Mucus glycoprotein secretion by duodenal mucosa in response to luminal arachidonic acid

The effect of luminal application of arachidonic acid on the alkaline secretion, prostaglandin generation, and mucus glycoprotein output and composition was studied in proximal and distal duodenum of conscious dogs. Surgically prepared duodenal loops were instilled in vivo for up to 2 h with saline (control) followed by various concentrations (12.5–100 μg/ml) of arachidonic acid. The experiments were conducted with and without intravenous pretreatment with indomethacin. The recovered instillates were assayed for the content of prostaglandin and HCO₃⁻, and used for the isolation of mucus glycoprotein. Exposure of duodenal mucosa to arachidonic acid led to concentration-dependent increase in the output of HCO₃⁻ and prostaglandin generation. In both cases this response was greater in the proximal duodenum. Pretreatment with indomethacin caused reduction in the basal HCO₃⁻ and prostaglandin output, and prevented the increments evoked by arachidonic acid. The proximal and distal duodenum displayed similar basal output and composition of mucus glycoprotein. Comparable increases in these glycoproteins were also obtained with arachidonic acid, the effect of which was abolished by indomethacin. Compared to basal conditions, mucus glycoproteins elaborated in response to arachidonic acid exhibited higher contents of associated lipids and covalently bound fatty acids, and contained less protein. The associated lipids of mucus glycoproteins elaborated in the presence of arachidonic acid showed enrichment in phospholipids and decrease in neutral lipids. The carbohydrate components in these glycoproteins also exhibited higher proportions of sialic acid and sulfate. The changes brought about by arachidonic acid were prevented by indomethacin pretreatment, and in both cases the glycoprotein composition returned to that obtained under basal conditions. The enrichment of mucus glycoprotein in lipids, sialic acid and sulfate in response to endogenous prostaglandin may be of significance to the function of this glycoprotein in the hostile environment of the duodenum.     

The effects of lipoxygenase metabolites of arachidonic acid on human myometrial contractility

The effects of the lipoxygenase products of arachidonic acid, 5- and 12-hydroxyeicosatetraenoic acid (5- and 12-HETE) and leukotriene B₄ (LTB₄), on the spontaneous contractility of lower uterine segment human myometrial strips obtained prior to labour have been studied . 5-HETE gave a dose- dependent (10–500ng) increase in both the rate of contractions and overall contractility of myometrial strips while 12-HETE and LTB₄ had no effect at the same concentrations. Prostaglandin F₂ (50ng) contracted all myometrial strips in a similar pattern to 5-HETE but was approximately 10 times more potent. The effect of 5-HETE may be direct or perhaps indirect via interaction with the cyclo-oxygenase pathway. The findings do not disprove the contention that the onset of parturition may be characterized by a switch in arachidonic acid metabolism in intra-uterine tissues from lipoxygenase to cyclo-oxygenase products.     

Parallel inhibition of neutrophil arachidonic acid metabolism and lysosomal enzyme secretion by nordihydroguaiaretic acid

Arachidonic acid at concentrations from 0.2 to 2.0 × 10?⁶M induces the secretion of lysosomal enzymes from cytochalasin B-treated rabbit neutrophils. These concentrations of arachidonic acid are metabolized primarily to hydroxyeicosatetraenic acids rather than to cyclooxygenase products. A good correlation is observed between the extent of arachidonic acid metabolism and the secretion of lysosomal enzymes. Nordihydroguaiaretic acid (1?10μM) inhibits both lysosomal enzyme secretion and the production of lipoxygenase products by neutrophils.     

Effect of Bicuculline-Induced Status Epilepticus on Prostaglandins and Hydroxyeicosatetraenoic Acids in Rat Brain Subcellular Fractions

Abstract: Rat cerebrum, prelabeled in vivo by intraventric-ular injection of [1-¹⁴C]arachidonic acid, was used to assess cyclooxygenase and lipoxygenase reaction products in total homogenates, cytosol, synaptosomes, and microsomes. Effects of bicuculline-induced status epilepticus on arachi-donic acid metabolism in synaptosomes and microsomes were also measured. Lipoxygenase activity, resulting in the synthesis of hydroxyeicosatetraenoic acids (HETEs), and cyclooxygenase activity, resulting in the synthesis of prostaglandins (PGs), were measured by reverse-phase and normal-phase HPLC with flow scintillation detection. Endogenous lipoxygenase products in synaptosomes were identified by capillary gas chromatography-mass spectrometry. PGs and HETEs were detected in all subcellular fractions. The synaptosomal fraction showed the highest lipoxygenase activity, with 5-HETE, 12-HETE, and leukotriene B₄ as the major products. Following bicuculline-induced status epilepticus, endogenous free arachidonic acid and other fatty acids accumulated in synaptosomes, but not in microsomes. Incorporation of [1-^l4C]arachidonic acid into synaptosomal and microsomal phospholipids was decreased after bicuculline treatment. Bicuculline-induced status epilepticus resulted in increased synthesis of HETEs in synaptosomes. PG synthesis increased in the microsomal fraction. When [1-¹⁴C]arachidonic acid-labeled synaptosomes and microsomes were incubated for 1 h at 37°C the synthesis of eicosa-noids, particularly PGD₂, was increased significantly in bi-cuculline-treated rats, as compared with untreated rats. Depolarization (45 mM K⁺) of synaptosomes induced a loss of [1-¹⁴C]arachidonic acid from phosphatidylinositol, and increased the synthesis of PGD₂ and HETEs, an effect that was enhanced in bicuculline-treated rats. This study localizes changes in arachidonic acid metabolism and lipoxygenase activity resulting from bicuculline-induced status epilepticus in the brain subcellular fraction enriched in nerve endings.     

Effects of prostaglandin E2, 16,16-dimethyl prostaglandin E2 and a prostaglandin endoperoxide analogue (U-46619) on gastric secretory volume, [H] and mucus synthesis and secretion in the rat

The effects of orally administered prostaglandin E₂, 16,16-dimethyl prostaglandin E₂ and U-46619, an analogue of the prostaglandin endoperoxide PGH₂, on gastric secretory volume, acid and mucus were studied in the rat. All of the compounds significantly increased the volume of gastric secretion, mucus secretion, measured as N-acetylneuraminic acid and mucus synthesis measured as the incorporation of [³H]-glucosamine into mucosal glycoprotein; however, only PGE₂ and 16,16-dimethyl PGE₂ inhibited acid secretion. U-46619, 1.5 mg/kg provided significant protection against ethanol-induced gastric ulcers, an effect that has been previously shown for the other two compounds. These studies provide additional evidence that prostaglandin induced mucosal protection may by related to an effect on mucus and on stimulation of nonparietal cell gastric secretion. Further study of these parameters may be important in the development of antiulcer drugs for long term clinical use.     

Metabolism of 7,10,13,16-docosatetraenoic to dihomo-thromboxane 14-hydroxy-7,10,12-nonadecatrienoic acid hydroxy fatty acids by human platelets

Human platelets metabolize 7,10,13,16-docosatetraenoic acid (22:4(n−6) into dihomo-thromboxane B₂ and 14-hydroxy-7,10,12-nonadecatrienoic acid at about twenty percent of the rate they convert arachidonic to thromboxane B₂ and 12-hydroxy-5,8,10-heptadecatrienoic acid. 14-Hydroxy-7,10,12,16-docasatetraenoic was the major metabolite produce via the lipoxygenase pathway. Several other hydroxy were also produced in small amounts via an indomethacin-insensitive pathway. Incubation of 20 μM arachidonic acid with various levels of 22:4(n−6) resulted In a dose-dependent inhibition of both thromboxane B₂ and 12-hydroxy-5,8,10-heptadecatrienoic acid production. Coversely, 12-hydroxy-5,8,10,14-eicosatetraenoic acid synthesis was stimulated because of substrate shunting to the lipoxygenase pathway. These results show that 22:4(n−6) may modify platelet function both by serving as a precursor for a 22-carbon thromboxane and by suppressing the synthesis of thromboxane A₂ from arachidonic acid. In addition, our results suggest that simultaneous release of 22:4(n−6) and arachidonic acid from platelet phospholipids will result in an elevation of both 12-hydroxy-5,8,10,14-eicosatetraenoic acid levels as well as simultaneous synthesis of 14-hydroxy-7,10,12,16-docosatetraenoic acid.     

Stimulation of human T-lymphocyte chemokinesis by arachidonic acid.

The migration of human T lymphocytes, assessed in modified Boyden chambers, was chemokinetically stimulated by arachidonic acid in a dose-related manner that achieved a peak level of 127 ± 34% enhancement (mean ± SD) at 8 μM arachidonic acid. The chemokinetic effect was dependent on the metabolism of the arachidonic acid by the T lymphocytes as derivatives of arachidonic acid that do not serve as prostaglandin and thromboxane precursors were without effect, while the cyclo-oxygenase inhibitors indomethacin (ID₅₀ = 10 μM) and 5,8,11,14-eicosatetraynoic acid (ETYA) (ID₅₀ = 20 μM) suppressed the stimulation of migration by arachidonic acid. The cyclo-oxygenase product 12-l-hydroxy-5,8,10-heptadecatrienoic acid (HHT) reproduced part of the chemokinetic effect of arachidonic acid, but the lipoxygenase product 12-l-hydroxy-5,8,10,14-eicosatetraenoic acid (HETE) as well as PGE₂, PGF_2α, and thromboxane B₂ had no stimulatory activity. The ability of ETYA, but not indomethacin, to suppress the migration of unstimulated T lymphocytes suggested that a lipoxygenase metabolite of endogenous arachidonic acid contributes to the maintenance of their normal levels of spontaneous migration.     

Demonstration and maintenance of mucus secretion in cultured human gallbladder epithelial cells

Summary The method of human gallbladder epithelial cell culture has been developed successfully with active mucus secretory function. Human gallbladder epithelial cells were dissociated by Dispase digestion from the specimens obtained by cholecystectomy for uncomplicated gallbladder stone cases. The dissociated cells formed a monolayer in Eagle’fs minimum essential medium supplemented with 10% fetal bovine serum within 24 h after the inoculation. These cells were maintained for at least 2 wk without fibroblastic overgrowth. Cultured cells contained periodic acid Schiff-positive material in cellular cytoplasm for 3 d. On transmission electron microscopy these materials were identified as mucous secretory granules. Mucous secretory function was determined by [³H]glucosamine incorporation. Sixty percent of the secreted glycoproteins labeled with [³H]glucosamine was eluted in excluded fractions of Sepharose 4B gel filtration, which were considered to be mucous glycoprotein, because they were found to be resistant to proteoglycan-specific enzymes such as hyaluronidase, chondroitinase ABC, heparitinase, and heparinase. The mucous glycoprotein secretion was maintained for 3 d and found to be inhibited in a dose-dependent manner by monensin (10⁻⁷ to 10⁻⁵ M) which is a known blocker of secretory function.     

Arachidonic acid metabolim in rat pancreatic acinar cells: Calcium-mediated stimulation of the lipoxygenase system

Isolated rat pancreatic acini were employed to demonstrate that the exocrine pancreas can metabolize [¹⁴C]-arachidonic acid by way of the lipoxygenase pathway as well as the cyclooxygenase pathway. Analysis by high performance liquid chromtography delineated a monohydroxy acid, presumably 12-L-hydroxy-5,8–10,14-eicosatetraenoic acid (12-HETE) as the major lipoxygenase product. The formation of this hydroxy arachidonic derivative was stimulated by the calcium ionophore ionomycin. Stimulation of lipoxygenase pathway by ionomycin was confirmed by thin layer chromatography. In addition, 6-keto-PGF_1α, PGF_2α, and PGE₂ were identified; and ionomycin, carbamylcholine, and caerulein enhanced the formation of these metabolites of the cyclooxygenase pathway. Ionomycin induced stimulation of HETE formation was inhibited by ETYA and nordihydroguaiaretic acid, but spontaneous and evoked enzyme secretion was unaffected. Thus, although ionomycin, a pancreatic secretagogue, stimulates the lipoxygenase pathway, the precise role of these arachidonate metabolites in the physiology of the exocrine pancreas is still obscure.     

Effect of caffeine on mucus secretion and agonist-dependent Ca2+ mobilization in human gastric mucus secreting cells

Caffeine is known to stimulate gastric acid secretion, but, the effects of caffeine on gastric mucus secretion have not been clarified. To elucidate the action of caffeine on gastric mucin-producing cells and its underlying mechanism, the effects of caffeine on mucus glycoprotein secretion and agonist-induced [Ca²⁺]_i mobilization were examined in human gastric mucin secreting cells (JR-I cells). The measurement of [Ca²⁺]_i using Indo-1 and the whole cell voltage clamp technique were applied. Mucus glycoprotein secretion was assessed by release of [³H]glucosamine. Caffeine by itself failed to increase [Ca²⁺]_i and affect membrane currents, while it dose-dependently inhibited agonist (acetylcholine (ACh) or histamine)-induced [Ca²+]_i rise, resulting in inhibiting activation of Ca²⁺-dependent K⁺ current (I_K.Ca) evoked by agonists. The effect of caffeine was reversible, and the half maximal inhibitory concentration was about 0.5 mM. But, caffeine did not suppress [Ca²⁺]_i rise and activation of I_K.Ca induced by A23187 or inositol trisphosphate (IP₃). Theophylline or 3-isobutyl-1-methyl-xanthine (IBMX) did not mimic the effect of caffeine. Caffeine failed to stimulate mucus secretion, while it significantly decreased ACh-induced mucus secretion. These results indicate that caffeine selectively inhibits agonist-mediated [Ca²⁺]_i rise in human gastric epithelial cells, probably through the blockade of receptor-IP₃ signaling pathway, which may affect the mucin secretion. © 1997 Elsevier Science B.V. All rights reserved.     

Increased pulmonary vascular resistance and permebility due to arachidonate metabolism in isolated rabbit lungs

Liberation and metabolism of arachidonic acid may be the common final pathway of different stimuli on the pulmunary vascular bed. In a model of isolated, ventilated rabbit lungs, perfused with krebs Henseleit albumin buffer in a recirculating system, changes of pulmonary vascular resistance and of vascular permeability are monitored continously. The addition of free arachidonic acid or of the Ca-ionophore A 23187 to the perfusion fluid consistently evokes a biphasic increases in vascular resistance as well as an initially reversible increase in vascular permeability, followed by pulmonary edema. Both phases of increased vascular resistance are completely suppressed by inhibition of the cyclooxygenase, decreased to a large degree by inhibitors of thromnoxane synthetase, and markedly augmented by short preincubation of arachidonic acid with ram seminal vescular microsomes and by sulfhydryl reagents. The increased pulmonary vascular permeability is augmented by inhibition of cyclooxygenase and reduced by simulteneous lipoxygenase inhibition. Antagonists of histamine, serotonin and sympathic or parasympathic activity do not have any influence.PG F_2α, TxB E₂ and PG I₂ alter the pulmonary vascular resistance, but do not increase vascular permeability.In inclusion, increased availability of free arachidonic acid evokes a rise in pulmonary vascular resistance, which can be ascribed to cyclooxygenase products, especially to thromboxane, and causes a rise in vascular permeability which can be ascribed to lipoxygenase products.The findings may be related to acute pulmonary lesions with increase in vascular resistance and with vascular leakage.     

The effect of arachidonic acid and prostanoids on collagen dissolution in human uterine cervix in vitro

Explants of human non-pregnant cervix produce collagenolytic enxymes which degrade collagen over a 10 day period in culture. This is significantly enhanced by the presence of very low concentrations of arachidonic acid (10⁻¹⁶−10⁻¹¹M). Prostaglandin E₂, F_2α and 6-keto-F_1α were synthesised in declining amounts over the 10 day period and synthesis was not increased by adding arachidonic acid (10−11M). Meclofenamic acid (10⁻⁶M) and indomethacin (10⁻⁵M), but not tranylcypromine (10⁻⁵) suppressed prostaglandin synthesis yet all reduced collagen dissolution. Mepacrine (phospholipase A₂ inhibitor) also suppressed collagen dissolution. Remodelling of the structure of the cervix matrix may, in part, depend upon arachidonic acid or one of its cyclo-oxygenase or lipoxygenase derived products.     

Effect of prolactin on the secretion of milk casein: metabolism of arachidonic acid

Mammary gland fragments were incubated in the presence of prolactin and arachidonic acid which stimulate casein secretion. The effects of these stimuli in the presence of agents that influence arachidonic acid metabolism were investigated. Chloroquine, a blocker of phospholipase A2 activity, decreased prolactin but not arachidonic acid stimulation of casein secretion. Phospholipase A2 markedly stimulated casein secretion. Nordihydroguaiaretic acid (NDGA), an antioxidant that inhibits lipoxygenase, blocked the stimulating effect of prolactin and arachidonic acid. Ultrastructural studies indicated that phospholipase A2-induced stimulation of secretion was comparable to that of prolactin but that arachidonic acid-induced stimulation did not involve the same Golgi membrane modifications. These studies suggest that prolactin and phospholipase A2 stimulate secretion by a common way, and that arachidonic acid interferes with secretion by metabolic products of the lipoxygenase pathway.     

Effects of iron on rainbow trout gill cells in primary culture

This study investigated the effects of iron in the form of iron sulphate (FeSO₄·7H₂O), over the range 0.01–1 mM on rainbow trout primary gill cells cultured on semi-permeable membranes. The endpoints measured were cell proliferation, mucous cell numbers, area of mucus in mucous cells, ultrastructural analysis and transepithelial resistance. Regardless of the concentration, FeSO₄ did not modify the apical surface of pavement cells (microridge) and mucous cells. However, at 1 mM, this metal reduced cell numbers, by inhibiting cell proliferation and causing cell death, and induced a decrease in transepithelial resistance. It is interesting to note that cell numbers were also reduced in the presence of 0.5 mM iron salt, although this reduction did not modify transepithelial resistance. FeSO₄ reduced mucous cell number but did not change mucus area in mucous cells suggesting that this metal could induce a discharge of mucous cells, but mucus secretion would be total and not partial. In conclusion, our in vitro model has allowed to study some toxic effect but also resistance of gill epithelium in presence of iron.     

15-Hydroxyeicosatetraenoic Acid Inhibits Superoxide Anion Generation by Human Neutrophils: Relationship to Lipoxin Production

Human neutrophils can aggregate, degranulate, and release mediators of inflammation including oxygen radicals and lipoxygenase (LO)-derived products of arachidonic acid. The regulation of 5– and 15-lipoxy-genases appears to be important since their products (e.g. leukotrienes and lipoxins) display unique spectra of bioactions. Addition of 15-HETE. a product of the 15-LO, to neutrophils in suspension dramatically shifted the LO products generated and led to a dose-dependent increase in lipoxins, while the production of leukotriene B₄ and its μ-oxidation products (i.e. 20-COOH-LTB₄ and 20-OH-LTB₄) was inhibited. Exogenous 15-HETE also dose-dependently inhibited the generation of superoxide anions induced by either the chemotactic peptide f-met-leu-phe or the divalent cation ionophore A23187. Neither lipoxin A, nor lipoxin B₄ (10^?8?10^?6M) inhibited O₂^?_? generation induced by either f-met-leu-phe or A23187. These results indicate that in addition to serving as a substrate for lipoxin generation, 15-HETE also inhibits superoxide anion generation by human neutrophils. Together they provide further evidence to suggest that products of the 15-lipoxygenase may serve a regulatory role at inflammatory loci.     

Teteraene and pentaene leukotrienes: Selective production from murine mastocytomo cells after dietary manipulation

A neoplastic mast cell tumor was grown in mice which had been raised since birth on a diet enriched with eicosapentaenoic acid. Intact harvest mastocytoma cells were stimulated with calcium ionophpore A23187 to produce lipoxygenase products from the polyunsaturated fatty acids liberated from the cellular membranes. Leukotriene B₄, B₅, C₄ and C₅ were isolated and characterized by HPLC retention time, ultraviolet absorption spectrometry and mass spectrometry. The arachidonic acid content of the mast cell tumor lipids was altered from 9.2 to 3.9 mole% while eicosapentaenoic acid increased from 0.5 to 4.5 mole % in response to the fish oil-supplement diet.The relative amount of arachidonic and eicosapentaenoic acids (3.9 and 4.5 mole % respectively) were associated with similar amounts of LTB₄ and LTP₅ synthesized by the cells. These results suggest that the epoxide leukotrine (LTA) derivative can be made efficiently from either arachidonic or eicosapentaenoic acids when both are present in cellular lipids. In contrast, the ratio of LTC₄ to LTC₅ (10 to 1) indicates that the reaction of LTA with glutathione may be critically dependent upon the structure of the unsaturated fatty acid with the ratio of LTC₄/LTB₄ (2.0) more than 10 times greater than that (0.16) for LTC₅/LTP₅.     

Effects of inhibitors of eicosanoid synthesis on insulin release by neonatal pancreatic islets

In the pancreatic islet, eicosanoids may arise from both cyclooxygenase- and lipoxygenase-dependent metabolism of arachidonic acid. The inclusion of inhibitors of selective steps in these pathways indicated that in cultured neonatal rat islets, arachidonic acid may be metabolised through both pathways, concurrent with insulin release stimulated by D-glucose, D-glyceraldehyde and 2-ketoisocaproate. The effects of the inhibitors suggested that the products of the lipoxygenase pathway were necessary for the stimulatory effects of nutrients to be observed. In contrast to glucose, where insulin release was stimulated in the presence of inhibitors of cyclooxygenase, the stimulatory action of D-glyceraldehyde, 2-ketoisocaproate and melittin was only minimally affected by these inhibitors, although it was inhibited by lipoxygenase inhibition. These findings support a major stimulatory role for products of the lipoxygenase pathway of arachidonic acid metabolism in nutrient-induced secretion, and a negative or modulatory role of cyclooxygenase pathway products on glucose-stimulated insulin release in the neonatal islet.     

In vivo metabolism of dihomo-γ-linolenic acid to bronchoactive products in the canine lung

We investigated the effects of dihomo-γ-linolenic acid (DGLA), the fatty acid precursor to the monoenoic prostaglandins, on pulmonary mechanics in the intact chest, artificially ventilated dog. Under conditions of normal airway tone, intravenously administered DGLA produced a modest, dose-related increase in lung resistance and decrease in dynamic lung compliance. These responses were approximately 30–100 times less than those produced by arachidonic acid. The bronchoconstrictive responses to DGLA were abolished by prior inhibition of the cyclooxygenase with indomethacin. PGD₁ was equal in activity to PGF_1α in causing constriction of central and peripheral airways of the dog. PGE₁ was without significant airway effects in these animals under conditions of resting airway tone. We conclude that DGLA is a moderately good substrate for the cyclooxygenase pathway enzymes of the canine lung, producing products having a predominantly bronchoconstrictive effect. This bronchoconstriction is most likely due to the synthesis of PGD₁ and PGF_1α and is not due to the synthesis of lipoxygenase pathway products.     

Glomeruli cooperate with macrophages in converting arachidonic acid to prostaglandins and hydroxyeicosatetraenoic acids

The interaction of proliferating macrophages with the glomerulus may play an important role in certain forms of glomerulonephritis. This interaction could involve metabolites of arachidonic acid (C20:4) such as prostaglandins (PG) and lipoxygenase products. We therefore investigated the conversion of exogenous |³H| C20:4 into hydroxyeicosatetraenoic acids (HETE) and PG by isolated glomeruli and macrophages from rats, alone and in combination. As demonstrated by HPLC, glomeruli converted C20:4 predominantly to lipoxygenase products -mainly 12-HETE- and, to a lesser extent, to PG. Resident macrophages converted C20:4 to equivalent amounts of HETE and PG, mainly 12-HETE and 6 keto-PGF_1α. When macrophages and glomeruli were studied in combination, a striking interaction was detected in both pathways of C20:4 metabolism. Production of 6 keto-PGF_1α was stimulated and considerable amounts of TXB₂, PGD₂ and hydroxyheptadecatrienoic acid (HHT) were also produced Total 12-HETE production was unchanged. When a lipid extract of glomeruli, containing oxygenated metabolites of C20:4, was added to macrophages, stimulation of 12-HETE occurred without any change in HHT or PG formation. When, on the contrary, a lipid extract from macrophages was added to glomeruli, 12-HETE production by the glomeruli was nearly completely abolished. Thus the unchanged total 12-HETE production by coincubated glomeruli and macrophages resulted from its increased production by macrophages and its decreased production by glomeruli. These data suggest that interaction between glomeruli and macrophages results in activation of C20:4 metabolism by macrophages followed by inhibition of C20:4 metabolism by glomeruli. Such a regulatory process could play a role in the inflammatory response to immunological injuries during macrophage-dependent human and experimental glomerulonephritis.     

Control of mucin molecular forms expression by salivary protease: Differences with caries

• 1.1. A protease activity capable of degradation of the high mol. wt salivary mucus glycoprotein to a low mol. wt glycoprotein form was identified in human submandibular gland secretion.
• 2.2. The protease exhibited optimum activity at pH 7.0–7.4, and gave on SDS-PAGE under reducing conditions two major protein bands of 48 and 53 kDa. The enzyme showed susceptibility to PMSF, α₁antitrypsin, and egg white and soybean inhibitors, a characteristic typical to serine proteases.
• 3.3. The activity of the protease towards the high mol. wt mucus glycoprotein was found to be 3.8-fold higher in submandibular gland secretion of caries-resistant individuals than that of caries-susceptible. Furthermore, the enzyme from both groups displayed greater activity against the mucus glycoprotein of caries-resistant subjects.
• 4.4. Since the low mol. wt salivary mucus glycoprotein form is more efficient in bacterial clearance than the high mol. wt mucin, the enhanced expression of this indigenous salivary protease activity towards mucin may be the determining factor in the resistance to caries.