Interactions of chlorhexidine with salivary films adsorbed at solid/liquid and air/liquid interfaces

Chlorhexidine is a cationic compound which has been shown to bind to salivary proteins and enamel through electrostatic interactions. The aims of this study were to investigate the interaction of chlorhexidine molecules with salivary films absorbed on solid surfaces with varying physico-chemical characteristics and to investigate the effect of different concentrations of chlorhexidine on the surface tension of saliva. The interactions between 0.2% chlorhexidine digluconate with films adsorbed from whole saliva were monitored by a Rudolph Thin-film ellipsometer equipped with a He-Ne laser (632.8 nm). The films were adsorbed on hydrophilic silica surfaces which were plasma cleaned or on methylated hydrophobic surfaces. Experiments of chlorhexidine adsorption on bare surfaces were also performed. The surface tension of mixtures of whole saliva with various concentrations of (0.1%, 0.2%, 1%) chlorhexidine was monitored with a tensiometer. The results show that chlorhexidine adsorbs on both types of studied substrates. Addition of the substance followed by rinsing caused a partial desorption of the adsorbed pellicles. Furthermore, at all studied concentrations chlorhexidine reduced the interfacial tension. There are indications that the amphiphilic characteristics of the molecule play an important role in the retention of the substance in the oral cavity.     

AUF1 and HuR proteins stabilize interleukin-8 mRNA in human saliva

Human saliva contains thousands of mRNAs, some of which have translational value as diagnostic markers for human diseases. We have found that more than 30% of the mRNAs detected in human saliva contain AU-rich elements (ARE) in their 3' untranslated regions (3'UTR). Since AREs are known to contribute to RNA turnover by forming complexes with ARE-binding proteins, we hypothesized that salivary mRNA stability is mediated by ARE-binding proteins in human saliva. To test this hypothesis, we monitored the in vitro degradation of a radiolabeled ARE-containing salivary mRNA (IL-8) in salivary protein extracts. The degradation of IL-8 mRNA was accelerated by competition for saliva ARE-binding proteins through the addition of excess unlabeled IL-8 mRNA fragments containing 4 tandem AREs. UV cross-linking and immunoprecipitation experiments revealed 2 ARE-binding proteins, AUF1 and HuR, associated with IL-8 mRNA in saliva. These results demonstrate that ARE-binding proteins contribute to the stability of ARE mRNAs in human saliva.     

On the adsorption behaviour of saliva and purified salivary proteins at solid/liquid interfaces

Salivary proteinaceous substances are known to play important roles in the formation of the salivary pellicle. The aim of this study was to investigate some aspects of the interfacial behaviour of selected purified salivary proteins, as well as human saliva secretions, using time-resolved in situ ellipsometry. Hydrophobic methylated silica and hydrophilic pure silica were used as test substrates. Experiments were performed in vitro, preferentially in the low concentration range, with samples of fresh human whole resting saliva, parotid resting saliva and submandibular/sublingual resting saliva. The protein fractions investigated were human MUC5B, PRP-1, PRP-3 and statherin, as well as bovine submaxillary mucin (BSM). The results indicate that the amount of material adsorbed was strongly related to the protein concentration in the range investigated for both pure proteins and secretions. Generally, a larger amount of material was adsorbed onto hydrophobic surfaces than onto hydrophilic ones. However, pure PRP-1 adsorbed in similar amounts to both hydrophilic and hydrophobic surfaces in the concentration range investigated and BSM adsorbed in larger amounts at high concentrations on hydrophilic surfaces. Comparison of the observed adsorption rates for salivary secretions and calculated diffusion rates for individual proteins suggested initial adsorption of low-molecular-weight proteins/peptides. On hydrophilic surfaces the data indicate adsorption of proteins with diffusion rates corresponding to those of statherin, PRP-3 and PRP-1. MUC5B adsorbs at a later stage from both HWS and the individual secretions, which can be explained by a "Vroman effect"-like phenomenon. On hydrophobic surfaces, adsorption rates were found to be faster than those calculated for any of the proteins, and thus smaller proteins/peptides appear to be involved. The similar adsorption behaviour of PRP-1 and parotid saliva (HPS) on hydrophilic surfaces may suggest that long aPRPs account for a substantial portion of the film-forming capacity of HPS. Effects of added electrolyte could be explained by general screening effects and specific Ca2+ binding to serine phosphates in aqueous solutions, but were complex in phosphate buffer. Inter-individual differences in amounts adsorbed from HWS, HPS and HSMSLS, respectively, were not found to be statistically significant.     

Determination of chlorhexidine in saliva and in aqueous solutions.

A new method is presented for the determination of chlorhexidine in centrifuged saliva and in aqueous solutions by means of fluorescence spectroscopy. The method relies on complex formation between chlorhexidine and eosin. The fluorescence value of the chlorhexidine-eosin system decreases with increasing chlorhexidine concentrations. In centrifuged saliva a linear relation was found between the fluorescence at 541 nm and the chlorhexidine content up to about 15 ppm; the reproducibility of the method was found to be better than 1 ppm chlorhexidine. The biological spreading for centrifuged saliva from different participants as well as the spreading due to the period of saliva collection are about 3%. In whole uncentrifuged saliva the fluorescence method has been a detection limit of about 8 ppm chlorhexidine, due to the binding of the compound to salivary constituents. Above 8 ppm chlorhexidine, the biological variation in saliva is such that chlorhexidine determinations by means of fluorescence can be done only with a reproducibility of +/- 10 ppm. Advantages and disadvantages of the ultraviolet spectroscopy method, the radioactive labelling technique, and the fluorescence method for chlorhexidine determinations are discussed.     

Inhibition of calcium phosphate precipitation by human salivary secretions.

Although human saliva is supersaturated with respect to most basic calcium phosphate salts, it lacks the properties which are characteristic of metastable solutions. The purpose of this investigation was to elucidate the reasons for this anomaly. Saliva, salivary ultrafiltrates and the protein fraction of saliva were examined for their effect on the precipitation of calcium phosphate from supersaturated solutions. Salivary ultrafiltrates had little or no effect, but saliva and its macromolecular fraction strongly inhibited precipitation. Serum proteins were inactive, even at 50 times active salivary protein concentrations. The specific inhibitory activity, per unit of protein, of the parotid saliva from 15 subjects varied by a factor of three. Comparison of the unstimulated and stimulated parotid and submandibular secretions of one subject showed activity to be present in the unstimulated salivas and to increase substantially in the stimulated secretions.     

Interaction between surface protein antigens of Streptococcus mutans and human salivary components

The potential involvement of surface antigens (Ags) I/II and III of Streptococcus mutans in its adherence to salivary pellicle-coated tooth surfaces was investigated. The binding of radiolabelled Ag I/II to hydroxyapatite was increased by pretreating the mineral with human parotid saliva, and binding was maintained in the continuous presence of saliva. Binding of Ag III to hydroxyapatite was inhibited by pretreatment with, or in the presence of, saliva. Various aminohexoses, and also tris, inhibited the binding of Ag I/II. When Ags I/II and III were tested for their ability to bind to salivary components separated by SDS gel electrophoresis, several proteins capable of binding Ag I/II were identified, notably 2 proteins of apparent relative molecular mass 28,000 and 38,000. Analysis of these proteins, isolated by micro-preparative electrophoresis, indicated high proportions of proline, glycine, and glutamic acid, and overall compositions similar to basic proline-rich salivary proteins.     

Saliva and dental caries.

A study of saliva and its tooth-protective components reveals at least four important functions of saliva: (1) buffering ability, (2) a cleansing effect, (3) antibacterial action, and (4) maintenance of a saliva supersaturated in calcium phosphate. Several salivary constituents subserve one or more of these functions. Research has yielded important information about organic and inorganic secretory products. It is also clear that saliva as a unique biologic fluid has to be considered in its entirety to account fully for its effects on teeth. Saliva is greater then the sum of its parts. One reason for this is that salivary components display redundancy of function, each often having more than one function. This redundancy, however, does not imply that proteins that share functional roles all contribute to the same degree. For instance, when comparing proteins that inhibit calcium phosphate precipitation, statherin and acidic proline-rich proteins are most potent, whereas histatins, cystatins, and mucins appear to play lesser roles. The complex interaction between proteins is another major factor contributing to saliva's function. In this regard, heterotypic complexes of various proteins have been shown to form on hydroxyapatite. Mucin binding to other salivary proteins, including proline-rich proteins, histatins, cystatins, and statherin, is well documented. The complexes, whether adsorbed to the tooth surface or in saliva, have important implications for bacterial clearance, selective bacterial aggregation on the tooth surface, and control of mineralization and demineralization. Finally, proteolytic activity of saliva generates numerous products whose biologic activities are often different from their parent compounds. Fluoride is another important component of saliva that is discussed separately in other articles in this issue. The ability of saliva to deliver fluoride to the tooth surface constantly makes salivary fluoride an important player in caries protection largely by promoting remineralization and reducing demineralization. Some key properties of salivary components discussed in this article are listed in Table 1. Saliva is well adapted to protection against dental caries. Saliva's buffering capability; the ability of the saliva to wash the tooth surface, to clear bacteria, and to control demineralization and mineralization; saliva's antibacterial activities; and perhaps other mechanisms all contribute to its essential role in the health of teeth. The fact that the protective function of saliva can be overwhelmed by bacterial action indicates the importance of prevention and therapy as in other infectious diseases. The knowledge of functional properties of saliva as well as those of its separate components may permit a better assessment of dental caries susceptibility. Future research is essential to characterize more fully salivary components and their interactions and how these affect the caries process. With such knowledge, the use of modified oral molecules as therapeutic agents may become a reality. Equally intriguing is the prospect of influencing the secretion of salivary components by greater knowledge and control over the secretory processes responsible for the delivery of those components.     

The effects of prolonged gum chewing on salivary flow rate and composition

OBJECTIVE: To determine the effect of gum chewing for 2 h on salivary flow rate and composition. DESIGN: Five male and five females each collected whole saliva at intervals over a 2 h period on three separate days, prior to which they collected unstimulated saliva for 5 min. For one 2 h session they continued to collect only unstimulated saliva while for the others one tablet of Wrigley's Extra peppermint- or fruit-flavoured (peach) gum was chewed continuously. Flow rates were calculated and the saliva was assayed for pH and for Na, K, Ca, Cl, inorganic P and protein concentrations. The data were subjected to repeated-measures ANOVA and Duncan tests. RESULTS: When only unstimulated saliva was collected, there was no significant change in salivary flow rate over the 2 h. With the chewing gums the flow rate increased initially and then, after 35-40 min, fell to similar plateau values which remained significantly higher than the initial unstimulated flow rate and significantly higher than the flow rate at the corresponding time intervals when only unstimulated saliva was collected. With both gums the salivary pH from 2 min to 2 h was significantly higher than that of unstimulated saliva. The changes in the salivary electrolyte and protein concentrations due to the flow rate increase elicited by the chewing gum were largely as expected from previous studies on parotid and submandibular saliva. CONCLUSION: During prolonged chewing gum use, both salivary flow rates and pH remained significantly above the values for unstimulated saliva.     

The interaction of human parotid salivary proteins with hydroxyapatite

The interactions responsible for the formation of the acquired integuments of the human dental enamel surface take place in a complex and poorly, understood milieu. One interaction of particular interest is the selective adsorption of certain salivary proteins on to the hydroxyapatite surface. The purpose of this study was to examine this process in isolation from other events occurring in vivo on the tooth surface and in the saliva. To this end the interaction between various hydroxyapatites and parotid saliva was examined.

Hydroxyapatite and dental enamel powders were found to selectively adsorb seven parotid salivary components from the stimulated and the unstimulated secretion. The two components showing the greatest affinity for these mineral surfaces have been shown, in parallel studies, to be acidic peptides or small proteins of unusual composition. A further four of these components were a group of previously described proline rich proteins, these being characterized by high levels of dicarboxylic amino acids, proline and glycine. The seventh component showed a marked resemblance to the latter materials and appeared to be a sub-group of related proteins. Possible mechanisms underlying the observed interactions are discussed.     

唾液蛋白质组学唾液样本制备的方法学初探

目的：通过对唾液淀粉酶单克隆抗体的制备和鉴定,寻找蛋白质组学研究中样本制备关键环节的可行性方案,为日后唾液蛋白质组学研究奠定基础。方法：将腮腺液经超滤法浓缩蛋白后,进行SDS-PAGE分析,切取分子量为50～65kDa的高丰度蛋白条带,研磨后注射BALB/c小鼠,诱导产生免疫应答并制备人唾液淀粉酶单克隆抗体,应用酶联免疫吸附试验和蛋白免疫印记等方法鉴定。结果：成功制备和鉴定了人唾液淀粉酶单克隆抗体。结论：成功制备和鉴定了唾液淀粉酶单克隆抗体,为唾液蛋白质组学研究中样本制备提供解决方案。     

Salivary proteins: protective and diagnostic value in cariology?

Saliva is essential for a lifelong conservation of the dentition. Various functions of saliva are implicated in the maintenance of oral health and the protection of our teeth: (i) The tooth surface is continuously protected against wear by a film of salivary mucins and proline-rich glycoprotein. (ii) The early pellicle proteins, proline-rich proteins and statherin, promote remineralization of the enamel by attracting calcium ions. (iii) Demineralization is retarded by the pellicle proteins, in concert with calcium and phosphate ions in saliva and in the plaque fluid. (iv) Several salivary (glyco)proteins prevent the adherence of oral microorganisms to the enamel pellicle and inhibit their growth. (v) The salivary bicarbonate/carbonate buffer system is responsible for rapid neutralization of acids. An overview is presented on the major antimicrobial systems in human saliva. Not only the well-known major salivary glycoproteins, including mucins, proline-rich glycoprotein and immunoglobulins, but also a number of minor salivary (glyco)proteins, including agglutinin, lactoferrin, cystatins and lysozyme, are involved in the first line of defense in the oral cavity. Besides, small cationic antimicrobial peptides, e.g. defensins, cathelicidin and the histatins, have come into focus. These are potentially suited as templates for the design of a new generation of antibiotics, since they kill a broad spectrum of microorganisms, while hardly evoking resistance, in contrast to the classical antibiotics.     

Multiple components contribute to ability of saliva to inhibit influenza viruses

Introduction:  Saliva is a potentially important barrier against respiratory viral infection but its mechanism of action is not well studied.
Methods:  We tested the antiviral activities of whole saliva, specific salivary gland secretions, and purified salivary proteins against strains of influenza A virus (IAV) in vitro .
Results:  Whole saliva or parotid or submandibular/sublingual secretions from healthy donors inhibited IAV based on hemagglutination inhibition and neutralization assays. This differs from human immunodeficiency virus (HIV), for which only submandibular/sublingual secretions are reported to be inhibitory. Among purified salivary proteins, MUC5B, scavenger receptor cysteine-rich glycoprotein 340 (salivary gp-340), histatins, and human neutrophil defensins (HNPs) inhibited IAV at the concentrations present in whole saliva. In contrast, some abundant salivary proteins (acidic proline-rich proteins and amylase) had no activity, nor did several other less abundant salivary proteins with known activity against HIV (e.g. thrombospondin or serum leukocyte protease inhibitor). Whole saliva and MUC5B did not inhibit neuraminidase activity of IAV and viral neutralizing and aggregating activity of MUC5B was potentiated by the neuraminidase inhibitor oseltamivir. Hence, MUC5B inhibits IAV by presenting a sialic acid ligand for the viral hemagglutinin. The mechanism of action of histatins requires further study.
Conclusions:  These findings indicate that saliva represents an important initial barrier to IAV infection and underline the complexity of host defense activity of oral secretions. Of interest, antiviral activity of saliva against IAV and HIV differs in terms of specific glandular secretions and proteins that are inhibitory.     

X-ray photoelectron spectroscopic studies of the adsorption of salivary constituents on enamel

Time-dependent change of adsorption of salivary components on the outermost surface layer of enamel was studied by x-ray photoelectron spectroscopy. Adsorption of proteinaceous components, as monitored in terms of the relative mass of nitrogen, was detected within 30 min, increased with time, and reached a plateau at 90 min. Thus, the ratio of nitrogen to calcium in the two-hour sample increased to about 240 times that in the control sample. The ratio of carbon to nitrogen on the surface decreased to about one-half of that in the control sample. The data established the time required for equilibrium between the proteinaceous component in saliva and the amount of material adsorbed onto the tooth surface.     

Effect of salivary biofilm on the adherence of oral bacteria to bleached and non-bleached restorative material.

OBJECTIVE: The objective of this work was to examine the effect of in vitro salivary biofilm on the adherence of oral bacteria to bleached and non-bleached restorative material (Charisma). METHODS: Charisma samples, prepared in silicon models, were treated with either 10% carbamide peroxide (CP) or 10% hydrogen peroxide (HP). After incubation with the bleaching agent for a period of one, two or three days, the samples were coated with freshly collected human saliva. The adsorption pattern of the saliva to the restorative material was determined using gel electrophoresis coupled with computerized densitometry techniques. The amount of salivary proteins adsorbed onto the treated surfaces was measured using the Bradford method. Sucrose-dependent bacterial adhesion to the salivary-coated Charisma was tested using radio-labeled Streptococcus mutans, Streptococcus sobrinus and Actinomyces viscosus. Adhesion of each bacterium to surfaces pretreated with the bleaching agents was compared with saliva coated bleached surfaces. RESULTS: The profile of salivary proteins adsorption followed a similar pattern in Charisma samples pretreated with either CP or HP or untreated samples. However, the total amount of salivary proteins adsorbed onto the samples decreased after bleaching with CP or HP. Salivary biofilm, coating the surface of the restorative material, significantly decreased sucrose-dependent adhesion of Streptococcus sobrinus and Streptococcus mutans to the bleached and non-bleached surfaces, compared to non-coated specimens (p < 0.05). Saliva had a minor effect on adhesion of Actinomyces viscosus. SIGNIFICANCE: Our study demonstrates the importance of salivary biofilm in controlling adhesion of oral bacteria to restorative material pretreated with bleaching agents or untreated.     

Protein composition of whole and parotid saliva in healthy and periodontitis subjects

Cystatins are physiological inhibitors of cysteine proteinases and they are widely distributed in human tissues and body fluids including saliva. We previously reported an increased cystatin activity in whole saliva of gingivitis and periodontitis subjects. Based on this result we decided to investigate the type and origin of cystatins involved in this increased cystatin activity by collecting both whole and parotid saliva of 25 healthy and 30 periodontitis subjects. Saliva samples were quantified for cystatins S and C by enzyme-linked immunosorbent assay and cystatin activities were measured toward papain. Besides, three other salivary proteins were determined: the plasma protein albumin, the typical parotid derived amylase and the salivary immunoglobulin IgA. The present investigation shows that levels of total protein and cystatin activity as well as the levels of glandular derived proteins amylase and cystatin C were significantly higher in whole and parotid saliva of subjects with periodontitis than in healthy controls. Cystatin S, the major salivary cystatin. however was higher in the whole saliva of the healthy group. Whole saliva concentrations of albumin and IgA, originating from sources other than the glandular cells, were not different between healthy and periodontitis subjects and were also not correlated with the typical salivary gland proteins. In conclusion, this study provides additional evidence that the human salivary glands may respond to an inflammatory disease of the oral cavity, periodontitis, by enhanced synthesis of some acinar proteins.     

Zinc and copper play a role in coaggregation inhibiting action of Porphyromonas gingivalis

Background/aim:  We investigated the mechanisms of adherence of salivary and serum proteins, which mimic gingival crevicular fluid (GCF), to Porphyromonas gingivalis , and the effects of these adhered proteins on coaggregation and hemagglutination properties.
Methods:  The amounts of salivary and serum proteins adhering to P. gingivalis were determined using ³H-labeled and non-labeled proteins. The coaggregation between P. gingivalis and Streptococcus oralis or Streptococcus gordonii was observed. Hemagglutination was evaluated using sheep erythrocytes. Proteins that interacted with zinc or copper in saliva and serum and on P. gingivalis were examined using sodium dodecyl sulfate–polyacrylamide gel electrophoresis.
Results:  The amount of salivary or serum proteins that adhered to the surface of P. gingivalis strains was increased by cations, especially zinc and copper ions. The pretreatment of bacterial cells with salivary or serum proteins before the assay inhibited coaggregation with gram-positive bacteria and hemagglutination. These phenomena were enhanced by the presence of zinc or copper ions during the pretreatment of P. gingivalis with proteins. We detected protein bands that were related to these cations in saliva and serum and on P. gingivalis.
Conclusions:  Our findings suggest that zinc and copper ions markedly enhanced the adhesion and accumulation of salivary and serum proteins on cells of P. gingivalis and inhibited the coaggregation and hemagglutination of P. gingivalis . These cations might be useful for limiting the settlement of P. gingivalis in the gingival sulcus with the goal of preventing periodontal disease.     

Adsorbed salivary acidic proline-rich proteins contribute to the adhesion of Streptococcus mutans JBP to apatitic surfaces

Experimental pellicles formed on hydroxyapatite (HA) beads from parotid or submandibular saliva promoted the adhesion of Streptococcus mutans JBP cells to a greater extent than did pellicles prepared from buffer, human plasma, or serum. The nature of the salivary components responsible was studied by the preparation of pellicles from fractions of parotid saliva obtained by chromatography on Trisacryl GF 2000 columns. Two groups of fractions promoted attachment of the organism. Components migrating in the high-molecular-weight mucin fraction were most effective, but a later-eluting fraction also possessed adhesion-promoting activity. Subfractionation of the latter material indicated that the adhesion-promoting activity was associated with the acidic proline-rich proteins (PRPs). Pellicles prepared from 10-20-micrograms/mL solutions of pure PRP-1 were effective in promoting attachment of S. mutans JBP cells. PRP-3 was less effective, while human salivary statherin, fibrinogen, fibronectin, type 1 collagen, and the amino-terminal tryptic peptide derived from PRP-1 were ineffective. The quantities of 150-residue and 106-residue PRPs and of statherin, which became incorporated into experimental pellicles prepared from saliva, were estimated with use of radiolabeled protein tracers. The data obtained suggest that these proteins compete for similar binding sites on HA, and that their ratios in saliva would therefore influence the quantity of the larger PRPs that become incorporated into the pellicle. Such competition may contribute to the variability observed in the adhesion-promoting activities of different saliva samples.     

Effect of critical surface tension on retention of oral microorganisms

Abstract – The effect of critical surface tension on the initial retention of microorganisms from unstimulated human saliva was tested in a flow cell system. Prior to each experiment the total numbers and the morphotypes of microorganisms present in saliva were recorded. The test surfaces were prepared to display known increasing critical surface tensions, as verified and standardized by contact angle measurements. Surfaces of initially low (20–22 mN/m), medium (35–38 mN/m) and high (>50 mN/m) critical surface tension were exposed to saliva at a flow rate of 1 ml/min. Microbiota and biofilm material associated with the test surface after 15 min of salivary exposure, were then subjected to standard detachment forces, by introducing a cell-free rinsing fluid at two different shear rates. Both the attachment and the detachment phases were executed at room temperature or 37°C. The retained population was counted in three different zones of the test surfaces with a light microscope and statistically tested for correlation to the main variables (critical surface tension, flow rate and temperature) and interactions. Retention success was significantly dependent on the initial critical surface tension and the flow rate. Surfaces of medium critical surface tension, representative of human tooth surfaces and most restorative dental materials, retained the highest numbers of microorganisms in comparison with the other surfaces tested, with no statistically verified selectivity in proportions of retained coccoid and rodshaped microorganisms for any surface. A 30-fold increase of the flow rate resulted in a 70–80% reduction of the retention success, with a higher relative number of cocci present on all the test surfaces. These results demonstrate that initial retention of microorganisms to surfaces is non-specific with regard to morphotypes, but is strongly related both to the mechanical removal forces and the surface energetic state of the solid surface exposed. Retention of microbial populations at interfaces might, advance selection of the critical surface tensions and predicted if shear forces at given sites are known.     

Surgical treatment of dry eye syndrome: conjunctival graft of the minor salivary gland

Despite the availability of efficient tear substitutes, many patients with dry eye syndrome experience severe corneal injuries and a subsequent loss of vision. Surgical techniques using mayor salivary glands to provide a substitute for tears have been reported; with this technique the drainage of saliva goes into the conjunctival fornix, permitting corneal and conjunctival humidification. The authors describe a new surgical approach in which minor salivary glands are autotransplanted into the conjunctival fornix by means of a graft of the intraoral mucosa-transporting salivary glands. This approach was used in a 56-year-old woman with a 2-year history of refractory and pharmacologically untreatable dry eye syndrome caused by Sj?gren's syndrome. The right eye had more severe corneal and conjunctival lesions than did the contralateral one, so the treatment was planned in the right eye only. A weekly follow-up during the first 6 months confirmed the significant improvement of dry eye symptoms in the surgically treated eye. Three months after surgery, a biopsy was performed in the minor salivary gland graft, and the histologic findings revealed the presence of glandular acinus, duct with mucin content, and lymphocyte infiltration. The significant improvement obtained in this patient suggests that the secretion from the grafted salivary minor glands was better in promoting homeostasis of the ocular surface than are artificial tears. This may be explained by: (1) The lacrimal and salivary secretions contain biologically active constituents that may protect from infection and promote normal growth epithelium; (2) The secreted mucin is thought to coat the epithelial surface, reducing the high surface tension of the eye wetted by aqueous tears; (3) The thick secretions of the minor gland might act in reducing the evaporation of the underlying tear layer and form a hydrophobic barrier along the lid margin that can retain the lid margin tear string and prevent its flow onto the skin. Minor gland salivary autotransplant is a new surgical technique with effectiveness demonstrated in one patient, but the scientific explanation is not clear; additional experience with more cases could confirm the initial success.     

Fractionation of salivary micelle-like structures by gel chromatography

Globular structures have been demonstrated in human parotid saliva by transmission electron microscopy and photon correlation spectroscopy. The aim of this study was to fractionate these salivary globular structures for analytical and preparative purposes using a gel-filtration material capable of separating spherical particles up to 300-400 nm in diameter. Freshly obtained parotid saliva was applied to a Sephacryl S-1000 column. Peak fractions were collected and prepared for transmission electron microscopy (TEM) or for amino acid analysis. Bovine milk was included as the casein micelles by TEM appear to be similar to the salivary aggregates and their elution profiles are known. The salivary globular structures were eluted in one major peak. TEM of negatively stained samples from the peak fractions demonstrated globular protein aggregates consistent with the salivary structures in parotid saliva. Amino acid analysis showed characteristic amino acid profiles with unusual high levels of proline, 40-45%. The casein micelles were eluted in one major peak and separated from the whey proteins. This study indicates that the salivary globular structures can be isolated by gel chromatography. The amino acid analysis indicates that proline-rich proteins may be an important fraction of the salivary globular structures.