Prostate-specific antigen: An unfamiliar protein in the human salivary glands.

OBJECTIVES: The presence of prostate-specific antigen (PSA) in saliva and salivary glands has been reported. Nevertheless, its release pathway in these glands remains to be elucidated. Here, we showed PSA subcellular distribution focusing on its plausible route in human salivary parenchyma. MATERIALS AND METHODS: Sections of parotid and submandibular glands were subjected to the immunohistochemical demonstration of PSA by the streptavidin-biotin method revealed by alkaline phosphatase. Moreover, ultrathin sections were collected on nickel grids and processed for immunocytochemical analysis, to visualize the intracellular distribution pattern of PSA through the observation by transmission electron microscopy. RESULTS: By immunohistochemistry, in both parotid and submandibular glands PSA expression was detected in serous secretory acini and striated ducts. By immunocytochemistry, immunoreactivity was retrieved in the cytoplasmic compartment of acinar and ductal cells, often associated with small cytoplasmic vesicles. PSA labeling appeared also on rough endoplasmic reticulum and in the acini's lumen. A negligible PSA labeling appeared in most of the secretory granules of both glands. CONCLUSIONS: Our findings clearly support that human parotid and submandibular glands are involved in PSA secretion. Moreover, based on the immunoreactivity pattern, its release in oral cavity would probably occur by minor regulated secretory or constitutive-like secretory pathways.

Integrated Bioinformatics and Validation Reveal Potential Biomarkers Associated With Progression of Primary Sjögren's Syndrome.

Background: Primary Sjögren's syndrome (pSS) is a chronic systemic autoimmune disease of the exocrine glands characterized by specific pathological features. Previous studies have pointed out that salivary glands from pSS patients express a unique profile of cytokines, adhesion molecules, and chemokines compared to those from healthy controls. However, there is limited evidence supporting the utility of individual markers for different stages of pSS. This study aimed to explore potential biomarkers associated with pSS disease progression and analyze the associations between key genes and immune cells. Methods: We combined our own RNA sequencing data with pSS datasets from the NCBI Gene Expression Omnibus (GEO) database to identify differentially expressed genes (DEGs) via bioinformatics analysis. Salivary gland biopsies were collected from 14 pSS patients, 6 non-pSS patients, and 6 controls. Histochemical staining and transmission electron micrographs (TEM) were performed to macroscopically and microscopically characterize morphological features of labial salivary glands in different disease stages. Then, we performed quantitative PCR to validate hub genes. Finally, we analyzed correlations between selected hub genes and immune cells using the CIBERSORT algorithm. Results: We identified twenty-eight DEGs that were upregulated in pSS patients compared to healthy controls. These were mainly involved in immune-related pathways and infection-related pathways. According to the morphological features of minor salivary glands, severe interlobular and periductal lymphocytic infiltrates, acinar atrophy and collagen in the interstitium, nuclear shrinkage, and microscopic organelle swelling were observed with pSS disease progression. Hub genes based on above twenty-eight DEGs, including MS4A1, CD19, TCL1A, CCL19, CXCL9, CD3G, and CD3D, were selected as potential biomarkers and verified by RT-PCR. Expression of these genes was correlated with T follicular helper cells, memory B cells and M1 macrophages. Conclusion: Using transcriptome sequencing and bioinformatics analysis combined with our clinical data, we identified seven key genes that have potential value for evaluating pSS severity.

Microanatomical and secretory characterization of the salivary gland of the Rhodnius prolixus (Hemiptera, Reduviidae, Triatominae), a main vector of Chagas disease.

Rhodnius prolixus is the principal vector of Trypanosoma cruzi, the aetiological agent of Chagas disease in American countries. This insect is haematophagous during all life cycles and, to antagonize its haemostatic, inflammatory and immune systems, it secretes saliva while feeding on the vertebrate host's blood. Here, we investigated characteristic changes of the salivary glands (SG) that occur during insect development. Two pairs of lobules and ducts comprise the SG of R. prolixus. The organ's size increases over time, but the microanatomical structures are preserved during insect development. Both lobules have a single layer epithelium formed by binucleated cells, which surrounds the saliva reservoir. The principal lobule presents higher polysaccharide and total protein contents than the accessory lobe. A network of external muscle layers is responsible for organ contraction and saliva release. Apocrine, merocrine and holocrine secretion types occur in the secretory epithelium. Dopamine, serotonin and tyrosine-hydroxylase are neural-related molecules that regulate SG function both during and after feeding.

Mutation in Bombyx mori fibrohexamerin (P25) gene causes reorganization of rough endoplasmic reticulum in posterior silk gland cells and alters morphology of fibroin secretory globules in the silk gland lumen.

Larvae of many lepidopteran species produce a mixture of secretory proteins, known as silk, for building protective shelters and cocoons. Silk consists of a water-insoluble silk filament core produced in the posterior silk gland (PSG) and a sticky hydrophilic coating produced by the middle silk gland (MSG). In Bombyx mori, the fiber core comprises three proteins: heavy chain fibroin (Fib-H), light chain fibroin (Fib-L) and fibrohexamerin (Fhx, previously referred to as P25). To learn more about the role of Fhx, we used transcription activator-like effector nuclease (TALEN) mutagenesis and prepared a homozygous line with a null mutation in the Fhx gene. Our characterization of cocoon morphology and silk quality showed that the mutation had very little effect. However, a detailed inspection of the secretory cells in the posterior silk gland (PSG) of mid-last-instar mutant larvae revealed temporary changes in the morphology of the endoplasmic reticulum. We also observed a morphological difference in fibroin secretory globules stored in the PSG lumen of Fhx mutants, which suggests that their fibroin complexes have a slightly lower solubility. Finally, we performed an LC-MS-based quantitative proteomic analysis comparing mutant and wild-type (wt) cocoon proteins and found a high abundance of a 16 kDa secretory protein likely involved in fibroin solubility. Overall, our study shows that whilst Fhx is dispensable for silk formation, it contributes to the stability of fibroin complexes during intracellular transport and affects the morphology of fibroin secretory globules in the PSG lumen.

The Effect of the Hypertrophy Virus (MdSGHV) on the Ultrastructure of the Salivary Glands of Musca domestica (Diptera: Muscidae).

The salivary glands of insects play a key role in the replication cycle and vectoring of viral pathogens. Consequently, Musca domestica (L.) (Diptera: Muscidae) and the Salivary Gland Hypertrophy Virus (MdSGHV) serve as a model to study insect vectoring of viruses. A better understanding of the structural changes of the salivary glands by the virus will help obtain a better picture of the pathological impact the virus has on adult flies. The salivary glands are a primary route for viruses to enter a new host. As such, studying the viral effect on the salivary glands is particularly important and can provide insights for the development of strategies to control the transmission of vector-borne diseases, such as dengue, malaria, Zika, and chikungunya virus. Using scanning and transmission electron microscopic techniques, researchers have shown the effects of infection by MdSGHV on the salivary glands; however, the exact location where the infection was found is unclear. For this reason, this study did a close examination of the effects of the hypertrophy virus on the salivary glands to locate the specific sites of infection. Here, we report that hypertrophy is present mainly in the secretory region, while other regions appeared unaffected. Moreover, there is a disruption of the cuticular, chitinous lining that separates the secretory cells from the lumen of the internal duct, and the disturbance of this lining makes it possible for the virus to enter the lumen. Thus, we report that the chitinous lining acts as an exit barrier of the salivary gland.

The salivary glands of Brontocoris tabidus (Heteroptera: Pentatomidae): Morphology and secretory cycle.

Brontocoris tabidus (Signoret) (Heteroptera: Pentatomidae) is a zoophytophagous insect used for biological control in agriculture and forest systems because its nymphs and adults feed on insects and plants. The predatory Pentatomidae insert the mouthparts into the prey, releasing saliva to paralysis and kills the insect, as well as digest body parts to be sucked in a preliminary extra-oral digestion. In a short period of time, this insect shows the ability to feed again, suggesting the existence of a constant and abundant secretory cycle in the salivary glands. This study evaluated the morphological, histochemical and ultrastructural changes of the salivary glands of B. tabidus in fed and starved insects. The salivary complex of this predatory bug has a pair of bilobed salivary glands and a pair of tubular accessory salivary glands. The accessory glands have the lumen lined by a thick non-cuticular layer rich in glycoproteins. The secretory cells of the B. tabidus principal salivary glands have constant secretory activity, with each lobe producing different substances. The physiological processes that occur in the salivary gland of B. tabidus indicate that the insect needs to feed constantly, corroborating the potential of this insect to be used in biological control programs.

The histology and ultrastructure of the salivary glands of Neopanorpa longiprocessa (Mecoptera: Panorpidae).

The salivary glands of Panorpidae usually exhibit distinct sexual dimorphism and are closely related to the nuptial feeding behavior. In this study, the salivary glands of Neopanorpa longiprocessa were investigated using light microscopy and transmission electron microscopy. The salivary glands are tubular labial glands and consist of a scoop-shaped salivary pump, a common salivary duct, and a pair of salivary tubes. The male and female salivary glands are remarkably different in the bifurcation position of the common salivary duct and the length and shape of the secretory tubes. Compared with the simple female salivary glands, the male's are more developed as their paired elongated salivary tubes can be divided into two parts, the glabrate anterior tube and the posterior tube with many secretory tubules. The ultrastructural study shows that the male salivary tubes have strong secretory function. The existence of different secretion granules indicates that there are some chemical reactions or mixing occurring in the lumen. Based on the ultrastructural characteristics, the functions of the different regions of the salivary tube have been speculated. The relationship between the salivary glands and nuptial feeding behavior of N. longiprocessa has been briefly discussed based on the structure of the salivary glands.

Gross morphology and ultrastructure of the salivary glands of the stink bug predator Eocanthecona furcellata (Wolff).

Eocanthecona furcellata Wolff (Hemiptera: Pentatomidae) is a native generalist predator which attacks and kills its prey by first inserting its stylet into the prey's body and then injecting saliva into it. Here, we describe the histology and ultrastructure of its salivary glands. The study showed that the salivary glands were made up of pairs of principal and tubular accessory salivary glands. The principal salivary glands were bilobed and consisted of a smaller anterior lobe and a larger elongated posterior lobe. The ducts of the principal and accessory salivary glands were located in a narrow region between the anterior and posterior lobe known as the hilum. The principal salivary gland was lined with a single-layered epithelium. The cells cytoplasm was enriched with rough endoplasmic reticulum and secretory, and the nucleus showed a higher level of uncondensed chromatin. The basal region of the cell had plasma membrane infoldings. The cytoplasm of the accessory gland was rich in rough endoplasmic reticulum and many large cavities. The ducts of the principal salivary gland were made up of a single layer of flattened cells which had a thin cuticle lining the apical portion. Variation in the lumen content of the different lobes, which made up the principal gland suggested that their chemical products also varied. These results indicate that these two salivary glands produce the proteins found in the saliva.

Transmission Electron Microscopy as a Tool to Study the Toxicological Effects of Thiamethoxam in Workers of Atta sexdens (Myrmicinae, Attini).

Thiamethoxam is a neonicotinoid that has been used to control insect pests. The literature reports a few behavioral studies evaluating the toxic effect of thiamethoxam in ants; however, there are scarce studies at the cellular level. The present research evaluated the effects of thiamethoxam in labial (LG) and mandibular glands (MG), fat bodies (FB), and Malpighian tubules (MT) of workers of Atta sexdens, using transmission electron microscopy. The duct and secretory cells of LG were profoundly affected, then the production of saliva can be compromised, as well as its quality and subsequent use. In MG, reservoir and canaliculi cells presented slight alterations; however, MG secretory cells presented vacuoles containing lamellar structures, increased lipid production, and a large amount of mitochondria, which may lead to organ's malfunctioning. The FB cell alterations do not seem enough to cause significant changes that lead to cell death. Prominent changes in MT, such as loss of the electron-dense concentric ring, increased smooth endoplasmic reticulum, loss of basal infolds, vacuoles containing mineralized granules, and lamellar structures associated with mitochondria, suggest that their excretory function is compromised. In conclusion, thiamethoxam acts not only in the nervous system but also contributes to systemic toxicity on the target organism.

Conserved and distinct morphological aspects of the salivary glands of sand fly vectors of leishmaniasis: an anatomical and ultrastructural study.

BACKGROUND: Sand flies are vectors of Leishmania spp., the causative agents of leishmaniasis in vertebrates, including man. The sand fly saliva contains powerful pharmacologically active substances that prevent hemostasis and enhance Leishmania spp. infections. On the other hand, salivary proteins can protect vaccinated mice challenged with parasites. Therefore, sand fly salivary proteins are relevant for the epidemiology of leishmaniasis and can be a potential target for a vaccine against leishmaniasis. Despite this, studies on sand fly salivary glands (SGs) are limited. METHODS: The present study analyzes, in detail, the morphology, anatomy and ultrastructure of the SGs of sand fly vectors of the genera Lutzomyia and Phlebotomus. We used histology, transmission and scanning electron microscopy and lectin labeling associated with confocal laser microscopy. RESULTS: The SGs have conserved and distinct morphological aspects according to the distinct sand fly species. Each SG has a single rounded lobe constituting of c.100-120 secretory cells. The SG secretory cells, according to their ultrastructure and lectin binding, were classified into five different subpopulations, which may differ in secretory pathways. CONCLUSIONS: To the best of our knowledge, these morphological details of sand fly salivary glands are described for the first time. Further studies are necessary to better understand the role of these different cell types and better relate them with the production and secretion of the saliva substances, which has a fundamental role in the interaction of the sand fly vectors with Leishmania.

Inhibition of Alk signaling promotes the induction of human salivary-gland-derived organoids.

Hyposalivation and xerostomia are the cause of several morbidities, such as dental caries, painful mucositis, oral fungal infections, sialadenitis and dysphagia. For these reasons, preservation of normal saliva secretion is critical for the maintenance of functionally normal oral homeostasis and for keeping good health. Several strategies for restoring salivary gland function have been reported, from different points of view, based on the use of salivary-gland-derived epithelial stem/progenitor cells and tissue engineering approaches to induce organoids that mimic in vivo salivary glands. In this study, we clarified that inhibition of activin receptor-like kinase (Alk) signaling was essential for the induction of human salivary-gland-derived organoids, and demonstrated the usefulness of such organoids as an inflammatory disease model. In inflammatory conditions like sialadenitis, in general, pro-inflammatory cytokines such as tumor necrosis factor-α (TNF-α, also known as TNF) are upregulated, but their function is still unclear. In our established human salivary-gland-derived organoid culture system, we successfully induced organoid swelling by stimulation with carbachol, a non-selective cholinergic agonist, and forskolin, an activator of cystic fibrosis transmembrane conductance regulator (CFTR). Furthermore, we found that this organoid swelling was inhibited by TNF-α. From these results, we could clarify the inhibitory function of TNF-α on saliva secretion in vitro Thus, our established human salivary-gland-derived organoids would be useful for in vitro analyses of the morphological and functional changes involved in salivary gland dysfunctions in several research fields, such as pathobiology, inflammation and regenerative medicine.This article has an associated First Person interview with the first author of the paper.

Effects of short- and long-term exposure to cadmium on salivary glands and fat body of soil centipede Lithobius forficatus (Myriapoda, Chilopoda): Histology and ultrastructure.

Cadmium (Cd) is the most widely studied heavy metal in terms of food-chain accumulation and contamination because it can strongly affect all environments (e.g., soil, water, air). It can accumulate in different tissues and organs and can affect the organism at different levels of organization: from organs, tissues and cells though cell organelles and structures to activation of mechanisms of survival and cell death. In soil-dwelling organisms heavy metals gather in all tissues with accumulation properties: midgut, salivary glands, fat body. The aim of this study was to describe the effects of cadmium on the soil species Lithobius forficatus, mainly on two organs responsible for gathering different substances, the fat body and salivary glands, at the ultrastructural level. Changes caused by cadmium short- and long-term intoxication, connected with cell death (autophagy, apoptosis, necrosis), and the crosstalk between them, were analyzed. Adult specimens of L. forficatus were collected in a natural environment and divided into three experimental groups: C (the control group), Cd1 (cultured in soil with 80 mg/kg of CdCl2 for 12 days) and Cd2 (cultured in soil with 80 mg/kg of CdCl2 for 45 days). Transmission electron microscopy revealed ultrastructural alterations in both of the organs analyzed (reduction in the amount of reserve material, the appearance of vacuoles, etc.). Qualitative analysis using TUNEL assay revealed distinct crosstalk between autophagy and necrosis in the fat body adipocytes, while crosstalk between autophagy, apoptosis and necrosis in the salivary glands was detected in salivary glands of the centipedes examined here. We conclude that different organs in the body can react differently to the same stressor, as well as to the same concentration and time of exposure. Different mechanisms at the ultrastructural level activate different types of cell death and with different dynamics.

Detailed ultrastructure of the Hirudo (Annelida: Hirudinea) salivary gland.

Medical leeches have been widely used in medical applications and treatments for millennia. Studies on the salivary glands of blood-sucking leeches have focused on their bioactive secretions and mechanisms of action, with little attention to ultrastructure. In this study, we examined dissected embryonic and adult Hirudo verbana salivary glands by scanning electron microscopy (SEM). Gland cells of embryos were physically separated while adults displayed highly developed cell bunches in which each cell was connected to others by fine channels. Channels from each bunch combined to form a larger canal that opened to the jaw. Secreted material from these glands prevent blood from clotting and allow the adult to feed while sucking blood.

Mechanisms of programmed cell death in the midgut and salivary glands from Bradysia hygida (Diptera: Sciaridae) during pupal-adult metamorphosis.

Programmed cell death is involved with the degeneration/remodeling of larval tissues and organs during holometabolous development. The midgut is a model to study the types of programmed cell death associated with metamorphosis because its structure while degenerating is a substrate for the formation of the adult organ. Another model is the salivary glands from dipteran because their elimination involves different cell death modes. This study aimed to investigate the models of programmed cell death operating during midgut replacement and salivary gland histolysis in Bradysia hygida. We carried out experiments of real-time observations, morphological analysis, glycogen detection, filamentous-actin localization, and nuclear acridine orange staining. Our findings allow us to establish that an intact actin cytoskeleton is required for midgut replacement in B. hygida and nuclear condensation and acridine orange staining precede the death of the larval cells. Salivary glands in histolysis present cytoplasmic blebbing, nuclear retraction, and acridine orange staining. This process can be partially reproduced in vitro. We propose that the larval midgut death involves autophagic and apoptotic features and apoptosis is a mechanism involved with salivary gland histolysis.

Crumbs organizes the transport machinery by regulating apical levels of PI(4,5)P2 in Drosophila.

An efficient vectorial intracellular transport machinery depends on a well-established apico-basal polarity and is a prerequisite for the function of secretory epithelia. Despite extensive knowledge on individual trafficking pathways, little is known about the mechanisms coordinating their temporal and spatial regulation. Here, we report that the polarity protein Crumbs is essential for apical plasma membrane phospholipid-homeostasis and efficient apical secretion. Through recruiting ßHeavy-Spectrin and MyosinV to the apical membrane, Crumbs maintains the Rab6-, Rab11- and Rab30-dependent trafficking and regulates the lipid phosphatases Pten and Ocrl. Crumbs knock-down results in increased apical levels of PI(4,5)P2 and formation of a novel, Moesin- and PI(4,5)P2-enriched apical membrane sac containing microvilli-like structures. Our results identify Crumbs as an essential hub required to maintain the organization of the apical membrane and the physiological activity of the larval salivary gland.

Anatomy, Histology, and Ultrastructure of Salivary Glands of the Burrower Bug, Scaptocoris castanea (Hemiptera: Cydnidae).

The burrower bug Scaptocoris castanea Perty, 1830 (Hemiptera: Cydnidae) is an agricultural pest feeding on roots of several crops. The histology and ultrastructure of the salivary glands of S. castanea were described. The salivary system has a pair of principal salivary glands and a pair of accessory salivary glands. The principal salivary gland is bilobed with anterior and posterior lobes joined by a hilus where an excretory duct occurs. The accessory salivary gland is tubular with a narrow lumen that opens into the hilus near the excretory duct, suggesting that its secretion is stored in the lumen of the principal gland. The cytoplasm of the secretory cells is rich in the rough endoplasmic reticulum, secretory vesicles with different electron densities and mitochondria. At the base of the accessory gland epithelium, there were scattered cells that do not reach the gland lumen, with the cytoplasm rich in the rough endoplasmic reticulum, indicating a role in protein production. Data show that principal and accessory salivary glands of S. castanea produce proteinaceous saliva. This is the first morphological description of the S. castanea salivary system that is similar to other Hemiptera Pentatomomorpha, but with occurrence of basal cells in the accessory salivary gland.

Functional Fluorescence Microscopy Imaging: Quantitative Scanning-Free Confocal Fluorescence Microscopy for the Characterization of Fast Dynamic Processes in Live Cells.

Functional fluorescence microscopy imaging (fFMI), a time-resolved (21 µs/frame) confocal fluorescence microscopy imaging technique without scanning, is developed for quantitative characterization of fast reaction-transport processes in solution and in live cells. The method is based on massively parallel fluorescence correlation spectroscopy (FCS). Simultaneous excitation of fluorescent molecules in multiple spots in the focal plane is achieved using a diffractive optical element (DOE). Fluorescence from the DOE-generated 1024 illuminated spots is detected in a confocal arrangement by a matching matrix detector comprising 32 × 32 single-photon avalanche photodiodes (SPADs). Software for data acquisition and fast auto- and cross-correlation analysis by parallel signal processing using a graphic processing unit (GPU) allows temporal autocorrelation across all pixels in the image frame in 4 s and cross-correlation between first- and second-order neighbor pixels in 45 s. We present here this quantitative, time-resolved imaging method with single-molecule sensitivity and demonstrate its usefulness for mapping in live cell location-specific differences in the concentration and translational diffusion of molecules in different subcellular compartments. In particular, we show that molecules without a specific biological function, e.g., the enhanced green fluorescent protein (eGFP), exhibit uniform diffusion. In contrast, molecules that perform specialized biological functions and bind specifically to their molecular targets show location-specific differences in their concentration and diffusion, exemplified here for two transcription factor molecules, the glucocorticoid receptor (GR) before and after nuclear translocation and the Sex combs reduced (Scr) transcription factor in the salivary gland of Drosophila ex vivo.

The Anatomy and Ultrastructure of the Digestive Tract and Salivary Glands of Hishimonus lamellatus (Hemiptera: Cicadellidae).

In recent years, we found that Hishimonus lamellatus Cai et Kuoh is a potential vector of jujube witches'-broom phytoplasma. However, little is known about the anatomy and histology of this leafhopper. Here, we examined histology and ultrastructure of the digestive system of H. lamellatus, both by dissecting and by semi- and ultrathin sectioning techniques. We found that the H. lamellatus digestive tract consists of an esophagus, a filter chamber, a conical midgut and midgut loop, Malpighian tubules, an ileum, and a rectum. Furthermore, both the basal region of the filter chamber epithelium and the apical surface of the midgut epithelium have developed microvilli. We also identify the perimicrovillar membrane, which ensheaths the microvilli of midgut loop enterocyte, and the flame-like luminal membrane, which covers the microvilli of the conical midgut epithelium. In addition, H. lamellatus has the principal and accessory salivary glands. Our observations also showed that the endoplasmic reticulum, mitochondria, and secretory granules were all highly abundant in the secretory cells of the principal salivary glands, while the accessory glands consist of only one ovate or elbow-like acinus. We also briefly contrast the structure of the gut of H. lamellatus with those of other leafhopper species. These results intend to offer help for the future study on the histological and subcellular levels of phytopathogen-leafhopper relationships, including transmission barriers and the binding sites of pathogens and other microorganisms within their leafhopper vectors.

Fine infrastructure of released and solidified Drosophila larval salivary secretory glue using SEM.

The Golgi-derived large secretory granules of Drosophila salivary glands (SGs) constitute the components of the salivary glue secretion (Sgs). The Sgs represents a highly special and unique extracellular composite glue matrix that has not yet been identified outside of Cyclorrhaphous Dipterans. For over half a century, the only major and unambiguously documented function of the larval salivary glands was to produce a large amount of mucinous glue-containing secretory granules that, when released during pupariation, serves to affix the freshly formed puparia to a substrate. Besides initial biochemical characterization of the Sgs proteins and cloning of their corresponding Sgs genes, very little is known about other properties and functions of the Sgs glue. We report here observations on the fine SEM-ultrastructure of the Sgs glue released into to the lumen of SGs, and after it has been expectorated and solidified into the external environment. Surprisingly, in contrast to long held expectations, it appears to be a highly structured bioadhesive mass with an internal spongious to trabecular infrastructure, reflecting the state of its hydratation. We also found that in addition to its cementing properties, it is highly efficient at glueing and trapping microorganisms, and thus may serve a potentially very important immune and defense role. High hydration capacity, the speed by which this glue can dry, uniqueness of its protein composition and spongious infrastructure can provide inspiration for development of potential biomimetics that can attach completely different or incompatible surfaces with high efficiency and strength.

Cell culture of differentiated human salivary epithelial cells in a serum-free and scalable suspension system: The salivary functional units model.

Saliva aids in digestion, lubrication, and protection of the oral cavity against dental caries and oropharyngeal infections. Reduced salivary secretion, below an adequate level to sustain normal oral functions, is unfortunately experienced by head and neck cancer patients treated with radiotherapy and by patients with Sjögren's syndrome. No disease-modifying therapies exist to date to address salivary gland hypofunction (xerostomia, dry mouth) because pharmacotherapies are limited by the need for residual secretory acinar cells, which are lost at the time of diagnosis, whereas novel platforms such as cell therapies are yet immature for clinical applications. Autologous salivary gland primary cells have clinical utility as personalized cell therapies, if they could be cultured to a therapeutically useful mass while maintaining their in vivo phenotype. Here, we devised a serum-free scalable suspension culture system that grows partially digested human salivary tissue filtrates composing of acinar and ductal cells attached to their native extracellular matrix components while retaining their 3D in vivo spatial organization; we have coined these salivary spheroids as salivary functional units (SFU). The proposed SFU culture system was sub-optimal, but we have found that the cells could still survive and grow into larger salivary spheroids through cell proliferation and aggregation for 5 to 10 days within the oxygen diffusion rates in vitro. In summary, by using a less disruptive cell isolation procedure as the starting point for primary cell culture of human salivary epithelial cells, we demonstrated that aggregates of cells remained proliferative and continued to express acinar and ductal cell-specific markers.