Q. Perveen Ghani

Interleukin-1β Activity and Collagen Synthesis in Human Dental Pulp Fibroblasts

Immunopathologic reactions play a significant role in inflammatory diseases of dental pulp. Interleukin-1beta (IL-1beta) is recognized as a key player in mediating cellular immune response. In this study, we measured the content of IL-1beta and its effect on collagen synthesis in cultures of fibroblasts derived from healthy and diseased dental pulps. We found that diseased pulp fibroblasts contain 2.5-fold greater amounts of IL-1beta and synthesized 80% greater amounts of collagen compared with healthy pulp fibroblasts. However, exogenous IL-1beta failed to stimulate collagen synthesis by diseased fibroblasts, whereas collagen synthesis by healthy pulp fibroblasts was stimulated by more than 2-fold. These observations imply that pulp disease induces abnormalities associated with fibroblast response toward IL-1beta.

Role of ADP-ribosylation in wound repair. The contributions of Thomas K. Hunt, MD

Nearly 36 years ago Thomas K. Hunt, with Patrick Twomey, was the first to report that the level of lactate significantly increases in healing wounds. This observation convinced him that lactate, besides being the by‐product of glycolysis, must have a regulatory role in the healing process. He set out to investigate this observation and found it to be so. This article is written in recognition of his foresight. It summarizes the salient findings emanating from this fundamental observation and describes the biochemical principles by which most of the lactate action may be explained. Down‐regulation of the ubiquitous protein modification reaction called ADP‐ribosylation turned out to be a basic signal behind the role of lactate in wound healing. (WOUND REP REG 2003;11:439–444)

Regulatory role of lactate in wound repair

Publisher Summary This chapter discusses the role of lactate in wound repair. Healing wounds produce and accumulate large concentrations of lactate. Wound-healing studies have revealed that hypoxia and high lactate are the characteristics of healing. High concentrations of lactate force the lactate dehydrogenase (LDH)-catalyzed conversion of nicotinamide adenine dinucleotide nicotinamide adenine dinucleotide (NAD +) to nicotine adenine dinucleotide (reduced)(NADH). The decline of the NAD + pool and the subsequent downregulation of NAD-mediated polyadenosine diphosphoribose (pADPR) and adenosine diphosphoribose (ADPR) are crucial mediators that stimulate collagen and vascular endothelial growth factor (VEGF) production. ADP-ribosylation regulates many enzyme activities and functions of nuclear proteins, including those involved in DNA repair, differentiation, and transcription through the modification of transcription factors, such as Sp1, p53, Yin Yang 1, and cyclic AMP response element-binding protein (CREB). In this chapter, the effect of lactate on collagen synthesis, effect of lactate on angiogenesis, methods for assessing the lactate effect, and immunohistochemical analysis of pADPR are discussed.

Stimulation of poly(ADP-ribose) synthetase activity in the lungs of mice exposed to a low level of ozone

Toxic effects of O3 are mediated through the formation of free radicals, which can cause DNA strand breaks. Cellular DNA repair is dependent upon the formation of poly(ADP-ribose) (polyADPR) catalyzed by polyADPR synthetase. In order to evaluate whether O3 exposure inflicted DNA damage in lung tissue, we measured the activity of polyADPR synthetase (known to be activated in response to DNA damage) in mouse lungs after exposure to 0.45 ppm (882 micrograms/m3) O3 for up to 7 days. The enzyme activity was stimulated with O3 exposure relative to unexposed controls, showing a 20% (P less than 0.05) increase at Day 5 and 42% (P less than 0.001) at Day 7 of O3 exposure. In addition, the activity of superoxide dismutase (SOD), known to be stimulated in response to production of superoxide anion (.O2-), was measured as an indicator of free radical involvement. Relative to unexposed controls, the SOD activity in exposed animal lungs increased to the peak level at Day 5 (48%, P less than 0.001) and then declined at Day 7 of O3 exposure but was still higher than controls (17%, P less than 0.05). When animals, after 5 days of O3 exposure, were allowed to recover in filtered room air, the activities of both enzymes declined to their respective control values in 6 days. These results suggest a possible temporal relationship between O3 injury and the activities of polyADPR synthetase and a free radical scavenging enzyme, SOD. The stimulation of polyADPR synthetase activity with O3 exposure, reflecting a response to lung cellular DNA repair, may be a sensitive indicator for assessing DNA damage in oxidant injury.

Control of Procollagen Gene Transcription and Prolyl Hydroxylase Activity by Poly(ADP-Ribose)

The synthesis of collagen increases during induced injury, growth and wound healing. There is an emerging consensus that collagen synthesis is controlled at the level of collagen gene transcription. The synthesis of specific collagen types in keloids (Bauer et al, 1986), virus transformed fibroblasts (Adams et al, 1982; Parker et al, 1979) and transforming growth factor-s (TGF-s) or interleukin-1 (IL-1) treated fibroblasts (Ignotz et al, 1987; Postlethwaite et al, 1988) is accompanied by an increase in the levels of the respective procollagen mRNAs. However, molecular events leading to this stimulation is not clear. Poly(ADPribose) is closely associated with the differentiation of several eukaryotic cells (Farzaneh et al, 1982; Ohashi et al, 1985) and is also implicated in the expression of individual genes (Kun et al, 1986; Poirer et al, 1982). (Tanuma et al, 1983) showed a clear relationship between ADP-ribosylation of HMG 14/HMG 17 and glucocorticoid regulated expression of MMTV gene. The use of 3-aminobenzamide (3-AB), lowered poly(ADP-ribose) levels on these nuclear proteins and enhanced the level of MMTV mRNA. Similarly, the expressions of c-myc and c-fos proto-oncogenes were stimulated by the treatment of fibroblasts with 3-methoxybenzamide (McNerney et al, 1987). Earlier, we found that exposure of fibroblasts to 20 mM lactate lowered [NAD+], total ADP-ribosylation and activated prolyl hydroxylase causing increased collagen synthesis (Hussain et al, 1989).

Regulatory Aspects of Neovascularization

Angiogenesis is a critical component of tissue repair and involves a series of interconnected, interdependent events, among which are generation and reception of angiogenic signals, chemotaxis, proteolysis of extracellular matrix, cell replication, cell-matrix adhesion, tube formation, and ligation of the newly formed vascular sprouts. The discovery of angiogenic cytokines and growth factors has greatly contributed to understanding ofthe regulation of angiogenesis (1–3). Although many angiogenic substances have been found that potentially upregulate most of the above mentioned angiogenic events, no unifying postulate for their synthesis or their mode of action has emerged. Perhaps a search for how these substances are elicited may prove to be a fruitful alternative. Only recently have research efforts touched on how the initiation of angiogenesis could be linked to the metabolic state (4).