News Medical Life Sciences, September 3 , 2024
A combination of precise and efficient growth factors and cytokine detection is key to better understanding disease mechanisms, cellular processes, and drug efficacy throughout the rapidly evolving fields of stem cell and immunology research.
This article introduces high-quality, rapid, and dependable immunoassay tools from ACROBiosystems’ ClinMax™ product line and highlights their applications in mesenchymal stem cell (MSC) research and immune response studies.
The promise of mesenchymal stem cells
Stem cells hold immense therapeutic potential due to their self-renewal capacity and ability to differentiate into various cell types. Mesenchymal stem cells (MSCs) are multipotent stem cells derived from the mesoderm, exhibiting multiple differentiation potentials and high self-renewal capability.
These important members of the stem cell family are found in a range of tissues throughout the body, and MSCs can be cultured and expanded to differentiate into neuron cells, chondrocytes, muscle cells, osteoblasts, adipocytes, and other cell types depending on the conditions they are exposed to in vitro.
MSCs are regarded as an extremely promising tool in regenerative medicine, with 12 globally approved MSC-based stem cell drugs already available. These drugs’ therapeutic effects are primarily exerted via paracrine signaling, secreting various chemokines, cytokines, and growth factors.
Understanding and quantifying these secreted factors is key to assessing MSCs’ therapeutic potential and successfully optimizing their clinical applications.
Hepatocyte Growth Factor (HGF)
Hepatocyte Growth Factor (HGF) plays a multifaceted role in stem cell paracrine signaling. One of the key factors secreted by MSCs, HGF exhibits potent immunoregulatory properties due to its capacity to inhibit effector T cell proliferation. This, in turn, prompts the downregulation of cyclin D2 and the upregulation of the p27kip1 protein, eventually leading to cell cycle arrest at the G1 phase.
In terms of angiogenesis, HGF stimulates the formation of new blood vessels by promoting the migration and proliferation of smooth muscle and vascular endothelial cells. It also facilitates ischemic blood flow recovery and vascular network remodeling.
HGF also demonstrates anti-apoptotic effects on several different cell types, including cardiomyocytes, renal tubular cells, endothelial cells, and hepatocytes. It also inhibits TGF-β1 expression to help counteract fibrosis.
Vascular Endothelial Growth Factor (VEGF)
Vascular Endothelial Growth Factor (VEGF) is also a key factor secreted by stem cells. Most notably, VEGF stimulates endothelial cell proliferation and survival, promoting vascular network remodeling, angiogenesis, and tissue regeneration.
VEGF secretion is a key indicator of stem cells’ regenerative capacity, as well as taking part in a range of anti-apoptotic processes. For example, researchers discovered that VEGF gene-modified MSCs significantly inhibited apoptosis and promoted normal cell proliferation in an acute kidney injury model, achieving this by enhancing microcirculation.
Interleukin-6 (IL-6)
MSCs secrete interleukin-6 (IL-6), which has the potential to inhibit inflammation and exert immunoregulatory effects. The majority of studies have confirmed that MSCs favorably regulate neutrophils. By secreting IL-6, MSCs can significantly inhibit neutrophil apoptosis, even at low MSC/neutrophil ratios.
Other secreted growth factors and cytokines
Stem cells can secrete various growth factors and cytokines, such as:
- Angiopoietin 1/2 (Ang 1/2)
- Epidermal Growth Factor (EGF)
- Fibroblast Growth Factor (FGF)
- Growth Hormone (GH)
- Insulin-Like Growth Factor (IGF)
- Interleukin-10 (IL-10)
- Monocyte Chemoattractant Protein-1 (MCP-1)
- Placental Growth Factor (PGF)
- Platelet-Derived Growth Factor (PDGF)
- Stem Cell Factor (SCF)
- Transforming Growth Factor (TGF-β1)
Bioactive substances like these are produced by stem cells and can function synergistically or antagonistically. They can also participate in inflammation response, immune response regulation, and tissue injury repair processes.
Rapid and fully validated cytokine detection for immune response research
The body recognizes and eliminates pathogens or abnormal cells via its immune response, and cytokines play a vital role in regulating and mobilizing this process.
Immune cells (such as B cells, T cells, and macrophages) and other cells (such as endothelial and epithelial cells) release cytokines when stimulated by external factors, enabling them to modulate the intensity and direction of the immune response.
Cytokines enable communication between immune cells, encouraging their proliferation, differentiation, and functional activation.
For example, interleukin-2 (IL-2) improves T cell proliferation and activation to improve the robustness of cell-mediated immune responses, while interferons (IFNs) augment antiviral immunity. Other cytokines regulate immune cell chemotaxis, directing cells to infection or injury sites to support localized immune defense.
Inappropriate or excessive cytokine release can prompt abnormal immune responses, potentially instigating inflammatory or autoimmune diseases. This risk emphasizes the vital role of balanced cytokine regulation in immune responses.
Appropriate cytokine use can improve immune defense while circumventing the risks of immune damage. Cytokine detection is, therefore, essential in the evaluation of immune responses.
Numerous cytokines, including interleukins (e.g., IL-1β, IL-2, IL-6, IL-10), tumor necrosis factors (TNFs), and IFNs, can be measured in body fluids, blood, and cell culture supernatants.
These measurements reflect the immune response’s activity level and the immune system’s current state, offering vital information on effective disease diagnosis, treatment, and monitoring.
Cytokine detection is frequently employed in exploring disease mechanisms, evaluating drug efficacy, and developing new therapies in the fields of antibody drug development, cell therapies, and other therapeutics.
Th1/Th2 cytokine profiling: flow cytometry multiplex bead assay
It is possible to categorize T helper (Th) cells into subtypes based on their cytokine secretion profiles, for example, Th1 and Th2.
Th cells rarely differentiate into Th1 or Th2 cells under normal physiological conditions, but their differentiation capacity significantly increases when stimulated by specific antigens, such as stimulants or pathogens.
Th1 and Th2 cells have distinct roles in the immune system and disease processes. Th1 cells primarily stimulate and mediate cellular immunity, macrophage activation, cytotoxic T lymphocyte activity, and delayed-type hypersensitivity.
Th2 cells, on the other hand, primarily mediate humoral immunity, with IL-4 secretion responsible for promoting B cell proliferation, differentiation, and antibody production.
Under normal conditions, Th1 and Th2 cells are in a state of homeostasis. A disruption of this balance is known as ‘Th1/Th2 imbalance’ and can potentially result in various conditions, including allergies, tumors, and autoimmune diseases.
Studying the cytokines secreted by Th1/Th2 cells enables a better understanding of the balance shift occurring. This allows for targeted regulation of Th1 or Th2 levels to restore balance and achieve therapeutic effects.
FDA regulations mandate pharmacological cytokine detection and the strict evaluation of immunomodulatory drugs’ potential to trigger cytokine release syndrome (CRS).
The FDA’s 2020 guidance on immunogenicity studies states that conducting in vitro cytokine release assays to assess the potential for CRS is vital, especially for biological therapeutics.
The guidance confirms that assays should be undertaken using human cells to evaluate immune activation and cytokine release. The studies should also focus on key cytokines such as IL-2, IL-6, IL-10, IFN-γ, and TNF-α.
Clinical safety considerations should be reported via these test results, including the selection of starting doses and appropriate monitoring strategies used during clinical trials.
It can be difficult to adequately predict CRS risk via conventional in vivo toxicity studies, prompting the regulatory framework to emphasize the importance of in vitro assessments.
The detection of cytokines secreted by Th1/Th2 cells is also essential. Cytokine levels in normal serum and cell culture supernatants are low and present specific detection challenges, with traditional ELISA methods often proving inadequate for simultaneous multi-cytokine detection.
A combination of the development of flow cytometry and solid-phase technology and the advent of bead-based multiplex detection techniques has afforded flow cytometry the capacity to analyze soluble proteins both qualitatively and quantitatively.
Conclusion
ACROBiosystems’ powerful and reliable ClinMax™ Cytokine Detection Tools are ideally placed to support stem cell and immunology research advances.
These tools boast the specificity, sensitivity, and accuracy required for precise and rapid cytokine detection, allowing researchers to gain an in-depth understanding of immune responses, cellular processes, and potential therapeutic interventions.
A comprehensive portfolio of high-quality ClinMax™ Cytokine ELISA kits, all of which have been validated for stem cell and immunology research, is available to support drug development efforts.
The ClinMax™ Th1/Th2 Cytokine Multiplex Bead Assay Kit has also been validated and developed via flow cytometry-based multiplex bead assay technology. This kit uses a single assay to quantitatively detect IL-2, IL-4, IL-6, IFN-γ, TNF-α, and IL-10 in cell culture media and serum, significantly improving research efficiency and enabling multiplex analysis of valuable samples.
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