TNF a Rabbit
Description
Meningitis and Blood-Brain Barrier Dysfunction
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Mycobacterial Infections: Rabbits infected with TNF-alpha-secreting recombinant BCG developed severe meningitis due to blood-brain barrier (BBB) disruption. TNF-alpha levels >10,000 pg/mL in cerebrospinal fluid (CSF) correlated with leukocytosis and brain edema .
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Vascular Effects: TNF-alpha induces procoagulant activity and nitric oxide synthase in cerebral vasculature, leading to thrombosis and reduced cerebral blood flow .
Tuberculosis Granuloma Formation
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Etanercept Impact: Blocking TNF-alpha with etanercept (TNFR2-Fc) in Mycobacterium tuberculosis-infected rabbits exacerbated lung pathology, reduced collagen deposition in granulomas, and impaired bacterial control .
Antibodies and ELISA Kits
Table 2: Antibodies for Rabbit TNF-alpha Detection
Neutralizing Antibodies
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A rabbit polyclonal antibody against human TNF-alpha (Genzyme) neutralized cytotoxicity in bioassays, confirming TNF-alpha’s role in CHF pathophysiology .
Recombinant TNF-alpha Effects
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Leukocytosis Induction: Intrathecal injection of 4 µg murine TNF-alpha in rabbits caused CSF leukocytosis (2,095 ± 1,900 cells/mm³), mimicking endogenous TNF-alpha activity .
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Apoptosis and Inflammation: TNF-alpha regulates lymphoid tissue development and promotes endothelial activation, contributing to diseases like rheumatoid arthritis and insulin resistance .
Therapeutic and Pathological Implications
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Dual Role: While essential for controlling infections, dysregulated TNF-alpha exacerbates tissue damage. Neutralizing TNF-alpha in rabbits impaired granuloma integrity, increasing Mtb dissemination .
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Biomarker Potential: Serum soluble TNF receptors (sTNF-Rs) modulate TNF-alpha activity, with elevated levels linked to worsened heart failure outcomes in clinical models .
Product Specs
The ED50, which represents the effective concentration required to achieve 50% of the maximum response, is determined by measuring the cytolysis (cell death) of murine L929 cells in the presence of Actinomycin D. The ED50 is less than 0.03ng/ml. This corresponds to a Specific Activity of 30,000,000 IU/mg.
Tumor necrosis factor, Cachectin, TNF-alpha, Tumor necrosis factor ligand superfamily member 2, TNF-a, TNF, TNFA, TNFSF2.
The sequence of the first five N-terminal amino acids was determined and was found to be Met-Ser-Ala-Ser-Arg.
Q&A
What is TNF and what are its primary functions in rabbit models?
TNF is a polypeptide hormone produced primarily by activated macrophages in rabbits. It exists in two main forms: TNF-α (cachectin) and TNF-β (lymphotoxin-alpha). In rabbit models, TNF functions as a key mediator of inflammation, immune response, and cell death regulation . It binds to specific receptors (TNFRSF1A/TNFR1 and TNFRSF1B/TNFBR) to exert biological effects including inducing cell death in tumor cell lines, stimulating fever through direct action or interleukin-1 secretion induction, and mediating glomerular damage .
Rabbit studies have demonstrated that TNF plays pivotal roles in:
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Endotoxin-induced glomerular damage
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Cytokine network regulation in inflammatory conditions
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Mediating macrophage-dependent damage in glomerulonephritis
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Anti-tumor responses in BCG-primed animals
How does TNF production differ between normal and BCG-primed rabbits?
TNF production exhibits significant differences between normal and BCG-primed rabbits:
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Normal rabbits produce negligible amounts of TNF after endotoxin injection in vivo, though their monocytes can produce TNF after endotoxin challenge in vitro
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BCG-primed rabbits release substantial TNF into serum after intravenous endotoxin injection
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BCG-injected rabbits contain more mononuclear phagocytes than normal rabbits, and these cells demonstrate enhanced capacity for TNF production
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Calculations indicate that BCG-primed rabbits produce over 20 times more TNF than normal rabbits after endotoxin challenge, assuming production primarily from lungs, blood, spleen, and liver
This differential response makes BCG-primed rabbits particularly valuable for studying TNF-mediated pathologies and therapeutic interventions.
What are the main cellular sources of TNF in rabbit models?
Research has identified a hierarchy of TNF-producing cells in rabbits, with production capacity following this order (from highest to lowest):
The liver is also recognized as an important site of TNF synthesis. TNF is newly synthesized by these cells, primarily within the first 7 hours of culture after stimulation. Interestingly, the TNF produced has similar biochemical properties (gel-filtration and ion exchange behavior) regardless of its cellular source .
What dose-dependent effects does TNF induce in rabbit kidney models?
TNF administration in rabbits produces dose-dependent effects on renal function and structure:
| TNF Dose (μg/kg/h) | Hematological Changes | Renal Effects | Histological Findings |
|---|---|---|---|
| 0.08 (Low) | Minimal changes | No significant renal dysfunction | Occasional PMNs in glomerular capillaries |
| 0.8 (Medium) | Anemia (15% Ht decrease) Leukopenia (47% decrease) Thrombocytopenia (45% decrease) | Mild alterations | Significant inflammatory cell presence in glomerular capillaries |
| 8.0 (High) | Anemia (16% Ht decrease) Leukopenia (59% decrease) Thrombocytopenia (57% decrease) | Renal failure (serum creatinine increase from 1.02±0.15 to 1.64±0.34 mg/dl) | Extensive inflammatory cell accumulation Fibrin deposition in glomerular capillaries (25% of animals) Dose-dependent endothelial cell damage visible on electron microscopy |
These findings demonstrate that TNF directly induces glomerular functional and structural changes that mirror those seen in endotoxemia, confirming TNF as a mediator of endotoxin-induced glomerular damage .
How does TNF participate in cytokine networks during rabbit inflammatory responses?
In rabbit models of acute inflammation (particularly acute gout), TNF operates within a complex cytokine network involving IL-1β, IL-8, and IL-1 receptor antagonist (IL-1Ra). The temporal dynamics and interactions follow specific patterns:
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TNF-α production peaks at 2 hours after inflammatory stimulus (e.g., monosodium urate crystal injection)
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IL-1β and IL-8 exhibit biphasic production patterns:
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First peak at 2 hours (coinciding with TNF peak)
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Second peak at 9 hours (IL-1β) or 12 hours (IL-8)
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Different cell types contribute to this cytokine network:
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Synovial cells primarily produce TNF-α and the first phase of IL-8
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Infiltrating leukocytes generate the second phase of IL-8, as well as IL-1β and IL-1Ra
Inhibition studies reveal regulatory relationships:
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TNF-α production remains unaffected by anti-IL-8 or IL-1Ra
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The first IL-1β peak requires both TNF-α and IL-8 for maximal production
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The second IL-1β peak can be reduced by either anti-TNF-α or anti-IL-8 alone
This intricate network highlights TNF's role as an initiator of inflammatory cascades in rabbit models.
What are the optimal conditions for in vitro TNF production by rabbit cells?
For optimal in vitro TNF production by rabbit mononuclear phagocytes, researchers should consider:
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Cell source: Alveolar and peritoneal macrophages yield highest production, followed by blood monocytes, spleen macrophages, and marrow cells
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Stimulation: Endotoxin (LPS) provides optimal stimulation for TNF production
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Timing: TNF is primarily synthesized within the first 7 hours of culture
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Cell condition: Cells from BCG-injected rabbits demonstrate enhanced TNF production capacity compared to those from normal rabbits
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Culture conditions: Standard tissue culture conditions with appropriate media (typically RPMI 1640 with serum)
For specialized applications such as TNF production in THP-1 and Raw264.7 cells:
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LPS treatment (1 μg/ml) for 6 hours stimulates TNF production
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Addition of Brefeldin A (300 ng/ml) in the final 3 hours inhibits Golgi traffic, enhancing detection of intracellular TNF
These parameters provide a methodological framework for researchers seeking to produce and study rabbit TNF in vitro.
What detection methods are most suitable for rabbit TNF quantification?
Several detection methods are available for rabbit TNF quantification, each with specific advantages:
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ELISA (Enzyme-Linked Immunosorbent Assay):
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Western Blotting:
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Immunocytochemistry/Immunofluorescence:
For recovery validation, matrices should be spiked with known TNF-β concentrations and recovery rates calculated by comparing measured values to expected amounts. Linearity assessment requires testing samples spiked with appropriate TNF-β concentrations to confirm proportional measurement across the detection range .
How should experimental designs incorporate TNF blockade in rabbit models?
When designing experiments to evaluate TNF's role through blockade approaches:
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Blocking agents selection:
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Anti-TNF monoclonal antibodies (mAbs) specific to rabbit TNF
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Soluble TNF receptors or receptor fusion proteins
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Small molecule TNF inhibitors
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Control considerations:
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Include isotype-matched control antibodies
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Consider both preventive (pre-treatment) and therapeutic (post-induction) blockade
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Evaluate dose-dependent effects of blocking agents
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Combined blockade approaches:
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Outcome measurements:
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Monitor cytokine production patterns (including second-wave cytokines)
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Assess leukocyte infiltration (initial phase occurs within 2 hours; maximal phase at 9 hours)
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Evaluate tissue damage parameters specific to model (e.g., glomerular changes)
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Compare with established treatments (e.g., colchicine in gout models inhibits neutrophil infiltration without affecting TNF or initial IL-8 production)
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What considerations are crucial for translating rabbit TNF findings to human applications?
When translating rabbit TNF research to human applications, researchers should consider:
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Species differences:
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Disease model relevance:
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Rabbit TNF-induced glomerular damage closely resembles human pathologies
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TNF's role in rabbit inflammatory networks parallels human cytokine cascades
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Consider disease-specific differences in TNF's role between species
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Biomarker correlations:
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Establish correlations between TNF levels and pathological findings
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Validate TNF as a biomarker across species for specific conditions
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Consider companion biomarkers that may enhance diagnostic or prognostic value
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Therapeutic implications:
Understanding these translational considerations enhances the value of rabbit TNF research for human medicine.
Product Science Overview
Tumor Necrosis Factor-Alpha (TNF-α) is a cytokine involved in systemic inflammation and is part of the body’s immune response. It is a member of the TNF superfamily, which consists of various proteins that can cause cell death (apoptosis). TNF-α is produced mainly by macrophages, but it can also be produced by other cell types such as lymphocytes, natural killer cells, and neurons.
Recombinant Rabbit TNF-α is produced using an E. coli expression system. The target gene encoding TNF-α is expressed, resulting in a protein that consists of three identical polypeptide chains, each containing 158 amino acids. These chains combine to form a compact, bell-shaped homotrimer . The molecular mass of the individual subunit is approximately 17.4 kDa .
TNF-α is a pleiotropic cytokine, meaning it has multiple effects on different types of cells. It functions by binding to two types of receptors: TNF-R1 (TNF receptor type 1, p55R) and TNF-R2 (TNF receptor type 2, p75R) . The binding of TNF-α to these receptors can lead to various cellular responses, including:
- Promotion of inflammation: TNF-α is a potent inflammatory cytokine that can induce the production of other pro-inflammatory cytokines, such as interleukin-1 (IL-1) and interleukin-6 (IL-6).
- Induction of apoptotic cell death: TNF-α can trigger apoptosis in certain cell types, which is a crucial mechanism for eliminating cancerous or infected cells.
- Inhibition of tumorigenesis: TNF-α has anti-tumor properties and can inhibit the growth and spread of tumors.
- Inhibition of viral replication: TNF-α can interfere with the replication of certain viruses, thereby helping to control viral infections .
Recombinant Rabbit TNF-α is widely used in research to study its various biological functions and its role in diseases. Some of the key applications include:
- Cancer research: TNF-α is studied for its potential use in cancer therapy due to its ability to induce apoptosis in tumor cells.
- Inflammatory diseases: Researchers investigate the role of TNF-α in inflammatory diseases such as rheumatoid arthritis, Crohn’s disease, and psoriasis.
- Infectious diseases: TNF-α is studied for its role in the immune response to infections, particularly in the context of viral infections .
Recombinant Rabbit TNF-α is typically lyophilized and should be stored at temperatures below -20°C for long-term stability. Once reconstituted, the protein solution can be stored at 2-8°C for short-term use (2-7 days) or at -20°C for longer periods (up to 3 months). It is important to avoid repeated freeze-thaw cycles to maintain the protein’s stability and activity .
