Effects of Pilocarpine Hydrochloride on the Hyposalivation Induced by Psychotropic Drugs

The aim was to study the secretagogue action of pilocarpine on the murine parotid glands submitted to chronic treatment with psychotropic drugs by salivary flow rate determinations and histological alterations. Fifty four male Wistar rats were equally divided in three groups： C group （control） received saline solution for 30 days; AD group （n = 18） received AmitriptylineR and DiazepamR for 30 days, and ADP group （n = 18） received Amitriptyline R and DiazepamR for 30 days and AmitriptylineR, DiazepamR and pilocarpine for further 30 days, resulting in 60 days of treatment. Saliva samples were collected 30 h after the end of treatment. Parotids were removed and processed for hematoxylin-eosin histological analysis. Dedicated software for image processing allowed the determination of cell number and volume. Significant differences between paired-groups C-AD （P 〈 0.01） and AD-ADP （P 〈 0.01） were observed for glands size and weight. The volume of serous cells was greater in AD, suggesting a hypertrophy of the salivary glands. For salivary flow rate, C group showed the highest average. The number of serous cells was similar between groups ADP and C, with the lowest average being found in AD group （P 〈 0.05）.

Hyperglycemia Induced Apoptosis Changes in Salivary Gland Cells of Mice

If the function of salivary glands that secrete saliva is degraded, saliva secretion will decrease and symptoms of xerostomia will appear. Although symptoms of xerostomia are often felt by patients with diabetes, the relationship between diabetes and xerostomia is currently unclear. In relation to DM, there are studies on salivation flow and saliva composition, but there are not studies of apoptosis of salivary gland cells. The objective of this study was to investigate whether apoptosis in salivary glands of mice with hyperglycemic, a symptom of diabetes, might be altered based on immunohistochemical analysis. This study used mice with hyperglycemia. Immunohistochemistry and Western blot analyses were performed using Fas, Bax, and cleaved caspase-3 antibodies. These antibodies are used not only as death receptors, but also are antibodies that activate upstream and downstream signals of apoptosis. TUNEL assay was performed to detect apoptosis by immunofluorescence using TdT enzyme. It was observed that the expression level of apoptosis signaling molecules and TUNEL positive cells were increased in hyperglycemia group (HG). As a result, there are many apoptosis cells in the HG groups of the salivary gland. The results of this study, the function of salivary gland could occur deteriorated due to apoptosis on salivary gland cells by hyperglycemic, a characteristic of diabetes.

Magnetic bioassembly platforms towards the generation of extracellular vesicles from human salivary gland functional organoids for epithelial repair

Salivary glands(SG)are exocrine organs with secretory units commonly injured by radiotherapy.Bio-engineered organoids and extracellular vesicles(EV)are currently under investigation as potential strategies for SG repair.Herein,three-dimensional(3D)cultures of SG functional organoids(SGo)and human dental pulp stem cells(hDPSC)were generated by magnetic 3D bioassembly(M3DB)platforms.Fibroblast growth factor 10(FGF10)was used to enrich the SGo in secretory epithelial units.After 11 culture days via M3DB,SGo displayed SG-specific acinar epithelial units with functional properties upon neurostimulation.To consistently develop 3D hDPSC in vitro,3 culture days were sufficient to maintain hDPSC undifferentiated genotype and phenotype for EV generation.EV isolation was performed via sequential centrifugation of the conditioned media of hDPSC and SGo cultures.EV were characterized by nanoparticle tracking analysis,electron microscopy and immunoblotting.EV were in the exosome range for hDPSC(diameter:88.03±15.60 nm)and for SGo(123.15±63.06 nm).Upon ex vivo administration,exosomes derived from SGo significantly stimulated epithelial growth(up to 60%),mitosis,epithelial progenitors and neuronal growth in injured SG;however,such biological effects were less distinctive with the ones derived from hDPSC.Next,these exosome biological effects were investigated by proteomic arrays.Mass spectrometry profiling of SGo exosomes predicted that cellular growth,development and signaling was due to known and undocumented molecular targets downstream of FGF10.Semaphorins were identified as one of the novel targets requiring further investigations.Thus,M3DB platforms can generate exosomes with potential to ameliorate SG epithelial damage.

Aquaporin 5 is degraded by autophagy in diabetic submandibular gland

Autophagy is a catabolic process which is involved in the development of many diseases including diabetes mellitus and its complications. Hyposalivation is a common complication of diabetes mellitus, whereas its mechanism remains unclear. Here, we observed that the stimulated salivary flow rate of SMG was significantly decreased in db/db mice, a diabetic mice model. The expressions of aquaporin 5(AQP5), a water channel protein, were decreased, whereas the m RNA level of AQP5 was increased in SMGs of both diabetic patients and mice. Under transmission electron microcope, more autophagosomes were detected in diabetic SMGs. Expressions of autophagy related proteins LC3 II, Beclin-1 and ATG5 were increased, meanwhile autophagy substrate p62 was decreased in SMGs of diabetic patients and mice, indicating that autophagy was activated in diabetic SMG.Double immunofluorescence staining showed that the colocalization of AQP5 and LC3 was increased in SMGs of diabetic mice.In cultured SMG-C6 cells, high glucose(HG), but not high osmotic pressure, reduced AQP5 protein expression and induced autophagy. Moreover, inhibition of autophagy by 3-methyladenin, an autophagy inhibitor, or by autophagy-related gene 5 siRNA, decreased HG-induced AQP5 reduction in SMG-C6 cells. Additionally, the expression of p-p85, p-Akt and p-mTOR were decreased in HG-treated SMG-C6 cells. Pretreatment with 740 Y-P, a PI3 K agonist, significantly suppressed HG-induced autophagy and AQP5 degradation. Taken together, these results indicate that autophagy plays a crucial role in AQP5 degradation in diabetic SMG via PI3 K/Akt/mTOR signaling pathway, which contributes to the dysfunction of diabetic SMG. Our study provides a novel mechanism of diabetic hyposalivation.