurological complications are usually late manifestations of microvascular injury that begins years earlier and is often influenced by other cardiometabolic disease factors (e.g. hypertension, obesity, dyslipidemia). Microvascular dysfunction in heart, muscle, and brain precedes anatomical microvascular disease. Early on, diabetes and/or cardiometabolic disease can cause reversible microvascular injury with dysfunction, which may or may not become irreversible and anatomically visible over time (e.g., Thickening of vascular basement membrane, sparse capillaries, loss of pericytes, etc.). Consequences may include common vision loss, renal insufficiency, and neuropathy, as well as escalating heart failure, sarcopenia, cognitive impairment, and metabolic dysfunction5,6. Platelet endothelial cell adhesion molecule-1 (PECAM-1), also known as CD31, is an important component of endothelial cell junctions and plays a key role in mediating cell-cell interactions, especially between endothelial cells and platelets effect. It is expressed on the surface of platelets, leukocytes and endothelial cells. PECAM-1 is involved in a variety of physiological and pathological processes, including leukocyte migration, platelet aggregation, inflammation and angiogenesis. PECAM-1 has been shown to inhibit endothelial cell proliferation, migration and tube formation, processes that are important in angiogenesis. It plays a role in regulating angiogenesis (the formation of new blood vessels) and endothelial cell function, helping maintain endothelial barrier integrity. PECAM-1 has a protective role in certain pathological conditions such as inflammation and atherosclerosis by maintaining endothelial barrier function and limiting vascular leakage7,8,9.

Based on the above background data, we hypothesize that the abnormal expression of PECAM-1 in T2DM patients may be one of the potential pathological mechanisms causing T2DM vascular injury. To this end, we first collected the baseline information and plasma samples from T2DM patients and healthy controls, detected the plasma PECAM-1 levels. The results of the multivariate logistic regression model revealed that PECAM-1 was an independent risk factor for vascular injury in T2DM patients. Further, bioinformatics analysis suggested that the alteration of PECAM-1 transcription factors, CREB3, TEAD3, GATAD1, RELA, and CUX1, may be the mechanism of T2DM plasma PECAM-1 reduction. Finally, network pharmacological analysis indicated that fostamatinib is a drug with high affinity for PECAM-1. This discovery provides novel insights into the pathological mechanism of T2DM and also identifies a new drug target for the treatment of T2DM.

Materials and methods

T2DM patients and health controls

Plasma samples were collected from 66 T2DM patients who were admitted to the Endocrinology Department of Putian University Affiliated Hospital from February 2021 to January 2023. Plasma samples were also obtained from 66 healthy adults who participated in outpatient adult health exams. Briefly, in the morning, a 5Ā ml forearm blood was drawn into a tube with sodium citrate anticoagulant from both the T2DM patients and the healthy controls. The tubes were then spun at 3000 x g for 20Ā min. The yellow supernatant plasma was collected and stored in a -80Ā Ā°C freezer until it was used for ELISA assay. This study was reviewed and approved by the Ethics Committee of Affiliated Hospital of Putian University (approval number: 2022043). All patients and healthy controls participating in the survey agreed and signed informed consent forms, agreeing to provide plasma samples. This study is carried out in full accordance with the Declaration of Helsinki.

T2DM diagnostic criteria and exclusion criteria

In this study, the diagnostic criteria for T2DM were based on the definition provided by the World Health Organization (WHO)10. Patients who met the following criteria were diagnosed with T2DM and included in the study:

  1. 1.

    Patients with symptoms of diabetes, such as polydipsia, polyphagia, polyuria, and unexplained weight loss caused by hyperglycemia, with a random blood glucose level of ā‰„ā€‰11.1 mmol/L (200Ā mg/dL).

  2. 2.

    Or, patients with symptoms of diabetes, as mentioned above, along with a fasting blood glucose level of ā‰„ā€‰7 mmol/L (126Ā mg/dL).

  3. 3.

    Or, patients with symptoms of diabetes, as mentioned above, along with a blood sugar level of ā‰„ā€‰11.1 mmol/L (200Ā mg/dL) two hours after a 75Ā g glucose tolerance test.

  4. 4.

    Or, patients with symptoms of diabetes, as mentioned above, along with a glycosylated hemoglobin (HbA1c) level of ā‰„ā€‰6.5 DCCT%.

Patients without typical symptoms of diabetes were required to undergo rechecking and confirmation on another day.

Diagnostic criteria for vascular injury

  1. 1.

    Detection via vascular ultrasound reveals an increase in the intima-media thickness (IMT) of blood vessels (carotid artery IMTā€‰ā‰„ā€‰1.0Ā mm or femoral artery IMTā€‰ā‰„ā€‰1.2Ā mm) or the presence of atherosclerotic plaques.

  2. 2.

    Or, there is a decrease in the blood flow velocity in the main renal artery and its branches, along with a reduction in the peak systolic velocity (PSV), accompanied by a urinary microalbumin-to-creatinine ratio (UACR)ā€‰ā‰„ā€‰30Ā mg/g or a serum creatinine level elevated beyond the normal reference range11,12.

The exclusion criteria for this study were as follows: (1) Pregnancy-related diabetes and other specific types of diabetes; (2) Secondary diabetes; (3) Acute complications of diabetes observed during hospitalization; (4) Severe complications, including concurrent tumors, infections, and multiple organ failure; (5) Missing data; (6) Presence of mental illnesses such as depression, mania, and anxiety; (7) Patients who did not sign the informed consent form.

ELISA assay

The level of plasma PECAM-1 was analyzed using a Human CD31 ELISA Kit (Cat.No.: JN23015) from Jining Biotechnology (Shanghai, China), by following the manufacturerā€™s instructions. For each sample, 20 ĀµL of plasma was diluted to 10, 40, or 160 times, respectively, and underwent the steps of the manufactureā€™s instruction to detect PECAM-1. The optical density (OD) value of each well at 450Ā nm was then examined by a Multiscan Spectrum (Thermo Fisher Scientific, Waltham, MA, USA). Based on a standard PECAM-1 curve, the concentration of PECAM-1 was calculated from the OD value. Each sample was repeat for three times.

Transcription factor analysis by networkanalyst

Using NetworkAnalyst (https://www.networkanalyst.ca/NetworkAnalyst/) analysis was applied to find out the PECAM-1 gene transcription factors. Based on the ENCODE database source, peak intensity signalā€‰<ā€‰500 and predicted regulatory potential scoreā€‰<ā€‰1 were used as screening coefficients to identify transcription factors related to PECAM-1 regulation.

Analysis of deferential expressed genes in T2DM blood mRNA from GEO database

Based on GEO (https://www.ncbi.nlm.nih.gov/geo/) database, a blood mRNA data of T2DM (GSE26168) was obtained, which comes from gene chip data from the Illumina HumanRef-8 v3.0 expression platform. A total of 8 control and 9 T2DM blood samples are included. The expression level of the PECAM-1 transcription factors found in NetworkAnalyst analysis were then obtained from the GSE26168 dataset (Fig.Ā 2). All significantly up-regulated and down-regulated genes were represented by a volcanic map (Fig.Ā 3A). Top 20 up-regulated and down-regulated proteins in the T2DMs were listed in a heat map.

The PECAM-1 interaction networks and function enrichment analysis

The proteins related to PECAM-1 function were analyzed using an online network database STRING (https://string-db.org/). The PECAM-1 function and interacting proteins in the human body was analyzed using nodes in the protein protein interaction (PPI) network (Fig.Ā 4A). Using the ClusterProfiler package in R language, the PECAM-1 functional related genes were divided into three parts through GO (Gene Ontology): cellular component (CC), molecular function (MF), and biological process (BP) (Fig.Ā 4B). KEGG (Kyoto Encyclopedia of Genes and Genomes) was employed to annotate the functions of the PECAM-1 gene and to determine which functional pathways of the PECA1 gene will involve in the human biological and pathological processes. (Fig.Ā 4C).

PECAM-1-targeted drug prediction analysis

The PECAM-1 targeted drug was detected with Targeted Medicinal Drugbank (https://go.drugbank.com/). A 3D structure of PECAM-1 was derived from PDB (https://www.rcsb.org/). The 3D structure of the small molecule of the predicated drugs was obtained from PubChem (https://pubchem.ncbi.nlm.nih.gov/). AutoDockTools 1.5.7 was employed to dock drug molecules that coordinate with PECAM-1 and predict their binding energy. Discovery Studio 4.5 was applied to visualize docking.

Statistical methods

Statistical Package for the Social Sciences 26.0 software (SPSS Inc., Chicago, IL, USA) was used for data analysis. Continuous variables were expressed as the meanā€‰Ā±ā€‰standard deviation, while categorical variables were presented as percentages. Logistic regression was employed to analyze the independent risk factors for vascular injury. Pā€‰<ā€‰0.05 on both sides was considered to indicate a significant difference.

Results

Significant decrease of plasma PECAM-1 level in T2DMs

A total of 66 patients with T2DM (mean age 59.0ā€‰Ā±ā€‰13.4 years, 45 males and 21 females) and 66 healthy individuals undergoing physical examinations (mean age 54.5ā€‰Ā±ā€‰10.4 years, 42 males and 24 females) were enrolled in this study. Plasma samples and relevant clinical and biochemical data were collected, and plasma levels of PECAM-1 in the T2DM and healthy control groups were compared via ELISA. The results showed that compared with the healthy control group, T2DM patients had higher glycated hemoglobin (HbA1c) (9.3% vs. 5.3%), cholesterol (6.9 mmol/L vs. 4.9 mmol/L), triglyceride (1.2 mmol/L vs. 0.8 mmol/L), platelet count (Plt) (217.2ā€‰Ć—ā€‰109/L vs. 177.0ā€‰Ć—ā€‰109/L), and body mass index (BMI) (24.3Ā kg/m2 vs. 22.0Ā kg/m2), and fewer engaged in aerobic exercise (53.0% vs. 71.2%). Significantly, plasma PECAM-1 levels in T2DM patients were remarkably lower than those in healthy control group (18.8 ng/mL vs. 36.3 ng/mL) (TableĀ 1), suggesting that PECAM-1 may be implicated in the pathological mechanism of vascular injury in T2DM.

Table 1 Demographic and pathological characteristics of patients with T2DM and and healthy controls.
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Decreased PECAM-1 is an independent risk factor for vascular injury in T2DM

T2DM patients were classified into a vascular injury group and a non - vascular injury group according to the presence of vascular injury. TableĀ 2 compares the PECAM-1 levels and relevant clinical and biochemical data between the two groups. The results show that the HbA1c level in the vascular injury group is significantly higher than that in the non - vascular injury group, more patients in the vascular injury group are smokers, and the PECAM-1 level in the vascular injury group is markedly lower (Pā€‰<ā€‰0.05). Univariate analysis indicates that elevated HbA1c, smoking habit, and decreased PECAM-1 level affect vascular injury in T2DM patients. Further multivariate regression analysis reveals that HbA1c and PECAM-1 are independent risk factors for vascular injury in T2DM patients. Meanwhile, collinearity analysis and visualization were performed for the variables in TableĀ 2. By calculating the Pearson correlation coefficient matrix of all variables, results showed that the absolute values of correlations between variables were all below 0.3, indicating no significant collinearity among variables (Fig.Ā 1A). Additionally, Variance Inflation Factor (VIF) was calculated as a supplementary analysis, with all variables demonstrating VIF valuesā€‰<ā€‰2.5, further confirming the robustness of the model (Fig.Ā 1B).

Table 2 Multifactor logistic analysis for vascular injury in patients with T2DM.
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Fig. 1
figure 1

Collinearity analysis of variables.

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PECAM-1 transcription gene variation in T2DMs

Transcription factors play an important regulatory role in protein expression. In order to analyze the possible mechanism of PECAM-1 reduction in T2DM patients at the transcription level, we first detected the transcription factors involved in PECAM-1 transcription regulation. Based on the criteria of peak intensity signalā€‰<ā€‰500 and the predicted regulatory potential scoreā€‰<ā€‰1, the transcription factors involved in PECAM-1 regulation were analyzed through NetworkAnalyst. A total of 10 PECAM-1 regulating transcription factors, including responsive element binding protein 3 (CREB3), interferon Regulatory Factor 1 (IRF1), general transcription factor IIA subunit 2 (GTF2A2), nuclear receptor subfamily 2 group F member 6 (NR2F6), RELA proto-oncogene, NF-KB subunit (RELA), GATA zinc finger domain containing 1 (GATAD1), nuclear factor IA (NFIA ), REST corepressor 2 (RCOR2), cut like homeobox 1 (CUX1), and TEA domain transcription factor 3 (TEAD3), were found out (Fig.Ā 2A).

A mRNA dataset from the blood of T2DM patients (GSE26168; it includes the data from 9 T2DM patients and 8 normal controls) was obtained through the GEO public database. The results showed that comparing to the control group, the PECAM-1 transcription factors CREB3, GATAD1, and TEAD3 were significantly reduced (pā€‰<ā€‰0.01ā€“0.05) in T2DM, while CUX1 and RELA were significantly increased (pā€‰<ā€‰0.01) in T2DMs. There was no significant difference in other five transcription factors between the groups (Fig.Ā 2B). The results suggest that the variation of these transcription factors may be a regulatory mechanism for the decreased expression of PECAM-1 in T2DM plasma.

Fig. 2
figure 2

Changes in transcription factors of the PECAM-1 gene in T2DM patients.

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Differentially expressed genes in T2DM patients by GEO database analysis

Based on GSE26168 data analysis, a total of 100 significantly up-regulated genes (LogFC>1 ; p<0.05)(red) and 106 significantly down-regulated genes (LogFC<1 ; p<0.05) (blue) were obtained (Fig.Ā 3A). Among them, the top 20 significantly up-regulated genes (red) and the top 20 significantly down-regulated genes (green) were listed on a hot map (Fig.Ā 3B). OLFM4, CEACAM6 (CEA adhesion molecule 6), ITGB3 (integratin subunit beta 3), CEACAM8, and TMPRSS9 (transmembrane serine protease 9) genes which are related to vascular stability and permeability were significantly increased (LogFC>1; p<0.05). While, ARFIP2 (ADP ribosylation factor interacting protein 2) and TMCO5A (transmembrane and coiled-coil domains 5Ā A) were significantly reduced (LogFC<1; p<0.05). Noticeably, these significantly differential expressed proteins included those involving cellular adherent, cellular migration, cellular skeleton and membrane proteins, such as: CEACAM8 and 6, ITGB3, TMPRSS9, and ARFIP2. The results suggested that these abnormal expressed genes may involve the pathological mechanism of vascular injury in T2DMs.

Fig. 3
figure 3

Genetic difference analysis of T2DM patients based on the GEO database.

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Speculation of PECAM-1 bio-activities

The biological and pathological functional roles of PECAM-1 was analyzed using the STRING online network database. To easy understand the protein-protein interaction (PPI) related to PECAM-1, cytoscape was used to visualize the obtained PECAM-1-PPI proteins as a network diagram (Fig.Ā 4A). It revealed that kinase insert domain Receptor (KDR), cadherin 5 (CDH5), CDH17, selectin E (SELE), CD44, catenin beta 1 (CTNNB1), protein tyrosine phosphatase non-receptor type 1 (PTPN1), PTPN6, CD38, and CD177 tightly interact with PECAM-1, playing important roles in the regulation of vascular stability, endothelial development, and vascular permeability. The GO (Fig.Ā 4B) enrichment analysis was further performed using the ClusterProfiler package in R language. The results showed that PECAM-1 was involved in processes of plasma membrane adhesion, cell membrane adhesion via plasma membrane adhesion molecules, endometrial cell differentiation, endometrial development, peptidyl tyrosine modification, and cell cell junction organization, which closely relate to vascular permeability, endothelial development, endothelial cell functional information transmission, etc., including plasma-membrane adhesion, cell-cell adhesion via plasma-membrane adhesion molecules, endothelial cell differentiation, endothelium development, peptidylā€‰āˆ’ā€‰tyrosine phosphorylation, cellā€‰āˆ’ā€‰cell junction organization, basolateral plasma membrane, cadherin binding, etc. KEGG (Fig.Ā 4C) enrichment analysis also showed that PECAM-1 plays an important role in fluid shear stress and atherosclerosis, adherens junction, and cell adhesion molecules activities, all of them participate in vascular pathological and biological processes.

Fig. 4
figure 4

Proteins and functional pathways related to PECAM-1.

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PECAM-1 targeted drug speculation

Given the above data, PECAM-1 may play an important role in vascular injury in T2DM. In order to explore the therapeutic medicine for vascular injuries in clinical treatment of T2DM, we employed Drugbank (https://go.drugbank.com/) analysis, a database that combines bioinformatics and chemical informatics resource analysis, to analyze and retrieve targeted drugs related to PECAM-1. The results showed that fostamatinib can dock with PECAM-1 molecule. Utilizing the small molecule 3D structure of fostamatinib (Fig.Ā 5A and B) obtained from PubChem (https://pubchem.ncbi.nlm.nih.gov/) and the PECAM-1 3D structure (Fig.Ā 5B) obtained from PDB (https://www.rcsb.org/), the molecular docking of them was verified using AutoDockTools 1.5.7, by which he predicted binding energy between them was āˆ’ā€‰7.1 (kcal/mol). FigureĀ 5 showed a 3D visualized images of PECAM-1-fostamatinib docking using Discovery Studio 4.5. This result indicated that fostamatinib could be a potential candidate drug for the treatment of vascular injuries in T2DMs.

Fig. 5
figure 5

Fostamatinib can be used as a regulatory drug for PECAM-1.

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Discussion

In this study, we initially detected the levels of plasma PECAM-1 in individuals with T2DM and those of healthy controls. We discovered that plasma PECAM-1 was significantly reduced in T2DM patients. Subsequently, we investigated the transcription factors involved in the regulation of PECAM-1 transcription using NetworkAnalyst. By analyzing T2DMsā€™ blood mRNA data from the GEO database, we identified changes in certain PECAM-1 transcription factors and vascular regulatory factors in T2DM patients. Additionally, we explored the biological activity of PECAM-1 in vascular regulation using the STRING online network database. These findings and data indicated that PECAM-1 may have an important role in vascular injury in individuals with T2DM. Finally, we conducted a targeted drug matching analysis using the Drugbank database and found that fostamatinib may be a potential regulatory drug for PECAM-1, which could be applied in T2DMs with microvascular injuries.

Abnormal angiogenesis commonly occurs in almost all organs and tissues of either diabetic patients or experimental models. It includes the conditions such as diabetic retinopathy, nephropathy, microvascular injury to the axon of nerves, impaired wound healing, increased risk of heart disease, heart attacks, strokes, and narrowing of blood vessels. Recent research has shown that angiogenesis defects are also present in the lungs13, bone marrow14,15, and placenta16,17 of individuals with diabetes. Therefore, abnormal angiogenesis is a common feature of diabetic microvascular disease18,19,20. It is worth noting that abnormal angiogenesis is prevalent in diabetic complications, indeed, both excessive and defective angiogenesis can be observed in different tissues. Furthermore, diabetes is also associated with risks of coronary and peripheral artery diseases. Impaired angiogenesis can also affect the function of pancreatic islets and adipose tissue, worsening the damage caused by diabetes. The mechanisms of microvascular injury involve defective responses to hypoxia and pro-angiogenic factors, altered vasomotor tone, impaired shear stress sensing, inflammation, impaired nitric oxide bioavailability, lack of pro-angiogenic cells, and advanced glycation accumulation of end products, and pericyte loss18. Understanding the molecular factors that drive tissue-specific changes in diabetic microvascular injury is crucial for developing effective drugs, prevention strategies, and treatment methods for diabetic vascular injury.

PECAM-1 exists on the surface of platelets, monocytes, neutrophils and T cells, being an important protein in the inter-cellular connection of endothelial cells8,21.The adhesive properties of PECAM-1 largely depend on the Ī±2,3-sialylated glycans participating PECAM-1/PECAM-1 formation. It regulates the vascular permeability barrier and leukocyte trafficking7. PECAM-1 is an important factor for vascular integrity as well7,9. Disruption of vascular integrity will lead to the accumulation of plasma, proteins and cells into the interstitial space, which is a signs of inflammation9.Thus, reducing PECAM-1 will induce a series of vascular injuries. Anti-PECAM-1 antibodies inhibit the tubular structures formation on Matrigel, reduce tumor-induced angiogenesis in mice. Tumor angiogenesis is impaired in PECAM-1-deficient mice7,8,22. Injection of PECAM-1 monoclonal antibody into mice resulted in fluid leakage into the liver and renal vasculature7. Inhibition of PECAM-1 reduced atherosclerotic lesions in mice23.In addition, PECAM-1 can interfere with the normal vascular function through inflammatory regulation. The plasma membrane fragment of PECAM-1 contains serines and tyrosines residues, both of which are present within the immunoreceptor tyrosine inhibitory motif (ITIM). The ITIM of PECAM-1 recruits the Src homology 2 containing protein tyrosine phosphatase2 when phosphorylated, thereby forming the PECAM-1/SHP-2 complex. This complex inhibits platelet activation, cytokine production and reactivity and stimulating prostacyclin production in the vessel wall, therefore prohibiting the activation of neutrophils, monocytes, and leukocytes7.Reduction of PECAM-1 will weaken its inhibitory effect on the activation of platelets and leukemia, leading to the activation of platelets and leukemia, inducing intravascular plaques, thrombus, and endothelial cell inflammation, resulting in vascular injury9,24. In this study, we found for the first time that plasma PECAM-1 level was significantly reduced in T2DM patients. According to the above mechanism, decreased PECAM-1 will cause vascular injury, including vascular instability, angiogenesis disorders, increased vascular permeability, local inflammation, thrombosis, etc.

Five regulatory factors, including CREB3, CUX1, GATAD1, RELA, and TEAD3, respectively, were found to be significant changed compared to those of the normal controls (p<0.05) from a T2DM blood mRNA sequencing data obtained from GEO public database (GSE26168) (Fig.Ā 2). In 2021, Gu et al. found that lead compound AG1296 had a vascular protective effect by activating factors such as CREB3 in a network pharmacology study25.Dong et al. demonstrated that sodium formononectin-3ā€™-sulphonate (Sul-F) promotes angiogenesis through regulation of cytokines including CREB3 in their study on a pharmalogical mechanism of Sul-F26. These data indicate that CREB3 exerts vascular protective and pro - angiogenic effects through certain mechanisms. GATAD1 gene mutation on the 7q21 chromosome arm leads to dilated cardiomyopathy27. Ma et al. reported that a significant decrease in the levels of GETAD1 and its 3 (prime) region DNA methylation in the placenta are positively correlated with the occurrence of preeclamptic placentas28. These results suggest that GATAD1 is an important cytokine in smooth muscle and vascular regulation. In a 9p21.3 Coronary Artery Disease study, Almontashiri et al. found that knocking down TEAD3 gene expression inhibited the expression of p16 that is a risk factor of Coronary Artery Disease, indicating that TEDAD3 is also closely related to vascular smooth muscle growth29.Based on our analysis, CREB3, GATAD1, and TEAD3 may exert their pathological effects in T2DM via the regulation of PECAM-1 gene transcription. A significant reduction of CREB3, GATAD1 and TEAD3 in T2DM patients is not conducive to vascular protection. CUX1 has a dual nature in tumor growth, either promoting or inhibiting tumor growth activities. The detailed mechanism is still unclear30.In a study on the activity of dipalmitoylphosphatidic acid (DPPA), Chen et al. reported that the anti-angiogenic activity of DPPA is created by suppression of CUX1 that reduce fibroblast growth factor 1 (FGF1) transcription31. It is suggested that CUX1 has a promoting effect on angiogenesis. RELA is functional domain of nuclear factor kappa B (NF-ĪŗB). In terms of vascular pathology, NF-ĪŗB can promote the occurrence and progression of arteriosclerosis in various ways and participates in cell proliferation in multiple ways32.Combining with our experimental data, increased CUX1 and RELA levels in T2DMs may promote abnormal angiogenesis and atherosclerosis through PECAM-1 transcription regulation.

A T2DM sequencing data (GSE26168) provided from the GEO database showed that CEACAM8, CEACAM6, ITGB3, and TMPRSS9 were significantly increased in T2DM blood, while ARFIP2 and TMCO5A were significantly reduced (Fig.Ā 3B). Carcinoembryonic antigen (CEA) is identified as a member of the CD66 differentiation cluster in immunology. These proteins include CD66b (CEACAM8), CD66c (CEACAM6), etc33.CEA is usually produced in the gastrointestinal tissue during fetal development, however it stops producing before birth and is elevated in many tumors, thus, being used as a tumor marker clinically. CEA can trigger endothelial cell behavior to promote angiogenesis, including adhesion, diffusion, proliferation, and migration in vitro, as well as tumor microvascularization in vivo34. Although we have not found any research report on TMPRSS9, however, its family member, TMPRSS4, is closely related to the vascular density of hepatocellular carcinoma (HCC), which can promote angiogenesis and HCC growth35, suggesting that TMPRSS9 may also have a similar activity. The increase of CEACAM6, CEACAM8, and TMPRSS9 in T2DMs may involve the induction of abnormal angiogenesis similar to cancer tissue. ITGB3 is the main integrin on platelets and crucial for platelet aggregation. It primarily involved in hemostasis and thrombosis36. The increase of ITGB3 in T2DMs is also consistent with the atherosclerotic lesions in T2DM patients, which should attract the attention of researchers to further explore the mechanism of ITGB3 in T2DM vascular injury. In a study of intracranial aneurysm (IA), Qin et al. found that the circ-ARFIP2 levels were significantly reduced in IA patients. Cellular experiments have revealed that circ-ARFIP2 enhances the proliferation, migration, and invasion ability of smooth muscle cells by regulating miR-338-3p37.Reduction of ARFIP2 in T2DM patients will affect the complete formation of blood vessels. Combining the angiogenic effects caused by elevated levels of CEACAM6, CEACAM8, and TMPRSS9, as well as the dysfunction of smooth muscle cells caused by decreased levels of ARFIP2, the formed novel blood vessels in T2DM patients might be dysfunctional blood vessels.

Interestingly, all proteins (Fig.Ā 4A) associating to PECAM-1 obtained from the STRING online network database analysis are involved in angiogenesis and vascular function regulation. Among them, KDR is the receptor of VEGF (VEGFR-2), which mediates the regulation of endothelial cell function by VEGF38. CDH5 is the main endothelial adhesion molecule that controls cell connectivity and angiogenesis, which plays an important role in tumor vascular growth39. SEL and CD44 are involved in regulating inflammatory responses. Inflammation of the inner wall of blood vessels will increase vascular adhesion, leading to the accumulation of white blood cells, therefore, participating in the regulation of the pathogenesis of atherosclerosis40,41. CTNNB1 also takes part in inflammation and cellular oxidative responses, controlling various vascular regulatory factors42.CD38 participates in vascular remodeling via regulating vascular smooth muscle cells43. PTPN6 and PTPN11 are key regulatory components in cell cycle control, playing important roles in cell growth, proliferation, differentiation, and transformation control, including vascular endothelial cells44.CD177 also plays a positive role in tumor angiogenesis45. The above-mentioned and PECAM-1-involving vascular regulatory effects have also been validated in GO functional enrichment analysis. PECAM-1 participates in the following biological processes: plasma membrane adhesion, cell membrane adhesion via plasma membrane adhesion molecules, endometrial cell differentiation, endometrial development, peptidyl tyrosine modification, and cell cell junction organization (Fig.Ā 4B). Furthermore, the functional enrichment analysis of KEGG also showed that PECAM-1 is involved in fluid shear stress and atmospheric clearance, adhesion junction, and cell adhesion molecules (Fig.Ā 4C). Vascular endothelial injury and inflammation are central to the pathogenesis of vascular complications in T2DM, with PECAM-1 playing a critical role by regulating vascular barrier integrity, inflammatory cell migration, and endothelial mechanotransduction46. Previous studies have shown that PECAM-1 maintains endothelial barrier function by stabilizing adherens junctions, specifically through promoting the interaction between Ī²-catenin and VE-cadherin, and its loss significantly enhances endothelial permeability to albumin9. During inflammation, PECAM-1 collaborates with CD99 and JAM (junctional adhesion molecule) to mediate leukocyte transendothelial migration: JAM facilitates leukocyte adhesion to endothelial cells via LFA-1 (lymphocyte function-associated antigen 1), PECAM-1 mediates subsequent extravasation, and CD99 completes the migration cascade47. Dysregulation of this process exacerbates vascular wall inflammation and promotes atherosclerotic plaque formation. In the context of T2DM, reduced PECAM-1 expression impairs endothelial cell response to shear stress, a key mechanical signal in vascular homeostasis, thereby disrupting vascular integrity48. Collectively, these findings indicate that PECAM-1 deficiency in T2DM contributes to vascular injury by compromising the endothelial barrier, promoting inflammatory cell infiltration, and inducing aberrant angiogenesis. Future intervention strategies targeting PECAM-1 may provide new directions for the treatment of vascular complications in T2DM.

Fostamatinib (commercial names as Tavalisse and Tavlese) is a tyrosine kinase inhibitor drug used clinically to treat chronic immune thrombocytopenia (ITP)49. Our study revealed that fostamatinib has a strong affinity for PECAM-1 (Fig.Ā 5). Platelets are the main source of PECAM-1 production, indicating the important role of PECAM-1 in platelet function regulation7. Based on the logical idea of using fostamatinib in clinical ITP treatment to enhance platelets50, combined with the special affinity of fostamatinib to PECAM-1 discovered in this study, it is suggested that the mechanism of fostamatinib in treating ITP to enhance platelets may be mediated via PECAM-1 regulation. This viewpoint needs to be validated by pharmacological research in future. Ingo et al. demonstrated that oral fostamatinib reduces atherosclerotic lesion size in low-density lipoprotein receptor-deficient mice, accompanied by decreased macrophage infiltration, increased smooth muscle cells and collagen content, and stabilized plaques51. Notably, fostamatinib has shown safety in clinical trials for rheumatoid arthritis52, a chronic inflammatory disease, which aligns with its proposed role in modulating inflammation and vascular remodeling in T2DM. Given that T2DM is often associated with platelet hyperreactivity and endothelial dysfunction, fostamatinibā€™s dual inhibition of platelet activation and inflammatory pathways may confer pleiotropic benefits for microvascular protection. Combined with the important vascular regulation function of PECAM-1 described above, it is necessary to explore the possible therapeutic effect of fostamatinib in diabetes vascular injury, tumor, thrombocytopenia, atherosclerosis, and other vascular diseases in clinical practice. Meanwhile, this study also provides research ideas for the pharmacological mechanism of fostamatinib.

One limitation of this study is the lack of multivariate adjustment for common comorbidities (e.g., hypertension) and diabetes medications (e.g., insulin, statins, metformin). These factors may indirectly confound the results by influencing endothelial function, inflammatory responses, or PECAM-1 transcriptional regulation. Additionally, laboratory validation of key PECAM-1 signaling pathways was not performed, which limits the mechanistic understanding of our findings. Future studies should comprehensively collect data on comorbidities and medication use, coupled with experimental verification, to validate these results and further refine the mechanistic analysis.

Conclusion

The serum PECAM-1 levels in T2DM patients were significantly reduced compared with healthy controls. The decrease in transcription factors CREB3, TEAD3, and GATAD1, as well as the increase in RELA and CUX1, obtained from a T2DM blood test database, can explain the serum test findings. Further analysis of PECAM-1 functional enrichment revealed that PECAM-1 is involved in angiogenesis, vascular integrity, and endothelial functional regulation from multiple directions. The T2DM blood test data provided by GEO showed that the functional proteins CEACAM6/8, ITGB3, and TMPRSS9 related to PECAM-1 in the blood of T2DM patients were significantly increased, while ARFIP2 and TMCO5A were significantly reduced. These functional protein alterations may participate in the vascular injury mechanism of T2DM patients. Finally, network pharmacological analysis showed that fostamatinib is a drug with high affinity for PECAM-1. The treatment effect of fostamatinib for vascular injury complications in T2DM and other vascular injury diseases still requires further research and verification.