Molecular and Cellular Toxicology

An Introduction

Lesley Stanley

• Publisher: Wiley
• Year: 2014
• ISBN:
  9781118526019
• Edition: 1
• Topic: Medicine
• Subtopic: Toxicology

Toxicology is the study of the adverse effects of chemical, physical, or biological agents on people, animals, and the environment. Toxicologists are trained to investigate, interpret, and communicate the nature of those effects.

Over the last ten years the subject of toxicology has changed dramatically, moving from a discipline which was once firmly wedded to traditional methods to one which is keen to embrace the innovative techniques emerging from the developing fields of cell culture and molecular biology. There is an acute need for this to be reflected in a paradigm shift which takes advantage of the opportunities offered by modern developments in the life sciences, including new in vitro and in silico approaches, alternative whole organism (non-mammalian) models and the exploitation of 'omics methods, high throughput screening (HTS) techniques and molecular imaging technologies.

This concise, accessible introduction to the field includes the very latest concepts and methodologies. It provides MSc, PhD and final year undergraduate students in pharmacy, biomedical and life sciences, as well as individuals starting out in the cosmetics, consumer products, pharmaceutical and testing industries, with everything they need to know to get to grips with the fast moving field of toxicology and the current approaches used in the risk assessment of drugs and chemicals.

Chapter 1
Background to Molecular and Cellular Toxicology

1.1 What do we mean by molecular and cellular toxicology?

The Society of Toxicology¹ defines Toxicology as ‘the study of the adverse effects of chemical, physical or biological agents on people, animals and the environment’ and toxicologists as ‘scientists trained to investigate, interpret and communicate the nature of those effects’. In the disciplines of molecular and cellular toxicology, toxicologists make use of the many new techniques which are becoming available in the molecular life sciences to understand the underlying mechanisms by which these agents damage cells, tissues and entire organisms. The main aims of toxicity testing, whether during pre-clinical drug development, in the course of safety assessment of cosmetic ingredients and consumer products or while evaluating the potential consequences of exposure to industrial and environmental chemicals, are to construct a toxicological profile of the chemical and to identify a threshold dose (if any).

The topic of this book is how molecular and cellular techniques can be used to study the toxicity of exogenous chemicals, referred to in the trade as xenobiotics. The primary target organs for xenobiotic toxicity are usually those which are exposed to xenobiotics and their metabolites because of the roles they play as portals of entry, sites of metabolism and/or organs of excretion. The molecular and cellular consequences of exposure are summarised in Figure 1.1.

Figure 1.1 Consequences of exposure to a toxic insult (source: Dr Cliff Elcombe, CXR Biosciences Ltd. Reproduced with permission of Dr Cliff Elcombe)

Despite the many scientific advances made in the life sciences over the last couple of decades, which include spectacular advances in the fields of molecular biology, biotechnology and bioinformatics, the basic concepts of regulatory toxicology have hardly changed over the same period. For example, although the classical LD₅₀ (dose giving 50% lethality) test for oral toxicity and the Draize tests for eye or skin irritancy are widely considered to cause unacceptable suffering to laboratory animals, they are still widely used and the development of non-animal alternatives has been slow, to say the least. However, the implementation of both the 7th Amendment to the European Union (EU) Cosmetic Ingredient Directive and the Registration, Evaluation, Authorisation and restriction of Chemicals (REACH) regulations during the early 2000s has provided a strong stimulus for further developments.

There is an acute need for this to be reflected in a paradigm shift in the field of toxicology to take advantage of the new opportunities offered by modern developments in the life sciences, including new in vitro models, alternative whole organism (non-mammalian) models and the exploitation of ‘omics methods, high throughput screening (HTS) technologies and molecular imaging technologies.²

1.2 Tissues and their maintenance

Tissues are made up of cells of various types plus the extracellular space which surrounds them. The extracellular space is filled with extracellular matrix, the proportion and structure of which depends on the tissue type. Epithelia, for example, consist mainly of sheets of epithelial cells with very little extracellular matrix whereas connective tissue contains few cells and a lot of extracellular matrix. The proteins of the extracellular matrix are linked to cytoskeletal proteins through the plasma membrane and are able to influence cell development, migration, proliferation, shape and function.

All tissues have certain basic requirements including mechanical strength, access to nutrients and removal of waste, connection to the nervous system, removal of debris and protection against infection. Specialised (differentiated) cells provide these and other functions.

During the process of embryonic development, the fertilised ovum proliferates and the resulting daughter cells differentiate to form three germ layers:

• The endoderm gives rise to the epithelia of the gut and its associated organs (lung, liver and pancreas).
• The ectoderm gives rise to the outer surface epithelia (epidermis, buccal epithelium and outer cervical epithelium) and neuroectodermal tissues.
• The mesoderm gives rise to the embryonic mesenchyme and thence to the connective tissue and supporting tissues including bone, cartilage, muscle, vascular tissue and haematopoietic system.

The products of this process are the various differentiated tissues of the body. Even when removed from their normal environment, differentiated cells retain their specialised characteristics; for example, glandular cells still secrete mucin, fibroblasts still make extracellular matrix and macrophages still carry out phagocytosis. Differentiated cells can still respond to the environment and some cell types can adapt quite dramatically: for example, fibroblasts can convert into cartilage cells, liver cells can express different enzymes and mammary cells can switch milk proteins on or off. Some cells, however, are terminally differentiated, having become so specialised that they have lost the ability to divide.

1.2.1 Stem cells

Terminally differentiated tissues are maintained by stem cells, precursors which are not themselves differentiated but are committed to produce a particular type of terminally differentiated cell. A stem cell can be defined as ‘a cell which can proliferate either symmetrically or asymmetrically in response to an appropriate external signal’;³ in other words, under one set of circumstances it will divide to produce two stem cells and under other circumstances it will divide to generate one stem cell and one progenitor cell which can give rise to a differentiated cell lineage. The signals to which stem cells can respond include growth factors, levels of oxygen and antioxidants and growth substrates (e.g. feeder layers, extracellular matrix).

Stem cells can divide without limit and on division the daughter cells have a choice either to remain as a stem cell or embark on terminal differentiation. The final differentiated state of the majority of stem cells is pre-determined (e.g. muscle satellite cell, spermatogonium), although some stem cells are pluripotent (can differentiate into many cell types). Organ-specific stem cells have two defining properties, the ability to self-renew and the potential to differentiate into organ-specific cell types. The various types of stem cells have different potencies (i.e. abilities to generate different classes of progeny):

• A totipotent stem cell can generate an entire new organism. The definitive totipotent stem cell is the fertilised egg; following implantation, the totipotent fertilised egg becomes committed to form an embryonic pluripotent stem cell.
• A pluripotent stem cell can give rise to any other type of cell but not to an entire new organism. Pluripotent cells give rise to committed progenitor cells which can only mature into one type of cell (i.e. each one is unipotent) and this maturation process involves differentiation, which is controlled by growth factors and the surrounding environment.
• Multi-potent stem cells can produce a limited number of cell types and are committed to become part of a particular organ. They give rise to lineages of progenitor cells.
• Progenitor cells are committed to a particular lineage (e.g. the haematopoietic system) and give rise to terminally differentiated cells, which do not divide further.

1.3 Tissue damage

Living tissues are constantly exposed to environmental changes to which they respond with modifications of metabolism and growth.

• Primary (direct) injury involves an interaction between the chemicals and the components of the cell. Toxic cell injury requires high concentrations of toxic compounds and, in some cases, metabolic activation. It may involve membrane damage (e.g. lipid peroxidation induced by carbon tetrachloride in the liver).
• Secondary (indirect) injury involves changes in the cellular environment (e.g. oxygen tens...