PTH (7-34) Human

What is PTH (7-34) Human and how does it differ from the full-length PTH hormone?

PTH (7-34) Human is a 28-amino acid fragment of the human Parathyroid Hormone that lacks the first six N-terminal amino acids of the full hormone. While full-length PTH (1-34) is a potent agonist of the PTH1 receptor, PTH (7-34) functions primarily as a PTH1R antagonist. It is naturally secreted by chief cells of the parathyroid glands when extracellular calcium levels are high, offsetting the effects of the full PTH hormone . Structurally, PTH (7-34) has a molecular weight of approximately 3.4 kDa and interacts with the PTH receptor through regions that are not homologous to the full-length PTH, which contributes to its distinctive pharmacological profile .

What are the primary biological functions of PTH (7-34) Human in research contexts?

In research contexts, PTH (7-34) Human serves as an important tool for understanding PTH receptor signaling and function. It acts as a PTH1R antagonist capable of stimulating proliferation and differentiation of hair follicles and epidermis while promoting cell survival through activation of the Akt pathway . Unlike the full PTH hormone, PTH (7-34) exhibits minimal effects on bone architecture and vascular calcification in experimental models . Its ability to inhibit PTH-stimulated cAMP production makes it valuable for studying receptor-mediated signaling pathways and for developing potential therapeutic applications targeting PTH-dependent processes .

How should PTH (7-34) Human be stored and handled in laboratory settings?

For optimal stability and activity, PTH (7-34) Human should be stored desiccated at -20°C . The lyophilized powder formulation maintains integrity during storage, but care should be taken during reconstitution to avoid repeated freeze-thaw cycles. When working with PTH (7-34), researchers should note that it is purified and tested specifically for cell culture applications, with purity typically >97% as determined by SDS-PAGE and HPLC analyses . Standard laboratory safety protocols should be followed, and it's important to note that research-grade materials are manufactured for research use only and cannot be used for human consumption.

What are the optimal cell culture systems for studying PTH (7-34) Human antagonist activity?

The most widely validated cell culture system for studying PTH (7-34) Human antagonist activity is the bone-derived rat osteosarcoma cell line ROS 17/2.8, which expresses functional PTH receptors . This system is particularly valuable because it can detect both antagonism and weak agonism of PTH analogs. For examining receptor binding, competition assays using radiolabeled PTH and varying concentrations of PTH (7-34) can effectively demonstrate receptor interaction dynamics.

Other relevant cellular models include:

When designing experiments, researchers should consider that PTH (7-34) exhibits approximately 8-fold higher potency than [Tyr34]bPTH-(7-34)NH2 for inhibiting PTH-stimulated cAMP production, though they are equipotent for inhibition of radiolabeled PTH-binding .

How should dose-response studies with PTH (7-34) Human be designed and interpreted?

When designing dose-response studies with PTH (7-34) Human, researchers should:

• Establish a concentration range that spans at least 5 orders of magnitude (typically 10⁻¹⁰ to 10⁻⁵ M)

• Include both PTH (1-34) and PTH (7-34) treatments to directly compare agonist versus antagonist effects

• Measure multiple endpoints, as PTH (7-34) may show different potencies for different signaling pathways

For interpretation, it's crucial to recognize that PTH (7-34) exhibits weak partial agonist activity at high concentrations (approximately 8 μM), producing a modest 2.4-fold increase in cAMP (about 5% of the maximal response) in ROS cells . This dual antagonist/weak agonist property must be considered when interpreting results, especially at higher concentrations. IC₅₀ values should be calculated for antagonistic effects, and potential partial agonism at higher concentrations should be carefully documented.

What methodological approaches are recommended for studying PTH (7-34) effects on calcium homeostasis in vivo?

For studying PTH (7-34) effects on calcium homeostasis in vivo, the following methodological approach is recommended based on established protocols:

• Animal selection: Use appropriate animal models, considering that rodent and human PTH receptors differ in sequence (91% identity), potentially affecting binding affinities .

• Blood collection: Collect blood samples via tail vein puncture into heparinized capillary tubes at baseline and multiple timepoints post-injection (typically 1, 2, 4, and 8 hours) .

• Calcium measurement: Analyze blood samples immediately using a calcium/pH analyzer (e.g., Siemens RapidLab 348) .

• Phosphate analysis: For phosphate measurements, collect blood into microcentrifuge tubes containing EDTA, centrifuge at 8000 rpm (4°C) for 15 minutes, and analyze plasma using a colorimetric phosphate assay .

• Comparative design: Include both PTH (7-34) and PTH (1-34) treatment groups, with appropriate vehicle controls.

When conducting antagonist studies, administer PTH (1-34) at a standard dose (e.g., 40 nmol/kg) alone or together with PTH (7-34) at a higher dose (e.g., 500 nmol/kg) to demonstrate antagonistic effects . Current evidence suggests minimal effects of PTH (7-34) on bone or vascular calcification compared to PTH (1-34) , but experimental design should account for potential species-specific differences in receptor binding.

How can humanized mouse models improve the translational potential of PTH (7-34) research?

Humanized mouse models expressing the human PTH1R offer significant advantages for translational PTH (7-34) research. The PTH (7-34) antagonist fragment binds with approximately 50-fold lower affinity to the rat PTH1R than to the human PTH1R, with the binding differences attributed to divergences in the N-terminal extracellular domain of the receptor . This species-based difference in receptor-binding affinities can lead to discrepant outcomes when evaluating PTH analogs in traditional rodent models versus humans.

The homologous recombination-based knockin approach has successfully generated mice expressing the human PTH1R instead of the endogenous murine receptor, with expression directed by the endogenous mouse promoter . These hPTH1R-KI mice:

• Remain healthy over multiple generations

• Show functional responses to injected PTH analog peptides consistent with human PTH1R function

• Provide a more accurate platform for evaluating the pharmacology of PTH (7-34) and other PTH analogs

These models are particularly valuable for determining the true potency of PTH (7-34) antagonism in a physiologically relevant context with the human receptor, potentially revealing effects that might be underestimated in conventional rodent models .

What are the molecular mechanisms behind the differential effects of PTH (7-34) on receptor binding versus signaling inhibition?

The molecular mechanisms underlying the differential effects of PTH (7-34) on receptor binding versus signaling inhibition are complex and reveal important insights about PTH receptor pharmacology:

• PTH (7-34) lacks the first six N-terminal amino acids of PTH that are critical for receptor activation but retains regions necessary for binding to the extracellular domain of PTH1R.

• The peptide exhibits equipotent inhibition of radiolabeled PTH-binding compared to [Tyr34]bPTH-(7-34)NH2, but is 8-fold more potent at inhibiting PTH-stimulated cAMP production .

• This differential potency suggests that PTH (7-34) induces a receptor conformation that more effectively blocks G-protein coupling while still allowing receptor binding.

Research indicates that PTH (7-34) interacts with PTH receptors based largely on regions that are not homologous to PTH , which may explain its unique pharmacological profile. At high concentrations (8 μM), PTH (7-34) demonstrates weak partial agonist activity, producing approximately 5% of the maximal cAMP response , suggesting that it can induce a minimally active receptor conformation at high occupancy levels. These nuanced effects highlight the importance of biased signaling analysis when studying PTH receptor pharmacology.

What experimental approaches can help resolve contradictory findings regarding PTH (7-34) effects on bone metabolism?

To resolve contradictory findings regarding PTH (7-34) effects on bone metabolism, researchers should implement the following comprehensive experimental approach:

• Species-specific receptor considerations: Use humanized mouse models expressing human PTH1R to account for the approximately 50-fold difference in binding affinity between rat and human receptors .

• Dose-response relationship assessment: Conduct detailed dose-response studies across a wide concentration range to identify potential biphasic effects, as PTH (7-34) can function as both an antagonist and weak partial agonist depending on concentration .

• Temporal analysis: Compare acute versus chronic administration effects, as PTH (7-34) may demonstrate different actions with intermittent versus continuous exposure.

• Comprehensive bone analysis methodology:

  • Micro-CT analysis for bone microarchitecture

  • Histomorphometry for cellular changes and bone formation rates

  • Biomechanical testing for functional bone strength

  • Serum biochemistry for systemic mineral metabolism markers

• Signaling pathway delineation: Investigate multiple downstream signaling pathways beyond cAMP, including calcium mobilization and β-arrestin recruitment.

Current evidence suggests minimal bone effects of PTH (7-34) in CKD animal models , but results may vary depending on experimental conditions, animal models, and dosing regimens. By systematically addressing these variables while accounting for species-specific receptor pharmacology, researchers can better reconcile contradictory findings in the literature.

How does PTH (7-34) Human compare with other PTH antagonists in research applications?

PTH (7-34) Human offers distinct advantages and limitations compared to other PTH antagonists used in research applications:

PTH (7-34) Human is particularly valuable for studies requiring high potency inhibition of PTH-stimulated cAMP production, being 8-fold more potent than [Tyr34]bPTH-(7-34)NH₂ in this regard, while both compounds are equipotent for inhibition of radiolabeled PTH-binding . This differential potency profile makes PTH (7-34) Human especially useful for investigating biased antagonism of PTH receptor signaling pathways.

What are the key experimental considerations when comparing PTH (7-34) versus PTH (1-34) in the same study?

When designing studies comparing PTH (7-34) and PTH (1-34) in the same experimental paradigm, researchers should consider several critical factors:

• Receptor species differences: The human PTH1R binds PTH (7-34) with approximately 50-fold higher affinity than the rat PTH1R, making species selection crucial for accurate comparison . Using humanized mouse models expressing the human PTH1R can provide more translatable results.

• Dose selection rationale: Due to differences in potency, appropriate dose ranges for each peptide must be established:

  • PTH (1-34): Typically effective at 40 nmol/kg for in vivo studies

  • PTH (7-34): Higher doses (e.g., 500 nmol/kg) are typically required to demonstrate antagonism

• Temporal considerations: PTH (1-34) typically shows rapid effects on calcium mobilization, while antagonistic effects of PTH (7-34) may require pre-administration before PTH (1-34) challenge.

• Endpoint selection: Include both:

  • Early signaling events (cAMP production, calcium mobilization)

  • Downstream physiological outcomes (blood calcium/phosphate, bone formation markers)

• Partial agonism awareness: At high concentrations, PTH (7-34) exhibits weak partial agonist activity (~5% of maximal response) , which may complicate interpretation of results.

Studies comparing these compounds should always include appropriate controls and concentration gradients to fully characterize their differential effects on receptor binding, signaling, and physiological outcomes.

What emerging applications of PTH (7-34) Human are being explored in translational research?

Several promising translational research directions for PTH (7-34) Human are emerging:

• Dermatological applications: PTH (7-34) has demonstrated capacity to stimulate proliferation and differentiation of hair follicles and epidermis while promoting cell survival through Akt activation . This suggests potential applications in hair loss treatments and skin regeneration therapies.

• Targeted bone metabolism modulation: Unlike full-length PTH, PTH (7-34) shows minimal effects on bone and vascular calcification , suggesting it might be used to selectively antagonize specific PTH effects while preserving others, potentially useful in disorders of mineral metabolism.

• Biased signaling exploitation: The differential potency of PTH (7-34) in receptor binding versus signaling inhibition points to its potential as a prototype for developing biased PTH receptor modulators that could selectively influence specific downstream pathways.

• Combination therapies: Preliminary research suggests potential for combining PTH (7-34) with other bone-active agents to fine-tune therapeutic responses in metabolic bone diseases, though more research is needed in this area.

• Novel delivery systems: Encapsulation of PTH (7-34) in nanoparticles or other advanced delivery systems is being explored to enhance stability, targeting, and duration of action for research and potential therapeutic applications.

These emerging applications highlight the need for continued mechanistic studies of PTH (7-34) pharmacology in physiologically relevant model systems.

How might advanced structural biology techniques enhance our understanding of PTH (7-34) interaction with PTH1R?

Advanced structural biology techniques offer promising avenues to enhance our understanding of PTH (7-34) interaction with PTH1R:

• Cryo-electron microscopy (Cryo-EM): This technique can reveal the conformational changes in PTH1R upon binding of PTH (7-34) compared to PTH (1-34), potentially explaining the molecular basis for antagonism versus agonism. Recent advances in Cryo-EM resolution now allow visualization of subtle conformational differences in class B GPCRs like PTH1R.

• Hydrogen-deuterium exchange mass spectrometry (HDX-MS): This approach can map the dynamic interaction surfaces between PTH (7-34) and the receptor, identifying regions involved in both binding and conformational changes that influence signaling.

• Molecular dynamics simulations: Based on crystal structures, these computational approaches can predict how PTH (7-34) alters receptor flexibility and conformational ensembles compared to the full agonist.

• Site-directed fluorescence spectroscopy: By introducing fluorescent probes at specific positions in the receptor and ligand, researchers can track conformational changes upon binding in real-time.

• NMR spectroscopy of labeled peptides: This can provide detailed information about the structure and dynamics of PTH (7-34) both free in solution and when receptor-bound.

These techniques would be particularly valuable for understanding why PTH (7-34) shows equipotent binding compared to other antagonists but is 8-fold more potent at inhibiting cAMP signaling , potentially revealing novel mechanisms of biased antagonism.

What methodological innovations could improve the specificity and sensitivity of PTH (7-34) research?

Several methodological innovations could significantly enhance the specificity and sensitivity of PTH (7-34) research:

• CRISPR-engineered cellular models: Precise genome editing to create cell lines with fluorescently tagged PTH1R or modified signaling components could enable real-time visualization of receptor trafficking and signaling in response to PTH (7-34).

• Biosensor development: Development of FRET or BRET-based biosensors specific for conformational changes induced by PTH (7-34) binding would allow more sensitive detection of subtle effects on receptor activation states.

• Single-molecule imaging techniques: These approaches could reveal heterogeneity in receptor responses to PTH (7-34) at the individual molecule level, potentially uncovering subpopulations of receptors with distinct signaling properties.

• Biased signaling quantification methods: Development of mathematical models that comprehensively capture the multidimensional aspects of PTH (7-34) signaling would enable more precise characterization of its pharmacological profile.

• Tissue-specific conditional expression systems: In vivo models with tissue-specific and inducible expression of human PTH1R would allow for more precise delineation of PTH (7-34) effects in different target tissues.

• Improved pharmacokinetic tracking: Development of minimally modified, traceable PTH (7-34) analogs would enhance our understanding of its distribution, metabolism, and elimination in vivo.

These methodological innovations would address current limitations in studying the complex pharmacology of PTH (7-34) and potentially reveal novel aspects of its biological activity that current approaches might miss.