TRPC expression in human periodontal ligament cells and the periodontal tissue of periodontitis mice: a preliminary study / 한국실험동물학회지

Background@#Transient receptor potential canonical (TRPC) channels are non-selective cationic channels with perme‑ ability to ­Ca2+ and ­Na+ . Despite their importance, there are currently few studies on TRPC in the periodontal ligament (PDL) and bone cells in the dental field. To provide biological information regarding TRPC in PDL cells and periodontal tissue, we evaluated TRPC channels expression in the osteoblast differentiation of PDL cells and periodontitis-induced tissue. Human PDL cells were cultured in osteogenic differentiation media for 28 days, and the expression of Runx2, osteocalcin (OCN), and TRPC1, 3, 4, and 6 was evaluated by real-time PCR. In ligature-induced periodontitis mice, the alveolar bone and osteoid areas, the osteoclast number, and the expression of Runx2, OCN, TRPC3, and TRPC6 was evaluated by H&E staining, TRAP staining, and immunohistochemistry, respectively. @*Results@#In the PDL cell differentiation group, TRPC6 expression peaked on day 7 and TRPC3 expression generally increased during differentiation. During the 28 days of periodontitis progression, alveolar bone loss and osteoclast numbers increased compared to the control group during the experimental period and the osteoid area increased from day 14. TRPC6 expression in the periodontitis group increased in the PDL area and in the osteoblasts compared to the control group, whereas TRPC3 expression increased only in the PDL area on days 7 and 28. @*Conclusions@#These results indicate changes of TRPC3 and TRPC6 expression in PDL cells that were differentiating into osteoblasts and in periodontitis-induced tissue, suggesting the need for research on the role of TRPC in osteo‑ blast differentiation or periodontitis progression.

Predialysis Urea Nitrogen Is a Nutritional Marker of Hemodialysis Patients / 전남의대학술지

End-stage renal disease (ESRD) patients on hemodialysis have poor nutritional status and associated problems such as inflammation and sarcopenia. Blood urea nitrogen (BUN) is an important measure of uremic toxins, and urea reduction is a marker of hemodialysis efficacy. However, a low protein diet for lower BUN could aggravate malnutrition in patients, and optimal pre-dialysis BUN is not defined. We investigated the association of pre-dialysis BUN with patients’ comorbidities and the relationship between pre-dialysis BUN and serum albumin as a nutrient marker. Among the 67 patients, the average pre- and post-dialysis BUN were 59.2 and 15.0 mg/dL, respectively, serum creatinine was 10.1 mg/dL, and the average serum albumin was 4.0 g/dL. Patients’ age was negatively correlated with serum creatinine (r=−0.277, p<0.05) and albumin (r=−0.453, p<0.001). Predialysis BUN showed a significant positive correlation with serum albumin (r=0.287, p<0.05) and creatinine (r=0.454, p<0.001). However, the predialysis BUN was not significantly related to diabetes, coronary artery disease, congestive heart failure, or cerebrovascular disease. Hemodialysis patients with high pre-dialysis BUN and high serum creatinine could be regarded as having good nutritional status. The significance of this study lies in the potential utility of pre-dialysis blood urea nitrogen as an indicator of the nutritional status of patients. Liberal protein intake might be recommended to adequately dialyzed patients.

Regulation of the expression and function of TRPCs and Orai1 by Homer2 in mouse pancreatic acinar cells

Under physiological conditions, calcium (Ca2+ ) regulates essential functions of polarized secretory cells by the stimulation of specific Ca2+ signaling mechanisms, such as increases in intracellular Ca2+ concentration ([Ca2+]i ) via the store-operated Ca2+ entry (SOCE) and the receptor-operated Ca2+ entry (ROCE). Homer proteins are scaffold proteins that interact with G protein-coupled receptors, inositol 1,4,5-triphosphate (IP3) receptors, Orai1-stromal interaction molecule 1, and transient receptor potential canonical (TRPC) channels. However, their role in the Ca2+ signaling in exocrine cells remains unknown. In this study, we investigated the role of Homer2 in the Ca2+ signaling and regulatory channels to mediate SOCE and ROCE in pancreatic acinar cells. Deletion of Homer2 (Homer2–/– ) markedly increased the expression of TRPC3, TRPC6, and Orai1 in pancreatic acinar cells, whereas these expressions showed no difference in whole brains of wild-type and Homer2–/– mice. Furthermore, the response of Ca2+ entry by carbachol also showed significant changes to the patterns regulated by specific blockers of SOCE and ROCE in pancreatic acinar cells of Homer2–/– mice. Thus, these results suggest that Homer2 plays a critical role in the regulatory action of the [Ca2+]i via SOCE and ROCE in mouse pancreatic acinar cells.

Regulation of the expression and function of TRPCs and Orai1 by Homer2 in mouse pancreatic acinar cells

Under physiological conditions, calcium (Ca2+ ) regulates essential functions of polarized secretory cells by the stimulation of specific Ca2+ signaling mechanisms, such as increases in intracellular Ca2+ concentration ([Ca2+]i ) via the store-operated Ca2+ entry (SOCE) and the receptor-operated Ca2+ entry (ROCE). Homer proteins are scaffold proteins that interact with G protein-coupled receptors, inositol 1,4,5-triphosphate (IP3) receptors, Orai1-stromal interaction molecule 1, and transient receptor potential canonical (TRPC) channels. However, their role in the Ca2+ signaling in exocrine cells remains unknown. In this study, we investigated the role of Homer2 in the Ca2+ signaling and regulatory channels to mediate SOCE and ROCE in pancreatic acinar cells. Deletion of Homer2 (Homer2–/– ) markedly increased the expression of TRPC3, TRPC6, and Orai1 in pancreatic acinar cells, whereas these expressions showed no difference in whole brains of wild-type and Homer2–/– mice. Furthermore, the response of Ca2+ entry by carbachol also showed significant changes to the patterns regulated by specific blockers of SOCE and ROCE in pancreatic acinar cells of Homer2–/– mice. Thus, these results suggest that Homer2 plays a critical role in the regulatory action of the [Ca2+]i via SOCE and ROCE in mouse pancreatic acinar cells.

Deficiencies of Homer2 and Homer3 accelerate aging-dependent bone loss in mice

Homer proteins are scaffold proteins that regulate calcium (Ca2+) signaling by modulating the activity of multiple Ca2+ signaling proteins. In our previous report, Homer2 and Homer3 regulated NFATc1 function through its interaction with calcineurin, which then acted to regulate receptor activator of nuclear factor-kappa B ligand (RANKL)-induced osteoclastogenesis and bone metabolism. However, to date, the role of Homers in osteoclastogenesis remains unknown. In this study, we investigated the roles of Homer2 and Homer3 in aging-dependent bone remodeling. Deletion of Homer2/Homer3 (Homer2/3 DKO) markedly decreased the bone density of the femur. The decrease in bone density was not seen in mice with Homer2 (Homer2−/−) and Homer3 (Homer3−/−) deletion. Moreover, RANKL treatment of bone marrow-derived monocytes/macrophages in Homer2/3 DKO mice significantly increased the formation of multinucleated cells and resorption areas. Finally, Homer2/3 DKO mice decreased bone density in an aging-dependent manner. These findings suggest a novel potent mode of bone homeostasis regulation through osteoclasts differentiation during aging by Homer proteins, specifically Homer2 and Homer3.

Homer2 regulates amylase secretion via physiological calcium oscillations in mouse parotid gland acinar cells

The salivary glands secrete saliva, which plays a role in the maintenance of a healthy oral environment. Under physiological conditions, saliva secretion within the acinar cells of the gland is regulated by stimulation of specific calcium (Ca2+) signaling mechanisms such as increases in the intracellular Ca2+ concentration ([Ca2+]i) via storeoperated Ca2+ entry, which involves components such as Orai1, transient receptor potential (TRP) canonical 1, stromal interaction molecules, and inositol 1,4,5-triphosphate (IP3) receptors (IP3Rs). Homer proteins are scaffold proteins that bind to G protein-coupled receptors, IP3Rs, ryanodine receptors, and TRP channels. However, their exact role in Ca2+ signaling in the salivary glands remains unknown. In the present study, we investigated the role of Homer2 in Ca2+ signaling and saliva secretion in parotid gland acinar cells under physiological conditions. Deletion of Homer2 (Homer2−/− markedly decreased the amplitude of [Ca2+]i oscillations via the stimulation of carbachol, which is physiologically concentrated in parotid acinar cells, whereas the frequency of [Ca2+]i oscillations showed no difference between wild-type and Homer2−/− mice. Homer2−/− mice also showed a significant decrease in amylase release by carbachol in the parotid gland in a dose-dependent manner. These results suggest that Homer2 plays a critical role in maintaining [Ca2+]i concentration and secretion of saliva in mouse parotid gland acinar cells.

Clonorchis sinensis Infection Presenting as Acute Cholangitis and Acute Cholecystitis / 대한췌담도학회지

A 59-year-old woman presented with abdominal pain. Abdominal computerized tomography was suggestive of biliary stones. During endoscopic retrograde cholangiopancreatography, adult worms resembling Clonorchis sinensis (C. sinensis) were drained. Eggs were detected in stool using the formalin-ether concentration method and C. sinensis-specific antibody was detected in the serum. A diagnosis of C. sinensis infection was made. The symptoms of the patient gradually resolved after treatment with anti-parasite medication. The patient lived in a non-endemic region for C. sinensis infection and had no history of intake of raw or undercooked freshwater fishes. South Korea is one of the endemic countries for C. sinensis infection and people can be infected via indirect routes of transmission such as cooking utensils. Therefore, the possibility of C. sinensis infection should be considered in patients presenting with biliary diseases in South Korea. We describe the clinical findings of this case with a review of literature.

DA-6034 Induces Ca2+i Increase in Epithelial Cells

DA-6034, a eupatilin derivative of flavonoid, has shown potent effects on the protection of gastric mucosa and induced the increases in fluid and glycoprotein secretion in human and rat corneal and conjunctival cells, suggesting that it might be considered as a drug for the treatment of dry eye. However, whether DA-6034 induces Ca2+ signaling and its underlying mechanism in epithelial cells are not known. In the present study, we investigated the mechanism for actions of DA-6034 in Ca2+ signaling pathways of the epithelial cells (conjunctival and corneal cells) from human donor eyes and mouse salivary gland epithelial cells. DA-6034 activated Ca2+-activated Cl- channels (CaCCs) and increased intracellular calcium concentrations ([Ca2+]i) in primary cultured human conjunctival cells. DA-6034 also increased [Ca2+]i in mouse salivary gland cells and human corneal epithelial cells. [Ca2+]i increase of DA-6034 was dependent on the Ca2+ entry from extracellular and Ca2+ release from internal Ca2+ stores. Interestingly, these effects of DA-6034 were related to ryanodine receptors (RyRs) but not phospholipase C/inositol 1,4,5-triphosphate (IP3) pathway and lysosomal Ca2+ stores. These results suggest that DA-6034 induces Ca2+ signaling via extracellular Ca2+ entry and RyRs-sensitive Ca2+ release from internal Ca2+ stores in epithelial cells.

Activation of G Proteins by Aluminum Fluoride Enhances RANKL-Mediated Osteoclastogenesis

Receptor activator of NF-kappaB ligand (RANKL)-induced osteoclastogenesis is accompanied by intracellular Ca2+ mobilization in a form of oscillations, which plays essential roles by activating sequentially Ca2+/calmodulin-dependent protein kinase, calcineurin and NFATc1, necessary in the osteoclast differentiation. However, it is not known whether Ca2+ mobilization which is evoked in RANKL-independent way induces to differentiate into osteoclasts. In present study, we investigated Ca2+ mobilization induced by aluminum fluoride (AlF4-), a G-protein activator, with or without RANKL and the effects of AlF4- on the osteoclastogenesis in primary cultured mouse bone marrow-derived macrophages (BMMs). We show here that AlF4- induces intracellular Ca2+ concentration ([Ca2+]i) oscillations, which is dependent on extracellular Ca2+ influx. Notably, co-stimulation of AlF4- with RANKL resulted in enhanced NFATc1 expression and formation of tartrate-resistant acid phosphatase (TRAP) positive multinucleated cells. Additionally, we confirmed that mitogen-activated protein kinase (MAPK) is also activated by AlF4-. Taken together, these results demonstrate that G-protein would be a novel modulator responsible for [Ca2+]i oscillations and MAPK activation which lead to enhancement of RANKL-mediated osteoclastogenesis.

TRPM7 Is Essential for RANKL-Induced Osteoclastogenesis

The transient receptor potential melastatin type 7 (TRPM7) channel is a widely expressed non-selective cation channel with fusion to the C-terminal alpha kinase domain and regarded as a key regulator of whole body Mg2+ homeostasis in mammals. However, the roles of TRPM7 during osteoclastogenesis in RAW264.7 cells and bone marrow-derived monocyte/macrophage precursor cells (BMMs) are not clear. In the present study, we investigate the roles of TRPM7 in osteoclastogenesis using methods of small interfering RNA (siRNA), RT-PCR, patch-clamp, and calcium imaging. RANKL (receptor activator of NF-kappaB ligand) stimulation did not affect the TRPM7 expression and TRPM7-mediated current was activated in HEK293, RAW264.7, and BMM cells by the regulation of Mg2+. Knock-down of TRPM7 by siTRPM7 reduced intracellular Ca2+ concentration ([Ca2+]i) increases by 0 mM [Mg2+]e in HEK293 cells and inhibited the generation of RANKL-induced Ca2+ oscillations in RAW264.7 cells. Finally, knock-down of TRPM7 suppressed RANKL-mediated osteoclastogenesis such as activation and translocation of NFATc1, formation of multinucleated cells, and the bone resorptive activity, sequentially. These results suggest that TRPM7 plays an essential role in the RANKL-induced [Ca2+]i oscillations that triggers the late stages of osteoclastogenesis.

Fast temporal detection of intracellular hydrogen peroxide by HyPer

HyPer is the genetically encoded biosensor of intracellular hydrogen peroxide (H2O2), the most stable of the reactive oxygen species (ROS) generated by living cells. HyPer has a high sensitivity and specificity for detecting intracellular H2O2 by confocal laser microscopy. However, it was not known whether high speed ratiometric imaging of H2O2 by HyPer is possible. We thus investigated the sensitivity of HyPer in detecting changes to the intracellular H2O2 levels in HEK293 and PC12 cells using a microfluorometer imaging system. Increase in the HyPer ratio were clearly evident on stimulations of more than 100 microM H2O2 and fast changes in the HyPer ratio were observed on ratiometric fluorescent images after H2O2 treatment. These results suggest that HyPer is a potent biosensor of the fast temporal production of intracellular H2O2.

The Transcription Factor Mist1 Regulates the Cellular Polarity in Mouse Pancreatic Acinar Cells

Pancreatic acinar cells exhibit a polarity that is characterized by the localization of secretory granules at the apical membrane. However, the factors that regulate cellular polarity in these cells are not well understood. In this study, we investigated the effect of Mist1, a basic helix-loop-helix transcription factor, on the cellular architecture of pancreatic acinar cells. Mist1-null mice displayed secretory granules that were diffuse throughout the pancreatic acinar cells, from the apical to basolateral membranes, whereas Mist1 heterozygote mice showed apical localization of secretory granules. Deletion of the Mist1 gene decreased the expression of type 3 inositol 1,4,5-triphosphate receptors (IP3R) but did not affect apical localization and expression of IP3R2. Mist1-null mice also displayed an increase in luminal areas and an increase in the expression of zymogen granules in pancreatic acinar cells. These results suggest that Mist1 plays a critical role in polar localization of cellular organelles and in maintaining cellular architecture in mouse pancreatic acinar cells.

Changes of periodontopathogens and clinical parameters of periodontal tissue after debanding / 대한치과교정학회지

OBJECTIVE: The purpose of this study was to evaluate clinical and microbiological changes in periodontal tissue around the banded molars after debanding. METHODS: This study included 17 young adult patients treated with fixed orthodontic appliances including bands on the last molars more than 1 years. Probing depth and bleeding frequency were measured and plaque samples were collected from the last banded molars in all quadrants of each patient. All the data were collected immediately after debanding and 4 weeks after debanding. RESULTS: Using polymerase chain reaction based on 16S rDNA, the presence of Porphyromonas gingivalis, Tannerella forsythia and Treponema denticola was detected. After debanding, probing depth, bleeding frequency, and prevalance of periodontopathogens were reduced. Probing depth and bleeding frequency were most decreased in the buccal site of the mandibular left molar and were least decreased in the lingual site of the maxillary right molar. CONCLUSION: The results of this study indicated that proper management of oral hygiene after debanding can recuperate unfavorable periodontal condition caused by orthodontic treatment.

Changes of afferent transmission to the SI cortex by transient co-stimulation of receptive field center and outside in anesthetized rats

We have characterized the aftereffects of impulse activities on the transmission of afferent sensory to the primary somatosensory (SI) cortex of the anesthetized rats (n=22). Following conditioning stimulation (CS, 10 sec, either 5 Hz or 200 Hz) to the receptive field (RF), quantitative determination of the changes of afferent sensory transmission was done by generating post-stimulus time histogram of unit response to the testing stimulation (TS, at 0.5 Hz) to the RF center (RFC) for 60 min. In one group of experiments, CS was delivered to the RF center (RFC). In another group of experiments, CSs were simultaneously given to both RFC and RF outside (RFO, either forepaw or hindpaw). CS of 5 Hz to RFC exerted irreversible facilitation of sensory transmissions evoked by TS. Simultaneous CSs of 5 Hz to RFC and hindpaw RFO exerted reversible suppression of afferent transmission. However, CSs of 5 Hz to RFC and forepaw RFO did not significantly altered afferent sensory transmission to SI cortex neurons. CS of 200 Hz to RFC exerted irreversible suppression of sensory transmissions up to 60 min of experimental period. Simultaneous CSs of 200 Hz to RFC and RFO did not significantly altered afferent sensory transmission to SI cortex neurons. The profiles of CS-induced modulation of afferent sensory transmission were significantly different between two CS conditions. Thus, this study suggests that activity-dependent modulation of afferent transmission from a RF center to the SI cortex may be significantly altered when remote body part was simultaneously activated.