CD4 Human (203-317)

What is CD4 Human (203-317) and what are its key structural properties?

CD4 Human (203-317) is a recombinant protein fragment encoding amino acids 203-317 of the human CD4 molecule. This specific domain contains important binding regions critical for immune cell interactions. CD4 is also known by several synonyms including gp55, HLA-2, L3/T4, Ly-4, T-cell antigen T4/LEU3, and sCD4 .

The recombinant version typically includes a His-tag to facilitate purification and detection in experimental settings. The protein fragment maintains specific binding capabilities while offering advantages in stability and experimental flexibility compared to the full-length protein .

How should researchers properly reconstitute and store CD4 Human (203-317)?

For optimal reconstitution, it is recommended to perform a quick spin on the vial followed by reconstitution in sterile water to achieve a concentration of at least 100 μg/ml. This solution can then be further diluted in other aqueous solutions as needed for specific experimental applications. Importantly, researchers should avoid vortexing the solution during reconstitution to prevent protein denaturation .

For storage, the protein should be kept at temperatures between -20°C and -80°C. When maintained under these recommended conditions, the protein remains stable for approximately 12 months. To preserve protein integrity, researchers should minimize freeze-thaw cycles as these can compromise structural stability and biological activity .

What techniques are most effective for studying CD4+ T cell subsets?

High-throughput approaches to profile αβ TCR repertoires have proven particularly effective for studying CD4+ T cell subset differentiation and relationships. These techniques allow researchers to track clonal lineages across naive and effector/memory CD4+ T cell subsets regardless of antigen specificity .

Polychromatic flow cytometry represents another essential methodology, enabling the identification and sorting of functionally distinct CD4+ T cell subsets (including Tfh, Th1, Th1-17, Th17, Th22, Th2a, Th2, and Treg) from peripheral blood. This approach typically uses multiple surface markers in a hierarchical gating strategy to isolate pure populations for downstream analysis .

For transcriptional profiling, single-cell RNA sequencing has proven invaluable, revealing that CD4+ T cells exist along a transcriptional continuum rather than as discrete populations. This technology can identify gene expression signatures associated with different stages of T cell differentiation and activation states .

How can researchers detect and measure HGF production in CD4+ T cells?

Detection of hepatocyte growth factor (HGF) in CD4+ T cells presents technical challenges due to its relatively low expression levels. Standard ELISAs often lack sufficient sensitivity, with values typically falling below detection limits. Researchers have developed specialized HGF ELISpot assays that offer enhanced sensitivity for comparing HGF production across different in vitro differentiated CD4+ T cell subsets .

Flow cytometry methods using anti-HGF antibodies have proven challenging for detecting surface-bound or intracellular HGF protein, despite attempts with various antibody clones, host species, and staining protocols. For transcriptional analysis, nested amplification methods specifically targeting HGF mRNA can be employed to detect HGF-positive cells, overcoming the limitations of standard scRNA-seq approaches for detecting low-abundance transcripts .

Surface marker enrichment strategies can enhance detection efficiency, with CD30 identified as a marker that can be used to enrich for HGF-positive cells in the CD4+ T cell population .

What is the current understanding of the relationship between naive and memory CD4+ T cell subsets?

Recent research has revealed that CD4+ T cells form a transcriptional continuum rather than existing as discrete populations. This continuum progresses from naive to central memory and finally to effector memory phenotypes, forming what researchers describe as an "effectorness gradient." This gradient is characterized by progressive increases in the expression of chemokines and cytokines as cells acquire more effector-like properties .

The differentiation pathways of CD4+ T cells are more complex than previously understood. Memory T cells exhibit different cytokine response patterns compared to naive cells, with memory cells showing limited capacity to differentiate into certain phenotypes. For example, memory CD4+ T cells demonstrate an inability to differentiate into the Th2 phenotype, while unexpectedly acquiring Th17-like characteristics when exposed to iTreg polarizing conditions .

This evolving model challenges the traditional view of stable CD4+ T cell fates and highlights the plasticity of memory T cell responses, which has significant implications for understanding inflammatory processes and immune regulation .

How do TCR repertoire characteristics influence CD4+ T cell subset fate decisions?

The T-cell receptor (TCR) repertoire exhibits distinct physicochemical and recombinatorial features that are encoded on a subset-specific basis within the effector/memory compartment. High-throughput TCR profiling has revealed that each effector/memory CD4+ T cell subset possesses characteristic repertoire features that are consistently observed across genetically unrelated donors, suggesting predetermined fate biases at the level of the somatically rearranged TCR .

Interestingly, some of these TCR characteristics are already present in the corresponding naive repertoires, most notably in regulatory T cells (Tregs). This indicates that certain CD4+ T cell fates may be influenced by TCR structural properties even before antigen encounter .

Clonal tracking has identified both forbidden and permitted transition pathways between subsets, effectively mapping relationships among effector/memory subsets based on interconversion potential or shared ontogeny. Public TCR sequences (those shared across individuals) tend to be confined to particular effector/memory subsets, suggesting conserved antigen recognition properties within functional CD4+ T cell populations .

What are the biological applications of CD4 in HIV research and therapeutic development?

CD4 plays a crucial role in HIV research, particularly for studying viral entry mechanisms and developing potential therapeutics. By modifying the HIV-1 envelope protein structure to enhance binding with rhesus CD4 (rhCD4), researchers have improved the replication capacity of simian human immunodeficiency virus (SHIV) in Rhesus monkey lymphocytes. This advancement has important implications for studying HIV-1 transmission dynamics and facilitating vaccine testing in non-human primate models .

The CD4 protein is also being utilized in immunotherapeutic approaches, particularly in activating CD4+ T cells to enhance anti-tumor immune responses. CD4+ T cells not only support CD8+ T cell cytotoxic functions but can also directly kill tumor cells, providing novel targets and strategies for cancer immunotherapy development .

How does CD4 actively regulate T-cell receptor signaling?

Recent research has overturned the historical view that CD4 serves merely a supporting role in T-cell functions. Studies now demonstrate that CD4 plays an active and regulatory role in T-cell receptor (TCR) signaling pathways. Rather than functioning as a passive co-receptor, CD4 actively modulates signal strength and specificity, influencing downstream activation events and subsequent T cell responses .

This regulatory function likely involves specific protein-protein interactions between CD4 and components of the TCR signaling complex, as well as effects on membrane organization and immune synapse formation. Understanding these interactions provides important insights into fundamental immunological processes and potential targets for therapeutic intervention in conditions involving dysregulated T cell activation .

What are the primary CD4-related diseases and their underlying mechanisms?

CD4-related diseases encompass a spectrum of conditions affecting the immune system, particularly involving CD4+ T cells. HIV/AIDS represents the most prominent CD4-related disease, where the human immunodeficiency virus selectively targets and destroys CD4+ T cells, leading to progressive immune system deterioration. The mechanistic basis involves viral binding to CD4, entry into cells, and subsequent depletion of these critical immune regulators .

Autoimmune disorders including lupus and rheumatoid arthritis also involve CD4+ T cell dysfunction, though through different mechanisms. In these conditions, dysregulated CD4+ T cell responses lead to inappropriate immune activation against self-tissues. This can occur through breaks in central or peripheral tolerance mechanisms, altered TCR signaling thresholds, or aberrant cytokine production patterns .

Treatment approaches for CD4-related diseases focus on either restoring CD4+ T cell numbers and function (as in HIV/AIDS) or modulating inappropriate CD4+ T cell responses (as in autoimmune conditions). Understanding the specific CD4+ T cell subsets and their dysregulation in different disease contexts remains an active area of research with therapeutic implications .

How does the heterogeneity of CD4+ T cell responses influence inflammation and disease progression?

The heterogeneity of CD4+ T cell responses significantly impacts inflammation and disease progression through multiple mechanisms. The effectorness gradient observed in CD4+ T cells directly influences activation thresholds and cytokine response patterns, with cells further along this gradient demonstrating enhanced inflammatory potential .

Memory CD4+ T cells show distinct cytokine response profiles compared to naive cells, which has important implications for secondary immune responses and chronic inflammatory conditions. Their inability to differentiate into certain phenotypes (like Th2) while readily acquiring others (like Th17-like characteristics) can skew immune responses in ways that either promote resolution or exacerbate pathology .

The plasticity of memory CD4+ T cells, particularly their ability to be reprogrammed by the cytokine microenvironment, means that previously committed cells can adopt different functional profiles depending on local tissue conditions. This plasticity contributes to the dynamic nature of inflammatory responses and presents both challenges and opportunities for therapeutic intervention in inflammatory diseases .