Elevated mitochondrial DNA copy number and POL-γ expression but decreased expression of TFAM in murine intestine following therapeutic dose irradiation.

Mitochondria play pivotal roles in cellular handling of oxygen and in apoptosis, the ordered suicide response of cells to irradiation. The involvement of expression products from the 16.5 kb human mitochondrial genome in these activities has been studied widely. However, little is known about effects of irradiation on mammalian mitochondrial DNA (mtDNA). The relative lack of mtDNA repair mechanisms compared with nuclear DNA (nDNA) predicts particular vulnerability to irradiation. Using a technique developed to ascertain mtDNA:nDNA ratios, we previously showed that this ratio increases dramatically in murine small bowel within 48 hours following whole body irradiation. We now report that those levels continue to rise for four days and remain elevated at close to that level beyond 30 days after 5 Gy of irradiation.We further demonstrate that levels of the mtDNA-specific DNA polymerase-γ (Pol-γ ) also show a sharp and sustained increase during this time course after a 2-Gy dose. Paradoxically, transcription factor A (TFAM), exhibited the directly opposite response.

Molecular markers of radiation-related normal tissue toxicity.

Over the past five decades, those interested in markers of radiation effect have focused primarily on tumor response. More recently, however, the view has broadened to include irradiated normal tissues-markers that predict unusual risk of side-effects, prognosticate during the prodromal and therapeutic phases, diagnose a particular toxicity as radiation-related, and, in the case of bioterror, allow for tissue-specific biodosimetry. Currently, there are few clinically useful radiation-related biomarkers. Notably, levels of some hormones such as thyroid-stimulating hormone (TSH) have been used successfully as markers of dysfunction, indicative of the need for replacement therapy, and for prevention of cancers. The most promising macromolecular markers are cytokines: TGFbeta, IL-1, IL-6, and TNFalpha being lead molecules in this class as both markers and targets for therapy. Genomics and proteomics are still in nascent stages and are actively being studied and developed.

Wyman's equation and oxygen flux through the red cell.

Wyman's equation of 1966 describes the facilitation of flux of a reversibly bound substrate such as oxygen, consequent on the translational diffusion of the binding protein (the carrier). While Wyman's equation, or some modification of it such as that by Murray 2, may provide a realistic description of the flux of oxygen through a dilute solution of haemoglobin (see also Wittenburg), it is unlikely to be the complete explanation, nor even the basis, for oxygen transport through the intact red cell. The mature erythrocyte contains approximately 350 g/l haemoglobin, and while this suggests that only 35% of the available water volume is actually occupied by the protein, the remaining 65% is unavailable for protein translational diffusion due to the mutual exclusion of the haemoglobin molecules. For this reason we have examined other possible mechanisms whereby haemoglobin may facilitate the translational diffusion of oxygen within the erythrocyte. Possible alternatives include rotational diffusion by the haemoglobins, intracellular shuffling of haemoglobins due to shape changes by the erythrocyte, and haemoglobin rotations and oxygen exchange consequent on the charge change which accompanies substration and desubstration of the haemoglobin molecule. Finally the dipole interactions are shown to generate significant intermolecular attractions between adjacent haemoglobins.

Effects of temperature on oxygen transport in sheets and spheres of respiring tissues.

The effect of temperature upon the oxygen partial pressure profiles (and hence upon flux) of oxygen through respiring tissues of differing architecture is examined. We have considered the two situations of respiring sheets of tissue and of respiring spheres. Sheets of respiring tissue can model to some extent the behaviour of skin (which abandons its own temperature stasis in response to its obligations in the control of overall body temperature). The oxygen profiles of spheres of respiring tissues subject to temperature shifts is investigated since it is a model for solid tumour oxygen kinetics where a spherical tumour, inadequately supplied with a capillary network, is being treated by one or another form of hyperthermia during cancer therapy.

The anomalous Einstein-Stokes behaviour of oxygen and other low molecular weight diffusants.

Almost a century ago, Einstein and Sutherland independently derived equations that describe the relationship between diffusion of solutes and the molecular parameters of those solutes. In that time it has been recognized that, although the equations adequately describe the diffusion of large and medium-sized molecules, there is deviation from this relationship for small molecules. Many authors have attempted to redefine the equations for diffusion, with varying degrees of success, but generally have not attempted to consider the fundamental events that may be occurring at the molecular level during the diffusion of small molecules. In this presentation, we attempt to provide such an explanation, particularly with respect to the diffusion of oxygen through water. We consider the possibility of a random rotational model that complements the (slower) translational process of traditional diffusion and thereby provides accelerated diffusion of small molecules. It is hoped that our description of this model may provide a basis for the development of mathematical modelling of the process.

Focusing on genomic and phenomic correlations in respiration of non-melanotic skin cancers.

In recent years, with the development of techniques in modem molecular biology, it has become possible to study the genetic basis of carcinogenesis down to the level of DNA sequence. Major advances have been made in our understanding of the genes involved in cell cycle control and descriptions of mutations in those genes. These developments have led to the definition of the role of specific oncogenes and tumour suppressor genes in several cancers, including, for example, colon cancers and some forms of breast cancer. Work reported from our laboratory has led to the identification of a number of candidate genes involved in the development of non-melanotic skin cancers. In this chapter, we attempt to further explain the observed (phenomic) alterations in metabolic pathways associated with oxygen consumption with the changes at the genetic level.

Chromosomal aberrations in squamous cell carcinoma and solar keratoses revealed by comparative genomic hybridization.

OBJECTIVE: To identify chromosomal copy numbers of frequent genetic aberrations within squamous cell carcinomas (SCCs) and solar keratoses (SKs), and provide further evidence to support or challenge current dogma concerning the relationship between these lesions. DESIGN: Retrospective analysis of genetic aberrations in DNA from SK and SCC biopsy specimens by comparative genomic hybridization. SETTING: University-based research laboratory in Queensland, Australia. PATIENTS: Twenty-two biopsy specimens from patients with diagnosed SKs (n = 7), cutaneous SCCs (n = 10), or adjoining lesions (n = 5). MAIN OUTCOME MEASURES: Identification of frequent genetic aberrations both specific to SK and SCC and shared by these lesions to investigate their clonal relationship. RESULTS: Shared genomic imbalances were identified in SK and SCC. Frequent gains were located at chromosome arms 3q, 17q, 4p, 14q, Xq, 5p, 9q, 8q, 17p, and 20q, whereas shared regional losses were observed at 9p, 3p, 13q, 17p, 11p, 8q, and 18p. Significant loss of 18q was observed only in SCC lesions. CONCLUSIONS: Our results demonstrate that numerous chromosomal aberrations are shared by the 2 lesions, suggesting a clonal relationship between SK and SCC. Additionally, the genomic loss of 18q may be a significant event in SK progression to SCC. Finally, the type and frequency of aberrations suggests a common mode of tumorigenesis in SCC-derived tumors.

How proton translocation across mitochondrial inner membranes drives the Fo rotor of ATP synthase.

It has been believed for some time that there are two major but alternative models for the selective transport of ions across membranes generally. On the one hand this transport is by way of transmembrane channels. These channels exist within macromolecular complexes which span the membrane and provide a hydrophilic pathway through which the ions can be translocated. Alternatively, carriers have been postulated which can dissolve in the lipid moiety of the membrane, are able to selectively co-ordinate ions, and then move from one side of the membrane to the other, before unloading the ion. Proton translocation across the inner mitochondrial membrane is intensely interesting, firstly because the process is tightly coupled to the synthesis of ATP, but additionally because the emerging picture of proton translocation incorporates features from both the classical mechanisms of ion transport. Thus there are two channels, one from either side of the membrane, both of which penetrate to the centre of the membrane. However neither of them individually spans the membrane, but they remain separated by a short distance in the plane of the membrane. Transport across this remaining gap involves a carrier that reversibly binds the ion. The mechanism for transport across this remaining region is not carrier-facilitated diffusion, nor any "flip flop" change of shape by the carrier. Rather it is an electrically driven rotation of the carrier, and the source of the electric field that drives this rotor is the transmembrane electric potential.

The flux of oxygen within tissues.

Diffusive flux of oxygen through tissues which are essentially connective and have few cells, display reduced diffusion coefficients when compared to that through an equivalent lamina of water. In general even significant reductions can be explained in terms of the exclusions imposed on small molecular weight diffusates by the large hydrodynamic domains of the connective tissue components. An alternative way of explaining this large exclusion is to point to the very large microscopic viscosities which large interacting polymers impose upon the solvent (water). By contrast, the diffusive flux of oxygen through tissues composed of contiguously packed and actively respiring cells, shows an increased diffusive flux for oxygen when compared to that through an equivalent water lamina. This increase can be explained in terms of the substantial solubility of oxygen within the membrane phase of the cells. This high oxygen partition coefficient into cell lipids has several consequences. Firstly oxygen diffusion will be directed and two dimensional rather than random and three dimensional. Secondly this diffusion will be directed towards the oxygen-consuming sites which are located at lipid surfaces. Thirdly the aqueous oxygen partial pressure will be kept low (since re-supply is constrained while consumption is continuous). This low aqueous environment permits all of the cell soluble redox systems to be maintained efficiently at low metabolic cost, as well as minimising the risk of unscheduled oxidations. Viewed from this perspective, the high value found for oxygen partition coefficient into the erythrocyte membrane suggests that evolution of membrane structure and components may have been driven in part by the selective advantages of high oxygen solubility.