Non-Lethal Concentration of MeHg Causes Marked Responses in the DNA Repair, Integrity, and Replication Pathways in the Exposed Human Salivary Gland Cell Line

In Brazilian northern Amazon, communities are potentially exposed and vulnerable to methylmercury (MeHg) toxicity through the vast ingestion of fish. In vivo and in vitro studies demonstrated that the salivary glands as a susceptible organ to this potent environmental pollutant, reporting alterations on physiological, biochemical, and proteomic parameters. However, the alterations caused by MeHg on the gene expression of the exposed human salivary gland cells are still unknown. Therefore, the goal was to perform the transcriptome profile of the human salivary gland cell line after exposure to MeHg, using the microarray technique and posterior bioinformatics analysis. The cell exposure was performed using 2.5 µM MeHg. A previously published study demonstrated that this concentration belongs to a range of concentrations that caused biochemical and metabolic alterations in this linage. As a result, the MeHg exposure did not cause lethality in the human salivary gland cells line but was able to alter the expression of 155 genes. Downregulated genes (15) are entirety relating to the cell metabolism impairment, and according to KEGG analysis, they belong to the glycosphingolipid (GSL) biosynthesis pathway. On the other hand, most of the 140 upregulated genes were related to cell-cycle progression, DNA repair, and replication pathway, or cellular defenses through the GSH basal metabolism. These genomic changes revealed the effort to the cell to maintain physiological and genomic stability to avoid cell death, being in accordance with the nonlethality in the toxicity test. Last, the results support in-depth studies on nonlethal MeHg concentrations for biomarkers identification that interpret transcriptomics data in toxicological tests serving as an early alert of physiological changes in vitro biological models.

Membrane association of importin α facilitates viral entry into salivary gland cells of vector insects

The importin α family belongs to the conserved nuclear transport pathway in eukaryotes. However, the biological functions of importin α in the plasma membrane are still elusive. Here, we report that importin α, as a plasma membrane–associated protein, is exploited by the rice stripe virus (RSV) to enter vector insect cells, especially salivary gland cells. When the expression of three importin α genes was simultaneously knocked down, few virions entered the salivary glands of the small brown planthopper, Laodelphax striatellus. Through hemocoel inoculation of virions, only importin α2 was found to efficiently regulate viral entry into insect salivary-gland cells. Importin α2 bound the nucleocapsid protein of RSV with a relatively high affinity through its importin β–binding (IBB) domain, with a dissociation constant KD of 9.1 μM. Furthermore, importin α2 and its IBB domain showed a distinct distribution in the plasma membrane through binding to heparin in heparan sulfate proteoglycan. When the expression of importin α2 was knocked down in viruliferous planthoppers or in nonviruliferous planthoppers before they acquired virions, the viral transmission efficiency of the vector insects in terms of the viral amount and disease incidence in rice was dramatically decreased. These findings not only reveal the specific function of the importin α family in the plasma membrane utilized by viruses, but also provide a promising target gene in vector insects for manipulation to efficiently control outbreaks of rice stripe disease.

Interactions between the flavescence dorée phytoplasma and its insect vector indicate lectin-type adhesion mediated by the adhesin VmpA

The flavescence dorée phytoplasma undergoes a propagative cycle in its insect vectors by first interacting with the insect cell surfaces, primarily in the midgut lumen and subsequently in the salivary glands. Adhesion of flavescence dorée phytoplasma to insect cells is mediated by the adhesin VmpA. We hypothesize that VmpA may have lectin-like activity, similar to several adhesins of bacteria that invade the insect gut. We first demonstrated that the luminal surface of the midgut and the basal surface of the salivary gland cells of the natural vector Scaphoideus titanus and those of the experimental vector Euscelidius variegatus were differentially glycosylated. Using ELISA, inhibition and competitive adhesion assays, and protein overlay assays in the Euva-6 insect cell line, we showed that the protein VmpA binds insect proteins in a lectin-like manner. In conclusion, the results of this study indicate that N-acetylglucosamine and mannose present on the surfaces of the midgut and salivary glands serve as recognition sites for the phytoplasma adhesin VmpA.

Qualitative ultrastructural analysis of the submandibular salivary glands after administration of khat: in vivo study

Objective Khat (Catha edulis Forssk) plant has been widely chewed for its psychostimulatory effects in the African and Arabian Peninsula, particularly in Yemen. Considering the khat leaves are gradually chewed without swallowing, while its active constituents are extracted into saliva, studying the effect of khat on salivary glands is necessary. This work is an extension of the previously published work that studied the effect of khat extract on the rats' submandibular salivary glands in terms of histological and immunohistochemical evaluations. The current research note aimed to better understand this effect on the ultrastructure of submandibular salivary gland cells by using transmission electron microscope. Results Oral administration of khat extract produced degenerative changes in the secretory and ductal cells of rats' submandibular salivary glands. These changes involved irregular boundaries of variable sized-nuclei, dilated RER, cytoplasmic vacuoles as well as swollen and degenerated mitochondria.

Muscarinic Receptors and BK Channels Are Affected by Lipid Raft Disruption of Salivary Gland Cells

Activity-dependent fluid secretion is the most important physiological function of salivary glands and is regulated via muscarinic receptor signaling. Lipid rafts are important for G-protein coupled receptor (GPCR) signaling and ion channels in plasma membranes. However, it is not well understood whether lipid raft disruption affects all membrane events or only specific functions in muscarinic receptor-mediated water secretion in salivary gland cells. We investigated the effects of lipid raft disruption on the major membrane events of muscarinic transcellular water movement in human salivary gland (HSG) cells. We found that incubation with methyl-β-cyclodextrin (MβCD), which depletes lipid rafts, inhibited muscarinic receptor-mediated Ca2+ signaling in HSG cells and isolated mouse submandibular acinar cells. However, MβCD did not inhibit a Ca2+ increase induced by thapsigargin, which activates store-operated Ca2+ entry (SOCE). Interestingly, MβCD increased the activity of the large-conductance Ca2+-activated K+ channel (BK channel). Finally, we found that MβCD did not directly affect the translocation of aquaporin-5 (AQP5) into the plasma membrane. Our results suggest that lipid rafts maintain muscarinic Ca2+ signaling at the receptor level without directly affecting the activation of SOCE induced by intracellular Ca2+ pool depletion or the translocation of AQP5 into the plasma membrane.

Frequent and Persistent Salivary Gland Ectasia and Oral Disease After COVID-19

The clinical picture of coronavirus disease 2019 (COVID-19) in various target organs has been extensively studied and described. However, relatively little is known about the characteristics of oral cavity involvement. This is surprising, considering that oral mucosal and salivary gland cells are known targets for the direct replication of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and that the presence of the virus in saliva is a source of transmission of the infection. The aim of our study was to investigate the presence and prevalence of oral manifestations in COVID-19 survivors. We profiled the oral involvement in 122 COVID-19 survivors that were hospitalized and followed up at a single-referral university hospital in Milan, Italy, between July 23, 2020 and September 7, 2020, after a median (interquartile range) time from hospital discharge of 104 (95 to 132) d. We found that oral manifestations, specifically salivary gland ectasia, were unexpectedly common, with oral manifestations being detected in 83.9% while salivary gland ectasia in 43% of COVID-19 survivors. Salivary gland ectasia reflected the hyperinflammatory response to SARS-CoV-2, as demonstrated by the significant relationship with C-reactive protein (CRP) and lactate dehydrogenase (LDH) levels at hospital admission, and with the use of antibiotics during acute disease. Both LDH levels and antibiotic administration survived as independent predictors of salivary gland ectasia at multivariable analysis. Temporomandibular joint abnormalities, facial pain, and masticatory muscle weakness were also common. Overall, this retrospective and prospective cohort study of COVID-19 survivors revealed that residual damage of the oral cavity persists in the vast majority of patients far beyond clinical recovery, and suggests that the oral cavity represents a preferential target for SARS-CoV-2 infection. Further studies are needed to clarify the connection between SARS-CoV-2 infection and oral disorders.

Viral Release Threshold in the Salivary Gland of Leafhopper Vector Mediates the Intermittent Transmission of Rice Dwarf Virus

Numerous piercing-sucking insects can persistently transmit viral pathogens in combination with saliva to plant phloem in an intermittent pattern. Insect vectors maintain viruliferous for life. However, the reason why insect vectors discontinuously transmit the virus remains unclear. Rice dwarf virus (RDV), a plant reovirus, was found to replicate and assemble the progeny virions in salivary gland cells of the leafhopper vector. We observed that the RDV virions moved into saliva-stored cavities in the salivary glands of leafhopper vectors via an exocytosis-like mechanism, facilitating the viral horizontal transmission to plant hosts during the feeding of leafhoppers. Interestingly, the levels of viral accumulation in the salivary glands of leafhoppers during the transmitting period were significantly lower than those of viruliferous individuals during the intermittent period. A putative viral release threshold, which was close to 1.79 × 104 copies/μg RNA was proposed from the viral titers in the salivary glands of 52 leafhoppers during the intermittent period. Thus, the viral release threshold was hypothesized to mediate the intermittent release of RDV from the salivary gland cells of leafhoppers. We anticipate that viral release threshold-mediated intermittent transmission by insect vectors is the conserved strategy for the epidemic and persistence of vector-borne viruses in nature.