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Immunoproteomics approach revealed elevated autoantibody levels against ANXA1 in early stage gallbladder carcinoma.

ABSTRACT: Background:Early diagnosis is important for the timely treatment of gallbladder carcinoma (GBC) patients and may lead to increased survival outcomes. Here, we have applied serological proteome analysis (SERPA), an immunoproteomics approach, for the detection of 'tumor-associated antigens (TAAs) that elicit humoral response' in early stage GBC patients.

Methods:Total protein from pooled tumor tissue of GBC patients (n?=?7) was resolved by two-dimensional gel electrophoresis (2-DE) followed by immunoblotting using pooled blood plasma from healthy volunteers (n?=?11) or gallstone disease (GSD) cases (n?=?11) or early stage GBC (Stage I and II) (n?=?5) or GBC stage IIIA (n?=?9). 2-D gel and immunoblot images were acquired and analyzed using PDQuest software to identify immunoreactive spots in GBC cases in comparison to controls. Proteins from immunoreactive spots were identified by liquid chromatography- tandem mass spectrometric analysis (LC-MS/MS). Autoantibody levels for two of the functionally relevant proteins were investigated in individual plasma samples (52 cases and 89 controls) by dot blot assay using recombinant proteins.

Results:Image analysis using PDQuest software identified 25 protein spots with significantly high or specific immunoreactivity in GBC cases. Mass spectrometric analysis of 8 corresponding protein spots showing intense immunoreactivity (based on densitometric analysis) in early stage GBC or GBC stage IIIA cases led to the identification of 27 proteins. Some of the identified proteins include ANXA1, HSPD1, CA1, CA2, ALDOA and CTSD. Among the two proteins, namely ANXA1 and HSPD1 verified using a cohort of samples, significantly elevated autoantibody levels against ANXA1 were observed in early stage GBC cases in comparison to healthy volunteers or GSD cases (unpaired t-test, p?
Conclusions:The study suggests that the ANXA1 autoantibody levels against ANXA1 may be potentially employed for early stage detection of GBC patients. Other proteins could also be explored and verified in a large cohort of clinical samples.

SUBMITTER: Akhtar J

PROVIDER: S-EPMC7709428 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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Immunoproteomics approach revealed elevated autoantibody levels against ANXA1 in early stage gallbladder carcinoma.

Akhtar Javed J Priya Ratna R Jain Vaishali V Sakhuja Puja P Agarwal Anil Kumar AK Goyal Surbhi S Polisetty Ravindra Varma RV Sirdeshmukh Ravi R Kar Sudeshna S Gautam Poonam P

BMC cancer 20201201 1

<h4>Background</h4>Early diagnosis is important for the timely treatment of gallbladder carcinoma (GBC) patients and may lead to increased survival outcomes. Here, we have applied serological proteome analysis (SERPA), an immunoproteomics approach, for the detection of 'tumor-associated antigens (TAAs) that elicit humoral response' in early stage GBC patients.<h4>Methods</h4>Total protein from pooled tumor tissue of GBC patients (n = 7) was resolved by two-dimensional gel electrophoresis (2-DE) ...[more]

PMID: 33261560

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Project description:Objective:Beta 1-adrenergic receptor autoantibodies (?1ARAbs) have been identified as a pathogenic factor in atrial fibrillation (AF), but the underlying pathogenetic mechanism is not well understood. We assessed the hypothesis that elevated ?1ARAb levels increase AF susceptibility by promoting atrial fibrosis. Methods:A total of 70 patients with paroxysmal AF were continuously recruited. The serum levels of ?1ARAb and circulating fibrosis biomarkers were analyzed by ELISA. Linear regression was used to examine the correlations of ?1ARAb levels with left atrial diameter (LAD) and circulating fibrosis biomarker levels. Furthermore, we established a rabbit ?1ARAb overexpression model. We conducted electrophysiological studies and multielectrode array recordings to evaluate the atrial effective refractory period (AERP), AF inducibility and electrical conduction. AF was defined as irregular, rapid atrial beats > 500 bpm for > 1000 ms. Echocardiography, hematoxylin and eosin staining, Masson's trichrome staining, and picrosirius red staining were performed to evaluate changes in atrial structure and detect fibrosis. Western blotting and PCR were used to detect alterations in the protein and mRNA expression of TGF-?1, collagen I and collagen III. Results:Patients with a LAD ? 40 mm had higher ?1ARAb levels than patients with a smaller LAD (8.87 ± 3.16 vs. 6.75 ± 1.34 ng/mL, P = 0.005). ?1ARAb levels were positively correlated with LAD and circulating biomarker levels (all P < 0.05). Compared with the control group, the rabbits in the immune group showed the following: (1) enhanced heart rate, shortened AERP (70.00 ± 5.49 vs. 96.46 ± 3.27 ms, P < 0.001), increased AF inducibility (55% vs. 0%, P < 0.001), decreased conduction velocity and increased conduction heterogeneity; (2) enlarged LAD and elevated systolic dysfunction; (3) significant fibrosis in the left atrium identified by Masson's trichrome staining (15.17 ± 3.46 vs. 4.92 ± 1.72%, P < 0.001) and picrosirius red staining (16.76 ± 6.40 vs. 4.85 ± 0.40%, P < 0.001); and (4) increased expression levels of TGF-?1, collagen I and collagen III. Conclusion:Our clinical and experiential studies showed that ?1ARAbs participate in the development of AF and that the potential mechanism is related to the promotion of atrial fibrosis.

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Immunoproteomics approach revealed elevated autoantibody levels against ANXA1 in early stage gallbladder carcinoma.

Publications

Immunoproteomics approach revealed elevated autoantibody levels against ANXA1 in early stage gallbladder carcinoma.

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