Search

Saliva plays a major role in maintaining oral health. Patients with salivary hypofunction exhibit difficulty in chewing and swallowing foods, tooth decay, periodontal disease, and microbial infections. At this time, treatments for hyposalivation are limited to medications (e.g., muscarinic receptor agonists: pilocarpine and cevimeline) that induce saliva secretion from residual acinar cells as well as artificial salivary substitutes. Therefore, advancement of restorative treatments is necessary to improve the quality of life in these patients. Our previous studies indicated that salivary cells are able to form polarized 3-dimensional structures when grown on growth factor-reduced Matrigel. This basement membrane is rich in laminin-III (L1), which plays a critical role in salivary gland formation. Mitotically inactive feeder layers have been used previously to support the growth of many different cell types, as they provide factors necessary for cell growth and organization. The goal of this study was to improve salivary gland cell differentiation in primary cultures by using a combination of L1 and a feeder layer of human hair follicle-derived mesenchymal stem cells (hHF-MSCs). Our results indicated that the direct contact of mouse submandibular (mSMG) cell clusters and hHF-MSCs was not required for mSMG cells to form acinar and ductal structures. However, the hHF-MSC conditioned medium enhanced cell organization and multilumen formation, indicating that soluble signals secreted by hHF-MSCs play a role in promoting these features., (© International & American Associations for Dental Research 2015.)

In humans, approximately 90% of saliva is secreted by the 3 major salivary glands: the parotid (PG), the submandibular (SMG), and the sublingual glands (SLG). Even though it is known that all 3 major salivary glands secrete saliva by a Cl(-)-dependent mechanism, salivary secretion rates differ greatly among these glands. The goal of this study was to gain insight into the properties of the ion-transporting pathways in acinar cells that might account for the differences among the major salivary glands. Pilocarpine-induced saliva was simultaneously collected in vivo from the 3 major salivary glands of mice. When normalized by gland weight, the amount of saliva secreted by the PG was more than 2-fold larger than that obtained from the SMG and SLG. At the cellular level, carbachol induced an increase in the intracellular [Ca(2+)] that was more than 2-fold larger in PG and SMG than in SLG acinar cells. Carbachol-stimulated Cl(-) efflux and the protein levels of the Ca(2+)-activated Cl(-) channel TMEM16A, the major apical Cl(-) efflux pathway in salivary acinar cells, were significantly greater in PG compared with SMG and SLG. In addition, we evaluated the transporter activity of the Na(+)-K(+)-2Cl(-) cotransporters (NKCC1) and anion exchangers (AE), the 2 primary basolateral Cl(-) uptake mechanisms in acinar cells. The SMG NKCC1 activity was about twice that of the PG and more than 12-fold greater than that of the SLG. AE activity was similar in PG and SLG, and both PG and SLG AE activity was about 2-fold larger than that of SMG. In summary, the salivation kinetics of the 3 major glands are distinct, and these differences can be explained by the unique functional properties of each gland related to Cl(-) movement, including the transporter activities of the Cl(-) uptake and efflux pathways, and intracellular Ca(2+) mobilization., (© International & American Associations for Dental Research 2015.)

NFIB (nuclear factor I B) is a NFI transcription factor family member, which is essential for the development of a variety of organ systems. Salivary gland development occurs through several stages, including prebud, bud, pseudoglandular, canalicular, and terminal. Although many studies have been done to understand mouse submandibular gland (SMG) branching morphogenesis, little is known about SMG cell differentiation during the terminal stages. The goal of this study was to determine the role of NFIB during SMG development. We analyzed SMGs from wild-type and Nfib-deficient mice (Nfib (-/-)). At embryonic (E) day 18.5, SMGs from wild-type mice showed duct branching morphogenesis and differentiation of tubule ductal cells into tubule secretory cells. In contrast, SMGs from Nfib (-/-) mice at E18.5 failed to differentiate into tubule secretory cells while branching morphogenesis was unaffected. SMGs from wild-type mice at E16.5 displayed well-organized cuboidal inner terminal tubule cells. However, SMGs from Nfib (-/-) at E16.5 displayed disorganized inner terminal tubule cells. SMGs from wild-type mice at E18.5 became fully differentiated, as indicated by a high degree of apicobasal polarization (i.e., presence of apical ZO-1 and basolateral E-cadherin) and columnar shape. Furthermore, SMGs from wild-type mice at E18.5 expressed the protein SMGC, a marker for tubule secretory cells. However, SMGs from Nfib (-/-) mice at E18.5 showed apicobasal polarity, but they were disorganized and lost the ability to secrete SMGC. These findings indicate that the transcription factor NFIB is not required for branching morphogenesis but plays a key role in tubule cell differentiation during mouse SMG development., (© International & American Associations for Dental Research 2014.)

Salivary gland epithelial cells (SGEC) release several cytokines that play important roles in the inflammatory process. In this study, we examined whether capsaicin can modulate cytokine release in SGEC. After cells were stimulated with polyinosinic-polycytidylic acid [poly(I:C)] or lipopolysaccharide (LPS), mRNA transcript and protein levels were detected by reverse-transcriptase-polymerase chain-reaction (RT-PCR), real-time PCR, and enzyme-linked immunosorbent assay (ELISA). These findings demonstrated that the increases in TNFα and IL-6 mRNA transcripts were highest at 3 hrs and 1 hr after incubation with poly(I:C) and LPS, respectively. Pre-treatment of the cells with 10 μµ capsaicin, however, significantly inhibited mRNA transcripts and its protein levels. The simultaneous application of 10 μµ capsazepine with capsaicin did not block the inhibitory effect of capsaicin. Furthermore, the inhibitory effect of capsaicin was also shown in primary cultured cells from TRPV1(-/-) mice. We found that both poly(I:C) and LPS induced IκB-α degradation and phosphorylation, which resulted in NF-κB activation, and capsaicin inhibited this NF-κB pathway. These results demonstrate that SGEC release pro-inflammatory cytokines mediated by TLR, and capsaicin inhibits this process through the NF-κB pathway. This study suggests that capsaicin could potentially alleviate inflammation in salivary glands.

Vanilloid receptor-1 (VR1) was originally found in the nervous system. Recent evidence indicates that VR1 is also expressed in various cell types. We hypothesized that VR1 exists in the human submandibular gland (SMG) and is involved in regulating salivary secretion. VR1 mRNA and protein were expressed in human SMGs and a human salivary intercalated duct cell line. VR1 was mainly located in serous acinar and ductal cells, but not in mucous acinar cells. Capsaicin, an agonist of VR1, increased intracellular free calcium, enhanced phosphorylation of extracellular signal-regulated kinase, and induced the trafficking of aquaporin 5 (AQP5) from the cytoplasm to the plasma membrane. These effects were abolished by pre-treatment with the VR1 antagonist capsazepine. Furthermore, capsaicin cream applied to the skin covering the submandibular area increased salivary secretion. These findings indicated that a functional VR1 is expressed in the human SMG and is involved in regulating salivary secretion by mediating AQP5 trafficking.

Three members of the SIBLING family of integrin-binding phosphoglycoproteins (bone sialoprotein, BSP; osteopontin, OPN; and dentin matrix protein-1, DMP1) were recently shown to bind with high affinity (nM) and to activate 3 different matrix metalloproteinases (MMP-2, MMP-3, and MMP-9, respectively) in vitro. The current study was designed to document the possible biological relevance of the SIBLING-MMP activation pathway in vivo by showing that these 3 SIBLINGs and their known MMP partners are co-expressed in normal adult tissue. BSP, OPN, and DMP1 were invariably co-expressed with their partner MMPs in salivary glands of humans and mice. The 2 SIBLING proteins without known MMP partners, dentin sialophosphoprotein (DSPP) and matrix extracellular phosphoglycoprotein (MEPE), were also expressed in salivary glands. Expression of all SIBLINGs in this normal, non-mineralizing epithelial tissue suggests that they serve at least one function in vivo other than directly promoting matrix mineralization--a function we hypothesize involves local activation of MMPs.

Studies of Drosophila salivary gland formation have elucidated the regulatory pathway by which the salivary gland fate is determined and the morphogenetic processes by which the primordial cells are internalized to form the tubular glands. Both the position of the salivary primordia and the number of cells recruited to a salivary gland fate are established through a combination of the localized expression of the transcription factors SEX COMBS REDUCED (SCR), TEASHIRT (TSH) and ABDOMINAL-B (ABD-B), and localized DPP-signaling. Similarly, the distinction between the two major cell types, duct and secretory, is determined by spatially limited EGF-signaling. Salivary gland formation also requires the function of two transcription factors expressed in nearly all cells of the developing embryo, EXTRADENTICLE (EXD) and HOMOTHORAX (HTH). Once the salivary gland fate is determined, cells of the secretory primordia are internalized by an apical constriction mode of invagination. We have characterized three genes encoding transcription factors, trachealess (trh), hückebein (hkb), and fork head (fkh), that are downstream targets of the salivary gland regulators. Mutations in these transcription factors profoundly affect salivary gland morphogenesis. trh is required for the formation of the salivary duct tubes. hkb determines the order of secretory cell invagination, a regulated process critical for determining the final shape of the salivary gland. fkh has two early roles in salivary gland formation. fkh both promotes secretory cell survival and facilitates secretory cell internalization. trh, hkb, and fkh are involved in the formation of not only the salivary duct and secretory tubes, but also of other tubular structures, such as the trachea and the gut endoderm. We propose that trh, hkb, and fkh may serve as "morphogenetic cassettes" responsible for forming tubular structures in a variety of tissues.