PepGenexShop our research products

In vitro vs in vivo

In vitro versus in vivo is the distinction between experiments performed on cells, tissues or purified molecules outside a living organism (in vitro) and experiments performed in a living organism (in vivo). A review by Duval and colleagues describes cell cultures in vitro as frequently used to advance understanding of the mechanisms that underlie cell behavior in vivo, and the ARRIVE guidelines for reporting research using laboratory animals take their name from the phrase Animals in Research: Reporting In Vivo Experiments.[1],[2]

What kinds of studies count as in vitro?

PepGenex Science classifies as in vitro any study in cells, tissues or purified molecules outside a living organism, including binding, signalling and structural studies.

A cell-based example is Mayo's 1992 report on a cloned receptor for growth hormone-releasing hormone (GHRH): the receptor was expressed in human kidney 293 cells, membrane fractions from those cells bound GHRH with high affinity and specificity, and GHRH stimulated intracellular cAMP production in the transfected cells.[3]

A structural example is the cryo-electron microscopy structure reported by Zhang and colleagues in 2017 of the peptide-activated GLP-1 receptor in complex with the G protein Gs.[4]

What does a cell culture leave out?

Duval and colleagues write that two-dimensional (2D) cell cultures have been used for over a century as in vitro models, and that growing evidence shows that under some circumstances 2D systems can result in cell bioactivities that deviate appreciably from the in vivo response.[1]

The same review describes conventional 2D culture as relying on adherence to a flat surface, typically glass or polystyrene. Cells in 2D monolayers have access to similar amounts of nutrients and growth factors, which results in homogeneous growth, and most 2D methods do not control cell shape, which determines biophysical cues affecting cell behavior in vivo.[1]

Three-dimensional (3D) culture aims to reproduce in vivo conditions more closely, but the review lists remaining challenges including the tissue-tissue interface, the mechanical microenvironment and the spatiotemporal distributions of oxygen, nutrients and metabolic wastes. It also notes that 2D approaches can still recapitulate in vivo behavior for many bioactivities.[1]

What happens to a peptide in a living organism that does not happen in a dish?

A review by Werle and Bernkop-Schnürch states that short plasma half-lives of peptides and proteins are commonly due to fast renal clearance and to enzymatic degradation during systemic circulation. It gives the example of octreotide, a shortened derivative of somatostatin containing D-amino acids, with a plasma half-life of 1.5 hours compared with only a few minutes for somatostatin.[5]

The same review compiles data on the most important proteolytic enzymes of human blood, liver and kidney and their cleavage specificity, for predicting the enzymatic cleavage of peptides and proteins during systemic circulation.[5]

Rang describes studies on signal transduction as revealing great complexity in the events linking ligand binding to the physiological response, with the current emphasis on unravelling the pathways that link receptors to responses.[6]

Mayo also reported that rat GHRH receptor mRNA is expressed predominantly, if not exclusively, in the anterior pituitary gland, a finding about where the receptor occurs in the animal that was reported separately from the binding measured in transfected cells.[3]

Why can the same ligand look different in two assays?

Kenakin describes inverse agonism as a phenotypic behavior that can only be observed in an appropriate assay, one in which constitutive receptor activity is present. In the absence of constitutive activity, inverse agonists behave as simple competitive antagonists.[7]

In a survey of 105 articles covering 380 antagonists at 73 G protein-coupled receptor targets, Kenakin found 322 inverse agonists and 58 neutral antagonists. Whether that distinction can be seen at all depends on the assay system in which a ligand is measured.[7]

What details define an in vivo experiment?

The ARRIVE guidelines define a laboratory animal as any species of animal undergoing an experimental procedure in a research laboratory or formal test setting. Their checklist of 20 items describes the minimum information that publications reporting research using animals should include.[2]

Those items include the number and specific characteristics of the animals used, including species, strain, sex and genetic background; details of housing and husbandry; and the experimental, statistical and analytical methods, including methods used to reduce bias such as randomisation and blinding.[2]

How does PepGenex Science keep in vitro and in vivo evidence apart?

Each PepGenex Science compound profile groups its studies by evidence type, so an in vitro finding is never presented beside an animal or human finding as if they were equivalent. A result from a cell assay is described as a cell assay result, and a result in an animal model is described as an animal model result.

Structural and mechanism studies explain how a molecule interacts with its target in the system studied. PepGenex Science does not present them as evidence of an effect in a whole organism.

Limitations

An in vitro result describes the system in which it was measured. Duval and colleagues state that the question at hand determines whether a 2D or 3D culture method is more suitable, and that a universal 3D platform does not currently exist.[1]

An in vivo result in an animal is not a human result. Van der Worp and colleagues write that animal studies do not predict with sufficient certainty what will happen in humans.[8]

This page describes what the cited documents say. It is not a statement that any PepGenex material has been manufactured, tested or released under any of them.

Compound profiles

Compound profiles whose recorded evidence includes both laboratory (in vitro) and animal studies.

References

  1. Duval K, Grover H, Han LH, et al. Modeling Physiological Events in 2D vs. 3D Cell Culture. Physiology (Bethesda). 2017;32(4):266-277. PMID 28615311 · DOI 10.1152/physiol.00036.2016 · PMC5545611
  2. Kilkenny C, Browne WJ, Cuthill IC, Emerson M, Altman DG. Improving bioscience research reporting: the ARRIVE guidelines for reporting animal research. PLoS Biol. 2010;8(6):e1000412. PMID 20613859 · DOI 10.1371/journal.pbio.1000412 · PMC2893951
  3. Mayo KE. Molecular cloning and expression of a pituitary-specific receptor for growth hormone-releasing hormone. Mol Endocrinol. 1992;6(10):1734-1744. PMID 1333056 · DOI 10.1210/mend.6.10.1333056
  4. Zhang Y, Sun B, Feng D, et al. Cryo-EM structure of the activated GLP-1 receptor in complex with a G protein. Nature. 2017;546(7657):248-253. PMID 28538729 · DOI 10.1038/nature22394 · PMC5587415
  5. Werle M, Bernkop-Schnürch A. Strategies to improve plasma half life time of peptide and protein drugs. Amino Acids. 2006;30(4):351-367. PMID 16622600 · DOI 10.1007/s00726-005-0289-3
  6. Rang HP. The receptor concept: pharmacology's big idea. Br J Pharmacol. 2006;147 Suppl 1:S9-S16. PMID 16402126 · DOI 10.1038/sj.bjp.0706457 · PMC1760743
  7. Kenakin T. Efficacy as a vector: the relative prevalence and paucity of inverse agonism. Mol Pharmacol. 2004;65(1):2-11. PMID 14722230 · DOI 10.1124/mol.65.1.2
  8. van der Worp HB, Howells DW, Sena ES, et al. Can animal models of disease reliably inform human studies? PLoS Med. 2010;7(3):e1000245. PMID 20361020 · DOI 10.1371/journal.pmed.1000245 · PMC2846855