Jung-Joon Cha

Effects of HB-EGF and epiregulin on wound healing of gingival cells in vitro

OBJECTIVE Gingival wound healing is important to periodontal disease and surgery. This in vitro study was conducted to assess the manner in which heparin-binding epidermal growth factor-like growth factor (HB-EGF) and epiregulin cooperatively participate in the wound-healing process in the gingival epithelial and fibroblast cells of the oral mucosa. MATERIAL AND METHODS Gingival epithelium and fibroblast were separated from gingival tissue biopsies and prepared to primary cultures. The changes in the mRNA expression were evaluated via real-time PCR. The effects on cell proliferation, migration, and repopulation were evaluated in vitro. RESULTS The different regulation of expressions of HB-EGF, epiregulin, and epidermal growth factor receptors was observed over time and with different gingival cell types. HB-EGF exerted a cell migration-inducing effect on both epithelial and fibroblast cells, whereas epiregulin did not. Both growth factors functioned as mitogens for epithelial cell proliferation, but not for fibroblast proliferation. HB-EGF strongly promoted epithelial cell repopulation and mildly promoted fibroblast repopulation, whereas epiregulin promoted only fibroblast repopulation. CONCLUSION These results indicated that both growth factors might function importantly in the wound-healing process of human gingival tissue via the different regulation of the expression, cell migration, proliferation, and repopulation.

Td92, an outer membrane protein of Treponema denticola, induces osteoclastogenesis via prostaglandin E2‐mediated RANKL/osteoprotegerin regulation

BACKGROUND AND OBJECTIVE Periodontitis is a chronic inflammatory disease of the periodontium that causes significant alveolar bone loss. Osteoclasts are bone-resorbing multinucleated cells. Osteoblasts regulate osteoclast differentiation by expression of RANKL and osteoprotegerin (OPG). Td92 is a surface-exposed outer membrane protein of Treponema denticola, a periodontopathogen. Although it has been demonstrated that Td92 acts as a stimulator of various proinflammatory mediators, the role of Td92 in alveolar bone resorption remains unclear. Therefore, in this study, we investigated the role of Td92 in bone resorption. MATERIAL AND METHODS Mouse bone marrow cells were co-cultured with calvariae-derived osteoblasts in the presence or absence of Td92. Osteoclast formation was assessed by TRAP staining. Expressions of RANKL, osteoprotegerin (OPG) and prostaglandin E(2) (PGE(2) ) in osteoblasts were estimated by ELISA. RESULTS Td92 induced osteoclast formation in the co-cultures. In the osteoblasts, RANKL and PGE(2) expressions were up-regulated, whereas OPG expression was down-regulated by Td92. The addition of OPG inhibited Td92-induced osteoclast formation. The prostaglandin synthesis inhibitors NS398 and indomethacin were also shown to inhibit Td92-induced osteoclast formation. The effects of Td92 on the expressions of RANKL, OPG and PGE(2) in osteoblasts were blocked by NS398 or indomethacin. CONCLUSION These results suggest that Td92 promotes osteoclast formation through the regulation of RANKL and OPG production via a PGE(2) -dependent mechanism.

Effect of Abciximab on the Levels of Circulating Microparticles in Patients with Acute Myocardial Infarction Treated by Primary Angioplasty

Background and Objectives We investigated the effect of the additional use of abciximab during percutaneous coronary intervention (PCI) on the level of procoagulant microparticles (MPs) in patients with ST-segment elevation myocardial infarction (STEMI) who had undergone primary PCI. Subjects and Methods In this study, we studied 86 patients with STEMI (72 men, age 58±13) who had undergone primary PCI. The decision to administer abciximab immediately prior to PCI was left to the discretion of the operator. Blood samples for analysis of MPs were obtained from the femoral artery before and after PCI. MPs with procoagulant potential were measured using a commercial kit. The cellular origins of MPs were determined by antigenic capture with specific antibodies. Results Procoagulant MPs captured onto annexin V were not changed significantly after PCI {13.4±13.2 nM vs. 13.2±16.1 nM phosphatidylserine equivalent (PS eq), p=0.479}. Abciximab was used in 30 of 86 patients (35%) immediately prior to PCI. In patients who had undergone PCI without abciximab, no significant change in the level of MPs was observed after PCI. However, in the abciximab group, the level of circulating MPs was significantly decreased after PCI (12.0±10.7 nM vs. 7.8±11.7 nM PS eq, p=0.018). Levels of endothelial- and platelet-derived MPs also showed a significant reduction after PCI in the abciximab group. Conclusion Primary PCI with additional abciximab significantly reduced the level of procoagulant MPs regardless of their cellular origins in patients with STEMI.

Ex vivo characterization of age-associated impedance changes of single vascular endothelial cells using micro electrical impedance spectroscopy with a cell trap

We aimed to characterize aging of single vascular endothelial cells, which are indicators of senescence, using micro electrical impedance spectroscopy (μEIS) for the first time. The proposed μEIS was equipped with two barriers under the membrane actuator near the sensing electrodes, increasing its cell-trapping capability and minimizing the interference between the target cell and subsequent cells. The cell-trapping capability in μEIS with barriers was considerably improved (90%) with a capture time of 5 s or less, compared to μEIS without barriers (30%). Cells were extracted from transgenic zebrafish to minimize an initial discrepancy originating from genetic differences. In order to estimate useful parameters, cytoplasm resistance and membrane capacitance were estimated by fitting an electrical equivalent circuit to the data of ex vivo sensor output. The estimated cytoplasm resistance and membrane capacitance in the younger vascular endothelial cells were 20.16 ± 0.79 kΩ and 17.46 ± 0.76 pF, respectively, whereas those in the older cells were 17.81 ± 0.98 kΩ and 20.08 ± 1.38 pF, respectively. Discrimination of each group with different aging showed statistical significance in terms of cytoplasm resistance (p < 0.001) and membrane capacitance (p < 0.001). Considering both of the sensor and cellular level, the optimal frequency was determined as 1 MHz at which the electrical impedance of each group was clearly discriminated (p < 0.001).

Periodontitis mainly increases osteoclast formation via enhancing the differentiation of quiescent osteoclast precursors into osteoclasts.

BACKGROUND AND OBJECTIVES Tartrate-resistant acid phosphatase (TRAP)-positive multinucleated osteoclasts are formed in sequential steps: proliferation and differentiation of hematopoietic progenitors into quiescent osteoclast precursors (QOPs), followed by fusion of QOPs. In this study, we investigated whether enhancement of osteoclast formation by periodontitis is derived from the stimulation of proliferation of hematopoietic progenitors or the differentiation of QOPs into osteoclasts. MATERIAL AND METHODS Ligatures were placed around the first molars in the left mandibles of Fischer 344 inbred rats. The rats received drinking water containing bromodeoxyuridine (BrdU) (which can be incorporated into dividing nuclei) after ligation during the experimental period. The number of inflammatory cells in the distal area was counted. Alveolar bone loss was histologically estimated by measuring the distance from the cementoenamel junction to the alveolar bone crest in the distal area and determining the percentage of periodontal ligament area in the furcation. The number of osteoclasts and percentage of BrdU(+) nuclei in total osteoclasts nuclei were counted after TRAP and BrdU double labeling. RESULTS The number of polymorphonuclear cells increased on day 1 and then rapidly decreased. The number of mononuclear cells increased in a time-dependent manner up to day 5 and remained the same until day 10. Alveolar bone loss of ligatured teeth increased in a time-dependent manner. The number of osteoclasts peaked on day 3 then gradually decreased. At peak, the percentage of BrdU(+) nuclei in total osteoclasts nuclei in the distal and furcation areas were 7.9% and 4.1%, respectively. CONCLUSION These results indicate that most of the osteoclasts formed after periodontitis induction are derived from preformed QOPs, suggesting that enhancement of osteoclast formation by periodontitis might be mainly caused by stimulating the differentiation of QOPs into osteoclasts.

Spontaneous Coronary Artery Dissection Mimicking Coronary Spasm Diagnosed by Intravascular Ultrasonography

Spontaneous coronary artery dissection (SCAD) is a rare and occasionally life-threatening cause of acute coronary syndrome. Patients may present with clinical scenarios ranging from angina pectoris to cardiogenic shock to sudden cardiac death, and it may be a potentially life-threatening condition if not recognized. However, its etiology, pathophysiology and optimal therapeutic strategies have not been well understood. SCAD is diagnosed on the basis of coronary angiography, but complementary techniques as such intravascular ultrasound (IVUS) and optical coherence tomography should be considered for diagnostic clarification where appropriate. Likewise, the selection of treatment strategy depends upon the clinical manifestation, location and the extent of dissection and amount of ischemic myocardium at risk. Herein, we present the case of a 35-year-old woman who presented with acute myocardial infarction. She was diagnosed by IVUS with spontaneous diffuse dissection of the left anterior descending artery without atheroma, treated with percutaneous coronary stenting, and had a favorable clinical course and was discharged on medical therapy.

Effects of heparin-binding epidermal growth factor-like growth factor on cell repopulation and signal transduction in periodontal ligament cells after scratch wounding in vitro.

BACKGROUND AND OBJECTIVE A growing amount of attention has been placed on periodontal regeneration and wound healing for periodontal therapy. This study was conducted in an effort to determine the effects of heparin-binding epidermal growth factor-like growth factor on cell repopulation and signal transduction in periodontal ligament cells after scratch wounding in vitro. MATERIAL AND METHODS Human periodontal ligament cells were acquired from explant tissue of human healthy periodontal ligament. After the wounding of periodontal ligament cells, the change in expression of heparin-binding epidermal growth factor-like growth factor and epidermal growth factor receptors 1-4 mRNA was assessed. The effects of heparin-binding epidermal growth factor-like growth factor on periodontal ligament cell proliferation and repopulation were assessed in vitro via the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and by photographing the injuries, respectively. Extracellular signal-regulated kinase (Erk)1/2, p38 and Akt phosphorylation was characterized via western blotting. RESULTS Scratch wounding resulted in a significant up-regulation of heparin-binding epidermal growth factor-like growth factor mRNA expression, whereas wounding had no effect on the expression levels of epidermal growth factor receptors 1-4. Interestingly, no expression of epidermal growth factor receptors 2 and 4 was detectable prior to or after wounding. Heparin-binding epidermal growth factor-like growth factor treatment promoted the proliferation and repopulation of periodontal ligament cells. The scratch wounding also stimulated the phosphorylation of Erk1/2 and p38, but not of Akt, in periodontal ligament cells, and heparin-binding epidermal growth factor-like growth factor treatment applied after wounding amplified and extended the activations of Erk1/2 and p38, but not of Akt. Furthermore, Erk1/2 inhibition blocked the process of cell repopulation induced by heparin-binding epidermal growth factor-like growth factor, whereas the inhibition of p38 delayed the process. CONCLUSION These results indicate that heparin-binding epidermal growth factor-like growth factor may constitute a critical factor in the wound healing of human periodontal ligament cells by a mechanism that requires the activation of Erk1/2 via specific interaction with epidermal growth factor receptor 1.

LBPS 01-14 MICRO-ELECTROCHEMICAL IMPEDANCE SPECTROSCOPY WITH A PRESSURE-DEPENDENT MEMBRANE ACTUATOR FOR CHARACTERIZING AGE-RELATED CHANGES IN SINGLE VASCULAR ENDOTHELIAL CELL

Objective: Directly detect age-related changes in electrical impedance of single vascular endothelial cell without using biomarkers. Design and Method: Aging is one of the major factors in cardiovascular disease (CVD). However, it is difficult to differentiate changes in cardiovascular system between the aged and the diseased. If the difference could be characterized clearly, it would be potential help to understand pathophysiology. In this study, micro-electrochemical impedance spectroscopy (&mgr;EIS) with a pressure-dependent membrane actuator for cell-capturing was used to characterize age-related changes in electrical impedance of sorted single vascular endothelial cell from dissected hearts of transgenic zebrafish (fli1a:EGFP). The electrical impedance for each group (3, 4, and 18-month-old) was measured 30 times from 1 kHz to 1 MHz under 250, 300, and 350 kPa of capturing pressures, respectively. Results: Maximum differences in the average electrical impedance among the three cell groups were observed at 1 MHz, and these differences were statistically significant for all capturing pressures (p < 0.05, one-way ANOVA). Figure 1 shows the changes in the electrical impedance depending on different ages at different capturing pressure. Our results show that both electrical parameters (resistance and reactance) similarly change with aging for all capturing pressures. The resistance increased monotonously with aging, whereas the reactance decreased in 4-month-old and then increased in 18-month-old cells. Especially, 350kPa of capturing pressure could clearly discriminate electrical impedance of different aging groups. Conclusions: We applied &mgr;EIS to zebrafish vascular endothelial cells at different ages and found that resistance increased monotonously with aging, while reactance decreased during early ages and increased in aged cells. Age-related changes in resistance and reactance can be attributed to the changes in fluidity and permeability of cells due to metabolic remnants such as hydrogen ion and reactive oxygen species. In this context, the proposed &mgr;EIS could be considered as a potential diagnostic tool for CVD to detect age-related changes.