Expression of BAFF and BAFF receptors in primary Sjögren's syndrome patients with ectopic germinal center-like structures.

B cell-activating factor (BAFF) is an essential cytokine in primary Sjögren's syndrome (pSS) physiopathology. It has been reported that pSS patients develop germinal center-like (GC-like) structures in their minor salivary glands (MSGs). BAFF, BAFF-R, TACI, and BCMA expression was analyzed in MSGs from 29 subjects (nonspecific chronic sialadenitis and focal lymphocytic sialadenitis with the presence [pSS-GC(+)] or absence [pSS-GC(-)] of GC-like structures). Twenty-four percent of patients showed ectopic GC-like structures and a high focus score [p < 0.001 vs pSS-GC(-)]. BAFF serum levels (sBAFF) were high in pSS patients (p = 0.025 vs healthy subjects). However, the pSS-GC(-) group showed higher sBAFF levels than pSS-GC(+) patients. BAFF and BAFF-R glandular expression levels were higher in pSS-GC(+) patients, without significant differences compared to pSS-GC(-) patients. Soluble levels of BAFF correlated with anti-La/SSB antibodies and disease duration. Our results showed that BAFF could contribute to focal lymphocytic infiltration. The role of BAFF-binding receptors in MSGs is proposed as a mechanism for the possible establishment of ectopic GC-like structures and disease progression in some patients. In conclusion, this study supports previous evidence that considers the active BAFF system role in the pathogenesis of pSS and the need for strong biomarkers in this disease.

Spatial organization of the excretory ducts and sections of microcirculatory blood flow of the labial salivary glands in older adults.

OBJECTIVE: Introduction: Salivary glands have a significant impact on the state of the homeostasis of the human body, oral cavity in particular, sensitively responding to pathological processes. The reactivity of the salivary glands in response to pathological processes that are organically linked to morphology and functions of the organ's structures, and particularly the excretory ducts of the glands and their microcirculatory blood flow, is one of the problems which have not been solved to date. The aim of the paper was to elucidate the features of the stereomicroscopic structure of the excretory ducts and sectors of the microcirculatory blood flow in labial glands of older adults. PATIENTS AND METHODS: Materials and methods: The object of the study was the labial mucosa of the older adults, which was cut into 3х3 mm pieces and fixed in the buffered 4% glutaraldehyde solution with subsequent 2-hour fixation in osmium. Once the pieces were washed and dehydrated they were embedded into the Epon-812. The series of the semi-thin sections, made from the obtained epoxy blocks, were stained in phosphate buffered 0,1% toluidine blue solution. The serial semi-thin sections were subjected to histological and cytological studies and multilayered plastic reconstruction. RESULTS: Results and conclusions: The series of histological epoxy semi-thin sections, as well as graphic and plastic reconstruction of the sectors of microcirculatory blood flow and excretory ducts of the human labial glands have demonstrated a range of morphological facts that can be used to clarify the intertissue stereological relationships. They also determined the syntopic proximity of the capacitive sectors of microcirculatory blood flow to the excretory ducts of the gland. Such pattern is especially notable in the collecting venules and intralobular ducts. It has been shown that the biggest venule is the collecting venous vessel. Anastomoses between the intralobular arterioles and collecting venules have been found in the microcirculatory blood flow of the labial glands.

Effects and Mechanisms of Tastants on the Gustatory-Salivary Reflex in Human Minor Salivary Glands.

The effects and mechanisms of tastes on labial minor salivary gland (LMSG) secretion were investigated in 59 healthy individuals. Stimulation with each of the five basic tastes (i.e., sweet, salty, sour, bitter, and umami) onto the tongue induced LMSG secretion in a dose-dependent manner. Umami and sour tastes evoked greater secretion than did the other tastes. A synergistic effect of umami on LMSG secretion was recognized: a much greater increase in secretion was observed by a mixed solution of monosodium glutamate and inosine 5'-monophosphate than by each separate stimulation. Blood flow (BF) in the nearby labial mucosa also increased following stimulation by each taste except bitter. The BF change and LMSG secretion in each participant showed a significant positive correlation with all tastes, including bitter. Administration of cevimeline hydrochloride hydrate to the labial mucosa evoked a significant increase in both LMSG secretion and BF, while adrenaline, atropine, and pirenzepine decreased LMSG secretion and BF. The change in LMSG secretion and BF induced by each autonomic agent was significantly correlated in each participant. These results indicate that basic tastes can induce the gustatory-salivary reflex in human LMSGs and that parasympathetic regulation is involved in this mechanism.

Minor Salivary Gland Transplantation for Severe Dry Eyes.

Dry eye is a multifactorial disease comprising a wide spectrum of ocular surface alterations and symptoms of discomfort. In most patients with aqueous-deficient dry eye, pharmaceutical tear substitutes are used to control symptoms and prevent ocular surface damage. However, in severe dry eye conditions caused by cicatricial disorders, such as Stevens-Johnson syndrome and ocular cicatricial mucous membrane pemphigoid, noninvasive treatments are insufficient, and patients are at risk of developing complications that can lead to blindness. The use of salivary glands as a source of lubrication to treat severe cases of dry eye has been proposed by different authors. The first reports proposed parotid or submandibular gland duct transplantation into the conjunctival fornix. However, complications limited the functional outcomes. Minor salivary gland autotransplantation together with labial mucosa has been used as a complex graft to the conjunctival fornix in severe dry eye with a good outcome. Our group demonstrated significant improvements in best-corrected visual acuity, Schirmer I test score, corneal transparency, and neovascularization after using this technique. A symptoms questionnaire applied to these patients revealed improvements in foreign body sensation, photophobia, and pain. Similar to tears, saliva has a complex final composition comprising electrolytes, immunoglobulins, proteins, enzymes, and mucins. We demonstrated the viability of minor salivary glands transplanted into the fornix of patients with dry eye by performing immunohistochemistry on graft biopsies with antibodies against lactoferrin, lysozyme, MUC1, and MUC16. The findings revealed the presence of functional salivary gland units, indicating local production of proteins, enzymes, and mucins.

Role of minor salivary glands in developing patient-rated xerostomia and sticky saliva during day and night.

PURPOSE: The purpose of this prospective study was to investigate the relationship between xerostomia during the day (XERday) and night (XERnight) and sticky saliva during the day (STICday) and night (STICnight) and dose distributions in different major and minor salivary glands among head and neck cancer (HNC) patients treated with primary radiotherapy (RT) or chemoradiation (CHRT). METHODS AND MATERIALS: The study population was composed of 201 consecutive HNC patients treated with intensity modulated radiotherapy (IMRT) or 3-dimensional conformal radiotherapy (3D-CRT). All patients were included in a standard follow up programme in which acute and late side effects and quality of life (QoL) were prospectively assessed, prior to, during and after treatment. The primary endpoints were XERday, XERnight, STICday, STICnight as assessed by the Groningen Radiotherapy Induced Xerostomia questionnaire (GRIX) six months after completion of treatment. Organs at risk (OARs) potentially involved in salivary function were delineated on planning-CT, including the parotid, submandibular and sublingual glands and the minor glands in the soft palate, buccal mucosa and lips. Patients with moderate-to-severe xerostomia or moderate-to-severe sticky saliva, respectively, at baseline were excluded. In order to determine which salivary glands were most important, a multivariate logistic regression analysis with an extended bootstrapping technique was used. RESULTS: In total, 29% and 19% of the cases suffered from XERday and XERnight, respectively. The multivariate analysis showed that baseline xerostomia and the mean parotid gland dose were the most important predictors for XERday and XERnight. At 6months after (CH)RT, 10% and 12% of the cases reported STICday and STICnight respectively. We were not able to identify prognostic factors related to dose distributions with regard to STICday. The mean submandibular gland dose was associated with STICnight. Baseline xerostomia and sticky saliva scores on the GRIX were associated with XERday, XERnight, STICday. Increasing age was correlated with both XERnight and STICnight. CONCLUSION: Organs at risk for XERday and STICday are similar to organs at risk for XERnight and STICnight.

Fosmid-based structure-function analysis reveals functionally distinct domains in the cytoplasmic domain of Drosophila crumbs.

The evolutionarily conserved transmembrane protein Crumbs is required for epithelial polarity and morphogenesis in the embryo, control of tissue size in imaginal discs and morphogenesis of photoreceptor cells, and prevents light-dependent retinal degeneration. The small cytoplasmic domain contains two highly conserved regions, a FERM (i.e., protein 4.1/ezrin/radixin/moesin)-binding and a PDZ (i.e., postsynaptic density/discs large/ZO-1)-binding domain. Using a fosmid-based transgenomic approach, we analyzed the role of the two domains during invagination of the tracheae and the salivary glands in the Drosophila embryo. We provide data to show that the PDZ-binding domain is essential for the maintenance of cell polarity in both tissues. In contrast, in embryos expressing a Crumbs protein with an exchange of a conserved Tyrosine residue in the FERM-binding domain to an Alanine, both tissues are internalized, despite some initial defects in apical constriction, phospho-Moesin recruitment, and coordinated invagination movements. However, at later stages these embryos fail to undergo dorsal closure, germ band retraction, and head involution. In addition, frequent defects in tracheal fusion were observed. These results suggest stage and/or tissue specific binding partners. We discuss the power of this fosmid-based system for detailed structure-function analyses in comparison to the UAS/Gal4 system.

Abnormal basement membrane type IV collagen alpha-chain composition in labial salivary glands in Sjögren's syndrome.

OBJECTIVE: Sjögren's syndrome (SS) is characterized by atrophy and malfunction of the acinar cells. The aim of this study was to investigate whether type IV collagen alpha-chain composition of acinar cell compartments could be abnormal in diseased glands. METHODS: Messenger RNA (mRNA) from human submandibular gland (HSG) cells, cultured with or without growth factor-depleted Matrigel, was analyzed using quantitative reverse transcription-polymerase chain reaction (RT-PCR). Labial salivary glands were analyzed using quantitative RT-PCR and immunohistochemistry. RESULTS: HSG cells of both the ductal and acinar phenotypes synthesized all alpha-chain mRNA, in particular those of the alpha1 and alpha2 chains. Labial salivary glands (LSGs) contained alpha1/2 chains but also contained mRNA of all the other alpha-chains, although the mRNA copy numbers for the alpha3 and alpha4 chains were low, and the corresponding proteins were absent. Type IV collagen alpha1/2-chains were observed in all tubuloalveolar basement membranes. In healthy glands, alpha5 and alpha6 chains were continuous around ducts but discontinuous around acini. In SS glands, these chains were absent or patchy around the ducts and absent around the acini. CONCLUSION: Ductal and acinar epithelial cells are able to locally produce mRNA for all 6 different alpha-chains. Type IV collagen alpha1/2-chains seem to form the backbone in the tubuloalveolar basement membrane in salivary glands. Type IV collagen alpha3 and alpha4 chain mRNA were found in cultured salivary epithelial cells and LSG explants but were not translated to the corresponding alpha-chains in LSGs. Both alpha5 and alpha6 mRNA were observed in salivary epithelial cells and glands. In healthy glands, immunolabeling always disclosed corresponding alpha-chains around ducts, but their synthesis and/or degradation seemed to be locally regulated around acinar cells.

Acinar epithelial cell laminin-receptors in labial salivary glands in Sjögren's syndrome.

OBJECTIVE: To analyze the epithelial cell-basement membrane attachment, in particular in the secretory end pieces (responsible for secretion of saliva) and in Sjögren's syndrome (SS) characterized by acinar cell failure. METHOD: Immunohistochemistry with laminin receptor chain-specific monoclonal antibodies to integrin (Int) subunits, Lutheran blood group antigen and alpha-dystroglycan. RESULTS: Only acinar cells contained Int alpha1 and alpha2 subunits. This staining was interrupted but strong in controls, but very weak in SS. Both acinar and ductal cells contained Int alpha3, alpha6, b1 and b4 and Lutheran blood group antigen and ductal cells also contained alpha-dystroglycan. These staining patterns were similar in SS and controls. CONCLUSIONS: Binding of the acinar and ductal cells to the basement membrane laminins seems to be mediated by Int alpha3b1, alpha6b1 and alpha6b4 integrin-receptors and Lutheran blood group antigen and alpha-dystroglycan non-integrin receptors. This structure-supporting system is intact in SS, compatible with the maintenance of the tubuloalveolar architecture of the SS glands. The irregular staining pattern of the acinus-specific Int alpha1b1 and alpha2b1 was compatible with a regulated signaling role, which was apparently impaired in SS. Indeed, their laminin counterparts (Lm -1/111 and -2/211) are also aberrant in SS revealing this as the central cell-matrix defect in the syndrome.

Minor salivary glands morphology in xerostomia patients / Morfología de las Glándulas Salivales Menores en Pacientes Xerostomizados

Las glándulas salivales menores son encontradas distribuidas a través de la mucosa oral, especialmente en los labios y en la mucosa del paladar blando. Varios factores pueden causar xerostomía, donde las características histológicas de las glándulas salivales son también consideradas como factores para definir la etiología. Así, las biopsias de las glándulas salivales menores representan una herramienta fundamental para alcanzar los criterios diagnósticos requeridos en la clasificación de pacientes con síndrome de Sjõgren, ya que no representa riesgo para los pacientes. El objetivo de este estudio es determinar las características histológicas de las glándulas mencionadas, obtenidas de biopsias de pacientes con xerostomía y clasificar los aspectos histológicos de las glándulas en el síndrome de Sjõgren. Estudiamos 40 láminas de pacientes con xerostomía, cuyas glándulas salivales menores fueron sometidas a biopsia en el Servicio de Estomatología de la Santa Casa de São Paulo, Brasil. Se observaron las variaciones de su aspecto histológico, desde la normalidad hasta la presencia de focos inflamatorios, los cambios del tejido conjuntivo entre los acinos y conductos, como también el parénquima. En 15 casos, el infltrado de células inflamatorias invadió el foco, es decir, grupos de al menos 50 células inflamatorias alrededor de acinos o conductos, lo cual es un aspecto característico del síndrome de Sjõgren. Por lo tanto, el hallazgo de al menos un foco inflamatorio de 4 mm2 de tejido glandular, representa un buen criterio, aunque no es uno de los criterios a considerar cuando se trata de clasificar a los pacientes con el Síndrome de Sjõgren.

The minor salivary glands are found scattered throughout the oral mucosa, especially in the lips and soft palate mucosa. Several factors can cause xerostomia, whereas the salivary glands histological characteristics are also considered as factors for defining the etiology. Thus, the minor salivary glands biopsy represents an essential tool for attending the required diagnosis criterion in the classification of Sjögren's Syndrome patients, since it does not present risk for the patient. The objective of this study is to determine the histological description of the minor salivary glands obtained from the biopsies of xerostomia patients and to classify the minor salivary gland histological aspect as the Sjögren's Syndrome. Forty laminas of xerostomia patients that were submitted to minor salivary glands biopsy at the Santa Casa de Sao Paulo Stomatology ambulatory were retrospectively studied. The variation in the glands histological aspect was observed, from the normality up to the presence of inflammatory focus, replacing the conjunctive between acini and ducts, as well as the parenchyma. In 15 cases, the infiltrated inflammatory cells amounted to focus, that is to say, groups of at least 50 inflammatory cells around the acini or ducts, which is a characteristic aspect of the Sjögren's Syndrome. Therefore, the finding of at least one inflammatory focus of 4 mm2 of glandular tissue represents a set criterion, although, not the only one in order to classify this patient as having the Sjögren's Syndrome.

A method of measuring salivary flow rate in the lower labial mucosal region.

To measure resting whole saliva, draining and spitting methods have been commonly used. However, these methods require the patients' understanding and cooperation. The aim of this study was to establish a method for measuring the salivary flow rate that does not rely on patient's cooperation or the state of the mouth and does not cause any discomfort to patients when performing the measurements. The salivary flow rate in the lower labial mucosal region was measured by means of strips that incorporated the iodo-starch reaction. The salivary flow rate was then calculated on the basis of the number of spots and area of discoloration on the strip. In order to test the validity of these methods, the correlation between these results and resting whole saliva measured by the draining method was also investigated. A positive correlation was found between the salivary flow rate estimated from the area of discoloration on the strip and the resting whole saliva (r=0.678, P=0.01). Therefore, these findings indicate that this is a valid method of measuring the salivary flow rate in the lower labial mucosal region.

The superficial mandibular gland of the rabbit: a new experimental model for scintigraphic evaluation of salivary glands.

The aim of this study was to provide an appropriate experimental model to study functional changes in salivary glands using scintigraphy. Although the rabbit was frequently used for laboratory experiments, there are only a few studies that describe the exact position of its different salivary glands on the sialoscintigram. Twenty rabbits were used for the study; ten of them were anatomically dissected to provide the required topographic anatomy on the different salivary and lacrimal glands. The remaining ten animals underwent a static scintigraphy after extirpation of a particular salivary gland. Changes in the pattern of tracer uptake indicated the exact position of every gland allowing its evaluation. The results show that the 99mTcO4- uptake can only be selectively evaluated in two salivary glands, the superficial mandibular gland and the parotid gland. The superficial mandibular gland was proved to be a particularly useful model to evaluate functional changes of salivary gland parenchyma due to its well defined and high enhanced structure which allows a precise detection and measurement of the tracer uptake. Additionally, the good surgical accessibility of this gland and the existence of a well defined capsule facilitate associated histological studies of its parenchyma.

Minor salivary gland duct-associated lymphoid tissue (DALT) in monkeys changes with age.

The duct-associated lymphoid tissue (DALT) of minor salivary glands (MSG) is accessible to oral antigens by retrograde passage. DALT responds immunologically to such purely local and duct-derived antigenic stimuli. This study addressed the question as to whether DALT, like other mammalian lymphoid tissues, would change with age. Labial and buccal mucosae of eight Macaca fascicularis animals of known age were processed for light microscopy by Epon embedding. Sections of approximately 1-2 microns thick were prepared, and a total of 144, more or less equidistant, labial and 63 buccal sections were selected and analyzed for various tissue components, by means of morphometric point-counting. The raw data were transformed into stereological parameters. The results showed that DALT and interacinar plasma cells are totally absent in the newborn monkey. They begin to appear early in life and reach a maximum volume density by one year of age. With further increasing age, the DALT volume showed a sharp decline, followed by a plateauing from year 3 onward, while the plasma cell concentration remained stable. This change was more pronounced in the labial mucosa. These observations on the age-related changes of simian DALT are suggestive of its antigen-induced and antigen-dependent nature. DALT, being part of the gut-associated lymphoid tissue, may play a substantial role in the local immuno-surveillance of the oral cavity.