Characteristics of Diseased Peripheral Blood Mononuclear Cells (PBMCs) and their Applications

Brief Introduction to PBMCs

Peripheral Blood Mononuclear Cells (PBMC), also called Human Mononuclear Cells from Peripheral Blood (HMNC-PB), are cells with a single, round nucleus that are collected from peripheral or circulating blood. These blood cells are a key component in the immune system to fight infections and disease, adapting to invaders. PBMCs respond selectively to the immune system and are a key component of human immunity. PBMCs are a mixed population of monocytes composed primarily of lymphocytes, including T cells, B cells, and natural killer (NK) cells.

PMBCs are also a rich source of monocytes, such as macrophages, dendritic cells. Further cell isolations are performed by selecting key protein surface markers such as CD4⁺ (T helper cells), CD8⁺ (cytotoxic T cells), CD14⁺ (monocytes), etc.

PBMCs undergo characteristic changes in various disease states and represent a way to monitor disease progression and understand disease pathology. For example, using transcriptomics to analyze the entire set of RNA transcripts produced by a cell or cell population, examining changes in PBMC gene expression allows researchers to better understand how disease pathology is triggered and which specific subsets of immune cells may be involved. A better understanding of autoimmune changes in specific diseases could also help scientists identify genes and proteins that could be targeted for therapeutic effects. A variety of diseased PBMCs have been widely used in scientific research, such as Human non-Small Cell Lung Carcinoma (NSCLC) Peripheral Blood Mononuclear Cells (PBMCs), Human Multiple Sclerosis PBMCs, Celiac Disease PBMCs, Human Systemic Lupus Erythematosus (SLE) PBMCs, Type-1 Diabetes PBMCs, and Human Ankylosing Spondylitis PBMCs, etc ^[1].

Cell Culture Protocol

Take Human non-Small Cell Lung Carcinoma PBMC as an example to introduce the culture method of diseased PBMCs.

How to isolate PBMCs from NSCLC Patients?

Peripheral blood was obtained from responsible institutions with many years of experience in sample collection. The blood is quickly transported to laboratory for processing.

Once in the laboratory, PBMCs from non-small cell lung carcinoma patients are isolated by density centrifugation with using a high molecular weight sucrose solution, Ficoll, to remove red blood cells under sterile conditions. Depending on the species of blood (human or non-human), use 1.077 or 1.084 g/ml Ficoll to form a perfect mononuclear cell layer separated from plasma, platelets, granulocytes and red blood cells. Take additional quality control steps to isolate and prepare PBMCs to ensure maximum viability for cryopreservation and downstream experimental applications.

Cryopreservation and Storage

PBMCs are in many ways ideal starting materials for studying the pathology of autoimmune diseases; they are an important part of the immune system, can be used to isolate a variety of immune cell types, and are relatively easy to obtain. The application of PBMCs is an ideal method to study disease pathology and monitor disease progression, as it undergoes characteristic changes in various disease states.

For example, some researchers have applied celiac disease PBMCs cells to study the effect of in vivo gluten challenge on the gene expression profile of PBMCs in diet-treated celiac disease (CD). The project is designed to research the molecular effects of brief exposure of gluten in GFD-treated CD patients and to study the biological pathways maintain constitutive changes in celiac disease despite of treatments. The researchers explored the peripheral transcriptome of CD patients after short gluten exposure by RNAseq analysis of PBMC samples (collected from treated CD groups， gluten-challenged groups and healthy controls). Surprisingly, gluten exposure altered genome-wide PBMC transcriptome of the patient after only 3 days. In the study of related mechanisms, pathway analysis showed upregulation of crucial immune response-related pathways induced by gluten in CD patients, covering innate and adaptive. They assessed perturbation of biological pathways in a sample-specific way.

The results suggested that related pathways associated with tight junctions, olfactory transduction, unsaturated fatty acid (e.g. arachidonic acid) metabolism, amino acid metabolism (e.g. cysteine and glutamate), and microbial infection were constitutively changed in CD groups exposed to gluten treatment compared with healthy controls,  whereas GFD treatment tended to normalize immune response pathways to a health condition. Besides, upstream regulator prediction analysis using differentially expressed genes manifested constitutively activated regulators relatively close to previously reported CD-related loci, especially SMARCA4 on 19p13.2 and CSF2 on 5q31. In a nutshell, RNAseq revealed obvious effects of short-term oral gluten challenge on whole PBMC fractions, and constitutively altered pathways in CD PBMCs, suggesting key factors other than gluten in CD pathogenesis ^[2].

Lupus is an autoimmune disease affecting multiple systems. Likewise, the researchers used SLE (lupus) PBMC cells to analyze about 276,000 peripheral blood mononuclear cells from 33 SLE children with varying degrees of disease activity and 11 matched controls also by single-cell RNA sequencing. It laid the foundation for addressing the origin of the SLE transcriptional signatures and disease heterogeneity for precision medicine applications that systematic mapping of lupus erythematosus heterogeneity at the horizontal level ^[3].

Recently, researchers also explored the differential expression of microRNAs in PBMCs of multiple sclerosis (MS) patients. They aimed to analyze the role of miRNAs in MS especially the relapse stage, which highlighted the significance of miRNA expression in molecular mechanisms involved in MS. Furthermore, the study that these small molecules are involved in MS opened up a new ground to explore candidate biomarker targets ^[4].

Immunological research of disease

PBMCs from NSCLC patients can be used in many immunology-based applications like in vitro applications to characterize immune cell populations based on immunophenotyping. These PBMCs were also an ideal source of effector cells for functional experiments such as cytokine-producing effector cells to determine whether the immune function of lymphocytes in NSCLC patients is altered. To determine the presence of NSCLC-driven alterations, specific immune cell types could also be isolated from NSCLC patients’ PBMC for genomic or transcriptome sequencing ^[5].

Conclusion

In short, different diseased PBMCs were used for a wide variety of immunology-based applications, including in vitro immunoassays, population characterization, sequencing studies, and biologic testing as above.

However, in the application process, attention should be paid to the quality stability, ethical issues of traceability, whether the cells are fresh, the selection of appropriate cells for different studies, etc.

Where to Get diseased PBMCs for research?

PBMCs give selective responses to the immune system and are the major cells in the human body immunity. AcceGen provides Diseased Human Peripheral Blood Mononuclear Cells for research use.

Here are our full list of Diseased PBMCs products:

It is our pleasure to help relative researches to move forward. All the products of AcceGen are strictly comply with international standards. For more detailed information, please visit our product portfolio or contact inquiry@accegen.com.

Reference

[1] Cojocaru M., et al. Multiple autoimmune syndrome. Maedica (Bucur). 5(2): 132–134. Apr 2010.

[2] Yohannes D A, Kauwe A D,  Kaukinen K, et al. Effects of In Vivo Gluten Challenge on PBMC Gene Expression Profiles in Diet Treated Celiac Disease[J]. Frontiers in Immunology, 2020, 11.

[3] Nehar-Belaid D, Hong S, R Marches, et al. Mapping systemic lupus erythematosus heterogeneity at the single-cell level[J]. Nature Immunology, 2020, 21(9).

[4] Otaegui D, Baranzini S E, Armananzas R, et al. Differential micro RNA expression in PBMC from multiple sclerosis patients[J]. PLoS One, 2009, 2009, 4(7).

[5] Chang D H, Rutledge J R, Patel A A, et al. The Effect of Lung Cancer on Cytokine Expression in Peripheral Blood Mononuclear Cells[J]. Plos One, 2013, 8.