Inflammatory changes in the dental pulp are accompanied by release of a wide variety of chemical mediators. Nitric oxide, an oxidative free radical produced by the enzyme nitric oxide synthase (NOS), has been implicated in multiple inflammatory processes, which makes it a suitable marker for changes which likely occur following tooth pulp insult. Since limited information on nitric oxide in the pulp is available, it is necessary first to examine relative distributions of NOS in uninflamed and inflamed rat pulp. We accomplished this by characterizing regions of nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) activity and the distribution of both macrophage NOS (macNOS) and neuronal NOS (nNOS) immunoreactivity in normal and inflamed rat molar pulp at multiple time points. The results showed that: (1) deep cavity preparation on the mesial surface of the molar produced a time-dependent inflammation, with acute inflammation early progressing to chronic, granulomatous inflammation with necrosis later that spread preferentially down the mesial root; (2) control (non-prepared) teeth showed a relatively faint and homogeneous distribution of NADPH-d and macNOS reactivity but no discernible nNOS reactivity; (3) inflamed teeth displayed localized increased intensity of NADPH-d and macNOS reactivity surrounding the inflamed area of pulp, but no increased nNOS activity; (4) pulp vessels supplying the inflamed area showed increased NADPH-d reactivity, but no increased macNOS or nNOS reactivity; and (5) neither NADPH-d, macNOS, nor nNOS reactivity was observed in pulpal nerves. Therefore, nitric oxide may mediate the pulpal inflammatory response through its effects on the paralesional pulp tissue and surrounding endothelial/vascular structures.
Ahlberg KF (1978). Influence level of noxious heat stimulation on sensory nerve activity in the feline tooth pulp. Acta Physiol Scand103:71-80.
2.
Aimi Y., Fujimura M., Vincent SR, Kimura H. (1991). Localization of NADPH-diaphorase-containing neurons in sensory ganglia of the rat. J Comp Neurol306:382-392.
3.
Albina JE, Henry WL Jr (1991). Suppression of lymphocyte proliferation through the nitric oxide synthetase pathway. J Surg Res50:403-409.
4.
Ashwal S., Cole DJ, Osborne TN, Pearce WJ (1994). Dual effects of L-NAME during transient focal cerebral ischemia in spontaneously hypertensive rats. Am J Physiol267:H276-H284.
5.
Bergenholtz G., Ahlsted S., Lindhe J. (1977). Experimental pulpitis in immunized monkeys. Scand J Dent Res85:396-406.
6.
Bergenholtz G., Nagaoka S., Jontell M. (1991). Class II antigen presenting cells in experimentally induced pulpitis. Int Endodont J24:8-14.
7.
Blanco CE, Sieck GC, Edgerton VR (1988). Quantitative histochemical determination of succinic dehydrogenase activity in skeletal muscle fibbers. Histochem J20:230-243.
8.
Bredt DS, Hwang PM, Snyder SH (1990). Localization of nitric oxide synthase indicating a neural role for nitric oxide. Nature347:768-770.
9.
Bredt DS, Glatt CE, Hwang PM, Fotuhi M., Dawson TM, Snyder SH (1991). Nitric oxide synthase protein and mRNA are discretely localized in neuronal populations of the mammalian CNS together with NADPH diaphorase. Neuron7:615-624.
10.
Brodin E., Gazelius B., Olgart L., Nilsson G. (1981). Tissue concentration and release of substance P-like immunoreactivity in the dental pulp. Acta Physiol Scand111:141-149.
11.
Busse R., Mulsch A. (1990). Induction of nitric oxide synthase by cytokines in vascular smooth muscle cells. FEBS Lett275 :87-90.
12.
Cole M., Bond CP ( 1972). Recent advances in automatic image analysis using a television system. J Microsc96:89-96.
13.
D'Souza RN, Bachman T., Baumgardner KR, Butler WT, Litz M. ( 1995). Characterization of cellular responses involved in reparative dentinogenesis in rat molars. J Dent Res74 :702-709.
14.
Dawson TW, Bredt DS, Fotuhi M., Hwang PM, Snyder SH (1991). Nitric oxide synthase and neuronal NADPH diaphorase are identical in brain and peripheral tissues. Proc Natl Acad Sci USA88:7797-7801.
15.
DelBalso AM, Nishimura RS, Setterstrom JA (1976). The effects of thermal and electrical injury on pulpal histamine levels. Oral Surg41:110-113.
16.
Diaz-Araya CM, Provis JM, Billson FA (1993). NADPH-diaphorase histochemistry reveals cone distributions in adult human retinae. Aust NZ J Ophthalmol21:171-179.
17.
Fischer A., Mundell P., Mayer B., Preissler U., Philippin B., Kummer W. (1993). Nitric oxide synthase in guinea pig lower airway innervation. Neurosci Lett149:157-160.
18.
Gabbott PLA, Bacon SJ (1993). Histochemical localization of NADPH dependent diaphorase (nitric oxide synthase) activity in vascular endothelial cells in the rat brain. Neuroscience57:79-95.
19.
Gazelius B., Edwall B., Olgart L., Lundberg JM, Hokfelt T., Fischer JA (1987). Vasodilatory effects and coexistence of calcitonin gene-related peptide (CGRP) and substance P in sensory nerves of cat dental pulp. Acta Physiol Scand130:33-40.
20.
Grozdanovic Z., Baumgarten HG, Bruning G. (1992). Histochemistry of NADPH-diaphorase, a marker for neuronal nitric oxide synthase, in the peripheral autonomic nervous system of the mouse. Neuroscience48:225-235.
21.
Grutzner EH, Garry MG, Hargreaves KM (1992). Effect of injury on pulpal levels of immunoreactive substance P and immunoreactive calcitonin gene-related peptide. J Endodont18:553-577.
22.
Hahn CL, Falkler WA Jr, Siegel MA (1989). A study of T and B cells in pulpal pathosis. J Endodont15:20-26.
23.
Hibbs JB Jr, Taintor RR, Vavrin Z., Rachlin EM (1987). Nitric oxide; a cytotoxic activated macrophage effector molecule. Biochem Biophys Res Commun157:87-94.
24.
Hirafuji M., Ogura Y. (1987). 5-Hydroxytryptamine stimulates the release of prostacyclin but not thromboxane A2 from isolated rat dental pulp. Eur J Pharmacol136:433-436.
25.
Hirafuji M., Satoh S., Ogura Y. (1980). Prostaglandins in rat pulp tissue. J Dent Res59:1535-1540.
26.
Hirafuji M., Terashima K., Satoh S., Ogura Y. (1982). Stimulation of prostaglandin E2 biosynthesis in rat dental pulp explants in vitro by 5-hydroxytryptamine. Arch Oral Biol27:961-964.
27.
Hope BT, Michael GJ, Knigge KM, Vincent SR (1991). Neuronal NADPH diaphorase is a nitric oxide synthase . Proc Natl Acad Sci USA88:2811-2814.
28.
Iadecola C., Beitz AJ, Renno W., Xu X., Mayer B., Zhang F. (1993). Nitric oxide synthase-containing neural processes on large cerebral arteries and cerebral microvessels. Brain Res606:148-155.
29.
Ignarro LJ, Buga GM, Wood KS, Byrns RE, Chaudhuri G. (1987). Endothelium-derived relaxing factor produced and released from artery and vein is nitric oxide. Proc Natl Acad Sci USA84:9265-9269.
30.
Janssens SP, Shimouchi A., Quertermous T., Bloch DB, Bloch KD (1992). Cloning and expression of a cDNA encoding human endothelium-derived relaxing factor/nitric oxide synthase. J Biol Chem267:14519-14522.
31.
Kerezoudis NP, Olgart L., Edwall L. (1993a). Differential effects of nitric oxide synthesis inhibition on basal blood flow and antidromic vasodilation in rat oral tissues . Eur J Pharmacol241:209-219.
32.
Kerezoudis NP, Olgart L., Fried K. (1993b). Localization of NADPH-diaphorase activity in the dental pulp periodontium and alveolar bone of the rat. Histochemistry100:319-322.
33.
Khayat BG, Byers MR, Taylor PE, Mecifi K., Kimberly CL (1988). Responses of nerve fibers to pulpal inflammation and peri-apical lesions in rat molars demonstrated by calcitonin gene-related peptide immunocytochemistry. J Endodont14:577-587.
34.
Kilbourn RG, Belloni P. (1990). Endothelial cell production of nitrogen oxides in response to interferon in combination with tumor necrosis factor, interleukin-1, or endotoxin. J Natl Canc Inst82:772-776.
35.
Kilbourn RG, Jubran A., Gross SS, Griffith OW, Levi R., Adams J., et al. (1990). Reversal of endotoxin-mediated shock by NGmethyl-L-arginine, an inhibitor of nitric oxide synthase. Biochem Biophys Res Commun172:1132-1138.
36.
Kummer W., Fischer A., Mundel P., Mayer B., Hoba B., Philippin B., et al. (1992). Nitric oxide synthase in VIP-containing vasodilator nerve fibres in the guinea-pig. Neuroreport3:653-655.
37.
Lamas S., Marsden PA, Li G-K., Tempst P., Michel T. (1992). Endothelial nitric oxide synthase: molecular cloning and characterization of a distinct constitutive enzyme form. Proc Natl Acad Sci USA89:6348-6352.
38.
Lohinai Z., Balla I., Marczis J., Vass Z., Kovach AG (1995). Evidence for the role of nitric oxide in the circulation of the dental pulp. J Dent Res74:1501-1506.
39.
Lohinai Z., Szekely AD, Benedek P., Csillag A. (1997). Nitric oxide synthase containing nerves in the cat and dog dental pulp and gingiva. Neurosci Lett227:91-94.
40.
MacNaul KL, Hutchinson NI (1993). Differential expression of iNOS and cNOS in human vascular smooth muscle cells and endothelial cells under normal and inflammatory conditions. Biochem Biophys Res Commun196:1330-1334.
41.
Madison S., Whitsel EA, Suarez-Roca H., Maixner W. (1992). Sensitizing effects of leukotriene B4 on intradental primary afferents. Pain49:99-104.
42.
Marletta MA (1989). Nitric oxide: biosynthesis and biological significance . Trends Biochem Sci14:488-492.
43.
McCall TB, Palmer RM, Moncada S. (1991). Induction of nitric oxide synthase in rat peritoneal neutrophils and its inhibition by dexamethasone. Eur J Immunol21:2523-2527.
44.
Moncada S., Palmer Rmj, Higgs EA (1991). Nitric oxide: physiology, pathology, and pharmacology . Pharmacol Rev43 :109-142.
45.
Mulligan MS, Hevel JM, Marletta MA, Ward PA ( 1991). Tissue injury caused by deposition of immune complexes is L-arginine dependent. Proc Natl Acad Sci USA88:6338-6342.
46.
Narhi Mvo, Hirvonen T. (1983). Functional changes in cat pulp nerve activity after thermal and mechanical injury to the pulp. Proc Finn Dent Soc79:162-167.
47.
Nathan CB, Hibbs JB Jr (1991). The role of nitric oxide synthesis in macrophage antimicrobial activity. Curr Opin Immunol3:65-70.
Okiji T., Morita I., Sunada I., Murota S. (1989). Involvement of arachidonic acid metabolites in increases in vascular permeability in experimental dental pulpal inflammation in the rat. Arch Oral Biol34:523-528.
50.
Olgart L., Edwall L., Gazelius B. (1991). Involvement of afferent nerves in pulpal blood-flow reactions in response to clinical and experimental procedures in the cat. Arch Oral Biol36:575-581.
51.
Oswald RJ, Byers MR (1993). The injury response of pulpal NPY-IR sympathetic fibers differs from that of sensory afferent fibers. Neurosci Lett164:190-194.
52.
Panopoulos P., Mejare B., Edwall L. (1983). Effects of ammonia and organic acids on the intradental sensory nerve activity. Acta Odontol Scand41:209-215.
53.
Parks DA, Bulkley GB, Granger DN, Hamilton SR, McCord JM (1982). Ischemic injury in the cat small intestine: role of superoxide radicals. Gastroenterol82:9-15.
54.
Picton W., Clark C. (1978). The measurement of histochemicallystained NADP dependent iscitrate dehydrogenase in the subcutaneous glands of hairless hamsters . Histochem J10:191-199.
55.
Poeggel G., Muller M., Seidel I., Rechardt L., Bernstein HG (1992). Histochemistry of guanylate cyclase, phosphodiesterase, and NADPH-diaphorase (nitric oxide synthase) in rat brain vasculature. J Cardiovasc Pharmacol20(12):S76-S79.
56.
Pohto P., Antila R. (1970). Assay of histamine in dental pulps. Acta Odontol Scand28:691-699.
57.
Pulver WH, Taubman MA, Smith DJ (1977). Immune components in normal and inflamed human dental pulp. Arch Oral Biol22:103-111.
58.
Rees DD, Palmer RM, Schulz R., Hodson HF, Moncada S. (1990). Characterization of three inhibitors of endothelial nitric oxide synthase in vitro and in vivo. Br J Pharmacol101:746-752.
59.
Sessa WC, Harrison JK, Barber CM, Zeng D., Durieux ME, D'Angelo DD, et al. (1992). Molecular cloning and expression of a cDNA encoding endothelial cell nitric oxide synthase. J Biol Chem267:15274-15276.
60.
Silverman JD, Kruger L. (1987). An interpretation of dental innervation based upon the pattern of calcitonin gene-related peptide (CGRP) immuno-reactive thin sensory axons. Somatosens Res5:157-175.
61.
Stuehr DJ, Cho HJ, Kwon NS, Weise MF, Nathan CF (1991). Purification and characterization of the cytokine-induced macrophage nitric oxide synthase: an FAD- and FMN-containing flavoprotein . Proc Natl Acad Sci USA88:7773-7777.
62.
Takahashi K. (1990). Changes in pulpal vasculature during inflammation . J Endodont16:92-97.
63.
Uddman R., Bjorlin G., Möller B., Sundler F. (1980). Occurrence of VIP nerves in mammalian dental pulps . Acta Odontol Scand38:325-328.
64.
Wakisaka S., Ichikawa H., Nishimoto T., Matsuo S., Yamamoto K., Nakata T., et al. (1984). Substance P-like immunoreactivity in the pulp-dentine zone of human molar teeth demonstrated by indirect immunofluorescence . Arch Oral Biol29:73-75.
65.
Ward SM, Xue C., Shuttleworth CW, Bredt DS, Snyder SH, Sanders KM (1992). NADPH diaphorase and nitric oxide synthase colocalization in enteric neurons of canina proximal colon. Am J Physiol263:G277-G284.
66.
Werner-Felmayer G., Werner ER, Fuchs D., Hausen A., Reibnegger G., Wachter H. (1990). Tetrahydrobiopterin-dependent formation of nitrite and nitrate in murine fibroblasts. J Exp Med172:1599-1607.
67.
Wilcox JN, Subramanian RR, Sundell CL, Tracey WR, Pollock JS, Harrison DG, et al. (1997). Expression of multiple isoforms of nitric oxide synthase in normal and atherosclerotic vessels. Arterioscl Thromb Vasc Biol17:2479-2488.
68.
Willis RA, Nussler AK, Fries KM, Geller DA, Phipps RP (1994). Induction of nitric oxide synthase in subsets of murine pulmonary fibroblasts: effect on interleukin-6 production. Clin Immunol Immunopathol71:231-239.
