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International audience; Lipid digestion is a complex process that takes place at the lipid-water interface and involves various lipolytic enzymes present predominantly in the stomach and the small intestine [34]. These enzymes catalyse the hydrolysis of a variety of dietary lipids from animal and plant sources, such as triacylglycerols (TAGs), phospholipids, galactolipids, cholesterol and vitamin esters. They include gastric lipase, colipase-dependent pancreatic lipase, pancreatic lipase-related proteins 2 (PLRP2), carboxyl ester hydrolase or bile salt-stimulated lipase (CEH, BSSL), and pancreatic phospholipase A2. A debate still exist about the existence of a lingual lipase in human [30, 86, 140], an enzyme that has been demonstrated to be present and active in rat and mice tongue only and which is the product of a gene ortholog [53] to the gene of gastric lipase [24] in humans and many other species. Bakala N’Goma et al. [12] have reviewed the key findings that support the existence of lingual or gastric lipases in several species in term of gene expression, enzyme immunocytolocalization and lipase activity. So far, no supporter of the existence of a lingual lipase in humans has been able to provide similar data.

Objectives This systematic literature review aims to summarize the existing scientific evidence on the association of oral lipolysis with diet and with the perception and digestion of lipids in humans and rodents. Methods A validated search strategy of two databases (PubMed and ISI Web of Knowledge) was carried out and the contents were screened by two independent reviewers. The quality of the included studies was critically evaluated on the basis of the Quality Assessment Criteria for Evaluating Primary Research Papers. Results From the originally identified studies (n = 2295), 17 articles met the eligibility criteria for inclusion in the analysis. Among them, only 6 articles received the maximum assessment score. The main reason for this finding was the absence of a control for the confounding bias between lipases and esterases. In rodents, oral lipolysis was principally due to the activity of lingual lipase, which was associated with the 3 selected parameters. In humans, the association parameters were principally established through indirect evidence without a clear demonstration of cause. Moreover, no specific lipase, such as lingual lipase in the case of rodents, was identified at the oral level. Conclusions Future research efforts should focus on (i) establishing a standard procedure for oral lipolytic activity evaluation and, in particular, a methodological control to address the lipase vs esterase confounding bias and (ii) identifying the main lipases that contribute to the lipolytic activity in humans at the oral level and their respective contribution to the association parameters defined in this review.

Mounting evidence shows that hedonic eating, leading to overeating just for pleasure, can be driven by oro-sensory factors through the activation of reward processing and learning in the brain. Foods rich in sugars and fats are potent rewards and saliva composition influences oral taste, texture and aroma perception. A role for salivary α-amylase and lipase in the gustatory system and a link between salivary α-amylase activity and dietary habits were recently hypothesized. The objective of this study was to verify the relationship among salivary lipase and α-amylase activities as well as zinc concentration with food preference and choice of people with different body mass indices. Forty-two (23 normal weight and 19 overweight) healthy subjects participated in the study. Data showed that α-amylase and lipase were 1.8 and 2.4 folds higher in overweight than in normal weight subjects, respectively. On the other hand, overweight subjects showed a 33% reduced salivary zinc concentration compared to normal weight subjects. Only lipase activity positively correlated with individual preference for high-fat foods and with fat content of the diets. All in all data suggested that high salivary lipase activity in overweight subjects could be an adaptive response to the low fat-taste perception related to the reduced zinc concentration. It cannot be ruled out that other factors but diet might influence salivary α-amylase activity in overweight subjects.

Normal posterior deep and superficial salivary glands of tongue were examined in male mice by means of light microscopical histochemistry and neurohistology. Both glands showed acini and simple ducts. Demilunes were present in the superficial gland. Disulphides and neutral mucosubstances occurred in acini and demilunes. Tryptophan staining was seen in acini of the deep gland and demilunes, whereas acid mucosubstances were exclusively localised in the superficial gland. Dehydrogenase activities were widespread. Strong esterase activity occurred throughout the parenchyma of the deep gland and in demilunes; it was variably inhibited by E600, apart from acinar apical regions in the deep gland. Lipase was confined to acini of the deep gland and demilunes. Acid phosphatase staining was similarly localised; it was also seen in periluminal ductal rims of the deep gland, in which ouabain-sensitive Na,K-ATPase was localised basolaterally. Staining for alkaline phosphatase decorated occasional myoepithelial-like arrangements and interstitial capillaries. Acetylcholinesterase was associated with nerve fibres embracing glandular parenchyma. Adrenergic fibres were not seen. The results suggest that the acini of the posterior deep lingual gland secrete neutral glycoproteins, whereas the ducts transport ions and absorb luminal material. The posterior superficial lingual gland mainly secretes acid glycoproteins. Both glands produce lingual lipase, receive cholinergic-type innervation and have inconspicuous myoepithelium.

Part of medium chain fatty acids (MCFAs) coming from dietary triglycerides (TGs) can be directly absorbed through the gastric mucosa after the action of preduodenal lipase (lingual lipase in the rat). MCFA gastric absorption, particularly that of octanoic acid (C8:0), may have a physiological importance in the octanoylation of ghrelin, the orexigenic gastric peptide acting as an endogenous ligand of the hypothalamic growth hormone secretagogue receptor la (GHSR-1a). However, the amount of C8:0 absorbed in the stomach and its metabolic fate still haven't been clearly characterized. The purpose of the present study was to further characterize and quantify the importance of preduodenal lipase activity on the release and gastric absorption of dietary C8:0 and on the subsequent ghrelin octanoylation in the stomach mucosa. Fifteen days old rats received fat emulsions containing triolein or [1,1,1-13C]-Tri-C8:0 and a specific inhibitor of preduodenal lipase, 5-(2-(benzyloxy)ethoxy)-3-(3-phenoxyphenyl)-1,3,4-oxadiazol-2(3H)-one or BemPPDX. The fate of the C-13-C8:0 was followed in rat tissues after 30 and 120 min of digestion and octanoylated ghrelin was measured in the plasma. This work (1) demonstrates that part of C8:0 coming from Tri-C8:0 is directly absorbed at the gastric level, (2) allows the estimation of C8:0 gastric absorption level (1.3% of the C-13-C8:0 in sn-3 position after 30 min of digestion), as well as (3) the contribution of rat lingual lipase to total lipolysis and to duodenal absorption of dietary FAs (at least 30%), (4) shows no short-term effect of dietary Tri-C8:0 consumption and subsequent increase of C8:0 gastric tissue content on plasma octanoylated ghrelin concentration. (C) 2016 Elsevier B.V. All rights reserved.

Ontogeny and fatty acid chain-length specificity of gastrointestinal lipases in neonatal piglets were examined to explore the basis for variations in postnatal use of medium-chain triacylglycerols (MCT). Twenty-four newborn pigs were studied at 4 ages: 0, 6, 18, and 48 h postpartum (n = 6 pigs/age). Piglets were gastrically intubated and given 3.0 mmol/kg of BW(0.75) each of emulsified tri-C6:0 and tri-C8:0. One hour after intubation, the plasma concentration of C6:0 was 7.5-fold greater than that of C8:0 (P 0.05). The lowest lipase activity was observed at 18 h for both tri-C6:0 and tri-C8:0. Chain-length specificity of pancreatic lipase was measured with tri-C4:0, tri-C6:0, tri-C8:0, and tri-C10:0 as combined or separate substrates. As separate substrates, the lipase activity decreased progressively as chain length increased from tri-C4:0 to tri-C10:0. As combined substrates, tri-C6:0 was hydrolyzed fastest (P < 0.05), followed by C4:0, C8:0, and C10:0. Gastric and lingual lipase activities averaged 2.7 nmol/min per mg of protein for the medium-chain fatty acid substrates, with hydrolysis of C6:0 being 7-fold greater than that of C8:0. In conclusion, pancreatic lipase dominates the preduodenal lipases in the neonatal pig, and greater activity of the gastrointestinal lipases toward tri-C6:0 underlies its increased rate of use.

The present study investigated the metabolic fate of dietary TAG and DAG and also their digestion products in the stomach and small intestine. A diet containing 10% TAG or DAG oil, enriched in 1,3-DAG, was fed to Wistar rats ad libitum for 9 d. After 18 h of fasting, each diet was re-fed ad libitum for 1 h. The weights of the contents of the stomach and small intestine were measured, and the acylglycerol and FFA levels were analyzed by GC at 0, 1, and 4 h after the 1-h re-feeding. The amounts of re-fed diet ingested and the gastric and small intestinal content were not different between the two diet groups. In the TAG diet group, the main products were TAG and DAG, especially 1(3),2-DAG. In addition, 1,3-DAG and 1(3)-MAG were present in the stomach, and the 1,3-DAG levels increased over time after the re-feeding period. In the DAG diet group, the main products in the stomach were DAG, MAG, FFA, and TAG. There were significantly greater amounts of 1,3-DAG, 1(3)-MAG, and FFA in the DAG diet group in the stomach compared with the TAG diet group. The amount of FFA in the stomach relative to the amount of ingested TAG plus DAG in the DAG diet group was higher than that in the TAG diet group. Acylglycerol and FFA levels were considerably lower in the small intestine than in the stomach. These results indicate that, in the stomach, where acyl migration might occur, the digestion products were already different between TAG and DAG oil ingestion, and that DAG might be more readily digested by lingual lipase compared with TAG. Furthermore, almost all of the dietary lipid was absorbed, irrespective of the structure of the acylglycerol present in the small intestine.

Oral administration of epsilon-polylysine to rats reduced the peak plasma triacylglycerol concentration. In vitro, epsilon-polylysine and polylysine strongly inhibited the hydrolysis, by either pancreatic lipase or carboxylester lipase, of trioleoylglycerol (TO) emulsified with phosphatidylcholine (PC) and taurocholate. The epsilon-polylysine concentration required for complete inhibition of pancreatic lipase, 10 microg/ml, is 1,000 times lower than that of BSA required for the same effect. Inhibition requires the presence of bile salt and, unlike inhibition of lipase by other proteins, is not reversed by supramicellar concentrations of bile salt. Inhibition increases with the degree of polylysine polymerization, is independent of lipase concentration, is independent of pH between 5.0 and 9.5, and is accompanied by an inhibition of lipase binding to TO-PC emulsion particles. However, epsilon-polylysine did not inhibit the hydrolysis by pancreatic lipase of TO emulsions prepared using anionic surfactants, TO hydrolysis catalyzed by lingual lipase, or the hydrolysis of a water-soluble substrate. In the presence of taurocholate, epsilon-polylysine becomes surface active and adsorbs to TO-PC monomolecular films. These results are consistent with epsilon-polylysine and taurocholate forming a surface-active complex that binds to emulsion particles, thereby retarding lipase adsorption and triacylglycerol hydrolysis both in vivo and in vitro.

Lingual lipase is usually secreted from von Ebner's glands, although there is great variation between species. Lingual lipase is thought to be an auxiliary enzyme for fat digestion and absorption in mammals; however, the reason for lipolysis in the oral cavity is not known. We focused on the gustatory sense and investigated the significance of lingual lipase in the perception of a fat taste by using orlistat, a potent lipase inhibitor. Five-minute two-bottle preference tests demonstrated that the addition of orlistat diminished the preference for triacylglycerides but not for free fatty acids. Radioactive triolein applied on rats' circumvallate papilla revealed that lingual lipase was released continuously to generate significant amounts of fatty acids and other lipolytic products within 1–5 s, which was enough time to taste fat. These findings suggest that lingual lipase is released to perceive the taste of triacylglycerides and to find nutritive lipids in food.

Enzymatic hydrolysis of phthalate esters in human saliva was investigated to characterize salivary esterase in the formation of monoesters from their diesters. The monoesters formed were analyzed by GC/MS after incubation of phthalate diesters in the saliva. Hydrolytic activity in the supernatant obtained by centrifugation of the saliva at 1350 × g was equivalent to that in whole saliva, and the activity was inhibited by the addition of denaturing protein. The hydrolytic activity was dependent on the protein concentration in the supernatant. The optimum temperature and pH of the hydrolysis was 50°C and 8.2, respectively. In addition, the 80% acetone powder of the supernatant showed high substrate specificity for straight-chain alkyl group of phthalate diesters, especially the butyl group, whereas almost no specificity was seen for the 2-ethylhexyl and benzyl groups. These results indicate that not the oral flora but salivary esterase, such as lingual lipase, is involved in phthalate monoester formation from the diesters in human saliva, and do not act on the hydrolysis of monoester.

Catalytic activities of lingual lipase were investigated by enzyme histochemistry in post-mortem tongues from male rats. Sections of fresh-frozen or formalin-calcium fixed tissue were incubated with naphthol-AS-nonanoate and α-naphthyl acetate substrate mixtures. The effects of pH level, sodium taurocholate activator and E600 inhibitor were also examined. The use of cryostat sections of tissues fixed in formalin-calcium and of nonanoate substrate within the range of pH 4.4–6.4, were optimal for localizing maximum reaction product, captured by Fast Blue BB, in acini and demilunes of the posterior deep and superficial lingual glands respectively. The reaction product corresponded with the distribution of secretory granules and failed to develop when taurocholate was omitted from the incubation medium. Similarly localized E600-resistant reaction product occurred with the acetate substrate and hexazotized New Fuchsin at pH 7.4, in the absence of taurocholate. Lipase and conventional esterase activities appear to be superimposed in posterior lingual glands of rat. The ability of their acini and demilunes to hydrolyse nonanoate substrate at an acidic pH optimum, when activated by sodium taurocholate, seems attributable to lipase destined for secretion into saliva – hence convenient for routine histochemical identification of the enzyme.

Lingual lipase generates nonesterified fatty acids (NEFA) from dietary fats during oral processing by lipolysis. Lingual lipase in rodents has strong lipolytic activity and plays a critical role in oral detection of fats. The functional activity of lingual lipase during oral processing of high-fat foods in humans remains poorly characterized. Five commonly consumed high-fat foods varying in physical states and fatty acid composition (almond, almond butter, olive oil, walnut, and coconut) were masticated by 15 healthy human subjects at the rate of one chew per second with and without lipase inhibitor orlistat. Salivary NEFA concentrations were measured. To determine the role of lingual lipase in oral fat detection, sensory ratings were obtained from the same 15 human subjects for almond butter with and without orlistat. Lingual lipase was active during oral processing of almond and coconut. No activity of lingual lipase was detected during processing of almond butter. There was only weak evidence lingual lipase is a determinant of oral fat detection. Lingual lipase may only contribute to NEFA generation and oral fat detection of fatty foods that require stronger oral processing effort.

Obesity is characterized by excessive adipose tissue mass and cause for a variety of pathologic conditions like cardiovascular disease, type 2 diabetes and cancer [1]. One important factor in the development of obesity is the overconsumption of high calorie, fat-rich diets. Prior to their uptake, dietary fats mostly composed of mixed triacylglycerols (TAGs) are processed in the digestive tract by extracellular lipases. While small portions of ingested lipids are hydrolyzed by lingual lipase and gastric lipase in the stomach, the vast majority of ingested TAGs are metabolized in the small intestine. There they are emulsified by bile acids and hydrolyzed by the action of pancreatic lipase (PL), co-lipase, PL-related protein 2, and carboxyl ester lipase [1,2]. The resulting free fatty acids (FFA) and monoacylglycerols are subsequently taken up by enterocytes, re-esterified to TAGs, packed into chylomicrons and released into the lymphatic system where they enter the circulation via the thoracic duct. Within the blood stream chylomicrons are transported to peripheral organs like adipose tissue (AT), skeletal muscle and heart, where the lipoprotein-associated TAGs are hydrolyzed again by lipoprotein lipase (LPL) and FFA are absorbed by the underlying tissues. Accordingly, LPL serves as a “gate-keeper” for the uptake of FFA in peripheral tissues. Altered LPL activities or enzyme defects are associated with various forms of hypertriglyceridemias and associated diseases such as cardiovascular disease [1,3]. Various members of the angiopoietin-like (Angptl) protein family inhibit LPL enzyme activity by a post-translational mechanism [4]. The Angptl-protein family consists of eight identified members (Angptl1–8) of which at least Angptl3, Angptl4, and Angptl8 affect LPL activity and plasma TAG concentrations [5,6]. Gene knockout of each of these Angptl-proteins decreases plasma TAG levels whereas the transgenic overexpression of Angptl4 raises plasma TAG concentrations. Angptl3 is predominantly expressed in liver, whereas Angptl4 is expressed in a variety of tissues and cell types, including adipose tissue, liver, heart and macrophages [4,5]. Interestingly, Kersten et al. recently reported that Angptl4 is also expressed and secreted from human entero-endocrine cells into the intestinal tract [7]. This suggested that Angptl4 expression and release might have a dual function in the intestine: It could act as endocrine hormone to regulate vascular lipoprotein metabolism or it could affect lipid absorption and turnover directly in the gut [7]. In this issue of Molecular Metabolism the same group provides strong evidence for a role of Angptl4 in intestinal lipid digestion. Angptl4−/− mice exhibited increased intestinal lipid uptake and decreased fat excretion resulting in increased weight gain and an obese phenotype [8]. These results pointed towards a specific inhibitory role of Angptl4 on lipid hydrolysis by digestive lipases. Indeed, enzymatic assays of fecal water of Angptl4−/− mice demonstrated that PL activity was significantly increased compared to wild-type (WT) controls. Mattijssen et al. [8] also showed that recombinant Angptl4 inhibits PL activity in vitro in a dose dependent manner. These findings were also translatable to humans as recombinant Angptl4 inhibited PL activity in human feces samples. Taken together, these results assign a novel role to Angptl4 affecting intestinal lipid catabolism via inhibition of PL [8]. This is exciting but also raises a number of new questions. For example, it remains unclear how increased intestinal lipid uptake translates into weight gain and obesity observed in Angptl4−/− mice [8]. Despite increased lipid tracer abundance in the intestine of Angptl4−/− mice in lipid tolerance tests, the synthesis and secretion rates of chylomicrons are not different between Angptl4−/− animals and controls. This suggests that obesity results from changes in FFA partitioning rather than increased fat delivery to adipose tissue. It is conceivable that the increased activity of adipose-specific LPL in global Angptl4 knockout mice leads to a more efficient uptake of chylomicron-associated TAG in adipose tissue. Unfortunately, it is currently not known whether adipose-specific overexpression or deletion of Angptl4 affects adipose tissue mass. Alternatively, Angptl4 may affect intracellular lipolysis of fat stores in adipose tissue. Gray et al. showed that Angptl4−/− mice had decreased glycerol release of fat pad explants and decreased plasma FFA levels when fasted [9]. Accordingly, decreased lipolysis in adipocytes lacking Angptl4 may result in enhanced lipid accumulation. Finally, it is conceivable that the obese phenotype in Angptl4−/− mice results from changes in total body energy expenditure by a currently unknown mechanism similarly as previously reported for acyl CoA:monoacylglycerol acyltransferase-2 deficient mice. In this mouse model, increased energy expenditure and body temperature leads to resistance to diet induced obesity despite the fact that fat resorption in the gut and mucosal chylomicron synthesis are unaltered compared to WT mice [10]. Clarifying which of the mechanisms mentioned above affects the body composition in Angptl4 deficiency will require a detailed characterization of mice overexpressing or lacking Angptl4 in specific tissues. The present study emphasizes the idea that Angptl4-mediated inhibition of PL may serve as protective mechanism against nutritional lipid overload [8]. The concept is based on the observation that intestinal Angptl4 expression significantly increases when mice were fed a high fat diet. Increasing Angptl4 levels may result in decreased PL activity and via a protective feedback loop prevent lipid overload. A similar role for Angptl4 in the protection from lipotoxicity has been reported for macrophages [11]. Another important aspect relates to the finding that Angptl4 expression in the intestine is regulated by gut microbiota [12–14]. Although the species of microorganisms and the involved mechanisms that regulate Angptl4 levels in the gut are unknown, changes in the gut microbiome during the course of obesity may affect intestinal lipid uptake via Angptl4 expression and PL inhibition. Future studies should focus on the biochemical mechanisms by which Angptl4 inhibits PL. What is the role of co-lipase? Does Angptl4 also affect the activity of PL-related protein 2? Once these questions are answered, intestinal Angptl4 could serve as a potential pharmacological target to selectively decrease lipid uptake and thereby affect energy balance.

In examining the secretory mechanism of exocrine glands, we focused on the small GTP-binding proteins, Rab3D and Rab3B, which function in the final steps of exocytosis in non-neuronal tissues. These proteins were observed in von Ebner's glands by 32P-GTP overlay. mRNA isolated from von Ebner's glands, the pancreas, parotid glands, and liver was subjected to reverse transcription PCR and probed with primers and nested primers for Rab3D and Rab3B. Rab3D was found in all three exocrine glands and the liver, while Rab3B was found in the liver. Utilizing immunofluorescence histochemistry, Rab3D was localized to hepatocytes of the liver and to secretory granules of the exocrine glands, and Rab3B to liver and pancreatic islets. Isoproterenol evoked decreases in α-amylase- and Rab3D-labelled parotid secretory granules, and pilocarpine stimulated decreases in secretory granules labelled for lingual lipase and Rab3D from von Ebner's glands, and amylase and Rab3D from pancreas. Neither secretagogue affected Rab3B in pancreatic islets. These observed parallel decreases in response to β-adrenergic (parotid) or cholinergic (von Ebner's and pancreas) secretagogues indicate that the function of Rab3D in exocytosis in these exocrine organs is similar and that the type of secretagogue does not determine the function.

Field, R. B., S. J. Chirtel and R. S. Redman. Effect of substance P and receptor antagonists on secretion of lingual lipase and amylase from rat von Ebner's gland. Peptides 18(2) 277–285, 1997.— Substance P (SP, 1 μ M) when incubated with minced von Ebner's glands for 15, 30, and 60 min, stimulated secretion of lingual lipase (12.14%±0.90) and amylase (8.30%±0.42). Only 10 μ M of the SP receptor antagonist CP-96,345 significantly inhibited SP-evoked secretion. d -Pro 2 - d -Phe 7 - d -Trp 9 -SP (Ia), d -Pro 2 - d -Trp 7,9 -SP (Ib), d -Arg 1 - d -Trp 7,9 - d -Leu 11 -SP (Ic), or 1 μ M CP-96,345 were not effective, suggesting that the SP receptor of von Ebner's gland might be an isoform. Propranolol and timolol, β 1 / β 2 -adrenergic receptor antagonists were not effective and the cholinergic receptor antagonist, atropine, was effective in only slightly reducing amylase secretion but not lingual lipase. Differential secretion of the two enzymes was observed for basal and stimulated secretion. Thus, exocytosis may not be the only pathway involved in SP-evoked protein secretion.

Oligodeoxyribonucleotide ligation to single—stranded cDNA (SLIC) and polymerase chain reaction (PCR) techniques were used to clone an entire dog gastric lipase (DGL) cDNA. The size of the cDNA is confirmed by Northern blot analysis. The DGL is synthesized as a 379–amino acid mature polypeptide with a molecular mass of 43176 Da which is preceded by a 19–amino acid signal sequence located at the NH2--terminus. Comparison of the signal sequences reveals a high degree of similitude between the DGL, the human gastric lipase (HGL), the rabbit gastric lipase (RGL) and the rat lingual lipase (RLL).

Human lysosomal acid lipase/cholesteryl ester hydrolase (HLAL), human gastric lipase (HGL), and rat lingual lipase (RLL) constitute a family of mammalian lipases characterized by an acidic pH optimum. HGL and RLL are secreted by the chief cells of the stomach and by the serous von Ebner's glands of the tongue, respectively, and hydrolyze dietary longchain triglycerides in the gastrointestinal tract. HLAL, in contrast, catalyzes the intralysosomal degradation of both triglycerides and cholesteryl esters in virtually all cells except erythrocytes. All three enzymes are proposed to be serine esterases with a catalytic Ser-Asp-His triad similar to other lipases, despite their sensitivity towards sulfhydryl modifying reagents. To investigate the role of conserved serine, aspartic acid, and histidine residues in HLAL and HGL, we constructed 24 mutant lipases with single amino acid substitutions using the site-directed mutagenesis approach. Our combined data strongly support the conclusion that Ser153, Asp324, and His355 are components of the catalytic triad of HLAL and HGL. Structural integrity of the conserved His-Gly dipeptide of lipases also appears to be important for neutral lipid hydrolysis, as replacement of His65 by glutamine abolished HLAL and HGL enzymic activity. Substitution of HLAL residues Asp93, Asp130, and Asp328 with glycine, in contrast, had a more pronounced impact on cholesteryl oleate hydrolysis than on triglyceride hydrolysis. These results provide new insights into the structural basis of HLAL and HGL function.

Human gastric lipase (HGL; triacylglycerol lipase; EC 3.1.1.3) plays an important role in the digestion of dietary triglycerides in the gastrointestinal tract, especially in patients suffering from pancreatic lipase deficiencies. The enzyme is secreted by the fundic mucosa of the stomach and hydrolyzes the ester bonds of triglycerides under acidic pH conditions, while cholesteryl esters are not attacked. The 379-amino acid protein is highly homologous to two other acidic lipases, rat lingual lipase (RLL; triacylglycerol lipase; EC 3.1.1.3) and human lysosomal acid lipase (HLAL; cholesteryl esterase; EC 3.1.1.13). To determine whether this remarkable similarity is also present at the genomic level, we have elucidated the respective gene structures by screening three bacteriophage lambda libraries and by polymerase chain reaction-based intron amplification. The genes encoding HGL, RLL, and HLAL are composed of 10 exons interrupted by nine introns and span about 14 kb, 18.7 kb, and 38.8 kb of genomic DNA, respectively. The HGL and RLL gene organizations are identical, suggesting that RLL is the rat gastric lipase expressed in the serous von Ebner glands of the tongue. The positions of the HLAL intervening sequences are also absolutely conserved, except for the location of intron 1. Our results support the concept that HLAL and HGL/RLL are members of a gene family of lipases that most likely have evolved by duplication of an ancestral gene and subsequently assumed distinct roles in neutral lipid metabolism due to sequence divergence and different expression patterns.

There is mounting evidence that, in addition to texture and olfaction, taste plays a role in the detection of long chain fatty acids. Triglycerides, the main components of oils and dietary fat, are hydrolyzed in the mouth by a lingual lipase secreted from the von Ebner gland and the released free fatty acids are detected by the taste system. GPR40 and GPR120, two fatty acid responsive G-protein-coupled receptors (GPCRs), are expressed in taste bud cells, and knockout mice lacking either of those receptors have blunted taste nerve responses to and reduced preference for fatty acids. Here we investigated whether activation of those GPCRs is sufficient to elicit fat taste and preference. Five non-fatty acid agonists of GPR40 and two non-fatty acid agonists of GPR120 activated the glossopharyngeal nerve of wild-type mice but not of knockout mice lacking the cognate receptor. In human subjects, two-alternative forced choice (2-AFC) tests, triangle tests and sensory profiling showed that non fatty acid agonists of GPR40 dissolved in water are detected in sip and spit tests and elicit a taste similar to that of linoleic acid, whereas 2-AFC tests showed that two agonists of GPR120 in water are not perceived fattier than water alone. Wild-type mice did not show any preference for five agonists of GPR40, two agonists of GPR120 and mixtures of both agonists over water in two-bottle preference tests. Together these data indicate that GPR40 mediated taste perception is not sufficient to generate preference.

The precise orosensory inputs engaged for dietary lipids detection in humans are unknown. We evaluated whether a common single nucleotide polymorphism (rs1761667) in the CD36 gene that reduces CD36 expression and the addition of orlistat, a lipase inhibitor, to reduce FA release from triacylglycerols (TGs), the main component of dietary fats, would attenuate fat orosensory sensitivity in humans. Twenty-one obese subjects with different rs1761667 genotypes (6 AA, 7 AG, and 8 GG) were studied on two occasions in which oleic acid and triolein orosensory detection thresholds were measured using emulsions prepared with and without orlistat. Subjects homozygous for the G-allele had 8-fold lower oral detection thresholds for oleic acid and triolein than subjects homozygous for the A allele, which associates with lower CD36 expression (P = 0.03). Thresholds for heterozygous subjects were intermediate. The addition of orlistat increased detection thresholds for triolein (log threshold = −0.3 ± 0.2 vs. 0.3 ± 0.1; P < 0.001) but not oleic acid (log threshold = −1.0 ± 0.2 vs. −0.8 ± 0.2; P > 0.2). In conclusion, this is the first experimental evidence for a role of CD36 in fat gustatory perception in humans. The data also support involvement of lingual lipase and are consistent with the concept that FA and not TG is the sensed stimulus.

The polymerase chain reaction (PCR) was used to amplify specific parts of the gene encoding calf pregastric esterase (PGE). Primers based on conserved regions in human gastric lipase (HGL) and rat lingual lipase (RLL) were used to screen a cDNA library prepared from calf tongue tissue. This resulted in the cloning of the entire coding sequence for PGE, which exists as a mature 378-amino-acid (aa) polypeptide with a molecular mass of 42960 Da. The PGE, HGL and RLL genes all share a high degree of identity at both the nucleotide and amino-acid sequence levels. Except for the Gly-Xaa-Ser-Xaa-Gly sequence containing the active site Ser, there is little identity with non-preduodenal lipases.

Lysosomal acid lipase (LAL) is a hydrolase essential for the intracellular degradation of cholesteryl esters and triacylglycerols. This report describes a multi-step procedure for the purification of LAL from human liver. After solubilization with non-ionic detergent, acid hydrolase activity was purified 17000-fold to apparent homogeneity by sequential chromatography on Concanavalin A Sepharose, carboxymethyl-cellulose, phenyl Superose, Mono S cation exchange and Superose 12 gel-filtration columns. This procedure yielded two silver-staining protein bands of 56 kDa and 41 kDa on SDS/PAGE. Size-exclusion chromatography of the 41-kDa protein indicated that the enzyme was catalytically competent as a monomer of approximately 38 kDa. When assayed in the presence of cholesteryl oleate or trioleoylglycerol, purified acid lipase had Vmax values of 4390 nmol fatty acid.min-1.mg protein and 4756 nmol fatty acid.min-1.mg protein-1, and apparent Km values of 0.142 mM and 0.138 mM, respectively. The purified enzyme was most active at low pH (4.5-5.0) and required non-ionic detergent and ethylene glycol for optimal stability. Incubation of the 41-kDa acid lipase with endoglucosaminidase H reduced the molecular mass by 4-6 kDa, demonstrating Asn-linked glycosylation with high-mannose oligosaccharides. Deglycosylation did not affect enzymic activity, indicating that carbohydrates are not required for LAL activity. Based on partial peptide sequence, an oligonucleotide was synthesized and utilized to isolate LAL cDNA clones from a human liver cDNA library. A full-length LAL cDNA contained 2626 nucleotides and coded for a predicted protein of 372 amino acids, preceded by a 27 residue hydrophobic signal peptide. Hepatic LAL differed from fibroblast acid lipase at the N-terminus and revealed extensive similarities with human gastric lipase and rat lingual lipase, confirming a gene family of acid lipases. Northern hybridization using the complete LAL cDNA as a radiolabeled probe indicated striking differences in mRNA expression among human tissues. LAL mRNA was most abundant in brain, lung, kidney and mammary gland. Placenta and HeLa cells expressed intermediate amounts of LAL mRNA, while RNA extracted from liver and heart showed low levels of expression.

Background Oil pulling has been used extensively as a traditional Indian folk remedy without scientific proof for many years for strengthening teeth, gums and jaws and to prevent decay, oral malodor, bleeding gums and dryness of throat and cracked lips. Aim The aim of this study was to evaluate the antibacterial activity of sesame oil and lignans isolated from sesame oil on oral microorganisms and to check whether saponification or emulsification occurs during oil-pulling therapy. Materials and methods The in vitro study was carried out in three different phases: (1) Antibacterial activity of the lignans and sesame oil were tested by minimum inhibitory concentration assay by agar dilution method and agar well diffusion method, respectively. (2) Increase in free fatty acid level of oil and the quantity of sodium hydroxide (NaOH) used up in the titration are good indicators of saponification process. This was assessed using analytical tests for vegetable oils. (3) Swished oil was observed under light microscope to assess the status of the oil, presence of microorganisms, oral debris and foreign bodies. Results Sesamin and sesamolin isolated from sesame oil did not have any antibacterial effect against oral microorganisms like Streptococcus mutans, Streptococcus mitis and Streptococcus viridans. Emulsification of sesame oil occurs during oil-pulling therapy. Increased consumption of NaOH in titration is a definite indication of a possible saponification process. Conclusion The myth that the effect of oil-pulling therapy on oral health was just a placebo effect has been broken and there are clear indications of possible saponification and emulsification process, which enhances its mechanical cleaning action.

The lingual lipases from calf and lamb were used as catalysts for the hydrolysis of the lipid, tributyrin, and the relatively hydrophilic ester, 4-nitrophenylacetate. The effects of pH on these reactions and of temperature on the reaction against 4-nitrophenylacetate were measured. Calf lingual lipase was used also to catalyze the hydrolysis of triolein. Under comparable conditions, calf lingual lipase was a more effective catalyst than lamb lingual lipase. Michaelis-Menten constants were evaluated and showed that the preferred substrates for the enzyme were tributyrin, then 4-nitrophenylacetate, and then triolein. For tributyrin and 4-nitrophenylacetate, the optimal pH were 6.9 and 8.0, respectively, for the calf enzyme and 6.6 and 6.8, respectively, for the lamb enzyme. The temperature optima against 4-nitrophenylacetate were 37.5 and 30°C for calf and lamb lingual lipases, respectively.

Recent evidence in the literature suggests that signals carried by the glossopharyngeal nerve (GL), which supplies sensory and parasympathetic innervation of the posterior tongue, might be essential in the maintenance of normal gustatory responses to fat stimuli. Here, we report that GL transection (GLX) significantly decreased corn oil intake and preference in 23-h two-bottle tests relative to sham-operated controls (Sham). Drinking-pattern analysis of corn oil licking revealed that bout size, rather than the number of bouts initiated, was smaller in GLX than Sham rats. We also tested a range of glucose concentrations and found that total licks over daily 23-h sessions significantly decreased in GLX compared with Sham rats, but this difference failed to reach significance when intake or any bout parameter was measured. These results show that the signals in the GL normally contribute to processes involved with corn oil bout termination as opposed to bout initiation. GL-derived signals could potentially provide input to “reward” circuits in the ventral forebrain that could serve to maintain ingestion during a meal or, alternatively, could act at the level of the brain stem to attenuate the inhibitory potency of vagal signals, thus delaying the onset of satiation, or perhaps contribute to a cephalic phase reflex modulation of the gut. Parasympathetic efferents in the GL innervating the von Ebner's glands, which secrete lingual lipase, which is thought to break down corn oil into detectable ligands, could also be playing a role in driving corn oil intake. Whatever the mechanism, an intact GL is clearly necessary in maintaining normal intake of corn oil.

Minced von Ebner's glands of rat tongue were incubatedin vitro with histamine and histamine receptor antagonists. At various time intervals, media and homogenates of the tissue were assayed for lingual lipase and amylase activity and percentage secretion calculated. Histamine elicited moderate secretion (≈10%) of lingual lipase and amylase. In contrast, pyrilamine, and H1 receptor antagonist, elicited>60% secretion. There were statistically significant differences between the percentage secretion of lingual lipase and amylase for basal secretion, as well as for histmine-and pyrilamine-evoked secretion of lingual lipase and amylase for basal secretion, as well as for histamine-and pyrilamine-evoked secretion above basal. The H2 receptor inhibitors, cimetidine and ranitidine, stimulated secretion of only amylase, but not lingual lipase. When combined with histamine, these antagonists partially inhibited only the secretion of histamine-evoked lingual lipase, but not amylase. The differences in percentage secretion between the two enzymes indicate that exocytosis may not be the only process involved in protein secretion. The anomalous effects of the H1 and H2 receptor antagonists necessitate a more detailed characterization of the receptors of von Ebner's glands.

Cheddar and goats' milk cheese bases that were lipolyzed by crude kid goat lingual lipase were stored at 1 or 21°C with the water activity ( a w ) controlled between 0·33 and 0·97. Concentrations of volatile n - and branched-chain free fatty acids in lipolyzed cheese bases after 4 weeks were the lowest in samples stored at a w 0·85, indicating possible resynthesis of fatty acids into acylglycerols. However, samples held at the extremes of the a w range did not exhibit evidence indicating a likelihood of acylglycerol resynthesis under these conditions. Storage at low temperature (1°C), as well as addition of ethanol to lipolyzed cheese bases reduced the volatile free fatty acid content.

The sections in this article are: 1 Enzymes Related to Intraluminal Digestion of Dietary Fat 1.1 Lingual Lipase and Gastric Lipase 1.2 Pancreatic Lipase and Colipase 1.3 Carboxyl Ester Lipase 1.4 Bile Salts as Cofactors of Carboxyl Ester Lipase 1.5 Phospholipase A2 2 Bile 3 Product Phases of Lipid Digestion: Experience From in vitro Studies 4 Lipid-Phase Products Found in Luminal Contents During Digestion 5 Cooperative Effects in Lipid Digestion 6 Fate of Fat-Soluble Molecules During Fat Digestion 7 Conclusions

Molecular cloning of a full-length cDNA for human lysosomal acid lipase/cholesteryl ester hydrolase (EC 3.1.1.13) reveals that it is structurally related to previously described enteric acid lipases, but lacks significant homology with any characterized neutral lipases. The lysosomal enzyme catalyzes the deacylation of triacylglyceryl and cholesteryl ester core lipids of endocytosed low density lipoproteins; this activity is deficient in patients with Wolman disease and cholesteryl ester storage disease. Its amino acid sequence, as deduced from the 2.6-kilobase cDNA nucleotide sequence, is 58 and 57% identical to those of human gastric lipase and rat lingual lipase, respectively, both of which are involved in the preduodenal breakdown of ingested triglycerides. Notable differences in the primary structure of the lysosomal lipase that may account for discrete catalytic and transport properties include the presence of 3 new cysteine residues, in addition to the 3 that are conserved in this lipase gene family, and of two additional potential N-linked glycosylation sites. Transfection of the cDNA into Cos-1 cells resulted in the expression of acid lipase activity with the substrate range of the native enzyme at a level that was greater than 40 times the endogenous activity.

The serous lingual glands of von Ebner secrete lingual lipase, an enzyme that begins fat digestion in the stomach. The objective of this study was to characterize the neuromodulators in the rat tongue and von Ebner glands using immunocytochemical techniques. Rat lingual tissues were fixed in formalin, embedded in paraffin and sectioned at 4 microns for light microscopic studies. Immunocytochemical localization of neuromodulators was performed with monospecific anti-rat neuromodulator IgG or control (preimmune) IgG as the primary antibody, using the peroxidase-antiperoxidase (PAP) technique. No staining was seen with control anti-rat IgG. Immunospecific staining for vasoactive intestinal peptide (VIP), tyrosine hydroxylase and choline acetyltransferase (CHAT) was observed in nerves in the tongue, and cells containing immunospecific staining for serotonin (5-hydroxytryptamine) were seen in the stroma between the lingual glands. Selected cells in the serous glands stained positively for the presence of substance P and somatostatin. Adrenergic, VIP-containing and cholinergic nerves appear to innervate the tongue and serous glands. Substance P and somatostatin were identified in cells of the lingual serous glands and may be additional local modulators regulating lingual lipase release.

Lingual lipase in the rat is present in the neonatal period and undergoes developmental increase during postnatal life. To evaluate the role of glucocorticoid in the control of lingual lipase during development, suckling rats were adrenalectomized at d 10 and various hormone replacements were performed. Adrenalectomy abolished the developmental increase of lingual lipase. Low doses of dexamethasone (0.2 and 0.5 microgram/100 g body wt) restored the lingual lipase to near normal level in adrenalectomized animals. High doses of dexamethasone (20 micrograms/100 g body wt), when given to similarly adrenalectomized animals, however, led to a reduction of lingual lipase levels. Inhibition by dexamethasone is through the action of the hormone inasmuch as the coadministration of RU38486, a glucocorticoid type II receptor antagonist, completely abolished the inhibitory action. Inhibition is also steroid specific, with dexamethasone and triamcinolone acetonide being more effective. The results suggest a unique bimodal regulation of lingual lipase by dexamethasone in the rat serous glands. Because of the possible importance of lingual lipase as an alternative enzyme for fat digestion in neonates, the inhibitory action of high doses of glucocorticoid on lingual lipase development may have important implications. The use of steroidal compounds in the hastening of lung maturation and treatment of inflammatory disease might conceivably compromise their lingual lipase development, hence their capacities of fat digestion and malabsorption in the same period.

Lingual serous glands (von Ebner) contain a potent lipase that hydrolyses triglycerides to a mixture of partial glycerides (di- and monoglycerides), glycerol and free fatty acids. Studies in man and in rat have shown that similar lipolytic activity is present in oesophageal and gastric aspirates and suggest that the intragastric digestion of dietary fat is initiated by lingual lipase. In the rat the lingual serous glands are initiated in 19- to 20-day fetuses; lipase activity is first detected in 20-day-old fetuses and increases 14-fold by birth. The data suggest that in the fetus lipase activity originates predominantly in the serous demilune cells of mucous glands whereas after birth the enzyme is synthesized and stored in the rapidly differentiating serous glands. In man lipolytic activity is present in gastric aspirates as early as 26 weeks of gestational age. Our studies suggest that the lipolytic activity has characteristics similar to those of adult human and rat lingual lipase; the enzyme is present in the oesophageal pouch of infants with oesophageal atresia indicating that in infants, as in adults, it probably originates in the lingual serous glands. Since the enzyme is active in the absence of bile salts and has a low pH optimum it is ideally suited to act in the stomach and probably compensates in the premature for low pancreatic lipase activity. The lipolytic activity could be important, not only in the digestion of dietary fat, but in helping to overcome the temporary bile salt deficiency and to solubilize dietary fat through the formation of amphiphilic reaction products.

We report here for the first time a morphological description and observations on some of the secretory proteins of the von Ebner's lingual salivary glands (VEG) of the Syrian hamster. Hamster VEG were macroscopically less distinct, but histologically similar to rat VEG. VEG extracts of hamster and rat were assayed for lipase, alpha-amylase and peroxidase activities. Unlike rat VEG, which is rich in lipase activity, hamster VEG extract had no detectable lipase activity and did not react with antibodies to either rat lingual lipase or human gastric lipase in Western blots. Immunohistochemical reactions with the anti-rat lingual lipase antibody were very weak in hamster VEG and strong in rat VEG. Moderate alpha-amylase enzyme activities and immunohistochemical reactions were demonstrated in both hamster and rat VEG. Peroxidase activity was negligible in the VEG, unlike the high activity in the submandibular glands of both species. An 18 kDa von Ebner's gland protein (VEGP), a member of the lipocalin superfamily of hydrophobic ligandbinding proteins, was abundant in rat VEG, but not detected in hamster VEG. Thus, hamster VEG differs from rat VEG in macroscopic appearance and the absence of lipase and VEGP. It is similar to rat VEG histologically and with regard to the presence of alpha-amylase and absence of peroxidase.

The oral administration of pectin to rats reduced and delayed the peak plasma triacylglycerol concentration. Pectin inhibited the hydrolysis of trioleoylglycerol emulsified with soybean phosphatidylcholine by pancreatic, carboxylester, and lingual lipases in a concentration-dependent manner. However, the effective concentration of pectin for lingual lipase was 100 times lower than that for pancreatic lipase. Pectin did not inhibit the tributyrin- and p-nitrophenylbutyrate-hydrolyzing activities by pancreatic and carboxylester lipase. When low molecular weight pectin was assayed, pectin at a molecular weight of 90,000 (MW 90) most strongly inhibited three lipase activities. When the effect of pH on pectin inhibition was analyzed using pancreatic lipase, strong inhibition was observed at an acidic pH (below pH 7.0). In the assay system, the pancreatic lipase protein levels in the supernatant and fat layer were estimated by Western blotting with an anti-pancreatic lipase antibody. Pectin reduced the amount of pancreatic lipase protein in the fat layer in a concentration-dependent manner and concomitantly increased that in the supernatant. These results suggest that pectin may interact with emulsified substrates and inhibit the adsorption of lipase to the surface of substrate emulsion.

The experiments reported here were designed to study the orosensory factors contributing to the ingestion of sucrose/corn oil mixtures. When a flavor aversion was conditioned to the sucrose/corn oil mixture, the subsequent aversion to the mixture strongly generalized to the corn oil but very little to the sucrose. Rats conditioned with corn oil show a more profound aversion to the sucrose/corn oil mixture than rats conditioned with sucrose, indicating that the salient feature of the sucrose/corn oil mixture is the oil. Aversion to the sucrose/corn oil mixture does not generalize to a sucrose/mineral oil mixture, giving evidence that the textural aspects of the oil do not play a major role in its perception. This flavor aversion to the mixture is further illustrated in very short-term tests where postingestive factors are minimized, indicating a role for the gustatory system in the detection of the sucrose/corn oil mixture. Preliminary experiments are reported where conditioning tests were run with mixtures of sucrose and linoleic acid, one of the fatty acids that is possibly derived from a breakdown of the corn oil in the oral cavity by lingual lipase from von Ebner's Gland.

Von Ebner's glands of the rat are minor salivary serous glands in the posterior portion of the tongue. They secrete two digestive enzymes, lingual lipase and amylase. In this investigation, circadian rhythm in feeding was established under a normal 12 h light/12 h dark cycle, with the rats eating primarily during the dark period. At lights on, the size of the acinar cells and the area of the inclusive secretory granules, and the amount of digestive enzyme activity (lingual lipase and amylase) remaining in the gland was significantly less than in the mid-afternoon, after very little daylight food consumption. However, after 7 days of continuous light the circadian rhythm was altered: the food consumption during the normal night-time hours (5 p.m. to 8 a.m.) went from 88% of total 24 h food consumption to 45%, and during normal daylight hours (8 a.m. to 5 p.m.) from 12% to 55%. These changes were correlated with histometric findings of a near reversal of the areas of acinar cells and secretory granules of a.m. and p.m. samples under continuous light. Lingual lipase activity in the glands went from 35% under 12 h light to 61% under continuous light in the a.m. and from 65% to 39% in the p.m. Amylase activity also showed nearly a reversal in activity remaining in the gland, from 36% at 12 h light to 58% at 24 h light in the a.m. and 64% to 41% for the p.m. samples. These results indicate that the von Ebner's glands of the rat have a circadian rhythm of secretion and storage of secretory proteins that is subject to light entrainment similar to that seen in other exocrine glands such as the parotid and pancreas.

Olestra is a mixture of compounds comprising sucrose esterified with 6-8 long-chain fatty acids. It is not hydrolyzed by pancreatic lipase and as a result is not absorbed from the small intestine. Olestra in general has physical properties similar to those of a triacylglycerol with the same fatty acid composition. Foods made with olestra are virtually identical in taste and texture to those made with typical triacylglycerols. Olestra consumption does not generate hydrolytic products in the small intestine and, therefore, does not generate some of the signals that alter motility in the gastrointestinal tract. A reduction in gastroesophageal reflux with olestra, in contrast to triacylglycerols, is consistent with a lack of effect on stomach emptying. Unlike triacylglycerols that are absorbed in the proximal small intestine, olestra is distributed throughout the small intestine during transit and passes into the colon. In the colon, olestra's effects depend on its physical properties. Liquid nondigestible lipids result in separation of oil from the fecal matrix. Olestra formulations made with specific fatty acid compositions, particularly those containing a solid sucrose polyester component including behenic acid, possess appropriate rheology to hinder separation of oil from the rest of the fecal matrix, thereby reducing gastrointestinal symptoms.

The aim of the present study was to determine, histochemically, the onset and location of production of preduodenal lipase in fetal, suckling, weaned and adult cats. Strong enzymatic activity was localized in the surface mucous cells of the gastric mucosa in animals at postpartal day 1 after ingestion of milk. Activity of gastric lipase persisted as long as animals were nursed. No gastric lipase could be demonstrated in weaned and adult cats. Lingual lipase was not found at any developmental stage examined. Thus, in the newborn cat, lipase of the gastric mucosa is responsible for milk fat lipolysis.

Lipases play a central role in the metabolism of triglyceride-rich lipoproteins in plasma and have therefore been extensively studied at the biochemical and molecular level. Primary structures from various species have been established for lipoprotein lipase (LPL), hepatic lipase (HL), pancreatic lipase (PL), and lysosomal acid lipase (LAL). Comparison of cDNA and genomic structures revealed a gene family of neutral lipases consisting of LPL, HL, and PL. Another gene family of acid lipases comprised LAL, gastric lipase, and lingual lipase.

The changes in contents of pancreatic carboxyl ester lipase, phospholipase A2, and lingual lipase in rats with streptozotocin (STZ)-induced diabetes have been studied. The contents of pancreatic carboxyl ester lipase and phospholipase A2 decreased by 40% and 45%, respectively, 5 days after injection of STZ, whereas pancreatic lipase steadily increased to 100% over control. The content of lingual lipase decreased sharply by more than 90% 2 days after STZ injection, followed by a tendency to recover slightly. Insulin treatment at a dose abolishing the urine glucose in diabetic rats for 3 days restored the contents of pancreatic lipase, carboxyl ester lipase, and lingual lipase but not pancreatic phospholipase A2. The results indicate that lack of insulin action induces an anticoordinate change in gastrointestinal lipolytic enzymes, with decreases in pancreatic carboxyl ester lipase, phospholipase A2, and lingual lipase contents and an increase in pancreatic lipase content.

Rat lingual lipase undergoes maturational increases during postnatal development. The role of thyroxine (T4) in the control of lingual lipase during development was evaluated. T4 given at an early suckling stage (starting day 4 or 5) moderately increased lingual lipase (20–30%) compared to age-matched controls. A similar dose of T4 given later (age > 2 weeks) was ineffective. The T4-sensitive period coincides with a time of low circulating T4, suggesting a role of T4 in modulating the development of lingual lipase in rat pups. Since simultaneous treatment with U486, a type II glucocorticoid receptor antagonist only partially blocked the T4 induction of lingual lipase, T4 appeared to have a direct action on the lingual gland. Pups of propylthiouracil (PTU)-treated dams (previously found to be hypothyroid) showed a delay in the maturation of lingual lipase compared to age-matched pups whose dam was not given PTU. Pups were most sensitive to PTU in the early suckling stage. PTU-induced delayed maturation of lingual lipase was a result of hypothyroidism, since T4 replacement when given early (at the age of 5 days) abolished most of the effect of PTU. When T4 was given later (at the age of 10 days), recovery was much less. This suggests the presence of an early period that is critically dependent on T4 for the full expression of lingual lipase in the rat tongue serous glands. In the same animals following various treatments, their pancreatic lipase and amylase were regulated by the thyroidal status as previously reported, i.e. hyperthyroid increased and hypothyroid decreased the enzyme levels. The magnitude of changes of pancreatic lipase as a result of hypo- or hyperthyroidism, however, was far greater than those observed with lingual lipase indicating a difference in tissue responsiveness to T4.

Rat lingual lipase increases during postnatal development. To evaluate the role of the sympathetic nervous system in the control of lingual lipase during development, suckling rats were chemically sympathectomized by chronic treatment with guanethidine. This treatment was found to be effective in suppressing the developmental increase of lingual lipase. The effect was age dependent and also related to the dose of guanethidine given (i.e., the higher the dose, the more effective the suppression is, up to 40 micrograms/g body wt). The effect of guanethidine on lingual lipase suppression was not a result of induced stress, since simultaneous treatment with RU-38486, a known glucocorticoid receptor antagonist, did not prevent the decrease in lingual lipase activity. Ephedrine, a known sympathomimetic agent, restored the lingual lipase to a near normal level in guanethidine-treated animals, confirming that guanethidine acts through the sympathetic nerves. Furthermore, histochemical studies showed that guanethidine-treatment resulted in the reduction or elimination of catecholaminergic fibers in the von Ebner's glands. The effect of guanethidine was found to be transient, in that the lingual lipase activity showed complete recovery upon withdrawal of the treatment for 1 week. Together, the results indicated that sympathetic nerves have an important regulatory role in lingual lipase in rat pups during development.